[1]戴旭芳,闫小晶,谢鹏,等.丙戊酸钠通过激活miR-34c-5p/ATG4B信号通路抑制SH-SY5Y细胞自噬[J].南方医科大学学报,2018,(12):1415.[doi:10.12122/j.issn.1673-4254.2018.12.03]
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丙戊酸钠通过激活miR-34c-5p/ATG4B信号通路抑制SH-SY5Y细胞自噬()
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《南方医科大学学报》[ISSN:1673-4254/CN:44-1627/R]

卷:
期数:
2018年12期
页码:
1415
栏目:
出版日期:
2018-12-31

文章信息/Info

Title:
Sodium valprovate suppresses autophagy in SH-SY5Y cells via activating miR-34c-5p/ ATG4B signaling pathway
作者:
戴旭芳闫小晶谢鹏连继勤
关键词:
丙戊酸钠SH-SY5Y细胞ATG4BmiR-34c-5p自噬
Keywords:
sodium valproate SH-SY5Y cells ATG4B miR-34c-5p autophagy
DOI:
10.12122/j.issn.1673-4254.2018.12.03
摘要:
目的探讨丙戊酸钠(VPA)对SH-SY5Y细胞中miR-34c-5p/ATG4B信号通路的激活作用及对自噬的影响。方法人神经 母细胞瘤细胞SH-SY5Y用含10%胎牛血清的DMEM培养基常规培养,然后使用不同剂量VPA处理SH-SY5Y细胞24 h,采用 定量PCR分析VPA处理对ATG4B mRNA与miR-34c-5p表达的影响,免疫印迹法分析VPA处理对ATG4B蛋白表达的影响,将 含ATG4B启动子片段的报告质粒转染SH-SY5Y细胞,然后通过荧光素酶报告基因系统分析VPA处理对ATG4B启动子活性的 影响,用VPA单独或联合转录抑制剂放线菌素D分别处理SH-SY5Y细胞不同时间,定量PCR分析检测不同处理对ATG4B mRNA稳定性的影响,用VPA单独或联合miR-34c-5p抑制剂处理细胞24 h,通过检测不同处理对ATG4B表达变化的影响,进 而判断miR-34c-5p在VPA下调ATG4B表达中的作用,同时通过Western blot检测不同处理对LC3-II表达的影响,分析细胞自 噬水平变化。结果VPA可剂量依赖性降低SH-SY5Y细胞中ATG4B的mRNA和蛋白水平(P<0.05)。VPA处理不影响SHSY5Y 细胞中ATG4B的启动子活性(P>0.05),但显著下调其mRNA的稳定性(P<0.05)。同时,VPA处理明显增强SH-SY5Y细 胞中miR-34c-5p的表达水平(P<0.05)。使用miR-34c-5p抑制剂与VPA联合处理SH-SY5Y细胞,可逆转VPA对ATG4B表达的 下调作用。VPA处理还下调了SH-SY5Y细胞中LC3-II的表达水平(P<0.05)。结论VPA可能通过激活miR-34c-5p/ATG4B信 号通路而抑制SH-SY5Y细胞自噬。
Abstract:
Objective To investigate the effect of sodium valproate (VPA) on activation of miR-34c-5p/ATG4B signaling pathway and autophagy in SH-SY5Y cells. Methods Routinely cultured SH-SY5Y cells were treated with VPA at different doses for 24 h, and the changes in the mRNA levels of ATG4B and miR-34c-5p and the protein expression of ATG4B were assessed using qRTPCR and immunoblotting, respectively. The effect of transfection with a plasmid containing ATG4B promoter on the promoter activity of ATG4B in VPA-treated SH-SY5Y cells was assessed using the reporter gene assay. The stability of ATG4B mRNA was analyzed with qPCR in SH-SY5Y cells treated with VPA alone or with VPA combined with the transcription inhibitor actinomycin D. The expression level of miR-34c-5p was detected using qPCR in SH-SY5Y cells treated with VPA alone or with VPA combined with miR-34c-5p mimics or antagonist, and the role of miR-34c-5p in VPA-induced ATG4B down-regulation was evaluated. The changes in the level of autophagy were evaluated by detecting LC3-II expression in the cells after treatment with VPA or VPA combined with miR-34c-5p antagonist. Results VPA dose-dependently down-regulated the expression of ATG4B at both the mRNA and protein levels in SH-SY5Y cells. VPA treatment did not significantly affect the promoter activity of ATG4B, but obviously lowered the mRNA stability of ATG4B in SH-SY5Y cells. VPA treatment up-regulated the expression of miR-34c-5p, and the miR-34c-5p antagonist reversed VPA-induced down-regulation of ATG4B in SH-SY5Y cells. VPA also down-regulated the expression level of LC3-II in SH-SY5Y cells. Conclusion VPA suppresses autophagy in SH-SY5Y cells possibly via activating miR-34c-5p/ATG4B signaling pathway.

相似文献/References:

[1]郑超,董淑英,童旭辉,等.丙戊酸钠对乳腺癌细胞缝隙连接功能的增强作用及其机制[J].南方医科大学学报,2013,(01):66.
[2]童旭辉,郑超,蒋国君,等.丙戊酸钠增强阿霉素的体外抗乳腺癌作用[J].南方医科大学学报,2015,(01):62.
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更新日期/Last Update: 1900-01-01