The Regulation Of Circular RNA PRKD3 On Osteogenic Differentiation Of Periodontal Stem Cells

Background:The treatment of periodontal tissue defects caused by periodontitis is a serious medical and economic challenge in modern society.In order to accelerate and strengthen the regeneration of periodontal tissues,scholars have been committed to relevant researches,such as the regenerative surgery,the development of transplant materials,the use of recombinant growth factors and so on.In recent years,with the development of tissue engineering and stem cell therapy technology,scholars have found that regeneration of periodontal tissues by using stem cells is a promising treatment method for periodontal defects.Periodontal ligaments are dense connective tissues that surround the tooth root and connect the tooth root and the alveolar bone.Periodontal ligament stem cells(PDLSCs)with the potential of self-renewal and multi-directional differentiation are important cells in the regeneration and remodeling of the periodontal tissues,and essential in the maintenance of the periodontal homeostasis.In order to better apply PDLSCs in promoting the regeneration of periodontal tissues,scholars have been paying attention to studying the regulatory mechanisms in the process of PDLSCs'osteogenic differentiation.Circluar RNAs(circRNAs)are special noncoding RNAs(ncRNAs)which are widely found in various organisms.They were first found in plant viroids and then in viruses and eukaryotic cells in the following decades.Since it was considered to be the product of a mistake splicing form at first,circRNAs did not attract people's attention.However,with the emergence of the new sequencing technologies and the development of bioinformatics in recent years,circRNAs have been found to be a common expressional form of the cellular genes.CircRNAs are a kind of conservative ncRNAs of high abundance&stability and diversity,which can act as sponges of microRNAs(miRNAs)and participate in the regulation of gene expression in transcription and translation.They can also be used as ideal biomarkers for the diagnosis of tumor,cardiovascular&cerebrovascular diseases,neuropsychiatric diseases,diabetes mellitus,bone diseases and other diseases.Meanwhile,circRNAs are widely involved in the tissue regeneration and the differentiation of stem cells,such as liver regeneration in rats,nerve regeneration in rats,and osteogenic differentiation of human bone marrow mesenchymal stem cells.Currently,circRNAs are mainly studied in tumors.Although there have been some studies on circRNAs in stem cell's osteogenesis,there are few studies on the effect of circRNAs on PDLSCs.And the roles of circRNA PRKD3 in the osteoblastic differentiation of PDLSCs has not been reported.Combined with the previous studies and bioinformatics analysis of the research group,we aimed to explore the effect of circRNA PRKD3 on the osteogenic differentiation of PDLSCs.PurposeThe differential expression of circRNA PRKD3 during the osteogenic differentiation of PDLSCs was detected,and the role of circRNA PRKD3 in the osteogenic differentiation of PDLSCs was studied.Methods1.PDLSCs were isolated and cultured by tissue block method,and the biological characteristics of PDLSCs were identified.2.Through raw material information analysis method,7 circRNAs(RSF1,HECA,DDB2,PRKD3,KIAA0146,C3orf23,DNMT3B)were selected.The gene with the most pronounced expression changes by real-time fluorescent quantitative polymerase chain reaction(qRT-PCR)were choosed for the following experiments.3.The most significantly differentially expressed circRNA was detected during the osteogenic differentiation of PDLSCs.The effect of the circRNA on the osteogenic differentiation of PDLSCs was tested by lentiviral vector preparation.Results1.PDLSCs were successfully isolated and cultured.Flow cytometry detection showed that PDLSCs had mesenchymal stem cell phenotype,and alkaline phosphatase staining,alizarin red staining and oil red O staining showed that PDLSCs had multi-directional differentiation ability.2.QRT-PCR results showed that circRNA PRKD3 was the most significantly different genes among the selected circRNAs.3.During the osteogenic differentiation of PDLSCs,circRNA PRKD3 expression was significantly increased.After knockdown of circRNA PRKD3,the osteogenic differentiation ability of PDLSCs was significantly reduced.ConclusionCircRNA PRKD3 promoted the osteogenic differentiation of PDLSCs.CircRNA PRKD3 is involved in the regulation of the osteogenic differentiation of PDLSCs.