CircLphn3 Protects The Blood-brain Barrier By Binding MiR-185-5p To Upregulate Tight Junction Protein ZO1

Objective: To investigate the protective effect of circ Lpn3(mmu＿circ＿0001358)on the blood brain barrier(BBB)containing tight junction protein(TJPs)after TBI.Methods: Twenty C57BL/6 male mice were randomly divided into control group and TBI group.Firstly,TBI model was established by controlling cortical injury(CCI)of mice,and the cirrna differentially expressed in normal and damaged brain tissues was screened by RNA-Seq technique.The expression level of circ Lphn3 in normal and damaged brain tissue and the expression level of brain microvascular endothelial cells(b End3)were detected by real-time fluorescence quantitative PCR(q RT-PCR).The cyclization of circ Lphn3 was identified by PCR and FISH.The bmend3 cells were transfected with plasmid and mi RNA,and then the microvascular endothelial cells were screened.Then the relationship between circlphn3,mir-185-5p and ZO1 was analyzed by bioinformatics analysis,RT-PCR,double luciferase and Western blotting.In vitro monolayer BBB model was established.Cell permeability,immunofluorescence and Western blotting were used to verify that circ Lphn3 can up regulate the expression of ZO1 and up regulate the tight junction protein after TBI by acting as mi R-185-5p sponge.Results: A total of 2015 differentially expressed circ RNAs were identified by RNA-Seq.Bioinformatics analysis showed that many mi RNAs targeting mmu＿circ＿0001358 were closely related to tight junction protein genes.The expression of circ Lphn3 was significantly reduced in the treatment of TBI and heme chloride.After overexpression of circ Lphn3,the expression of ZO1,ZO2 and Occludin was up regulated,and mir-185-5p level decreased.On the contrary,the expression of ZO1 decreased after transfection of mir-185-5p mimics.Finally,we found that circ Lphn3 targets mir-185-5p,regulates the expression of ZO1 and improves the blood-brain barrier function.Conclusion: Circ Lphn3 is an important potential gene.It can play a protective role in blood brain barrier by regulating the tight junction protein after the up regulation of TBI by mir-185-5p/ZO1 axis,and provides a new direction for TBI treatment.