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目的:探讨五灵胶囊对脂多糖(LPS)诱导的大鼠枯否细胞(Kupffer cells,KC)p38MAPK信号转导通路的影响。方法:分离纯化KCs,60ng/ml LPS刺激建立LPS的肝细胞损伤模型;40只SD大鼠药物处理后,分离制备含药血清。实验分为四组:空白血清组、LPS+空白血清组、含药血清Ⅰ组(10.0g/kg)+LPS、含药血清Ⅱ组(6.25g/kg)+LPS。KCs产生促炎因子(I125放免法测定TNF-α、IL-6、IL-8,比色法测定NO生成量),采用Western blot法检测ERK、p-ERK、p38、p-p38、TNF-α和STAT3的蛋白水平。结果:1、空白血清+LPS组,TNF-α、IL-6、IL-8和NO浓度明显高于空白血清组;2、同空白血清+LPS组比较,含药血清Ⅰ、Ⅱ组TNF-α、IL-6、IL-8和NO水平明显降低;3、与空白血清组比较,空白血清+LPS组能上调KCs对p-ERK、P38、p-P38、STAT3和TNF-α表达(p<0.01,p<0.05),对ERK表达无影响(p>0.05)4、同空白血清+LPS组比较,含药血清Ⅰ+LPS、含药血清Ⅱ+LPS组p-p38、STAT3、TNF-α表达显著下调。结论:五灵胶囊对LPS诱导的大鼠KCs p38MAPK信号转导通路及KCs分泌促炎细胞因子TNF-a、IL-6、IL-8和NO具有抑制作用,其作用机制为防治慢性肝炎提供新的理论依据,具有潜在的临床应用前景。
Objective: To investigate the effect of Wuling capsule on the p38 MAPK signal transduction pathway induced by lipopolysaccharide (LPS) in Kupffer cells (KC) in rats. Methods: KCs were isolated and purified, and stimulated with 60ng / ml LPS to establish LPS-induced hepatocyte injury model. Forty SD rats were treated with the drug and the serum was separated and prepared. The experiment was divided into four groups: blank serum group, LPS + blank serum group, drug-containing serum groupⅠ (10.0g / kg) + LPS, serum-containing group Ⅱ (6.25g / kg) + LPS. KCs produced proinflammatory cytokines (I125, TNF-α, IL-6 and IL-8 by radioimmunoassay), and NO production by colorimetric assay. Western blot was used to detect the expression of ERK, p-ERK, p38, p-p38 and TNF- α and STAT3 protein levels. The levels of TNF-α, IL-6, IL-8 and NO in blank serum + LPS group were significantly higher than those in blank serum group.2. Compared with blank serum + LPS group, IL-8, IL-8 and NO were significantly decreased.3. Compared with the blank serum group, the blank serum + LPS group could upregulate the expression of p-ERK, p38, p-P38, STAT3 and TNF- 0.01, p <0.05), but had no effect on ERK expression (p> 0.05) .4 Compared with the blank serum + LPS group, the expressions of p-p38, STAT3 and TNF- α expression was significantly down-regulated. CONCLUSION: Wuling capsule can inhibit the p38 MAPK signal transduction pathway induced by LPS and the secretion of proinflammatory cytokines TNF-a, IL-6, IL-8 and KO in KCs, and its mechanism of action is to provide new prevention and treatment of chronic hepatitis The theoretical basis, with potential clinical applications.