乙型脑炎病毒感染的神经元中干扰素诱导蛋白-10与肿瘤坏死因子-α的相关性及意义

Correlation and Significance of Interferon Gamma-induced Protein-10 and Tumor Necrosis Factor-αin JEV-Infected Neurons

乙型脑炎病毒(Japanese encephalitis virus,JEV)感染引起血脑屏障损伤与干扰素诱导蛋白-10(Interferon gamma-induced protein-10,IP-10)表达上调及下游肿瘤坏死因子-α(Tumor necrosis factor-α,TNF-α)释放增加有关,但IP-10及TNF-α在JEV感染引起神经元损伤中的作用并不清楚。为了研究JEV感染的神经元中IP-10与TNF-α的相关性及生物学意义,本研究以SH-SY5Y细胞为实验对象,建立JEV感染的细胞模型,转染阴性对照(Negative control,NC)siRNA或IP-10 siRNA,给予溶剂或重组人TNF-α干预。干预后,检测细胞中IP-10、p-JNK的表达水平,培养基中TNF-α的含量,细胞存活率及凋亡率,JEV的滴度及拷贝数。结果显示,JEV组细胞中IP-10的表达水平及培养基中TNF-α的含量均高于对照组(P<0.05)且IP-10的表达水平与培养基中TNF-α的含量具有正相关关系;敲低IP-10后,si-IP-10+JEV组细胞中IP-10、p-JNK的表达水平、培养基中TNF-α的含量、细胞凋亡率均低于siNC+JEV组,细胞存活率高于si-NC+JEV组(P<0.05),JEV的滴度及拷贝数与si-NC+JEV组比较无差异(P>0.05);加用重组人TNF-α后,TNF-α+si-IP-10+JEV组的细胞存活率低于溶剂+si-IP-10+JEV组,p-JNK的表达水平及细胞凋亡率高于溶剂+si-IP-10+JEV组(P<0.05)。以上结果表明,JEV感染神经元的损伤与IP-10表达上调及下游TNF-α释放增加有关;高表达的IP-10能够增加TNF-α释放并激活JNK,进而引起细胞凋亡、加重细胞损伤。本研究初步探究了JEV感染引起神经元损伤的分子机制,发现IP-10通过增加TNF-α释放、激活JNK的方式在JEV感染神经元的过程中促进细胞凋亡、加重细胞损伤,这也为今后深入认识流行性乙型脑炎的发病机制提供了实验依据。

Japanese encephalitis virus(JEV)infection-induced injury to the blood–brain barrier is associated with upregulation of interferon gamma-induced protein(IP)-10 expression and increased downstream release of tumor necrosis factor(TNF)-α.However,the role of IP-10 and TNF-αin injury to JEV-infected neurons is not clear.To study the correlation and biological significance of IP-10 and TNF-αin JEV-infected neurons,SH-SY5Y cells were used as experimental objects,and we established a model of JEV infection.Then,negative control(NC)small interfering(si)RNA or IP-10 siRNA was transfected into SH-SY5Y cells,and solvent or recombinant human TNF-αwas given.After intervention,expression of IP-10 and phosphorylated c-Jun N-terminal kinase(p-JNK)in cells,the content of TNF-αin the medium,cell-survival rate,apoptosis rate,JEV titer,and copy number were measured.Expression of IP-10 and TNF-αcontent in the medium of the JEV group were higher than those of the control group(P<0.05)and there was a positive correlation between IP-10expression and TNF-αexpression.After knockdown of IP-10 expression,we found that expression of IP-10 and p-JNK,the TNF-αcontent in the medium,and apoptosis rate in the si-IP-10+JEV group were lower than those in the si-NC+JEV group,the survival rate was higher that than that in the si-NC+JEV group(P<0.05),and there was no difference in JEV titer or copy number between the si-NC+JEV group and si-IP-10+JEV group(P>0.05).After addition of recombinant human TNF-α,the survival rate in the TNF-α+si-IP-10+JEV group was lower than that of the solvent+si-IP-10+JEV group,and p-JNK expression and apoptosis rate were higher than those in the solvent+si-IP-10+JEV group(P<0.05).These results suggest that the injury to JEV-infected neurons is related to the upregulation of IP-10 expression and increased release of TNF-α.High expression of IP-10 can increase TNF-αexpression,which then activates JNK,induces apoptosis,and aggravates cell damage.We preliminarily explored the molecular mechanism of neuron damage caused by JEV infection,and found that IP-10 has a role in promoting apoptosis and aggravating cell damage in JEV-infected neurons by increasing TNF-αrelease and activating JNK.These results also provide an experimental basis for further understanding of the pathogenesis of Japanese encephalitis.