基于TLR4/MyD88/NF-κB信号通路探讨芪黄解毒化瘀饮治疗脓毒症急性肾损伤的作用机制

Exploring the mechanism of Qihuang Jiedu Huayu Yin in treating acute kidney injury in sepsis based on TLR4/MyD88/NF-κB signal pathway

关键词:  脓毒症  急性肾损伤  芪黄解毒化瘀饮  TLR4/MyD88/NF-κB信号通路

Key words:  sepsis  acute kidney injury  Qihuang Jiedu Huayu Yin  TLR4/MyD88/NF-κB signal pathway

摘要:

[目的] 基于Toll样受体4（TLR4）/髓样分化因子88（MyD88）/核转录因子-κB（NF-κB）信号通路探讨芪黄解毒化瘀饮治疗脓毒症急性肾损伤（AKI）的影响及作用机制。[方法] 将SD大鼠随机分为假手术组、模型组、芪黄解毒化瘀饮低、中、高剂量组，每组 10 只。通过盲肠结扎穿刺的方法构建脓毒症AKI大鼠模型，各组分别于造模前每12 h灌胃，连续3 d，造模后12 h灌胃。记录各组大鼠的一般状态，酶联免疫吸附法（ELISA）试剂盒检测各组大鼠血清肌酐（Scr）、尿素氮（BUN）、中性粒细胞明胶酶相关脂质释放素（NGAL）、肾损伤分子（KIM）-1、白细胞介素（IL）-1β、IL-6、IL-10、肿瘤坏死因子（TNF）-α、过氧化氢酶（CAT）、还原性谷胱甘肽过氧化物酶（GSH-PX）、丙二醛（MDA）、总超氧化物歧化酶（T-SOD）水平。苏木素-伊红（HE）染色观察大鼠肾组织病理形态变化。蛋白免疫印迹法（Western blot）检测大鼠肾组织中TLR4、MyD88、NF-κB蛋白表达水平。[结果] 与模型组相比，芪黄解毒化瘀饮低、中、高剂量组大鼠的一般状态均已改善；高剂量组血清Scr、BUN水平明显降低（P<0.05），中、高剂量组NGAL和KIM-1水平明显降低（P<0.05，P<0.01）；低、中、高剂量组大鼠肾脏组织病理损伤明显改善；芪黄解毒化瘀饮低、中、高剂量组IL-1β、IL-6、TNF-α的水平降低（P<0.01或P<0.05），IL-10水平升高（P<0.01）；芪黄解毒化瘀饮低、中、高剂量组MDA的水平明显降低（P 0.01），CAT水平明显升高（P<0.05或P<0.01）；中、高剂量组GSH-PX水平明显升高（P<0.05）；高剂量组T-SOD水平明显升高（P<0.05）；芪黄解毒化瘀饮低、中、高剂量组TLR4、MyD88、NF-κBp65 蛋白表达水平明显降低，差异具有统计学意义（P<0.01或P<0.05）。[结论] 芪黄解毒化瘀饮可以改善脓毒症AKI大鼠的一般状态，改善肾功能障碍和肾脏病理损伤，减轻炎症水平和氧化应激反应，其作用机制可能与抑制TLR4/MyD88/NF-κB信号通路相关。

Abstract:

[Objective] To explore the mechanism of Qihuang Jiedu Huayu Yin in treating sepsis-induced acute kidney injury（AKI） based on TLR4/MyD88/NF-κB signal pathway. [Methods] SD rats were randomly divided into sham group，model group，and low，medium，and high dose groups of Qihuang Jiedu Huayu Yin，with 10 rats in each group. The sepsis-induced AKI rat model was constructed by cecal ligation and puncture. Each group was orally administered every 12 hours before modeling for 3 consecutive days，and 12 hours after modeling. Record the general status of each group of rats，and use an ELISA kit to detect serum creatinine（Scr），urea nitrogen（BUN），neutrophil gelatinase-associated lipid releasing hormone（NGAL），kidney injury molecule-1（KIM-1），and interleukin-1β（IL-1β）、IL-6，IL-10，tumor necrosis factor（TNF）-α，catalase（CAT），glutathione peroxidase（GSH-PX），malondialdehyde（MDA），and total superoxide dismutase（T-SOD）. Observation of pathological morphological changes in rat kidney tissue using hematoxylin-eosin（HE） staining. The expression levels TLR4，MyD88，NF-κB in rat kidney tissue protein were detected by Western blot. [Results] ACompared with the model group，the general state of rats in the low，medium，and high-dose groups of Qihuang Jiedu Huayu Yin was improved. The serum levels of Scr and BUN were significantly reduced in the high-dose group（P<0.05），while the levels of NGAL and KIM-1 were significantly reduced in the medium and high-dose groups（P<0.05 or P<0.01）. The pathological damage of renal tissue in low，medium，and high dose groups of Qihuang Jiedu Huayu Yin was significantly improved；the level of IL-1β，IL-6，TNF-α in Low，medium，and high-dose groups of Qihuang Jiedu Huayu Yin were decreased（P<0.01 or P<0.05），while the level of IL-10 was increased（P<0.01）. The levels of MDA in the low，medium，and high dose groups of Qihuang Jiedu Huayu Yin were significantly reduced（P<0.01），while the level of CAT was significantly increased（P<0.05 or P<0.01）. The levels of GSH-PX significantly increased in the medium and high dose groups（P<0.05）. The T-SOD level in the high-dose group significantly increased（P<0.05）. In low，medium，and high-dose groups of Qihuang Jiedu Huayu Yin，the expression level of TLR4，MyD88，and NF-κB were significantly reduced（P<0.01 or P<0.05）. [Conclusion] Qihuang Jiedu Huayu Yin can improve the general state of sepsis-induced AKI rats，improve renal dysfunction and pathological damage，reduce inflammation levels and oxidative stress response，and its mechanism of action may be related to the inhibition of TLR4/MyD88/NF-κB signal pathway.