| Objective:Osteoarthritis(OA)It belongs to the progressive degenerative joint disease with the highest incidence ratei。The clinical manifestations of osteoarthritis are mainly joint pain,localized pressure,joint stiffness and limitation of movement.The high disability and mortality rates of this disease have imposed a heavy health and economic burden on humans and society.Although research on the pathogenesis of OA has been a hot topic in the academic community,its pathogenesis is complex and involves many signaling pathways,which are still not fully understood.Currently,the treatment of OA is still limited to oral medication,physical exercise and surgery,and the only effective solution for end-stage OA is joint replacement surgery.Therefore,only by further exploring the pathogenesis of OA can we provide a solid theoretical basis for finding new specific therapeutic targets or developing new treatments.Deleted in malignant brain tumors(DMBT1)was first identified as an oncogene in malignant brain tumors,and is widely expressed in various tissues throughout the body,with the highest expression in the mucosal epithelium and related secretory glands of the respiratory and digestive systems.DMBT1 glycoprotein is widely expressed in various tissues throughout the body,with the highest expression in the mucosal epithelium and related secretory glands of the respiratory and digestive systems.The expression of DMBT1 expressed by cells in local tissues can be upregulated by stimulation of various pro-inflammatory factors.However,the relevance of DMBT1 to OA and the mechanism of DMBT1 in OA have not been reported.Lipid metabolism is now an important risk factor in the pathology of OA.Peroxisome proliferator-activated receptor α(PPARα)is a transcription factor that is important in the physiological process of maintaining lipid metabolism homeostasis in chondrocytes.However,there is still no report on whether DMBT1 has a direct regulatory relationship with PPARα.We intend to explore the potential role and molecular mechanism of DMBT1 in the pathological process of OA through this study,and provide new therapeutic ideas for clinical research on the treatment of OA.METHODS:We downloaded and analyzed publicly published transcriptomic data GSE114007 about human normal cartilage and OA cartilage samples to initially observe whether there were differences in DMBT1 expression levels between normal and OA cartilage.Immunohistochemistry,RT-qPCR and Western-blot techniques were applied to examine damaged cartilage samples and non-damaged cartilage samples obtained from OA patients to verify the results derived in the transcriptomic data.Cholesterol was extracted from human cartilage samples to compare the difference in cholesterol levels in damaged and undamaged cartilage.After it was clear that DMBT1 expression was elevated in OA,the changes in DMBT1 expression levels in SW1353 cells were detected by RTqPCR and Western-blot techniques at different times and concentrations of IL-1βstimulation.The expression levels of MMP13 and COL2A1 were detected by immunofluorescence technique in IL-1β-induced SW1353 cells to construct an in vitro OA model.After inhibition of DMBT1 using small interfering RNA(siRNA)in an IL-1βinduced inflammatory environment,the expression levels of matrix metalloproteinase 3(MMP3),MMP9,and COL2A1 in chondrocytes were detected by Western-blot.MMP13 and COL2A1 expression levels in chondrocytes by Western-blot,cell activity by CCK8 and chondrocyte apoptosis by loss cytometry,and cholesterol levels by special kits.After clarifying the above phenomena,siRNA was applied and the expression levels of PPARa and its target-regulated ACOX1 and CPT1A were detected by Western-blot and RT-qPCR.To explore the regulatory relationship between DMBT1 and PPARα,the expression levels of P38,ERK and JNK expression changes.Finally,in the IL-1β-induced inflammatory environment,DMBT 1 was knocked down while using agonists of PPARα,and the expression levels of COL2A1 and MMP13 were detected by Western-blot,apoptosis levels were detected by flow cytometry,and cholesterol levels were detected by a special kit.The expression levels of PPARα were detected by Western-blot using P38 MAPK inhibitor in DMBT1 knockdown cells to clarify the regulatory mechanism of DMBT1 on PPARαunder OA environment.To verify that knockdown of DMBT1 has a chondroprotective effect in the pathological process of OA,we detected MMP13 and COL2A1 expression levels and apoptosis levels in mouse knee joints using immunohistochemical staining and TUNEL after siRNA injection in a mouse OA model,and cholesterol levels in cartilage using a special kit.RESULTS:(1)Analysis of transcriptomic data and clinical cartilage tissue samples revealed that the expression level of DMBT1 was significantly increased during the development of OA.(2)Cholesterol levels in OA cartilage were higher than those in normal cartilage.(3)Knockdown of DMBT1 in chondrocytes effectively inhibited IL-1βinduced elevated levels of catabolism and apoptosis.(4)Knockdown of DMBT1 in chondrocytes effectively inhibited IL-1β-induced elevation of total cholesterol levels.(5)DMBT1 can inhibit the function of PPARα in regulating lipid metabolism in chondrocytes by activating the P38-MAPK signaling pathway,which ultimately leads to imbalance of catabolism and apoptosis in chondrocytes.(6)Knockdown of DMBT1 in the mouse knee joint can effectively inhibit DMM-induced articular cartilage degeneration.(7)Knockdown of DMBT1 in the knee joint of mice effectively inhibited the DMM-induced increase in MMP13 expression,decrease in COL2A1 expression,chondrocyte apoptosis and increase in cholesterol level in chondrocytes.CONCLUSIONS:(1)DMBT1 expression level correlates with the development of OA,plays an important role in the pathological process of OA,Perhaps this will become a potential molecular target to be developed for clinical treatment of OA.(2)DMBT1 can regulate PPARα by activating the P38 MAPK signaling pathway,thereby controlling lipid metabolism in chondrocytes,ultimately leading to increased levels of cartilage catabolism and apoptosis.(3)Knockdown of DMBT1 in mouse knee cartilage has a therapeutic effect on DMM-induced OA and can significantly improve articular cartilage degeneration. |
