Screening And Verifying The Target Genes Of MiR-23a, MiR-30a In Regulating The PDLCs

Periodontitis is a chronic inflammatory disease which has a very wide range of hazards for causing irreversible bone destruction. Currently, the common methods used in the treatment of periodontitis are the basic treatment, drug therapy and surgical treatment, but the ideal effect of periodontal bone tissue regeneration is difficult to achie ve. Coordination of osteoblasts and osteoclasts maintain the balance of bone metabolism. The inhibition of osteoblast function and the secretion of matrix mineralization blocked is a key link in the process of bone metabolic imbalances in inflammatory cond itions. Osteoblast differentiation can be regulated through different signaling pathways by a lot of inflammatory cytokines which include miRN As. MiRNAs are endogenous non-coding small RN As which are evolutionary highly conserved with the length of about 2 2nt involved in post-transcriptional gene repression. MiRN As have been associated with diverse biological processes and diseases. Relevant research reveals that the occurrence of periodontitis and regulation of osteoblast differentiation is related to certain miRNAs expression changes. However, the study remains discovery of a cluster of certain miRNAs. We found that miR-23 a and miR-30 a is common molecule in regulation of the periodontitis and osteoblast differentiation. Based on the above background, we will forecast the potential target genes by bioinformatics approaches and intervene the expression level of miR-23 a and miR-30 a in PDLCs in vitro, subsequently detect the expression of the forecasted potential target genes and related protein in order to screen the target genes of miR-23 a and miR-30 a in regulation of PDLCs. Objective:(1) To confirm the candidate target genes of miR-23 a and miR-30 a.(2) To screen and verify the target genes of miR-23 a among the candidate target genes.(3) To screen and verify the target genes of miR-30 a among the candidate target genes. Methods(1) miRanda, TargetScan and Pic Tar, these bioinformatics approaches were used to do the bio- informatic prediction for the candidate target genes of both miR-23 a and miR-30 a. The common forecasted potential genes of these softwares will include the potential target genes of both miR-23 a and miR-30 a.(2) Real-time PCR and Western Blot were used to detect the changed expression of related genes and protein after cell transfection of both miR-23 a mimic and inhibitor.(3) Real-time PCR and Western Blot were used to detect the changed expression of related genes and protein after cell transfection of miR-30 a mimic or inhibitor. Results:(1) The predicted result of these bioinformatics softwares is that genes include CAMTA1, CNTTBIP, PPPAR, Runx2, STAT1, STAT5 B and SATB1 were predicted as the putative target genes of miR-23 a and miR-30 a.(2) Real-time PC R and Western Blot showed the expression of the related gene STAT5 B and related protein of PDLC was attenuated by increasing the expression of miR-23 a. While, the opposite had happened by decreasing the expression of miR-23 a, which confirmes that STAT5 B is the target gene of miR-23 a.(3) Real-time PCR and Western Blot showed the expression of the related gene Runx2 and related protein of PDLC was attenuated by upregulating the expression of miR-30 a. While, the opposite had happened by downregulating the expression of miR-30 a, which confirmes that Runx2 is the target gene of miR-30 a. Conclusions:(1) MiR-23 a and miR-30 a are involved in the regulation of PDLCs in the inflammatory microenvironment.(2) The osteogenesis of PDLCs can be inhibited when the expression of miR-23 a and miR-30 a is attenuated.