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目的研究不同剂量“化瘀方”对内毒素(LPS)诱导脓毒症大鼠心肌损伤的保护作用,并探讨其作用机制。方法 6周龄SPF级雄性Wistar大鼠80只,随机分为正常组、模型组、中药(丹参30g,大黄15g)高[8g/(kg·d)]、中[4g/(kg·d)]、低[2g/(kg·d)]剂量预处理组,每组又分为注射LPS后3h和6h两个时间点。透射电镜观察大鼠心肌超微结构;ELISA法检测大鼠血清TNF-α、IL-1β、IL-6等炎性因子水平的变化;荧光定量聚合酶链式反应法和免疫印迹法检测心肌组织TNF-α、NF-κB和TLR4的基因表达。结果超微结构显示,各中药治疗剂量组心肌细胞均有不同程度损伤,但损伤程度明显轻于模型组。与模型组同时间点比较,中药各剂量组大鼠血清IL-6、IL-1β、TNF-α水平明显降低(P<0.01),高剂量组下降尤为显著(P<0.05 vs.低剂量)。与正常组比较,模型组和“化瘀方”各治疗组心肌组织中TNF-α、NF-κB和TLR4的mRNA转录和蛋白质翻译水平明显升高,模型6h组蛋白水平更高;而与模型组比较,中药治疗组相关基因转录和翻译水平则明显下降,其中高剂量组下降尤为显著。结论 “化瘀方”可以明显改善脓毒症大鼠的心肌损伤,并且证实该保护作用是通过抑制TLR4介导的炎症反应实现的。
Objective To study the protective effects of different doses of “Hua Yu Fang” on myocardial injury induced by endotoxin (LPS) in rats and its mechanism. Methods Eighty male Wistar rats of 6 weeks old were randomly divided into 4 groups: normal group, model group, high dose of Radix Salviae Miltiorrhizae (30g, Rhubarb 15g) [8g / (kg · d) ], Low [2g / (kg · d)] dose pretreatment group, each group is divided into three hours after injection of LPS and 6h two time points. The ultrastructure of myocardium was observed by transmission electron microscopy. The levels of TNF-α, IL-1β, IL-6 and other inflammatory cytokines in serum of rats were detected by ELISA. The changes of myocardial tissue were detected by fluorescence quantitative polymerase chain reaction and Western blotting. TNF-α, NF-κB and TLR4 gene expression. Results The ultrastructure showed that the cardiomyocytes in each dose group of Chinese traditional medicine had different degrees of injury, but the degree of injury was obviously lighter than that of the model group. Compared with the model group, the serum levels of IL-6, IL-1β and TNF-α in the Chinese medicine groups were significantly decreased (P <0.01), especially in the high-dose group (P <0.05 vs. low dose) . Compared with the normal group, mRNA transcription and protein translation of TNF-α, NF-κB and TLR4 in the model group and each treatment group were significantly increased Compared with model group, the transcription and translation level of related genes in Chinese medicine treatment group decreased significantly, especially in high-dose group. Conclusion “Huayu Recipe” can significantly improve myocardial injury in septic rats, and confirm that this protective effect is achieved by inhibiting the TLR4-mediated inflammatory reaction.