Danggui Buxuetang Alleviates Oxidative Stress and Inflammation in Diabetic Kidney Disease Rats by Improving Mitochondrial Dysfunction of Podocytes

He-chao JIN; Jia-wei QIANG; Guan-wen ZHANG; Sheng-ran LIANG; Deng-zhou GUO

doi:10.13422/j.cnki.syfjx.20212442

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Danggui Buxuetang Alleviates Oxidative Stress and Inflammation in Diabetic Kidney Disease Rats by Improving Mitochondrial Dysfunction of Podocytes

更新时间：2021-12-31

- Danggui Buxuetang Alleviates Oxidative Stress and Inflammation in Diabetic Kidney Disease Rats by Improving Mitochondrial Dysfunction of Podocytes
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 28, Issue 3, Pages: 31-40(2022)
- 作者机构：
  
  1.河北中医学院，石家庄 050200
  2.河北省中医院，石家庄 050011
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.20212442
  CLC： R2-0;R289;R33;R587.1
- Received：02 August 2021，
  
  Published Online：10 November 2021，
  
  Published：05 February 2022
- 稿件说明：
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靳贺超,强家维,张冠文等.当归补血汤通过改善足细胞线粒体功能障碍减轻糖尿病肾病大鼠氧化应激及炎症反应[J].中国实验方剂学杂志,2022,28(03):31-40.

JIN He-chao,QIANG Jia-wei,ZHANG Guan-wen,et al.Danggui Buxuetang Alleviates Oxidative Stress and Inflammation in Diabetic Kidney Disease Rats by Improving Mitochondrial Dysfunction of Podocytes[J].Chinese Journal of Experimental Traditional Medical Formulae,2022,28(03):31-40.
靳贺超,强家维,张冠文等.当归补血汤通过改善足细胞线粒体功能障碍减轻糖尿病肾病大鼠氧化应激及炎症反应[J].中国实验方剂学杂志,2022,28(03):31-40. DOI： 10.13422/j.cnki.syfjx.20212442.

JIN He-chao,QIANG Jia-wei,ZHANG Guan-wen,et al.Danggui Buxuetang Alleviates Oxidative Stress and Inflammation in Diabetic Kidney Disease Rats by Improving Mitochondrial Dysfunction of Podocytes[J].Chinese Journal of Experimental Traditional Medical Formulae,2022,28(03):31-40. DOI： 10.13422/j.cnki.syfjx.20212442.

摘要

目的

探讨当归补血汤通过改善足细胞线粒体功能障碍对糖尿病肾病（DKD）大鼠氧化应激及炎症反应的干预作用。

方法

SD大鼠随机分为正常组和造模组，造模组采用高糖高脂喂养联合小剂量链脲佐菌素（STZ）一次性腹腔注射制备2型糖尿病大鼠模型。成模大鼠随机分为模型组、当归补血汤低剂量（0.72 g·kg

-1

）组、当归补血汤高剂量（1.44 g·kg

-1

）组、厄贝沙坦（0.017 g·kg

-1

）组进行灌胃，正常组及模型组予等体积生理盐水灌胃。药物干预20周后检测各组大鼠尿微量白蛋白与尿肌酐比值（UACR），血清丙二醛（MDA）含量，血清锰超氧化物歧化酶（MnSOD）活性；马松（Masson）染色、高碘酸希夫（PAS）染色分别观察肾组织病理形态学变化；透射电镜（TEM）观察足细胞线粒体超微结构变化；二氢乙锭（DHE）荧光探针检测大鼠肾组织活性氧（ROS）表达水平；免疫组化法（IHC）检测大鼠肾组织过氧化物酶体增殖物激活受体

辅助激活因子-1

（PGC-1

），核苷酸结合域样受体蛋白3（NLRP3），肿瘤蛋白-1（WT-1）蛋白表达水平；免疫荧光（IF）三标法检测NLRP3，白细胞介素-1

（IL-1

）与WT-1在足细胞的表达水平；实时荧光定量聚合酶链式反应（Real-time PCR）检测大鼠肾组织PGC-1

，NLRP3 mRNA表达水平；蛋白免疫印迹法（Western blot）检测大鼠肾组织PGC-1

，MnSOD，NLRP3，IL-1

蛋白表达水平。

结果

与正常组比较，模型组大鼠UACR，MDA含量显著升高，MnSOD活性显著降低（

0.01）；病理表现为肾小球肥大、基底膜增厚、系膜增生、系膜外基质增多、出现K-W结节、足细胞线粒体肿胀、线粒体嵴走向紊乱、模糊或消失，可见空泡形成；ROS表达量显著增多（

0.01）；NLRP3及IL-1

在足细胞表达显著升高，WT-1在足细胞表达显著减少；PGC-1

mRNA表达水平显著降低（

0.01），PGC-1

，MnSOD蛋白表达水平显著降低（

0.01），NLRP3 mRNA表达水平显著升高，NLRP3，IL-1

蛋白表达水平显著升高（

0.01）。与模型组比较，当归补血汤高剂量组明显降低UACR，MDA含量，升高MnSOD活性（

0.05，

0.01）；明显改善肾组织病理学，改善足细胞线粒体超微结构；减少ROS表达（

0.05，

0.01）；NLRP3及IL-1

在足细胞表达减少且WT-1在足细胞表达增多；升高PGC-1

，MnSOD mRNA及蛋白表达水平，降低NLRP3，IL-1

mRNA及蛋白表达水平（

0.05，

0.01）。

结论

当归补血汤可能通过改善DKD大鼠足细胞线粒体功能障碍，缓解氧化应激，减轻炎症反应，起到明显的肾脏保护及延缓病程进展作用。

Abstract

Objective

To investigate the intervention effect of Danggui Buxuetang on oxidative stress and inflammatory response in diabetic kidney disease （DKD） rats from its improvement of podocyte mitochondrial dysfunction.

Method

SD rats were randomly divided into the control group and modeling group， and the ones in the latter group rats were fed a high-glucose and high-fat diet and then intraperitoneally injected with a small dose of streptozotocin （STZ） for inducing type 2 diabetes. The successfully modeled rats were randomized into the model group， high- and low-dose （1.44 and 0.72 g·kg

-1

） Danggui Buxuetang groups， and irbesartan （0.017 g·kg

-1

）group and gavaged with the corresponding drugs， while those in the normal and model groups with an equal volume of normal saline. After 20 weeks of drug intervention， the urinary microalbumin-to-urine creatinine ratio （UACR） and serum malondialdehyde （MDA） content and manganese superoxide dismutase （MnSOD） activity in each group were measured. The pathological changes in renal tissue were observed by Masson trichrome staining， and periodic acid-silver metheramine （PASM） staining， followed by the observation of ultrastructural changes in podocytes under the transmission electron microscope （TEM）. The expression level of reactive oxygen species （ROS） in rat kidney tissue was detected using a fluorescent probe dihydroethidium （DHE）. The protein expression levels of peroxisome proliferator-activated receptor

-coactivator -1

（PGC-1

）， nucleotide-binding domain like receptor protein 3 （NLRP3）， and Wilms tumor protein-1 （WT-1） were measured by immunohistochemistry （IHC）， and the expression levels of NLRP3， interleukin-1

（IL-1

），and WT-1 in podocytes by immunofluorescence （IF） assay. The mRNA expression levels of PGC-1

and NLRP3 in the renal tissues were determined by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）， and the protein expression levels of PGC-1

， MnSOD， NLRP3， and IL-1

were assayed by Western blot.

Result

Compared with the normal group， the model group exhibited elevated UACR and MDA content， weakened MnSOD activity （

0.01）， glomerular hypertrophy， thickened basement membrane， mesangial hyperplasia， increased extracellular matrix， K-W nodules， podocyte mitochondrial swelling， disordered mitochondrial cristae， foot process fusion or loss， vacuolization， increased ROS （

0.01）， enhanced NLRP3 and IL-1

but diminished WT-1 expression in podocytes， down-regulated PGC-1

mRNA expression （

0.01） and PGC-1

and MnSOD protein expression （

0.01）， and up-regulated NLRP3 mRNA expression and NLRP3 and IL-1

protein expression （

0.01）. Compared with the model group， Danggui Buxuetang high-dose group significantly decreased UACR and MDA， enhanced MnSOD activity （

0.05，

0.01）， improved renal histopathology and podocyte mitochondrial ultrastructure， decreased ROS （

0.05，

0.01） and NLRP3 and IL-1

expression in podocytes， enhanced WT-1 expression in podocytes， up-regulated the mRNA and protein levels of PGC-1

and MnSOD， and down-regulated the mRNA and protein levels of NLRP3 and IL-1

（

0.05，

0.01）.

Conclusion

Danggui Buxuetang alleviates oxidative stress， reduces inflammatory response， protects kidney， and delays the progression of DKD possibly by improving the mitochondrial dysfunction in podocytes of DKD rats.

关键词

Keywords

references

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