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目的:探讨谷氨酰胺(Gln)抑制人外周血单个核细胞(PBMC)细胞因子过度表达的分子机制。方法:用Ficoll密度梯度离心法提取健康志愿者新鲜外周血单个核细胞,在培养液中加入Gln、P38MAPK通路阻滞剂(SB203580)和HSP70阻断剂(Quercetin),用内毒素刺激4 h后,留取细胞和上清液,ELISA法测定内毒素刺激下单个核细胞肿瘤坏死因子(TNF-α)和IL-10的表达,Western blot方法检测细胞内磷酸化P38的表达情况。结果:Gln能明显增加HSP70的表达,同时TNF-α和IL-10表达受抑;使用Quercetin和SB203580后,磷酸化P38减少的同时TNF-α和IL-10也减少了。结论:Gln下调LPS刺激下PBMC细胞因子的过度表达依赖于P38MAPK信号蛋白的抑制。
AIM: To investigate the molecular mechanism by which Gln inhibits the over-expression of cytokines in human peripheral blood mononuclear cells (PBMCs). Methods: Fresh peripheral blood mononuclear cells from healthy volunteers were extracted by Ficoll density gradient centrifugation. Gln, P38MAPK channel blockers (SB203580) and HSP70 blockers (Quercetin) were added to the culture medium and stimulated with endotoxin for 4 h , And the cells and supernatants were collected. The expression of TNF-α and IL-10 in mononuclear cells stimulated by endotoxin were measured by ELISA, and the phosphorylated P38 expression was detected by Western blot. Results: Gln significantly increased the expression of HSP70, while the expression of TNF-α and IL-10 was inhibited. After treatment with Quercetin and SB203580, the phosphorylated P38 decreased and TNF-α and IL-10 decreased at the same time. Conclusion: The overexpression of cytokines in PBMCs induced by Gln down-regulated LPS stimulation depends on the inhibition of P38MAPK signal protein.