Nitrogen mustard inhibits keratinocyte proliferation by activating AMPK/autophagy pathway

Xunhu DONG; Mingliang CHEN; Feng YE; Guorong DAN; Zhongmin ZOU

doi:10.16016/j.1000-5404.201901061

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Nitrogen mustard inhibits keratinocyte proliferation by activating AMPK/autophagy pathway

Basic Medicine | 更新时间：2026-01-09

- Nitrogen mustard inhibits keratinocyte proliferation by activating AMPK/autophagy pathway
- Journal of Third Military Medical University Vol. 41, Issue 14, Pages: 1295-1300(2019)
- 作者机构：
  
  400038 重庆, 陆军军医大学(第三军医大学)军事预防医学系防化医学教研室
- 作者简介：
- 基金信息：
  
  the National Natural Science Foundation for Young Scholars of China(81703268);the National Natural Science Foundation for Young Scholars of China(81803279)
- DOI：10.16016/j.1000-5404.201901061
  CLC： R322.991; R827.12; R827.174
- Received：08 January 2019，
  
  Revised：2019-04-07，
  
  Published：30 July 2019
- 稿件说明：
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Xunhu DONG, Mingliang CHEN, Feng YE, et al. Nitrogen mustard inhibits keratinocyte proliferation by activating AMPK/autophagy pathway[J]. Journal of Third Military Medical University, 2019, 41(14): 1295-1300.
DOI：

Xunhu DONG, Mingliang CHEN, Feng YE, et al. Nitrogen mustard inhibits keratinocyte proliferation by activating AMPK/autophagy pathway[J]. Journal of Third Military Medical University, 2019, 41(14): 1295-1300. DOI： 10.16016/j.1000-5404.201901061.

摘要

目的

探讨腺苷酸活化蛋白激酶(AMP-activated protein kinase，AMPK)/自噬通路在氮芥(nitrogen mustard，NM)诱导角质形成细胞增殖活力下降中的作用。

方法

用不同浓度(1、5、10、20 μmol/L)的氮芥单独或加入5 mmol/L自噬特异性抑制剂(3-methyladenine

3-MA)、20 nmol/L自噬特异性激动剂(rapamycin

RAPA)、10 μmol/L AMPK特异性抑制剂(compound C

CC)处理人角质形成细胞株(HaCaT细胞)24 h后，CCK-8法检测细胞增殖活力，Western blot检测AMPK、pAMPK、LC3-B2/B1和p62的表达水平，共聚焦显微镜观察自噬体形成。

结果

NM处理后HaCaT细胞增殖活力被显著抑制(

＜0.05)，且处理浓度越高抑制效应越明显；当NM浓度为20 μmol/L时，角质形成细胞增殖活力约下降40%。同时，NM处理能力显著提高pAMPK、LC3-B2的表达水平并下调p62的表达，且随处理浓度的增加，表达变化越明显；同时，NM可明显促进HaCaT细胞自噬体的形成。加入3-MA或CC干预后能显著抑制NM的上述效应(

＜0.05)，而RAPA干预对NM处理的效应无明显影响(

0.05)。

结论

氮芥能够激活AMPK/自噬通路诱导角质形成细胞的毒性损伤，抑制细胞的增殖活性。

Abstract

Objective

To explore the role of AMP-activated protein kinase (AMPK)/ autophagy signaling pathway in nitrogen mustard (NM)-induced cytotoxicity in cultured keratinocytes.

Methods

HaCaT cells were treated with different concentrations of NM(1

10 and 20 μmol/L) in the presence or absence of 5 mmol/L 3-methyladenine (3-MA

a specific inhibitor of autophagy)

20 nmol/L rapamycin (RAPA

a specific activator of autophagy)

or 10 μmol/L compound C (a specific inhibitor of AMPK) for 24 h. CCK-8 assay was used to assess the cell viability

and the expression of p62

LC3

AMPK and pAMPK in the cells were detected using Western blotting; laser confocal microscopy was performed to detect the formation of autophagosomes in the cells after the treatments.

Results

Treatment with NM significantly decreased the viability of HaCaT cells in a dose-dependent manner

and NM at 20 μmol/L markedly decreased the cell viability to 60% of the control level. NM also dose-dependently up-regulated the phosphorylation of AMPK and LC3-B2

down-regulated the expression of p62

and significantly promoted the formation of autophagosomes in HaCaT cells. The application of 3-MA and compound C both abolished the effects of NM (

0.05)

whereas RAPA did not produce significant changes in the effect of NM (

0.05).

Conclusion

NM induces cytotoxicity in cultured keratinocytes by activating AMPK/autophagy signaling pathway to inhibit the cell proliferation.

关键词

Keywords

references

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