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.