Background & Aims: Directed migration of hepatic myofibroblasts (hMFs) contributes to the development of liver fibrosis. However, the signals regulating the motility of these cells are incompletely understood. We have recently shown that sphingosine 1-phosphate (S1P) and S1P receptors (S1PRs) are involved in mouse liver fibrogenesis. Here, we investigated the role of S1P/S1PRs signals in human liver fibrosis involving motility of human hMFs. Methods: Si P level in the liver was examined by high-performance liquid chromatography. Expression of S1PRs was characterized, in biopsy specimens of human liver and cultured hMFs, by immunofluorescence and real-time RT-PCR or Western blot analysis. Cell migration was determined in Boyden chambers, by using the selective S1P receptor agonist or antagonist and silencing of S1PRs expression with small interfering RNA. Results: Si P level in the human fibrotic liver was increased through up-regulation of sphingosine kinase (SphK), irrespective of the etiology of fibrosis. S1P receptors type 1, 2, and 3 (S1P(1,2,3)) were expressed in human hMFs in vivo and in vitro. Interestingly, S1P(1,3) were strongly induced in human fibrotic samples, whereas expression of S1P(2) was massively decreased. S1P exerted a powerful migratory action on human hMFs. Furthermore, the effect of S1P was mimicked by SEW2871 (an S1P(1) agonist), and blocked by suramin (an S1P(3) antagonist) and by silencing S1P(1,3) expression. In contrast, JTE-013 (an S1P(2) antagonist) and silencing of S1P(2) expression enhanced S1P-induced migration. Conclusions: SphK/S1P/S1PRs signaling axis plays an important role in human liver fibrosis and is involved in the directed migration of human hMFs into the damaged areas. (C) 2010 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.