Mechanisms Of MiRNA-106b-5p On Autophagy And Apoptosis Of SH-SY5Y Cells

Objective: The expression of miR-106b-5p in spinal cord injury was analyzed in rat spinal cord injury model.Then,the expression of miR-106b-5p was overexpressed or inhibited by cell experiments to analyze the effects of miR-106b-5p on Beclin-1,P62,Atg7,Caspase-3 and Bax in SH-SY5 Y cells.To further explore whether miR-106b-5p plays a role in the pathophysiological process of spinal cord injury based on the regulation of autophagy and apoptosis of spinal cord neurons.It provides a new therapeutic target and thought for the treatment of spinal cord injury.Methods: The model of spinal cord injury in rats was established by modified Allen percussion method.The rats were randomly divided into the Sham group(Sham group)and the SCI group(SCI group).The spinal cord injury of the rats was evaluated by BBB score and spinal cord water content.The expression level of miR-106b-5p in SCI rats was detected by q RT-PCR on days 1,3,7 and 14.SH-SY5 Y cell lines were cultured in vitro and divided into 4 groups for transfection: silent negative control group(inhibitor control group),miR-106b-5p silent group(miR-106b-5p inhibitor group),mimic negative control group(mimic control group)and miR-106b-5p mimic group(miR-106b-5p mimic group).The transfection efficiency of miR-106b-5p was verified by q RT-PCR,and the protein levels of Beclin-1,P62,Atg7,Caspase-3 and Bax were determined by Western blot.Results:(1)Animal experiments showed that,compared with the Sham group,BBB scores of 1d,3d,7d and 14 d after spinal cord injury were significantly decreased,with significant differences(P<0.01).The measurement of spinal cord moisture content showed that the spinal cord moisture content of Sham group was about 75%,while the spinal cord moisture content of Sham group on day 1,3,7 and 14 after modeling was all higher than that of Sham group,the difference was significant(P< 0.001).q RT-PCR showed that,compared with the Sham group,the expression level of miR-106b-5p was significantly up-regulated on day 1,3,7 and 14 after spinal cord injury(P < 0.01),and the expression level of miR-106b-5p was the highest on day 1 after spinal cord injury.(2)Compared with inhibitor control group,the expression of miR-106b-5p in inhibitor control group was significantly decreased(P < 0.01);Compared with mimic control group,the expression of miR-106b-5p in mimic group increased significantly(P < 0.01).This indicated that miR-106b-5p was successfully transfected into SH-SY5 Y cells.Compared with inhibitor control group,the expressions of Beclin-1 and ATG-7 in miR-106b-5p inhibitor group were significantly up-regulated(P < 0.01),while the proteins of P62,Caspase-3 and Bax were significantly decreased(P < 0.01).Compared with mimic control group,the expressions of Beclin-1 and ATG-7 in miR-106b-5p mimic group were significantly down-regulated(P < 0.01),while the expressions of P62,Caspase-3 and Bax were significantly increased(P <0.01).Conclusions:(1)The m RNA expression of miR-106b-5p was increased in the rat model of spinal cord injury,suggesting that miR-106b-5p may be related to the pathophysiology and prognosis of SCI.(2)Silencing miR-106b-5p in SH-SY5 Y cells can promote the expression of Beclin-1 and ATG-7 proteins,and inhibit the expression of P62,caspase-3 and Bax proteins;Overexpression of miR-106b-5p inhibited the expression of Beclin-1 and ATG-7 proteins,and promoted the expression of P62,Caspase-3 and Bax proteins.The possible reason is that silencing miR-106b-5p can promote autophagy and inhibit apoptosis in SH-SY5 Y cells.(3)In this study,the regulatory effects of miR-106b-5p on Beclin-1,ATG-7,P62,caspase-3 and Bax were briefly verified by cell experiments.Meanwhile,the expression of miR-106b-5p was detected in the rat spinal cord injury model,and the mechanism of miR-106b-5p’s influence on autophagy and apoptosis of neurons in spinal cord injury was preliminarily discussed.The possible mechanism of spinal cord injury recovery was revealed from the perspective of miRNA-autophagy-apoptosis-disease.However,in-depth studies have not been conducted on the animal level and the regulation of downstream target genes by miRNA.In the next step,we will explore the mechanism of miR-106b-5p promoting spinal cord injury recovery from multiple perspectives,levels and dimensions.To provide a new target and theoretical basis for the clinical treatment of spinal cord injury.