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Preview: Journal of Orthopaedic Research

Journal of Orthopaedic Research

Wiley Online Library : Journal of Orthopaedic Research

Published: 2017-12-01T00:00:00-05:00


Inflammatory and degenerative phases resulting from anterior cruciate rupture in a non-invasive murine model of post-traumatic osteoarthritis


Joint injury is the predominant risk factor for post-traumatic osteoarthritis development (PTOA). Several non-invasive mouse models mimicking human PTOA investigate molecular mechanisms of disease development; none have characterised the inflammatory response to this acute traumatic injury. Our aim was to characterise the early inflammatory phase and later degenerative component in our in vivo non-invasive murine model of PTOA induced by anterior cruciate ligament (ACL) rupture. Right knees of 12-week-old C57Bl6 mice were placed in flexion at a 30° offset position and subjected to a single compressive load (12N, 1.4mm/s) to induce ACL rupture with no obvious damage to surrounding tissues. Tissue was harvested 4 hours post-injury and on days 3, 14 and 21; contralateral left knees served as controls. Histological, immunohistochemical and gene analyses were performed to evaluate inflammatory and degenerative changes. Immunohistochemistry revealed time-dependent expression of mature (F4/80 positive) and inflammatory (CD11b positive) macrophage populations within the sub-synovial infiltrate, developing osteophytes and inflammation surrounding the ACL in response to injury. Up-regulation of genes encoding acute pro-inflammatory markers, inducible nitric oxide synthase, interleukin-6 and interleukin-17, and the matrix degrading enzymes, ADAMTS-4 and MMP3 was detected in femoral cartilage, concomitant with extensive cartilage damage and bone remodelling over 21-days post-injury. Our non-invasive model describes pathologically distinct phases of the disease, increasing our understanding of inflammatory episodes, the tissues/cells producing inflammatory mediators and the early molecular changes in the joint, thereby defining the early phenotype of PTOA. This knowledge will guide appropriate interventions to delay or arrest disease progression following joint injury. This article is protected by copyright. All rights reserved

Biomechanical Influence of Deficient Posterolateral Corner Structures on Knee Joint Kinematics: A Computational Study


The posterolateral corner (PLC) structures including the popliteofibular ligament (PFL), popliteus tendon (PT) and lateral collateral ligament (LCL) are important soft tissues for posterior translational, external rotational and varus angulation knee joint instabilities. The purpose of this study was to determine the effects of deficient PLC structures on the kinematics of the knee joint under gait and squat loading conditions. We developed subject-specific computational models with full 12-degree-of-freedom tibiofemoral and patellofemoral joints for four male subjects and one female subject. The subject-specific knee joint models were validated with computationally predicted muscle activation, electromyography data and experimental data from previous study. According to our results, deficiency of the PFL did not significantly influence knee joint kinematics compared to an intact model under gait loading conditions. Compared with an intact model under gait and squat loading conditions, deficiency of the PT led to significant increases in external rotation and posterior translation, while LCL deficiency increased varus angulation. Deficiency of all PLC structures led to the greatest increases in external rotation, varus angulation, and posterior translation. These results suggest that the PT is an important structure for external rotation and posterior translation, while the LCL is important for varus angulation under dynamic loading conditions. This article is protected by copyright. All rights reserved

The contribution of facet joints, axial compression, and composition to human lumbar disc torsion mechanics


Stresses applied to the spinal column are distributed between the intervertebral disc and facet joints. Structural and compositional changes alter stress distributions within the disc and between the disc and facet joints. These changes influence the mechanical properties of the disc joint, including its stiffness, range of motion, and energy absorption under quasi-static and dynamic loads. There have been few studies evaluating the role of facet joints in torsion. Furthermore, the relationship between biochemical composition and torsion mechanics is not well understood. Therefore, the first objective of this study was to investigate the role of facet joints in torsion mechanics of healthy and degenerated human lumbar discs under a wide range of compressive preloads. To achieve this, each disc was tested under four different compressive preloads (300-1200 N) with and without facet joints. The second objective was to develop a quantitative structure-function relationship between tissue composition and torsion mechanics. Facet joints have a significant contribution to disc torsional stiffness (∼60%) and viscoelasticity, regardless of the magnitude of axial compression. The findings from this study demonstrate that annulus fibrosus GAG content plays an important role in disc torsion mechanics. A decrease in GAG content with degeneration reduced torsion mechanics by more than an order of magnitude, while collagen content did not significantly influence disc torsion mechanics. The biochemical-mechanical and compression-torsion relationships reported in this study allow for better comparison between studies that use discs of varying levels of degeneration or testing protocols and provide important design criteria for biological repair strategies. This article is protected by copyright. All rights reserved

Characterization and Comparison of Rat Monosodium Iodoacetate and Medial Meniscal Tear Models of Osteoarthritic Pain


Osteoarthritis (OA) is a degenerative form of arthritis that can result in loss of joint function and chronic pain. The pathological pain state that develops with OA disease involves plastic changes in the peripheral and central nervous systems, however, the cellular mechanisms underlying OA are not fully understood. We characterized the medial meniscal tear (MMT) surgical model and the intra-articular injection of monosodium iodoacetate (MIA) chemical model of OA in rats. Both models produced histological changes in the knee joint and associated bones consistent with OA pathology. Both models also increased p38 activation in the L3, but not L4 dorsal root ganglia (DRG), increased tyrosine hydroxylase immunostaining in the L3 DRG indicating sympathetic sprouting, and increased phosphorylated (p)CREB in thalamic neurons. In MIA-OA, but not MMT-OA rats, p38 and pERK were increased in the spinal cord, and pCREB was enhanced in the prefrontal cortex. Using in vivo electrophysiology, elevated spontaneous activity and increased responsiveness of wide dynamic range neurons to stimulation of the knee was found in both models. However, a more widespread sensitization was observed in the MIA-OA rats as neurons with paw receptive fields spontaneously fired at a greater rate in MIA-OA than MMT-OA rats. Taken together, the MIA and MMT models of OA share several common features associated with histopathology and sensitization of primary somatosensory pathways, but, observed differences between the models highlights unique consequences of the related specific injuries, and these differences should be considered when choosing an OA model and when interpreting data outcomes. This article is protected by copyright. All rights reserved

The utility of the ankle SPECT/CT scan to predict functional and clinical outcomes in supramalleolar osteotomy patients


Combined single-photon emission computed tomography and conventional computed tomography (SPECT/CT) is a hybrid imaging modality that shows a combination of metabolic and structural information about the ankle, including arthritis. We hypothesize that uptake in specific locations within the ankle joint can be associated with both clinical outcomes and may help predict which patients will have a successful SMO. Eighty-five pre-operative SMO patients with varus (37), valgus (41), or neutral (7) alignment of the hindfoot were assessed using SPECT/CT. The level of activation on SPECT/CT scans was measured. Pre and Post-operative functional scores were recorded. Patients with medial gutter activation had significantly worse (p <.05) AOFAS alignment (AOFAS-A) scores pre-operatively. Patients with varus or valgus alignment did not have any difference in VAS pain scores, but those in valgus did have worse AOFAS-P (pain) scores. Those with cystic lesions had a worse FAOS score pre-operatively. Ten patients (12.5%) had a treatment failure. Pre or post-operative alignment did not correlate to a treatment failure. The only statistically significant (p =.036) poor prognostic indicator was a bipolar lesion. Pre-operative SPECT/CT evaluation of an ankle before a SMO can be used to clinically correlate patient-specific factors such as pain and function in the pre and post-operative period. We caution against performing a SMO in patients with bipolar activation on a pre-operative SPECT-CT scan. This article is protected by copyright. All rights reserved

Highly Metastatic K7M2 Cell Line: A Novel Murine Model Capable of In Vivo Imaging via Luciferase Vector Transfection


Osteosarcoma is rare and little improvement in survival rates has occurred in the last 25 years despite modern chemotherapeutic treatment. Bioluminescent cell lines for the modeling of osteosarcoma have shown success in tracking metastases in vivo, but commonly use adenoviral vectors to transfect the native cell line with bioluminescent reporters. The purpose of this study was to develop an orthotopic model for metastatic osteosarcoma capable of in vivo monitoring of metastatic and primary tumor burden in an immunocompetent mouse and compare that model to its wild type pathogenesis. K7M2 cells were transfected using a plasmid vector and were stable after 12 weeks. Thirty-four female BALB/c mice aged four to five weeks underwent orthotopic implantation of either wild type (n = 12) or transfected (n = 22) K7M2 cells in the proximal tibia. Mice were monitored for tumor growth and weekly In Vivo Imaging System (IVIS) imaging was performed to monitor for pulmonary metastasis. Although tumors developed sooner in the wild type group, no significant differences were seen compared to Transfected Group 1 in rate of inoculation, growth rates after first detection, metastatic rate, and time between inoculation and death. This study establishes a new murine model for metastatic osteosarcoma using the K7M2-wt cell line transfected with a non-viral plasmid luciferase vector. The benefits of this preclinical model include an intact immune system and orthotopically driven metastatic disease; this model appears comparable to its wild type counterpart. In the future, the model may be used to examine promising immunomodulatory therapies using bioluminescence in vivo. This article is protected by copyright. All rights reserved

Evidence and Mechanism by which Upper Partial Fibulectomy Improves Knee Biomechanics and Decreases Knee Pain of Osteoarthritis


To investigate the change in short-term clinical outcomes and biomechanical properties of the knee in response to upper partial fibulectomy and to probe into the biomechanical mechanism underlying the clinical benefits of upper partial fibulectomy for medial compartment knee osteoarthritis (KOA). 29 patients with medial compartment KOA underwent upper partial fibulectomy. Visual analog scale (VAS) pain, the hospital for special surgery knee score (HSS), hip-knee-ankle (HKA) angle (measured in the frontal plane), and flexion/extension range of motion of the knee were assessed before and up to 6 months after surgery. Patients and 20 healthy controls were evaluated by 3D gait analysis and dynamic lower limb musculoskeletal analysis. Both VAS pain and HSS score were significantly improved (P <0.001) one day after surgery and steadily improved during the subsequent 6 months. HKA angle improved (P = 0.025) immediately and remained stable by 3 months after surgery. The decreased overall peak KAM (decreased by 11.1%) and increased HKA angle (increased by 1.80 degrees from a more varus to more neutral alignment) of affected and operated side by 6 months after surgery were observed. Muscle activity of biceps femoris caput longum of affected and operated side increased immediately and was equivalent to healthy controls by 6 months after surgery (P = 0.007). This pilot study provides biomechanical evidence of benefit from partial upper fibulectomy and indicates a plausible rationale for the improvement in clinical symptoms. Long-term clinical outcomes and precise biomechanical mechanism of partial upper fibulectomy should be further investigated. This article is protected by copyright. All rights reserved

Altering spacer material affects bone regeneration in the Masquelet Technique in a Rat Femoral Defect


The Masquelet technique depends on pre-development of a foreign-body membrane to support bone regeneration with grafts over three times larger than the traditional maximum. To date, the procedure has always used spacers made of bone cement, which is the polymer polymethyl methacrylate (PMMA), to induce the foreign-body membrane. This study sought to compare (i) morphology, factor expression, and cellularity in membranes formed by PMMA, titanium, and polyvinyl alcohol sponge (PVA) spacers in the Masquelet milieu and (ii) subsequent bone regeneration in the same groups. Ten-week-old, male Sprague-Dawley rats were given an externally stabilized, 6mm femur defect, and a pre-made spacer of PMMA, titanium, or PVA was implanted. All animals were given 4 weeks to form a membrane, and those receiving an isograft were given 10 weeks post-implantation to union. All samples were scanned with microCT to measure phase 1 and phase 2 bone formation. Membrane samples were processed for histology to measure membrane morphology, cellularity, and expression of the factors BMP2, TGFβ, VEGF, and IL6. PMMA and titanium spacers created almost identical membranes and phase 1 bone. PVA spacers were uniformly infiltrated with tissue and cells and did not form a distinct membrane. There were no quantitative differences in phase 2 bone formation. However, PMMA induced membranes supported functional union in 6 of 7 samples while a majority of titanium and PVA groups failed to achieve the same. Spacer material can alter the membrane enough to disrupt phase 2 bone formation. The membrane's role in bone regeneration is likely more than just as a physical barrier. This article is protected by copyright. All rights reserved

In Vivo Comparison of Medialized Dome and Anatomic Patellofemoral Geometries using Subject-specific Computational Modeling


Successful outcome following total knee arthroplasty (TKA) with patella resurfacing is partly determined by the restoration of patellofemoral (PF) function and recovery of the quadriceps mechanism. The current study compared two patellar TKA geometries (medialized dome and anatomic) to determine their impact on PF mechanics and quadriceps function. In-vivo, subject-specific patellar mechanics were evaluated using a sequential experimental and modeling approach. First, stereo radiography, marker-based motion capture, and force plate data were collected for TKA patients (10 dome, 10 anatomic) performing a knee extension and lunge. Second, subject-specific, whole-body, musculoskeletal models, including 6 degrees-of-freedom (DOF) knee joint kinematics, were created for each subject and activity to predict quadriceps forces. Lastly, finite element models of each subject and activity were created to predict PF kinematics, patellar loading, moment arm, and patellar tendon angle. Differences in mechanics between dome and anatomic patients were highlighted during load-bearing (lunge) activity. Anatomic subjects demonstrated greater PF flexion angles (avg. 11 ± 3°) compared to dome subjects during lunge. Similar to the natural knee, contact locations on the patella migrated inferior to superior as the knee flexed in anatomic subjects, but remained relatively superior in dome subjects. Differences in kinematics and contact location likely contributed to altered mechanics with anatomic subjects presenting greater load transfer from the quadriceps to the patellar tendon in deep flexion (>75°), and dome subjects demonstrating larger contact forces during lunge. Although there was substantial patient variability, evaluations of PF mechanics suggested improved quadriceps function and more natural kinematics in the anatomic design. This article is protected by copyright. All rights reserved

Gene Expression Profiles of the Meniscus Avascular Phenotype: A Guide for Meniscus Tissue Engineering


Avascular (Avas) meniscus regeneration remains a challenge, which is partly a consequence of our limited knowledge of the cells that maintain this tissue region. In this study, we utilized microarrays to characterize gene expression profiles of intact human Avas meniscus tissue and of cells following culture expansion. Using these data, we examined various 3D culture conditions to redifferentiate Avas cells toward the tissue phenotype. RNA was isolated from either the tissue directly or following cell isolation and 2 weeks in monolayer culture. RNA was hybridized on human genome arrays. Differentially expressed (DE) genes were identified by ranking analysis. DAVID pathway analysis was performed and visualized using STRING analysis. Quantitative PCR (qPCR) on additional donor menisci (tissues and cells) were used to validate array data. Avas cells cultured in 3D were subjected to qPCR to compare with the array-generated data. A total of 387 genes were DE based on differentiation state (>3-fold change; p < 0.01). In Avas cultured cells, the upregulated pathways included focal adhesion, ECM-receptor interaction, regulation of actin cytoskeleton, and PDGF Signaling. In 3D-cultured Avas cells, TGFβ1 or combinations of TGFβ1 and BMP6 were most effective to promote an Avas tissue phenotype. THBS2 and THBS4 expression levels were identified as a means to denote meniscus cell phenotype status. We identified the key gene expression profiles, new markers and pathways involved in characterizing the Avas meniscus phenotype in the native state and during in vitro dedifferentiation and redifferentiation. These data served to screen 3D conditions to generate meniscus-like neotissues. This article is protected by copyright. All rights reserved

The Influence of Evolution on Cam Deformity and its Impact on Biomechanics of the Human Hip Joint


Anatomy and biomechanics of the human hip joint are a consequence of the evolution of permanent bipedal gait. Habitat and behaviour have an impact on hip morphology and significant differences are present even within the same biological family. The forces acting upon the hip joint are mainly a function of gravitation and strength of the muscles. Acetabular and femoral anatomy ensure an inherently stable hip with a wide range of motion. The femoral head in first human ancestors with upright gait was spherical (coxa rotunda). Coxa rotunda is also seen in close human relatives (great apes) and remains the predominant anatomy of present-day humans. High impact sport during adolescence with open physis however can activate an underlying genetic predisposition for reinforcement of the femoral neck, causing an epiphyseal extension and the formation of an osseous asphericity at the antero-superior femoral neck (cam deformity). The morphology of cam deformity is similar to the aspherical hips of quadrupeds (coxa recta), with the difference that in quadrupeds the asphericity is posterior. It has been postulated that this is due to the fact that humans bear weight on the extended leg, while quadrupeds bear weight at 90 − 100° flexion. The asphericity alters the biomechanical properties of the joint and as it is forced into the acetabulum leading to secondary cartilage damage. It is considered a risk factor for later development of osteoarthritis of the hip. Clinically this presents as reduced range of motion, which can be an indicator for the structural deformity of the hip. This article is protected by copyright. All rights reserved

Computational evaluation of TKR stability using feedback-controlled compressive loading


Pre-clinical assessment of stability in total knee replacement is crucial for developing preferred implant performance. Current total knee replacement patients often experience joint instability that the human body addresses with compensatory strategies. Specifically, an increased quadriceps-hamstrings co-contraction serves to increase joint stability through an increased compressive force across the tibiofemoral joint. The aim of this study is to introduce a novel method to evaluate total knee replacement by determining the compressive loading required to achieve natural knee stability. Four current total knee replacement geometries in both their cruciate-retaining and posterior-stabilized forms are modeled in a finite-element framework. The finite-element model is initially validated experimentally using traditional knee laxity testing with a constant compressive load and anterior-posterior displacement or internal-external rotation. Model predictions of constraint are in reasonable agreement with experimental results (average root mean square errors: 0.46 Nm, 62.5 N). The finite-element model is subsequently interfaced with a feedback controller to vary the compressive force that the implant requires in order to match experimental natural knee internal-external and anterior-posterior stability at different flexion angles. Results show that the lower constraint total knee replacement designs require on average 66.7% more compressive load than the higher constraint designs to achieve natural knee constraint. As expected, total knee replacement stability and compressive load requirements to replicate natural kinematics vary with inclusion of tibiofemoral ligaments. The current study represents a novel approach to evaluate stability in existing total knee replacement geometries and to design implants that better restore natural knee mechanics. This article is protected by copyright. All rights reserved

Complete tear of the lateral meniscus posterior root is associated with meniscal extrusion in anterior cruciate ligament deficient knees


This study aimed to evaluate the relationship between preoperative lateral meniscal extrusion (LME) and arthroscopic findings of lateral meniscus posterior root tear (LMPRT) in knees with anterior cruciate ligament (ACL) tear. Thirty-five knees that had LMPRTs with concomitant ACL tears on arthroscopy were evaluated. Patients were divided into two groups, partial and complete root tears, via arthroscopic findings at the time of ACL reconstruction. For comparison, we added two groups, using the same database; 20 normal knees (normal group) and 20 ACL-injured knees without LM injury (intact LM group). We retrospectively measured preoperative LMEs using magnetic resonance imaging (MRI). Twenty-three knees had partial LMPRTs. Complete LMPRTs were observed in 12 knees. The average LME was −0.1 ± 0.4 mm in the normal group, 0.2 ± 0.5 mm in the intact LM group, 0.4 ± 0.8 mm in the partial LMPRT group and 2.0 ± 0.6 mm in the complete LMPRT group. A significant difference in preoperative LMEs was observed between the complete LMPRT group and the other groups (P < 0.001). The receiver operating curve analysis, which distinguishes a partial tear from a complete tear, identified an optimal cutoff point of 1.1 mm for preoperative LME. This LME cutoff had a sensitivity of 100% and specificity of 83% for complete LMPRT. We found that preoperative LMEs were larger in complete LMPRTs associated with ACL injuries than in partial LMPRTs. Our results suggest that preoperative MRI-detected LME may be a useful indicator for estimating LMPRT severity in ACL-injured knees. This article is protected by copyright. All rights reserved

Differential Proteomic Analysis of Synovial Fluid from Hip Arthroplasty Patients with a Pseudotumor vs. Periprosthetic Osteolysis


Adverse tissue reactions to metal implants, including pseudotumors, can compromise implant functionality and survivorship. The identification of specific proteins in the synovial fluid (SF) of hip arthroplasty patients with a pseudotumor may lead to a better understanding of the underlying pathomechanisms. The objective of the present study was to compare the protein content of SF from patients with a short-term metal-on-metal hip implant associated with a pseudotumor and patients with a long-term metal-on-polyethylene hip implant associated with periprosthetic osteolysis. Discovery proteomics was used to identify differentially abundant proteins in albumin-depleted SF. In toto, 452 distinct proteins were identified. Thirty (30) of these 452 proteins were differentially abundant between the two groups, including two potential biomarkers: 6-phosphogluconate dehydrogenase (which plays a major protective role against oxidative stress) for the pseudotumor group, and scavenger receptor cysteine-rich type-1 protein M130 (which is involved in low-grade inflammation) for the periprosthetic osteolysis group. Other differentially abundant proteins identified suggest the presence of an adaptive immune response (particularly a type-IV hypersensitivity reaction), necrosis, and greater oxidative stress in patients with a pseudotumor. They also suggest the presence of an innate immune response, oxidative stress, tissue remodeling, and apoptosis in both patient groups, although differences in the specific proteins identified in each group point to differences in the pathomechanisms. Overall, results provide insights into the molecular mechanisms underlying metal-related pseudotumors and periprosthetic osteolysis, and may ultimately help elucidate pseudotumor etiology and assess the risk that asymptomatic pseudotumors will develop into an aggressive lesion. This article is protected by copyright. All rights reserved

The Challenges of Promoting Osteogenesis in Segmental Bone Defects and Osteoporosis


Conventional clinical management of complex bone healing scenarios continues to result in 5-10% of fractures forming non-unions. Additionally, the aging population and prevalence of osteoporosis related fractures necessitate the further exploration of novel ways to augment osteogenesis in this special population. This review focuses on the current clinical modalities available, and the ongoing clinical and pre-clinical research to promote osteogenesis in segmental bone defects, delayed unions, and osteoporosis. In summary, animal models of fracture repair are often small animals as historically significant large animal models, like the dog, continue to gain favor as companion animals. Small rodents have well-documented limitations in comparing to fracture repair in humans, and few similarities exist. Study design, number of studies, and availability of funding continue to limit large animal studies. Osteoinduction with rhBMP-2 results in robust bone formation, although long-term quality is scrutinized due to poor bone mineral quality. PTH 1-34 is the only FDA approved osteo-anabolic treatment to prevent osteoporotic fractures. Limited to 2 years of clinical use, PTH 1-34 has further been plagued by dose-related ambiguities and inconsistent results when applied to pathologic fractures in systematic human clinical studies. There is limited animal data of PTH 1-34 applied locally to bone defects. Gene therapy continues to gain popularity among researchers to augment bone healing. Non-integrating viral vectors and targeted apoptosis of genetically modified therapeutic cells is an ongoing area of research. Finally, progenitor cell therapies and the content variation of patient-side treatments (e.g. PRP and BMAC) are being studied. This article is protected by copyright. All rights reserved

Bone Marrow Lesions in Osteoarthritis: What Lies Beneath


Osteoarthritis (OA) is the most common joint disease in the United States, affecting more than 30 million people, and is characterized by cartilage degeneration in articulating joints. OA can be viewed as a group of overlapping disorders, which result in functional joint failure. However the precise cellular and molecular events within which lead to these clinically observable changes are neither well understood nor easily measurable. It is now clear that multiple factors, in multiple joint tissues, contribute to degeneration. Changes in subchondral bone are recognized as a hallmark of OA, but are normally associated with late-stage disease when degeneration is well established. However, early changes such as Bone Marrow Lesions (BMLs) in OA are a relatively recent discovery. BMLs are patterns from magnetic resonance images (MRI) that have been linked with pain and cartilage degeneration. Their potential utility in predicting progression, or as a target for therapy, is not yet fully understood. Here we will review the current state-of-the-art in this field under three broad headings: (1) BMLs in symptomatic OA: malalignment, joint pain and disease progression (2) biological considerations for bone-cartilage crosstalk in joint disease and (3) mechanical factors that may underlie BMLs and drive their communication with other joint tissues. Thus this review will provide insights on this topic from a clinical, biological and mechanical perspective. This article is protected by copyright. All rights reserved

Canine Hip Dysplasia: A Natural Animal Model for Human Developmental Dysplasia of the Hip


Developmental dysplasia of the hip (DDH) in humans is a common condition that is associated with hip pain, functional limitations, and secondary osteoarthritis (OA). Surgical treatment of DDH has improved in the last decade, allowing excellent outcomes at short- and mid-term follow-up. Still, the etiology, mechanobiology, and pathology underlying this disease are not well understood. A pre-clinical animal model of DDH could help advance the field with a deeper understanding of specific pathways that initiate hip joint degeneration secondary to abnormal biomechanics. An animal model would also facilitate different interventional treatments that could be tested in a rigorous, controlled environment. The dog model exhibits several important characteristics that make it valuable as a pre-clinical animal model for human DDH. Dogs are naturally prone to develop canine hip dysplasia (CHD), which is treated in a similar manner as in humans. Comparable to human DDH, CHD is considered a pre-OA disease; if left untreated it will progress to OA. However, progression to OA is significantly faster in dogs than humans, with progression to OA within one to two years of age, due to their shorter life span compared to humans. Animal studies could potentially reveal the underlying biochemical pathway(s), which can inform refined treatment modalities and provide opportunities for new treatment and prevention targets. Herein, we review the similarities and differences between the two species and outline the argument supporting CHD as an appropriate pre-clinical model of human DDH. This article is protected by copyright. All rights reserved

The pathological kinematic patterns of the tarsal complex in stage II adult-acquired flatfoot deformity


The in vivo kinematic characteristics of the tarsal joints during gait stance phase were still unclear in adult-acquired foot deformity (AAFD). This study included seven healthy subjects (fourteen feet) and twelve stage II AAFD patients (fourteen feet). The 3D models of tarsal bones were reconstructed based on CT scan. Each subject took standard gait on the single fluoroscopy system. Continuous lateral fluoroscopic images were collected. The key postures during the stance phase were selected. The 2D-3D registration technique was applied to explore the spatial motions of the tarsal joints in 6 degrees of freedom (DOF). During the whole stance phase, the AAFD talo-navicular joint (TNJ) exhibited ROM of 13±6° in the sagittal plane while the normal subjects showed ROM of 7±3° (P=0.004). In AAFD, the subtalar joint (STJ) demonstrated 19±8° and 7±3° of motion in coronal and horizontal plane respectively while the normal subjects showed 14±4°(p=0.031) and 11±3°(p=0.014) of motion respectively. Additionally STJ of AAFD patients showed significantly less dorsiflexion during the weight acceptance and showed significantly less external rotation both during the weight acceptance and single limb support of stance phase. In conclusion, for stage II AAFD patients, the talonavicular joint and the subtalar joint showed hypermobility in dorsi/planterflexion and inversion/eversion respectively during the gait stance phase while the internal/external rotation of the subtalar joint was reduced. The current study improves our understanding of the pathological kinematics of the tarsal complex in AAFD patients. Notice should be taken about these tarsal joints mobility in AAFD during clinical practice. This article is protected by copyright. All rights reserved

Analysis of skeletal muscle microcirculation in a porcine polytrauma model with haemorrhagic shock


Polytraumatised patients with haemorrhagic shock are prone to develop systemic complications, such as SIRS (systemic inflammatory response syndrome), ARDS (acute respiratory distress syndrome) and MOF (multiple organ failure). The pathomechanism of severe complications following trauma is multifactorial, and it is believed that microcirculatory dysfunction plays an important role. The aim of this study was to determine the changes in the microcirculation in musculature over time during shock and subsequent resuscitation in a porcine model of haemorrhagic shock and polytrauma. Twelve pigs (German Landrace) underwent femur fracture, liver laceration, blunt chest trauma and haemorrhagic shock under standard anaesthesia and intensive care monitoring. Microcirculation data were measured from the vastus lateralis muscle using a combined white light spectrometry and laser spectroscopy system every 15 minutes during the shock and resuscitation period, and at 24, 48 and 72 hours. Oxygen delivery and oxygen consumption were calculated and compared to baseline. The relative haemoglobin, local oxygen consumption and saturation values in the microcirculation were observed significantly lower during shock, however, no changes in the microcirculatory blood flow and microcirculatory oxygen delivery were observed. After resuscitation, the microcirculatory blood flow and relative haemoglobin increased and remained elevated during the whole observation period (72 hours). In this study, we observed changes in microcirculation during the trauma and shock phases. Furthermore, we also measured persistent dysfunction of the microcirculation over the observation period of three days after resuscitation and haemorrhagic shock. This article is protected by copyright. All rights reserved

Animal models for studying the etiology and treatment of low back pain


Chronic low back pain is a major cause of disability and health care costs. Effective treatments are inadequate for many patients. Animal models are essential to further understanding of the pain mechanism and testing potential therapies. Currently, a number of preclinical models have been developed attempting to mimic aspects of clinical conditions that contribute to low back pain. This review focused on describing these animal models and the main behavioral tests for assessing pain in each model. Animal models of low back pain can be divided into the following five categories: discogenic low back pain, radicular back pain, facet joint osteoarthritis back pain, muscle-induced low back pain, and spontaneous occurring low back pain models. These models are important not only for enhancing our knowledge of how low back pain is generated, but also for the development of novel therapeutic regimens to treat low back pain in patients. This article is protected by copyright. All rights reserved

The seating mechanics of head-neck modular tapers in vitro: Load-displacement measurements, moisture, and rate effects


The mechanically-assisted crevice corrosion performance of head-neck modular tapers is a significant concern in orthopedic biomaterials. Fretting crevice corrosion processes in modular tapers are thought to be influenced by a wide array of factors including seating mechanics of the junction, hence there is a need for in vitro test methods that can assess their performance. This study presented a test method to directly measure the load-displacement seating mechanics of modular tapers and used this method to compare the seating mechanics for different tapers, moisture, seating loads and seating rates. Seating mechanics were explored whereby the instantaneous load-displacement behavior of the head seating onto the neck is captured and used to define the mechanics of seating. Two distinct taper design/material combinations were assembled wet or dry using axially applied loads (500, 1000, 2000 and 4000 N) at two loading rates of 100 and 104 N/s (n = 5 for each condition) using a servohydraulic test frame. The results showed that pull-off strength scaled with seating load and ranged between 43-68% of seating load depending on sample and wetness. Tapers seated wet had higher pull-off strengths (2200 N +/− 300 N) than those seated dry (1800 N +/− 200 N, P < 0.05). Seating mechanics (load-displacement plots) varied due to sample type and due to wetness with differences in seating energy, seating stiffness and seating displacement. These results show the detailed mechanics of seating during assembly and provide significant insight into the complex interplay of factors associated with even “ideal” seating (axial, quasistatic) loading. This article is protected by copyright. All rights reserved

Muscle Stem Cell Activation in a Mouse Model of Rotator Cuff Injury


Rotator cuff (RC) tears are frequently complicated by muscle atrophy. Muscle stem cells (MuSCs) repair damaged myofibers following injury, but their role in the prevention or pathogenesis of atrophy following RC tears remains undefined. We hypothesized that the RC MuSC population would be affected by supraspinatus (SS) and infraspinatus (IS) tendon transection (TT) compared to uninjured muscle in a mouse model of RC tear. C57BL6/J mice underwent unilateral SS and IS TT and contralateral sham surgery. At 3, 8, or 14 weeks after injury, mice were euthanized and SS and IS were harvested for FACS sorting of CD31-/CD45-/Sca1-/ITGa7 + /VCAM+ MuSCs or histological analysis. Ki-67+ MuSCs from injured muscle increased 3.4 fold at 3 weeks (p = 0.03) and 8.1 fold at 8 weeks (p = 0.04) following TT injury, but returned to baseline by 14 weeks (p = 0.91). Myod1 remained upregulated 3.3 fold at 3 weeks (p = 0.03) and 2.0 fold at 14 weeks (p = 0.0003), respectively. Myofiber cross-sectional area was decreased at both 3 and 14 weeks after injury, but the number of MuSCs per fiber remained relatively constant at 3 (p = 0.3) and 14 (p = 0.6) weeks after TT. In this study, we characterized the longitudinal effect of RC tendon injury on the MuSC population in supraspinatus and infraspinatus muscles. MuSCs are transiently activated, and are not depleted, in spite of persistent muscle atrophy. This article is protected by copyright. All rights reserved

Effect of increased mechanical knee joint loading during running on the serum concentration of cartilage oligomeric matrix protein (COMP)


The purpose of the study was to investigate the effect of an increase in mechanical knee joint loading during running on the serum COMP level. On two different test days, 20 healthy men ran with knee orthoses for 30 min on a treadmill (v = 2.2 m/s). On day 1, the orthoses were passive, whereas on day 2 they were pneumatically driven (active) and thus increased the external knee flexion moments (+30.9 Nm) during stance phase. Lower-limb mechanics and serum COMP levels (baseline; 0, 0.5, 1, 2 h post running) were analyzed. COMP levels increased immediately after running with passive (+35%; pre: 7.5 U/l, 95%CI: 6.4, 8.7, post: 9.8 U/l, 95%CI: 8.8, 10.8, p < 0.001) and active orthoses (+45%; pre: 7.6 U/l; 95%CI: 6.4, 8.8, post: 10.3 U/l, 95%CI: 9.2, 11.5, p < 0.001), but they did not differ between interventions. While running with active orthoses, greater ankle dorsiflexion angles, knee flexion angles, and moments occurred (p < 0.05). Comparing both interventions, the Δ COMP pre–post, meaning the difference (Δ) between running with active and passive orthoses in pre to post COMP level change (=level after (post) running minus level before (pre) running), correlated negatively with Δ COMP baseline (difference between the baseline COMP level before running with active and passive orthoses, r = −0.616; p = 0.004), and with a positive tendence with the Δ maximum knee flexion (r = 0.388; p = 0.091). Therefore, changes in COMP concentration after physical activity seem to be highly influenced by the COMP baseline level. In addition, correlation analysis indicates that modifications in knee joint kinematics have a greater effect on cartilage metabolism than an increase in joint moments. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Non-enzymatic cross-linking of collagen type II fibrils is tuned via osmolality switch


An important aspect in cartilage ageing is accumulation of advanced glycation end products (AGEs) after exposure to sugars. Advanced glycation results in cross-links formation between the collagen fibrils in articular cartilage, hampering their flexibility and making cartilage more brittle. In the current study, we investigate whether collagen cross-linking after exposure to sugars depends on the stretching condition of the collagen fibrils. Healthy equine cartilage specimens were exposed to l-threose sugar and placed in hypo-, iso-, or hyper-osmolal conditions that expanded or shrank the tissue and changed the 3D conformation of collagen fibrils. We applied micro-indentation tests, contrast enhanced micro-computed tomography, biochemical measurement of pentosidine cross-links, and cartilage surface color analysis to assess the effects of advanced glycation cross-linking under these different conditions. Swelling of extracellular matrix due to hypo-osmolality made cartilage less susceptible to advanced glycation, namely, the increase in effective Young's modulus was approximately 80% lower in hypo-osmolality compared to hyper-osmolality and pentosidine content per collagen was 47% lower. These results indicate that healthy levels of glycosaminoglycans not only keep cartilage stiffness at appropriate levels by swelling and pre-stressed collagen fibrils, but also protect collagen fibrils from adverse effects of advanced glycation. These findings highlight the fact that collagen fibrils and therefore cartilage can be protected from further advanced glycation (“ageing”) by maintaining the joint environment at sufficiently low osmolality. Understanding of mechanochemistry of collagen fibrils provided here might evoke potential ageing prohibiting strategies against cartilage deterioration. © 2018 The Authors. Journal of Orthopaedic Research Published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. J Orthop Res

Stainless steel wear debris of a scoliotic growth guidance system has little local and systemic effect in an animal model


Options to treat early-onset scoliosis include guided-growth systems with sliding action between rods and pedicle screws. The wear was previously measured in an in vitro test, and in this in vivo rabbit model, we evaluated the local and systemic biological response to the stainless steel debris. Compared to the previous study, a relatively higher volume of representative wear particles with a median particle size of 0.84 μm were generated. Bolus dosages were injected into the epidural space at L4-L5 for a minimum of 36 rabbits across three treatment groups (negative control, 1.5 mg, and 4.0 mg) and two timepoints (12 and 24 weeks). Gross pathology evaluated distant organs and the injection site with a dorsal laminectomy to examine the epidural space and dosing site. Peri-implanted particle tissues were stained for immunohistochemical and quantitatively analyzed for IL-6 and TNF-α cytokines. Based on ISO 10993-6:2007 scoring, particles in the high-dose group were primarily non-irritant (12 weeks) with one slightly irritant. At 24 weeks, inflammatory cell infiltration was non-existent to minimal with all groups considered non-irritant at the injection site. Material characterization confirmed that particles detected in distant organs were stainless steel or contaminants. At 12 weeks, stainless steel groups demonstrated statistically increased amounts of cytokine levels compared to control but there was a statistical decrease for both at 24 weeks. These findings indicate that stainless steel wear debris, comparable to the expected usage from a simulated growth guidance system, had no discernible untoward biological effects locally and systemically in an animal model. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Limb asymmetry during recovery from anterior cruciate ligament reconstruction


There is limited literature that follows a population of Anterior Cruciate Ligament Reconstruction (ACLR) patients through recovery. Our aim was to examine differences in movement and loading patterns across time and between limbs over four visits during 12 months post-ACLR. We hypothesized that kinematic and kinetic data during a stop-jump would have time- and limb-dependent differences through 12 months post-surgery. Twenty-three ACLR athletes performed five vertical stop-jumps at 4, 5, 6, and 12 months post-op with motion capture and force plate data collection. The peak knee flexion (PKF) was different between the 4 and 12, 5 and 6, and the 5 and 12 month visits with earlier months exhibiting higher PKF. The peak vertical ground reaction force (vGRF) was lower at 4 than at 5 and 6 months. The peak posterior ground reaction force (pGRF) was lower at 4 months than all other visits. Frontal knee and sagittal hip range of motion (ROM) were different between 12 months and each previous visit. Asymmetries were present in peak vGRF, peak knee extension moment and impulse up to 12 months. The loading rate and peak pGRF demonstrated between limb differences up to 6 months; limb stiffness demonstrated differences up to 5 months post-ACLR. PKF was only asymmetric at the 4 month visit. While some variables improved in the 12 months post-ACLR, limb asymmetries in peak knee extension moment, peak vGRF and impulse persisted up to 12 months. Additionally, frontal plane knee and sagittal hip ROM had not normalized at 12 months. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Influence of varying stem and metaphyseal sleeve size on the primary stability of cementless revision tibial trays used to reconstruct AORI IIA defects. A simulation study


Traditionally, diaphyseal stems have been utilized to augment the stability of revision total knee replacement (rTKR) implants. More recently metaphyseal augments, such as sleeves, have been introduced to further augment component fixation. The effect of augments such as stems and sleeves have on the primary stability of a rTKR implant is poorly understood, however it has important implications on the complexity, costs and survivorship of the procedure. Finite element analysis was used to investigate the primary stability and strain distribution of various size stems and sleeves used in conjunction with a cementless revision tibial tray. The model was built from computer tomography images of a single healthy tibia obtained from an 81-year-old patient to which an Anderson Orthopaedic Research Institute (AORI) IIA defect was virtually added. The influences of varying body mass index (BMI) and bone modulus were also investigated. Stemless sleeves were found to provided adequate primary implant stability (average implant micro-motion <50 μm) for the studied defect. Addition of a stem did not enhance the primary stability. Furthermore, this study found that varying BMI and bone modulus had a considerable effect on strain distribution but negligible effect on micro-motion in the sleeve area. In conclusion, the addition of diaphyseal stem to a metaphyseal sleeve had little benefit in enhancing the primary stability of tibial trays augmented when simulating reconstructions of AORI IIA tibial defects. Additional studies are required to determine the relative benefit of the diaphyseal stem when using metaphyseal sleeves defects with more extensive bone loss. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res Recently metaphyseal sleeves have been introduced to further augment revision tibial tray fixation. In this study, Finite element analysis was used to investigate the primary stability and strain distribution of various size stems and sleeves used in conjunction with a cementless revision tibial tray. This study found that, the addition of diaphyseal stem to a metaphyseal sleeve had little benefit in enhancing the primary stability of revision tibial trays in an AORI IIA defects.

Quantitative evaluation of retrieved reverse total shoulder arthroplasty liner surface deviation and volumetric wear


Polyethylene wear is a known complication in total joint arthroplasty, however, in vivo wear rates in reverse total shoulder arthroplasty (RTSA) remain largely unknown. This study aimed to quantify volumetric and surface deviation changes in retrieved RTSA humeral liners using a novel micro-computed tomography (μCT)-based technique. After IRB-approval, 32 humeral liners (single manufacturer and model) with term-of-service greater than 90 days were analyzed. Clinical demographics and surgical data were collected via chart review. Unworn liners were used as geometric controls. Retrieved and unworn liners underwent μCT scanning. Retrieved liner volumes were isolated, co-registered to controls of matching geometry, and surface deviations of the articulation surface and rim were computed. Differences in total volume loss (TVL), volumetric wear rate (VWR), and surface deviation were reported. Semi-quantitative grading evaluated rim damage presence and severity. Mean term-of-service for all liners was 2.07 ± 1.33 years (range: 0.30–4.73). Mean TVL and VWR were 181.3 ± 208.2 mm3 and 114.5 ± 160.3 mm3/year, respectively. Mean articulation and rim surface deviations were 0.084 ± 0.065 and 0.177 ± 0.159 mm, respectively. Articulation surface deviation was positively correlated to term-of-service. Rim damage was present on 63% of liners and correlated significantly to rim surface deviation. This study reports in vivo wear rates of retrieved RTSA implants. Our results demonstrate volumetric and articulation surface wear in select RTSA liners that is correlated to term-of-service. Calculation of in vivo wear rates can help bridge the gap between clinical outcomes and experimental models such as wear simulations and computational models. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Fixed-bearing medial unicompartmental knee arthroplasty restores neither the medial pivoting behavior nor the ligament forces of the intact knee in passive flexion


Medial unicompartmental knee arthroplasty (UKA) is an accepted treatment for isolated medial osteoarthritis. However, using an improper thickness for the tibial component may contribute to early failure of the prosthesis or disease progression in the unreplaced lateral compartment. Little is known of the effect of insert thickness on both knee kinematics and ligament forces. Therefore, a computational model of the tibiofemoral joint was used to determine how non-conforming, fixed bearing medial UKA affects tibiofemoral kinematics, and tension in the medial collateral ligament (MCL) and the anterior cruciate ligament (ACL) during passive knee flexion. Fixed bearing medial UKA could not maintain the medial pivoting that occurred in the intact knee from 0° to 30° of passive flexion. Abnormal anterior–posterior (AP) translations of the femoral condyles relative to the tibia delayed coupled internal tibial rotation, which occurred in the intact knee from 0° to 30° of flexion, but occurred from 30° to 90° of flexion following UKA. Increasing or decreasing tibial insert thickness following medial UKA also failed to restore the medial pivoting behavior of the intact knee despite modulating MCL and ACL forces. Reduced AP constraint in non-conforming medial UKA relative to the intact knee leads to abnormal condylar translations regardless of insert thickness even with intact cruciate and collateral ligaments. This finding suggests that the conformity of the medial compartment as driven by the medial meniscus and articular morphology plays an important role in controlling AP condylar translations in the intact tibiofemoral joint during passive flexion. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Nondestructive, indirect assessment of the biomechanical properties of the rat intervertebral disc using contrast-enhanced μCT


Mechanical characterization of the intervertebral disc involves labor-intensive and destructive experimental methodology. Contrast-enhanced micro-computed tomography is a nondestructive imaging modality for high-resolution visualization and glycosaminoglycan quantification of cartilaginous tissues. The purpose of this study was to determine whether anionic and cationic contrast-enhanced micro-computed tomography of the intervertebral disc can be used to indirectly assess disc mechanical properties in an ex vivo model of disc degeneration. L3/L4 motion segments were dissected from female Lewis rats. To deplete glycosaminoglycan, samples were treated with 0 U/ml (Control) or 5 U/ml papain. Contrast-enhanced micro-computed tomography was performed following incubation in 40% Hexabrix (anionic) or 30 mg I/ml CA4+ (cationic) for 24 h (n = 10/contrast agent/digestion group). Motion segments underwent cyclic mechanical testing to determine compressive and tensile modulus, stiffness, and hysteresis. Glycosaminoglycan content was determined using the dimethylmethylene blue assay. Correlations between glycosaminoglycan content, contrast-enhanced micro-computed tomography attenuation, and mechanical properties were assessed via the Pearson correlation. The predictive accuracy of attenuation on compressive properties was assessed via repeated random sub-sampling cross validation. Papain digestion produced significant decreases in glycosaminoglycan content and corresponding differences in attenuation and mechanical properties. Attenuation correlated significantly to glycosaminoglycan content and to all compressive mechanical properties using both Hexabrix and CA4+. Predictive linear regression models demonstrated a predictive accuracy of attenuation on compressive modulus and stiffness of 79.8–86.0%. Contrast-enhanced micro-computed tomography was highly predictive of compressive mechanical properties in an ex vivo simulation of disc degeneration and may represent an effective modality for indirectly assessing disc compressive properties. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Acetabular and spino-pelvic morphologies are different in subjects with symptomatic cam femoro-acetabular impingement


Acetabular and spino-pelvic (SP) morphological parameters are important determinants of hip joint dynamics. This prospective study aimed to determine whether acetabular and SP morphological differences exist between hips with and without cam morphology and between symptomatic and asymptomatic hips with cam morphology. A cohort of 67 patients/hips was studied. Hips were either asymptomatic with no cam (Controls, n = 18), symptomatic with cam (n = 26) or asymptomatic with cam (n = 23). CT-based quantitative assessments of femoral, acetabular, pelvic, and spino-pelvic parameters were performed. Measurements were compared between controls and those with a cam deformity, as well as between the three groups. Morphological parameters that were independent predictors of a symptomatic cam were determined using a regression analysis. Hips with cam deformity had slightly smaller subtended angles superior-anteriorly (87° vs. 84°, p = 0.04) and greater pelvic incidence (53° vs. 48°, p = 0.003) compared to controls. Symptomatic cams had greater acetabular version (p < 0.01), greater subtended angles superiorly and superior-posteriorly (p = 0.01), higher pelvic incidence (p = 0.02), greater alpha angles and lower femoral neck-shaft angles compared to asymptomatic cams (p < 0.01) and controls (p < 0.01). The four predictors of symptomatic cam included antero-superior alpha angle, femoral neck-shaft angle, acetabular depth, and pelvic incidence. In conclusion, this study illustrates that symptomatic hips had a greater amount of supero-posterior coverage; which would be the contact area between a radial cam and the acetabulum, when the hip is flexed to 90°. Furthermore, individuals with symptomatic cam morphology had greater PI. Acetabular- and SP parameters should be part of the radiological assessment of femoro-acetabular impingement. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

In-vitro oxidation model for UHMWPE incorporating synovial fluid lipids


Post-irradiation melting of ultra-high molecular weight polyethylene (UHMWPE) reduced the oxidation potential of UHMWPE in vivo. After mid-term (5–10 years) use in vivo, there is detectable oxidation in irradiated and melted joint implant retrievals. The absorption of the synovial fluid lipid squalene was identified as a possible factor initiating oxidation. We investigated the role of lipids in UHMWPE oxidation by asking: (1) Do other synovial fluid lipids initiate oxidation in irradiated and melted UHMWPE?; (2) What is the effect of the absorption of multiple lipids on UHMWPE oxidation?; (3) How does lipid-initiated oxidation in vitro compare to what is observed in long-term retrievals? We diffused emulsified single and mixed lipids into irradiated and melted UHMWPE and accelerated aged them. We analyzed the oxidation in these samples and in four long-term highly crosslinked, irradiated, and melted Longevity™ UHMWPE liner retrievals (in vivo for up to 190 months) using Fourier Transform Infrared Spectroscopy (FTIR). We showed that lipids other than squalene could initiate oxidation in UHMWPE and that the types of absorbed lipids determined the amount of resultant oxidation. Although mixed lipids doping and accelerated aging reproduced the average and maximum oxidation values and oxidation products observed in vivo, the oxidation depth profile and its effect on cross-link density was different. One reason for this was the variability of oxidation in retrievals, suggesting additional factors contributing to oxidation. The understanding of oxidative processes in vivo and the development of clinically relevant in vitro protocols to evaluate implant materials is crucial for their long-term performance. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Immunotherapy synergizes with debridement and antibiotic therapy in a murine 1-stage exchange model of MRSA implant-associated osteomyelitis


Methicillin-resistant Staphylococcus aureus (MRSA) reinfection following revision surgery remains a major orthopaedic problem. Toward the development of immunotherapy with anti-glucosaminidase monoclonal antibodies (anti-Gmd), we aimed to: (i) develop a murine 1-stage exchange model of bioluminescent MRSA (USA300LAC::lux) contaminated femoral implants; and (ii) utilize this model to demonstrate the synergistic effects of combination vancomycin and anti-Gmd therapy on reinfection and bone healing. Following an infection surgery, the original plate and two screws were removed on day 7, and exchanged with sterile implants. Mice were randomized to five groups: (i) no infection control; (ii) infected placebo; (iii) anti-Gmd; (iv) vancomycin; and (v) combination therapy. Bioluminescent imaging (BLI) was performed on days 0, 1, 3, 5, 7, 8, 10, 12, and 14. Mice were euthanized on day 14 (day 7 post-revision), and efficacy was assessed via colony forming units (CFU) on explanted hardware, micro-CT, and histology. As monotherapies, anti-Gmd inhibited Staphylococcus abscess communities, and vancomycin reduced CFU on the implants. However, only combination therapy prevented increased BLI post-revision surgery, with a significant 6.5-fold reduction on day 10 (p < 0.05 vs. placebo), and achieved sterile implant levels by day 12. Synergistic effects were also apparent from reduced osteolysis and increased new bone formation around the screws only observed following combination therapy. Taken together, we find that: (i) this murine femoral plate 1-stage revision model can efficiently evaluate therapies to prevent reinfection; and (ii) immunotherapy plays a distinct role from antibiotics to reduce reinfection following revision surgery, such that synergy to achieve osseointegration is possible. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 9999:1–9, 2018.

Metabolomic serum profiling after ACL injury in rats: A pilot study implicating inflammation and immune dysregulation in post-traumatic osteoarthritis


ACL rupture is a major risk factor for post-traumatic osteoarthritis (PTOA) development. Little information exists on acute systemic metabolic indicators of disease development. Thirty-six female Lewis rats were randomized to Control or noninvasive anterior cruciate ligament rupture (ACLR) and to three post-injury time points: 72 h, 4 weeks, 10 weeks (n = 6). Serum was collected and analyzed by 1H nuclear magnetic resonance (NMR) spectroscopy and combined direct injection and liquid chromatography (LC)-mass spectrometry (MS)/MS (DI-MS). Univariate and multivariate statistics were used to analyze metabolomic data, and predictive biomarker models were analyzed by receiver operating characteristic (ROC) analysis. Topological pathway analysis was used to identify perturbed pathways. Two hundred twenty-two metabolites were identified by 1H NMR and DI-MS. Differences in the serum metabolome between ACLR and Control were dominated by medium- and long-chain acylcarnitine species. Further, decreases in several tryptophan metabolites were either found to be significantly different in univariate analysis or to play important contributory roles to multivariate model separation. In addition to acylcarnitines and tryptophan metabolites, glycine, carnosine, and D-mannose were found to differentiate ACLR from Control. Glycine, 9-hexadecenoylcarnitine, trans-2-Dodecenoylcarnitine, linoelaidyl carnitine, hydroxypropionylcarnitine, and D-Mannose were identified as biomarkers with high area under ROC curve values and high predictive accuracies. Our analysis provides new information regarding the potential contribution of inflammatory processes and immune dysregulation to the onset and progression of PTOA following ACL injury. As these processes have most commonly been associated with inflammatory arthropathies, larger-scale studies elucidating their involvement in PTOA development and progression are necessary. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Comparing damage on retrieved total elbow replacement bushings with lab worn specimens subjected to varied loading conditions


Complication rates following total elbow replacement (TER) with conventional implants are relatively high due to mechanical failure involving the UHMWPE bushings. Unfortunately, there are no standardized pre-clinical durability testing protocols for assessing the durability of TER components. This study examines the damage observed on retrieved humeral bushings, and then uses in vitro durability testing with two different loading protocols to compare resulting damage. Damage on 25 pairs of retrieved humeral bushings was characterized using micro-computed tomographic imaging techniques. The damage was compared with that of in vitro test specimens which were subjected to 200 K cycles of either high joint reaction force (high JRF) or high varus moment (high VM) loading. Material removal (mass loss) from bushing components was measured using gravimetric techniques. Thinning was less for retrieved bushings which were still assembled in their humeral component, versus bushings which were loose (0.3 ± 0.3 mm vs. 0.6 ± 0.3 mm, p = 0.02). Comparing in vitro test specimens, thinning due to high VM loading was 0.9 ± 0.3 mm, versus 0.2 ± 0.0 mm for high JRF loading (p = 0.08); however, the actual material removal rates from the humeral bushings were not different between the two protocols (48 ± 5 mm3/Mc vs. 43 ± 2 mm3/Mc, p = 1). Neither loading protocol could produce damage patterns fully representative of the spectrum of damage patterns observed on clinical retrievals. Pre-clinical testing should employ multiple loading protocols to characterize implant performance under a broader spectrum of usage. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res This study examines retrieved humeral bushings versus bushings subjected to two different in vitro durability tests. Bushing thinning was worse for specimens subjected to high varus moments versus high joint reaction force loading; however, the actual material removal rates were not different between the two protocols. Neither loading protocol could produce damage patterns fully representative of the spectrum of damage patterns observed on clinical retrievals, therefore pre-clinical testing should employ multiple loading protocols to consider a broad spectrum of usage.

Effect of connective tissue growth factor delivered via porous sutures on the proliferative stage of intrasynovial tendon repair


Recent growth factor, cell, and scaffold-based experimental interventions for intrasynovial flexor tendon repair have demonstrated therapeutic potential in rodent models. However, these approaches have not achieved consistent functional improvements in large animal trials due to deleterious inflammatory reactions to delivery materials and insufficient induction of targeted biological healing responses. In this study, we achieved porous suture-based sustained delivery of connective tissue growth factor (CTGF) into flexor tendons in a clinically relevant canine model. Repairs with CTGF-laden sutures were mechanically competent and did not show any evidence of adhesions or other negative inflammatory reactions based on histology, gene expression, or proteomics analyses at 14 days following repair. CTGF-laden sutures induced local cellular infiltration and a significant biological response immediately adjacent to the suture, including histological signs of angiogenesis and collagen deposition. There were no evident widespread biological effects throughout the tendon substance. There were significant differences in gene expression of the macrophage marker CD163 and anti-apoptotic factor BCL2L1; however, these differences were not corroborated by proteomics analysis. In summary, this study provided encouraging evidence of sustained delivery of biologically active CTGF from porous sutures without signs of a negative inflammatory reaction. With the development of a safe and effective method for generating a positive local biological response, future studies can explore additional methods for enhancing intrasynovial tendon repair. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res Porous suture-based sustained delivery of connective tissue growth factor (CTGF) into canine flexor tendons induced local cellular infiltration and significant biological activity immediately adjacent to the suture at 14 days following repair. Repairs with CTGF-laden sutures were mechanically competent and did not show any evidence of adhesions or other negative inflammatory reactions based on histology, gene expression, or proteomics analyses. This study developed a safe and effective method for generating a local biological response to enhance intrasynovial tendon repairs.

Sclerostin antibody enhances bone formation in a rat model of distraction osteogenesis


Neutralizing monoclonal sclerostin antibodies are effective in promoting bone formation at a systemic level and in orthopedic scenarios including closed fracture repair. In this study we examined the effects of sclerostin antibody (Scl-Ab) treatment on regenerate volume, density, and strength in a rat model of distraction osteogenesis. Surgical osteotomy was performed on 179 Sprague Dawley rats. After 1 week, rats underwent distraction for 2 weeks, followed by 6 weeks for consolidation. Two treatment groups received biweekly subcutaneous Scl-AbIII (a rodent form of Scl-Ab; 25 mg/kg), either from the start of distraction onward or restricted to the consolidation phase. These groups were compared to controls receiving saline. Measurement modalities included longitudinal DXA, ex vivo QCT, and microCT, tissue histology, and biomechanical four-point bending tests. Bone volume was increased in both Scl-Ab treatments regimens by the end of consolidation (+26–38%, p < 0.05), as assessed by microCT. This was associated with increased mineral apposition. Importantly, Scl-Ab led to increased strength in united bones, and this reached statistical significance in animals receiving Scl-Ab during consolidation only (+177%, p < 0.01, maximum load to failure). These data demonstrate that Scl-Ab treatment increases bone formation, leading to regenerates with higher bone volume and improved strength. Our data also suggest that the optimal effects of Scl-Ab treatment are achieved in the latter stages of distraction osteogenesis. These findings support further investigation into the potential clinical application of sclerostin antibody to augment bone distraction, such as limb lengthening, particularly in the prevention of refracture. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Osteochondral repair using an acellular dermal matrix—pilot in vivo study in a rabbit osteochondral defect model


The aim of this pilot project was to introduce a novel use of acellular dermal matrix (ADM) in combination with infrapatellar fat pad mesenchymal stromal cells (IPFP-MSCs) to effect repair in a rabbit osteochondral defect model. ADM, in a range of surgical procedures, has been shown to promote remodelling of tissue at the site of implantation. Rabbit-derived ADM (rabADM) was prepared from the skin of donor rabbits. Autologous IPFP-MSCs were obtained at the time of knee surgery. Osteochondral defects (4 mm cartilage outer/2 mm central bone defect) were drilled into distal femoral condyles of 12 New Zealand White rabbits. Treatments groups: (i) defect only; (ii) rabADM alone; (iii) IPFP-MSCs alone; and (iv) rabADM with IPFP-MSCs. Condyles were harvested at 12 weeks, and analyzed using histology, immunohistochemistry (types I and II collagen) and histomorphometry to evaluate osteochondral repair. The rabADM only group achieved the highest ratio of type II to non-type II collagen (77.3%) using areal measures (similar to normal cartilage), which indicated a higher quality of cartilage repair. The addition of IPFP-MSCs, with or without rabADM, formed a fibrous collagen cap above the lesion site not seen with rabADM alone. Macroscopically, there was no joint erosion, inflammation, swelling or deformity, and all animals maintained full range of motion. Conclusions: RabADM alone resulted in neocartilage formation similar to native cartilage. IPFP-MSCs limited osteochondral repair and contributed to fibrosis, even in combination with the rabADM. Further studies using ADM for osteochondral repair are warranted in a more appropriate pre-clinical model of osteochondral repair. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res Rabbit-derived acellular dermal matrix (rabADM) in combination with/without infrapatellar fat pad mesenchymal stromal cells (IPFP-MSCs) was utilized in a 12-week pilot study in a rabbit osteochondral defect model. Treatments were: defect only, rabADM alone, IPFP-MSCs alone, and with rabADM. RabADM alone achieved the highest ratio of type II to non-type II collagen (77.3%) (immunohistochemical assessment), using areal measures, similar to native cartilage. IPFP-MSCs limited osteochondral repair and contributed to fibrosis. Further studies using ADM for osteochondral repair are warranted.

Cartilage quantitative T2 relaxation time 2–4 years following isolated anterior cruciate ligament reconstruction


Cartilage T2 relaxation time in isolated anterior cruciate ligament reconstruction (ACLR) without concomitant meniscal pathology and their changes over time remain unclear. The purpose of this exploratory study was to: (i) compare cartilage T2 relaxation time (T2 values) in people with isolated ACLR at 2–3 years post-surgery (baseline) and matched healthy controls and; (ii) evaluate the subsequent 2-year change in T2 values in people with ACLR. Twenty-eight participants with isolated ACLR and nine healthy volunteers underwent knee magnetic resonance imaging (MRI) at baseline; 16 ACLR participants were re-imaged 2 years later. Cartilage T2 values in full thickness, superficial layers, and deep layers were quantified in the tibia, femur, trochlear, and patella. Between-group comparisons at baseline were performed using analysis of covariance adjusting for age, sex, and body mass index. Changes over time in the ACLR group were evaluated using paired sample t-tests. ACLR participants showed significantly higher (p = 0.03) T2 values in the deep layer of medial femoral condyle at baseline compared to controls (mean difference 4.4 ms [13%], 95%CI 0.4, 8.3 ms). Over 2 years, ACLR participants showed a significant reduction (p = 0.04) in T2 value in the deep layer of lateral tibia (mean change 1.4 ms [−7%], 95%CI 0.04, 2.8 ms). The decrease in T2 values suggests improvement in cartilage composition in the lateral tibia (deep layer) of ACLR participants. Further research with larger ACLR cohorts divided according to meniscal status and matched healthy cohorts are needed to further understand cartilage changes post-ACLR. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 9999:1–8, 2018. Twenty-eight individuals with isolated ACL reconstruction (ACLR) exhibited significantly (p < 0.05) higher T2 relaxation time values—suggestive of cartilage degeneration—in the deep layer of the medial femoral condyle compared with controls (n = 9) at 2–3 years post-surgery. Somewhat surprisingly, ACLR individuals (n = 16) showed a significant decrease in T2 values in the deep layer of the lateral tibia from 2 to 4 years post-surgery, suggesting a partial improvement in cartilage composition.

Biomarkers for equine joint injury and osteoarthritis


We report the results of a symposium aimed at identifying validated biomarkers that can be used to complement clinical observations for diagnosis and prognosis of joint injury leading to equine osteoarthritis (OA). Biomarkers might also predict pre-fracture change that could lead to catastrophic bone failure in equine athletes. The workshop was attended by leading scientists in the fields of equine and human musculoskeletal biomarkers to enable cross-disciplinary exchange and improve knowledge in both. Detailed proceedings with strategic planning was written, added to, edited and referenced to develop this manuscript. The most recent information from work in equine and human osteoarthritic biomarkers was accumulated, including the use of personalized healthcare to stratify OA phenotypes, transcriptome analysis of anterior cruciate ligament (ACL) and meniscal injuries in the human knee. The spectrum of “wet” biomarker assays that are antibody based that have achieved usefulness in both humans and horses, imaging biomarkers and the role they can play in equine and human OA was discussed. Prediction of musculoskeletal injury in the horse remains a challenge, and the potential usefulness of spectroscopy, metabolomics, proteomics, and development of biobanks to classify biomarkers in different stages of equine and human OA were reviewed. The participants concluded that new information and studies in equine musculoskeletal biomarkers have potential translational value for humans and vice versa. OA is equally important in humans and horses, and the welfare issues associated with catastrophic musculoskeletal injury in horses add further emphasis to the need for good validated biomarkers in the horse. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Altered erector spinae activity and trunk motion occurs with moderate and severe unilateral hip OA


People with hip osteoarthritis (OA) demonstrate altered movement patterns in the hip joint, as well as the pelvis and spine. While kinematic changes have been described in the literature, little is known about the associated erector spinae (ES) activity. Increased or prolonged ES activity may contribute to the low back pain often associated with hip OA. Using a cross-sectional cohort study, 3D trunk motions and ES surface electromyography were recorded on 19 individuals with severe OA (SOA), 20 with moderate hip OA (MOA), and 19 asymptomatic (ASYM) individuals during treadmill walking, using standardized collection and processing procedures. Principal component analysis was used to derive electromyographic amplitude and temporal waveform features. Three-dimensional thoracic motion in a global system, and thoraco-lumbar motion was calculated. Various statistical analyses determined between group differences (α = 0.05). In the sagittal plane, thoracic motion was greater in the SOA group (p < 0.001), whereas the ASYM group used less thoraco-lumbar motion than either OA group (p ≤ 0.002). Greater frontal plane angular excursion during early stance was found in the thoracic region in the SOA group (p ≤ 0.001) . With increasing OA severity, bilateral ES activity increased during the swing phase of gait (p < 0.001), whereas during stance, the SOA ipsilateral ES activity was higher than other groups (p < 0.001). Statement of clinical significance: with moderate and severe OA, sagittal, and frontal trunk motion increases during gait. ES activity during the entire gait cycle is more sustained with increased disease severity, which may aide our understanding of low back pain associated with hip OA. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Subchondral bone fragility with meniscal tear accelerates and parathyroid hormone decelerates articular cartilage degeneration in rat osteoarthritis model


The aims of this study were to investigate the influence of subchondral bone fragility (SBF) on the progression of the knee osteoarthritis by using a novel rat model, and to examine the preventive effect of parathyroid hormone (PTH) on cartilage degeneration. First, 40 rats were assigned to the following four groups: Sham, SBF, Medial meniscal tear (MMT), and MMT + SBF groups. In SBF and MMT + SBF groups, we induced SBF by microdrilling the subchondral bone. Second, 10 additional rats were randomly assigned to the following two groups: MMT + SBF + saline and MMT + SBF + PTH groups. Osteoarthritic changes in the articular cartilage and subchondral bone were evaluated using safranin-O/fast green staining, matrix metalloproteinase-13 (MMP-13), and type X collagen immunohistochemistry, toluidine blue staining, and micro-CT scanning. The combination of SBF and meniscal tear increased the number of mast cells in the subchondral bone, and led to the abnormal subchondral bone microarchitecture, such as abnormally decreased trabecular number and increased trabecular thickness, compared with meniscal tear alone. Moreover, SBF with meniscal tear enhanced articular cartilage degeneration and increased the expression of MMP-13 and type X collagen, compared with meniscal tear alone. The administration of PTH decreased the number of mast cells in the subchondral bone and improved the microstructural parameters of the subchondral bone, and delayed the progression of articular cartilage degeneration. These results suggest that SBF is one of the factors underlying the osteoarthritis development, especially in knees with traumatic osteoarthritis, and that the administration of PTH is a potential therapeutic treatment for preventing OA progression. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Effect of modulating dietary vitamin D on the general bone health of rats during posterolateral spinal fusion


Vitamin D plays a significant role in musculoskeletal health by regulating calcium, phosphate, and promoting new bone mineralization. The purpose of this study was to understand the effect of dietary vitamin D on general bone health during peri-operative bone healing via an in vivo dosing study of vitamin D in a rat posterolateral fusion model using autograft. Vitamin D Deficient (DD), vitamin D Insufficient (ID), Control vitamin D (CD), and Hyper-vitamin D (HD) groups were studied. Increasing dietary vitamin D improved quantitative measures of femoral geometry, including femoral strength, stiffness, and density. Femoral biomechanics, cortical thickness, moment of inertia, cross-sectional area, and measures from bone ashing were all greater in the HD group versus the CD. This suggests that additional dietary vitamin D above normal levels during spinal fusion may lead to improvement in bone health. Serum vitamin D levels were also observed to decrease during fusion healing. These results demonstrate that dietary vitamin D improves general bone health in the femur of a rat model during posterolateral spinal fusion. This suggests a role for further clinical evaluation of vitamin D dietary intake during the peri-operative period, with the possibility of avoiding adverse consequences to general bone health. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res This study evaluated dietary vitamin D's effect on bone health during peri-operative bone healing via an in vivo dosing study of vitamin D in a rat posterolateral fusion model. Dietary vitamin D improved femoral biomechanics, cortical thickness, moment of inertia, cross-sectional area, and bone ashing measures. Results indicate dietary vitamin D above normal levels during spinal fusion may lead to improvement in bone health. Study provides basis for a clinical study of vitamin D dietary intake during the peri-operative period.

In vivo correlates between daily physical activity and intervertebral disc health


Physical activity impacts health and disease in multiple body tissues including the intervertebral discs. Fluid flow within the disc is an indicator of disc health that can be observed using diffusion weighted magnetic resonance imaging. We monitored activity levels of 26 participants, age 35–55 yrs, using Actigraph accelerometers for 4 days to evaluate vigorous-intensity activity, moderate to vigorous intensity activity, and sedentary time. Participants underwent structural and diffusion weighted magnetic resonance imaging to evaluate intervertebral disc health and fluid flow. They also underwent bone density scans, carotid artery ultrasounds, a treadmill test, and a physical exam for pain, range of motion, and instability. These measures were used to correlate MRI indicators of intervertebral disc health with participant activity levels. Participants with any vigorous-intensity physical activity compared with no vigorous-intensity activity had significantly greater L5/S1 apparent diffusion coefficient values (p = 0.002), corresponding to higher freedom of diffusive movement for cellular nutrients and metabolic waste. Sagittal T2 values in the L5/S1 were also higher (p = 0.004), corresponding to a higher water content in the discs. Higher apparent diffusion coefficients were also found in participants with more than 30 min compared with less than 30 min of daily moderate to vigorous physical activity (p = 0.03), and in participants with less than 67% awake time as sedentary time compared with more than 67% sedentary time (p = 0.03). Increased dynamic loading through physical activity and decreased static loading from sedentary time benefit intervertebral disc health. Physical activity, particularly vigorous activity, is beneficial in helping maintain intervertebral disc health. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Effects of combined teriparatide and zoledronic acid on posterior lumbar vertebral fusion in an aged ovariectomized rat model of osteopenia


There has been no study regarding the effect of a combination of teriparatide (TPTD) and zoledronic acid (ZA) on vertebral fusion. In this study, we investigate the effect of single and combined TPTD and ZA treatment on lumbar vertebral fusion in aged ovariectomized (OVX) rats. Sixty two-month-old female Sprague-Dawley rats were ovariectomized and underwent bilateral L4–L5 posterolateral intertransverse fusion after 10 months. The OVX rats received vehicle (control) treatment, or ZA (100 µg/kg, once), or TPTD (60 µg/kg/2 d for 42 d), or ZA + TPTD until they were euthanized at 6 weeks following lumbar vertebral fusion. The lumbar spine was harvested. Bone mineral density (BMD), bone fusion, bone volume (BV), and bone formation rate (BFR)were analyzed by dual-energy X-ray absorptiometry (DXA), radiography, micro-computed tomography, and histomorphometry. Compared with vehicle (control) treatment, ZA and TPTD monotherapy increased bone volume (BV) at fusion site, and ZA + TPTD combined therapy had an additive effect. Treatment with TPTD and ZA + TPTD increased the bone fusion rate when compared with the control group. ZA monotherapy did not alter the rate of bone fusion. The TPTD and ZA + TPTD treatment groups had increased mineral apposition rate (MAR), mineralizing surfaces/bone surface ((MS/BS), and BFR/BS compared with the OVX group. Our experiment confirm that the monotherapy with TPTD and combination therapy with ZA + TPTD in an OVX rat model of osteopenia following lumbar vertebral fusion surgery increased bone fusion mass and bone fusion rate, and ZA + TPTD combined therapy had an additive effect on bone fusion mass. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res In this study, we investigate the effect of single and combined teriparatide (TPTD) and zoledronic acid (ZA) treatment on lumbar vertebral fusion in aged ovariectomized (OVX) rats. Our experiment confirm that the monotherapy with TPTD and combination therapy with ZA+TPTD in an OVX rat model of osteopenia following lumbar vertebral fusion surgery increased bone fusion mass and bone fusion rate, and ZA+TPTD combined therapy had an additive effect on bone fusion mass.

Multiple calcium patterns of rat osteoblasts under fluidic shear stress


The intracellular calcium ([Ca2+]i) response induced by external forces can be diverse and complex. Using primary osteoblasts from Wistar rats, we found multiple patterns of [Ca2+]i responses induced by fluidic shear stress (Fss), including homogeneous non-oscillation and heterogeneous oscillations. These multiple-patterned [Ca2+]i responses could be influenced by Fss intensity, cell density, and cell differentiation. Our real-time measurements with free calcium, ATP, ATP without calcium, suramin, apyrase, and thapsigargin confirmed homogeneous [Ca2+]i patterns and/or heterogeneous [Ca2+]i oscillations with respect to the combined degree of external Ca2+ influx, and intracellular Ca2+ release. Our theoretical model supported diverse Fss-induced calcium activities as well. We concluded that a singular factor of Ca2+ influx or release dominated to produce smooth homogeneous patterns, but combined factors produced oscillatory heterogeneous patterns. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

RNA sequencing identifies gene regulatory networks controlling extracellular matrix synthesis in intervertebral disk tissues


Degenerative disk disease of the spine is a major cause of back pain and disability. Optimization of regenerative medical therapies for degenerative disk disease requires a deep mechanistic understanding of the factors controlling the structural integrity of spinal tissues. In this investigation, we sought to identify candidate regulatory genes controlling extracellular matrix synthesis in spinal tissues. To achieve this goal we performed high throughput next generation RNA sequencing on 39 annulus fibrosus and 21 nucleus pulposus human tissue samples. Specimens were collected from patients undergoing surgical discectomy for the treatment of degenerative disk disease. Our studies identified associations between extracellular matrix genes, growth factors, and other important regulatory molecules. The fibrous matrix characteristic of annulus fibrosus was associated with expression of the growth factors platelet derived growth factor beta (PDGFB), vascular endothelial growth factor C (VEGFC), and fibroblast growth factor 9 (FGF9). Additionally we observed high expression of multiple signaling proteins involved in the NOTCH and WNT signaling cascades. Nucleus pulposus extracellular matrix related genes were associated with the expression of numerous diffusible growth factors largely associated with the transforming growth signaling cascade, including transforming factor alpha (TGFA), inhibin alpha (INHA), inhibin beta A (INHBA), bone morphogenetic proteins (BMP2, BMP6), and others. Clinical significance: this investigation provides important data on extracellular matrix gene regulatory networks in disk tissues. This information can be used to optimize pharmacologic, stem cell, and tissue engineering strategies for regeneration of the intervertebral disk and the treatment of back pain. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Relationship between glenoid deformity and gait characteristics in a rat model of neonatal brachial plexus injury


Neonatal brachial plexus injury (NBPI) results in substantial postural and functional impairments associated with underlying muscular and osseous deformities. We examined the relationship between glenoid deformity severity and gait in a rat model of NBPI, an established model for studying the in vivo pathomechanics of NBPI. At 8 weeks post-operatively, we monitored the gait of 24 rat pups who exhibited varying degrees of glenoid deformity following unilateral brachial plexus neurectomy and chemodenervation interventions administered 5 days postnatal. Five basic stride and stance metrics were calculated for the impaired forelimbs over four consecutive gait cycles. Bilateral differences in glenoid version (ΔGAv) and inclination (ΔGAi) angles were computed from data for the same rats as reported in a previous study. A linear regression model was generated for each deformity-gait pair to identify significant relationships between the two. ΔGAv was not significantly correlated with any gait measurements, while ΔGAi significantly correlated with all five gait measurements. Specifically, ΔGAi was significantly positively correlated with stride length (R2 = 0.38, p = 0.001) and stance factor (R2 = 0.45, p < 0.001), and significantly negatively correlated with stance width (R2 = 0.24, p = 0.016), swing/stance ratio (R2 = 0.17, p = 0.046), and stride frequency (R2 = 0.33, p = 0.003). Rats with declined glenoids exhibited the most altered gait. Clinical significance: Our findings link musculoskeletal changes and functional outcomes in an NBPI rat model. Thus, gait analysis is a potentially useful, non-invasive, quantitative way to investigate the effects of injury and deformity on limb function in the NBPI rat model. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

The onset of fretting at the head-stem connection in hip arthroplasty is affected by head material and trunnion design under simulated corrosion conditions


Mechanically assisted crevice corrosion (MACC) is a mechanism for trunnion damage in total hip arthroplasties (THAs). Retrieval studies have shown reduced MACC-related damage for ceramic heads compared with cobalt–chromium (CoCr) heads. We propose that ceramic heads demonstrate fretting at higher cyclic compressive loads than CoCr heads on titanium alloy trunnions in a simulated corrosion model. A closed electrochemical chamber was used to measure fretting current onset loads for two modern titanium alloy trunnions (Zimmer 12/14 and Stryker V-40) in which trunnion failure has been reported. Ceramic and CoCr alloy 36 + 0 mm heads were impacted on each trunnion and cyclically loaded at 3 Hz with increasing magnitude from 100 to 3,400 N for 540 cycles. Onset load was the cyclic compressive load at which the slope of the average fretting current increased significantly. A CoCr head with V40 trunnion demonstrated the lowest onset load (1,400 N), while the V40 trunnion with a ceramic head showed the highest onset load (2,200 N). Significant differences occurred in average fretting current between head materials for V40 trunnions (p < 0.001) at loads over 2,000 N. CoCr-12/14 and ceramic-12/14 couples demonstrated similar onset loads (2,000 N). All head–trunnion combinations showed cyclical fretting response to loading at 100 N. Head material composition was observed to increase fretting at the taper junction but the effect was taper geometry dependent. Using ceramic heads may reduce the phenomena of trunnion fretting and corrosion but the effect of both trunnion geometry and metallurgy warrants further investigation. Statement of clinical significance: Trunnion corrosion may occur with titanium alloy stems regardless of the head material used. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res A closed electrochemical chamber was used to measure fretting current onset loads for two titanium alloy trunnions in which trunnion failure has been reported. A CoCr head with V40 trunnion demonstrated lowest onset load (1,400 N), while the V40 trunnion with a ceramic head showed the highest onset load (2,200 N). Average fretting current between head materials for V40 trunnions were differenct at loads over 2,000 N (p < 0.001). CoCr-12/14 and ceramic-12/14 couples demonstrated similar onset loads (2,000 N).

Crosstalk between substance P and calcitonin gene-related peptide during heterotopic ossification in murine Achilles tendon


Heterotopic ossification (HO) is abnormal bone formation within soft tissue, usually predisposed by neurogenic or musculoskeletal trauma. Inflammation resulting from trauma is considered to be the main trigger for HO by eliciting changes within the injury site, including elevation of bone morphogenetic proteins (BMPs). Recent research, however, has also associated changes in sensory neuropeptide expression with HO. Substance P (SP) and calcitonin gene-related peptide (CGRP) are two of those neuropeptides that have been implicated with various aspects of HO, including regulation of inflammation and BMP signaling. Despite discoveries associating SP and CGRP with soft tissue HO, it remains unclear whether SP and CGRP have a direct role in the induction of HO. Here, we investigated the effect of SP and CGRP in vivo with the aid of inkjet-based biopatterning technology to controllably deliver these neuropeptides onto a murine Achilles tendon. While we did not observe any significant effect with CGRP, SP alone promoted HO in vivo with increased expression of BMP2. Remarkably, when SP and CGRP were delivered together, CGRP counteracted the effect of SP and essentially blocked SP-induced HO. This report contributes to the understanding of the complex problem of HO pathophysiology and warrants more study to better elucidate the interplay between SP and CGRP in the induction of HO. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Integrating soft and hard tissues via interface tissue engineering


The enthesis, or interface between bone and soft tissues such as ligament and tendon, is prone to injury and often does not heal, even post surgical intervention. Interface tissue engineering represents an integrative strategy for regenerating the native enthesis by functionally connecting soft and hard tissues and thereby improving clinical outcome. This review focuses on integrative and cell-instructive scaffold designs that target the healing of the two most commonly injured soft tissue-bone junctions: tendon-bone interface (e.g., rotator cuff) and ligament-bone interface (e.g., anterior cruciate ligament). The inherent connectivity between soft and hard tissues is instrumental for musculoskeletal motion and is therefore a key design criterion for soft tissue regeneration. To this end, scaffold design for soft tissue regeneration have progressed from single tissue systems to the emerging focus on pre-integrated and functional composite tissue units. Specifically, a multifaceted, bioinspired approach has been pursued wherein scaffolds are tailored to stimulate relevant cell responses using spatially patterned structural and chemical cues, growth factors, and/or mechanical stimulation. Moreover, current efforts to elucidate the essential scaffold design criteria via strategic biomimicry are emphasized as these will reduce complexity in composite tissue regeneration and ease the related burden for clinical translation. These innovative studies underscore the clinical relevance of engineering connective tissue integration and have broader impact in the formation of complex tissues and total joint regeneration. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Increased oxidative protection by high active vitamin E content and partial radiation crosslinking of UHMWPE


Vitamin E stabilization successfully improved long-term oxidation resistance of wear-resistant ultra-high-molecular-weight polyethylene (UHMWPE) used for joint implants. Stabilization can be achieved by blending an antioxidant into the UHMWPE resin powder before consolidation and irradiation. Balancing the wear resistance and vitamin E content in the blend is the current challenge with this approach, because vitamin E hinders crosslinking of UHMWPE during irradiation, which decreases wear resistance. The vitamin E concentration in the blend is generally limited to less than 0.3 wt%. Wear- and oxidation-resistant UHMWPE has been obtained previously by consolidating blends of pre-irradiated UHMWPE powders (XPE) into an unmodified polyethylene matrix (PE), where the improvement in wear rate depended on the radiation dose and fraction of XPE. We hypothesized that increasing the vitamin E content in the unirradiated matrix would not compromise wear and would further improve the oxidative stability of XPE/PE blends. Pin-on-disk wear testing showed that the XPE/PE blends containing 0.1–1.0 wt% vitamin E in the matrix had comparable wear rates. We used an aggressive accelerated aging test in the presence of the pro-oxidant squalene and oxidation induction time (OIT) test and found that higher amounts of vitamin E resulted in stronger oxidation resistance for XPE/PE blends. The mechanical strength and toughness of the blends were not affected by changing the vitamin E content in the matrix. Stabilizing UHMWPE with higher vitamin E content may extend the service life of UHMWPE implants. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Obesity/type 2 diabetes increases inflammation, periosteal reactive bone formation, and osteolysis during Staphylococcus aureus implant-associated bone infection


Obese and type 2 diabetic (T2D) patients have a fivefold increased rate of infection following placement of an indwelling orthopaedic device. Though implant infections are associated with inflammation, periosteal reactive bone formation, and osteolysis, the effect of obesity/T2D on these complicating factors has not been studied. To address this question, C57BL/6J mice were fed a high fat diet (60% Kcal from fat) to induce obesity/T2D, or a control diet (10% Kcal from fat) for 3 months, and challenged with a transtibial pin coated with a bioluminescent USA300 strain of S. aureus. In the resulting infected bone, obesity/T2D was associated with increased S. aureus proliferation and colony forming units. RNA sequencing of the infected tibiae on days 7 and 14 revealed an increase in 635 genes in obese/T2D mice relative to controls. Pathways associated with ossification, angiogenesis, and immunity were enriched. MicroCT and histology on days 21 and 35 demonstrated significant increased periosteal reactive bone formation in infected obese/T2D mice versus infected controls (p < 0.05). The enhanced periosteal bone formation was associated with increased osteoblastic activity and robust endochondral ossification, with persistant cartilage on day 21 that was only observed in infected obesity/T2D. Osteolysis and osteoclast numbers in obesity/T2D were also significantly increased versus infected controls (p < 0.05). Consistent with an up-regulated immune transcriptome, macrophages were more abundant within both the periosteum and the new reactive bone of obese/T2D mice. In conclusion, we find that implant-associated S. aureus osteomyelitis in obesity/T2D is associated with increased inflammation, reactive bone formation, and osteolysis. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res Obese and diabetic patients are more susceptible to infection following orthopaedic surgery. Despite this, if and how obesity and diabetes complicate bone healing following infection is unknown. Here, we report increased inflammation, periosteal reactive bone formation, osteolysis, and macrophage recruitment in obese/diabetic mice in a model of orthopaedic S. aureus infection. Overall, these effects were more deleterious to the bone environment in obese/diabetics and controlling inflammation may represent a therapeutic strategy in treating the complications of osteomyelitis in this population.

Magnetic resonance measurements of tissue quantity and quality using T2* relaxometry predict temporal changes in the biomechanical properties of the healing ACL


The purpose of this study was to develop a magnetic resonance T2* relaxometry-based multiple linear regression model to predict the structural properties of the healing anterior cruciate ligament (ACL) over a 24-week healing period following ACL repair in Yucatan minipigs. Two hypotheses were tested: (i) that a regression model based on ACL sub-volumes containing short and long T2* relaxation times would outperform a competing model based on sub-volumes of short T2* relaxation times only; and (ii) that an optimized regression model would be capable of predicting ACL structural properties between 6 and 24 weeks post-repair. ACLs were imaged in 24 minipigs (8/group) at either 6, 12, or 24 weeks after ACL repair. The structural properties of the ACLs were determined from tensile failure tests. Four multiple linear regression models of increasing complexity were fitted to the data. Akaike Information Criterion values and Bland–Altman tests were used to compare model performance and to test the hypotheses. The structural properties predicted from the multiple linear regression model that was based on the change in ACL sub-volumes of both the short and long T2* relaxation times over the healing period were in closest agreement to the measured values, suggesting that the amounts of both organized and disorganized collagen, and the change in these quantities over time, are required to predict the structural properties of healing ACLs accurately. Clinical significance: our time-specific, T2*-based regression model may allow us to estimate the structural properties of ACL repairs in vivo longitudinally. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Progenitor cells from different zones of human cartilage and their correlation with histopathological osteoarthritis progression


Cell-based therapies development for the treatment of osteoarthritis (OA) requires an understanding of the disease progression and attributes of the cells resident in cartilage. This study focused on quantitative assessment of the concentration and biological potential of stem and progenitor cells resident in different zones of cartilage displaying macroscopic Outerbridge grade 1–2 OA, and their correlation with OA progression based on established histologic scoring system. Lateral femoral condyles were collected from 15 patients with idiopathic OA and varus knees undergoing total knee arthroplasty. Superficial(Csp, top ∼ 500 µm) and deep cartilage(Cdp) was separated. Chondrogenic Connective Tissue Progenitors (CTP-C) were assayed by standardized Colony-Forming-Unit assay using automated image analysis (ColonyzeTM) based on ASTM standard F-2944-12. Cell concentration (cells/mg) was significantly greater in Csp (median: 7,000; range: 3,440–17,600) than Cdp (median: 5,340; range: 3,393–9,660), p = 0.039. Prevalence (CTPs/million cells) was not different between Csp (median: 1,274; range: 0–3,898) and Cdp (median:1,365; range:0–6,330), p = 0.42. In vitro performance of CTP-C progeny varied widely within and between patients, manifest by variation in colony size and morphology. Mean histopathological Mankin score was 4.7 (SD = 1.2), representing mild to moderate OA. Tidemark breach by blood vessels was associated with lower Csp cell concentration (p = 0.02). Matrix degradation was associated with lower Cdp cell and CTP-C concentration (p = 0.015 and p = 0.095, respectively), independent of articular surface changes. These findings suggest that the initiation of OA may occur in either superficial or deep zones. The pathological changes affect CTP-Cs in Csp and Cdp cartilage zones differently. The heterogeneity among the available CTP-Cs in Csp and Cdp suggests performance-based selection to optimize cell-sourcing strategies for therapy. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Role of dexamethasone in the long-term functional maturation of MSC-laden hyaluronic acid hydrogels for cartilage tissue engineering


The purpose of study was to investigate the maturation of mesenchymal stem cells (MSC) laden in HA constructs with various combinations of chemically defined medium (CM) components and determine the impact of dexamethasone and serum on construct properties. Constructs were cultured in CM with the addition or withdrawal of media components or were transferred to serum containing media that partially represents an in vivo-like condition where pro-inflammatory signals are present. Constructs cultured in CM+ (CM with TGF-β3) and DEX− (CM+ without dexamethasone) conditions produced robust matrix, while those in ITS/BSA/LA− (CM+ without ITS/BSA/LA) and Serum+ (10% FBS with TGF-β3) produced little matrix. While construct properties in DEX− were greater than those in CM+ at 4 weeks, properties in CM+ and DEX− reversed by 8 weeks. While construct properties in DEX− were greater than those in CM+ at 4 weeks, the continued absence or removal of dexamethasone resulted in marked GAG loss by 8 weeks. Conversely, the continued presence or new addition of dexamethasone at 4 weeks further improved or maintained construct properties through 8 weeks. Finally, when constructs were converted to Serum (in the continued presence of TGF-β3 with or without dexamethasone) after pre-culture in CM+ for 4 weeks, GAG loss was attenuated with addition of dexamethasone. Interestingly, however, collagen content and type was not impacted. In conclusion, dexamethasone influences the functional maturation of MSC-laden HA constructs, and may help to maintain properties during long-term culture or with in vivo translation by repressing pro-inflammatory signals. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res MSCs were viable only in CM+ and DEX− and maintained their initial shape with robust matrix production. Conversely, cells in ITS/BSA/LA− and Serum+ did not remain viable, and these constructs contracted significantly over the culture period. Constructs cultured in Serum+ contracted very quickly, MSCs appeared to aggregate centrally, and this culminated in the formation of a central pellet-like structure that separated from the HA hydrogel.

Hydrogen peroxide induces programmed necrosis in rat nucleus pulposus cells through the RIP1/RIP3-PARP-AIF pathway


This study aimed to systematically investigate whether programmed necrosis contributes to H2O2-induced nucleus pulposus (NP) cells death and to further explore the underlying mechanism involved. Rat NP cells were subjected to different concentrations of H2O2 for various time periods. The cell viability was measured using a cell counting kit-8, and the death rate was detected by Hoechst 33258/propidium iodide (PI) staining. The programmed necrosis-related molecules receptor-interacting protein 1 (RIP1), receptor-interacting protein 3 (RIP3), poly (ADP-ribose) polymerase (PARP), and apoptosis inducing factor (AIF) were determined by real-time polymerase chain reaction and Western blotting, respectively. The morphologic and ultrastructural changes were examined by phasecontrast microscopy and transmission electron microscopy (TEM). In addition, the necroptosis inhibitor Necrostatin-1 (Nec-1), the PARP inhibitor diphenyl-benzoquinone (DPQ) and small interfering RNA (siRNA) technology were used to indirectly evaluate programmed necrosis. Our results indicated that H2O2 induced necrotic morphologic and ultrastructural changes and an elevated PI positive rate in NP cells; these effects were mediated by the upregulation of RIP1 and RIP3, hyperactivation of PARP, and translocation of AIF from mitochondria to nucleus. Additionally, NP cells necrosis was significantly attenuated by Nec-1, DPQ pretreatment and knockdown of RIP3 and AIF, while knockdown of RIP1 produced the opposite effects. In conclusion, these results suggested that under oxidative stress, RIP1/RIP3-mediated programmed necrosis, executed through the PARP-AIF pathway, played an important role in NP cell death. Protective strategies aiming to regulate programmed necrosis may exert a beneficial effect for NP cells survival, and ultimately retard intervertebral disc (IVD) degeneration. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Human mesenchymal stem cells induced to differentiate as chondrocytes follow a biphasic pattern of extracellular matrix production


Regenerative medicine and tissue engineering studies are actively developing novel means to repair adult articular cartilage defects using biological approaches. One such approach is the harnessing of adult human therapeutic cells such as those referred to as mesenchymal stem cells. Upon exposure to chondrogenic signals, these cells differentiate and initiate the production of a complex and voluminous cartilaginous matrix that is crucial to both the structure and function of cartilage. Furthermore, this complexity requires the time-sensitive activation of a large number of genes to produce the components of this matrix. The current study analyzed the kinetics of matrix production in an aggregate culture model where adult human mesenchymal stem cells were induced to differentiate as chondrocytes. The results indicate the existence of a biphasic mode of differentiation and maturation during which matrix genes and molecules are differentially activated and secreted. These results have important implications for developing novel approaches for the creation of tissue engineered articular cartilage. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res Human mesenchymal stem cells in aggregate culture form chondrogenic cells in a two-stage differentiation process. Pre-chondrogenic cells develop prior to day 3 in culture and mature chondrocytes develop between day 10 and day 14 in culture. Chondrogenic differentiation is demonstrated by a significant upregulation in mRNAs for multiple matrix molecules and this indicates a window for modulating chondrogenesis. The fold change data represent averages of two donors. See Supplemental Data section for more details.

CDK4 expression in chordoma: A potential therapeutic target


Chordomas are rare bone tumors and treatment is commonly based on a combination of surgery and radiotherapy. There is no standard chemotherapy treatment for chordoma. The aim of this study was to determine the expression of cyclin-dependent kinase 4 (CDK4) in chordoma and its therapeutic implications. We evaluated CDK4 expression both in chordoma cell lines and in chordoma tissues. Also, we investigated the functional roles of CDK4 in chordoma cell growth and proliferation. Furthermore, the therapeutic implications of targeting CDK4 in chordoma were evaluated. We found CDK4 highly expressed in chordoma cell lines and in a majority (97.7%) of chordoma tissues. Higher CDK4 expression correlated with metastasis and recurrence of chordoma. Treatment of chordoma cells using CDK4 inhibitor palbociclib could efficiently inhibit chordoma cells growth and proliferation. These data demonstrate that targeting CDK4 may be useful as a novel strategy in the treatment of chordoma. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Biomechanical tolerance of whole lumbar spines in straightened posture subjected to axial acceleration


Quantification of biomechanical tolerance is necessary for injury prediction and protection of vehicular occupants. This study experimentally quantified lumbar spine axial tolerance during accelerative environments simulating a variety of military and civilian scenarios. Intact human lumbar spines (T12-L5) were dynamically loaded using a custom-built drop tower. Twenty-three specimens were tested at sub-failure and failure levels consisting of peak axial forces between 2.6 and 7.9 kN and corresponding peak accelerations between 7 and 57 g. Military aircraft ejection and helicopter crashes fall within these high axial acceleration ranges. Testing was stopped following injury detection. Both peak force and acceleration were significant (p < 0.0001) injury predictors. Injury probability curves using parametric survival analysis were created for peak acceleration and peak force. Fifty-percent probability of injury (95%CI) for force and acceleration were 4.5 (3.9–5.2 kN), and 16 (13–19 g). A majority of injuries affected the L1 spinal level. Peak axial forces and accelerations were greater for specimens that sustained multiple injuries or injuries at L2–L5 spinal levels. In general, force-based tolerance was consistent with previous shorter-segment lumbar spine testing (3–5 vertebrae), although studies incorporating isolated vertebral bodies reported higher tolerance attributable to a different injury mechanism involving structural failure of the cortical shell. This study identified novel outcomes with regard to injury patterns, wherein more violent exposures produced more injuries in the caudal lumbar spine. This caudal migration was likely attributable to increased injury tolerance at lower lumbar spinal levels and a faster inertial mass recruitment process for high rate load application. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Standardization of hemipelvis alignment for in vitro biomechanical testing


Although in vitro biomechanical tests are regularly performed, the definition of a suitable reference frame for hemipelvic specimens is still a challenge. The aims of the present study were to: (i) define a reference frame for the human hemipelvis suitable for in vitro applications, based on robust anatomical landmarks; (ii) identify the alignment of a hemipelvis based on the alignment of a whole pelvis (including right/left and male/female differences); (iii) identify the relative alignment of the proposed in vitro reference frame with respect to a reference frame commonly used in gait analysis; (iv) create an in vitro alignment procedure easy, robust and inexpensive; (v) quantify the intra-operator repeatability and inter-operator reproducibility of the procedure. A procedure to univocally identify the anatomical landmarks was created, exploiting the in vitro accessibility of the specimen's surface. Through the analysis on 53 CT scans (106 hemipelvises), the alignment of the hemipelvis based on the alignment of a whole pelvis was analyzed: differences between male/female and right/left hemipelvises were not statistically significant To overcome the uncertainty in the identification of the acetabular rim, a standard acetabular plane was defined. An alignment procedure was developed to implement such anatomical reference frame. The intra-operator repeatability and the inter-operator reproducibility were quantified with four operators, on male and female hemipelvises. The intra-operator repeatability was better than 1.5°. The inter-operator reproducibility was better than 2.0°. Alignment in the transverse plane was the most repeatable. The presented procedure to align hemipelvic specimens is sufficiently robust, standardized, and accessible. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res In order to identify the alignment of the human hemipelvis based on the anatomical alignment of a whole pelvis, 53 CT scans (106 hemipelvises) were analyzed: differences among males and females were not statistically significant. Results were combined with the alignment of a standard acetabular plane to implement an alignment procedure for in vitro applications. The intra-operator repeatability was better than 1.5°; the inter-operator reproducibility better than 2.0°. The proposed alignment will help performing reproducible biome[...]

Preclinical models for orthopedic research and bone tissue engineering


In this review, we broadly define and discuss the preclinical rodent models that are used for orthopedics and bone tissue engineering. These range from implantation models typically used for biocompatibility testing and high-throughput drug screening, through to fracture and critical defect models used to model bone healing and severe orthopedic injuries. As well as highlighting the key methods papers describing these techniques, we provide additional commentary based on our substantive practical experience with animal surgery and in vivo experimental design. This review also briefly touches upon the descriptive and functional outcome measures and power calculations that are necessary for an informative study. Obtaining informative and relevant research outcomes can be very dependent on the model used, and we hope this evaluation of common models will serve as a primer for new researchers looking to undertake preclinical bone studies. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Interleukin-17 and Toll-like Receptor 10 genetic polymorphisms and susceptibility to large joint osteoarthritis


Primary osteoarthritis (OA) is the most common type of a joint disease. It has a polygenic risk inheritance pattern and affects older people. The etiology of this disease is not fully understood. The aim of this study was to investigate the associations between polymorphisms in pro-inflammatory interleukin-17 (IL17A and IL17F) and anti-inflammatory Toll-like Receptor 10 (TLR10) genes with the risk for development of advanced stage hip and knee primary OA in the Croatian population. A total of 500 OA patients and 597 controls were genotyped for IL17A SNP (rs2275913), IL17F SNPs (rs763780 and rs1889570), and TLR10 (rs11096957) genes. The allelic and genotypic frequencies of IL17F SNP (rs763780) showed statistically significant differences in comparisons of controls with hip—but not knee—OA patients. The major allele (T) of rs763780 was associated with the lower risk for developing hip OA (p = 7.9 × 10−4, OR = 0.45, 95%CI = 0.27–0.74), whereas the minor allele (C) was associated with susceptibility to hip OA (p = 7.9 × 10−4, OR = 2.24, 95%CI = 1.35–3.72). The genotype T/T was associated with the protection to hip OA (p = 3.9 × 10−4, OR = 0.41, 95%CI = 0.24–0.70), and, lastly, the genotype T/C was associated with the higher risk to acquiring hip OA (p = 2.6 × 10−4, OR = 2.50, 95%CI = 1.47–4.25). TLR10 SNP rs11096957 was found significantly associated with predisposition to hip OA (p = 0.04, OR = 1.41, 95%CI = 1.02–1.94) but not knee OA. Our findings suggest that hip OA in Croatian population might have a different genetic risk regarding the IL17 and TLR10 gene locus than knee OA. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 9999:1–10, 2017.

MicroRNAs in orthopaedic research: Disease associations, potential therapeutic applications, and perspectives


MicroRNAs (miRNAs) are small non-coding RNAs that function to control many cellular processes by their ability to suppress expression of specific target genes. Tens to hundreds of target genes may be affected by one miRNA, thereby resulting in modulation of multiple pathways in any given cell type. Therefore, altered expression of miRNAs (i.e., during tissue development or in scenarios of disease or cellular stress) can have a profound impact on processes regulating cell differentiation, metabolism, proliferation, or apoptosis, for example. Over the past 5–10 years, thousands of reports have been published on miRNAs in cartilage and bone biology or disease, thus highlighting the significance of these non-coding RNAs in regulating skeletal development and homeostasis. For the purpose of this review, we will focus on miRNAs or miRNA families that have demonstrated function in vivo within the context of cartilage, bone or other orthopaedic-related tissues (excluding muscle). Specifically, we will discuss studies that have utilized miRNA transgenic mouse models or in vivo approaches to target a miRNA with the aim of altering conditions such as osteoarthritis, osteoporosis and bone fractures in rodents. We will not discuss miRNAs in the context skeletal cancers since this topic is worthy of a review of its own. Overall, we aim to provide a comprehensive description of where the field currently stands with respect to the therapeutic potential of specific miRNAs to treat orthopaedic conditions and current technologies to target and modify miRNA function in vivo. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Transcutaneous carbon dioxide application with hydrogel prevents muscle atrophy in a rat sciatic nerve crush model


The acceleration of nerve regeneration remains a clinical challenge. We previously demonstrated that transcutaneous CO2 application using a novel hydrogel increases the oxygen concentration in local tissue via an “artificial Bohr effect” with the potential to prevent muscle atrophy. In this study, we investigated the effect of transcutaneous CO2 administration on limb function after peripheral nerve injury in a rat sciatic nerve injury model. In total, 73 Sprague–Dawley rats were divided into a sham group, a control group (crush injury to sciatic nerve and no treatment) or a CO2 group (crush injury with transcutaneous CO2 application). CO2 was administered percutaneously for 20 min five times per week. Scores for the sciatic function index and pinprick test were significantly higher in the CO2 group than control group. The muscle wet weight ratios of the tibialis anterior and soleus muscles were higher in the CO2 group than control group. Electrophysiological examination showed that the CO2 group had higher compound motor action potential amplitudes and shorter distal motor latency than the control group. Histological examination of the soleus muscle sections at postoperative week 2 showed shorter fiber diameter in the control group than in the CO2 group. The mRNA expression of Atrogin-1 and MuRF-1 was lower, mRNA expression of VEGF and myogenin and MyoD was higher in CO2 group at postoperative week 2 compared to the control group. Clinical significance: Transcutaneous CO2 application has the therapeutic potential to accelerate the recovery of muscle atrophy in peripheral nerve injury. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res The acceleration of nerve regeneration remains a clinical challenge. In this study, we investigated the effect of transcutaneous CO2 administration on limb function after peripheral nerve injury in a rat sciatic nerve injury model. Rats were divided into a sham group, a control group (crush injury to sciatic nerve and no treatment), or a CO2 group (crush injury with transcutaneous CO2 application). Scores for the sciatic function index and pinprick test were significantly higher in the CO2 group than control group. The muscle wet weight ratios were higher in the CO2 group. [...]

Frontal plane hip joint loading according to pain severity in people with hip osteoarthritis


The primary objective was to examine the hip adduction moment during walking in people with hip osteoarthritis (OA) according to pain severity. Sixty-eight participants with unilateral symptomatic hip OA were included. Pain during walking was assessed on a 5-point Likert item within the Western Ontario and McMaster Universities Index (no pain = 12; mild pain n = 37; moderate pain n = 19). Measures of the external hip adduction moment (peaks, Nm/BW × BH (%) and impulse, Nm.s/BW × BH (%)) were determined. Other measures included frontal plane hip, pelvis and trunk kinematics, walking speed and peak isometric hip abductor strength. Variables were compared according to pain severity using linear models and biomechanical variables were examined. Participants with moderate pain had a significantly higher second peak hip adduction moment and impulse compared to those with less pain. There was no difference in any measure of hip adduction moment between those with mild pain and no pain. There were no differences in kinematics across pain severity categories. Participants with moderate pain had a significantly slower walking speed compared to participants with mild and no pain. Participants with moderate pain had weaker peak isometric hip abductor strength compared to those with mild pain and no pain. The hip adduction moment during walking, hip abduction strength and walking speed differs according to pain severity during walking in people with hip OA. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Mechanical properties of Triclosan sutures


To avoid infections and wound healing disorders, Triclosan coated sutures have been invented. Little is known of these sutures regarding their tensile properties. Three different Triclosan coated sutures (Vicryl 1 plus, PDS 0 plus, Monocryl 3-0 plus) were tested at several time points over 42 days regarding load to failure, strain, and stiffness compared to their non-coated versions (Vicryl 1, PDS 0, Monocryl 3-0). Four different measurement points were made. Suture loops were fixed in a material testing machine over two metal bars which were moved apart creating a stress to the fiber. Unpaired, two-tailed t-test were performed for each group (untreated and treated) while level of significance was defined at a level of p < 0.05. Vicryl 1 was significantly stronger on day 14 than Vicryl 1 plus (p = 0.033). On day 28, significant changes were found in PDS 0 which was weaker compared to PDS 0 plus (p = 0.039) and Vicryl 1 which was stronger than Vicryl 1 plus (p = 0.032). We have seen that Vicryl 1 plus sutures are significantly weaker according to loading to failure after 14 and 28 days, which might cause incisional hernias. PDS 0 sutures are used to reconstruct tendons, therefore a longer durableness might be of interest as re-ruptures of tendons are problematic. Our in vitro findings support, the use of Triclosan coated PDS plus sutures and Vicryl sutures as they show a longer resistance. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res To avoid wound healing disorders Triclosan coated sutures were invented. Three different Triclosan coated sutures were tested over 42 days regarding load to failure, strain, and stiffness. Significant changes in load to failure could be found on days 14 and 28 in Vicryl 1 versus Vicryl 1 plus and on day 28 in PDS versus PDS 0 plus. Our in vitro findings support, the use of Triclosan coated PDS plus sutures and Vicryl sutures as they show a longer resistance.

New algorithm for simulation of proteoglycan loss and collagen degeneration in the knee joint: Data from the osteoarthritis initiative


Osteoarthritis is a harmful joint disease but prediction of disease progression is problematic. Currently, there is only one modeling framework which can be applied to predict the progression of knee osteoarthritis but it only considers degenerative changes in the collagen fibril network. Here, we have developed the framework further by considering all of the major tissue changes (proteoglycan content, fluid flow, and collagen fibril network) occurring in osteoarthritis. While excessive levels of tissue stresses controlled degeneration of the collagen fibril network, excessive levels of tissue strains controlled the decrease in proteoglycan content and the increase in permeability. We created four knee joint models with increasing degrees of complexity based on the depth-wise composition. Models were tested for normal and abnormal, physiologically relevant, loading conditions in the knee. Finally, the predicted depth-wise compositional changes from each model were compared against experimentally observed compositional changes in vitro. The model incorporating the typical depth-wise composition of cartilage produced the best match with experimental observations. Consistent with earlier in vitro experiments, this model simulated the greatest proteoglycan depletion in the superficial and middle zones, while the collagen fibril degeneration was located mostly in the superficial zone. The presented algorithm can be used for predicting simultaneous collagen degeneration and proteoglycan loss during the development of knee osteoarthritis. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 9999:1–11, 2017. Four knee joint models with increasing degrees of complexity based on the depth-wise composition. Models were tested for normal and abnormal, physiologically relevant, loading conditions in the knee. The predicted depth-wise compositional changes from each model were compared against experimentally observed compositional changes in vitro. The model incorporating typical depth-wise composition of cartilage produced the best match with experimental observations. Result revealed that mechanical overloading caused by tissue stresses and str[...]

Post-Traumatic Heterotopic Ossification: An Old Problem in Need of New Solutions


Heterotopic ossification (HO) is the formation of pathological bone in ectopic sites and it can have serious consequences for functional outcomes. For many years, its main clinical relevance was as a rare complication of elective joint arthroplasty or CNS injury and a number of prophylaxes were developed to mitigate against it in these settings. As a consequence of changes in patterns of wounding and survival in conflicts since the turn of the century, post-traumatic HO has become much more common and case severity has increased. It represents one of the main barriers to rehabilitation in a large cohort of combat-injured patients. However, extant prophylaxes have not been shown to be effective or appropriate in this patient cohort. In addition, the lack of reliable early detection or means of predicting which patients will develop HO is another barrier to effective prevention. This review examines the current state of understanding of post-traumatic HO including the historical context, epidemiology, pathophysiology, clinical issues, currently prophylaxis and detection, management, and potential future approaches. Our aims are to highlight the current lack of effective means of early detection and prevention of HO after major trauma and to stimulate research into novel solutions to this challenging problem. © 2017 The Authors. Journal of Orthopaedic Research Published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. J Orthop Res

Mechanosensitiviy of aged muscle stem cells


During aging, skeletal muscle tissue progressively declines in mass, strength, and regenerative capacity. Decreased muscle stem cell (MuSC) number and impaired function might underlie the aging-related muscle wasting and impaired regenerative capacity. As yet, the search for factors that regulate MuSC fate and function has revealed several biochemical factors within the MuSC niche that may be responsible for the decline in MuSC regenerative capacity. This decline cannot be explained by environmental factors solely, as the MuSC potential to regenerate muscle tissue is not reversed by changing the biochemical MuSC niche composition. Here we discuss the likeliness that during physical exercise, MuSCs within their niche are subjected to mechanical loads, in particular pressure and shear stress, as well as associated deformations. We postulate that these physical cues are involved in the activation and differentiation of MuSCs as these cells contain several transmembrane sensor proteins that have been shown to be mechanosensitive in other cell types, that is, endothelial cells and osteoprogenitors. We will specifically address age-related changes in mechanosensing in MuSCs and their niche. Insight in the physical cues applied to the MuSCs in vivo, and how these cues affect MuSC fate and function, helps to develop new therapeutic interventions to counterbalance age-related muscle loss. This requires an approach combining two- and three-dimensional live cell imaging of MuSCs within contracting muscle tissue, mathematical finite element modeling, and cell biology. © 2017 The Authors. Journal of Orthopaedic Research Published by Wiley Periodicals, Inc. on behalf of the Orthopaedic Research Society. J Orthop Res In this perspective we argue that, in addition to biochemical cues, muscle stem cells (MuSCs) within their niche during physical exercise are subjected to mechanical cues. These physical loads include shear stress, pressure and tensile stress. Here we show the plausibility that mechanotransduction via mechanosensitive protein complexes is essential in determining MuSC fate and function which is impaired in aged MuSCs[...]

Distribution of segmental foot kinematics in patients with degenerative joint disease of the ankle


Degenerative joint disease (DJD) of the ankle is a debilitating chronic disease associated with severe pain and dysfunction resulting in antalgic gait alteration. Little information is available about segmental foot and ankle motion distribution during gait in ankle osteoarthritis. The aim of the current study was to dynamically characterize segmental foot and ankle kinematics of patients with severe ankle arthrosis requiring total ankle replacement. This was a prospective study involving 36 (19 M, 17 F) adult patients with a clinical diagnosis of ankle arthrosis (“DJD” group) and 36 (23 M, 13 F) healthy subjects (“Control” group). Motion data were collected at 120 Hz using a 3-D motion camera system at self-selected speed along a 6-m walkway and processed using the Milwaukee Foot Model (MFM). The SF-36 Health Survey and Orthopaedic Foot and Ankle Society (AOFAS) ankle-hindfoot scale were administered to evaluate functional levels. Findings include decreases in walking speed, cadence, stride length and swing phase, and reduced outcomes scores (SF-36 and AOFAS). Multisegemental motion in patients with ankle DJD demonstrates significant changes in foot mechanics characterized by altered segment kinematics and significant reduction in dynamic ROM at the tibia, hindfoot, forefoot, and hallux when compared to controls. The results demonstrate decreased temporal-spatial parameters and low outcomes scores indicative of functional limitations. Statement of clinical significance: Altered segment kinematics and reduced overall range of motion demonstrate how a single joint pathology affects kinematic distribution in the other segments of the foot and ankle and alters patients’ overall gait. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Quantifying meniscal kinematics in dogs


The dog has been used extensively as an experimental model to study meniscal treatments such as meniscectomy, meniscal repair, transplantation, and regeneration. However, there is very little information on meniscal kinematics in the dog. This study used MR imaging to quantify in vitro meniscal kinematics in loaded dog knees in four distinct poses: extension, flexion, internal, and external rotation. A new method was used to track the meniscal poses along the convex and posteriorly tilted tibial plateau. Meniscal displacements were large, displacing 13.5 and 13.7 mm posteriorly on average for the lateral and medial menisci during flexion (p = 0.90). The medial anterior horn and lateral posterior horns were the most mobile structures, showing average translations of 15.9 and 15.1 mm, respectively. Canine menisci are highly mobile and exhibit movements that correlate closely with the relative tibiofemoral positions. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Early mechanical stimulation only permits timely bone healing in sheep


Bone fracture healing is sensitive to the fixation stability. However, it is unclear which phases of healing are mechano-sensitive and if mechanical stimulation is required throughout repair. In this study, a novel bone defect model, which isolates an experimental fracture from functional loading, was applied in sheep to investigate if stimulation limited to the early proliferative phase is sufficient for bone healing. An active fixator controlled motion in the fracture. Animals of the control group were unstimulated. In the physiological-like group, 1 mm axial compressive movements were applied between day 5 and 21, thereafter the movements were decreased in weekly increments and stopped after 6 weeks. In the early stimulatory group, the movements were stopped after 3 weeks. The experimental fractures were evaluated with mechanical and micro-computed tomography methods after 9 weeks healing. The callus strength of the stimulated fractures (physiological-like and early stimulatory) was greater than the unstimulated control group. The control group was characterized by minimal external callus formation and a lack of bone bridging at 9 weeks. In contrast, the stimulated groups exhibited advanced healing with solid bone formation across the defect. This was confirmed quantitatively by a lower bone volume in the control group compared to the stimulated groups.The novel experimental model permits the application of a well-defined load history to an experimental bone fracture. The poor healing observed in the control group is consistent with under-stimulation. This study has shown early mechanical stimulation only is sufficient for a timely healing outcome. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res A novel bone defect model, which isolates an experimental fracture from functional loading, was applied in sheep to investigate if stimulation limited to the early proliferative phase is sufficient for bone healing. Unstimulated controls were characterized by minimal amounts of callus formation and f[...]

Human prosthetic joint infections are associated with myeloid-derived suppressor cells (MDSCs): Implications for infection persistence


Prosthetic joint infection (PJI) is a devastating complication of joint arthroplasty surgery typified by biofilm formation. Currently, mechanisms whereby biofilms persist and evade immune-mediated clearance in immune competent patients remain largely ill-defined. Therefore, the current study characterized leukocyte infiltrates and inflammatory mediator expression in tissues from patients with PJI compared to aseptic loosening. CD33+HLA-DR−CD66b+CD14−/low granulocytic myeloid-derived suppressor cells (G-MDSCs) were the predominant leukocyte population at sites of human PJI compared to aseptic tissues. MDSCs inhibit T cell proliferation, which coincided with reduced T cells in PJIs compared to aseptic tissues. IL-10, IL-6, and CXCL1 were significantly elevated in PJI tissues and have been implicated in MDSC inhibitory activity, expansion, and recruitment, respectively, which may account for their preferential increase in PJIs. This bias towards G-MDSC accumulation during human PJI could account for the chronicity of these infections by preventing the pro-inflammatory, antimicrobial actions of immune effector cells. Clinical significance: Animal models of PJI have revealed a critical role for MDSCs and IL-10 in promoting infection persistence; however, whether this population is prevalent during human PJI and across distinct bacterial pathogens remains unknown. This study has identified that granulocytic-MDSC infiltrates are unique to human PJIs caused by distinct bacteria, which are not associated with aseptic loosening of prosthetic joints. Better defining the immune status of human PJIs could lead to novel immune-mediated approaches to facilitate PJI clearance in combination with conventional antibiotics. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Effects of lateral-offset sole shoes on knee adduction moment in women with medial compartment knee osteoarthritis


This study aimed to determine the impact of wearing a lateral-offset sole shoe (LOSS) on knee adduction moment (KAM) in patients with medial knee osteoarthritis (OA). From December 2012 to November 2016, patients with medial knee OA were recruited. Ninety-three knees (50 left, 43 right) of 93 female patients were analyzed. The first peak KAMs were measured with patients (i) walking barefoot; (ii) walking in conventional shoes; and (iii) walking in LOSSs. The patients had grade 1 (n = 19), grade 2 (n = 49), grade 3 (n = 20), and grade 4 (n = 5) knee OA. First peak KAMs differed significantly in all three conditions (p = 0.031). In the post hoc analysis, first peak KAMs were significantly lower during LOSS walking than during conventional shoe walking (p = 0.001), but there were no differences in peak KAMs between barefoot and LOSS walking (p = 0.784). In the subgroup analysis, patients with grades 2 and 3 OA showed significantly lower first peak KAMs during LOSS walking than during conventional shoe walking (p = 0.029 and p = 0.011, respectively). Both the peak eversion ankle angle and moment of barefoot walking showed a significant increase compared with LOSS and conventional shoe walking, while there was no significant difference between LOSS and conventional shoe walking (p = 0.612 and p = 0.197, respectively). Our results suggest that LOSS wearing caused significant KAM reductions compared with conventional shoe wearing. Since LOSS wearing does not cause changes in the peak eversion ankle angle and moment during the load response, it may be an effective method to reduce the KAM in women with knee OA. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Gait patterns, symptoms, and function in patients with isolated tibiofemoral osteoarthritis and combined tibiofemoral and patellofemoral osteoarthritis


The purpose of this study was to compare hip and knee biomechanics during walking in individuals with isolated tibiofemoral osteoarthritis (TFOA), combined TFOA and patellofemoral osteoarthritis (PFOA), and those without knee osteoarthritis (OA), and to compare patient-reported symptoms and function in individuals with isolated TFOA and those with combined TFOA and PFOA. Participants with and without knee OA were assessed and categorized into (i) no OA, (ii) isolated TFOA, and (iii) combined TFOA and PFOA, based on Kellgren and Lawrence diagnostic criteria. Quantitative motion analyses were conducted during walking, and hip and knee kinematics, and external moments were calculated. Peak values in the sagittal and frontal planes during stance phase were computed. Patient-reported symptoms and function data were obtained using the Western Ontario McMaster Universities Arthritis Index. Multivariate analyses of variance were conducted to compare between-group differences in gait and patient-reported symptoms and function data. The results showed no statistically significant differences in hip and knee kinematics and external moments between the three groups. Relative to those with isolated TFOA, individuals with combined TFOA and PFOA had greater pain (mean difference [95%CI]: 1.5 [0.05–3.1]), stiffness (0.8 [0.02–1.5]), and poorer function (5.4 [0.2–10.7]). In conclusion, the combined TFOA and PFOA radiographic disease pattern is associated with worse pain and function compared to the isolated TFOA disease pattern. The results of the present study provide no indications that treatments designed to change walking biomechanics should differ between individuals with isolated TFOA and those with combined TFOA and PFOA. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Low-intensity vibration increases cartilage thickness in obese mice


Obesity is associated with an elevated risk of osteoarthritis (OA). We examined here whether high fat diet administered in young mice, compromised the attainment of articular cartilage thickness. Further, we sought to determine if low-intensity vibration (LIV) could protect the retention of articular cartilage in a mouse model of diet-induced obesity. Five-week-old, male, C57BL/6 mice were separated into three groups (n = 10): Regular diet (RD), High fat diet (HF), and HF + LIV (HFv; 90 Hz, 0.2g, 30 min/d, 5 d/w) administered for 6 weeks. Additionally, an extended HF diet study was run for 6 months (LIV at 15 m/d). Articular cartilage and subchondral bone morphology, and sulfated GAG content were quantified using contrast agent enhanced μCT and histology. Gene expression within femoral condyles was quantified using real-time polymerase chain reaction. Contrary to our hypothesis, HF cartilage thickness was not statistically different from RD. However, LIV increased cartilage thickness compared to HF, and the elevated thickness was maintained when diet and LIV were extended into adulthood. RT-PCR analysis showed a reduction of aggrecan expression with high fat diet, while application of LIV reduced the expression of degradative MMP-13. Further, long-term HF diet resulted in subchondral bone thickening, compared to RD, providing early evidence of OA pathology—LIV suppressed the thickening, such that levels were not significantly different from RD. These data suggest that dynamic loading, via LIV, protected the retention of cartilage thickness, potentially resulting in joint surfaces better suited to endure the risks of elevated loading that parallel obesity. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res Obesity is associated with increased osteoarthritis risk. We examined whether high fat diet compromised the attainment of articular cartilage thickness, and if low-intensi[...]

Biomechanical investigation of four different fixation techniques in sacrum Denis type II fracture with low bone mineral density


With increasing life expectancy, fragility fractures of the pelvic ring are seen more frequently. Although their osteosynthesis can be very challenging, specific biomechanical studies for investigation of the fixation stability are still lacking. The aim of this study was to biomechanically evaluate four different fixation methods for sacrum Denis type II fractures in osteoporotic bone. Unstable Denis type II vertical sacrum fractures were created in 16 human pelves. Their osteosynthesis was performed with one sacro-iliac screw, posterior sacral plating, triangular fixation, or spino-pelvic fixation. For that purpose, each pelvis was randomly assigned to two paired groups for treatment with either SI-screw/posterior sacral plating or triangular fixation/spino-pelvic fixation. Each hemi-pelvis was cyclically tested under progressively increasing axial compression. Relative interfragmentary movements were investigated via optical motion tracking analysis. Axial stiffness of triangular fixation was significantly higher versus posterior sacral plating and spino-pelvic fixation (p ≤ 0.022), but not significantly different in comparison to SI-screw fixation (p = 0.337). Cycles to 2, 3, 5, and 8 mm fracture displacement, as well as to 3°, 5°, and 8° gap angle at the fracture site were significantly higher for triangular fixation compared to all other groups (p ≤ 0.041). Main failure mode for all osteosynthesis techniques was screw cutting through the bone, leading loss of fixation stability. From a biomechanical point of view, triangular fixation in sacrum Denis type II fractures demonstrated less interfragmentary movements and should be considered in unstable fragility fractures of the sacrum. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res Fixation of sacrum fragility fractures can be very challenging. The aim of this study was to biomechanically[...]

Towards engineering distinct multi-lamellated outer and inner annulus fibrosus tissues


The annulus fibrosus (AF) of the intervertebral disc (IVD) has a zonal distribution of phenotypically distinct cells. The outer AF (OAF) cells produce an extracellular matrix (ECM) rich in type I collagen with little proteoglycans, whereas the ECM of the inner AF (IAF) has abundant type II collagen and proteoglycans. The inhomogeneous distribution of the ECM in the AF may reflect the complex mechanical forces that the IVD experiences. A bioengineered AF tissue should recapitulate both the inner and outer zones in order to have proper functionality. The aim of this study is to generate multi-lamellated OAF and IAF tissues with ECM compositions that resemble their zonal origin using polycarbonate urethane (PU) scaffolds. It was observed that supplementation of the media with insulin-transferrin-selenium (ITS) and proline yielded tissues with good cellularity. However, IAF cells accumulated only type I collagen, similar to OAF cells. Addition of dexamethasone and sodium pyruvate induced the accumulation of IAF tissues rich in type II collagen and aggrecan, without altering the accumulation of type I collagen in OAF tissues. Dexamethasone stimulated mitochondrial membrane potential in both tissues in the presence of sodium pyruvate, suggesting a relationship between the mitochondrial aerobic respiratory state and dexamethasone signalling during the in vitro-tissue formation by OAF and IAF cells. Inhibition of the glucocorticoid receptor blocked the stimulation of mitochondrial membrane potentials and type II collagen accumulation. In summary, biologically distinct multi-lamellated OAF and IAF tissues can be generated, which will facilitate advancement towards the goal of engineering a biological IVD replacement. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

Degradation alters the lubrication of articular cartilage by high viscosity, hyaluronic acid-based lubricants


Hyaluronic acid (HA) is widely injected as a viscosupplement in the treatment of osteoarthritis. Despite its extensive use, it is not currently known if cartilage degradation alters how HA-based solutions lubricate the articular surface. In this study we utilized a model of cartilage degradation by IL-1β along with a recently developed framework to study role of cartilage degradation on lubrication by clinically-approved HA-based lubricants with high viscosities. Cartilage explants were cultured up to 8 days with 10 ng/ml IL-1β. After culture, samples were examined histologically, immunohistochemically, biochemically, mechanically, topographically, and tribologically. The tribological testing analyzed both boundary and mixed lubrication modes to assess individual effects of viscosity and boundary lubricating ability. Friction testing was carried out using PBS and two clinically approved HA-based viscosupplements in a cartilage-glass configuration. After culture with IL-1β, boundary mode friction was elevated after both 4 and 8 days. Additionally, friction in mixed mode lubrication, where HA is most effective as a lubricant, was significantly elevated after 8 days of culture. As cartilage became rougher, softer, and more permeable after culture, the boundary mode plateau was extended, and as a result, significantly increased lubricant viscosities or sliding speeds were necessary to achieve effective mixed lubrication. Overall, this study revealed that lubrication of cartilage by HA is degradation-dependent and coincides with changes in mechanics and roughness. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

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Issue Information - Editorial Board and TOC


JOR Virtual Issue on stem cells in orthopaedics


Arthrotomy-based preclinical models of particle-induced osteolysis: A systematic review


We completed a systematic literature review of in vivo animal models that use arthrotomy-based methods to study particle-induced peri-implant osteolysis. The purpose of the review was to characterize the models developed to date, to determine the questions addressed, to assess scientific rigor and transparency, and to identify gaps in knowledge. We probed three literature databases (Medline, Embase, and Scopus) and found 77 manuscripts that fit the search parameters. In the most recent 10 years, researchers mainly used rat and mouse models, whereas in the previous 20 years, large animal, canine, and rabbit models were more common. The studies have demonstrated several pathophysiology pathways, including macrophage migration, particle phagocytosis, increased local production of cytokines and lysosomal enzymes, elevated bone resorption, and suppressed bone formation. The effect of variation in particle characteristics and concentration received limited attention with somewhat mixed findings. Particle contamination by endotoxin was shown to exacerbate peri-implant osteolysis. The possibility of early diagnosis was demonstrated through imaging and biomarker approaches. Several studies showed that both local and systemic delivery of bisphosphonates inhibits the development of particle-induced osteolysis. Other methods of inhibiting osteolysis include the use of anabolic agents and altering the implant design. Few studies examined non-surgical rescue of loosened implants, with conflicting results with alendronate. We found that the manuscripts often lacked the methodological detail now advocated by the ARRIVE guidelines, suggesting that improvement in reporting would be useful to maximize rigor and transparency. © 2017 Orthopaedic Research Societ[...]

Bench-to-bedside: Bridge-enhanced anterior cruciate ligament repair


Anterior cruciate ligament (ACL) injuries are one of the most well-known orthopaedic injuries and are treated with one of the most common orthopaedic procedures performed in the United States. This surgical procedure, ACL reconstruction, is successful at restoring the gross stability of the knee. However, the outcomes of ACL reconstruction can be limited by short and long-term complications, including muscle weakness, graft rupture, and premature osteoarthritis. Thus, new methods of treating this injury are being explored. This review details the pathway of how a tissue engineering strategy can be used to improve the healing of the ACL in preclinical studies and then translated to patients in an FDA-approved clinical study. This review paper will outline the clinical importance of ACL injuries, history of primary repair, the pathology behind failure of the ACL to heal, pre-clinical studies, the FDA approval process for a high risk medical device, and the preliminary results from a first-in-human study. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:2606–2612, 2017.

Inflammatory cytokine response is greater in acute tibial plafond fractures than acute tibial plateau fractures


The purpose of the study was to compare the inflammatory cytokine and matrix metalloproteinase (MMP) concentrations in synovial fluid after acute plafond fracture with acute tibial plateau fracture. Between December 2011 and August 2014, we prospectively enrolled patients with acute tibial plateau and plafond fractures. Synovial fluid aspirations were obtained from injured and uninjured joints. The concentrations of IL-1β, IL-1RA, IL-6, IL-8, IL-10, MCP-1, TNF-α, MMP-1, -3, -9, -10, -12, and -13 were quantified using multiplex assays. A Bonferroni correction was used so that the adjusted alpha level for significance was p [...]

New opportunities for fracture healing detection: Impedance spectroscopy measurements correlate to tissue composition in fractures


Accurate evaluation of fracture healing is important for clinical decisions on when to begin weight-bearing and when early intervention is necessary in cases of fracture nonunion. While the stages of healing involving hematoma, cartilage, trabecular bone, and cortical bone have been well characterized histologically, physicians typically track fracture healing by using subjective physical examinations and radiographic techniques that are only able to detect mineralized stages of bone healing. This exposes the need for a quantitative, reliable technique to monitor fracture healing, and particularly to track healing progression during the early stages of repair. The goal of this study was to validate the use of impedance spectroscopy to monitor fracture healing and perform comprehensive evaluation comparing measurements with histological evidence. Here, we show that impedance spectroscopy not only can distinguish between cadaver tissues involved throughout fracture repair, but also correlates to fracture callus composition over the middle stages of healing in wild-type C57BL/6 mice. Specifically, impedance magnitude has a positive relationship with % trabecular bone and a negative relationship with % cartilage, and the opposite relationships are found when comparing phase angle to these same volume fractions of tissues. With this information, we can quantitatively evaluate how far a fracture has progressed through the healing stages. Our results demonstrate the feasibility of impedance spectroscopy for detection of fracture callus composition and reveals its potential as a method for early detection of bone healing and fracture nonunion. © 2017 Orthopaedic Resear[...]

Prediction of normal bone anatomy for the planning of corrective osteotomies of malunited forearm bones using a three-dimensional statistical shape model


Corrective osteotomies of the forearm based on 3D computer simulation using contralateral anatomy as a reconstruction template is an approved method. Limitations are existing considerable differences between left and right forearms, and that a healthy contralateral anatomy is required. We evaluated if a computer model, not relying on the contralateral anatomy, may replace the current method by predicting the pre-traumatic healthy shape. A statistical shape model (SSM) was generated from a set of 59 CT scans of healthy forearms, encoding the normal anatomical variations. Three different configurations were simulated to predict the pre-traumatic shape with the SSM (cross-validation). In the first two, only the distal or proximal 50% of the radius were considered as pathological. In a third configuration, the entire radius was assumed to be pathological, only the ulna being intact. Corresponding experiments were performed with the ulna. Accuracy of the prediction was assessed by comparing the predicted bone with the healthy model. For the radius, mean rotation accuracy of the prediction between 2.9 ± 2.2° and 4.0 ± 3.1° in pronation/supination, 0.4 ± 0.3° and 0.6 ± 0.5° in flexion/extension, between 0.5 ± 0.3° and 0.5 ± 0.4° in radial-/ulnarduction. Mean translation accuracy along the same axes between 0.8 ± 0.7 and 1.0 ± 0.8 mm, 0.5 ± 0.4 and 0.6 ± 0.4 mm, 0.6 ± 0.4 and 0.6 ± 0.5 mm, respectively. For the ulna, mean rotation accuracy between 2.4 ± 1.9° and 4.7 ± 3.8° in pronation/supination, 0.3 ± 0.3° and 0.8 ± 0.6° in flexion/extension, 0.3[...]

Low-dose BMP-2 and MSC dual delivery onto coral scaffold for critical-size bone defect regeneration in sheep


Tissue-engineered constructs (TECs) combining resorbable calcium-based scaffolds and mesenchymal stem cells (MSCs) have the capability to regenerate large bone defects. Inconsistent results have, however, been observed, with a lack of osteoinductivity as a possible cause of failure. This study aimed to evaluate the impact of the addition of low-dose bone morphogenetic protein-2 (BMP-2) to MSC-coral-TECs on the healing of clinically relevant segmental bone defects in sheep. Coral granules were either seeded with autologous MSCs (bone marrow-derived) or loaded with BMP-2. A 25-mm-long metatarsal bone defect was created and stabilized with a plate in 18 sheep. Defects were filled with one of the following TECs: (i) BMP (n = 5); (ii) MSC (n = 7); or (iii) MSC-BMP (n = 6). Radiographic follow-up was performed until animal sacrifice at 4 months. Bone formation and scaffold resorption were assessed by micro-CT and histological analysis. Bone union with nearly complete scaffold resorption was observed in 1/5, 2/7, and 3/6 animals, when BMP-, MSC-, and MSC-BMP-TECs were implanted, respectively. The amount of newly formed bone was not statistically different between groups: 1074 mm3 [970–2478 mm3], 1155 mm3 [970–2595 mm3], and 2343 mm3 [931–3276 mm3] for BMP-, MSC-, and MSC-BMP-TECs, respectively. Increased scaffold resorption rate using BMP-TECs was the only potential side effect observed. In conclusion, although the dual delivery of MSCs and BMP-2 onto a coral scaffold further increased bone formation and bone union when compared to single treatment, results were non-significant. Only 5[...]

Reproducibility of three dimensional digital preoperative planning for the osteosynthesis of distal radius fractures


A three-dimensional (3D) digital preoperative planning system for the osteosynthesis of distal radius fractures was developed for clinical practice. To assess the usefulness of the 3D planning for osteosynthesis, we evaluated the reproducibility of the reduction shapes and selected implants in the patients with distal radius fractures. Twenty wrists of 20 distal radius fracture patients who underwent osteosynthesis using volar locking plates were evaluated. The 3D preoperative planning was performed prior to each surgery. Four surgeons conducted the surgeries. The surgeons performed the reduction and the placement of the plate while comparing images between the preoperative plan and fluoroscopy. Preoperative planning and postoperative reductions were compared by measuring volar tilt and radial inclination of the 3D images. Intra-class correlation coefficients (ICCs) of the volar tilt and radial inclination were evaluated. For the implant choices, the ICCs for the screw lengths between the preoperative plan and the actual choices were evaluated. The ICCs were 0.644 (p [...]

Electromagnetic tracking for femoral derotation osteotomy—an in vivo study


Femoral derotation osteotomy delivers good to excellent results in the treatment of rotational gait abnormalities and especially in internal rotation gait. The outcome of the procedure has been evaluated in numerous short- and long-term studies. Although reasons for recurrence and over-/under-correction have been unveiled in earlier studies the mechanisms are still not fully understood. False intra-operative assessment of the derotation angle may contribute to imprecise outcomes. In a recent saw-bone study we evaluated an electromagnetic tracking system in comparison to conventional goniometer measurement and a CT reference measurement and found it to be extremely accurate, whereas the use of a conventional goniometer for derotation measurement showed a high inter- and intra-rater variability. The current study evaluates the electromagnetic tracking system for continuous intra-operative derotation control under real OR conditions. Adults (age: 18–40 years) with the diagnosis of internal rotation gait, independent of the underling pathology, undergoing a supracondylar deroation osteotomy were included. A rotational CT scan was conducted before and in close proximity after surgery and the difference served as reference for the electromagnetic tracking results. The results showed a mean deviation of 2.6° (1.2–5.5°) in comparison to the reference measurement of the pre- and post-operative CT scans. The system proved to be stable under OR conditions with a good usability and a small technical footprint. Electromagnetic tracking delivers a precise, relia[...]

Calreticulin inhibits inflammation-induced osteoclastogenesis and bone resorption


Osteoclasts play key roles in bone remodeling and pathologic osteolytic disorders such as inflammation, infection, bone implant loosening, rheumatoid arthritis, metastatic bone cancers, and pathological fractures. Osteoclasts are formed by the fusion of monocytes in response to receptor activators of NF-κB-ligand (RANKL) and macrophage colony stimulating factor 1 (M-CSF). Calreticulin (CRT), a commonly known intracellular protein as a calcium-binding chaperone, has an unexpectedly robust anti-osteoclastogenic effect when its recombinant form is applied to osteoclast precursors in vitro or at the site of bone inflammation externally in vivo. Externally applied Calreticulin was internalized inside the cells. It inhibited key pro-osteoclastogenic transcription factors such as c-Fos and nuclear factor of activated T cells, cytoplasmic 1 (NFATc1)-in osteoclast precursor cells that were treated with RANKL in vitro. Recombinant human Calreticulin (rhCRT) inhibited lipopolysaccharide (LPS)-induced inflammatory osteoclastogenesis in the mouse calvarial bone in vivo. Cathepsin K molecular imaging verified decreased Cathepsin K activity when rhCalreticulin was applied at the site of LPS application in vivo. Recombinant forms of intracellular proteins or their derivatives may act as novel extracellular therapeutic agents. We anticipate our findings to be a starting point in unraveling hidden extracellular functions of other intracellular proteins in different cell types of many organs for new therapeutic opportunities. © 2017 Orthopaedic[...]

Comparative investigation of bone mineral density using CT and DEXA in a canine femoral model


Bone density measurements using computed tomography (CT) instead of dual-energy X-ray absorptiometry (DEXA) are currently of great interest in human and veterinary medical research as it would be beneficial to use CT scans obtained for other indications also for determining bone density. For Hounsfield units (HU) measured with CT in specific regions of interests (ROIs) in one or several slice/s a correlation with bone mineral density (BMD) measured by DEXA in humans and dogs of between 0.44 and 0.77 is reported in the literature. In the present study, instead certain volumes of interest (VOIs) obtained by CT scan and the corresponding HU to the respective VOIs were compared with the bone mineral density of the corresponding areas measured by DEXA. The aim of the study was to investigate whether this procedure gives more accurate information about bone density of the bones as three-dimensional objects of the respective patient. Correlation between measured HU in the respective VOI and BMD measured with DEXA in the corresponding ROI showed a very good correlation of 0.93. Linear regression with R2 = 0.85 (p = 0.0262) was calculated. Except for VOI5, similar distribution of values and significant differences (p [...]

Intraoperative biomechanics of lumbar pedicle screw loosening following successful arthrodesis


Pedicle screw loosening has been implicated in recurrent back pain after lumbar spinal fusion, but the degree of loosening has not been systematically quantified in patients. Instrumentation removal is an option for patients with successful arthrodesis, but remains controversial. Here, we quantified pedicle screw loosening by measuring screw insertion and/or removal torque at high statistical power (beta = 0.02) in N = 108 patients who experienced pain recurrence despite successful fusion after posterior instrumented lumbar fusion with anterior lumbar interbody fusion (L2–S1). Between implantation and removal, pedicle screw torque was reduced by 58%, indicating significant loosening over time. Loosening was greater in screws with evoked EMG threshold under 11 mA, indicative of screw misplacement. A theoretical stress analysis revealed increased local stresses at the screw interface in pedicles with decreased difference in pedicle thickness and screw diameter. Loosening was greatest in vertebrae at the extremities of the fused segments, but was significantly lower in segments with one level of fusion than in those with two or more. Clinical significance: These data indicate that pedicle screws can loosen significantly in patients with recurrent back pain and warrant further research into methods to reduce the incidence of screw loosening and to understand the risks and potential benefits of instrumentation removal. © 2017 Orthopaedic Research Society. Published [...]

Age-related reduction in the expression of FOXO transcription factors and correlations with intervertebral disc degeneration


Aging is a main risk factor for intervertebral disc (IVD) degeneration, the main cause of low back pain. FOXO transcription factors are important regulators of tissue homeostasis and longevity. Here, we determined the expression pattern of FOXO in healthy and degenerated human IVD and the associations with IVD degeneration during mouse aging. FOXO expression was assessed by immunohistochemistry in normal and degenerated human IVD samples and in cervical and lumbar IVD from 6-, 12-, 24-, and 36-month-old C57BL/6J mice. Mouse spines were graded for key histological features of disc degeneration in all the time points and expression of two key FOXO downstream targets, sestrin 3 (SESN3) and superoxide dismutase (SOD2), was assessed by immunohistochemistry. Histological analysis revealed that FOXO proteins are expressed in all compartments of human and mouse IVD. Expression of FOXO1 and FOXO3, but not FOXO4, was significantly deceased in human degenerated discs. In mice, degenerative changes in the lumbar spine were seen at 24 and 36 months of age whereas cervical IVD showed increased histopathological scores at 36 months. FOXO expression was significantly reduced in lumbar IVD at 12-, 24-, and 36-month-old mice and in cervical IVD at 36-month-old mice when compared with the 6-month-old group. The reduction of FOXO expression in lumbar IVD was concomitant with a decrease in the expression of SESN3 and SOD2. These findings suggest that reduced FOXO expr[...]

Microcalcification of lumbar spine intervertebral discs and facet joints is associated with cartilage degeneration, but differs in prevalence and its relation to age


Cartilage calcification (CC) is associated with degeneration in non-vertebral joints, but little is known about CC and lumbar vertebral joints. The goal of this study was to analyze the prevalence of CC in lumbar facet joints (FJ) and intervertebral discs (IVD) and its relation to cartilage degeneration and age in a non-selected cohort of the general population. The segment L4/5 of 85 consecutive donors (mean age 61.9 years) was analyzed by high-resolution imaging digital-contact radiography (DCR). Quantification was achieved by measuring CC in % of total cartilage area. Histological degeneration of FJs and IVDs was determined by OARSI and Boos scores. Prevalence of CC was 36.5% for FJ (95%CI (0.26, 0.48)) and 100% for IVD (95%CI (0.96, 1.00)). The amount of IVD CC (3.36% SD ± 7.14) was 16.3 times higher (p [...]

Lumbar multifidus muscle degenerates in individuals with chronic degenerative lumbar spine pathology


Histological and cell-level changes in the lumbar musculature in individuals with chronic lumbar spine degenerative conditions are not well characterized. Although prior literature supports evidence of changes in fiber type and size, little information exists describing the tissue quality and biology of pathological features of muscle in this population. The purpose of this study was to quantify multifidus tissue composition and structure, inflammation, vascularity, and degeneration in individuals with chronic degenerative lumbar spine pathology. Human multifidus biopsies were acquired from 22 consecutive patients undergoing surgery for chronic degenerative lumbar spine pathology. Relative fractions of muscle, adipose, and extracellular matrix were quantified along with muscle fiber type and cross-sectional area (CSA) and markers of inflammation, vascularity, satellite cell density, and muscle degeneration. On average, multifidus biopsies contained 48.5% muscle, 11.7% adipose tissue, and 26.1% collagen tissue. Elevated inflammatory cell counts (48.5 ± 30.0 macrophages/mm2) and decreased vascularity (275.6 ± 69.4 vessels/mm2) were also observed compared to normative values. Satellite cell densities were on average 13 ± 9 cells per every 100 muscle fibers. Large fiber CSA (3,996.0 ± 1,909.2 µm2) and a predominance of type I fibers (61.8 ± 18.0%) were observed in a[...]

Collagen V haploinsufficiency in a murine model of classic Ehlers–Danlos syndrome is associated with deficient structural and mechanical healing in tendons


Classic Ehlers–Danlos syndrome (EDS) patients suffer from connective tissue hyperelasticity, joint instability, skin hyperextensibility, tissue fragility, and poor wound healing due to heterozygous mutations in COL5a1 or COL5a2 genes. This study investigated the roles of collagen V in establishing structure and function in uninjured patellar tendons as well as in the injury response using a Col5a1+/− mouse, a model for classic EDS. These analyses were done comparing tendons from a classic EDS model (Col5a1+/−) with wild-type controls. Tendons were subjected to mechanical testing, histological, and fibril analysis before injury as well as 3 and 6 weeks after injury. We found that Col5a1+/− tendons demonstrated diminished recovery of mechanical competency after injury as compared to normal wild-type tendons, which recovered their pre-injury values by 6 weeks post injury. Additionally, the Col5a1+/−tendons demonstrated altered fibril morphology and diameter distributions compared to the wild-type tendons. This study indicates that collagen V plays an important role in regulating collagen fibrillogenesis and the associated recovery of mechanical integrity in tendons after injury. In addition, the dysregulation with decreased collagen V expression in EDS is associated with a diminished injury response. The results presented herein have the potential to direct fu[...]