[Effects of Arg-Gly-Asp-Ser tetrapeptide on integrin signaling and apoptosis in hepatic stellate cells].
OBJECTIVE
To investigate the effects of Arg-Gly-Asp-Ser (RGDS) tetrapeptide on integrin signaling and apoptosis in fibronectin (FN) -stimulated hepatic stellate cells (HSCs).


METHODS
3H-thymidine incorporation, annexin-V/propidium iodide double-labeled flow cytometry (FCM) and transmission electron microscopy were employed to estimate the influence of RGDS on the proliferation and apoptosis of HSCs. And the adhesion rates were observed by toluidine blue colorimetric assay. The expression of focal adhesion kinase (FAK) mRNA and protein in HSCs was detected using RT-PCR and western blotting analysis, respectively.


RESULTS
RGDS tetrapeptide at the concentrations of 25 microg/ml, 50 microg/ml and 100 microg/ml inhibited the proliferation of HSCs and induced HSCs apoptosis in dose-dependent and time-dependent manners, with the apoptotic rates of 9.49%, 27.67%, 31.59%, and the necrotic rates of 3.47%, 5.38%, 9.10%, respectively. Both the rates were higher than those in FN group (apoptotic rate: 3.44%; necrotic rate: 2.39%), F=8.02, P<0.05. After adding RGDS tetrapeptide to HSCs for 2 hours, the adhesive inhibition rates were 8.82%, 29.41% and 45.59%, respectively, which were higher than that in FN group (F=20.58, P<0.01). After exposure of HSCs to RGDS tetrapeptide for 24 hours, FAK protein decreased, and FAK mRNA was down-regulated earlier, about 2 hours after exposure to RGDS tetrapeptide.


CONCLUSIONS
These results suggest that RGDS tetrapeptide may induce apoptosis of HSC in both dose-dependent and time-dependent manners in vitro, which may be related to the disruption of cell matrix adhesion and down-regulation of FAK expression.
Zhang
Xiao-lan
XL
Department of Gastroenterology, Second Hospital of Hebei Medical University, Shijiazhuang 050000, China.
Jiang
Hui-qing
HQ
Liu
Li
L
Bai
Yun
Y
Song
Mei
M
D017209:Apoptosis / Q000187:drug effects*; D005353:Fibronectins / Q000494:pharmacology; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D022781:Hepatocytes / Q000166:cytology / Q000502:physiology; D006801:Humans; D016023:Integrins / Q000378:metabolism* / Q000502:physiology; D008099:Liver / Q000166:cytology* / Q000502:physiology; D009842:Oligopeptides / Q000494:pharmacology*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction
D005353:Fibronectins; D016023:Integrins; D009842:Oligopeptides; D010975:Platelet Aggregation Inhibitors; C046617:arginyl-glycyl-aspartyl-serine; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human