Fascin, a novel marker of human hepatic stellate cells, may regulate their proliferation, migration, and collagen gene expression through the FAK-PI3K-Akt pathway

摘要

Fascin is a component of actin bundles and may regulate various cellular events. The expression and function of fascin in human hepatic stellate cells (HSCs) has remained largely uncharacterized. Fascin expression in human liver tissue was studied using immunohistochemistry. To identify cells expressing fascin, double immunofluorescent staining with vimentin, α-smooth muscle actin (α-SMA), or fibulin-2 was performed and analyzed with confocal microscopy. In culture experiments, fascin expression and the phosphorylation of focal adhesion kinase (FAK) and Akt in LX-2 cells, a cell line of human HSCs, were investigated using western blot. Specific siRNAs were used to reduce the expression of fascin in LX-2 cells. Proliferation and migration were assayed with a CyQuant assay kit and a Matrigel-coated culture insert system, respectively. Levels of matrix metalloproteinase (MMP)-2 and collagen mRNAs were examined using quantitative RT-PCR. Immunohistochemistry revealed the expression of fascin along sinusoids and overlapping with vimentin and α-SMA in both non-fibrotic and fibrotic liver tissue, but it was almost absent in periportal myofibroblastic cells and did not colocalize with fibulin-2, a marker of portal myofibroblasts. In addition, fascin immunoreactivity was almost undetectable in septa of fibrotic human liver tissue. The expression of fascin in LX-2 cells was confirmed using western blot. Two different specific siRNAs against fascin significantly reduced the number of viable LX-2 cells to 65% compared with control cultures and downregulated the mRNAs levels of types I and III collagen and MMP-2 to 62%, 65%, and 70% of control levels, respectively. This condition also reduced the migration activity of LX-2 cells to 46% of control cells and the phosphorylation level of both FAK and Akt. Fascin may be an excellent novel marker of human HSCs that distinguishes HSCs from periportal myofibroblasts. Fascin may regulate functions of human HSCs through the FAK-phosphoinositide 3-kinase-Akt pathway.