Arg-gly-asp-mannose-6-phosphate inhibits activation and proliferation of hepatic stellate cells in vitro.
OBJECTIVE
To investigate the effect of arg-gly-asp-mannose-6 phosphate (RGD-M6P) on the activation and proliferation of primary hepatic stellate cells in vitro.


METHODS
Hepatic stellate cells (HSCs) were isolated from rats by in situ collagenase perfusion of liver and 18% Nycodenz gradient centrifugation and cultured on uncoated plastic plates for 24 h with DMEM containing 10% fetal bovine serum (FBS/DMEM) before the culture medium was substituted with 2% FBS/DMEM for another 24 h. Then, HSCs were cultured in 2% FBS/DMEM with transforming growth factor beta1, M6P, RGD, or RGD-M6P, respectively. Cell morphology was observed under inverted microscope, smooth muscle alpha-actin (alpha-SMA) was detected by immunocytochemistry, type III procollagen (PC III) in supernatant was determined by radioimmunoassay, and the proliferation rate of HSCs was assessed by flow cytometry.


RESULTS
RGD-M6P significantly inhibited the morphological transformation and the alpha-SMA and PC III expressions of HSCs in vitro and also dramatically prevented the proliferation of HSCs in vitro. Such effects were remarkably different from those of RGD or M6P.


CONCLUSIONS
The new compound, RGD-M6P, which has a dramatic effect on primary cultured HSCs in vitro, can inhibit the transformation of HSCs in culture caused by TGFbeta1, suppresses the expression of PC III and decreases proliferation rate of HSC. RGD-M6P can be applied as a selective drug carrier targeting at HSCs, which may be a new approach to the prevention and treatment of liver fibrosis.
Digestive Disease Laboratory, Xinhua Hospital, Shanghai Second Medical University, Shanghai 200092, China.
Lian-Sheng
LS
Wang
Ying-Wei
YW
Chen
Ding-Guo
DG
Li
Han-Ming
HM
Lu
D000199:Actins / Q000032:analysis; D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D024061:Collagen Type III / Q000032:analysis; D005434:Flow Cytometry; D022781:Hepatocytes / Q000737:chemistry / Q000166:cytology* / Q000187:drug effects* / Q000502:physiology; D007150:Immunohistochemistry; D008103:Liver Cirrhosis / Q000188:drug therapy / Q000473:pathology / Q000517:prevention & control; D008297:Male; D008358:Mannose / Q000031:analogs & derivatives* / Q000494:pharmacology; D009842:Oligopeptides / Q000494:pharmacology*; D011863:Radioimmunoassay; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction; D016212:Transforming Growth Factor beta / Q000502:physiology; D053773:Transforming Growth Factor beta1
D000199:Actins; D024061:Collagen Type III; D009842:Oligopeptides; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C510074:argynylglycylaspartylmannose-6-phosphate; D008358:Mannose