The CH25H-CYP7B1-RORα axis of cholesterol metabolism regulates osteoarthritis.

Osteoarthritis-the most common form of age-related degenerative whole-joint disease1-is primarily characterized by cartilage destruction, as well as by synovial inflammation, osteophyte formation and subchondral bone remodelling2,3. However, the molecular mechanisms that underlie the pathogenesis of osteoarthritis are largely unknown. Although osteoarthritis is currently considered to be associated with metabolic disorders, direct evidence for this is lacking, and the role of cholesterol metabolism in the pathogenesis of osteoarthritis has not been fully investigated4-6. Various types of cholesterol hydroxylases contribute to cholesterol metabolism in extrahepatic tissues by converting cellular cholesterol to circulating oxysterols, which regulate diverse biological processes7,8. Here we show that the CH25H-CYP7B1-RORα axis of cholesterol metabolism in chondrocytes is a crucial catabolic regulator of the pathogenesis of osteoarthritis. Osteoarthritic chondrocytes had increased levels of cholesterol because of enhanced uptake, upregulation of cholesterol hydroxylases (CH25H and CYP7B1) and increased production of oxysterol metabolites. Adenoviral overexpression of CH25H or CYP7B1 in mouse joint tissues caused experimental osteoarthritis, whereas knockout or knockdown of these hydroxylases abrogated the pathogenesis of osteoarthritis. Moreover, retinoic acid-related orphan receptor alpha (RORα) was found to mediate the induction of osteoarthritis by alterations in cholesterol metabolism. These results indicate that osteoarthritis is a disease associated with metabolic disorders and suggest that targeting the CH25H-CYP7B1-RORα axis of cholesterol metabolism may provide a therapeutic avenue for treating osteoarthritis.

Pathological changes and expression of lysine oxidases and matrix metalloproteinases -1, -2, and -3 in ligaments of patients with haemophilic arthritis.

INTRODUCTION: Studying the pathological changes of ligaments in patients with haemophilic arthritis (HA) has important significance for guiding the release of ligaments during total knee arthroplasty (TKA) and exploring interventions to prevent ligament lesions. AIM: This study was conducted to show the pathological changes and investigate the lysine oxidase (LOX) and matrix metalloproteinase (MMP)-1, -2, and -3 levels in the ligaments of patients with HA compared with those of patients with osteoarthritis (OA). METHODS: Ligaments obtained during the TKA were stained with Masson trichrome, Verhoeff-Van Gieson and haematoxylin and eosin to show the basic pathological changes. Collagen I, elastin, LOXs and MMP-1, -2, and -3 expression levels were detected via western blot. LOX and MMP-1, -2, and -3 mRNA expression levels were analysed via quantitative real-time PCR. RESULTS: Compared with OA ligaments, HA ligaments were constructed more loosely with wider gaps, more breaks, haemocytodeposition and local hypertrophy among the fibres. LOXs and MMP mRNA expression levels were upregulated in the HA tissues, which was consistent with the western blot results. Collagen I and elastin levels were also higher in patients with HA. CONCLUSIONS: The metabolism of the ligaments in patients with HA is more complex than in those with OA, and the ligaments of patients with HA have stronger healing and destruction processes. This pathology is related to iron overload and imbalanced inflammatory factors due to repeated intra-articular bleeding.

PM2.5 facilitates IL-6 production in human osteoarthritis synovial fibroblasts via ASK1 activation.

Osteoarthritis (OA) is a progressive degenerative joint disorder characterized by synovial inflammation. Interleukin-6 (IL-6) is a key proinflammatory cytokine in OA progression. Particulate matter 2.5 (PM2.5) exposure increases the risk of different diseases, including OA. Up until now, no studies have described any association between PM2.5 and IL-6 expression in human OA synovial fibroblasts (OASFs). Here, our data show that PM2.5 concentration- and time-dependently promoted IL-6 synthesis in human OASFs. We also found that reactive oxygen species (ROS) generation potentiated the effects of PM2.5 on IL-6 production. ASK1, ERK, p38, and JNK inhibitors reduced PM2.5-induced increases of IL-6 expression. Treatment of OASFs with PM2.5 promoted phosphorylation of these signaling cascades. We also found that PM2.5 enhanced c-Jun phosphorylation and its translocation into the nucleus. Thus, PM2.5 increases IL-6 production in human OASFs via the ROS, ASK1, ERK, p38, JNK, and AP-1 signaling pathways. Our evidence links PM2.5 with OA progression.

High-Temperature Requirement A1 Protease as a Rate-Limiting Factor in the Development of Osteoarthritis.

Preserving the mature articular cartilage of joints is a critical focus in the prevention and treatment of osteoarthritis. We determined whether the genetic inactivation of high-temperature requirement A1 (HtrA1) can significantly attenuate the degradation of articular or condylar cartilage. Two types of mouse models of osteoarthritis were used, a spontaneous mutant mouse model [type XI collagen-haploinsufficient (Col11a1+/-) mice] and two post-traumatic mouse models [destabilization of the medial meniscus (DMM) on the knee and a partial discectomy (PDE) on the temporomandibular joint]. Three different groups of mice were generated: i) HtrA1 was genetically deleted from Col11a1+/- mice (HtrA1-/-;Col11a1+/-), ii) HtrA1-deficient mice (HtrA1-/-) were subjected to DMM, and iii) HtrA1-/- mice were subjected to PDE. Knee and temporomandibular joints from the mice were characterized for evidence of cartilage degeneration. The degradation of articular or condylar cartilage was significantly delayed in HtrA1-/-;Col11a1+/- mice and HtrA1-/- mice after DMM or PDE. The amount of collagen type VI was significantly higher in the articular cartilage in HtrA1-/-;Col11a1+/- mice, compared with that in Col11a1+/- mice. The genetic removal of HtrA1 may delay the degradation of articular or condylar cartilage in mice.

ADAMTS-5: A difficult teenager turning 20.

A Disintegrin And Metalloproteinase with ThromboSpondin motif (ADAMTS)-5 was identified in 1999 as one of the enzymes responsible for cleaving aggrecan, the major proteoglycan in articular cartilage. Studies in vitro, ex vivo and in vivo have validated ADAMTS-5 as a target in osteoarthritis (OA), a disease characterized by extensive degradation of aggrecan. For this reason, it attracted the interest of many research groups aiming to develop a therapeutic treatment for OA patients. However, ADAMTS-5 proteoglycanase activity is not only involved in the dysregulated aggrecan proteolysis, which occurs in OA, but also in the physiological turnover of other related proteoglycans. In particular, versican, a major ADAMTS-5 substrate, plays an important structural role in heart and blood vessels and its proteolytic processing by ADAMTS-5 must be tightly regulated. On the occasion of the 20th anniversary of the discovery of ADAMTS-5, this review looks at the evidence for its detrimental role in OA, as well as its physiological turnover of cardiovascular proteoglycans. Moreover, the other potential functions of this enzyme are highlighted. Finally, challenges and emerging trends in ADAMTS-5 research are discussed.

Chamazulene reverses osteoarthritic inflammation through regulation of matrix metalloproteinases (MMPs) and NF-kß pathway in in-vitro and in-vivo models.

This study was conducted to evaluate the protective effects of chamazulene against IL-1ß-induced rat primary chondrocytes and complete Freund's adjuvant (CFA)-induced osteoarthritic inflammation in rats. Oxidative stress markers, pro-inflammatory cytokines, and regulatory proteins were measured. Chamazulene significantly reverted (p < 0.05) the levels of lipid peroxidation and enhanced the activities of superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) enzymes against IL-1ß and CFA-induced oxidative stress. The levels of TNF-α and IL-6 were reduced (p < 0.05) in chamazulene treatment against IL-1ß and CFA-induced inflammation. Western blot analysis results on the expressions of MMP-3, MMP-9, p65 NF-kß, iNOS, and COX-2 showed chamazulene was able to protect the chondrocytes against IL-1ß-induced osteoarthritic inflammation. Histopathology of rat hind ankle showed chamazulene significantly protected against CFA-induced osteoarthritic inflammation. Therefore, chamazulene can be recommended as a therapeutic agent for clinical trials against osteoarthritic inflammation.

Molecular Targets of Natural Products for Chondroprotection in Destructive Joint Diseases.

Osteoarthritis (OA) is the most common type of arthritis that occurs in an aged population. It affects any joints in the body and degenerates the articular cartilage and the subchondral bone. Despite the pathophysiology of OA being different, cartilage resorption is still a symbol of osteoarthritis. Matrix metalloproteinases (MMPs) are important proteolytic enzymes that degrade extra-cellular matrix proteins (ECM) in the body. MMPs contribute to the turnover of cartilage and its break down; their levels have increased in the joint tissues of OA patients. Application of chondroprotective drugs neutralize the activities of MMPs. Natural products derived from herbs and plants developed as traditional medicine have been paid attention to, due to their potential biological effects. The therapeutic value of natural products in OA has increased in reputation due to their clinical impact and insignificant side effects. Several MMPs inhibitor have been used as therapeutic drugs, for a long time. Recently, different types of compounds were reviewed for their biological activities. In this review, we summarize numerous natural products for the development of MMPs inhibitors in arthritic diseases and describe the major signaling targets that were involved for the treatments of these destructive joint diseases.

Exercise as an Adjuvant to Cartilage Regeneration Therapy.

This article provides a brief review of the pathophysiology of osteoarthritis and the ontogeny of chondrocytes and details how physical exercise improves the health of osteoarthritic joints and enhances the potential of autologous chondrocyte implants, matrix-induced autologous chondrocyte implants, and mesenchymal stem cell implants for the successful treatment of damaged articular cartilage and subchondral bone. In response to exercise, articular chondrocytes increase their production of glycosaminoglycans, bone morphogenic proteins, and anti-inflammatory cytokines and decrease their production of proinflammatory cytokines and matrix-degrading metalloproteinases. These changes are associated with improvements in cartilage organization and reductions in cartilage degeneration. Studies in humans indicate that exercise enhances joint recruitment of bone marrow-derived mesenchymal stem cells and upregulates their expression of osteogenic and chondrogenic genes, osteogenic microRNAs, and osteogenic growth factors. Rodent experiments demonstrate that exercise enhances the osteogenic potential of bone marrow-derived mesenchymal stem cells while diminishing their adipogenic potential, and that exercise done after stem cell implantation may benefit stem cell transplant viability. Physical exercise also exerts a beneficial effect on the skeletal system by decreasing immune cell production of osteoclastogenic cytokines interleukin-1ß, tumor necrosis factor-α, and interferon-γ, while increasing their production of antiosteoclastogenic cytokines interleukin-10 and transforming growth factor-ß. In conclusion, physical exercise done both by bone marrow-derived mesenchymal stem cell donors and recipients and by autologous chondrocyte donor recipients may improve the outcome of osteochondral regeneration therapy and improve skeletal health by downregulating osteoclastogenic cytokine production and upregulating antiosteoclastogenic cytokine production by circulating immune cells.

Circ_0136474 and MMP-13 suppressed cell proliferation by competitive binding to miR-127-5p in osteoarthritis.

Osteoarthritis (OA) is a prevalent degenerative joint disease whose pathogenesis remains unclear. The research aims to investigate the roles of Circ_0136474/miR-127-5p/MMP-13 axis in OA. Differentially expressed circRNAs and miRNAs in OA cartilage tissue were screened out and visualized by R project based on RNA-seq data and microarray data respectively. qRT-PCR was carried out for detection of relative expression levels of Circ_0136474, miR-127-5p, MMP-13 and other inflammatory factors and Western blot analysis was conducted to detect the protein expression level of MMP-13. CCK-8 assay and flow cytometry were conducted to determine cell proliferation and cell apoptotic ability respectively. RNA-fluorescence in situ hybridization (RNA-FISH) experiments were conducted to confirm the immune-localization of the Circ_0136474 and MMP-13 in human tissues. Targeted relationships were predicted by bioinformatic analysis and verified by dual-luciferase reporter assay. Our findings revealed that the expression levels of both Circ_0136474 and MMP-13 in OA cartilage tissue were significantly higher than that in normal cartilage tissue. Circ_0136474 could suppress cell proliferation by facilitating MMP-13 expression and suppressing miR-127-5p expression in OA. Overexpression of miR-127-5p negatively regulated MMP-13 expression to enhance cell proliferation. Our study demonstrated that Circ_0136474 and MMP-13 suppressed cell proliferation, while enhanced cell apoptosis by competitive binding to miR-127-5p in OA, which may well provide us with a new therapeutic strategy for osteoarthritis.

microRNA-186 inhibition of PI3K-AKT pathway via SPP1 inhibits chondrocyte apoptosis in mice with osteoarthritis.

Chondrocyte apoptosis has been implicated as a major pathological osteoarthritis (OA) change in humans and experimental animals. We evaluate the ability of miR-186 on chondrocyte apoptosis and proliferation in OA and elucidate the underlying mechanism concerning the regulation of miR-186 in OA. Gene expression microarray analysis was performed to screen differentially expressed messenger RNAs (mRNAs) in OA. To validate the effect of miR-186 on chondrocyte apoptosis, we upregulated or downregulated endogenous miR-186 using mimics or inhibitors. Next, to better understand the regulatory mechanism for miR-186 governing SPP1, we suppressed the endogenous expression of SPP1 by small interfering RNA (siRNA) against SPP1 in chondrocytes. We identified SPP1 is highly expressed in OA according to an mRNA microarray data set GSE82107. After intra-articular injection of papain into mice, the miR-186 is downregulated while the SPP1 is reciprocal, with dysregulated PI3K-AKT pathway in OA cartilages. Intriguingly, miR-186 was shown to increase chondrocyte survival, facilitate cell cycle entry in OA chondrocytes, and inhibit chondrocyte apoptosis in vitro by modulation of pro- and antiapoptotic factors. The determination of luciferase activity suggested that miR-186 negatively targets SPP1. Furthermore, we found that the effect of miR-186 suppression on OA chondrocytes was lost when SPP1 was suppressed by siRNA, suggesting that miR-186 affected chondrocytes by targeting and depleting SPP1, a regulator of PI3K-AKT pathway. Our findings reveal a novel mechanism by which miR-186 inhibits chondrocyte apoptosis in OA by interacting with SPP1 and regulating PI3K-AKT pathway. Restoring miR-186 might be a future therapeutic strategy for OA.

Ubiquitin-specific protease 3 targets TRAF6 for deubiquitination and suppresses IL-1ß induced chondrocyte apoptosis.

Traditionally, the development of osteoarthritis (OA) is associated with factors such as aging and injure, but more and more epidemiological and biological evidence suggests that the disease is closely related to metabolic syndrome and metabolic components. Ubiquitin-specific protease 3(USP3), a member of the USPs family, is a specific protease capable of cleavage of ubiquitin chains linked by proline residues. In our presented study, we firstly found that USP3 expression level was decreased in OA. USP3 overexpression inhibited IL-1ß induced chondrocytes apoptosis and nuclear factor κB (NF-κB) activation. USP3 knockdown induced chondrocytes apoptosis and activated NF-κB pathway. USP3 interacts with TRAF6 (tumor necrosis factor-receptor-associated factor 6), which is an essential adaptor protein for the NF-κB (nuclear factor κB) signaling pathway and plays important roles in inflammation and immune response. IL-1ß treatment up-regulated the polyubiquitination of TRAF6 in chondrocytes, which was attenuated when USP3 was forced expression. Our study mechanistically links USP3 to TRAF6 in osteoarthritis development. Moreover, these data support the pursuit of USP3 and TRAF6 as potential targets for osteoarthritis therapies.

Critical role for arginase II in osteoarthritis pathogenesis.

OBJECTIVE: Osteoarthritis (OA) appears to be associated with various metabolic disorders, but the potential contribution of amino acid metabolism to OA pathogenesis has not been clearly elucidated. Here, we explored whether alterations in the amino acid metabolism of chondrocytes could regulate OA pathogenesis. METHODS: Expression profiles of amino acid metabolism-regulating genes in primary-culture passage 0 mouse chondrocytes were examined by microarray analysis, and selected genes were further characterised in mouse OA chondrocytes and OA cartilage of human and mouse models. Experimental OA in mice was induced by destabilisation of the medial meniscus (DMM) or intra-articular (IA) injection of adenoviruses expressing catabolic regulators. The functional consequences of arginase II (Arg-II) were examined in Arg2-/- mice and those subjected to IA injection of an adenovirus encoding Arg-II (Ad-Arg-II). RESULTS: The gene encoding Arg-II, an arginine-metabolising enzyme, was specifically upregulated in chondrocytes under various pathological conditions and in OA cartilage from human patients with OA and various mouse models. Adenovirus-mediated overexpression of Arg-II in mouse joint tissues caused OA pathogenesis, whereas genetic ablation of Arg2 in mice (Arg2-/-) abolished all manifestations of DMM-induced OA. Mechanistically, Arg-II appears to cause OA cartilage destruction at least partly by upregulating the expression of matrix-degrading enzymes (matrix metalloproteinase 3 [MMP3] and MMP13) in chondrocytes via the nuclear factor (NF)-κB pathway. CONCLUSIONS: Our results indicate that Arg-II is a crucial regulator of OA pathogenesis in mice. Although chondrocytes of human and mouse do not identically, but similarly, respond to Arg-II, our results suggest that Arg-II could be a therapeutic target of OA pathogenesis.

The "elusive DMOAD": Aggrecanase inhibition from laboratory to clinic.

From the time of their discovery in 1999, the aggrecanases, and ADAMTS-5 in particular, have been heavily investigated as targets for disease-modifying osteoarthritis drug (DMOAD) development. Here, we provide a brief narrative review of the discovery efforts to target these enzymes, and how this led to the current ongoing programmes that hold promise for the future. We discuss a comparison of inhibition of collagen breakdown versus inhibition of aggrecan breakdown. We then summarise existing programmes that target ADAMTS-5, including small molecule inhibitors, monoclonal neutralising antibodies and nanobodies, and gene editing technologies. We also briefly discuss the potential analgesic effects this strategy may offer in addition to its joint-protective effects.

Effect of luteolin on apoptosis, MAPK and JNK signaling pathways in guinea pig chondrocyte with osteoarthritis.

Osteoarthritis (OA) is a degenerative joint disease usually seen in the elderly, which incidence increases with age. Its pathogenesis and underlying mechanism are still unclear. The disease severely affects the physical health and life quality of patients, thereby constituting a huge economic burden to family and society. Luteolin (LUT) is a natural flavonoid with multiple pharmacological properties. Many plants containing LUT have been applied in the treatment of several inflammation-related diseases due the relatively strong anti-inflammatory effects of LUT. The present study investigated the influence of LUT on cell apoptosis and inflammatory reactions in cartilage of OA guinea pigs, and its underlying mechanism. It was found that LUT effectively inhibited proliferation of OA cartilage cells, down-regulated the expressions of JNK and p38MAPK in cartilage cells of OA, and downregulated NO, TNF-α and IL-6. Thus, it alleviated inflammatory reactions, protected cartilage cells, and delayed cartilage degeneration.

Hexokinase 2 as a novel selective metabolic target for rheumatoid arthritis.

OBJECTIVES: Recent studies indicate that glucose metabolism is altered in rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS). Hexokinases (HKs) catalyse the first step in glucose metabolism, and HK2 constitutes the principal HK inducible isoform. We hypothesise that HK2 contributes to the synovial lining hypertrophy and plays a critical role in bone and cartilage damage. METHODS: HK1 and HK2 expression were determined in RA and osteoarthritis (OA) synovial tissue by immunohistochemistry. RA FLS were transfected with either HK1 or HK2 siRNA, or infected with either adenovirus (ad)-GFP, ad-HK1 or ad-HK2. FLS migration and invasion were assessed. To study the role of HK2 in vivo, 108 particles of ad-HK2 or ad-GFP were injected into the knee of wild-type mice. K/BxN serum transfer arthritis was induced in HK2F/F mice harbouring Col1a1-Cre (HK2Col1), to delete HK2 in non-haematopoietic cells. RESULTS: HK2 is particular of RA histopathology (9/9 RA; 1/8 OA) and colocalises with FLS markers. Silencing HK2 in RA FLS resulted in a less invasive and migratory phenotype. Consistently, overexpression of HK2 resulted in an increased ability to migrate and invade. It also increased extracellular lactate production. Intra-articular injection of ad-HK2 in normal knees dramatically increased synovial lining thickness, FLS activation and proliferation. HK2 was highly expressed in the synovial lining after K/BxN serum transfer arthritis. HK2Col1 mice significantly showed decreased arthritis severity, bone and cartilage damage. CONCLUSION: HK2 is specifically expressed in RA synovial lining and regulates FLS aggressive functions. HK2 might be an attractive selective metabolic target safer than global glycolysis for RA treatment.

Tyrosine kinase Fyn promotes osteoarthritis by activating the ß-catenin pathway.

OBJECTIVES: To investigate the role of tyrosine kinase Fyn in the development of osteoarthritis (OA) and the underlying mechanisms, and to define whether targeting Fyn could prevent OA in mice. METHODS: Cartilage samples from normal and aged mice were analysed with proteome-wide screening. Fyn expression was examined with immunofluorescence in human and age-dependent or experimental mouse OA cartilage samples. Experimental OA in Fyn-knockout mice was induced by destabilisation of the medial meniscus. Primary cultured mouse chondrocytes were treated with proinflammatory cytokine interleukin-1ß. The inhibitor of Src kinase family, AZD0530 (saracatinib), and inhibitor of Fyn, PP1, were used to treat experimental OA in mice. RESULTS: Fyn expression was markedly upregulated in human OA cartilage and in cartilage from aged mice and those with post-traumatic OA. Fyn accumulates in articular chondrocytes and interacts directly with and phosphorylates ß-catenin at Tyr142, which stabilises ß-catenin and promotes its nuclear translocation. The deletion of Fyn effectively delayed the development of post-traumatic and age-dependent OA in mice. Fyn inhibitors AZD0530 and PP1 significantly attenuated OA progression by blocking the ß-catenin pathway and reducing the levels of extracellular matrix catabolic enzymes in the articular cartilage. CONCLUSIONS: Fyn accumulates and activates ß-catenin signalling in chondrocytes, accelerating the degradation of the articular cartilage and OA development. Targeting Fyn is a novel and potentially therapeutic approach to the treatment of OA.

Heparanase is expressed in adult human osteoarthritic cartilage and drives catabolic responses in primary chondrocytes.

OBJECTIVES: The chondrocytes' pericellular matrix acts as a mechanosensor by sequestering growth factors that are bound to heparan sulfate (HS) proteoglycans. Heparanase is the sole mammalian enzyme with HS degrading endoglycosidase activity. Here, we aimed to ascertain whether heparanase plays a role in modulating the anabolic or catabolic responses of human articular chondrocytes. METHODS: Primary chondrocytes were incubated with pro-heparanase and catabolic and anabolic gene expression was analyzed by quantitative polymerase chain reaction (PCR). MMP13 enzymatic activity in the culture medium was measured with a specific fluorescent assay. Extracellular regulated kinase (ERK) phosphorylation was evaluated by Western blot. Human osteoarthritis (OA) cartilage was assessed for heparanase expression by reverse-transcriptase PCR, by Western blot and by a heparanase enzymatic activity assay. RESULTS: Cultured chondrocytes rapidly associated with and activated pro-heparanase. Heparanase induced the catabolic genes MMP13 and ADAMTS4 and the secretion of active MMP13, and down-regulated the anabolic genes ACAN and COL2A1. PG545, a HS-mimetic, inhibited the effects of heparanase. Heparanase expression and enzymatic activity were demonstrated in adult human osteoarthritic cartilage. Heparanase induced ERK phosphorylation in cultured chondrocytes and this could be inhibited by PG545, by fibroblast growth factor 2 (FGF2) neutralizing antibodies and by a FGF-receptor inhibitor. CONCLUSIONS: Heparanase is active in osteoarthritic cartilage and induces catabolic responses in primary human chondrocytes. This response is due, at least in part, to the release of soluble growth factors such as FGF2.

Transcription factor Hes1 modulates osteoarthritis development in cooperation with calcium/calmodulin-dependent protein kinase 2.

Notch signaling modulates skeletal formation and pathogenesis of osteoarthritis (OA) through induction of catabolic factors. Here we examined roles of Hes1, a transcription factor and important target of Notch signaling, in these processes. SRY-box containing gene 9 (Sox9)-Cre mice were mated with Hes1(fl/fl) mice to generate tissue-specific deletion of Hes1 from chondroprogenitor cells; this deletion caused no obvious abnormality in the perinatal period. Notably, OA development was suppressed when Hes1 was deleted from articular cartilage after skeletal growth in type II collagen (Col2a1)-Cre(ERT);Hes1(fl/fl) mice. In cultured chondrocytes, Hes1 induced metallopeptidase with thrombospondin type 1 motif, 5 (Adamts5) and matrix metalloproteinase-13 (Mmp13), which are catabolic enzymes that break down cartilage matrix. ChIP-seq and luciferase assays identified Hes1-responsive regions in intronic sites of both genes; the region in the ADAMTS5 gene contained a typical consensus sequence for Hes1 binding, whereas that in the MMP13 gene did not. Additionally, microarray analysis, together with the ChIP-seq, revealed novel Hes1 target genes, including Il6 and Il1rl1, coding a receptor for IL-33. We further identified calcium/calmodulin-dependent protein kinase 2δ (CaMK2δ) as a cofactor of Hes1; CaMK2δ was activated during OA development, formed a protein complex with Hes1, and switched it from a transcriptional repressor to a transcriptional activator to induce cartilage catabolic factors. Therefore, Hes1 cooperated with CaMK2δ to modulate OA pathogenesis through induction of catabolic factors, including Adamts5, Mmp13, Il6, and Il1rl1. Our findings have contributed to further understanding of the molecular pathophysiology of OA, and may provide the basis for development of novel treatments for joint disorders.

Can photobiomodulation associated with implantation of mesenchymal adipose-derived stem cells attenuate the expression of MMPs and decrease degradation of type II collagen in an experimental model of osteoarthritis?

This study aimed to determine whether photobiomodulation therapy (PBMT) could improve the bioavailability and chondroprotective benefits of mesenchymal stem cells injected into the knees of rats used as an experimental model of osteoarthritis (OA) as well as reduce the expression of matrix metalloproteinases (MMPs) and degradation of type II collagen (COL2-1) in the cartilage. Adipose-derived stem/stromal cells (ADSCs) were collected from three male Fischer 344 rats and characterized by flow cytometry. Fifty female Fischer 344 rats were distributed into five groups of 10 animals each. These groups were as follows: control, OA, OA PBMT, OA ADSC, and OA ADSC PBMT. OA was induced in the animals using a 4% papain solution. Animals from the OA ADSC and OA ADSC PBMT groups received an intra-articular injection of 10 × 106 ADSCs and were treated with PBMT by irradiation (wavelength: 808 nm, power: 50 mW, energy: 42 J, energy density: 71.2 J/cm2, spot size: 0.028). Euthanasia was performed 7 days after the first treatment. The use of PBMT alone and the injection of ADSCs resulted in downregulation of pro-inflammatory cytokines and MPs in cartilage compared to the OA group. PBMT and ADSCs caused upregulation of tissue inhibitors of MPs 1 and 2 and mRNA and protein expression of COL2-1 in cartilage compared to the OA group. The intra-articular injection of ADSCs and PBMT prevented joint degeneration resulting from COL2-1 degradation and modulated inflammation by downregulating cytokines and MMPs in the OA group.

Mithramycin A Alleviates Osteoarthritic Cartilage Destruction by Inhibiting HIF-2α Expression.

Osteoarthritis (OA) is the most common and increasing joint disease worldwide. Current treatment for OA is limited to control of symptoms. The purpose of this study was to determine the effect of specificity protein 1 (SP1) inhibitor Mithramycin A (MitA) on chondrocyte catabolism and OA pathogenesis and to explore the underlying molecular mechanisms involving SP1 and other key factors that are critical for OA. Here, we show that MitA markedly inhibited expressions of matrix-degrading enzymes induced by pro-inflammatory cytokine interleukin-1β (IL-1β) in mouse primary chondrocytes. Intra-articular injection of MitA into mouse knee joint alleviated OA cartilage destruction induced by surgical destabilization of the medial meniscus (DMM). However, modulation of SP1 level in chondrocyte and mouse cartilage did not alter catabolic gene expression or cartilage integrity, respectively. Instead, MitA significantly impaired the expression of HIF-2α known to be critical for OA pathogenesis. Such reduction in expression of HIF-2α by MitA was caused by inhibition of NF-κB activation, at least in part. These results suggest that MitA can alleviate OA pathogenesis by suppressing NF-κB-HIF-2α pathway, thus providing insight into therapeutic strategy for OA.