Roles Of MicroRNA-195 In Hippocampal Neurons Apoptosis Induced By Intermittent Hypoxia

Objective: This study aims to investigate microRNA-195(miR-195)expression in normal mice hippocampal neurons under intermittent hypoxia(IH)condition and the roles of miR-195 in hippocampal neurons apoptosis though targeting bcl-2.CIH is associated with lots of biological process,including neurobehavioral impairments,increased apoptosis and oxidant stress and inflammation,and hippocampal neurons apoptosis.MicroRNAs,a class of 20–23 nucleotides,noncoding single-stranded RNAs that regulate gene function,leading to a significant negative regulation of relevant mRNAs to be translated into proteins.Recent studies turned their eyes on the role of miRNAs in IH and found several miRNAs could influence this process of IH-induced cell apoptosis.Methods: A mouse model of CIH was established.MiR-195 expression levels in hippocampal neurons were detected by real-time quantitative PCR(qPCR).The primaryhippocampal neurons cultures were performed and maitained by using C57BL/6 mice.Anti-miR-195 inhibitor,mi R-negative control and miR-195 mimic were used to tranfect the cells using Lipofectamine 2000 in serum-free Opti-MEM medium(Gibco)according to the manufacturer's instruction.We divided them intodifferent groups under IH condition,interference group(treated with miR-195 inhibitor);negative group(treated with miR-negative control),IH group(cells under IH condition with no treatment),and also a blank group(cells under normoxia).PCR conducted in an ABI7500 real-time PCR instrument were used to determine the expressions of bcl-2 and bax.Western blotting was used to test the expression of bcl-2,bax,caspase-3.GAPDH was used as a control.Cell apoptosis was evaluated by TUNEL assay.All calculations were made using SPSS 18.0.Results: MiR-195 is significantly upregulated after CIH treatment in hippocampal neurons.Expression of miR-195 was significantly up regulated in the CIH group and was apparently inhibited in the interference group.Downregulated MiR-195 increased bcl-2 mRNA expression,decreased bax mRNA and protein expression(all P < 0.05).Results showed that when treated under CIH condition,expression of caspase-3 and bax was significantly increased while expression of bcl-2 was apparently decreased compared with the blank group.However,when expression of miR-195 was interrupted,the above influence was inhibited significantly comparing the CIH group with interference group,in which expression of caspase-3 and bax was reduced and expression of bcl-2 increased,which reflected the same results in the detection of apoptotic rate by TUNEL assay.Conclusion: Our study identified that miR-195 expression was upregulated under intermittent hypoxia,and miR-195 overexpression may promote hippocampal neurons apoptosis by targeting bcl-2 and inducing apoptotic pathway.These results may be useful for further relative researches and contribute to development of a new therapy for CIH-associated hippocampal neurons dysfunction.