摘要
目的:观察TNF-α对脂多糖应答基因(lrg)在人HEK293和U937细胞中表达的影响.方法:正常培养人胚肾细胞(HEK293)和人单核细胞(U937),用TNF-α(终浓度1 ×106U/L)刺激2 h.提取刺激前后HEK293和U937细胞的总蛋白,用纯化后的兔抗人Lrg抗血清作一抗(1:1000),对TNF-α刺激前后的HEK293和U937细胞进行Western Blot分析.提取刺激前后HEK293和U937细胞的总RNA,用RT-PCB分析TNF-α对lrg在细胞中表达的影响.以B-actin为内参.结果:Western Blot分析显示,用TNF-α刺激2 h后,lrg在人HEK293和U937细胞内的蛋白含量明显上升;RT-PCR结果显示,用TNF-α刺激2 h后,人HEK293和U937细胞内的lrgmRNA水平明显上升.结论:TNF-α的刺激增强了人HEK293和U937细胞内lrg的表达,提示lrg可能参与了TNF-α诱导的炎症反应.
Abstract
AIM:To examine the effect of tumor necrosis factor α (TNF-α) on lipopolysaccharide response gene (lrg) expression in human cell lines HEK293 and U937. METHODS: Human embryonic cell line HEK293 and leukemia cell line U937 were cultured in RPMI 1640 with 100 mL/L fetal serum under 50 mL/L CO2 condition. The cells were treated with TNF-α at the concentration of 1 × 106 U/L or placebo for 2 h. Western Blot and RT-PCR were performed to detect the lrg expression. β-actin was applied as an internal control. RESULTS: The results of Western Blot indicated that the lrg expression in both HEK293 and U937 cells at protein levels were up-regulated significantly after stimulation by TNF-α for 2 h. The results of RT-PCR indicated that the lrg mRNA levels in both HEK293 and U937 cells increased after stimulation by TNF-α for 2 h. CONCLUSION: The lrg expression in human HEK293 and U937 cells is both up-graded remarkably by the stimulation of TNF-α, which suggests that lrg probably plays roles in the inflammation induced by TNF-α.
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