益肾化湿颗粒改善糖尿病小鼠早期肾小管损伤的实验研究

Experimental study of Yishen Huashi Granule inproving early renal tubular injury in diabetic mice

关键词:  益肾化湿颗粒  db/db小鼠  megalin  cubilin  NGAL  早期肾小管损伤

Key words:  Yishen Huashi Granule  db/db mice  megalin  cubilin  NGAL  early renal tubular injury

摘要:

[目的] 探讨益肾化湿颗粒对自发型2型糖尿病db/db小鼠早期肾小管损伤的作用。[方法] 6~7周龄SPF级雄性自发型2型糖尿病db/db小鼠18只，随机分为模型组、缬沙坦组[13. 33 mg/（kg·d）]、益肾化湿组[5 g/（kg·d）]，每组6只，另设雄性非糖尿病db/m小鼠6只为正常组。正常组及模型组予去离子水10 mL/（kg·d）]灌胃，连续干预12周。代谢笼收集24 h尿液行24 h尿蛋白定量（24 h-UTP）及中性粒细胞明胶酶相关脂质运载蛋白（NGAL）检测。12周后小鼠麻醉取血，全自动生化分析仪检测血清肌酐（Scr）、尿素氮（BUN）、尿酸（UA）、甘油三酯（TG）、低密度脂蛋白（LDL）水平。取肾组织行PAS染色后光镜观察病理改变；免疫组化（IHC）检测肾小管上皮细胞megalin、cubilin蛋白表达强度。[结果] 与模型组比较，益肾化湿颗粒干预后db/db小鼠血液UA、LDL水平下降（P<0.05），24 h-UTP及尿液NGAL水平下降（P<0.05，P<0.01），PAS染色提示系膜基质增生及小管损伤减轻，肾小管上皮细胞megalin、cubilin蛋白表达增加，Image Pro Plus 6.0 软件统计，益肾化湿组肾小球系膜基质指数下降（P<0.01），megalin、cubilin平均光密度值升高（P<0.05）。[结论] 益肾化湿颗粒可以上调db/db小鼠肾小管上megalin、cubilin蛋白的表达从而减少尿蛋白的形成，起到保护肾小管的作用。

Abstract:

[Objective] To investigate the effect of Yishen Huashi Granule on early renal tubular injury in spontaneous type 2 diabetic db/db mice. [Methods] The 18 SPF male spontaneous type 2 diabetic db/db mice aged 6~7 weeks were randomly divided into model group,valsartan group[13.33 mg/(kg·d)] and Yishen Huashi group[5 g/(kg·d)],with 6 mice in each group. Another 6 male non-diabetic db/m mice were set as normal group. The normal group and model group were given 10 mL/(kg·d) deionized water by gavage for 12 weeks. 24 h urine was collected by metabolic cage for 24 h urine protein (24 h-UTP) and neutrophil-gelatinase-associated lipid-carrying proteins (NGAL) detection. After 12 weeks,the mice were anesthetized and blood samples were taken. The levels of Scr,BUN,UA,TG and LDL were detected by automatic biochemical analyzer. The renal pathological morphology changes were observed by light microscope with PAS staining. The expression intensity of megalin and cubilin in renal tissues were detected by immunohistochemistry. [Results] After the intervention of Yishen Huashi Granule,the levels of UA and LDL in blood,24 h-UTP and NGAL in urine of db/db mice were decreased(P<0.05,P<0.01) compared with the model group. PAS staining showed that mesangial matrix proliferation and tubule injury were alleviated,and the protein expression of megalin and cubilin in renal tubular epithelial cells were increased. According to the statistics of Imagepro Plus 6.0 software,the glomerular mesangial matrix index decreased (P<0.01),and the mean optical density of megalin and cubilin increased (P<0.05). [Conclusion] Yishen Huashi Granule can reduce the formation of urine protein by increasing the expression of megalin and cubilin protein on the kidney tubes of db/db mice,and play a role in protecting the renal tube.