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

摘要

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 vs 0.38 ± 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.