Abstract
Rapamycin is an immunosuppressant with antiproliferative properties. We investigated whether rapamycin treatment of bile duct-ligated (BDL) rats is capable of inhibiting liver fibrosis and thereby affecting hemodynamics. Following BDL, rats were treated for 28 days with rapamycin (BDL SIR). BDL animals without drug treatment (BDL CTR) and sham-operated animals served as controls. After 28 days, hemodynamics were measured, and livers were harvested for histology/immunohistochemistry. Liver mRNA levels of transforming growth factor (TGF)-β1, connective tissue growth factor (CTGF), platelet-derived growth factor (PDGF)-β, cyclin-dependent kinase inhibitor p27kip (p27), and cyclin-dependent kinase inhibitor p21WAF1/CIP1 (p21) were quantified by real-time polymerase chain reaction. Liver protein levels of p27, p21, p70 S6 kinase (p70s6k), phosphorylated p70s6k (p-p70s6k), eukaryotic initiation factor 4E-binding protein (4E-BP1), p-4E-BP1 (Thr37/46), and p-4E-BP1 (Ser65/Thr70) were determined by Western blotting. Portal vein pressure was lower in BDL SIR than in BDL CTR animals. Volume fractions of connective tissue, bile duct epithelial, and desmin- and actin-positive cells were lower in BDL SIR than in BDL CTR rats. On the mRNA level, TGF-β1, CTGF, and PDGF were decreased by rapamycin. p27 and p21 mRNA did not differ. On the protein level, rapamycin increased p27 and decreased p21 levels. Levels of nonphosphorylated p70s6k and 4E-BP1 did not vary between groups, but levels of p-p70s6k were decreased by rapamycin. Rapamycin had no effect on p-4E-BP1 (Thr37/46) and p-4E-BP1 (Ser65/Thr70) levels. In BDL rats, rapamycin inhibits liver fibrosis and ameliorates portal hypertension. This is paralleled by decreased levels of TGF-β1, CTGF, and PDGF. Rapamycin influences the cell cycle by up-regulation of p27, down-regulation of p21, and inhibition of p70s6k phosphorylation.
Footnotes
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This study was supported by Grant 3200-063476 from the Swiss National Foundation for Scientific Research (to J.R.).
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doi:10.1124/jpet.104.079616.
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ABBREVIATIONS: HSC, hepatic stellate cell(s); PDGF, platelet-derived growth factor; TGF, transforming growth factor; CTGF, connective tissue growth factor; FK506, tacrolimus; FKBP, FK506-binding protein; mTOR, mammalian target of rapamycin; PI3K, phosphoinositide-3-kinase; p70s6k, p70 S6 kinase; eIF, eukaryotic initiation factor; 4E-BP1, eukaryotic initiation factor 4E-binding protein; pRb, retinoblastoma protein; p27, cyclin-dependent kinase inhibitor p27kip; p21, cyclin-dependent kinase inhibitor p21WAF1/CIP1; BDL, bile duct-ligated/bile duct ligation; BDL SIR, BDL rats with rapamycin treatment; BDL CTR, BDL rats without rapamycin treatment; PCR, polymerase chain reaction; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; PAGE, polyacrylamide gel electrophoresis; HEK, human embryonic kidney; SMA, smooth muscle actin; CAB, chromotrope-anilin blue; AST, aspartate aminotransferase; NRC, normal rat cholangiocyte(s).
- Received October 22, 2004.
- Accepted March 2, 2005.
- The American Society for Pharmacology and Experimental Therapeutics
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