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唐路军, 张稳, 曾阳, 袁柳媚, 刘慧慧, 贺海霞, 胡国恒.防己茯苓汤改善糖尿病肾病肾小球系膜细胞焦亡的机制研究[J].湖南中医药大学学报,2025,45(7):1231-1239[点击复制] |
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| 防己茯苓汤改善糖尿病肾病肾小球系膜细胞焦亡的机制研究 |
| 唐路军,张稳,曾阳,袁柳媚,刘慧慧,贺海霞,胡国恒 |
| (湖南航天医院, 湖南 长沙 410205;湖南中医药大学第一附属医院, 湖南 长沙 410007) |
| 摘要: |
| 目的 探讨防己茯苓汤对糖尿病肾病小鼠肾功能和肾小球系膜细胞焦亡的影响及其作用机制。方法 40只8周龄雄性C57BL/6小鼠随机分为对照组、模型组、防己茯苓汤组、抑制剂组和防己茯苓汤+抑制剂组，每组8只。对照组小鼠正常喂养，灌胃等量生理盐水，其余3组小鼠均通过高脂饲料喂养并注射链脲佐菌素构建糖尿病肾病模型。造模成功后，模型组灌胃等量生理盐水，防己茯苓汤组予防己茯苓汤灌胃，抑制剂组腹腔注射NLRP3抑制剂MCC950，防己茯苓汤+抑制剂组予防己茯苓汤灌胃+腹腔注射NLRP3抑制剂MCC950，每组连续干预8周。MES-13细胞随机分为对照组、模型组、防己茯苓汤组、抑制剂组和防己茯苓汤+抑制剂组。对照组即正常培养的肾小球系膜细胞MES-13细胞加入空白血清处理，其余3组细胞用含30 mmol/L葡萄糖的高糖培养基处理建立糖尿病高糖环境诱导的细胞损伤模型。模型组细胞予空白血清处理，防己茯苓汤组予10%防己茯苓汤含药血清处理，抑制剂组予NLRP3抑制剂MCC950处理，防己茯苓汤+抑制剂组予10%防己茯苓汤含药血清+NLRP3抑制剂MCC950处理，所有细胞均处理48 h。检测各组小鼠空腹血糖（FBS）、血清肌酐（Scr）、血尿素氮（BUN）、尿白蛋白/肌酐比值（UACR）、总胆固醇（TC）、甘油三酯（TG）、低密度脂蛋白胆固醇（LDL-C）；采用HE染色观察小鼠肾组织病理形态；采用Western blot检测小鼠肾组织和肾小球系膜细胞MES-13细胞中焦亡相关蛋白NOD样受体蛋白3（NLRP3）、胱天蛋白酶-1（Caspase-1）、裂解素D（GSDMD）及炎症因子白细胞介素（IL）-1β和IL-18的表达水平；采用透射电子显微镜进一步观察各组MES-13细胞的焦亡形态。结果 动物实验结果示：与对照组相比，模型组小鼠肾组织出现明显糖尿病肾病病理改变，FBS、Scr、BUN、UACR、TC、TG、LDL-C、NLRP3、GSDMD、Caspase-1、IL-1β和IL-18水平均升高（P<0.05，P<0.001）；与模型组相比，防己茯苓汤组和抑制剂组小鼠肾组织病理改变有所改善，FBS、Scr、BUN、UACR、TC、TG、LDL-C、NLRP3、GSDMD、Caspase-1、IL-1β和IL-18水平均降低（P<0.05，P<0.01，P<0.001）；与防己茯苓汤组相比，防己茯苓汤+抑制剂组小鼠肾组织病理形态改善，FBS、Scr、BUN、UACR、TC、TG、LDL-C、NLRP3、GSDMD、Caspase-1、IL-1β和IL-18水平均下降（P<0.05）。细胞实验结果示：与对照组相比，模型组细胞焦亡明显，且NLRP3、GSDMD、Caspase-1、IL-1β和IL-18水平均升高（P<0.01）；与模型组相比，防己茯苓汤组和抑制剂组细胞焦亡有所改善，NLRP3、GSDMD、Caspase-1、IL-1β和IL-18水平均降低（P<0.05）；与防己茯苓汤组相比，防己茯苓汤+抑制剂组细胞焦亡有所改善，NLRP3、GSDMD、Caspase-1、IL-1β和IL-18水平均降低（P<0.05，P<0.01）。结论 防己茯苓汤可能通过抑制NLRP3/Caspase-1通路介导的细胞焦亡，改善糖尿病肾病小鼠的肾功能，从而减少肾组织病理损伤及炎症反应。 |
| 关键词: 糖尿病肾病 防己茯苓汤 细胞焦亡 肾功能 炎症反应 NOD样受体蛋白3 胱天蛋白酶1 |
| DOI：10.3969/j.issn.1674-070X.2025.07.005 |
| 投稿时间：2025-02-17 |
| 基金项目:北京航天医科集团青年项目课题（2022YK15）；国家中医药管理局“全国名老中医药专家传承工作室”建设项目（国中医药人教函〔2022〕75号）；国家自然科学基金青年基金项目（82104842）；湖南省教育厅科学研究项目（19A379）。 |
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| Mechanism of Fangji Fuling Decoction in ameliorating glomerular mesangial cell pyroptosis in diabetic nephropathy |
| TANG Lujun, ZHANG Wen, ZENG Yang, YUAN Liumei, LIU Huihui, HE Haixia, HU Guoheng |
| (Hunan Aerospace Hospital, Changsha, Hunan 410205, China;The First Hospital of Hunan University of Chinese Medicine, Changsha, Hunan 410007, China) |
| Abstract: |
| Objective To investigate the effects of Fangji Fuling Decoction (FJFLD) on renal function and glomerular mesangial cell pyroptosis in diabetic nephropathy (DN) mice, and to explore its underlying mechanism of action. Methods Forty 8-week-old male C57BL/6 mice were randomly divided into control group, model group, FJFLD group, inhibitor group, and FJFLD+inhibitor group, with eight mice in each group. The control group was fed normally and administered an equivalent volume of normal saline by gavage, while the other groups were induced with a high-fat diet and streptozotocin (STZ) injection to establish DN models. After successful modeling, the model group was given an equivalent volume of normal saline by gavage, the FJFLD group was treated with FJFLD via gavage, the inhibitor group received intraperitoneal injection of NLRP3 inhibitor MCC950, and the FJFLD+inhibitor group received both FJFLD by gavage and intraperitoneal injection of MCC950. All groups were continuously treated for eight weeks. MES-13 cells were randomly divided into control group, model group, FJFLD group, inhibitor group, and FJFLD+inhibitor group. The control group (normal cells) was treated with blank serum, while the other groups were exposed to a high-glucose (30 mmol/L) medium to establish DN-induced cell injury models. The model group received blank serum, the FJFLD group received 10% FJFLD-containing serum, the inhibitor group received the NLRP3 inhibitor MCC950, and the FJFLD+inhibitor group received combined 10% FJFLD-containing serum and MCC950. All cells were treated for 48 h. The fasting blood glucose (FBG), serum creatinine (Scr), blood urea nitrogen (BUN), urinary albumin-to-creatinine ratio (UACR), total cholesterol (TC), triglycerides (TG), and low-density lipoprotein cholesterol (LDL-C) levels were measured in mice. Hematoxylin-eosin (HE) staining was performed to observe renal histopathological changes. Western blot was performed to determine the expression levels of pyroptosis-related proteins (NLRP3, Caspase-1, and GSDMD), and inflammatory factors (IL-1β and IL-18) in renal tissue and MES-13 cells. Transmission electron microscopy was used to further observe the pyroptotic morphology of MES-13 cells in each group. Results Animal experiements indicate that, compared with the control group, the model group showed significant DN pathological changes in renal tissue, along with elevated FBS, Scr, BUN, UACR, TC, TG, LDL-C, NLRP3, GSDMD, Caspase-1, IL-1β, and IL-18 levels (P<0.05, P<0.001). Compared with the model group, both FJFLD and inhibitor groups exhibited ameliorated renal histopathology, along with reduced FBS, Scr, BUN, UACR, TC, TG, LDL-C, NLRP3, GSDMD, Caspase-1, IL-1β, and IL-18 levels (P<0.05, P<0.01, P<0.001). Compared with the FJFLD group, the FJFLD+inhibitor group demonstrated improvement in renal histopathology, with lower levels of FBS, Scr, BUN, UACR, TC, TG, LDL-C, NLRP3, GSDMD, Caspase-1, IL-1β, and IL-18 (P<0.05). Cell experiments indicate that compared with the control group, the model group showed obvious pyroptosis and elevated NLRP3, GSDMD, Caspase-1, IL-1β, and IL-18 levels (P<0.01). Compared with the model group, both the FJFLD group and FJFLD+inhibitor group exhibited ameliorated pyroptosis and reduced NLRP3, GSDMD, Caspase-1, IL-1β, and IL-18 levels (P<0.05, P<0.01). Compared with the FJFLD group, the FJFLD+inhibitor group showed ameliorated pyroptosis and reduced NLRP3, GSDMD, Caspase-1, IL-1β, and IL-18 levels (P<0.05, P<0.01). Conclusion FJFLD may improve the renal function of DN mice by inhibiting NLRP3/Caspase-1-mediated pyroptosis, thereby reducing pathological damage and inflammatory response in renal tissue. |
| Key words: diabetic nephropathy Fangji Fuling Decoction pyroptosis renal function inflammatory reaction NOD-like receptor protein 3 Caspase-1 |
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