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Upregulation of COX-2 and PGE(2) Induced by TNF-α Mediated Through TNFR1/MitoROS/PKCα/P38 MAPK, JNK1/2/FoxO1 Cascade in Human Cardiac Fibroblasts
PURPOSE: Tumor necrosis factor-α (TNF-α) has been shown to exert as a pathogenic factor in cardiac fibrosis and heart failure which were associated with the up-regulation of cyclooxygenase (COX)-2/prostaglandin E(2) (PGE(2)) axis. However, whether TNF-α-induced COX-2/PGE(2) upregulation mediated thr...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Dove
2021
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8254141/ https://www.ncbi.nlm.nih.gov/pubmed/34234507 http://dx.doi.org/10.2147/JIR.S313665 |
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author | Yang, Chuen-Mao Yang, Chien-Chung Hsiao, Li-Der Yu, Chia-Ying Tseng, Hui-Ching Hsu, Chih-Kai Situmorang, Jiro Hasegawa |
author_facet | Yang, Chuen-Mao Yang, Chien-Chung Hsiao, Li-Der Yu, Chia-Ying Tseng, Hui-Ching Hsu, Chih-Kai Situmorang, Jiro Hasegawa |
author_sort | Yang, Chuen-Mao |
collection | PubMed |
description | PURPOSE: Tumor necrosis factor-α (TNF-α) has been shown to exert as a pathogenic factor in cardiac fibrosis and heart failure which were associated with the up-regulation of cyclooxygenase (COX)-2/prostaglandin E(2) (PGE(2)) axis. However, whether TNF-α-induced COX-2/PGE(2) upregulation mediated through ROS-dependent cascade remains elusive in human cardiac fibroblasts (HCFs). This study aims to address the underlying mechanisms of TNF-α-induced COX-2/PGE(2) expression. METHODS: Here, we used TNF receptor neutralizing antibody (TNFR nAb), pharmacologic inhibitors, and siRNAs to dissect the involvement of signaling components examined by Western blot and ELISA in TNF-α-mediated responses in HCFs. MitoSOX Red was used to measure mitoROS generation. Isolation of subcellular fractions was performed to determine membrane translocation of PKCα. Promoter luciferase assay and chromatin immunoprecipitation (ChIP) assay were used to determine the role of transcription factor. RESULTS: We found that TNF-α time- and concentration-dependently upregulated COX-2 protein and mRNA expression as well as PGE(2) synthesis which was attenuated by TNFR1 nAb, the inhibitor of mitochondrial ROS scavenger (MitoTEMPO), protein kinase C [(PKC)α, Gö6976], p38 MAPK [p38 inhibitor VIII, (p38i VIII)], JNK1/2 (SP600125), or forkhead box protein O1 [(FoxO1), AS1842856], and transfection with their respective siRNAs in HCFs. TNF-α-stimulated PKCα phosphorylation was inhibited by TNFR1 nAb, MitoTEMPO, or Gö6976. TNF-α stimulated phosphorylation of p38 MAPK and JNK1/2 was attenuated by TNFR1 nAb, MitoTEMPO, Gö6976, and their inhibitors p38i VIII and SP600125. Moreover, TNF-α-triggered FoxO1 phosphorylation was abolished by AS1842856, TNFR1 nAb, and its upstream inhibitors MitoTEMPO, Gö6976, p38i VIII, and SP600125. Phosphorylation of FoxO1 could enhance its interaction with the COX-2 promoter element revealed by ChIP assay, which was attenuated by AS1842856. CONCLUSION: Our results suggested that TNF-α-induced COX-2/PGE(2) upregulation is mediated through TNFR1-dependent MitoROS/PKCα/p38 MAPK and JNK1/2 cascade to activate FoxO1 binding with the COX-2 promoter in HCFs. |
format | Online Article Text |
id | pubmed-8254141 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | Dove |
record_format | MEDLINE/PubMed |
spelling | pubmed-82541412021-07-06 Upregulation of COX-2 and PGE(2) Induced by TNF-α Mediated Through TNFR1/MitoROS/PKCα/P38 MAPK, JNK1/2/FoxO1 Cascade in Human Cardiac Fibroblasts Yang, Chuen-Mao Yang, Chien-Chung Hsiao, Li-Der Yu, Chia-Ying Tseng, Hui-Ching Hsu, Chih-Kai Situmorang, Jiro Hasegawa J Inflamm Res Original Research PURPOSE: Tumor necrosis factor-α (TNF-α) has been shown to exert as a pathogenic factor in cardiac fibrosis and heart failure which were associated with the up-regulation of cyclooxygenase (COX)-2/prostaglandin E(2) (PGE(2)) axis. However, whether TNF-α-induced COX-2/PGE(2) upregulation mediated through ROS-dependent cascade remains elusive in human cardiac fibroblasts (HCFs). This study aims to address the underlying mechanisms of TNF-α-induced COX-2/PGE(2) expression. METHODS: Here, we used TNF receptor neutralizing antibody (TNFR nAb), pharmacologic inhibitors, and siRNAs to dissect the involvement of signaling components examined by Western blot and ELISA in TNF-α-mediated responses in HCFs. MitoSOX Red was used to measure mitoROS generation. Isolation of subcellular fractions was performed to determine membrane translocation of PKCα. Promoter luciferase assay and chromatin immunoprecipitation (ChIP) assay were used to determine the role of transcription factor. RESULTS: We found that TNF-α time- and concentration-dependently upregulated COX-2 protein and mRNA expression as well as PGE(2) synthesis which was attenuated by TNFR1 nAb, the inhibitor of mitochondrial ROS scavenger (MitoTEMPO), protein kinase C [(PKC)α, Gö6976], p38 MAPK [p38 inhibitor VIII, (p38i VIII)], JNK1/2 (SP600125), or forkhead box protein O1 [(FoxO1), AS1842856], and transfection with their respective siRNAs in HCFs. TNF-α-stimulated PKCα phosphorylation was inhibited by TNFR1 nAb, MitoTEMPO, or Gö6976. TNF-α stimulated phosphorylation of p38 MAPK and JNK1/2 was attenuated by TNFR1 nAb, MitoTEMPO, Gö6976, and their inhibitors p38i VIII and SP600125. Moreover, TNF-α-triggered FoxO1 phosphorylation was abolished by AS1842856, TNFR1 nAb, and its upstream inhibitors MitoTEMPO, Gö6976, p38i VIII, and SP600125. Phosphorylation of FoxO1 could enhance its interaction with the COX-2 promoter element revealed by ChIP assay, which was attenuated by AS1842856. CONCLUSION: Our results suggested that TNF-α-induced COX-2/PGE(2) upregulation is mediated through TNFR1-dependent MitoROS/PKCα/p38 MAPK and JNK1/2 cascade to activate FoxO1 binding with the COX-2 promoter in HCFs. Dove 2021-06-28 /pmc/articles/PMC8254141/ /pubmed/34234507 http://dx.doi.org/10.2147/JIR.S313665 Text en © 2021 Yang et al. https://creativecommons.org/licenses/by-nc/3.0/This work is published and licensed by Dove Medical Press Limited. The full terms of this license are available at https://www.dovepress.com/terms.php and incorporate the Creative Commons Attribution – Non Commercial (unported, v3.0) License (http://creativecommons.org/licenses/by-nc/3.0/ (https://creativecommons.org/licenses/by-nc/3.0/) ). By accessing the work you hereby accept the Terms. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed. For permission for commercial use of this work, please see paragraphs 4.2 and 5 of our Terms (https://www.dovepress.com/terms.php). |
spellingShingle | Original Research Yang, Chuen-Mao Yang, Chien-Chung Hsiao, Li-Der Yu, Chia-Ying Tseng, Hui-Ching Hsu, Chih-Kai Situmorang, Jiro Hasegawa Upregulation of COX-2 and PGE(2) Induced by TNF-α Mediated Through TNFR1/MitoROS/PKCα/P38 MAPK, JNK1/2/FoxO1 Cascade in Human Cardiac Fibroblasts |
title | Upregulation of COX-2 and PGE(2) Induced by TNF-α Mediated Through TNFR1/MitoROS/PKCα/P38 MAPK, JNK1/2/FoxO1 Cascade in Human Cardiac Fibroblasts |
title_full | Upregulation of COX-2 and PGE(2) Induced by TNF-α Mediated Through TNFR1/MitoROS/PKCα/P38 MAPK, JNK1/2/FoxO1 Cascade in Human Cardiac Fibroblasts |
title_fullStr | Upregulation of COX-2 and PGE(2) Induced by TNF-α Mediated Through TNFR1/MitoROS/PKCα/P38 MAPK, JNK1/2/FoxO1 Cascade in Human Cardiac Fibroblasts |
title_full_unstemmed | Upregulation of COX-2 and PGE(2) Induced by TNF-α Mediated Through TNFR1/MitoROS/PKCα/P38 MAPK, JNK1/2/FoxO1 Cascade in Human Cardiac Fibroblasts |
title_short | Upregulation of COX-2 and PGE(2) Induced by TNF-α Mediated Through TNFR1/MitoROS/PKCα/P38 MAPK, JNK1/2/FoxO1 Cascade in Human Cardiac Fibroblasts |
title_sort | upregulation of cox-2 and pge(2) induced by tnf-α mediated through tnfr1/mitoros/pkcα/p38 mapk, jnk1/2/foxo1 cascade in human cardiac fibroblasts |
topic | Original Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8254141/ https://www.ncbi.nlm.nih.gov/pubmed/34234507 http://dx.doi.org/10.2147/JIR.S313665 |
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