Sarcomeric gene expression and contractility in myofibroblasts

Abstract

Myofibroblasts are cells found in both normal and pathological states that have structural and functional features of both muscle and nonmuscle cells. The goal of this project was to determine how these cells achieve their intermediate phenotype and whether they have unique contractile properties. In these studies, permanent cell lines corresponding to myofibroblasts from liver (stellate cells) and kidney (mesangial cells) were used. The liver stellate cell line is derived from a cirrhotic rat liver. Our data suggest that the baby hamster kidney (BHK) cell line is a model of kidney mesangial cells. Characterization of both the liver stellate cell line and BHK cells revealed expression of a number of muscle structural and regulatory proteins. BHK cells express six distinct skeletal myosin heavy chains (MyHC) and several sarcomeric thin filament proteins. Liver stellate cells express a more limited program of muscle genes, restricted to three skeletal MyHC isoforms, but no sarcomeric thin filament proteins. Both cell types express two myogenic regulators, myogenin and MyoD, which normally result in cell cycle withdrawal and morphological differentiation. This is intriguing since these cells do not fuse to form multinucleated myotubes and do not withdraw from the cell cycle. Transfections of skeletal muscle promoters linked to reporter genes suggest that the myogenic regulatory program that normally regulates gene expression in skeletal muscle cells also activates sarcomeric genes in BHK and liver stellate cells.;In BHK cells, skeletal MyHC is partially associated with the stress fibers of the cytoskeleton. In the liver stellate cells, sarcomeric MyHC forms dense patches in the cytoplasm. Sarcomeric MyHC expressed by these two types of myofibroblasts is filamentous and shows a distinct organization when compared to nonmuscle MyHC. BHK and liver stellate cells also contract in response to endothelin 1, a vasoactive agent that is upregulated during wound healing. This contraction is accompanied by an increase in sarcomeric protein expression in both liver stellate cells and BHK cells. In addition, ET-1 seems to regulate the expression of sarcomeric MyHC at the translational level. Finally, transfection of a dominant negative inhibitor of sarcomeric MyHC assembly and of a truncated sarcomeric troponin T into BHK cells indicate that both sarcomeric thick and thin filament proteins are necessary for myofibroblast contractility.