title: Vitamin D and S-farnesylthiosalicylic acid have a synergistic effect on hepatic stellate cells proliferation.
abstract: BACKGROUND
  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.
  
  
  OBJECTIVE
  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.
  
  
  METHODS
  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.
  
  
  RESULTS
  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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Sackler Faculty of Medicine, Tel Aviv University, Tel-Aviv, Israel, rinaneeman1@gmail.com.
authors.forename: Rina
authors.initials: R
authors.lastname: Neeman
authors.forename: Shirley
authors.initials: S
authors.lastname: Abramovitch
authors.forename: Efrat
authors.initials: E
authors.lastname: Sharvit
authors.forename: Galit
authors.initials: G
authors.lastname: Elad-Sfadia
authors.forename: Roni
authors.initials: R
authors.lastname: Haklai
authors.forename: Yoel
authors.initials: Y
authors.lastname: Kloog
authors.forename: Shimon
authors.initials: S
authors.lastname: Reif
mesh_terms: D000818:Animals; D015415:Biomarkers; D002117:Calcitriol / Q000008:administration & dosage / Q000494:pharmacology*; D049109:Cell Proliferation; D019938:Cyclin D1; D004357:Drug Synergism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000378:metabolism; D005204:Farnesol / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D055166:Hepatic Stellate Cells / Q000187:drug effects* / Q000502:physiology; D008297:Male; D051381:Rats; D017208:Rats, Wistar; D012459:Salicylates / Q000008:administration & dosage / Q000494:pharmacology*; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D015415:Biomarkers; D012459:Salicylates; C093323:farnesylthiosalicylic acid; D019938:Cyclin D1; D005204:Farnesol; D048049:Extracellular Signal-Regulated MAP Kinases; D018631:ras Proteins; D002117:Calcitriol