The Mechanism Of Long Non-Coding RNA H19 Alleviates Osteoarthritis Progression By Targeting Insulin Like Growth Factor Binding Protein 3

Background and objectiveOsteoarthritis(OA)is a common and chronic joint degeneration disease,with a high prevalence and disability rate in the ageing population.Joint replacement surgery remains the main treatment for advanced OA patients.Articular cartilage degeneration,known as one of the important pathological features of OA,includes an increase in matrix degrading enzymes such as matrix metalloproteinases and a decrease in the matrix components such as type Ⅱ collagen,which results in an imbalance metabolism regulation of the cartilage extracellular matrix.However,the mechanism on mediating the articular cartilage degeneration in OA still unclear.Long non-coding RNAs(LncRNAs)are transcriptional products with a length more than 200 nucleotides.Although LncRNAs lack the function of encoding proteins,they can participate in a variety of biological processes through regulating transcription,translation and post-translational modifications after binding to DNA,mRNA,proteins and other molecules.It has been shown that LncRNA H19 alleviated the progression of OA by regulating chondrocyte apoptosis and inflammatory response.However,the molecular mechanism of LncRNA H19 in regulating the balance between synthesis and degradation of chondrocytes extracellular matrix in OA remains undetermined.Therefore,this study further to explore the mechanism of LncRNA H19 to alleviate OA progression by regulating the metabolic balance on synthesis and degradation chondrocytes extracellular matrix.To further elucidate the potential mechanism and significance of LncRNA H19 in regulating the pathology of OA.Methods1.Clinical experiments were conducted by collecting tibial plateau tissues from OA patients who underwent knee replacement surgery.Safranin O-fast green staining detected the pathological change of knee cartilage tissue from osteoarthritis patients,and RNA In situ hybridization detected the expression of LncRNA H19 in cartilage tissues from OA patients.2.In vivo experiments the 8-week-old SD rats were performed the ACLT operation to establish the OA animal model and then injected the overexpression of LncRNA H19 adenovirus into the knee articular cavity.Safranin O-fast green staining detected the severity of cartilage destruction in rat knee.RNA In situ hybridization detected the expression of LncRNA H19 and immunohistochemistry staining detected the expression of MMP3 and COL2 in rats knee cartilage on different groups.3.In vitro experiments the primary chondrocytes from rats were co-cultivated with H2O2 to mimic injury of chondrocytes during OA.WB and q-PCR detected the expression of MMP13,COL2,Aggrecan and ADAMTS5 in chondrocytes.RNA pulldown assay combined with mass spectrometry analysis detected the target proteins interacting with LncRNA H19.Results1.The expression of LncRNA H19 was significantly down-regulated in cartilage tissues with more serious destruction from OA patients compared to the control group;2.The expression of LncRNA H19 decreased significantly in the cartilage tissue from ACLT-OA groups compared to the sham group.Exogenous supplementation of LncRNA H19 alleviated the cartilage destruction and reduced OARSI score in ACLTOA rats,and inhibited the expression of MMP3 as well as increased the expression of COL2.3.Co-stimulated with H2O2 down-regulated the expression of LncRNA in a concentration dependent manner on chondrocyte.Co-stimulated with H2O2 promoted the expression of MMP13 and ADAMTS5 while inhibiting the expression of COL2 and Aggrecan,and the expression of these molecules could be reversed by overexpression of LncRNA H19 on chondrocyte.LncRNA H19 interacted with protein IGFBP3 and then negatively regulated the expression of IGFBP3 on chondrocyte.ConclusionsThe expression of LncRNA H19 expression was significantly downregulated in OA cartilage tissue,and the downregulation of LncRNA H19 broke the metabolic balance on chondrocyte extracellular matrix.LncRNA H19 negatively regulated the expression of IGFBP3 through binding to the protein IGFBP3,thus promoted the synthesis and inhibited the degradation of chondrocyte extracellular matrix.Exogenous supplementation of LncRNA H19 alleviated the progression of cartilage degeneration in OA rats.