Rosmarinic acid down-regulates NO and PGE2 expression via MAPK pathway in rat chondrocytes.

Rosmarinic acid (RosA) is a water-soluble polyphenol, which can be isolated from many herbs such as orthosiphon diffuses and rosmarinus officinalis. Previous studies have shown that RosA possesses various biological properties. In this study, we investigate the anti-osteoarthritic effects of RosA in rat articular chondrocytes. Chondrocytes were pre-treated with RosA, followed by the stimulation of IL-1ß. Real-time PCR and Western blot were performed to detect the expression of matrix metalloproteinase (MMP)-1, MMP-3 and MMP-13. Nitric oxide and PGE2 production were measured by Griess reagent and enzyme-linked immunosorbent assay (ELISA). The expression of mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) was also investigated by Western blot analysis. We found that RosA down-regulated the MMPs expression as well as nitric oxide and PGE2 production in IL-1ß-induced chondrocytes. In addition, RosA inhibited p38 and JNK phosphorylation as well as p65 translocation. The results suggest that RosA may be considered a possible agent in the treatment of OA.

Acacetin inhibits expression of matrix metalloproteinases via a MAPK-dependent mechanism in fibroblast-like synoviocytes.

It is well known that rheumatoid arthritis (RA) is an autoimmune joint disease in which fibroblast-like synoviocytes (FLSs) play a pivotal role. In this study, we investigated the anti-arthritic properties of acacetin in FLSs. The expression of matrix metalloproteinase (MMP)-1, MMP-3 and MMP-13 were investigated by quantitative RT-PCR and western blot at gene and protein levels. At the same time, the phosphorylation of mitogen-activated protein kinases (MAPK) was investigated. The DNA-binding activity of NF-κB was investigated by electrophoretic mobility shift assay. We found that acacetin inhibits p38 and JNK phosphorylation and reduces MMP-1, MMP-3 and MMP-13 expression in interleukin-1ß-induced FLSs. Our results suggest that acacetin has antiarthritic effects in FLSs. Thus, acacetin should be further studied for the treatment of arthritis.

Baicalein Inhibits MMPs Expression via a MAPK-Dependent Mechanism in Chondrocytes.

BACKGROUND: Baicalein is a flavonoid isolated from Scutellaria baicalensis Georgi. Here, we investigated the anti-osteoarthritic effect of baicalein in vitro and in vivo. METHODS: Interleukin-1 beta (IL-1ß)-induced chondrocytes were treated with different concentrations of baicalein, real-time PCR and ELISA were performed to detect the matrix metalloproteinases (MMPs) expression. Western blot was used to evaluate the mitogen-activated protein kinase (MAPK) expression. In experimental osteoarthritis (OA), rabbits were treated with baicalein, gross morphological and histological assessment was performed to evaluate the cartilage damage. RESULTS: Baicalein significantly reduced the expression of MMPs in vitro and in vivo. Moreover, baicalein significantly reduced the phosphorylation of p38 and extracellular signal regulated kinase (ERK), but not of c-Jun N-terminal kinase (JNK). In addition, intra-articular injection of baicalein ameliorated the cartilage damage in a rabbit model of OA induced by anterior cruciate ligament transection (ACLT). CONCLUSIONS: The results indicate that baicalein may be considered as a potential agent for OA treatment.

The disease-modifying effect of dehydroepiandrosterone in different stages of experimentally induced osteoarthritis: a histomorphometric study.

BACKGROUND: Osteoarthritis (OA) is likely to become an increasing burden in the coming decades. Various agents have been developed to slow the progression of OA, and are collectively known as 'disease-modifying drugs', however, there is still little reliable evidence that such agents will be successful. Dehydroepiandrosterone (DHEA), a sex hormone precursor, has been recently proven as protective agent against OA, but the exact mechanism is still unkown. In the current study, the effects of weekly intra-articular injections of DHEA in preventing the progression of existing cartilage degeneration in an OA rabbit model were evaluated. The aim of the current study is to demonstrate the feature of its disease-modifying efficacy during OA progression. METHODS: Thirty male New Zealand white rabbits were used in this study. An anterior cruciate ligament transection (ACLT) model was used to create a progressive OA model in twenty rabbits. The animals were treated with DHEA or a placebo and were necropsied at 9 and 16 weeks. Ten rabbits receiving sham operations served as controls. The articular cartilage of the medial femoral condyle (MFC), lateral femoral condyle (LFC), medial tibial plateau (MTP) and lateral tibial plateau (LTP) was evaluated macroscopically and histologically. RESULTS: In the joints of the sham-operated rabbits, few histological changes were detected on the articular surfaces of the femoral condyles and tibial plateaus. ACLT obviously induced erosive changes on the cartilage surfaces. Compared to the placebo group, the macroscopic and Mankin score analyses demonstrated that the DHEA treatment markedly reduced the cartilage lesions and delayed cartilage degeneration in the four regions of the knee at 9 weeks after operation (macroscopic score: MFC P = 0.013; LFC P = 0.048; MTP P = 0.045; LTP P = 0.02, Mankin score: MFC P = 0.012; LFC P = 0.034; MTP P = 0.016; LTP P = 0.002). At 16 weeks, DHEA demonstrated chondroprotective effects on the lateral compartment of the knee compared to the placebo group, whereas the cartilage degeneration at the medial compartment of the knee did not differ among the groups (macroscopic score: LFC P = 0.046; LTP = 0.034, Mankin score: LFC P = 0.005; LTP P = 0.002). CONCLUSION: The disease-modifying efficacy of DHEA aganist OA is time-specific and site-dependent. DHEA could be used as a disease-modifying strategy to limit the progression of OA, especially in the middle stage.

Lubricin: a novel potential biotherapeutic approaches for the treatment of osteoarthritis.

Osteoarthritis (OA) is a multi-factor disorder of sinovial joints, which characterized by escalated degeneration and loss of articular cartilage. Treatment of OA is a critical unmet need in medicine for regeneration of damaged articular cartilage in elderly. On the other hand, lubricin, a glycoprotein specifically synthesized by chondrocytes located at the surface of articular cartilage, has been shown to provide boundary lubrication of congruent articular surfaces under conditions of high contact pressure and near zero sliding speed. Lubrication of these surfaces is critical to normal joint function, while different gene expressions of lubricin had been found in the synovium of rheumatoid arthritis (RA) and OA. Moreover, mutations or lacking of lubricin gene have been shown to link to the joint disease such as camptodactyly-arthropathy-coxa vara-pericarditis syndrome (CACP), synovial hyperplasia and failure of joint function, suggesting an important role of lubricin in the pathogenesis of these joint disease. Recent studies demonstrate that administration with recombinant lubricin in the joint cavity would be effective in the prevention of cartilage degeneration in animal OA models. Therefore, a treatment with lubricin which would protect cartilage in vivo would be desirable. This article reviews recent findings with regard to the possible role of lubricin in the progression of OA, and further discusses lubricin as a novel potential biotherapeutic approaches for the treatment of OA.

The emerging role of adipokines in osteoarthritis: a narrative review.

Osteoarthritis (OA) is a most common multifactorial degenerative joint disease in elderly individuals. OA is affecting severely the quality of life of patients, while the causes of OA are not completely understood. Age, obesity, the female sex, and previous injury are considered as significant risk factors. Recently, increased levels of adipokines which are mainly produced by adipocytes have been detected in patients with osteoarthritis. Moreover, studies on different adipokines all reveal that they have played proinflammatory and catabolic/anabolic roles during the pathophysiology of OA. In the present review, we summarize current data on the effect of the adipose tissue-derived hormones leptin, adiponectin, resistin and visfatin on initiation and progression of OA.

Effect of dehydroepiandrosterone on aggrecanase expression in articular cartilage in a rabbit model of osteoarthritis.

To investigate the in vivo effect of dehydroepiandrosterone (DHEA) on the expression of aggrecanases and their endogenous inhibitor in a rabbit model of OA. Ten New Zealand white rabbits underwent bilateral anterior cruciate ligament transection (ACLT). One knee of each rabbit was randomly assigned to receive 100 µM DHEA dissolved in dimethylsulphoxide (DMSO) and the other was treated with DMSO only. The treatment was given once a week for 5 weeks, starting 4 weeks after transection. All rabbits were euthanized 9 weeks after ACLT treatment, and the knee joints were evaluated by gene expression analysis. Intra-articular administration of DHEA significantly reduced the gene expression of aggrecanases, while markly increasing that of tissue inhibitor of metalloproteinase-3 (TIMP-3), an endogenous inhibitor of aggrecanases. DHEA may have beneficial effects on OA by influencing the balance between aggrecanases and TIMP-3 through which DHEA may protect against OA.

Trichostatin A inhibits expression of cathepsins in experimental osteoarthritis.

The aim of this study was to investigate the effects of trichostatin A (TSA) on expression of cathepsins in cartilage in experimental osteoarthritis (OA). OA was induced in 18 rabbits by bilateral anterior cruciate ligament transection (ACLT). Four weeks after surgery, rabbits received intra-articular injection with TSA dissolved in the dimethylsulphoxide (DMSO) in the right knees and DMSO in the left knees once a week for 5 weeks. Rabbits were killed 7 days after the last injection. The knee joints were assessed by morphological and histological examination. Messenger RNA expression of cathepsins K, B, L, S and cystatin C was studied by real-time PCR. TSA inhibited the expression of cathepsins K, B, L, S and cystatin C accompanied with the less degradation in cartilage. The results suggest that TSA exhibits protective effects against cartilage degradation in rabbits with OA and the effects may be associated with the inhibition of cathepsins.

Protective effects of berberine in an experimental rat osteoarthritis model.

Berberine shows anticancer, antibacterial, antiinflammatory and antioxidant effects and may be useful in many clinical applications. The effects of berberine on articular cartilage metabolism remain unknown, so this study was performed to evaluate these effects in vitro and in vivo. For the in vitro work, rat articular chondrocytes were cultured in a monolayer and matrix metalloproteinase-1 (MMP-1), -3, -13 and tissue inhibitor of metalloproteinase (TIMP-1) expression was evaluated by real-time quantitative PCR. Nitric oxide (NO) levels were determined using the Griess reaction, and glycosaminoglycan (GAG) release was measured using the dimethylmethylene blue method. For the in vivo work, berberine was administered by intraarticular injection, and the effects on MMPs and TIMP-1 were examined at the gene and protein levels. Berberine was found to inhibit the expression of MMP-1, -3 and -13, and increased the level of TIMP-1 at the mRNA level in a dose-dependent manner. In IL-1ß-induced rat articular chondrocytes, berberine decreased IL-1ß-induced GAG release and NO production. Meanwhile, high-dose berberine exhibited an anticatabolic effect in an IL-1ß-induced rat osteoarthritis (OA) model. These findings suggest that berberine may play a protective role in the development of OA and may be useful in the treatment of OA.

Effects of diallyl sulphide in chondrocyte and cartilage in experimental osteoarthritis in rabbit.

Diallyl sulphide (DAS) is known for its antioxidant, anticancer and detoxifying properties. The aim of this study was to investigate the effect of DAS on rabbit articular chondrocytes and cartilage in experimental osteoarthritis (OA) induced by anterior cruciate ligament transection (ACLT). DAS inhibited matrix metalloproteinase-1 (MMP-1), MMP-3 and MMP-13 expression in interleukin-1beta (IL-1ß)-induced chondrocytes. In an in vivo study, DAS ameliorated cartilage degradation as assessed by morphological and histological examination. Messenger RNA expression of MMP-1, MMP-3, MMP-13 and IL-1ß was inhibited by DAS in cartilage. In addition, DAS increased the collagen II level in cartilage. The results suggest that DAS may protect cartilage in the development of OA.

Increased apelin serum levels and expression in human chondrocytes in osteoarthritic patients.

Apelin is a recently discovered hormone secreted by adipocytes. The aim of this study, therefore, was to evaluate the distribution of apelin in paired serum and synovial fluid (SF) of osteoarthritis (OA) patients, as compared to that in healthy controls, and to characterise the expression profile of apelin and its cognate receptor APJ in human chondrocytes. Apelin levels in serum and SF were analysed by enzyme-linked immunosorbent assay (ELISA). Expression of apelin and APJ in human chondrocytes was determined by real-time quantitative polymerase chain reaction (PCR). Apelin was found to be present in OA SF, and concentrations were positively correlated with the severity of OA. OA serum exhibited significantly elevated levels of apelin (2.18 ± 0.22 ng/ml) as compared to normal serum (1.31 ± 0.12 ng/ml) (p < 0.05), and serum apelin levels exceeded those in paired SF (p < 0.001). The apelin and APJ transcripts were identified in chondrocytes, and levels were significantly higher in OA cartilage than in healthy donors. These findings suggest that apelin may contribute to the onset and/or progression of OA, and may provide new insights into the pathophysiology of OA.

Increased serum concentrations of visfatin and its production by different joint tissues in patients with osteoarthritis.

BACKGROUND: The goal of this study was to investigate the distribution of visfatin in paired serum and synovial fluid (SF) samples from patients with osteoarthritis (OA), and in serum from healthy controls. In addition, we wished to determine the potential sources of visfatin in joint tissue. METHODS: Twenty-three patients with OA requiring total knee arthroplasty (TKA), 17 healthy subjects and ten donors requiring amputation were included in the study. Concentrations of visfatin in serum and SF were analyzed using an enzyme-linked immunosorbent assay (ELISA). The concentration of visfatin was also measured in conditioned media from cultured joint tissues. RESULTS: We found serum visfatin concentrations to be higher in patients with OA compared to healthy controls (p<0.05), and SF-visfatin concentrations exceeded those in paired serum (p=0.004). In addition, infrapatellar fat pad and synovium were important sources of visfatin, while osteophytes released largest amounts of visfatin. Therefore, osteophytes can be considered a major source of visfatin in the knee joint in patients with OA. CONCLUSIONS: These results suggest that visfatin may be involved in the pathophysiology of OA and may have local effects in the joint during the process of OA.

Alleviation of osteoarthritis by Trichostatin A, a histone deacetylase inhibitor, in experimental osteoarthritis.

This study investigated the effects of the histone deacetylase (HDAC) inhibitor trichostatin A (TSA) on cartilage degradation in an experimental model of osteoarthritis (OA). Thirty-two male New Zealand rabbits underwent unilateral anterior cruciate ligament transection (ACLT) on left knee joints to induce OA and were randomly divided into two groups (n = 16), the TSA group was injected intra-articularly with 0.3 ml TSA [250 ng/ml in the dimethylsulphoxide (DMSO)], the OA group received DSMO since 4 weeks after operation once a week for 5 weeks. Rabbits were killed seven days after the last injection. Left knee cartilage was harvested for morphological, histological and genetic analysis. Another ten rabbits were used for normal control and received no injection. The TSA group showed less cartilage degradation as compared to the OA group assessed by morphological and histological evaluation. Gene expression of matrix metalloproteinase-1 (MMP-1), MMP-3, MMP-13, and interleukin-1 (IL-1) was increased significantly in the OA group compared to the normal group. The elevated expression was reduced by TSA. Our results suggest that TSA could be considered as a potential agent for treatment for OA.

Leptin plays a catabolic role on articular cartilage.

Leptin has been shown to play a crucial role in the regulation of body weight. There is also evidence that this adipokine plays a key role in the process of osteoarthritis. However, the precise role of leptin on articular cartilage metabolism is not clear. We investigate the role of leptin on articular cartilage in vivo in this study. Recombinant rat leptin (100 µg) was injected into the knee joints of rats, 48 h later, messenger RNA (mRNA) expression and protein levels of basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), matrix metalloproteinases 2 and 9 (MMP-2, MMP-9), cathepsin D, and collagen II from articular cartilage were analyzed by real-time quantitative polymerase chain reaction (PCR) and western blot. Two important aggrecanases ADAMTS-4 and -5 (a disintegrin and metalloproteinase with thrombospondin motifs 4 and 5) were also analyzed by real-time quantitative PCR. Besides, articular cartilage was also assessed for proteoglycan/GAG content by Safranin O staining. Leptin significantly increased both gene and protein levels of MMP-2, MMP-9, cathepsin D, and collagen II, while decreased bFGF markedly in cartilage. Moreover, the gene expression of ADAMTS-4 and -5 were markedly increased, and histologically assessed depletion of proteoglycan in articular cartilage was observed after treatment with leptin. These results strongly suggest that leptin plays a catabolic role on cartilage metabolism and may be a disadvantage factor involve in the pathological process of OA.

Dehydroepiandrosterone and age-related musculoskeletal diseases: Connections and therapeutic implications.

Musculoskeletal disorders related to ageing are one of the most common causes of mortality and morbidity among elderly individuals worldwide. The typical constitutive components of the musculoskeletal system, including bone, muscle, and joints, gradually undergo a process of tissue loss and degeneration as a result of life-long mechanical and biological stress, ultimately leading to the onset of a series of age-related musculoskeletal diseases, including osteoporosis (OP), sarcopenia, and osteoarthritis (OA). Dehydroepiandrosterone (DHEA), a precursor of androgen secreted mainly by the adrenal gland, has attracted much attention as a marker for senescence due to its unique age-related changes. This pre-hormone has been publicly regarded as an "antidote for ageing" because of its favourable effect against a wide range of age-related diseases, such as Alzheimer disease, cardiovascular diseases, immunosenescence and skin senescence, though its effect on age-related musculoskeletal diseases has been explored to a lesser extent. In the present review, we summarized the action of DHEA against OP, sarcopenia and OA. Extensive detailed descriptions of the pathogenesis of each of these musculoskeletal disorders are beyond the scope of this review; instead, we aim to highlight the association of changes in DHEA with the processes of OP, sarcopenia and OA. A special focus will also be placed on the overlapping pathogeneses among these three diseases, and the molecular mechanisms underlying the action of DHEA against these diseases are discussed or postulated.

Tricetin Protects Rat Chondrocytes against IL-1ß-Induced Inflammation and Apoptosis.

Tricetin is a well-studied flavonoid with a wide range of pharmacological activities in cancer and inflammation. However, the ability of tricetin to ameliorate the inflammation that occurs in osteoarthritis (OA) has not been determined. This study explored the effects of tricetin on interleukin- (IL-) 1ß-induced rat chondrocytes. Chondrocytes harvested from rat cartilage were incubated in vitro with tricetin in the presence of IL-1ß. The expression of matrix metalloproteinase- (MMP-) 1, MMP-3, MMP-13, nitric oxide (NO), prostaglandin E2 (PGE2), Bax, and Bcl-2 was evaluated by real-time-PCR, ELISA, Griess reaction, and western blotting. Caspase-3 activity in chondrocytes was determined using a caspase-3 activity assay and MAPK pathway activity by western blotting. Tricetin decreased the expression of MMP-1, MMP-3, and MMP-13 at both the gene and protein level in IL-1ß-induced rat chondrocytes. It also inhibited IL-1ß-induced NO and PGE2 production, by modulating inducible NO synthase and cyclooxygenase 2 gene expression. An antiapoptotic role of tricetin involving the Bax/Bcl-2/caspase-3 pathway was also determined. The chondroprotective effect of tricetin was shown to be partly related to the suppression of the MAPK signaling pathway. The results of this study demonstrate the chondroprotective role of tricetin, based on its anticatabolic, anti-inflammatory, and antiapoptotic effects in chondrocytes. The therapeutic potential of tricetin in OA patients should be explored in future studies.

Dehydroepiandrosterone and Experimental Osteoarthritis.

Despite an increased understanding of the pathogenesis of osteoarthritis (OA) and the availability of a number of drugs designed to ameliorate its symptoms, a successful disease-modifying therapy remains elusive. Recent lines of evidence suggest that dehydroepiandrosterone (DHEA), a 19-carbon steroid hormone classified as an adrenal androgen, exerts a chondroprotective effect in OA patients, and it has been proven to be an effective DMOAD candidate that slows OA progression. However, the exact mechanisms underlying its anti-OA effect is largely unknown. This review summarizes emerging observations from studies of cell biology, preclinical animal studies, and preliminary clinical trials and describes the findings of investigations on this topic to develop an initial blueprint of the mechanisms by which DHEA slows OA progression. Presently, studies on DMOADs are increasing in importance but have met limited success. Encouragingly, the current data on DHEA are promising and may prove that DHEA-based treatment is efficacious for preventing and slowing human OA progression.

Paeoniflorin inhibits IL-1ß-induced chondrocyte apoptosis by regulating the Bax/Bcl-2/caspase-3 signaling pathway.

Apoptosis serves a pivotal role in the pathogenesis of osteoarthritis (OA). Increasing evidence has demonstrated that paeoniflorin exerts key properties (including anticancer, anti-inflammation and neuroprotective) for clinical applications. However, the precise role of paeoniflorin in articular cartilage apoptosis remains unknown. The present study explored the effects and potential molecular mechanism of paeoniflorin on rat chondrocyte apoptosis. Rat articular chondrocytes were cultured in monolayers. The lactate dehydrogenase (LDH) release rate of cells was determined by an LDH release assay. Annexin V-fluorescein isothiocyanate and propidium iodide staining were performed to detect early and advanced apoptotic cells by flow cytometry. The activity of caspase-3 in chondrocytes was determined using a caspase-3 activity assay. The expression of B-cell lymphoma 2 (Bcl-2)/Bcl-2-associated X protein (Bax) was examined by reverse transcription­quantitative polymerase chain and western blotting. The present study also examined the protein kinase B (Akt) signaling pathway by western blotting. Treatment with 25 or 50 µM paeoniflorin markedly decreased the release of LDH and the ratio of apoptotic cells in interleukin (IL)-1ß-induced rat chondrocytes. Paeoniflorin treatment decreased the mRNA and protein levels of Bax, and increased the level of Bcl-2. Paeoniflorin also reduced the activity of caspase-3 in chondrocytes. Furthermore, paeoniflorin was determined to regulate the Akt signaling pathway by increasing Akt phosphorylation. Therefore, paeoniflorin may exert its protective effect by inhibiting apoptosis in IL-1ß-induced rat chondrocytes and thus, may be an effective agent in the prevention and treatment of OA.

Paeoniflorin inhibits IL-1ß-induced MMP secretion via the NF-κB pathway in chondrocytes.

Paeoniflorin serves important cellular roles, exerting anti-cancer, anti-inflammatory and anti-pulmonary fibrosis effects and possesses immune-modulatory properties. However, the exact role of paeoniflorin in the pathogenesis of osteoarthritis (OA) remains unclear. The aim of the present study was to investigate the effects of paeoniflorin on articular surfaces in vitro. Rat chondrocytes were cultured in vitro and an MTT assay was performed to assess chondrocyte survival. Following treatment with interleukin (IL)-1ß and paeoniflorin, the production of matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases-1 (TIMP-1) was examined using reverse transcription-quantitative polymerase chain reaction and western blotting. The interleukin (IL)-1ß-induced nuclear factor (NF)-κB pathway activation was also investigated. The results demonstrated that paeoniflorin was able to downregulate the expression of MMP and increase the expression of TIMP-1ntmRNA and protein in IL-1ß-induced rat chondrocytes. Furthermore, treating chondrocytes with paeoniflorin blocked the activation of NF-κB. These results suggest that paeoniflorin may serve am anti-catabolic role in the progression of OA and may be an effective preventative treatment for OA.

Vaspin prevents leptin­induced inflammation and catabolism by inhibiting the activation of nuclear factor­κB in rat chondrocytes.

The present study aimed to investigate the function and possible underlying mechanism of various concentrations of visceral adipose tissue­derived serine protease inhibitor (vaspin) on leptin­induced inflammatory and metabolic responses in rat chondrocytes. Rat articular chondrocytes were isolated and treated with different concentrations of vaspin, which was followed by stimulation with leptin. The expression of genes, secretion of nitric oxide and tumor necrosis factor­α, and activation of the nuclear factor (NF)­κB pathway were analyzed by reverse transcription­quantitative polymerase chain reaction, ELISA and western blotting. The results demonstrated that vaspin inhibited the leptin­induced upregulated gene expression levels of leptin receptor (OB­Rb), a disintegrin and metalloprotease with thrombospondin motifs (ADAMTS)­4, ADAMTS­5, matrix metalloproteinase (MMP)­2 and MMP­9, and the secretion of NO and TNF­α, in a dose­dependent manner. The phosphorylation of inhibitor of NF­κB (IκB), IκB kinase (IKK)α, IKKß and NF­κB were also promoted by leptin in the chondrocytes, which were also suppressed by increased concentration of vaspin. Taken together, results demonstrated that vaspin prevented leptin­induced inflammation and catabolism by inhibiting the activation of NF­κB in rat chondrocytes.