摘要
目的探究糖尿病肾病(diabetic kidney disease,DKD)血瘀证大鼠肾损害与肾脏铁死亡的潜在机制。方法将50只SPF级雄性SD大鼠分为对照组、DKD组、DKD血瘀证组。采用腹腔注射链脲佐菌素的方法复制DKD大鼠模型,采用尾静脉注射右旋糖酐的方法复制DKD血瘀证模型。实验过程中观察大鼠血瘀证表现及检测24 h尿蛋白、血清肌酐、血尿素氮、血液流变学指标,采用苏木精-伊红染色、Masson染色、PAS染色观察肾脏的组织形态,采用透射电子显微镜观察铁死亡典型细胞的线粒体变化;采用免疫组织化学法、Western blot法检测肾组织铁死亡相关蛋白[长链酯酰辅酶A合成酶4(Acyl-CoA synthetase long chain family member 4,ACSL4)、谷胱甘肽过氧化物酶4(glutathione peroxidase 4,GPX4)]及肾脏纤维化指标[纤维连接蛋白(fibronectin,FN)、Ⅳ型胶原蛋白(typeⅣcollagen,Col-Ⅳ)]的表达水平。结果与对照组比较,DKD组大鼠肾脏病理变化加重,线粒体损伤明显,24 h尿蛋白含量,血清肌酐、血尿素氮水平及全血黏度、血浆黏度明显升高(P<0.05),肾脏ACSL4、FN和Col-Ⅳ及其mRNA表达水平明显升高(P<0.05),GPX4蛋白及其mRNA表达水平明显降低(P<0.05)。DKD血瘀证大鼠出现唇色黯淡、眼球黯红、耳廓紫红、舌下脉络紫黯等血瘀证表现,24 h尿蛋白和血清肌酐、血尿素氮水平明显高于DKD组,光学显微镜下可见肾脏出现明显的系膜基质增生、肾小球萎缩、肾间质胶原沉积和纤维化,电子显微镜下可见肾组织细胞线粒体损伤明显,嵴基本断裂,ACSL4、FN、Col-Ⅳ及其mRNA表达水平较DKD组明显上升(P<0.05),GPX4蛋白及其mRNA表达水平较DKD组明显下降(P<0.05)。结论DKD血瘀证大鼠肾脏损害更加严重,其机制可能与GPX4、ACSL4介导的铁死亡有关。
Objective To investigate the potential mechanisms of renal damage and renal ferroptosis in rats with diabetic kidney disease(DKD)with blood stasis syndrome.Methods Fifty male specific pathogen-free Sprague-Dawley rats were divided into control group,DKD group,and DKD with blood stasis syndrome group.A rat DKD model was established by intraperitoneal injection of streptozotocin.A rat model of DKD with blood stasis syndrome was established by injecting dextran into the tail vein.The rats were monitored for blood stasis syndrome as well as 24-h urinary protein,serum creatinine,urea nitrogen,and hemorheological indices.The histomorphological changes of renal tissues were observed by hematoxylin and eosin staining,Masson staining,and PAS staining.Transmission electron microscopy was used to observe the mitochondrial changes of cells undergoing typical ferroptosis.Immunohistochemical staining and Western blot were used to measure the expression of ferroptosis-related proteins[acyl-CoA synthetase long-chain family member 4(ACSL4)and glutathione peroxidase 4(GPX4)]and renal fibrosis markers[fibronectin(FN)and typeⅣcollagen(Col-Ⅺ)]in renal tissues.Results Compared with the control group,the DKD group showed more severe renal pathological changes and mitochondrial damage,as well as significantly higher levels of 24-h urinary protein,serum creatinine,urea nitrogen,blood viscosity,and plasma viscosity(all P<0.05),significantly higher expression of ACSL4,FN,and Col-Ⅳprotein and mRNA(all P<0.05),and significantly lower expression of GPX4 protein and mRNA(both P<0.05).The DKD with blood stasis syndrome group developed blood stasis syndrome:dark lips,dark red eyes,reddish-purple ear auricles,and bluish-purple vessels underneath the tongue.Compared with the DKD group,those rats had significantly higher levels of 24-h urinary protein,serum creatinine,and urea nitrogen;renal mesangial matrix expansion,glomerulus atrophy,and renal interstitium collagen deposition and fibrosis under the optical microscope;damaged mitochondria w
作者
韩佳瑞
彭紫凝
王学艺
邢玉凤
朱清
王慧丽
庞欣欣
HAN Jiarui;PENG Zining;WANG Xueyi;XING Yufeng;ZHU Qing;WANG Huili;PANG Xinxin(The Second Clinical Medical College of Henan University of Chinese Medicine,Henan Zhengzhou 450046,China;Department of Nephrology,Henan Provincial Hospital of Traditional Chinese Medicine,Henan Zhengzhou 450002,China)
出处
《安徽中医药大学学报》
CAS
2023年第2期78-84,共7页
Journal of Anhui University of Chinese Medicine
基金
河南省中医药科学研究专项重点课题(20-21ZY1003)
全国中医药创新骨干人才培养项目
河南省中医药拔尖人才培养项目专项课题(2019ZYBJ17)。
