Fluoride disrupts intestinal epithelial tight junction integrity through intracellular calcium-mediated RhoA/ROCK signaling and myosin light chain kinase
Fluoride is a common contaminant of groundwater and agricultural commodity, which poses challenges to animal and human health. A wealth of research has demonstrated its detrimental effects on intestinal mucosal integrity; however, the underlying mechanisms remain obscure. This study aimed to investi...
| Main Authors: | , , , , , , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2023-06-01
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| Series: | Ecotoxicology and Environmental Safety |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S014765132300444X |
| _version_ | 1797833100081758208 |
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| author | Lianxin Li Jinge Xin Hesong Wang Yadong Wang Weiqi Peng Ning Sun Haonan Huang Yanxi Zhou Xingmei Liu Yu Lin Jing Fang Bo Jing Kangcheng Pan Yan Zeng Dong Zeng Xiang Qin Yang Bai Xueqin Ni |
| author_facet | Lianxin Li Jinge Xin Hesong Wang Yadong Wang Weiqi Peng Ning Sun Haonan Huang Yanxi Zhou Xingmei Liu Yu Lin Jing Fang Bo Jing Kangcheng Pan Yan Zeng Dong Zeng Xiang Qin Yang Bai Xueqin Ni |
| author_sort | Lianxin Li |
| collection | DOAJ |
| description | Fluoride is a common contaminant of groundwater and agricultural commodity, which poses challenges to animal and human health. A wealth of research has demonstrated its detrimental effects on intestinal mucosal integrity; however, the underlying mechanisms remain obscure. This study aimed to investigate the role of the cytoskeleton in fluoride-induced barrier dysfunction. After sodium fluoride (NaF) treatment of the cultured Caco-2 cells, both cytotoxicity and cytomorphological changes (internal vacuoles or massive ablation) were observed. NaF lowered transepithelial electrical resistance (TEER) and enhanced paracellular permeation of fluorescein isothiocyanate dextran 4 (FD-4), indicating Caco-2 monolayers hyperpermeability. In the meantime, NaF treatment altered both the expression and distribution of the tight junction protein ZO-1. Fluoride exposure increased myosin light chain II (MLC2) phosphorylation and triggered actin filament (F-actin) remodeling. While inhibition of myosin II by Blebbistatin blocked NaF-induced barrier failure and ZO-1 discontinuity, the corresponding agonist Ionomycin had effects comparable to those of fluoride, suggesting that MLC2 serves as an effector. Given the mechanisms upstream of p-MLC2 regulation, further studies demonstrated that NaF activated RhoA/ROCK signaling pathway and myosin light chain kinase (MLCK), strikingly increasing the expression of both. Pharmacological inhibitors (Rhosin, Y-27632 and ML-7) reversed NaF-induced barrier breakdown and stress fiber formation. The role of intracellular calcium ions ([Ca2+]i) in NaF effects on Rho/ROCK pathway and MLCK was investigated. We found that NaF elevated [Ca2+]i, whereas chelator BAPTA-AM attenuated increased RhoA and MLCK expression as well as ZO-1 rupture, thus, restoring barrier function. Collectively, abovementioned results suggest that NaF induces barrier impairment via Ca2+-dependent RhoA/ROCK pathway and MLCK, which in turn triggers MLC2 phosphorylation and rearrangement of ZO-1 and F-actin. These results provide potential therapeutic targets for fluoride-induced intestinal injury. |
| first_indexed | 2024-04-09T14:19:18Z |
| format | Article |
| id | doaj.art-2f717ebba00b4fd99df907112a263aeb |
| institution | Directory Open Access Journal |
| issn | 0147-6513 |
| language | English |
| last_indexed | 2024-04-09T14:19:18Z |
| publishDate | 2023-06-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Ecotoxicology and Environmental Safety |
| spelling | doaj.art-2f717ebba00b4fd99df907112a263aeb2023-05-05T04:39:43ZengElsevierEcotoxicology and Environmental Safety0147-65132023-06-01257114940Fluoride disrupts intestinal epithelial tight junction integrity through intracellular calcium-mediated RhoA/ROCK signaling and myosin light chain kinaseLianxin Li0Jinge Xin1Hesong Wang2Yadong Wang3Weiqi Peng4Ning Sun5Haonan Huang6Yanxi Zhou7Xingmei Liu8Yu Lin9Jing Fang10Bo Jing11Kangcheng Pan12Yan Zeng13Dong Zeng14Xiang Qin15Yang Bai16Xueqin Ni17Animal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaBaiyun Branch, Nanfang Hospital, Southern Medical University, Guangzhou, ChinaGuangdong Provincial Key Laboratory of Gastroenterology, Department of Gastroenterology, Institute of Gastroenterology of Guangdong Province, Nanfang Hospital, Southern Medical University, Guangzhou, ChinaBaiyun Branch, Nanfang Hospital, Southern Medical University, Guangzhou, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaGuangdong Provincial Key Laboratory of Gastroenterology, Department of Gastroenterology, Institute of Gastroenterology of Guangdong Province, Nanfang Hospital, Southern Medical University, Guangzhou, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaAnimal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, ChinaSchool of Life Science and Technology, University of Electronic Science and Technology of China, Chengdu, China; Corresponding authors.Baiyun Branch, Nanfang Hospital, Southern Medical University, Guangzhou, China; Guangdong Provincial Key Laboratory of Gastroenterology, Department of Gastroenterology, Institute of Gastroenterology of Guangdong Province, Nanfang Hospital, Southern Medical University, Guangzhou, China; Corresponding author at: Baiyun Branch, Nanfang Hospital, Southern Medical University, Guangzhou, China.Animal Microecology Institute, College of Veterinary, Sichuan Agricultural University, Chengdu, Sichuan, China; Corresponding authors.Fluoride is a common contaminant of groundwater and agricultural commodity, which poses challenges to animal and human health. A wealth of research has demonstrated its detrimental effects on intestinal mucosal integrity; however, the underlying mechanisms remain obscure. This study aimed to investigate the role of the cytoskeleton in fluoride-induced barrier dysfunction. After sodium fluoride (NaF) treatment of the cultured Caco-2 cells, both cytotoxicity and cytomorphological changes (internal vacuoles or massive ablation) were observed. NaF lowered transepithelial electrical resistance (TEER) and enhanced paracellular permeation of fluorescein isothiocyanate dextran 4 (FD-4), indicating Caco-2 monolayers hyperpermeability. In the meantime, NaF treatment altered both the expression and distribution of the tight junction protein ZO-1. Fluoride exposure increased myosin light chain II (MLC2) phosphorylation and triggered actin filament (F-actin) remodeling. While inhibition of myosin II by Blebbistatin blocked NaF-induced barrier failure and ZO-1 discontinuity, the corresponding agonist Ionomycin had effects comparable to those of fluoride, suggesting that MLC2 serves as an effector. Given the mechanisms upstream of p-MLC2 regulation, further studies demonstrated that NaF activated RhoA/ROCK signaling pathway and myosin light chain kinase (MLCK), strikingly increasing the expression of both. Pharmacological inhibitors (Rhosin, Y-27632 and ML-7) reversed NaF-induced barrier breakdown and stress fiber formation. The role of intracellular calcium ions ([Ca2+]i) in NaF effects on Rho/ROCK pathway and MLCK was investigated. We found that NaF elevated [Ca2+]i, whereas chelator BAPTA-AM attenuated increased RhoA and MLCK expression as well as ZO-1 rupture, thus, restoring barrier function. Collectively, abovementioned results suggest that NaF induces barrier impairment via Ca2+-dependent RhoA/ROCK pathway and MLCK, which in turn triggers MLC2 phosphorylation and rearrangement of ZO-1 and F-actin. These results provide potential therapeutic targets for fluoride-induced intestinal injury.http://www.sciencedirect.com/science/article/pii/S014765132300444XFluorideIntestinal epithelial barrierTight junctionRho kinaseMyosin light chain kinaseIntracellular calcium level |
| spellingShingle | Lianxin Li Jinge Xin Hesong Wang Yadong Wang Weiqi Peng Ning Sun Haonan Huang Yanxi Zhou Xingmei Liu Yu Lin Jing Fang Bo Jing Kangcheng Pan Yan Zeng Dong Zeng Xiang Qin Yang Bai Xueqin Ni Fluoride disrupts intestinal epithelial tight junction integrity through intracellular calcium-mediated RhoA/ROCK signaling and myosin light chain kinase Ecotoxicology and Environmental Safety Fluoride Intestinal epithelial barrier Tight junction Rho kinase Myosin light chain kinase Intracellular calcium level |
| title | Fluoride disrupts intestinal epithelial tight junction integrity through intracellular calcium-mediated RhoA/ROCK signaling and myosin light chain kinase |
| title_full | Fluoride disrupts intestinal epithelial tight junction integrity through intracellular calcium-mediated RhoA/ROCK signaling and myosin light chain kinase |
| title_fullStr | Fluoride disrupts intestinal epithelial tight junction integrity through intracellular calcium-mediated RhoA/ROCK signaling and myosin light chain kinase |
| title_full_unstemmed | Fluoride disrupts intestinal epithelial tight junction integrity through intracellular calcium-mediated RhoA/ROCK signaling and myosin light chain kinase |
| title_short | Fluoride disrupts intestinal epithelial tight junction integrity through intracellular calcium-mediated RhoA/ROCK signaling and myosin light chain kinase |
| title_sort | fluoride disrupts intestinal epithelial tight junction integrity through intracellular calcium mediated rhoa rock signaling and myosin light chain kinase |
| topic | Fluoride Intestinal epithelial barrier Tight junction Rho kinase Myosin light chain kinase Intracellular calcium level |
| url | http://www.sciencedirect.com/science/article/pii/S014765132300444X |
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