Vitamin D and S-Farnesylthiosalicylic Acid Have a Synergistic Effect on Hepatic Stellate Cells Proliferation
- 311 Downloads
Hepatic stellate cells (HSCs) have a key role in the formation of hepatic fibrosis. The active form of vitamin D, 1,25(OH)2D3, has been found to have antiproliferative and antifibrotic effects in various tissues including liver. Farnesylthiosalicylic acid (FTS), a novel Ras antagonist, was also found to inhibit hepatic fibrosis.
The purpose of this study was to examine the antiproliferative and antifibrotic effects of the combined treatment of 1,25(OH)2D3 and FTS on primary cultured HSCs.
Primary HSCs, isolated from rat’s livers, were treated with 1,25(OH)2D3, FTS or a combination of both. Proliferation was assessed by bromodeoxyuridine. Expression of p-ERK, ERK, Ras-GTP, total-Ras, CyclinD1 and fibrotic markers was measured by western blotting analysis and real-time PCR. Cytotoxicity was assessed by lactate dehydrogenase method.
The combined treatment inhibited HSCs proliferation by threefold. The effect was synergistic and non-cytotoxic. In concordance, the combined treatment suppressed CyclinD1 expression by ~2-fold, whereas 1,25(OH)2D3 or FTS alone showed a significantly lower inhibitory effect. The effect of the combined treatment on CyclinD1 expression was mediated via Ras-GTP and p-ERK signal transduction pathway. The effect on fibrotic markers showed that 1,25(OH)2D3 decreased collagen Iα1 expression by ~40 %, FTS by ~50 % and the combined treatment by ~60 %. 1,25(OH)2D3 inhibited tissue inhibitor of metalloproteinases-1 (TIMP-1) expression by 20 %. FTS alone or 1,25(OH)2D3 + FTS inhibited TIMP-1 expression by 60 %. FTS inhibited transforming growth factor-β (TGF-β) expression by 25 %, while 1,25(OH)2D3 had no effect.
Although the combination of 1,25(OH)2D3 and FTS did not demonstrate an additive antifibrotic effect, it showed a synergistic antiproliferative effect on primary HSCs. Therefore, the combined treatment may have a potential therapeutic value in the initiation of fibrotic process.
KeywordsLiver 1,25(OH)2D3 FTS Proliferation Fibrosis
This work was performed in partial fulfillment of the M.D. thesis requirements of the Sackler Faculty of Medicine, Tel Aviv University.
Conflict of interest
- 2.Crawford JM. Cirrhosis. In: MacSween RNM, Anthony P, Scheuer PJ, Burt AD, Portmann BC, eds. Pathology of the Liver, 4th edn. Philadelphia, PA: WB Saunders; 2001:575–619.Google Scholar
- 3.Eng F, Friedman SL, Fibrogenesis I. New insight into hepatic stellate cell activation; the simple becomes complex. Am J Physiol. 2000;279:G7–G11.Google Scholar
- 6.Friedman S, Arthur MJ. Activation of cultured rat hepatic lipocytes by Kupffer cell conditioned medium. Direct enhancement of matrix synthesis and stimulation of cell proliferation via induction of platelet-derived growth factor receptors. J Clin Invest. 1989;84:1780–1785.PubMedCrossRefPubMedCentralGoogle Scholar
- 7.Starkel P, Lambotte L, Sempoux C, De Saeger C, et al. After portal branch ligation in the rat, cellular proliferation is associated with selective induction of Ha-Ras, P53, cycline E, and Cdk. Gut. 2000;49:119–130.Google Scholar
- 8.Lavoie JN, L’Allemain G, Brunet A, Muller R, Pouyssegur J. Cyclin D1 expression is regulated positively by the p42/p44 MAPK and negatively by the p38/HOG MAPK pathway. J Biol Chem. 1996;271:20608–20616.Google Scholar
- 9.Kato K, Cox AD, Hisaka MM, Graham SM, Buss JE, Der CJ. Isoprenoid addition to Ras protein is the critical modification for its membrane association and transforming activity. Proc Natl Acad Sci USA. 1992;89:6403–6407.Google Scholar
- 15.Spina C, Ton L, Yao M, Maehr H, et al. Selective vitamin D receptor modulators and their effects on colorectal tumor growth. J Steroid Biochem Mol Biol. 2007;103:757–762.Google Scholar
- 18.Abramovitch S, Dahan Bachar L, Sharvit E, Weisman Y, et al. Vitamin D inhibits proliferation and profibrotic marker expression in hepatic stellate cells and decreases thioacetamide-induced liver fibrosis in rats. Gut. 2011;60:1728–1737.Google Scholar
- 20.Marciano D, Ben-Baruch G, Marom M, Ego Y, Haklai R, Kloog Y. Farnesyl derivatives of rigid carboxylic acids-inhibitors of ras dependent cell growth. J Med Chem. 1995;38:1267–1272.Google Scholar
- 21.Ravid A, Koren R, Narinsky R, Rotem C, Novogrodsky A, Liberman A. 1,25-dihydroxyvitamin D3 and agents that increase intracellular adenosine 3’,5’-monophosphate synergistically inhibit the mitogenic stimulation of human lymphocytes. J Clin Endocrinol Metab. 1990;70:1687–1692.Google Scholar
- 22.Xu J, Kochanek KD, Murphy SL, Tejada-Vera B. Deaths: final data for 2007. Natl Vital Stat Rep. 2010;58:1–136.Google Scholar
- 25.Ding N, Yu RT, Subramaniam N, Sherman MH, Wilson C, Rao R, Leblanc M, Coulter S, He M, Scott C, Lau SL, Atkins AR, Barish GD, Gunton JE, Liddle C, Downes M, Evans RM. A vitamin D receptor/SMAD genomic circuit gates hepatic fibrotic response. Cell. 2013;153:601–613.Google Scholar
- 26.Bouillon R, Bischoff-Ferrari H, Willett W. Vitamin D and health: perspectives from mice and man. J Bone Miner Res. 2008;23: 974–979.Google Scholar
- 29.Benedetti A, DiSario A, Bendia E, Saccomanno, et al. H-Ras inhibition reduces rat liver fibrosis in vivo. Hepatology. 2000;32:302A.Google Scholar