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目的探讨TGF-β1刺激后对皮肤成纤维细胞中磷酸化ERK表达的影响及相互关系。方法以原代培养人正常皮肤成纤维细胞为研究对象。细胞分2组:浓度组:10 ng/ml、5 ng/ml、1.0 ng/ml、0 ng/ml。时间组:给予10 ng/ml的TGF-β1刺激不同时间,分别在0、15、30、60、120 min的时间点提取蛋白。采用Western blotting法检测成纤维细胞中磷酸化ERK蛋白的表达变化。结果浓度组及时间组p-ERK蛋白表达与对照组相比均有差异(P<0.05),随着TGF-β1剂量增加和作用时间延长,其磷酸化ERK蛋白表达水平逐渐增强,在TGF-β1 5 ng/ml时p-ERK水平达作用最强,在作用60 min时p-ERK达高峰。结论随着TGF-β1浓度增加及作用时间延长,大剂量TGF-β1时可上调体外培养成纤维细胞内的ERK信号蛋白活性。TGF-β1诱导的成纤维细胞的p-ERK水平增高可能是依赖ERK通路激活的,但ERK通路与TGF-β/Smad通路存在交互作用的机制,目前还无定论。
Objective To investigate the effect of TGF-β1 on the phosphorylation of ERK in skin fibroblasts and its relationship. Methods Primary cultured human normal skin fibroblasts were studied. Cells were divided into 2 groups: concentration group: 10 ng / ml, 5 ng / ml, 1.0 ng / ml, 0 ng / ml. Time group: 10 ng / ml of TGF-β1 stimulation at different times, respectively, at 0,15,30,60,120 min time point protein extraction. Western blotting was used to detect the expression of phosphorylated ERK protein in fibroblasts. Results Compared with the control group, the expression of p-ERK protein in the concentration group and the time group were significantly different (P <0.05). The phosphorylated ERK protein expression gradually increased with the increase of TGF-β1 dose and time, The p-ERK level was the strongest at β5 ng / ml, and peaked at 60 min. Conclusion With the increase of TGF-β1 concentration and prolongation of action time, high-dose TGF-β1 can up-regulate the activity of ERK signaling in cultured fibroblasts. TGF-β1-induced fibroblasts increased p-ERK levels may be dependent on ERK pathway activation, but ERK pathway and TGF-β / Smad pathway exists interaction mechanism, it is still inconclusive.