核因子κB抑制剂小白菊内酯改善2型糖尿病肾病肾小管白蛋白摄取

郝秋发; 王宝宝; 张薇; 邱维; 刘倩伶; 李雪梅

doi:10.24920/003573

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核因子κB抑制剂小白菊内酯改善2型糖尿病肾病肾小管白蛋白摄取

中国医学科学杂志（英文版） 2020年35卷第1期页码：31-42

Original Article | Updated：2024-04-10

- 核因子κB抑制剂小白菊内酯改善2型糖尿病肾病肾小管白蛋白摄取
- Affiliations：
  
  Department of Nephrology, Peking Union Medical College Hospital, Chinese Academy of Medicine Sciences & Peking Union Medical College, Beijing 100730, China
- Author bio：
  
  * Tel: 86-10-69154056, Fax: 86-10-69155058, E-mail: professorliunion@163.com
- Funds：
  
  This study was supported by the Natural Science Foundation of Beijing, China(Grant No. 7122143)
- DOI：10.24920/003573
  中图分类号：
- 收稿日期：2019-04-02，
  
  网络出版日期：2020-03-31，
  
  纸质出版日期：2020-02-29
- Accepted：
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郝秋发, 王宝宝, 张薇, 等. 核因子κB抑制剂小白菊内酯改善2型糖尿病肾病肾小管白蛋白摄取[J]. 中国医学科学杂志（英文版）, 2020,35(1):31-42.

Hao Qiufa, Wang Baobao, Zhang Wei, et al. NF-κB Inhibitor Parthenolide Promotes Renal Tubules Albumin Uptake in Type 2 Diabetic Nephropathy[J]. Chinese medical sciences journal, 2020, 35(1): 31-42.
郝秋发, 王宝宝, 张薇, 等. 核因子κB抑制剂小白菊内酯改善2型糖尿病肾病肾小管白蛋白摄取[J]. 中国医学科学杂志（英文版）, 2020,35(1):31-42. DOI： 10.24920/003573.

Hao Qiufa, Wang Baobao, Zhang Wei, et al. NF-κB Inhibitor Parthenolide Promotes Renal Tubules Albumin Uptake in Type 2 Diabetic Nephropathy[J]. Chinese medical sciences journal, 2020, 35(1): 31-42. DOI： 10.24920/003573.

摘要

目的

在糖尿病肾病早期

尿白蛋白增多由肾小管重吸收功能受损所致

然而其机制尚未完全阐明。本研究旨在探究减轻炎症反应并改善胰岛素信号受损能否改善糖尿病肾病肾小管对白蛋白的摄取。

方法

8周龄雄性db/db小鼠（

=8）作为2型糖尿病肾病模型

给予核因子κB（NF-κB）抑制剂小白菊内酯（PTN）治疗

按照1 mg/kg（小鼠体重）隔日1次（连续8周）腹腔注射给药。同时给予同龄db/m组（

=5）及db/db组（

=8）小鼠等剂量的生理盐水溶解液

作为正常对照组和2型糖尿病肾病组。8周后处死小鼠

留取血和尿标本检测稳态模型胰岛素抵抗指数（HOMA-IR）和尿白蛋白肌酐比。留取肾脏组织

利用PAS染色分析肾脏组织病理

利用免疫组化和免疫印迹检测NF-κB p65、p-AKT、amnionless和cubilin表达水平

并利用免疫荧光检测肾小管摄取白蛋白情况。HKC细胞被分为胰岛素组（胰岛素干预）、TNF-α组（胰岛素和TNF-α干预）、TNF-α+PTN组（胰岛素、TNF-α和PTN干预）。对HKC细胞的白蛋白摄取水平和NF-κB p65

p-IRS-1/IRS-1

p-AKT/AKT

amnionless及cubilin表达水平进行检测。

结果

与db/db组相比

db/db+PTN组HOMA-IR水平下降（36.83±14.09比31.07±28.05）

尿白蛋白肌酐比下降（190.3±7.3比143.0±97.6 mg/mmol）

但差异无统计学意义（

0.05）。PAS染色显示PTN能够减轻db/db小鼠肾小球肥大

减少系膜区基质。与db/m组相比

db/db组小鼠肾脏NF-κB p65表达增加

p-AKT（s473）水平减低（

0.05）。PTN能够显著降低db/db组NF-κB p65表达

改善p-AKT（s473）下降水平（

0.05）。与db/m组相比

db/db小鼠肾脏amnionless和cubilin的表达水平降低（

0.05）

肾小管白蛋白摄取减少

而PTN能够显著改善db/db小鼠cubilin的降低（

0.05）

改善肾小管白蛋白摄取。胰岛素可促进HKC细胞amnionless和cubilin表达增多

促进白蛋白摄取增加（

0.05）。TNF-α刺激HKC细胞NF-κB p65表达增加

p-IRS-1（s307）水平增高及p-AKT（s473）水平减低（

0.05）。与TNF-α组相比

TNF-α+PTN组HKC细胞NF-κB p65表达减低

并逆转p-IRS-1（s307）和p-AKT（s473）的表达变化（

0.05）。TNF-α减少HKC细胞amnionless和cubilin的表达水平（

0.05）

而PTN显著改善cubilin的降低（

0.05）。

结论

炎症损伤胰岛素信号通路

进而减少amnionless-cubilin表达和白蛋白摄取。PTN能够减轻炎症反应

修复胰岛素信号通路

从而改善cubilin表达和白蛋白摄取。本研究阐述了炎症在2型糖尿病肾病中降低肾小管摄取白蛋白的作用。

Abstract

Objective

Injured tubular reabsorption is highlighted as one of the causes of increased albuminuria in the early stage of diabetic nephropathy; however

the underlying mechanism has not been fully elucidated. In this study

we aimed to explore whether reducing inflammation and remodeling the insulin signaling pathway could improve albumin uptake of renal tubules.

Methods

8-week-old male db/db mice (

=8)

a type 2 diabetic nephropathy model

administered with nuclear factor kappa-B (NF-κB) inhibitor parthenolide (PTN

1 mg/kg) intraperitoneally every other day for 8 weeks

were as the treatment group. Meanwhile

the age-matched male db/m mice (

=5) and db/db mice (

=8) were treated with saline as the control group and type 2 diabetic nephropathy group. When the mice were sacrificed

blood and urine were collected to examine homeostasis model assessment of insulin resistance (HOMA-IR) and urine albumin creatinine ratio

and kidney samples were used to analyze histopathologic changes with periodic acid-Schiff (PAS) staining

NF-κB p65

phosphorylation of AKT (p-AKT)

amnionless and cubilin expressions with immunohistochemistry as well as western blot

and the albumin uptake of renal tubules by using immunofluorescence. In addition

HKC cells were divided into the insulin group treated with insulin alone

the TNF-α group treated with insulin and tumor necrosis factor (TNF-α)

and the TNF-α+PTN group exposed to PTN

insulin and TNF-α. The levels of albumin uptake and expression levels of NF-κB p65

p-IRS-1/IRS-1

p-AKT/AKT

amnionless and cubilin in HKC cells were measured.

Results

Compared with the db/db group

the db/db+PTN group demonstrated decreased levels of HOMA-IR (36.83±14.09

. 31.07±28.05) and urine albumin creatinine ratio (190.3±7.3

. 143.0±97.6 mg/mmol); however

the differences were not statistically significant (

0.05). Periodic acid-Schiff staining showed PTN could alleviate the glomerular hypertrophy and reduce the matrix in mesangial areas of db/db mice. The renal expression of NF-κB p65 was increased and p-AKT (s473) decreased in the db/db group compared with the db/m group (

0.05). PTN significantly reduced the renal expression of NF-κB p65 and ameliorated the decline of p-AKT (s473) compared with the db/db group (

0.05). Compared with the db/m group

the expression of amnionless and cubilin decreased and albumin uptake in tubules were reduced in the db/db group (

0.05)

and PTN could significantly increase the expression of cubilin (

0.05)

and improve albumin uptake in tubules. Insulin promoted albumin uptake and the expression of amnionless and cubilin in HKC cells (

0.05). TNF-α stimulated the expression of NF-κB p65

increased p-IRS-1 (s307) and reduced p-AKT (s473) in HKC cells (

0.05). In the TNF-α+PTN group

the expression of NF-κB p65 declined and p-IRS-1 (s307) and p-AKT (s473) were restored

compared with the TNF-α group (

0.05). The expression of amnionless and cubilin decreased in the TNF-α group (

0.05)

and PTN could significantly increase the expression of cubilin (

0.05).

Conclusions

Inflammation caused damage to insulin signaling

which reduced amnionless-cubilin expression and albumin uptake. PTN could reduce inflammation and remodel the impaired insulin signaling pathway

which promoted the expression of cubilin and albumin uptake. Our study can shed light on the role of inflammation in the reduction of albumin uptake of renal tubules in type 2 diabetic nephropathy.

关键词

Keywords

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