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Liver myofibroblasts of murine origins express mesothelin: Identification of novel rat mesothelin splice variants*

Liver myofibroblasts are specialized effector cells that drive hepatic fibrosis, a hallmark process of chronic liver diseases, leading to progressive scar formation and organ failure. Liver myofibroblasts are increasingly recognized as heterogeneous with regards to their origin, phenotype, and funct...

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Autores principales: Fausther, Michel, G. Lavoie, Elise, Dranoff, Jonathan A.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5595315/
https://www.ncbi.nlm.nih.gov/pubmed/28898276
http://dx.doi.org/10.1371/journal.pone.0184499
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author Fausther, Michel
G. Lavoie, Elise
Dranoff, Jonathan A.
author_facet Fausther, Michel
G. Lavoie, Elise
Dranoff, Jonathan A.
author_sort Fausther, Michel
collection PubMed
description Liver myofibroblasts are specialized effector cells that drive hepatic fibrosis, a hallmark process of chronic liver diseases, leading to progressive scar formation and organ failure. Liver myofibroblasts are increasingly recognized as heterogeneous with regards to their origin, phenotype, and functions. For instance, liver myofibroblasts express cell markers that are universally represented such as, ItgαV and Pdgfrβ, or restricted to a given subpopulation such as, Lrat exclusively expressed in hepatic stellate cells, and Gpm6a in mesothelial cells. To study liver myofibroblasts in vitro, we have previously generated and characterized a SV40-immortalized polyclonal rat activated portal fibroblast cell line called RGF-N2 expressing multiple mesothelin mRNA transcripts. Mesothelin, a cell-surface molecule expressed in normal mesothelial cells and overexpressed in several cancers such as, mesothelioma and cholangiocarcinoma, was recently identified as a key regulator of portal myofibroblast proliferation, and fibrosis progression in the setting of chronic cholestatic liver disease. Here, we identify novel mesothelin splice variants expressed in rat activated portal fibroblasts. RGF-N2 portal fibroblast cDNA was used as template for insertion of hemagglutinin tag consensus sequence into the complete open reading frame of rat mesothelin variant coding sequences by extension PCR. Purified amplicons were subsequently cloned into an expression vector for in vitro translation and transfection in monkey COS7 fibroblasts, before characterization of fusion proteins by immunoblot and immunofluorescence. We show that rat activated portal fibroblasts, hepatic stellate cells, and cholangiocarcinoma cells express wild-type mesothelin and additional splice variants, while mouse activated hepatic stellate cells appear to only express wild-type mesothelin. Notably, rat mesothelin splice variants differ from the wild-type isoform by their protein properties and cellular distribution in transfected COS7 fibroblasts. We conclude that mesothelin is a marker of activated murine liver myofibroblasts. Mesothelin gene expression and regulation may be critical in liver myofibroblasts functions and fibrosis progression.
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spelling pubmed-55953152017-09-15 Liver myofibroblasts of murine origins express mesothelin: Identification of novel rat mesothelin splice variants* Fausther, Michel G. Lavoie, Elise Dranoff, Jonathan A. PLoS One Research Article Liver myofibroblasts are specialized effector cells that drive hepatic fibrosis, a hallmark process of chronic liver diseases, leading to progressive scar formation and organ failure. Liver myofibroblasts are increasingly recognized as heterogeneous with regards to their origin, phenotype, and functions. For instance, liver myofibroblasts express cell markers that are universally represented such as, ItgαV and Pdgfrβ, or restricted to a given subpopulation such as, Lrat exclusively expressed in hepatic stellate cells, and Gpm6a in mesothelial cells. To study liver myofibroblasts in vitro, we have previously generated and characterized a SV40-immortalized polyclonal rat activated portal fibroblast cell line called RGF-N2 expressing multiple mesothelin mRNA transcripts. Mesothelin, a cell-surface molecule expressed in normal mesothelial cells and overexpressed in several cancers such as, mesothelioma and cholangiocarcinoma, was recently identified as a key regulator of portal myofibroblast proliferation, and fibrosis progression in the setting of chronic cholestatic liver disease. Here, we identify novel mesothelin splice variants expressed in rat activated portal fibroblasts. RGF-N2 portal fibroblast cDNA was used as template for insertion of hemagglutinin tag consensus sequence into the complete open reading frame of rat mesothelin variant coding sequences by extension PCR. Purified amplicons were subsequently cloned into an expression vector for in vitro translation and transfection in monkey COS7 fibroblasts, before characterization of fusion proteins by immunoblot and immunofluorescence. We show that rat activated portal fibroblasts, hepatic stellate cells, and cholangiocarcinoma cells express wild-type mesothelin and additional splice variants, while mouse activated hepatic stellate cells appear to only express wild-type mesothelin. Notably, rat mesothelin splice variants differ from the wild-type isoform by their protein properties and cellular distribution in transfected COS7 fibroblasts. We conclude that mesothelin is a marker of activated murine liver myofibroblasts. Mesothelin gene expression and regulation may be critical in liver myofibroblasts functions and fibrosis progression. Public Library of Science 2017-09-12 /pmc/articles/PMC5595315/ /pubmed/28898276 http://dx.doi.org/10.1371/journal.pone.0184499 Text en © 2017 Fausther et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Fausther, Michel
G. Lavoie, Elise
Dranoff, Jonathan A.
Liver myofibroblasts of murine origins express mesothelin: Identification of novel rat mesothelin splice variants*
title Liver myofibroblasts of murine origins express mesothelin: Identification of novel rat mesothelin splice variants*
title_full Liver myofibroblasts of murine origins express mesothelin: Identification of novel rat mesothelin splice variants*
title_fullStr Liver myofibroblasts of murine origins express mesothelin: Identification of novel rat mesothelin splice variants*
title_full_unstemmed Liver myofibroblasts of murine origins express mesothelin: Identification of novel rat mesothelin splice variants*
title_short Liver myofibroblasts of murine origins express mesothelin: Identification of novel rat mesothelin splice variants*
title_sort liver myofibroblasts of murine origins express mesothelin: identification of novel rat mesothelin splice variants*
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5595315/
https://www.ncbi.nlm.nih.gov/pubmed/28898276
http://dx.doi.org/10.1371/journal.pone.0184499
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