Structural Consequences of a Partial Anterior Cruciate Ligament Injury on Remaining Joint Integrity: Evidence for Ligament and Bone Changes Over Time in an Ovine Model.

BACKGROUND: Severe injury to the knee joint often results in accelerated posttraumatic osteoarthritis (PTOA). In an ovine knee injury model, altered kinematics and degradation of the cartilage have been observed at 20 and 40 weeks after partial anterior cruciate ligament (ACL) transection (p-ACL Tx) surgery. However, changes to the integrity of the remaining intact intra-articular ligaments (posterolateral [PL] band and posterior cruciate ligament [PCL]) as well as the subchondral bone after anteromedial (AM) band Tx remain to be characterized. PURPOSE: (1) To investigate histological alterations to the remaining intact intra-articular ligaments, the synovium, and the infrapatellar fat pad (IPFP) and (2) to quantify subchondral bone changes at the contact surfaces of the proximal tibia at 20 and 40 weeks after AM band Tx. STUDY DESIGN: Descriptive laboratory study. METHODS: Mature female Suffolk cross sheep were allocated into 3 groups: nonoperative controls (n = 6), 20 weeks after partial ACL transection (p-ACL Tx; n = 5), and 40 weeks after p-ACL Tx (n = 6). Ligament, synovium, and IPFP sections were stained and graded. Tibial subchondral bone microarchitecture was assessed using high-resolution peripheral quantitative computed tomography. RESULTS: p-ACL Tx of the AM band led to significant change in histological scores of the PL band and the PCL at 20 weeks after p-ACL Tx (P = .031 and P = .033, respectively) and 40 weeks after p-ACL Tx (P = .011 and P = .029) as compared with nonoperative controls. Alterations in inflammatory cells and collagen fiber orientation contributed to the greatest extent of the combined histological score in the PL band and PCL. p-ACL Tx did not lead to chronic activation of the synovium or IPFP. Trabecular bone mineral density was strongly inversely correlated with combined gross morphological damage in the top and middle layers of the subchondral bone in the lateral tibial plateau for animals at 40 weeks after p-ACL Tx. CONCLUSION: p-ACL Tx influences the integrity (biology and structure) of remaining intact intra-articular ligaments and bone microarchitecture in a partial knee injury ovine model. CLINICAL RELEVANCE: p-ACL Tx leads to alterations in structural integrity of the remaining intact ligaments and degenerative changes in the trabecular bone mineral density, which may be detrimental to the injured athlete's knee joint in the long term.

Molecular Response of Rabbit Menisci to Surgically Induced Hemarthrosis and a Single Intra-Articular Dexamethasone Treatment.

Anterior cruciate ligament reconstructive surgery can restore biomechanical stability, however, such surgery cannot reliably prevent the onset of post-traumatic osteoarthritis. The aim of this study was to elucidate the molecular response that occurs within the menisci following a surgical injury that allows bleeding into the joint space, and then to investigate the effect of dexamethasone (DEX) on this molecular response. Cell viability studies following acute controlled exposure to blood and blood plus DEX were also conducted. Forty-eight New Zealand white rabbits were randomly allocated into control, sham, surgical, and surgical + DEX groups (each group n = 6). Animals were sacrificed at 48 h and 9 weeks, and menisci were harvested. The messenger RNA (mRNA) expression levels for key inflammatory, and degradative proteins, as well as mRNA levels for autophagy pathway molecules were quantified, and statistically significant changes were described. Meniscal cell viability was calculated by incubating groups of medial and lateral menisci in autologous blood, or autologous blood plus DEX for 48 h (each group n = 4; total of eight medial and eight lateral menisci), and then conducting a histological live/dead assay. Results indicated a significant reduction in only medial meniscal cell viability when the tissue was exposed to blood in combination with DEX. A single administration of DEX following surgery significantly suppresses the elevated molecular expression for key inflammatory and degradative markers within menisci at 48 h and 9 weeks post-surgery. In vitro, autologous blood did not affect cell viability, but addition of DEX uniquely impacted the medial menisci. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:2043-2052, 2019.

The infrapatellar fat pad is affected by injury induced inflammation in the rabbit knee: use of dexamethasone to mitigate damage.

OBJECTIVE AND DESIGN: The health of the infrapatellar fat pad (IFP) has been linked to pain, joint inflammation, and the onset of post-traumatic osteoarthritis. Thus, early inflammation effects on the IFP could have long term sequelae on joint integrity. This study was designed to characterize the natural history of the IFP in a model of surgically induced knee injury and inflammation, and to test the efficacy of one intra-articular (IA) administration of dexamethasone (DEX) immediately following surgery. METHODS: An IA bone drill hole injury to the rabbit knee was conducted and immediately treated with DEX (n = 12). Early and late post-surgical time-points were investigated (48 h and 9 weeks) and the outcome measures were analysis of IFP histology, mRNA levels for relevant molecules, and protein levels for a subset of cytokines. Data were analyzed against a surgical control (injury without treatment; n = 12), a surgical sham (capsular incision only; n = 12), and normal control (n = 6). TREATMENT: Single IA injection of DEX (0.5 mg/kg), administered at the completion of surgery. RESULTS: IFPs from injured joints exhibited significantly increased cellularity and early fibrosis at 48 h post surgery. While the histological inflammation from a capsular incision alone resolved, knee injured animals progressed to a significantly more fibrotic IFP by 9 weeks. DEX significantly lowered histological scores at 48 h, but not at the 9 weeks. DEX did not influence mRNA levels for IL-1ß, 6, and 8, however, protein analysis indicated that IL-8 levels were lower in DEX treated joints. DEX resulted in significantly elevated expression of mRNA for MCP-1, leptin, and VEGF. CONCLUSION: One IA administration of a glucocorticoid appears to mitigate the initial inflammation within the joint, but is not sufficient to protect the joint to 9 weeks post-surgery.

Single intra-articular dexamethasone injection immediately post-surgery in a rabbit model mitigates early inflammatory responses and post-traumatic osteoarthritis-like alterations.

Despite surgical reconstruction of the anterior cruciate ligament, a significant number of patients will still develop post-traumatic osteoarthritis (PTOA). Our objective was to determine if mitigating aspects of the acute phase of inflammation following a defined knee surgery with a single administration of a glucocorticoid could prevent the development of PTOA-like changes within an established rabbit model of surgically induced PTOA. An early and late post-surgical time-point was investigated in this study (48 h and 9 weeks post-surgery) in which the following groups were repeated (each n=6, for a total of 24 rabbits per time-point, and 48 rabbits used in the study): control (age/sex matched), sham (arthrotomy), drill injury (arthrotomy+two drill holes to a non-cartilaginous area of the femoral notch), and drill injury+single intra-articular (IA) injection of dexamethasone (DEX). At 48 h post-surgery, DEX treatment significantly lowered the mRNA levels for a subset of pro-inflammatory mediators, and significantly lowered the histological grade. Nine weeks post surgery, DEX treatment significantly lowered the histological scores (presented as effect size) for synovium (3.8), lateral femoral condyle (3.9), and lateral tibial cartilage (5.1) samples. Thus, DEX likely acts to prevent injury induced inflammation that could contribute to subsequent joint damage.

Alterations in Hoffa's fat pad induced by an inflammatory response following idealized anterior cruciate ligament surgery.

OBJECTIVE: To determine whether inflammation following anterior cruciate ligament (ACL) reconstruction leads to long-term pathological changes in the infrapatellar fat pad (IPFP or Hoffa's fat pad) which could compromise the integrity of the knee joint. MATERIALS AND METHODS: Sixteen mature sheep underwent anatomic idealized ACL reconstruction surgery (ACL-R) and were sacrificed at 2 weeks (n = 9) and 20 weeks (n = 7) post-ACL-R. Five additional animals served as unoperated controls. A histological grading protocol was developed to quantify the changes in the IPFP post-injury. mRNA expression levels for key markers of inflammation, angiogenesis and tissue regeneration were assessed by qPCR. RESULTS: The IPFP exhibited altered cellularity and fibrosis at 2 and 20 weeks post-ACL-R. Immunohistochemistry detected macrophage-like cells in the IPFP which were increased at 20 weeks. Specific pro-inflammatory cytokines and IPFP specific adipokines exhibited changes indicating early inflammation mediated alterations. Elevations in CD105 mRNA levels at 2 weeks corroborated the increases in neovascularization observed in the IPFP following injury. CONCLUSIONS: Sustained long-term pathological changes stemming from inflammation are present in IPFP tissue after ACL-R surgery and may compromise the long-term integrity of the knee joint.

Metabolic profiling of synovial fluid in a unilateral ovine model of anterior cruciate ligament reconstruction of the knee suggests biomarkers for early osteoarthritis.

Joint injuries and subsequent osteoarthritis (OA) are the leading causes of chronic joint disease. In this work, we explore the possibility of applying magnetic resonance spectroscopy-based metabolomics to detect host responses to an anterior cruciate ligament (ACL) reconstruction injury in synovial fluid in an ovine model. Using multivariate statistical analysis, we were able to distinguish post-injury joint samples (ACL and sham surgery) from the uninjured control samples, and as well the ACL surgical samples from sham surgery. In all samples there were 65 metabolites quantified, of which six could be suggested as biomarkers for early post-injury degenerative changes in the knee joints: isobutyrate, glucose, hydroxyproline, asparagine, serine, and uridine. Our results raise a cautionary note indicating that surgical interventions into the knee can result in metabolic alterations that need to be distinguished from those caused by the early onset of OA. Our findings illustrate the potential application of metabolomics as a diagnostic and prognostic tool for detection of injuries to the knee joint. The ability to detect a unique pattern of metabolic changes in the synovial fluid of sheep offers the possibility of extending the approach to precision medicine protocols in patient populations in the future.

Normal sheep synovium has similar appearances and constitutive expression of inflammatory cytokines within and between knee joints: a baseline histological and molecular analysis.

Abstract Clinical evidence suggests that synovium can add to adjacent articular cartilage damage, potentially contributing to the development of osteoarthritis (OA). Inflammation of the synovium (synovitis) is dependent on the type of injury sustained, the time after injury and concomitant changes in other joint tissues. To define the role of synovitis in OA development, there is a need for baseline measures that can reliably distinguish synovial inflammation from normal synovium both within and between joints. This study tested the hypothesis that normal synovium from distinct anatomical locations in young and adult sheep is homogeneous with respect to consistently low molecular expression of the inflammatory mediators - tumour necrosis factor alpha (TNF-α) and interleukins (IL) such as IL-1ß, IL-1Ra, IL-6 and IL-8. Additionally, maturation will not influence the expression of these select inflammatory biomarkers. Samples of synovium from four anatomic locations (medial and lateral margins, suprapatellar pouch (patella region), posterior to the posterior cruciate ligament, from each joint of 5 adult and 4 immature animals were graded histologically or analyzed for mRNA expression of inflammatory cytokines. Histologically, no evidence of synovitis was noted although some variance in sub-intimal fibrosis was observed between sample locations in mature sheep. Molecular expression of all inflammatory mediators was low and homogeneously expressed at constitutive levels in all sample locations. These findings confirm the hypothesis that the normal sheep synovium is a homogeneous tissue throughout the joint and establishes the baseline expression levels for several pro-inflammatory mediators in both immature and mature sheep.

Dexamethasone inhibits inflammation and cartilage damage in a new model of post-traumatic osteoarthritis.

Corticosteroids are used in musculoskeletal diseases, and offer patient relief. Injections of corticosteroids are recommended for management of osteoarthritis (OA). Current data have shown the role of corticosteroids in ameliorating pain. We hypothesized that repeated intra-articular injections of high dose dexamethasone would protect the cartilage from damage in a post-traumatic model of OA. Eighteen female New Zealand White rabbits were used. Twelve underwent surgery to induce OA; six of them received intra-articular injections of dexamethasone every 3 days for 3 weeks. The other six rabbits served as operated controls. Six additional rabbits served as non-operated controls. All animals were euthanized 3 weeks post-surgery. Knees were assessed grossly. Cartilage, synovium, and fat pad were assessed histologically. Synovium and fat pad were analyzed with qPCR and Western blots. Surgical controls had cartilage damage which was supressed with dexamethasone. Dexamethasone significantly decreased synovial expression of interleukin-1ß and collagen I, and a trend to decrease synovial matrix metalloproteinase3 expression. There were also significantly lower levels of interleukin-1ß protein with dexamethasone treatment. Dexamethasone significantly decreased fat pad expression of matrix metalloproteinase13, basic fibroblast growth factor, and interleukin8, and a trend to decrease matrix metalloproteinase3 and transforming growth factorß expression. Dexamethasone decreased joint inflammation and joint tissue degradation and was chondroprotective in this unique model of PTOA.

Kinematic and kinetic interactions during normal and ACL-deficient gait: a longitudinal in vivo study.

The interactions between different tissues within the knee joint and between different kinematic DOF and joint flexion during normal gait were investigated. These interactions change following ACL transection, in both short (4 weeks) and long (20 weeks) term. Ten skeletally mature sheep were used in control (N = 5) and experimental (N = 5) groups. The 6-DOF stifle joint motion was first measured during normal gait. The control group were then euthanized and mounted on a unique robotic testing platform for kinetic measurements. The experimental group underwent ACL transection surgery, and kinematics measurements were repeated 4 and 20 weeks post-operatively. The experimental group were then euthanized and underwent kinetic assessment using the robotic system. Results indicated significant couplings between joint flexion vs. abduction and internal tibial rotation, as well as medial, anterior, and superior tibial translations during both normal and ACL-deficient gait. Distinct kinetic interactions were also observed between different tissues within the knee joint. Direct relationships were found between ACL vs. LM/MM, and PCL vs. MCL loads during normal gait; inverse relationships were detected between ACL vs. PCL and PCL vs. LM/MM loads. These kinetic interaction patterns were considerably altered by ACL injury. Significant inter-subject variability in joint kinematics and tissue loading patterns during gait was also observed. This study provides further understanding of the in vivo function of different tissues within the knee joint and their couplings with joint kinematics during normal gait and over time following ACL transection.

Estrogen receptor beta and truncated variants enhance the expression of transfected MMP-1 promoter constructs in response to specific mechanical loading.

BACKGROUND: Joint diseases such as osteoarthritis (OA) predominantly afflict post-menopausal women, suggesting a pertinent role for female hormones. Estrogen receptor beta (ER-ß) has been detected in connective tissues of the knee joint suggesting that these tissues are responsive to the hormone estrogen. Matrix metalloproteinase-1 (MMP-1) activity contributes to cartilage degradation, a key factor leading to OA development in synovial joints. Two polymorphic forms of MMP-1 exist due to a deletion/insertion of the guanine residue in the promoter, and the 2G allelic variant of MMP-1 exhibits more activity than the 1G allele. Previous studies have demonstrated that the polymorphic forms of the human MMP-1 are influenced by the modulating effects of estrogen receptor isoforms. In addition to hormonal influences, physiological factors such as altered mechanical loading are also contributory features of OA. In the present study, the combined influence of biomechanical and hormonal variables on the activity of MMP-1 isoforms was evaluated. We hypothesized that the combined effects of ER-ß and sheer stress will differentially activate the two allelic forms of MMP-1 in a hormone-independent manner. METHODS: HIG-82 synoviocytes were transiently transfected with 1G or 2G alleles (±) ER-ß and subjected to either shear or equibiaxial stress. Next, 1G/2G promoter activity was measured to determine the combined influence of physiological stimuli. Truncated ER-ß constructs were used to determine the importance of different domains of ER-ß on 1G/2G activation. RESULTS: The 2G allele exhibited a constitutively higher activity than the 1G allele, which was further increased when the transfected cells were subject to shear stress, but not equibiaxial stress. Moreover, the combination of ER-ß and shear stress further increased the activity levels of the 1G/2G allelic variants. Additionally, select AF-2 truncated ER-ß variants led to increased activity levels for the 2G allele, indicating the AF-1 domain was likely involved in the response to mechanical stimulation. CONCLUSIONS: These results suggest that the 1G/2G alleles of MMP-1 are influenced by specific mechanical stimuli like shear stress, as well as the ER-ß receptor. These findings contribute to the potential allelic involvement in connective tissue diseases such as OA in females compared to males.

Characterization of proteoglycan 4 and hyaluronan composition and lubrication function of ovine synovial fluid following knee surgery.

The objective of this study was to determine changes in (1) proteoglycan 4 (PRG4) and hyaluronan (HA) concentration, (2) HA molecular weight (MW) distribution, and (3) cartilage boundary lubricating ability of synovial fluid (SF) from surgical sham (SHAM), anterior cruciate ligament (ACL)/medial collateral ligament (MCL) transection, and lateral meniscectomy (MEN) in a post-knee surgery ovine model. Ovine SF (oSF) was collected at euthanization 20 weeks after surgery, with the contralateral joint serving as the non-operative control. PRG4 and HA concentration in oSF was measured by sandwich enzyme-linked immunosorbent assay, and HA MW distribution by agarose gel electrophoresis. Cartilage boundary lubricating ability of oSF was measured by a cartilage-cartilage friction test. PRG4 and HA concentration in SHAM, ACL/MCL, and MEN oSF were similar in comparison to the contralateral control (CTRL) oSF. The HA MW distribution in the operated oSF for all ranges were similar to the respective CTRL oSF. The kinetic coefficients of friction in operated and CTRL oSF were similar in all groups, and were significantly lower than saline. These results indicate oSF lubricant composition and function at 20 weeks post-knee surgery were similar to contralateral CTRL, and suggest earlier time points post surgery warrant further investigation.

Tendon mineralization is accelerated bilaterally and creep of contralateral tendons is increased after unilateral needle injury of murine achilles tendons.

Heterotopic mineralization may result in tendon weakness, but effects on other biomechanical responses have not been reported. We used a needle injury, which accelerates spontaneous mineralization of murine Achilles tendons, to test two hypotheses: that injured tendons would demonstrate altered biomechanical responses; and that unilateral injury would accelerate mineralization bilaterally. Mice underwent left hind (LH) injury (I; n = 11) and were euthanized after 20 weeks along with non-injured controls (C; n = 9). All hind limbs were examined by micro computed tomography followed by biomechanical testing (I = 7 and C = 6). No differences were found in the biomechanical responses of injured tendons compared with controls. However, the right hind (RH) tendons contralateral to the LH injury exhibited greater static creep strain and total creep strain compared with those LH tendons (p ≤ 0.045) and RH tendons from controls (p ≤ 0.043). RH limb lesions of injured mice were three times larger compared with controls (p = 0.030). Therefore, despite extensive mineralization, changes to the responses we measured were limited or absent 20 weeks postinjury. These results also suggest that bilateral occurrence should be considered where tendon mineralization is identified clinically. This experimental system may be useful to study the mechanisms of bilateral new bone formation in tendinopathy and other conditions.

New surgical model of post-traumatic osteoarthritis: isolated intra-articular bone injury in the rabbit.

Osteoarthritis (OA) is a leading cause of disability worldwide. We hypothesized that inflammation following isolated intra-articular bone injury can stimulate post-traumatic OA and developed a rabbit model to test that concept. Sixty female New Zealand White Rabbits were used. Twenty-six experimental animals had two holes drilled into their right femoral-notch, 18 rabbits had sham surgery, and 16 were un-operated controls. Rabbits were euthanized in subgroups at 72 h, 3, 6, 9, and 52 weeks. Knees were assessed grossly and tissues collected. Cartilage and synovium were analyzed with histology and qPCR and subgroups compared statistically. All surgical joints showed gross and histological (modified Mankin score) cartilage damage after surgery, with experimentals worsening with time (p < 0.05). Cartilage qPCR showed fivefold increases in TGFß (p < 0.05) expression at 72 h and 3 weeks with sixfold increases in MMP13 (p < 0.025) expression at 72 h. By 6 weeks, expression of these markers was similar to baseline levels. Synovial membrane thickening with increased cellularity was seen at both 9 and 52 weeks (p < 0.05). Short-term synovial inflammatory marker (IL-1ß, IL-Ra, IL-6, and IL-8) expression was three- to fourfold increase in experimentals at 72 h (p < 0.01) returning to baseline levels by 3 weeks. Intra-articular bone injury creates early joint inflammation with some chronic synovial changes and progressive cartilage damage consistent with OA in adult rabbits. This model provides an exciting new avenue to potentially explore some relevant inflammatory drivers of OA without major mechanical variables.

Osteoarthritis develops in the operated joint of an ovine model following ACL reconstruction with immediate anatomic reattachment of the native ACL.

We tested the hypothesis that immediate reattachment of the native anterior cruciate ligament (ACL) can prevent kinematic changes and the development of osteoarthritis (OA). Five sheep underwent anatomic unilateral ACL reconstruction (ACL-R). Animals from a previous study served as sham (n = 7) or non-operated (n = 17) controls. At 4 points of walking gait, 6 degrees of freedom stifle joint kinematics of ACL-R animals were compared with sham controls at 4 and 20 weeks post-surgery. Gross cartilage, bone, and meniscal changes were graded at euthanasia; paired and differential scores were compared. Inter-animal differences were noted in all groups. Of 48 points of gait comparison between ACL-R and sham operated groups, 42 points showed no difference (p > 0.05). Of the six significant differences (p < 0.05), internal rotation in ACL-R animals accounted for three. At 20 weeks, differential scores showed that sham operated joints were morphologically indistinguishable from non-operated controls (p ≥ 0.129) while ACL-R joints had significantly higher combined cartilage and osteophyte scores than those controls (p ≤ 0.003). This method of ACL reconstruction in sheep did not restore normal walking gait kinematics completely and allowed some OA to develop in operated joints. OA may result from relatively subtle mechanical abnormalities, apparently more so in some individuals than others.

Heterotopic mineralization (ossification or calcification) in tendinopathy or following surgical tendon trauma.

Heterotopic tendon mineralization (ossification or calcification), which may be a feature of tendinopathy or which may develop following surgical trauma (repair or graft harvest), has not received much attention. The purpose of this article is to review the prevalence, mechanisms and consequences of heterotopic tendon mineralization and to identify the gaps in our current understanding. We focus on endochondral heterotopic ossification and draw on knowledge of the mechanisms of this process in other tissues and conditions. Finally, we introduce a novel murine Achilles tendon needle injury model, which will enable us to further study the mechanisms and biomechanical consequences of tendon mineralization.

Investigating the potential value of individual parameters of histological grading systems in a sheep model of cartilage damage: the Modified Mankin method.

A total histological grade does not necessarily distinguish between different manifestations of cartilage damage or degeneration. An accurate and reliable histological assessment method is required to separate normal and pathological tissue within a joint during treatment of degenerative joint conditions and to sub-classify the latter in meaningful ways. The Modified Mankin method may be adaptable for this purpose. We investigated how much detail may be lost by assigning one composite score/grade to represent different degenerative components of the osteoarthritic condition. We used four ovine injury models (sham surgery, anterior cruciate ligament/medial collateral ligament instability, simulated anatomic anterior cruciate ligament reconstruction and meniscal removal) to induce different degrees and potentially 'types' (mechanisms) of osteoarthritis. Articular cartilage was systematically harvested, prepared for histological examination and graded in a blinded fashion using a Modified Mankin grading method. Results showed that the possible permutations of cartilage damage were significant and far more varied than the current intended use that histological grading systems allow. Of 1352 cartilage specimens graded, 234 different manifestations of potential histological damage were observed across 23 potential individual grades of the Modified Mankin grading method. The results presented here show that current composite histological grading may contain additional information that could potentially discern different stages or mechanisms of cartilage damage and degeneration in a sheep model. This approach may be applicable to other grading systems.

Complete ACL/MCL deficiency induces variable degrees of instability in sheep with specific kinematic abnormalities correlating with degrees of early osteoarthritis.

People are not equally disabled by combined anterior cruciate ligament (ACL)/medial collateral ligament (MCL) injuries, nor do they all develop osteoarthritis (OA). Although biological/biomechanical causes are not clear, some association presumably exists between joint instability and OA development. We hypothesized that degree of OA development following standardized complete ACL/MCL injuries will vary directly with the degree of biomechanical abnormality between individuals. Three groups of sheep were used to test the hypothesis: 17 normal, 9 ACL/MCL transected, and 7 sham animals. Normal joints were assessed morphologically while sham and experimental animals had gait assessment pre- and at 4 and 20 weeks post-surgery, with cartilage and bone changes being mapped and graded at sacrifice at 20 weeks. Sham joints were morphologically normal and had only one minor kinematic change at 20 weeks. Although variable, ACL/MCL deficient animals showed significant kinematic abnormalities in 4/6 degrees of freedom (DOFs), as well as cartilage/bone damage by 20 weeks (p < 0.05). Linear regression analysis revealed that changes in medial-lateral (ML) translation were related to the current level of joint degradation as represented by total gross OA score (p = 0.0044, R(2) = 0.71) in the ACL/MCL transected group. Even identical ACL/MCL injuries result in inter-animal variations in instability and OA, however significant kinematic abnormalities in ML translation do relate to early OA in sheep.

Early joint tissue changes are highly correlated with a set of inflammatory and degradative synovial biomarkers after ACL autograft and its sham surgery in an ovine model.

While impossible in humans, the mechanisms of early cartilage, bone and meniscal damage can be quantified after anterior cruciate ligament (ACL) injury in animal models. We utilized an ovine model to determine if the mRNA expression of inflammatory and degradative molecules (IL-1ß, IL-6, MMP-1, 2, 3, and 13) in the synovium correlated to changes in joint tissues 2 weeks post-ACL surgery, to test the hypothesis that synovial inflammation is a marker of these changes and possibly their originator. Nine "idealized" ACL autografts were performed and compared with three sham and six normal animals. Using validated protocols, early osteophyte formation, articular cartilage, and meniscal damage were quantified. Synovium was harvested and mRNA expression quantified using qPCR. Multiple linear regression analysis (MLRA) was utilized to correlate synovial mRNA expression in treated and contra-lateral limbs, from all treatment groups with corresponding joint scores. Synovial mRNA expression was significantly elevated in all experimental and sham joints. The MLRA model was a significant predictive tool (p = 0.001, R(2) = 0.70) of gross tissue scores with significant contributions from IL-1ß, IL-6, and MMP-3. Findings suggest that this set of synovial biomarkers is predictive (p < 0.009) of early gross changes of joint tissues after arthrotomy and likely directly involved in the relevant mechanisms, particularly early osteophyte formation, in vivo.

Meniscectomy causes significant in vivo kinematic changes and mechanically induced focal chondral lesions in a sheep model.

Significant meniscal loss with progression to osteoarthritis is common in humans. In vitro work suggests that meniscectomy causes increased joint contact stress, but what other alterations in dynamic joint actions actually occur remains unknown. In a sheep model, we tested the hypothesis that complete lateral meniscectomy increases joint abduction, shifting the in vivo locations of tibiofemoral contact to regions that qualitatively correspond to locations of chondral damage. Nine sheep underwent unilateral arthrotomy (n = 4) or arthrotomy plus complete lateral meniscectomy (n = 5). Kinematics were collected prior to surgery and serially up to 20 weeks post-surgery. Gross cartilage damage was mapped in each joint, graded using a published scoring scheme used in goats, and compared to the locations of minimum tibiofemoral distance. Over the 20 weeks, meniscectomy caused increased stifle abduction and medial tibial translation, shifting the points of minimum tibiofemoral distance 7.5 ± 2.1 mm laterally and 3.3 ± 1.1 mm anteriorly (mean ± SEM), which corresponded to the locations of focal chondral damage. Locations of new tibiofemoral contact in the meniscectomized compartment qualitatively correspond to subject-specific locations of early chondral damage in an ovine model.

Hydrostatic pressure stimulation of human mesenchymal stem cells seeded on collagen-based artificial extracellular matrices.

Human mesenchymal stem cells (hMSCs) from bone marrow are considered a promising cell source for bone tissue engineering applications because of their ability to differentiate into cells of the osteoblastic lineage. Mechanical stimulation is able to promote osteogenic differentiation of hMSC; however, the use of hydrostatic pressure (HP) has not been well studied. Artificial extracellular matrices containing collagen and chondroitin sulfate (CS) have promoted the expression of an osteoblastic phenotype by hMSCs. However, there has been little research into the combined effects of biochemical stimulation by matrices and simultaneous mechanical stimulation. In this study, artificial extracellular matrices generated from collagen and/or CS were coated onto polycaprolactone-co-lactide substrates, seeded with hMSCs and subjected to cyclic HP at various time points during 21 days after cell seeding to investigate the effects of biochemical, mechanical, and combined biochemical and mechanical stimulations. Cell differentiation was assessed by analyzing the expression of alkaline phosphatase (ALP) at the protein- and mRNA levels, as well as for calcium accumulation. The timing of HP stimulation affected hMSC proliferation and expression of ALP activity. HP stimulation after 6 days was most effective at promoting ALP activity. CS-containing matrices promoted the osteogenic differentiation of hMSCs. A combination of both CS-containing matrices and cyclic HP yields optimal effects on osteogenic differentiation of hMSCs on scaffolds compared with individual responses.