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Fluorochloridone induces autophagy in TM4 Sertoli cells: involvement of ROS-mediated AKT-mTOR signaling pathway
BACKGROUND: Fluorochloridone (FLC), a selective pyrrolidone herbicide, has been recognized as a potential endocrine disruptor and reported to induce male reproductive toxicity, but the underlying mechanism is unclear. The aim of this study was to investigate the mechanism of FLC-induced reproductive...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8073911/ https://www.ncbi.nlm.nih.gov/pubmed/33902598 http://dx.doi.org/10.1186/s12958-021-00739-8 |
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author | Ni, Zhijing Sun, Weiqi Li, Rui Yang, Mingjun Zhang, Fen Chang, Xiuli Li, Weihua Zhou, Zhijun |
author_facet | Ni, Zhijing Sun, Weiqi Li, Rui Yang, Mingjun Zhang, Fen Chang, Xiuli Li, Weihua Zhou, Zhijun |
author_sort | Ni, Zhijing |
collection | PubMed |
description | BACKGROUND: Fluorochloridone (FLC), a selective pyrrolidone herbicide, has been recognized as a potential endocrine disruptor and reported to induce male reproductive toxicity, but the underlying mechanism is unclear. The aim of this study was to investigate the mechanism of FLC-induced reproductive toxicity on male mice with particular emphasis on the role of autophagy in mice’ TM4 Sertoli cells. METHODS: Adult C57BL/6 mice were divided into one control group (0.5% sodium carboxymethyl cellulose), and four FLC-treated groups (3,15,75,375 mg/kg). The animals (ten mice per group) received gavage for 28 days. After treatment, histological analysis, sperm parameters, the microstructure of autophagy and the expression of autophagy-associated proteins in testis were evaluated. Furthermore, to explore the autophagy mechanism, TM4 Sertoli cells were treated with FLC (0,40,80,160 μM) in vitro for 24 h. Cell activity and cytoskeletal changes were measured by MTT assay and F-actin immunofluorescence staining. The formation of autophagosome, accumulation of reactive oxygen species (ROS), expression of autophagy marker proteins (LC3, Beclin-1 and P62) and AKT-related pathway proteins (AKT, mTOR) were observed. The ROS scavenger N-acetylcysteine (NAC) and AKT agonist (SC79) were used to treat TM4 cells to observe the changes of AKT-mTOR pathway and autophagy. RESULTS: In vivo, it showed that FLC exposure caused testicular injuries, abnormality in epididymal sperm. Moreover, FLC increased the formation of autophagosomes, the accumulation of LC3II/LC3I, Beclin-1 and P62 protein, which is related to the degradation of autophagy. In vitro, FLC triggered TM4 cell autophagy by increasing the formation of autophagosomes and upregulating of LC3II/LC3I, Beclin-1 and P62 levels. In addition, FLC induced ROS production and inhibited the activities of AKT and mTOR kinases. The Inhibition of AKT/mTOR signaling pathways and the activation of autophagy induced by FLC could be efficiently reversed by pretreatment of NAC. Additionally, decreased autophagy and increased cell viability were observed in TM4 cells treated with SC79 and FLC, compared with FLC alone, indicating that FLC-induced autophagy may be pro-death. CONCLUSION: Taken together, our study provided the evidence that FLC promoted autophagy in TM4 Sertoli cells and that this process may involve ROS-mediated AKT/mTOR signaling pathways. |
format | Online Article Text |
id | pubmed-8073911 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-80739112021-04-26 Fluorochloridone induces autophagy in TM4 Sertoli cells: involvement of ROS-mediated AKT-mTOR signaling pathway Ni, Zhijing Sun, Weiqi Li, Rui Yang, Mingjun Zhang, Fen Chang, Xiuli Li, Weihua Zhou, Zhijun Reprod Biol Endocrinol Research BACKGROUND: Fluorochloridone (FLC), a selective pyrrolidone herbicide, has been recognized as a potential endocrine disruptor and reported to induce male reproductive toxicity, but the underlying mechanism is unclear. The aim of this study was to investigate the mechanism of FLC-induced reproductive toxicity on male mice with particular emphasis on the role of autophagy in mice’ TM4 Sertoli cells. METHODS: Adult C57BL/6 mice were divided into one control group (0.5% sodium carboxymethyl cellulose), and four FLC-treated groups (3,15,75,375 mg/kg). The animals (ten mice per group) received gavage for 28 days. After treatment, histological analysis, sperm parameters, the microstructure of autophagy and the expression of autophagy-associated proteins in testis were evaluated. Furthermore, to explore the autophagy mechanism, TM4 Sertoli cells were treated with FLC (0,40,80,160 μM) in vitro for 24 h. Cell activity and cytoskeletal changes were measured by MTT assay and F-actin immunofluorescence staining. The formation of autophagosome, accumulation of reactive oxygen species (ROS), expression of autophagy marker proteins (LC3, Beclin-1 and P62) and AKT-related pathway proteins (AKT, mTOR) were observed. The ROS scavenger N-acetylcysteine (NAC) and AKT agonist (SC79) were used to treat TM4 cells to observe the changes of AKT-mTOR pathway and autophagy. RESULTS: In vivo, it showed that FLC exposure caused testicular injuries, abnormality in epididymal sperm. Moreover, FLC increased the formation of autophagosomes, the accumulation of LC3II/LC3I, Beclin-1 and P62 protein, which is related to the degradation of autophagy. In vitro, FLC triggered TM4 cell autophagy by increasing the formation of autophagosomes and upregulating of LC3II/LC3I, Beclin-1 and P62 levels. In addition, FLC induced ROS production and inhibited the activities of AKT and mTOR kinases. The Inhibition of AKT/mTOR signaling pathways and the activation of autophagy induced by FLC could be efficiently reversed by pretreatment of NAC. Additionally, decreased autophagy and increased cell viability were observed in TM4 cells treated with SC79 and FLC, compared with FLC alone, indicating that FLC-induced autophagy may be pro-death. CONCLUSION: Taken together, our study provided the evidence that FLC promoted autophagy in TM4 Sertoli cells and that this process may involve ROS-mediated AKT/mTOR signaling pathways. BioMed Central 2021-04-26 /pmc/articles/PMC8073911/ /pubmed/33902598 http://dx.doi.org/10.1186/s12958-021-00739-8 Text en © The Author(s) 2021 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data. |
spellingShingle | Research Ni, Zhijing Sun, Weiqi Li, Rui Yang, Mingjun Zhang, Fen Chang, Xiuli Li, Weihua Zhou, Zhijun Fluorochloridone induces autophagy in TM4 Sertoli cells: involvement of ROS-mediated AKT-mTOR signaling pathway |
title | Fluorochloridone induces autophagy in TM4 Sertoli cells: involvement of ROS-mediated AKT-mTOR signaling pathway |
title_full | Fluorochloridone induces autophagy in TM4 Sertoli cells: involvement of ROS-mediated AKT-mTOR signaling pathway |
title_fullStr | Fluorochloridone induces autophagy in TM4 Sertoli cells: involvement of ROS-mediated AKT-mTOR signaling pathway |
title_full_unstemmed | Fluorochloridone induces autophagy in TM4 Sertoli cells: involvement of ROS-mediated AKT-mTOR signaling pathway |
title_short | Fluorochloridone induces autophagy in TM4 Sertoli cells: involvement of ROS-mediated AKT-mTOR signaling pathway |
title_sort | fluorochloridone induces autophagy in tm4 sertoli cells: involvement of ros-mediated akt-mtor signaling pathway |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8073911/ https://www.ncbi.nlm.nih.gov/pubmed/33902598 http://dx.doi.org/10.1186/s12958-021-00739-8 |
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