Osteoarthritis (OA) is the most common degenerative joint disease and a major cause of pain and disability in adult individuals. The etiology of OA includes joint injury, obesity, aging, and heredity. However, the detailed molecular mechanisms of OA initiation and progression remain poorly understood and, currently, there are no interventions available to restore degraded cartilage or decelerate disease progression. The diathrodial joint is a complicated organ and its function is to bear weight, perform physical activity and exhibit a joint-specific range of motion during movement. During OA development, the entire joint organ is affected, including articular cartilage, subchondral bone, synovial tissue and meniscus. A full understanding of the pathological mechanism of OA development relies on the discovery of the interplaying mechanisms among different OA symptoms, including articular cartilage degradation, osteophyte formation, subchondral sclerosis and synovial hyperplasia, and the signaling pathway(s) controlling these pathological processes.

IntroductionOsteoarthritis (OA) is a degenerative joint disease affecting a large population of people. The mechanism of this highly prevalent disease is not fully understood. Currently there is no effective disease-modifying treatment for OA. The purpose of this study was two-fold: 1) to investigate the role of MMP13 in the development of OA; and 2) to evaluate the efficacy of the MMP13 inhibitor CL82198 as a pharmacologic treatment for preventing OA progression.MethodsTo investigate the role of the endogenous Mmp13 gene in OA development, tamoxifen was administered to two-week-old Col2CreER;Mmp13fx/fx (Mmp13Col2ER) and Cre-negative control mice for five days. OA was induced by meniscal-ligamentous injury (MLI) when the mice were 10 weeks old and MLI or sham-operated joints were harvested 4, 8, 12, or 16 weeks after surgery. To evaluate the efficacy of CL82198, MLI surgery was performed on 10-week-old wild type mice. CL82198 or saline was administered to the mice daily beginning immediately after the surgery for up to 16 weeks. The joint tissues collected from both experiments were evaluated by cartilage grading, histology/histomorphometry, immunohistochemistry (IHC), and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. The ability of CL82198 to inhibit MMP13 activity in vitro was confirmed by ELISA.ResultsThe OA progression was decelerated in Mmp13Col2ER mice 8, 12, and 16 weeks post-surgery. Cartilage grading by blinded observers confirmed decreased articular cartilage degeneration in Mmp13Col2ER mice at 8, 12 and 16 weeks compared to Cre-negative mice. Histomorphometric analysis demonstrated that Mmp13Col2ER mice had a higher articular cartilage area and thickness at 12 and 16 weeks post-surgery compared to the control mice. Results of IHC revealed greater type II collagen and proteoglycan expression in Mmp13Col2ER mice. Chondrocyte apoptosis, as determined by TUNEL staining, was higher in control mice compared to Mmp13Col2ER mice. CL82198 inhibited MMP13 activity in conditioned media from vehicle (> 85%) or bone morphogenetic protein 2 (BMP2)-treated (> 90%) primary murine sternal chondrocytes. Intraperitoneal injection of CL82198 decelerated MLI-induced OA progression, increased type II collagen and proteoglycan levels, and inhibited chondrocyte apoptosis compared to saline treatment as determined by OA grading, histology, histomorphometry, IHC, and TUNEL staining, respectively.ConclusionsMmp13 is critical for OA progression and pharmacologic inhibition of MMP13 is an effective strategy to decelerate articular cartilage loss in a murine model of injury-induced knee OA.

While mechanical loading is known to be essential in maintaining tendon homeostasis, repetitive mechanical loading has also been implicated in the etiology of tendon overuse injuries. The purpose of this study was to determine whether cyclic mechanical stretching regulates inflammatory responses induced by interleukin-1β (IL-1β) treatment in human patellar tendon fibroblasts (HPTFs). HPTFs were grown in microgrooved silicone dishes, where they became elongated in shape and aligned with the microgrooves, which is similar to the shape and organization of tendon fibroblasts in vivo. Cyclic uniaxial stretching was then applied to silicone culture dishes with a 4% or 8% stretch at a stretching frequency of 0.5 Hz for a duration of 4 h in the presence or absence of 10 pM IL-1β treatment. Non-stretched cells in the presence or absence of IL-1β were used for controls, respectively. The expression of cyclooxygenase-2 (COX-2), matrix metalloproteinase-1 (MMP-1), and the production of prostaglandin E 2 (PGE 2 ) were measured. In the absence of stretching, it was found that 10 pM of IL-1β markedly induced higher levels of COX-2, MMP-1 gene expression, and PGE 2 production than non-treated cells. Furthermore, cells with 4% stretching decreased the COX-2 and MMP-1 gene expression and PGE 2 production that were stimulated by IL-1β, whereas cells with 8% stretching further increased these gene products and/or expression levels in addition to the effects of IL-1β stimulation. Thus, the results suggest that repetitive, small-magnitude stretching is antiinflammatory, whereas large-magnitude stretching is pro-inflammatory. Therefore, moderate exercise may be beneficial to reducing tendon inflammation.

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Mechanical loads placed on connective tissues alter gene expression in fibroblasts through mechanotransduction mechanisms by which cells convert mechanical signals into cellular biological events, such as gene expression of extracellular matrix components (e.g., collagen). This mechanical regulation of ECM gene expression affords maintenance of connective tissue homeostasis. However, mechanical loads can also interfere with homeostatic cellular gene expression and consequently cause the pathogenesis of connective tissue diseases such as tendinopathy and osteoarthritis. Therefore, the regulation of gene expression by mechanical loads is closely related to connective tissue physiology and pathology. This article reviews the effects of various mechanical loading conditions on gene regulation in fibroblasts and discusses several mechanotransduction mechanisms. Future research directions in mechanoregulation of gene expression are also suggested.

Increased vascular endothelial growth factor (VEGF) levels are associated with OA progression. Indeed, VEGF appears to be involved in OA specific pathologies including cartilage degeneration, osteophyte formation, subchondral bone cysts and sclerosis, synovitis, and pain. Moreover, a wide range of studies suggests that inhibition of VEGF signaling reduces OA progression. This review highlights both the potential significance of VEGF in OA pathology and pain as well as potential benefits of inhibition of VEGF and its receptors as an OA treatment. With the emergence of the clinical use of anti-VEGF therapy outside of OA, both as high dose systemic treatments and low dose local treatments, these particular therapies are now more widely understood. Currently, there is no established disease-modifying drug available for patients with OA, which warrants continued study of the inhibition of VEGF in OA, as standalone or adjuvant therapy.

Improving the effectiveness of adipose-tissue derived human mesenchymal stromal/stem cells (AMSCs) for skeletal therapies requires a detailed characterization of mechanisms supporting cell proliferation and multi-potency. We investigated the molecular phenotype of AMSCs that were either actively proliferating in platelet lysate or in a basal non-proliferative state. Flow cytometry combined with high-throughput RNA sequencing (RNASeq) and RT-qPCR analyses validate that AMSCs express classic mesenchymal cell surface markers (e.g., CD44, CD73/NT5E, CD90/THY1 and CD105/ENG). Expression of CD90 is selectively elevated at confluence. Self-renewing AMSCs express a standard cell cycle program that successively mediates DNA replication, chromatin packaging, cyto-architectural enlargement and mitotic division. Confluent AMSCs preferentially express genes involved in extracellular matrix (ECM) formation and cellular communication. For example, cell cycle-related biomarkers (e.g., cyclins E2 and B2, transcription factor E2F1) and histone-related genes (e.g., H4, HINFP, NPAT) are elevated in proliferating AMSCs, while ECM genes are strongly upregulated (>10 fold) in quiescent AMSCs. AMSCs also express pluripotency genes (e.g., POU5F1, NANOG, KLF4) and early mesenchymal markers (e.g., NES, ACTA2) consistent with their multipotent phenotype. Strikingly, AMSCs modulate expression of WNT signaling components and switch production of WNT ligands (from WNT5A/WNT5B/WNT7B to WNT2/WNT2B), while up-regulating WNT-related genes (WISP2, SFRP2 and SFRP4). Furthermore, post-proliferative AMSCs spontaneously express fibroblastic, osteogenic, chondrogenic and adipogenic biomarkers when maintained in confluent cultures. Our findings validate the biological properties of self-renewing and multi-potent AMSCs by providing high-resolution quality control data that support their clinical versatility.

Platelet-rich plasma (PRP) containing hepatocyte growth factor (HGF) and other growth factors are widely used in orthopaedic/sports medicine to repair injured tendons. While PRP treatment is reported to decrease pain in patients with tendon injury, the mechanism of this effect is not clear. Tendon pain is often associated with tendon inflammation, and HGF is known to protect tissues from inflammatory damages. Therefore, we hypothesized that HGF in PRP causes the anti-inflammatory effects. To test this hypothesis, we performed in vitro experiments on rabbit tendon cells and in vivo experiments on a mouse Achilles tendon injury model. We found that addition of PRP or HGF decreased gene expression of COX-1, COX-2, and mPGES-1, induced by the treatment of tendon cells in vitro with IL-1β. Further, the treatment of tendon cell cultures with HGF antibodies reduced the suppressive effects of PRP or HGF on IL-1β-induced COX-1, COX-2, and mPGES-1 gene expressions. Treatment with PRP or HGF almost completely blocked the cellular production of PGE2 and the expression of COX proteins. Finally, injection of PRP or HGF into wounded mouse Achilles tendons in vivo decreased PGE2 production in the tendinous tissues. Injection of platelet-poor plasma (PPP) however, did not reduce PGE2 levels in the wounded tendons, but the injection of HGF antibody inhibited the effects of PRP and HGF. Further, injection of PRP or HGF also decreased COX-1 and COX-2 proteins. These results indicate that PRP exerts anti-inflammatory effects on injured tendons through HGF. This study provides basic scientific evidence to support the use of PRP to treat injured tendons because PRP can reduce inflammation and thereby reduce the associated pain caused by high levels of PGE2.