抗微小核糖核酸-21治疗对糖尿病肾病模型小鼠肾脏损伤作用的实验研究  

Experimental study on the effect of anti-microRNA-21 treatment on renal injury in a diabetic nephropathy mouse model

作　　者：孙佳 沈军 SUN Jia;SHEN Jun(Department of Nephrology,the First People's Hospital of Linping District,Hangzhou,Zhejiang 311100,China)

机构地区：[1]杭州市临平区第一人民医院肾内科,杭州311100

出　　处：《浙江中西医结合杂志》2025年第1期13-18,共6页Zhejiang Journal of Integrated Traditional Chinese and Western Medicine

基　　金：余杭区医疗卫生科研项目(No.2019006)。

摘　　要：目的 探讨抗微小核糖核酸-21 (microRNA-21,miR-21)治疗对糖尿病肾病(diabetic nephropathy,DN)模型小鼠肾脏损伤的作用。方法 8周龄C57BL/6小鼠随机分为对照组、DN组、DN+miR-21反义寡核苷酸(antisense oligonucleotides,ASO)组(DN+miR-21-ASO组)、DN+miR-21反义寡核苷酸错配碱基对(MM-ASO)组(DN+miR-21-MM-ASO组)。通过腹腔注射链脲佐菌素构建DN小鼠模型。全自动生化分析仪检测小鼠24 h尿蛋白(urine protein,Upro)、血清肌酐(serum creatinine,Scr)和血尿素氮(blood urea nitrogen,BUN)含量。PAS染色观察小鼠肾脏糖原沉积,苏木素-伊红(HE)染色观察小鼠肾脏病理学变化,TUNEL染色观察小鼠肾脏组织细胞凋亡情况。实时定量PCR检测小鼠肾脏组织miR-21的相对表达水平。蛋白质印迹(Western blot)检测B淋巴细胞瘤-2基因(Bcl-2)、Bcl-2相关X蛋白(Bax)和活化型半胱氨酸天冬氨酸蛋白酶-3(cleaved caspase-3)蛋白的表达水平。结果 与对照组比较,DN组小鼠肾脏组织miR-21表达显著上调[(3.23±0.24)比(1.00±0.11),P<0.01],24 h Upro、Scr和BUN[(53.87±5.87)mg比(13.83±1.80)mg、(89.35±3.34)μmol/L比(28.27±2.91)μmol/L、(25.79±2.15)mmol/L比(7.36±0.50)mmol/L,P<0.01]水平均显著增高;与DN+miR-21-MM-ASO组比较,DN+miR-21-ASO组小鼠肾脏组织miR-21的表达受到抑制[(0.77±0.11)比(3.52±0.27),P<0.01],小鼠24 h Upro、Scr和BUN[(29.92±6.06)mg比(50.69±6.93)mg、(68.39±3.75)μmol/L比(87.84±6.63)μmol/L、(17.66±1.04)mmol/L比(23.80±3.16)mmol/L,P<0.01]水平明显降低。PAS染色显示,与对照组比较,DN组小鼠肾脏组织系膜和基底膜区PAS染色阳性物质增多,颜色深;与DN+miR-21-MM-ASO比较,DN+miR-21-ASO组小鼠肾脏组织系膜和基底膜区PAS染色阳性物质明显减少且红染区域较少。HE染色表明,与对照组比较,DN组小鼠出现肾小球体积增大和肾小管刷状缘脱离等肾脏损伤表现;与DN+miR-21-MM-ASO组比较,DN+miR-21-ASO组小鼠肾脏损伤显著改善。TUNEL染色表明,与�Objective To investigate the effect of anti-microRNA-21(miR-21)treatment on renal injury in a diabetic nephropathy(DN)mouse model.Methods Eight-week-old C57BL/6 mice were randomly divided into the control group,DN group,DN+miR-21 antisense oligonucleotides(ASO)group(DN+miR-21-ASO group),and DN+miR-21 antisense oligonucleotides mismatch base pair(miR-21-mismatched-ASO)group(DN+miR-21-MM-ASO group).DN mouse model was established by intraperitoneal injection of streptozotocin.The 24 h urinary protein(Upro),serum creatinine(Scr),and blood urea nitrogen(BUN)levels in mice were detected using a fully automatic biochemical analyzer.Periodic acid-Schiff(PAS)staining was used to observe glycogen deposition in renal tissues of mice,hematoxylin-eosin(HE)staining was used to observe pathological changes in renal tissues of mice,and TUNEL staining was used to observe apoptosis in renal tissue cells of mice.The relative expression of miR-21 in renal tissue of mice was quantified using real-time quantitative PCR.The protein expression levels of B-cell lymphoma-2(Bcl-2),Bcl-2 associated X protein(Bax),and cleaved caspase-3 were detected using Western blotting.Results Compared with the control group,the DN group showed significantly upregulated expression of miR-21 in renal tissue[(3.23±0.24)vs.(1.00±0.11),P<0.01],increased 24 h Upro[(53.87±5.87)mg vs.(13.83±1.80)mg,P<0.01],Scr[(89.35±3.34)μmol/L vs.(28.27±2.91)μmol/L,P<0.01],and BUN levels[(25.79±2.15)mmol/L vs.(7.36±0.50)mmol/L,P<0.01].Compared with DN+miR-21-MM-ASO group,the DN+miR-21-ASO group showed inhibited expression of miR-21 in kidney tissue[(0.77±0.11)vs.(3.52±0.27),P<0.01],and significantly decreased 24 h Upro[(29.92±6.06)mg vs.(50.69±6.93)mg,P<0.01],Scr[(68.39±3.75)μmol/L vs.(87.84±6.63)μmol/L,P<0.01]and BUN levels[(17.66±1.04)mmol/L vs.(23.80±3.16)mmol/L,P<0.01].Compared with the control group,PAS staining showed that the DN group mice had increased PAS-positive substances in the mesangial and basement membrane regions of renal tissue,with a dark

关 键 词：小鼠 糖尿病肾病 肾脏损伤 微小核糖核酸-21 细胞凋亡 

分 类 号：R692.9[医药卫生—泌尿科学] R-332[医药卫生—外科学] R587.2[医药卫生—临床医学]