BMSCs-derived exosomes overexpressing TGM2 inhibit lactate-induced oxidative stress related apoptosis in nucleus pulposus cells

Jinhui LOU; Xiaoxin GAO; Yuyao ZHANG; Yu ZHAI; Changqing LI

doi:10.16016/j.2097-0927.202402033

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BMSCs-derived exosomes overexpressing TGM2 inhibit lactate-induced oxidative stress related apoptosis in nucleus pulposus cells

Basic Medicine | 更新时间：2026-01-09

- BMSCs-derived exosomes overexpressing TGM2 inhibit lactate-induced oxidative stress related apoptosis in nucleus pulposus cells
- Journal of Army Medical University Vol. 46, Issue 19, Pages: 2188-2196(2024)
- 作者机构：
  
  400037 重庆，陆军军医大学(第三军医大学)第二附属医院骨科
- 作者简介：
- 基金信息：
  
  the General Program of National Natural Science Foundation of China(82172488)
- DOI：10.16016/j.2097-0927.202402033
  CLC： R322.72;R329.28;R394.2
- Received：08 February 2024，
  
  Revised：2024-06-07，
  
  Published：15 October 2024
- 稿件说明：
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Jinhui LOU, Xiaoxin GAO, Yuyao ZHANG, et al. BMSCs-derived exosomes overexpressing TGM2 inhibit lactate-induced oxidative stress related apoptosis in nucleus pulposus cells[J]. Journal of Army Medical University, 2024, 46(19): 2188-2196.
DOI：

Jinhui LOU, Xiaoxin GAO, Yuyao ZHANG, et al. BMSCs-derived exosomes overexpressing TGM2 inhibit lactate-induced oxidative stress related apoptosis in nucleus pulposus cells[J]. Journal of Army Medical University, 2024, 46(19): 2188-2196. DOI： 10.16016/j.2097-0927.202402033.

摘要

目的

探究转谷氨酰胺酶2(transglutaminase2，TGM2)过表达的骨髓间充质干细胞(bone marrow mesenchymal stem cells，BMSCs)来源外泌体在乳酸介导的髓核细胞氧化应激性凋亡中的作用及机制。

方法

选用20只6周龄雄性SD大鼠[体质量(350±50)g

]

用于提取髓核细胞。利用乳酸构建原代髓核细胞氧化应激性凋亡模型；利用慢病毒载体在BMSCs中过表达TGM2蛋白，提取外泌体并进行鉴定；后为验证TGM2过表达的外泌体抗髓核细胞氧化应激性凋亡潜在机制，利用流式细胞术对髓核细胞活性氧水平进行检测，Western blot、免疫荧光对髓核细胞Bax、Bcl-2、Cleaved-Caspase3、磷酸化PI3K、磷酸化Akt、核内Nrf2、胞质Nrf2、HO-1蛋白表达水平及Nrf2核内荧光表达情况进行检测，分为4组(

=3)：对照组、乳酸处理组、乳酸+对照外泌体组、乳酸+TGM2过表达外泌体组，评估髓核细胞的氧化应激性凋亡程度及PI3K/Akt/Nrf2通路变化情况。

结果

10 mmol/L乳酸能够更好的诱导髓核细胞氧化应激性凋亡(

＜0.05)；对照BMSCs组和TGM2过表达BMSCs组均可产生外泌体且能够稳定过表达TGM2(

＜0.05

＜0.01)；且相较于乳酸处理组，乳酸+TGM2过表达外泌体组和乳酸+对照外泌体组的髓核细胞均表现为凋亡率降低(

＜0.05)，活性氧水平下降(

＜0.05)，乳酸+TGM2过表达外泌体组抗凋亡、活性氧蓄积效果更好(

＜0.05)，髓核细胞内Bax、Cleaved-Caspase3蛋白表达均下降(

＜0.05)，Bcl-2蛋白水平均增高(

＜0.05)，PI3K、Akt磷酸化程度均增强(

＜0.05)，Nrf2核内表达均增加，抗氧化应激因子HO-1表达均增多，乳酸+TGM2过表达外泌体组抗凋亡及PI3K/Akt/Nrf2通路激活程度更强(

＜0.05)。

结论

TGM2过表达的BMSCs外泌体可抑制乳酸介导的髓核细胞氧化应激性凋亡，其机制可能与PI3K/Akt/Nrf2通路激活有关。

Abstract

Objective

To explore the role and mechanism of exosomes derived from bone marrow mesenchymal stem cells (BMSCs) overexpressing transglutaminase 2 (TGM2) in lactate-mediated oxidative stress-induced apoptosis in nucleus pulposus cells.

Methods

Twenty 6-week-old male SD rats (350±50 g) were used to extract nucleus pulposus cells. Lactic acid was utilized to establish a model of oxidative stress-induced apoptosis in primary nucleus pulposus cells. Lentiviral vectors carrying TGM2 were employed to transfect BMSCs to overexpress the protein

and then exosomes were extracted from the BMSCs and identified. Subsequently

to investigate the potential mechanisms of TGM2-overexpressing exosomes against oxidative stress-induced apoptosis in nucleus pulposus cells

flow cytometry was used to detect the production of reactive oxygen species (ROS)

and Western blotting and immunofluorescence assay were applied to measure the expression levels of Bax

Bcl-2

Cleaved-Caspase3

p-PI3K

p-Akt

n-Nrf2

c-Nrf2 and HO-1

and Nrf2 nuclear translocation. The rate of oxidative stress-induced apoptosis in nucleus pulposus cells and changes in the PI3K/Akt/Nrf2 pathway were assessed in 4 groups of cells (

=3): control group

lactate treatment group

lactate+control exosome group

and lactate+TGM2-overexpressing exosomes group.

Results

Lactic acid of 10 mmol/L was found to be more effective in inducing oxidative stress-induced apoptosis in nucleus pulposus cells (

0.05). Both control BMSCs and TGM2-overexpressing BMSCs could produce exosomes and stably overexpress TGM2 (

0.05

0.01). Compared with the lactate treatment group

the nucleus pulposus cells in the lactate+TGM2-overexpressing exosome group and the lactate+control exosome group showed reduced apoptotic rates (

0.05)

as well as decreased ROS level (

0.05). The lactate+TGM2-overexpressing exosome group exhibited bette

r anti-apoptosis and ROS accumulation effects (

0.05). Additionally

the expression of Bax and Cleaved-Caspase3 was decreased (

0.05)

that of Bcl-2 protein was increased (

0.05)

the phosphorylation of PI3K/Akt was enhanced (

0.05)

and both Nrf2 nuclear translocation and the expression of the antioxidant stress factor HO-1 were increased in nucleus pulposus cells. The lactic acid + TGM2-overexpressing exosome group showed stronger anti-apoptotic effects and activation of the PI3K/Akt/Nrf2 pathway (

0.05).

Conclusion

Exosomes derived from BMSCs overexpressing TGM2 can inhibit lactate-mediated oxidative stress-induced apoptosis in nucleus pulposus cells

and the mechanism may be related to the activation of the PI3K/Akt/Nrf2 pathway.

关键词

Keywords

references

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