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目的:研究不同浓度TNF-α、IL-1β、TGF-β刺激对原代培养海马神经元CLC-3(voltage-gated chloride channel 3)mRNA及蛋白质水平的影响。方法:取原代培养8 d的大鼠海马神经元,随机分为对照组、TNF-α组、IL-1β组和TGF-β组。TNF-α、IL-1和TGF-β组分别加入加入含有浓度为10 ng/ml的相应炎症因子培养液,每组分别在孵育6、12、24 h后分别收集细胞提取总RNA和蛋白采用实时定量PCR、Western Blot技术检测CLC-3 mRNA及蛋白水平的表达。结果:TNF-α、IL-1β处理12 h后培养海马神经元CLC-3在mRNA及蛋白水平开始上调,高峰持续从12 h至24 h(P<0.05)。TNF-α刺激作用强于IL-1β。TGF-β处理海马神经元CLC-3 mRNA在6 h后即开始升高(P<0.05),但在孵育6 h到24 h时下降至正常对平(P>0.05)。结论:TNF-α、IL-1β、TGF-β可能通过刺激海马神经元CLC-3表达增加增强神经元存活能力。
Objective: To investigate the effects of different concentrations of TNF-α, IL-1β and TGF-β on the mRNA and protein level of CLC-3 in primary cultured hippocampal neurons. Methods: Primary cultured hippocampal neurons of 8-day-old rats were randomly divided into control group, TNF-α group, IL-1β group and TGF-β group. TNF-α, IL-1 and TGF-β groups were added to the medium containing the corresponding inflammatory cytokines at a concentration of 10 ng / ml, and the total RNA and protein were collected from the cells after incubation for 6, 12 and 24 h Real-time quantitative PCR and Western Blot were used to detect the expression of CLC-3 mRNA and protein. Results: The mRNA and protein levels of CLC-3 in hippocampal neurons increased from 12 h to 24 h after TNF-α and IL-1β treatment for 12 h (P <0.05). TNF-α stimulation stronger than IL-1β. The level of CLC-3 mRNA in hippocampal neurons increased at 6 h after TGF-β treatment (P <0.05), but decreased to normal level at 6 h to 24 h (P> 0.05). CONCLUSION: TNF-α, IL-1β and TGF-β may enhance neuronal viability by stimulating the expression of CLC-3 in hippocampal neurons.