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


CONCLUSIONS
FAK plays an important role in the survival of HSCs and the specific anti-FAK antibody could induce the apoptosis in rat HSCs.
Liu
Xiao-Jing
XJ
Laboratory of Department of Internal Medicine, West China Hospital, Sichuan University, 37 Wainan Guoxueshang, Chengdu 610041, Sichuan Province, China. xiaojingliu67@hotmail.com
Yang
Li
L
Wu
Hong-Bin
HB
Qiang
Ou
O
Huang
Ming-Hui
MH
Wang
Ying-Ping
YP
D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D053938:DNA Fragmentation; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D015870:Gene Expression; D022781:Hepatocytes / Q000166:cytology* / Q000201:enzymology*; D066298:In Vitro Techniques; D008297:Male; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D051381:Rats; D017208:Rats, Wistar; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics
D000906:Antibodies; D019715:Tissue Inhibitor of Metalloproteinase-1; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496052:Ptk2 protein, rat; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases