TAX1BP1调控B细胞自噬和凋亡的研究

TAX1BP1 regulates B cell autophagy and apoptosis

目的探究TAX1BP1在B细胞自噬与凋亡中的作用。方法慢病毒感染B淋巴瘤细胞(Raji细胞),敲低TAX1BP1表达。利用雷帕霉素处理Raji细胞,通过Western blot、细胞免疫荧光染色和流式细胞术,观察自噬流强弱以及细胞凋亡情况。通过检测自噬相关通路P53蛋白表达变化,初步探索调控机制。结果荧光显微镜观察慢病毒感染阳性率在90%以上。与阴性组比较,shTAX1BP1组细胞中TAX1BP1的mRNA表达水平降低80%、蛋白表达水平降低90%。经10 nmol/L雷帕霉素处理后,与阴性组相比较,shTAX1BP1组LC3II/I比值显著下降(P<0.01),凋亡相关蛋白Bcl-2表达显著升高,Bax、Caspase-3表达显著下降。敲低TAX1BP1降低Raji细胞自噬水平,减少Raji细胞凋亡。敲低TAX1BP1增加P53蛋白表达,推测TAX1BP1可能通过P53相关通路调控Raji细胞的自噬与凋亡。结论敲低TAX1BP1可抑制Raji细胞自噬与凋亡,初步预测这一生物学作用可能与P53信号通路有关。本研究为进一步探讨SLE的发病机制奠定了一定的实验基础。

This study was designed to explore the role of TAX1BP1 in autophagy and apoptosis of B cells.B lymphoma cells(Raji cells)were infected with lentivirus to knock down TAX1BP1 expression;Rapamycin was used to treat Raji cells,and the autophagy flow and apoptosis were observed by Western blotting,immunofluorescence staining and flow cytometry.The protein expression of autophagy-related pathway P53 was detected to preliminarily explored the regulation mechanism.Fluorescence microscopy showed that the positive rate of lentivirus infection was above 90%.Compared with the negative group,the mRNA expression level of TAX1BP1 in the shTAX1BP1 group was reduced by 80%and the protein expression level was reduced by 90%.After treatment with 10 nmol/L rapamycin,the LC3II/I ratio in shTAX1BP1 group was significantly decreased(P<0.01),the expression of apoptosisrelated protein Bcl-2 was significantly increased,and the expression of Bax and Caspase-3 were significantly decreased,as compared with the negative group.Knocking down TAX1BP1 reduced the autophagy and apoptosis levels of Raji cells.Furthermore,knockdown of TAX1BP1 increased the expression of P53 protein,suggesting that TAX1BP1 may regulate autophagy and apoptosis of Raji cells through P53-related pathways.In conclusion,the knockdown of TAX1BP1 can inhibit the autophagy and apoptosis of Raji cells,and the mechanism may be related to the P53 signaling pathway.This study lays certain experimental and theoretical basis for further exploring the pathogenesis of systemic lupus erythematosus.