AKT/mTOR and c-Jun N-terminal kinase signaling pathways are required for chrysotile asbestos-induced autophagy

Our plain-English summary of AKT/mTOR and c-Jun N-terminal kinase signaling pathways are required for chrysotile asbestos-induced autophagy, by Z. Lin, T. Liu, D.W. Kamp, et al (Free Radical Biology & Medicine, 2014).

The study

• Authors: Z. Lin, T. Liu, D.W. Kamp, Y. Wang, H. He, X. Zhou, D. Li, L. Yang, B. Zhao, G. Liu
• Journal: Free Radical Biology & Medicine
• Published: 2014;72:296-307
• Cited by: 30 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Chrysotile asbestos is closely associated with excess mortality from pulmonary diseases such as lung cancer, mesothelioma, and asbestosis. Although multiple mechanisms in which chrysotile asbestos fibers induce pulmonary disease have been identified, the role of autophagy in human lung epithelial cells has not been examined. In this study, we evaluated whether chrysotile asbestos induces autophagy in A549 human lung epithelial cells and then analyzed the possible underlying molecular mechanism. Chrysotile asbestos induced autophagy in A549 cells based on a series of biochemical and microscopic autophagy markers. We observed that asbestos increased expression of A549 cell microtubule-associated protein 1 light chain 3 (LC3-II), an autophagy marker, in conjunction with dephosphorylation of phospho-AKT, phospho-mTOR, and phospho-p70S6K. Notably, AKT1/AKT2 double-knockout murine embryonic fibroblasts (MEFs) had negligible asbestos-induced LC3-II expression, supporting a crucial role for AKT signaling. Chrysotile asbestos also led to the phosphorylation/activation of Jun N-terminal kinase (JNK) and p38 MAPK. Pharmacologic inhibition of JNK, but not p38 MAPK, dramatically inhibited the protein expression of LC3-II. Moreover, JNK2(-/-) MEFs but not JNK1(-/-) MEFs blocked LC3-II levels induced by chrysotile asbestos. In addition, N-acetylcysteine, an antioxidant, attenuated chrysotile asbestos-induced dephosphorylation of P-AKT and completely abolished phosphorylation/activation of JNK. Finally, we demonstrated that chrysotile asbestos-induced apoptosis was not affected by the presence of the autophagy inhibitor 3-methyladenine or autophagy-related gene 5 siRNA, indicating that the chrysotile asbestos-induced autophagy may be adaptive rather than prosurvival. Our findings demonstrate that AKT/mTOR and JNK2 signaling pathways are required for chrysotile asbestos-induced autophagy. These data provide a mechanistic basis for possible future clinical applications targeting these signaling pathways in the management of asbestos-induced lung disease.

Copyright © 2014 Elsevier Inc. All rights reserved.

In plain English

This was a laboratory study of cells grown in dishes, not people. The authors exposed human lung cancer cells (A549) and mouse embryo cells to chrysotile asbestos, mostly at 100 μg/cm2. They tested whether chrysotile triggers autophagy, in which cells break down their own contents. In A549 cells, chrysotile raised the autophagy marker LC3-II 1.5- to 2.2-fold from as early as 30 minutes. Chrysotile lowered active forms of the signaling proteins AKT, mTOR and p70S6K and activated JNK and p38. Blocking JNK, but not p38, stopped the LC3-II increase. The response was abolished in mouse cells lacking AKT1 and AKT2, and in cells lacking JNK2, but not JNK1. An antioxidant, NAC, partly reversed the AKT change and reduced JNK activation. Blocking autophagy with 3-MA did not significantly affect apoptosis or cell death after 24 hours of chrysotile exposure, although 3-MA alone modestly increased cell death. The authors concluded that the AKT/mTOR and JNK2 pathways are required for chrysotile-induced autophagy, which may be adaptive rather than protective. They acknowledged that cultured-cell results do not reflect conditions in the body.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: In vitro laboratory experiment on cultured cells (cell-line and knockout-cell experiments)
• Who was studied: Human lung carcinoma A549 cells, plus mouse embryonic fibroblasts (MEFs): AKT1/AKT2 double-knockout, JNK1-knockout and JNK2-knockout cells, compared with wild-type MEFs
• Exposure: Chrysotile asbestos mined in Qinghai, China; the International Union against Cancer standard reference sample (average length 7.8 μm, average diameter 0.2 μm). Cells were mainly treated at 100 μg/cm2 for various times; some tests used 50 to 300 μg/cm2.
• Main results:
  • In chrysotile-treated A549 cells, the autophagy marker LC3-II rose 1.5- to 2.2-fold from as early as 30 minutes, and the rise persisted for over 24 hours
  • Electron microscopy of chrysotile-treated A549 cells showed double-membrane structures characteristic of autophagosomes and autolysosomes
  • In mouse cells lacking AKT1 and AKT2, chrysotile-induced autophagy was abolished compared with wild-type mouse cells
  • A JNK inhibitor (SP600125) blocked the chrysotile-induced LC3-II increase in A549 cells; a p38 inhibitor (SB203580) did not
  • The chrysotile-induced LC3-II increase was abolished in JNK2-knockout mouse cells but not in JNK1-knockout mouse cells
  • The antioxidant NAC significantly diminished chrysotile-induced JNK activation in A549 cells, and partly reversed the drop in active AKT
  • A p53 inhibitor (pifithrin-α) reduced chrysotile-induced LC3-II formation in A549 cells
  • Given alone, without asbestos, 3-MA modestly increased cell death, but not apoptosis, in A549 cells compared with controls
  • Blocking autophagy with 3-MA did not significantly affect apoptosis or cell death in A549 cells after 24 hours of chrysotile exposure. With ATG5 knockdown, cell death after chrysotile was similar to control cells.
• The authors' conclusion: The authors concluded that the AKT/mTOR and JNK2 signaling pathways are required for chrysotile-induced autophagy in these cells, and that this autophagy may be adaptive rather than prosurvival
• Limitations the authors state:
  • Autophagy seen in A549 cells in the dish does not reflect conditions in the body, particularly in human disease
  • The detailed molecular mechanisms and the significance in living organisms remain to be studied
  • The study focused on A549 cells; primary lung epithelial cells and animal models were not examined

Topics

The National Library of Medicine indexes this study under subjects our site covers: Chrysotile.

Medical terms in this study, explained in our Lexicon: Chrysotile, Lung Cancer, Mesothelioma, A549 Cells, Fibroblast, Apoptosis.

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