Cr(VI)-Induced Autophagy Protects L-02 Hepatocytes from Apoptosis Through the ROS-AKT-mTOR Pathway
Background/Aims: Hexavalent chromium [Cr(VI)] pollution has become a global concern for both ecosystems and human health. Our previous study revealed Cr(VI) could induce both apoptosis and autophagy in L-02 hepatocytes. Here, we sought to explore the underlying mechanism of Cr(VI)-induced autophagy...
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| Format: | Article |
| Language: | English |
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Cell Physiol Biochem Press GmbH & Co KG
2018-11-01
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| Series: | Cellular Physiology and Biochemistry |
| Subjects: | |
| Online Access: | https://www.karger.com/Article/FullText/495713 |
| _version_ | 1828973432695422976 |
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| author | Qi Liang Yuanyuan Xiao Kaihua Liu Caigao Zhong Ming Zeng Fang Xiao |
| author_facet | Qi Liang Yuanyuan Xiao Kaihua Liu Caigao Zhong Ming Zeng Fang Xiao |
| author_sort | Qi Liang |
| collection | DOAJ |
| description | Background/Aims: Hexavalent chromium [Cr(VI)] pollution has become a global concern for both ecosystems and human health. Our previous study revealed Cr(VI) could induce both apoptosis and autophagy in L-02 hepatocytes. Here, we sought to explore the underlying mechanism of Cr(VI)-induced autophagy and its exact role in cell death. Methods: Autophagy ultrastructure was observed under transmission electron microscope (TEM), autophagy flux was measured with double-tagged mCherry-green fluorescent protein (GFP)-microtubule-associated protein 1 light chain 3 (LC3) assay, long-lived protein degradation assay, and LC3II expression assay in the presence of lysosomal inhibitor, bafilomycin A1 (BafA1). Reactive oxygen species (ROS) level was determined using fluorescent probe dichloro-dihydrofluorescein diacetate (DCFH-DA). The expression levels of Beclin-1, LC3, p62/ SQSTM1, and AKT-mammalian target of rapamycin (mTOR) pathway-related molecules including phosphorylation (p)-AKT, AKT, p-mTOR, and mTOR were examined using real-time polymerase chain reaction (RT-PCR) and western blotting. Apoptosis was determined using Annexin V- fluorescein isothiocyanate (FITC)/propidium iodide (PI) staining. Results: Our results demonstrated Cr(VI) exposure activated autophagy in L-02 hepatocytes, as evidenced by the accumulation of autophagosomes, the increase of LC3-II and degradation of p62/ SQSTM1, and the enhanced overall degradation of proteins. We also confirmed Cr(VI)-induced LC3-II elevation mainly came from autophagy induction rather than lysosomal degradation impairment. ROS-AKT-mTOR pathway was associated with Cr(VI)-induced autophagy, and ROS scavenger N-acetylcysteine (NAC) pretreatment inhibited Cr(VI)-induced autophagy by alleviating the inhibition of the AKT-mTOR pathway. Autophagy inhibitors 3-methyladenine (3-MA) and chloroquine diphosphate (CDP) promoted Cr(VI)-induced apoptotic death. Conclusion: These findings indicated Cr(VI)-induced autophagy protected L-02 hepatocytes from apoptosis through the ROS-AKT-mTOR pathway. |
| first_indexed | 2024-12-14T13:46:08Z |
| format | Article |
| id | doaj.art-56c3faeaa4944db19790c576b1db387e |
| institution | Directory Open Access Journal |
| issn | 1015-8987 1421-9778 |
| language | English |
| last_indexed | 2024-12-14T13:46:08Z |
| publishDate | 2018-11-01 |
| publisher | Cell Physiol Biochem Press GmbH & Co KG |
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| series | Cellular Physiology and Biochemistry |
| spelling | doaj.art-56c3faeaa4944db19790c576b1db387e2022-12-21T22:59:19ZengCell Physiol Biochem Press GmbH & Co KGCellular Physiology and Biochemistry1015-89871421-97782018-11-015141863187810.1159/000495713495713Cr(VI)-Induced Autophagy Protects L-02 Hepatocytes from Apoptosis Through the ROS-AKT-mTOR PathwayQi LiangYuanyuan XiaoKaihua LiuCaigao ZhongMing ZengFang XiaoBackground/Aims: Hexavalent chromium [Cr(VI)] pollution has become a global concern for both ecosystems and human health. Our previous study revealed Cr(VI) could induce both apoptosis and autophagy in L-02 hepatocytes. Here, we sought to explore the underlying mechanism of Cr(VI)-induced autophagy and its exact role in cell death. Methods: Autophagy ultrastructure was observed under transmission electron microscope (TEM), autophagy flux was measured with double-tagged mCherry-green fluorescent protein (GFP)-microtubule-associated protein 1 light chain 3 (LC3) assay, long-lived protein degradation assay, and LC3II expression assay in the presence of lysosomal inhibitor, bafilomycin A1 (BafA1). Reactive oxygen species (ROS) level was determined using fluorescent probe dichloro-dihydrofluorescein diacetate (DCFH-DA). The expression levels of Beclin-1, LC3, p62/ SQSTM1, and AKT-mammalian target of rapamycin (mTOR) pathway-related molecules including phosphorylation (p)-AKT, AKT, p-mTOR, and mTOR were examined using real-time polymerase chain reaction (RT-PCR) and western blotting. Apoptosis was determined using Annexin V- fluorescein isothiocyanate (FITC)/propidium iodide (PI) staining. Results: Our results demonstrated Cr(VI) exposure activated autophagy in L-02 hepatocytes, as evidenced by the accumulation of autophagosomes, the increase of LC3-II and degradation of p62/ SQSTM1, and the enhanced overall degradation of proteins. We also confirmed Cr(VI)-induced LC3-II elevation mainly came from autophagy induction rather than lysosomal degradation impairment. ROS-AKT-mTOR pathway was associated with Cr(VI)-induced autophagy, and ROS scavenger N-acetylcysteine (NAC) pretreatment inhibited Cr(VI)-induced autophagy by alleviating the inhibition of the AKT-mTOR pathway. Autophagy inhibitors 3-methyladenine (3-MA) and chloroquine diphosphate (CDP) promoted Cr(VI)-induced apoptotic death. Conclusion: These findings indicated Cr(VI)-induced autophagy protected L-02 hepatocytes from apoptosis through the ROS-AKT-mTOR pathway.https://www.karger.com/Article/FullText/495713Cr(VI)AutophagyL-02 hepatocytesApoptosisROSAKT-mTOR |
| spellingShingle | Qi Liang Yuanyuan Xiao Kaihua Liu Caigao Zhong Ming Zeng Fang Xiao Cr(VI)-Induced Autophagy Protects L-02 Hepatocytes from Apoptosis Through the ROS-AKT-mTOR Pathway Cellular Physiology and Biochemistry Cr(VI) Autophagy L-02 hepatocytes Apoptosis ROS AKT-mTOR |
| title | Cr(VI)-Induced Autophagy Protects L-02 Hepatocytes from Apoptosis Through the ROS-AKT-mTOR Pathway |
| title_full | Cr(VI)-Induced Autophagy Protects L-02 Hepatocytes from Apoptosis Through the ROS-AKT-mTOR Pathway |
| title_fullStr | Cr(VI)-Induced Autophagy Protects L-02 Hepatocytes from Apoptosis Through the ROS-AKT-mTOR Pathway |
| title_full_unstemmed | Cr(VI)-Induced Autophagy Protects L-02 Hepatocytes from Apoptosis Through the ROS-AKT-mTOR Pathway |
| title_short | Cr(VI)-Induced Autophagy Protects L-02 Hepatocytes from Apoptosis Through the ROS-AKT-mTOR Pathway |
| title_sort | cr vi induced autophagy protects l 02 hepatocytes from apoptosis through the ros akt mtor pathway |
| topic | Cr(VI) Autophagy L-02 hepatocytes Apoptosis ROS AKT-mTOR |
| url | https://www.karger.com/Article/FullText/495713 |
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