Mechanism of miR-370-3p/SESN2 regulating lipopolysaccharide-induced apoptosis of mouse alveolar epithelial cells

Wu Zhouyou; Li Ting; Zhang Tengwei; Fang Qiaoyan; Yang Liu; Li Qiao

doi:10.13406/j.cnki.cyxb.003590

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Mechanism of miR-370-3p/SESN2 regulating lipopolysaccharide-induced apoptosis of mouse alveolar epithelial cells

Basic research | 更新时间：2025-10-21

- Mechanism of miR-370-3p/SESN2 regulating lipopolysaccharide-induced apoptosis of mouse alveolar epithelial cells
- Journal of Chongqing Medical University Vol. 49, Issue 9, Pages: 1121-1128(2024)
- 作者机构：
  
  湖南省妇幼保健院新生儿一科，长沙 410008
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.003590
  CLC： R563
- Received：27 July 2023，
  
  Revised：2024-02-19，
  
  Accepted：01 March 2024，
  
  Published：28 September 2024
- 稿件说明：
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武周游,李婷,张腾伟,等.miR-370-3p/SESN2调控脂多糖诱导的小鼠肺泡上皮细胞凋亡的机制研究[J].重庆医科大学学报,2024,49(9):1121-1128.

Wu Zhouyou,Li Ting,Zhang Tengwei,et al.Mechanism of miR-370-3p/SESN2 regulating lipopolysaccharide-induced apoptosis of mouse alveolar epithelial cells[J].Journal of Chongqing Medical University,2024,49(9):1121-1128.
武周游,李婷,张腾伟,等.miR-370-3p/SESN2调控脂多糖诱导的小鼠肺泡上皮细胞凋亡的机制研究[J].重庆医科大学学报,2024,49(9):1121-1128. DOI： 10.13406/j.cnki.cyxb.003590.

Wu Zhouyou,Li Ting,Zhang Tengwei,et al.Mechanism of miR-370-3p/SESN2 regulating lipopolysaccharide-induced apoptosis of mouse alveolar epithelial cells[J].Journal of Chongqing Medical University,2024,49(9):1121-1128. DOI： 10.13406/j.cnki.cyxb.003590.

摘要

目的

探讨miR-370-3p/Sestrin2（SESN2）调控脂多糖（lipopolysaccharide，LPS）诱导的小鼠肺泡上皮细胞（alveolar epithelial cells，AECs）凋亡的机制。

方法

将C57BL/6J小鼠随机分为LPS+anti-NC组和LPS + miR-370-3p抑制剂（anti-miR-370-3p）组，每组12只。通过气管内注射LPS（5 mg/kg）来诱导急性肺损伤（acute lung injury，ALI）。分别通过小鼠尾静脉注射anti-miR-370-3p或anti-NC。MLE12细胞随机分为对照组（Con+anti-NC）、Con+anti-miR-370-3p组、LPS+anti-NC组、LPS+anti-miR-370-3p组、LPS+si-SESN2+anti-miR-370-3p组。使用定量实时PCR分析肺组织或细胞中miR-370-3p。通过测量线粒体形态学和线粒体膜电位考察miR-370-3p敲低对LPS诱导的线粒体功能障碍的保护作用。双荧光素酶报告分析证实了miR-370-3p和SESN2之间的靶向关系。

结果

与LPS+anti-NC组相比，LPS+anti-miR-370-3p组肺组织中miR-370-3p表达、丝氨酸苏氨酸激酶3（receptor-interacting serine/threonine-protein kinase 3，RIPK3）、混合系列蛋白激酶样结构域（mixed lineage kinase domain-Like，MLKL

）蛋白水平、肺部炎症评分明显降低（

0.05），和肺组织中肺表面活性蛋白C（surfactant protein C，SPC）蛋白水平明显增加（

0.001）。与LPS+anti-NC组相比，LPS+anti-miR-370-3p组MLE12细胞中RIPK3、MLKL蛋白表达、PI阳性细胞百分比明显降低（

0.05），并且线粒体片段化和线粒体膜电位明显增加（

0.05）。miR-370-3p对SESN2-WT报道基因的荧光素酶活性具有抑制作用。与LPS+anti-miR-370-3p组相比，LPS+si-SESN2+anti-miR-370-3p组线粒体片段化和线粒体膜电位均明显降低（

0.05）。

结论

miR-370-3p/SESN2轴通过介导线粒体断裂和损害线粒体功能促进LPS诱导的ALI期间AECs坏死性凋亡。

Abstract

Objective

To investigate the mechanism of miR-370-3p/Sestrin2（SESN2） regulating the apoptosis of mouse alveolar epithelial cells（AECs） induced by lipopolysaccharide（LPS）.

Methods

C57BL/6J mice were randomly divided into LPS+anti-NC group and LPS+miR-370-3p inhibitor（anti-miR-370-3p） group，with 12 mice in each group. LPS（5 mg/kg） was given by intratracheal injection to induce acute lung injury（ALI），and anti-miR-370-3p or anti-NC was injected via the caudal vein of mice. MLE12 cells were randomly divided into control group（Con+anti-NC），Con+anti-miR-370-3p group，LPS+anti-NC group，LPS+anti-miR-370-3p group，and LPS+si-SESN2+anti-miR-370-3p group. Quantitative real-time PCR was used to analyze miR-370-3p in lung tissue or cells. Mitochondrial morphology and mitochondrial membrane potential were measured to investigate the protective effect of miR-370-3p knockdown against LPS-induced mitochondrial dysfunction. Dual-luciferase reporter assay confirmed the targeting relationship between miR-370-3p and SESN2.

Results

Compared with the LPS+anti-NC group，the LPS+anti-miR-370-3p group had significant reductions in the expression of miR-370-3p and the protein expression levels of receptor-interacting serine/threonine-protein kinase 3（RIPK3） and mixed lineage kinase domain-like（MLKL） in lung tissue，as well as a significant reduction in lung inflammation score（

0.05） and a significant increase in the protein expression level of surfactant protein C（SPC） in lung tissue（

0.001）. Compared with the LPS+anti-NC group，the LPS+anti-miR-370-3p group had significant reductions in the protein expression levels of RIPK3 and MLKL in MLE12 cells and the percentage of PI-positive cells（

0.05），as well as significant increases in mitochondrial fragmentation and mitochondrial membrane potential（

0.05）. MiR-370-3p could inhibit the luciferase activity of SESN2-WT reporter gene. Compared with the LPS+anti-miR-370-3p group，the LPS+Si-SEN2+anti-miR-370-3p group had significant reductions in mitochondrial fragmentation and mitochondrial membrane potential（

0.05）.

Conclusion

The miR-370-3p/SESN2 axis promotes the necrotizing apoptosis of AECs during ALI induced by LPS by mediating mitochondrial breakage and damaging mitochondrial function.

关键词

Keywords

references

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