Apoptosis of rat hepatic stellate cells induced by anti-focal adhesion kinase antibody

AIM: To explore the role of focal adhesion kinase (FAI)in the apoptosis in culture-activated rat hepatic stellatecells (HSCs) using a specific anti-FAK antibody.METHODS: Rat HSCs were prepared from Wistar rats byin situ perfusion of collagenase and pronase and single-step density Nycodenze gradient. Culture-activatedHSCs were serum-starved and treated with the anti-FAK antibodies for 24, 48 or 72 h. The apoptosis of HSCwas detected by DNA-fragment assay, flow cytometryand caspase-3 activity determination. The expressionof tissue inhibitor of metalloproteinase-1 (TIMP-1)mRNA was assessed by reverse transcriptionpolymerase chain reaction (RT-PCR).RESULTS: The experiment showed that anti-FAKantibodies induced apoptosis of culture-activated ratHSCs. This phenomenon displayed the classical featuresof apoptotic cell death (DNA fragmentation, cell cycleanalysis) after treated with 30 mg@L-1 FAK antibody for72 h, and accompanied by a significant increase ofcespase-3 activity(1208±76) vs (309±28) nmol@min-1@ g-1, t=-208.5, P＜0.05. Meanwhile, treatment with theFAK antibody in HSCs could markedly decrease theTIMP-1 mRNA expression (0.07±0.01 vs 0.38±0.03, t=2.72, P＜0.05).CONCLUSION: FAK plays an important role in the survivalof HSCs and the specific anti-FAK antibody could inducethe apoptosis in rat HSCs.

AIM:To explore the role of focal adhesion kinase(FAK) in the apoptosis in culture-activated rat hepatic stellate cells(HSCs)using a specific anti-FAK antibody. METHODS:Rat HSCs were prepared from Wistar rats by in situ perfusion of collagenase and pronase and single- step density Nycodenze gradient.Culture-activated HSCs were serum-starved and treated with the anti- FAK antibodies for 24,48 or 72 h.The apoptosis of HSC was detected by DNA-fragment assay,flow cytometry and caspase-3 activity determination.The expression of tissue inhibitor of metalloproteinase-1(TIMP-1) mRNA was assessed by reverse transcription polymerase chain reaction(RT-PCR). RESULTS:The experiment showed that anti-FAK antibodies induced apoptosis of culture-activated rat HSCs.This phenomenon displayed the classical features of apoptotic cell death(DNA fragmentation,cell cycle analysis)after treated with 30 mg·L^(-1)FAK antibody for 72 h,and accompanied by a significant increase of caspase-3 activity(1208±76)vs(309±28)nmol·min^(-1) ·g^(-1),t=208.5,P<0.05.Meanwhile,treatment with the FAK antibody in HSCs could markedly decrease the TIMP-1 mRNA expression(0.07±0.01 vs0.38±0.03,t =2.72,P<0.05). CONCLUSION:FAK plays an important role in the survival of HSCs and the specific anti-FAK antibody could induce the apoptosis in rat HSCs.