Study On Effect Of MicroRNA-10a-5p On IL-6 Receptor On The Degeneration Of Human Cartilage Endplate Cells Based On Bioinformatics Analysis

OBJECTIVE: This study conducted in bioinformatics analysis by mining TCGA and NCBI GEO data sets to predictIL-6 and IL-6 receptor(IL-6R)regulate the expression of MicroRNA-10a-5p in chondrocyte degeneration,and validate hypotheses through cell biological experiments to evaluate the relationship between the expression of IL-6R and the clinical characteristics and prognosis prediction of chondrocyte degenerat ion and the mechanism of action of relevant miRNA regulators,and to explore the prognostic and predictive value of IL-6R as a potential biomarker of osteochondral degeneration.METHODS: 1.We retrieved gene data chips of chon drocyte degeneration model from GEO database,screened chip data that met the research purpose,analyzed the expression of microRNAs and other related genes in the data set to evaluate their expression regulation,screened gene expression data sets that met the research requirements,and analyzed and selected up-regulated and down-regulated IL-6 and IL-6.Genes related to IL-6R expression were then retrieved from the mirDB gene database to predict microRNAs potentially regulating IL-6R.The screened results were intersected to identify microRNAs that could be expected to target IL-6R,and a hypothesis was proposed that this microRNA might be potentially regulated.IL-6R expression to affect thesignal expression of IL-6 in osteoarthritic cartilage degeneration;2.Study the isolation,culture and identification of human normal and degenerated primary chondrocytes,and then detect the expression levels of target microRNA and IL-6R by PCR to verify the hypothesis and draw conclusions.RESULTS: 1.Two groups of gene expression datasets were screened out by NCBI GEO database: GSE93008 and GSE81139.MicoRNAs regulating IL-6R down-regulation were screened by microRNA intersection of two groups of gene chip data in 18 groups;3 groups of microRNAs that regulate increased expression;then predicted the potential micro RNAs that regulate IL-6R through mirDB and Tarscan7.2database,and continued the intersection analysis with the screened microRNAs.For the gene data of the intersection analysis,homology analysis was carried out by selecting a group as the research object,and the comparison found that it was consistent with the human and murine base sequences.Moreover,by analyzing the base sequence of IL-6R,we found that there were more than seven binding sites between the microRNA and IL-6R.Therefore,we can deduce the hypothesis that miR-10a-5p targets IL-6R and reduces IL-6-induced chondrocyte degenerative lesions.2.10 groups of normal primary chondrocytes and10 groups of degenerated primary chondrocytes were separated from normal cartilage tissue(from patients with scoliosis)and degeneratedcartilage tissue(from patients undergoing surgery for degenerative spinal lesions).At the same time,chondrocytes confirmed by cell identification test were used to detect miR-10a-5p by PCR.The expression of IL-6R and IL-6 in normal cartilage group was lower,the expression of miR-10a-5p was higher,the expression of IL-6R and IL-6 in degeneration group was higher,and the expression of mi R-10a-5p was lower.CONCLUSION: There may be a negative feedback relationship between microrna-10a-5p and IL-6R,which can affect the signal transmission of IL-6 and regulate the degeneration of chondrocytes.