Circular RNA CircBNC2 Reduces Renal Fibrosis After Injury By Inhibiting Cell Cycle Arrest In Renal Tubular Epithelial Cells

BackgroundChronic kidney disease is chronic kidney structure and dysfunction caused by various reasons.It has the characteristics of long course of disease,complex etiology,and lack of treatment methods.It is considered to be a major threat to human health.Fibrosis is a common pathological manifestation and an important driving factor of chronic kidney disease.When the body is severely or repeatedly injured,normal tissue repair can evolve into progressive fibrosis,excessive accumulation of extracellular matrix(ECM),leading to organ dysfunction,and finally lead to end-stage renal disease and even death.Studies have confirmed that renal fibrosis is related to renal tubular epithelial cell(TEC)dysfunction,but the molecules involved in this process and their mechanism of action have not been fully revealed,and further research is urgently needed to find promising biomarkers.Circular RNA(circRNA)is a type of non-coding RNA widely expressed in eukaryotes.Its circular structure is resistant to degradation by exonucleases and can exist relatively stable in vivo.Strong tissue specificity and cell specificity,so it is considered as an ideal potential biomarker.Recent literature has shown that circular RNAs are involved in the regulation of organ fibrosis,but the specific circular RNA molecules that regulate renal fibrosis and their mechanisms of action have not yet been elucidated.This topic intends to explore the regulatory effect and mechanism of circular RNA on renal fibrosis,aiming to provide new ideas for the diagnosis and treatment of chronic kidney disease.Methods1.Screening and identification of fibrosis-associated circular RNAs after renal injury1.1 Construct a mouse model of ischemia-reperfusion injury(IRI),and screen for significantly differentially expressed circular RNAs by high-throughput sequencing.1.2 Polymerase chain reaction(PCR)was used to detect the expression of circBNC2 in various human tissues to identify tissue specificity.1.3 Actinomycin D test and RNase test to identify the stability of circBNC2.1.4 Nucleocytoplasmic separation and Northern blot experiments to detect the localization of circBNC2 cells.2.CircBNC2 inhibits cell cycle arrest and fibrosis in human TECs after injury2.1 Down-regulate the expression of circBNC2 in TEC,and analyze the biological process that circBNC2 may participate in by genomics.2.2 Flow cytometry,immunofluorescence,and Western blot were used to detect cell cycle and fibrosis-related indicators.2.3 After overexpressing circBNC2 in TEC,cell injury experiments were performed to verify the effect of circBNC2 on the cell cycle.3.CircBNC2 inhibits fibrosis after renal injury in mice3.1 The mouse circBNC2 knockout and overexpression models were constructed to treat with ischemia-reperfusion injury.3.2 Western blot and pathological staining to detect cell cycle and fibrosis-related indicators.Results1.The expression of circBNC2 was significantly down-regulated in the injured mouse kidney tissue.It has a stable circular structure,is specifically expressed in kidney tissue,and is mainly localized in the TEC cytoplasm.2.circBNC2 can inhibit the G2/M arrest of TEC after injury.3.circBNC2 can inhibit the secretion of pro-fibrotic factors and G2/M arrest after renal injury in mice,and reduce the degree of renal fibrosis.ConclusionCircBNC2 attenuates renal fibrosis after kidney injury by inhibiting TEC G2/M arrest.