Blebbistatin inhibits contraction and accelerates migration in mouse hepatic stellate cells.
OBJECTIVE
Blebbistatin, an inhibitor of myosin-II-specific ATPase, has been used to inhibit contraction of invertebrate and mammalian muscle preparations containing non-muscle myosin. Activated hepatic stellate cells have contractile properties and play an important role in the pathophysiology of liver fibrosis and portal hypertension. Therefore, hepatic stellate cells are considered as therapeutic target cells. In the present study, we studied the effect of blebbistatin during the transition of mouse hepatic stellate cells into contractile myofibroblasts.


METHODS
Effects of blebbistatin on cell morphology were evaluated by phase contrast microscopy. Cell stress fibres and focal adhesions were investigated by dual immunofluorescence staining and visualized using fluorescence microscopy. Contractile force generation was examined by silicone wrinkle formation assays and collagen gel contraction assays. Intracellular Ca(2+) release in response to endothelin-1 was measured by using Fluo-4. Cell migration was measured by wound healing experiments.


RESULTS
In culture-activated hepatic stellate cells, blebbistatin was found to change both cell morphology and function. In the presence of blebbistatin, stellate cells became smaller, acquired a dendritic morphology and had less myosin IIA-containing stress fibres and vinculin-containing focal adhesions. Moreover, blebbistatin impaired silicone wrinkle formation, reduced collagen gel contraction and blocked endothelin-1-induced intracellular Ca(2+) release. Finally, it promoted wound-induced cell migration.


CONCLUSIONS
By inhibiting myosin II ATPase, blebbistatin has profound effects on the morphology and function of activated hepatic stellate cells. Our data suggest that myosin II could be a therapeutic target in the treatment of liver fibrosis and portal hypertension.
Liu
Zhenan
Z
Department of Cell Biology, Vrije Universiteit Brussel, Brussels, Belgium.
van Grunsven
Leo A
LA
Van Rossen
Elke
E
Schroyen
Ben
B
Timmermans
Jean-Pierre
JP
Geerts
Albert
A
Reynaert
Hendrik
H
D000199:Actins / Q000378:metabolism; D000818:Animals; D002465:Cell Movement / Q000187:drug effects; D048429:Cell Size; D054338:Cell Transdifferentiation / Q000187:drug effects; D002478:Cells, Cultured; D055166:Hepatic Stellate Cells / Q000187:drug effects* / Q000502:physiology / Q000648:ultrastructure; D006576:Heterocyclic Compounds, 4 or More Rings / Q000494:pharmacology*; D051379:Mice; D024462:Myosin Type II / Q000037:antagonists & inhibitors* / Q000502:physiology; D022502:Stress Fibers / Q000187:drug effects / Q000648:ultrastructure
D000199:Actins; D006576:Heterocyclic Compounds, 4 or More Rings; C472645:blebbistatin; D024462:Myosin Type II