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目的：探讨大黄甘草汤通过调控沉默信息调节因子3/PTEN诱导激酶1/E3泛素连接酶(Sirt3/Pink1/Parkin)通路，干预线粒体自噬以改善铁死亡，治疗顺铂诱导的急性肾损伤(AKI)的作用机制。方法：将36只C57BL/6小鼠随机分成正常对照组、模型对照组、大黄甘草汤1、2、4g/kg组、地塞米松0.5mg/kg组，每组6只。除正常对照组外，其余每组连续给予药物灌胃预处理3d后，以顺铂腹腔注射建立AKI模型。检测血清肌酐(SCr)、血尿素氮(BUN)评估肾功能；苏木素-伊红(HE)、过碘酸-雪夫(PAS)染色评估肾脏病理；酶联免疫吸附测定法(ELISA)检测血清炎症因子肿瘤坏死因子(TNF-α)、白细胞介素-1β(IL-1β)、IL-6以评估全身炎症反应；免疫组化检测肾组织肾损伤因子1(KIM-1)、中性粒细胞明胶酶相关载脂蛋白(NGAL)的表达，以评估肾小管损伤程度；比色法检测谷胱甘肽(GSH)、丙二醛(MDA)、超氧化物歧化酶(SOD)含量；免疫组化检测4-羟基壬烯醛(4-HNE)、普鲁士蓝染色评估脂质过氧化和铁代谢水平；利用透射电镜观察线粒体超微结构；采用Westernblot法检测Sirt3、Pink1、Parkin蛋白表达。结果：与正常对照组小鼠相比，模型对照组小鼠血清SCr、BUN、TNF-α、IL-1β、IL-6含量显著升高(P＜0.01或P＜0.05)；肾组织MDA含量显著升高，SOD、GSH含量显著降低(P<0.01)；肾组织病理损伤严重；肾小管胞质及管腔内KIM-1与NGAL表达显著上调；电镜下可见线粒体皱缩、双层膜密度增高及嵴减少或消失等铁死亡特征，且线粒体数量明显减少；肾组织Sirt3、Pink1、Parkin蛋白表达明显下调；与模型对照组相比较，大黄甘草汤各剂量组以及地塞米松0.5mg/kg组血清SCr、BUN、TNF-α、IL-1β、IL-6及肾组织MDA含量降低，SOD、GSH含量显著升高(P<0.05或P＜0.01)；小鼠肾组织病理损伤得到明显缓解；肾小管KIM-1与NGAL 蛋白阳性表达显著降低(P<0.05或P＜0.01)；电镜下线粒体铁死亡特征减轻，嵴结构恢复且数量增多；肾组织Sirt3、Pink1、Parkin蛋白表达上调 (P＜0.05或P＜0.01)。结论：大黄甘草汤对顺铂诱导的AKI小鼠具有改善作用，该保护作用可能通过激活Sirt3/Pink1/Parkin信号通路抑制肾脏细胞铁死亡发挥作用。
Abstract：Objective: To investigate the mechanism through which Dahuang Gancao (大黄甘草) Decoction (DHGC) treats cisplatin-induced acute kidney injury (AKI) by regulating the sirtuin 3 (Sirt3)/PTEN-induced kinase 1 (Pink1)/E3 ubiquitin-protein ligase (Parkin) pathway to modulate mitophagy and ferroptosis. Methods: Thirty-six C57BL/6 mice were randomized into a normal control group， a model control group， DHGC (1， 2， and 4 g/kg) groups， and a dexamethasone (0.5 mg/kg) group， with 6 mice in each group. Except the normal control group， the other groups were pretreated with corresponding drugs by gavage for 3 days. The AKI model was established by intraperitoneal injection of cisplatin. Serum creatinine (SCr) and blood urea nitrogen (BUN) levels were measured to evaluate the renal function. Renal pathology was evaluated by hematoxylin-eosin (HE) and periodic acid-Schiff (PAS) staining. Serum levels of inflammatory factors including tumor necrosis factor-α (TNF-α)， interleukin-1β (IL-1β)， and interleukin-6 (IL-6) were quantified by enzyme-linked immunosorbent assay (ELISA). The expression of kidney injury molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL) in the renal tissue was detected by immunohistochemistry (IHC) to evaluate the degree of renal tubular injury. The levels of glutathione (GSH)， malondialdehyde (MDA)， and superoxide dismutase (SOD) were determined by the colorimetric assay. The 4-hydroxynonenal (4-HNE) expression was assessed by IHC， while the levels of lipid peroxidation and iron metabolism were determined by Prussian blue staining. Mitochondrial ultrastructure was observed by transmission electron microscopy (TEM). The protein levels of Sirt3， Pink1， and Parkin were determined by Western blot. Results: Compared with the normal control group， the model control group showed elevated serum levels of SCr， BUN， TNF-α， IL-1β， and IL-6 (P<0.01 or P<0.05)， increased content of MDA and decreased content of SOD and GSH in the renal tissue (P<0.01)， severe pathological damage in the renal tissue， markedly upregulated expression of KIM-1 and NGAL in the tubular cytoplasm and lumina， distinct ferroptosis-like changes of mitochondria， including mitochondrial shrinkage and cristae dissolution， with reducing total number， and significantly down-regulated protein levels of Sirt3， Pink1， and Parkin in the renal tissue. Compared with the model control group， all doses of DHGC and 0.5 mg/kg dexamethasone lowered the serum levels of SCr， BUN， TNF-α， IL-1β， and IL-6 and the renal level of MDA， increased the renal levels of SOD and GSH (P<0.01 or P<0.05)， alleviated renal pathological damage， reduced the positive expression of KIM-1 and NGAL in renal tubules (P<0.01 or P<0.05)， alleviated ferroptotic morphology in mitochondria， characterized by restored crista integrity and increased mitochondrial number， and up-regulated the protein levels of Sirt3， Pink1， and Parkin in the renal tissue (P<0.01 or P<0.05). Conclusion: DHGC has ameliorative effect on cisplatin-induced AKI in mice. It may exert the protective effect by activating the Sirt3/Pink1/Parkin signaling pathway to inhibit renal cell ferroptosis.
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基本信息:
DOI：10.13412/j.cnki.zyyl.20260716.004
中图分类号:R285.5
引用信息:
[1]熊惠颖,崔梦迪,张子唯,等.大黄甘草汤通过调控Sirt3/Pink1/Parkin信号通路缓解急性肾损伤的机制研究[J].中药药理与临床().DOI:10.13412/j.cnki.zyyl.20260716.004.
基金信息:
国家自然科学基金青年基金项目（编号：82405273）; 湖北省自然科学基金青年项目（编号：2023AFB348）
2026-07-16
2026-07-16
2026-07-16