LncRNA GAS5 Regulates Nerve Cell Apoptosis through miR-21 after Spinal Cord Injury
Objective To clarify the relationship between long non coding RNA growth arrest-special transcript 5(LncRNA GAS5)and microRNA-21(miR-21),and explore the mechanism of neuronal apoptosis regulated by LncRNA GAS5 via miR-21 after spinal cord injury.Methods The models of spinal cord injury of rats and adrenal pheochromocytoma cells(PC-12)with oxygen glucose deprivation reoxygenation(OGD/R)were established.After silencing or overexpressing GAS5,a lentivirus with downregulated shGAS5 expression was constructed,and the binding site between LncRNA GAS5 and miR-21 was predicted through bioinformatics analysis.The binding sites of GAS5 and miR-21 were explored by dual luciferase reporter gene.The real-time fluorescence quantitative polymerase chain reaction(RT-qPCR)was used to detect the RNA expression levels of miR-21,GAS5,homologous phosphatase and tensin homolog(PTEN)genes,caspase 3,Bcl-2 related X genes(Bax),Bcl-2,and protein kinase B(AKT).Western blotting was used to detect the protein expression levels of Cleaved caspase 3,Bax,Bcl-2,AKT,and phosphorylated AKT(p-AKT).TUNEL technology was used to detect the degree of neuronal apoptosis.Results The expression levels of miR-21,Bcl-2,and p-AKT decreased(P＜0.05),while the expression of GAS5,PTEN,Bax,and Cleaved caspase-3 increased(P＜0.05)in rats with spinal cord injury.PC-12 cells showed similar results to the rat spinal cord injury model after in vitro culture and OGD/R injury,and the results were most significant 2 h after OGD/R treatment.GAS5 could bind to miR-21 through complementary base pairing,thereby regulating the downstream PTEN signaling pathway.Downregulation of GAS5 or overexpression of miR-21 could inhibit the apoptosis of PC-12 cells(P＜0.05).Conclusion GAS5 upregulates PTEN and inhibits AKT phosphorylation by binding to miR-21,thereby promoting neuronal apoptosis induced by spinal cord injury.
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