Sestrin2 protects rat cardiomyocytes from hypoxia-reoxygenation injury through Nrf2/HO-1 signaling pathway

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Sestrin2 protects rat cardiomyocytes from hypoxia-reoxygenation injury through Nrf2/HO-1 signaling pathway

Basic Medicine | 更新时间：2026-01-09

- Sestrin2 protects rat cardiomyocytes from hypoxia-reoxygenation injury through Nrf2/HO-1 signaling pathway
- Journal of Third Military Medical University Vol. 42, Issue 15, Pages: 1536-1542(2020)
- 作者机构：
  
  1.401120 重庆，重庆医科大学附属第三医院心血管疾病中心
  2.400038 重庆，陆军军医大学(第三军医大学)基础医学院生物医学分析测试中心
- 作者简介：
- 基金信息：
- DOI：10.16016/j.1000-5404.202003201
  CLC： R322.11;R341;R364.4
- Received：16 March 2020，
  
  Revised：2020-05-09，
  
  Published：15 August 2020
- 稿件说明：
移动端阅览
Yingcun LIU, Fei LI, Yi HUANG, et al. Sestrin2 protects rat cardiomyocytes from hypoxia-reoxygenation injury through Nrf2/HO-1 signaling pathway[J]. Journal of Third Military Medical University, 2020, 42(15): 1536-1542.
DOI：

Yingcun LIU, Fei LI, Yi HUANG, et al. Sestrin2 protects rat cardiomyocytes from hypoxia-reoxygenation injury through Nrf2/HO-1 signaling pathway[J]. Journal of Third Military Medical University, 2020, 42(15): 1536-1542. DOI： 10.16016/j.1000-5404.202003201.

摘要

目的

2

探讨Sestrin2蛋白在大鼠心肌细胞缺氧复氧损伤中的作用及其机制。

方法

2

体外培养H9C2心肌细胞，构建心肌细胞缺氧复氧模型。细胞分为对照组、缺氧4 h组、复氧2 h组和复氧4 h组。采用CCK-8法检测细胞存活率；流式细胞术结合Annexin V/7AAD荧光染色检测细胞凋亡；微量酶标法检测心肌细胞损伤标记物乳酸脱氢酶(lactate dehydrogenase

LDH)水平。构建针对Sestrin2蛋白沉默的siRNA与其阴性对照(si-NC)，利用脂质体转染法导入H9C2心肌细胞，后将H9C2心肌细胞分为对照组、缺氧4 h+si-NC组(阴性对照组)、缺氧4 h+siRNA组、复氧4 h+si-NC(阴性对照组)、复氧4 h+siRNA组。Western blot检测Sestrin2蛋白的表达；流式细胞术结合Annexin V/7AAD荧光染色检测细胞凋亡；Western blot检测Sestrin2蛋白、核转录相关因子2(nuclear factor erythroid2-related factor 2，Nrf2)及血红素加氧酶-1(heme oxygenase-1，HO-1)的表达。

结果

2

与对照组比较，复氧4 h组细胞存活率降低(

P

<

0.01)

细胞凋亡率增加(

P

<

0.05)

LDH水平明显升高(

P

<

0.01)。与对照组比较，Sestrin2蛋白表达在缺氧组和缺氧复氧组均显著增高(

P

<

0.01)，且缺氧复氧组Sestrin2蛋白表达量显著高于单纯缺氧组(

P

<

0.01)。转染siRNA特异有效地下调了Sestrin2蛋白表达(

P

<

0.05)，同时细胞凋亡率显著增加(

P

<

0.05)，Nrf2蛋白表达降低(

P

<

0.01)，HO-1蛋白表达降低(

P

<

0.05)。

结论

2

Sestrin2蛋白对缺氧复氧诱导的H9C2心肌细胞氧化损伤有保护作用，其机制可能与激活Nrf2/HO-1相关信号通路有关。

Abstract

Objective

2

To investigate the role and underlying mechanism of Sestrin2 in hypoxia/reoxygenation injury of rat cardiomyocytes.

Methods

2

H9C2 cardiomyocytes were cultured

in vitro

and inflicted to establish hypoxia/reoxygenation injury (H/R) model. Thus

the cells were divided into control group

hypoxia 4-hour group (H4h group)

and hypoxia 4-hour + reoxygenation 2- and 4-hour groups (H4hR2h and H4hR4h groups). Cell survival rate was measured by cell counting kit-8 (CCK8) assay; Annexin V/7AAD staining with flow cytometry was used to detect cell apoptosis. Micro-enzymatic method was used to detect the content of myocardial injury marker

lactate dehydrogenase (LDH). Then H9C2 cardiomyocytes were divided into normal control group

hypoxia 4-hour+si-NC group (negative control)

hypoxia 4-hour+Sestrin2 siRNA group

hypoxia/reoxygenation 4 hour+si-NC group and hypoxia/reoxygenation 4 h+Sestrin2 siRNA group. Recombinant siRNA for Sestrin2 silencing and negative control (si-NC) were constructed and transfected into H9C2 cardiomyocytes with liposomes. The expression of Sestrin2

Nrf2 and HO-1 proteins was detected by Western blotting. Flow cytometry was used to measure the cell apoptosis.

Results

2

The cell viability of cardiomyocytes was significantly lower (

P

<

0.05)

and the apoptosis (

P

<

0.05) as well as LDH level (

P

<

0.01) was obviously increased in the H4hR4h group than the control group. The expression of Sestrin2 protein was increased in both hypoxia and reoxygenation groups when compared with the control group (

P

<

0.01)

with the level in the H/R groups higher than that in the H groups (

P

<

0.01). Transfected of Sestrin2 siRNA significantly knocked down the protein expression (

P

<

0.

05)

enhanced the cell apoptosis (

P

<

0.05)

and decreased the expression levels of Nrf2 (

P

<

0.01) and HO-1 (

P

<

0.05).

Conclusion

2

Sestrin2 exerts protective effect on oxidative damage in H9C2 cardiomyocytes induced by H/R injury

which might be due to its activating Nrf2/HO-1 related signaling pathway.

关键词

Keywords

references

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