lncRNA-BC200调控Aβ_(25-35)诱导的神经细胞PC12炎症因子表达和细胞凋亡

LncRNA-BC200 regulating the expression of inflammatory factor and apoptosis of neuronal cell PC12 induced by Aβ_(25-35)

目的探讨lncRNA-BC200对Aβ_(25-35)诱导的神经细胞炎症和细胞凋亡的影响及可能机制。方法将神经细胞PC12分为正常对照组(细胞常规培养)和10、20、40μmol/L Aβ_(25-35)组(分别用10、20、40μmol/L Aβ_(25-35)干预细胞24 h),流式细胞术检测细胞凋亡,qRT-PCR法检测细胞中lncRNA-BC200表达。将PC12细胞分为正常对照组、Aβ_(25-35)组(用20μmol/L Aβ_(25-35)干预PC12细胞24 h)、si-NC+Aβ_(25-35)组(用20μmol/L Aβ_(25-35)干预转染si-NC的PC12细胞24 h)、si-lncRNA-BC200+Aβ_(25-35)组(用20μmol/L Aβ_(25-35)干预转染si-lncRNA-BC200的PC12细胞24 h)和TNF-α+si-lncRNA-BC200+Aβ_(25-35)组[用20μmol/L Aβ_(25-35)和20μg/L肿瘤坏死因子α(TNF-α)共同干预转染si-lncRNA-BC200的PC12细胞24 h],流式细胞术检测细胞凋亡,酶联免疫吸附法检测细胞培养上清液中TNF-α、白细胞介素6(IL-6)和γ干扰素(IFN-γ)表达,Western blot检测细胞中cleaved-Caspase-3、p-p65和p-IκBα蛋白表达。结果10、20、40μmol/L Aβ_(25-35)组PC12细胞凋亡率和lncRNA-BC200表达均高于正常对照组。Aβ_(25-35)组细胞中cleaved-Caspase-3、p-p65和p-IκBα蛋白表达,以及TNF-α、IL-6和IFN-γ水平均高于正常对照组。si-lncRNABC200+Aβ_(25-35)组PC12细胞凋亡率和细胞中cleaved-Caspase-3、p-p65和p-IκBα蛋白表达及TNF-α、IL-6和IFN-γ水平均低于Aβ_(25-35)组。TNF-α+si-lncRNA-BC200+Aβ_(25-35)组PC12细胞凋亡率和细胞中cleaved-Caspase-3、p-p65和p-IκBα蛋白表达及TNF-α、IL-6和IFN-γ水平均高于si-lncRNA-BC200+Aβ_(25-35)组。结论敲减lncRNA-BC200可能通过抑制NF-κB信号通路的激活减少Aβ_(25-35)诱导的神经细胞PC12分泌炎症因子及细胞凋亡。

Aim To investigate the effect of lncRNA-BC200 on the inflammation and apoptosis of nerve cells induced by Aβ_(25-35) and its possible mechanism.Methods The nerve cells PC12 were divided into NC group(conventional cell culture)and 10,20,40μmol/L Aβ_(25-35) groups(cells were treated with 10,20,40μmol/L Aβ_(25-35) for 24 h),and then cell apoptosis was detected by flow cytometry.qRT-PCR method was used to detect the expression of lncRNABC200 in cells.The PC12 cells were divided into NC group(cells were routinely cultured),Aβ_(25-35) group(PC12 cells were intervened with 20μmol/L Aβ_(25-35) for 24 h),si-NC+Aβ_(25-35) group(PC12 cells were transfected with si-NC and then intervened with 20μmol/L Aβ_(25-35) for 24 h),si-lncRNA-BC200+Aβ_(25-35) group(PC12 cells were transfected with si-lncRNA-BC200 and then intervened with 20μmol/L Aβ_(25-35) for 24 h)and TNF-α+si-lncRNA-BC200+Aβ_(25-35) group(PC12 cells were transfected with si-lncRNA-BC200 and then co-intervened with 20μmol/L Aβ_(25-35) and 20μg/L tumor necrosis factor(TNF-α)for 24 h).Flow cytometry was used to detect cell proliferation activity and apoptosis,ELISA was used to detect the expression of TNF-α, interleukin-6 (IL-6) and interferon-γ (IFN-γ) in the cell culture supernatant, and West-ern blot was used to detect the protein expression of cleaved-Caspase-3, p-p65 and p-IκBα in the cells. Results The apoptosis rate of PC12 cells and the expression of lncRNA-BC200 were higher in 10, 20, 40 μmol/ L Aβ_(25-35) groups than NC group. The protein expression of cleaved-Caspase-3, p-p65 and p-IκBα and the levels of TNF-α, IL-6 and IFN-γ in Aβ_(25-35) group cells were all higher than NC group. The apoptotic rate of PC12 cells, the protein expression of cleaved- Caspase-3, p-p65 and p-IκBα, and the levels of TNF-α, IL-6 and IFN-γ were all lower in the si-lncRNA-BC200+Aβ_(25-35) group than Aβ_(25-35) group. The apoptotic rate of PC12 cells, the protein expression of cleaved-Caspase-3, p-p65 and p- IκBα, and the levels of TNF-α, IL-6 and IFN-γ in the TNF-α+si-lncRNA-BC200+Aβ_(25-35) group were all higher than the si-lncRNA- BC200+Aβ_(25-35) group. Conclusion Knockdown of lncRNA-BC200 may inhibit Aβ_(25-35) -induced neuronal cell PC12 secretion of inflammatory factors and apoptosis by inhibiting the activation of NF-κB signaling pathway.