Objective:Circular RNAs(circRNAs)have been shown to involve in pathological processes of ischemic stroke(IS),including autophagy.This study was designed to explore the effect of circR-ZC3HC1 on neuronal autophagy in I...Objective:Circular RNAs(circRNAs)have been shown to involve in pathological processes of ischemic stroke(IS),including autophagy.This study was designed to explore the effect of circR-ZC3HC1 on neuronal autophagy in IS and the related mechanisms.Methods:Expression of circR-ZC3HC1 in blood samples of IS patients and healthy controls was detected.Hippocampal neurons were treated with oxygen and glucose deprivation(OGD)to establish IS in vitro model.The expression of LC3 and p62 and the number of autophagosomes were examined to evaluate the autophagy level of OGD induced neurons using western blotting and transmission electron microscope.Cell apoptosis rate and the expression of cleaved caspase-3,Bax,and Bcl-2 were assessed byflow cytometry and western blotting.The binding relationships among circR-ZC3HC1,miR-384-5p,and SIRT1 were predicted and verified.Results:Low expression of circR-ZC3HC1 was found in blood samples of IS patients and OGD-treated neurons.Overexpressed circR-ZC3HC1 or inhibited miR-384-5p expression promoted autophagy and inhibited apoptosis of OGD-treated neurons,which could be reversed by further 3-MA treatment.Mechanistically,circR-ZC3HC1 targeted miR-384-5p to mediate SIRT1 expression.miR-384-5p overexpression or SIRT1 knockdown in the presence of circR-ZC3HC1 overexpression in OGD-treated neurons lead to reduced autophagy and enhanced apoptosis.Conclusion:Collectively,circR-ZC3HC1 promoted neuronal autophagy to attenuate IS via miR-384-5p/SIRT1 axis.展开更多
目的:研究NDRG2基因与去泛素化酶Zc3h12d在NF-κB转导通路中的调控机制。方法:采用LPS刺激人支气管气道上皮16HBE细胞株来构建炎性刺激时气道黏液高分泌模型,用瞬时转染过表达NDRG2基因和Zc3h12d si RNA干扰下调16HBE细胞,将细胞分成LPS...目的:研究NDRG2基因与去泛素化酶Zc3h12d在NF-κB转导通路中的调控机制。方法:采用LPS刺激人支气管气道上皮16HBE细胞株来构建炎性刺激时气道黏液高分泌模型,用瞬时转染过表达NDRG2基因和Zc3h12d si RNA干扰下调16HBE细胞,将细胞分成LPS+瞬时转染过表达NDRG2组(A组)、LPS+瞬时转染过表达NDRG2+Zc3h12d si RNA组(B组)、LPS+瞬时转染Zc3h12d si RNA组(C组)、LPS+转染空白载体组(D组),以未做任何处理的16HBE细胞作为对照组(E组)。反转录PCR检测气道黏蛋白(mucin,MUC)5AC m RNA的表达水平,ELISA法检测IL-1β、IL-6等炎症因子及MUC5AC的分泌水平,Western印迹检测Zc3h12d和NF-κB p65的分泌水平,激光共聚焦法检测16HBE细胞内MUC5AC表达。结果:与B组比较,A组炎症因子及MUC5AC表达明显减少,差异有统计学意义(P<0.05);而B组与C组比较,差异无统计学意义(P>0.05)。与D组比较,A组炎症因子及MUC5AC的表达明显降低,差异均有统计学意义(均P<0.05)。D组与E组比较,LPS刺激后炎症因子及MUC5AC分泌增多,差异均有统计学意义(均P<0.05)。结论:NDRG2基因可以通过Zc3h12d而实现对NF-κB转导通路的调控。展开更多
基金Supported by Ningbo Health Technology Project,Nos.2020Y12 and 2022Y12.
文摘Objective:Circular RNAs(circRNAs)have been shown to involve in pathological processes of ischemic stroke(IS),including autophagy.This study was designed to explore the effect of circR-ZC3HC1 on neuronal autophagy in IS and the related mechanisms.Methods:Expression of circR-ZC3HC1 in blood samples of IS patients and healthy controls was detected.Hippocampal neurons were treated with oxygen and glucose deprivation(OGD)to establish IS in vitro model.The expression of LC3 and p62 and the number of autophagosomes were examined to evaluate the autophagy level of OGD induced neurons using western blotting and transmission electron microscope.Cell apoptosis rate and the expression of cleaved caspase-3,Bax,and Bcl-2 were assessed byflow cytometry and western blotting.The binding relationships among circR-ZC3HC1,miR-384-5p,and SIRT1 were predicted and verified.Results:Low expression of circR-ZC3HC1 was found in blood samples of IS patients and OGD-treated neurons.Overexpressed circR-ZC3HC1 or inhibited miR-384-5p expression promoted autophagy and inhibited apoptosis of OGD-treated neurons,which could be reversed by further 3-MA treatment.Mechanistically,circR-ZC3HC1 targeted miR-384-5p to mediate SIRT1 expression.miR-384-5p overexpression or SIRT1 knockdown in the presence of circR-ZC3HC1 overexpression in OGD-treated neurons lead to reduced autophagy and enhanced apoptosis.Conclusion:Collectively,circR-ZC3HC1 promoted neuronal autophagy to attenuate IS via miR-384-5p/SIRT1 axis.
文摘目的:研究NDRG2基因与去泛素化酶Zc3h12d在NF-κB转导通路中的调控机制。方法:采用LPS刺激人支气管气道上皮16HBE细胞株来构建炎性刺激时气道黏液高分泌模型,用瞬时转染过表达NDRG2基因和Zc3h12d si RNA干扰下调16HBE细胞,将细胞分成LPS+瞬时转染过表达NDRG2组(A组)、LPS+瞬时转染过表达NDRG2+Zc3h12d si RNA组(B组)、LPS+瞬时转染Zc3h12d si RNA组(C组)、LPS+转染空白载体组(D组),以未做任何处理的16HBE细胞作为对照组(E组)。反转录PCR检测气道黏蛋白(mucin,MUC)5AC m RNA的表达水平,ELISA法检测IL-1β、IL-6等炎症因子及MUC5AC的分泌水平,Western印迹检测Zc3h12d和NF-κB p65的分泌水平,激光共聚焦法检测16HBE细胞内MUC5AC表达。结果:与B组比较,A组炎症因子及MUC5AC表达明显减少,差异有统计学意义(P<0.05);而B组与C组比较,差异无统计学意义(P>0.05)。与D组比较,A组炎症因子及MUC5AC的表达明显降低,差异均有统计学意义(均P<0.05)。D组与E组比较,LPS刺激后炎症因子及MUC5AC分泌增多,差异均有统计学意义(均P<0.05)。结论:NDRG2基因可以通过Zc3h12d而实现对NF-κB转导通路的调控。
