基于TGF-β1/Smad3信号通路探究真武汤对风湿性心脏病大鼠心肌纤维化 和免疫紊乱的影响

作者：赵 鹏1，王洲力1，熊 鹏2，邓文清1

单位：1.西安交通大学医学部附属三二〇一医院，陕西 汉中 723000； 2.陕西中医药大学附属医院，陕西 咸阳 712000

引用：引用：赵鹏，王洲力，熊鹏，邓文清.基于TGF-β1/Smad3信号通路探究真武汤对风湿性心脏病大鼠心肌纤维化和免疫紊乱的影响[J].中医药导报,2025,31(1):20-26.

DOI：10.13862/j.cn43-1446/r.2025.01.004

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摘要：目的：研究真武汤对风湿性心脏病（RHD）大鼠转化生长因子-β1（TGF-β1）/Smads信号通路及免疫紊乱的影响。方法：将60只Lewis大鼠随机分为正常组、RHD组、曲美他嗪组及真武汤组，每组15只。建立灭活A组-β型溶血性链球菌（GSA）诱导的大鼠风湿性心脏病模型。各组大鼠每天以相应药物灌胃1次，持续6周。检测大鼠心率；HE染色观察心脏组织病理改变；苦味酸-天狼猩红染色检测心脏组织纤维化；酶联免疫吸附（ELISA）法检测血清脑尿钠肽（BNP）、肌酸激酶同工酶MB（CK-MB）、乳酸脱氢酶（LDH）、肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）、IL-6、IL-10水平；Western blotting法检测心脏组织中TGF-β1、p-Smad3、Ⅰ型胶原蛋白（COL-Ⅰ）、基质金属蛋白酶1（MMP-1）、基质金属蛋白酶抑制剂-1（TIMP-1）、NOD样受体热蛋白结构域相关蛋白3（NLRP3）、诱导型一氧化氮合酶（iNOS）、环氧合酶-2（COX-2）相对表达量；免疫荧光法检测心脏组织中TGF-β1、Smad3、α-平滑肌肌动蛋白(α-SMA)、波形蛋白（vimentin）阳性表达。结果：正常组大鼠心肌细胞形态正常；RHD组大鼠心肌细胞排列紊乱且松散，可见大量肌纤维断裂，胞浆染色不均；与RHD组比较，曲美他嗪组、真武汤组大鼠心肌损伤有所改善。RHD组大鼠心脏组织相对纤维化面积占比高于正常组（P＜0.05）；曲美他嗪组、真武汤组大鼠心脏组织纤维化面积占比均低于RHD组（P＜0.05）。RHD组大鼠心率低于正常组（P＜0.05）；曲美他嗪组、真武汤组大鼠心率均高于RHD组（P＜0.05）。RHD组大鼠血清BNP、LDH、CK-MB、TNF-α、IL-1β、IL-6水平高于正常组（P＜0.05），IL-10水平低于正常组（P＜0.05）；曲美他嗪组、真武汤组大鼠血清BNP、LDH、CK-MB水平均低于RHD组（P＜0.05）；真武汤组大鼠血清TNF-α、IL-1β、IL-6水平低于RHD组，IL-10水平高于RHD组（P＜0.05）；真武汤组大鼠血清LDH、CK-MB水平低于曲美他嗪组（P＜0.05）。RHD组大鼠心脏组织TGF-β1、Smad3、α-SMA、Vimentin阳性表达水平高于正常组（P＜0.05）；曲美他嗪组、真武汤组大鼠心脏组织TGF-β1、Smad3、α-SMA、Vimentin阳性表达水平均低于RHD组（P＜0.05）；真武汤组大鼠心脏组织Smad3阳性表达水平低于曲美他嗪组（P＜0.05）。RHD组大鼠心脏组织TGF-β1、p-Smad3、COL-Ⅰ、MMP-1、NLRP3、iNOS、COX-2蛋白相对表达量高于正常组（P＜0.05），Smad7、TIMP-1蛋白相对表达量低于正常组（P＜0.05）；曲美他嗪组大鼠心脏组织MMP-1、NLRP3蛋白相对表达量低于RHD组（P＜0.05），Smad7、TIMP-1蛋白相对表达量高于RHD组（P＜0.05）；真武汤组大鼠心脏组织TGF-β1、p-Smad3、COL-Ⅰ、MMP-1、NLRP3、iNOS、COX-2蛋白相对表达量低于RHD组（P＜0.05），Smad7、TIMP-1蛋白相对表达量高于RHD组（P＜0.05）；真武汤组大鼠心脏组织TGF-β1、p-Smad3、COL-Ⅰ蛋白相对表达量低于曲美他嗪组（P＜0.05）。结论：真武汤可能通过负反馈调节TGF-β1/Smad3信号通路，抑制免疫炎症反应，改善RHD大鼠心肌纤维化，减轻心肌病理损伤。

关键词：风湿性心脏病；真武汤；TGF-β1/Smad3；心肌纤维化；免疫紊乱；大鼠

Abstract：

Objective: To investigate the effects of Zhenwu decoction on the transforming growth factor-β1 (TGF-β1)/Smads signaling pathway and immune dysregulation in rats with rheumatic heart disease (RHD). Methods: Totally 60 Lewis rats were randomly divided into normal group, RHD group, trimetazidine group and Zhenwu decoction group, with 15 rats in each group. A rat model of rheumatic heart disease was established using inactivated group A β-hemolytic streptococcus (GSA). Rats were given the corresponding medication via gavage in each group, once daily for six weeks. Parameters such as heart rate were measured; Hematoxylin-eosin (HE) staining was performed to observe pathological changes in the cardiac tissue. Bitterness acid-cockroach red staining was used to detect fibrosis in the heart tissue. Enzyme-linked immunosorbent assay (ELISA) was employed to measure serum levels of brain natriuretic peptide (BNP), creatine kinase MB isoenzyme (CK-MB), lactate dehydrogenase (LDH), tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), IL-6, and IL-10. Western blotting was used to analyze the relative expression levels of TGF-β1, p-Smad3, type I collagen (COL-I), matrix metalloproteinase-1 (MMP-1), tissue inhibitor of metalloproteinases-1 (TIMP-1), NOD-like receptor family pyrin domain containing 3 (NLRP3), inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) in cardiac tissue. Immunofluorescence was performed to detect the positive expression of TGF-β1, Smad3, alpha-smooth muscle actin (α-SMA), and vimentin in the heart tissue. Results: The myocardial cells of the normal group of rats appeared normal in morphology. In the RHD group, the myocardial cells were disorganized and loose, with a significant number of muscle fibers ruptured and uneven cytoplasmic staining. Compared to the RHD group, the myocardial damage showed some improvement in trimetazidine group and Zhenwu decoction group. The relative area of fibrosis in the myocardial tissue of the RHD group was higher than that of the normal group (P<0.05). The fibrotic area in the trimethazine group and the Zhenwu Decoction group was lower than that of the RHD group (P<0.05). The RHD group showed lower heart rate than normal group (P<0.05). The trimebutine group and the Zhenwu decoction group showed higher heart rates than the RHD group (P<0.05). The RHD group showed higher serum levels of BNP, LDH, CK-MB, TNF-α, IL-1β, and IL-6 than normal group (P<0.05), while lower IL-10 level than normal group (P<0.05). The trimebutine group and Zhenwu decoction group showed lower serum levels of BNP, LDH, and CK-MB than RHD group (P<0.05). The Zhenwu decoction group showed lower serum levels of TNF-α, IL-1β, and IL-6 than RHD group, while higher IL-10 level than RHD group (P<0.05). The Zhenwu decoction group showed lower serum levels of LDH and CK-MB than trimebutine group (P<0.05). In cardiac tissue, the positive expression levels of TGF-β1, Smad3, α-SMA, and Vimentin in the RHD group were higher than that of the normal group (P<0.05). In contrast, the positive expression levels of TGF-β1, Smad3, α-SMA, and Vimentin in the trimebutine group and Zhenwu decoction group were lower than that of the RHD group (P<0.05). The Zhenwu decoction group showed lower positive expression level of Smad3 than trimebutine group (P<0.05). The relative expression levels of TGF-β1, p-Smad3, COL-I, MMP-1, NLRP3, iNOS, and COX-2 proteins in the cardiac tissue of the RHD group were higher than that of the normal group (P<0.05), while the relative expression levels of Smad7 and TIMP-1 proteins were lower than normal group (P<0.05). In the trimebutine group, the relative expression levels of MMP-1 and NLRP3 proteins in the cardiac tissue were lower than that of the RHD group (P<0.05), while the relative expression levels of Smad7 and TIMP-1 proteins were higher than RHD group (P<0.05). In the Zhenwu decoction group, the relative expression levels of TGF-β1, p-Smad3, COL-I, MMP-1, NLRP3, iNOS, and COX-2 proteins in cardiac tissue were lower than RHD group (P<0.05), while the relative expression levels of Smad7 and TIMP-1 proteins were higher than RHD group (P<0.05). The relative expression levels of TGF-β1, p-Smad3, and COL-I proteins in the cardiac tissue of the Zhenwu decoction group were lower than that of the trimebutine group (P<0.05). Conclusion: Zhenwu decoction may regulate the TGF-β1/Smad3 signaling pathway through negative feedback, inhibiting immune inflammatory responses, improving myocardial fibrosis in rats with rheumatic heart disease, and alleviating myocardial pathological damage.

Key words：rheumatic heart disease; Zhenwu decoction; TGF-β1/Smad 3; myocardial fibrosis; immune disorders; rat

发布时间：2025-11-28

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