sphingosine-kinase and Liver-Cirrhosis

Sphingolipids (SL) are a family of bioactive lipids and a major cellular membrane structural component. SLs include three main compounds: ceramide (Cer), sphingosine (Sp), and sphingosine-1-phosphate (S-1P), all of which have emerging roles in biological functions in cells, especially in the liver. They are under investigation in various liver diseases, including cirrhosis and end-stage liver disease. In this review, we provide an overview on the role of SLs in liver pathobiology and focus on their potential role in the development of hepatic fibrosis. We describe recent evidence and suggest SLs are a promising potential therapeutic target for the treatment of liver disease and fibrosis.

Topics: Animals; Cell Proliferation; Enzyme Inhibitors; Extracellular Matrix; Hepatic Stellate Cells; Humans; Kupffer Cells; Liver; Liver Cirrhosis; Phosphotransferases (Alcohol Group Acceptor); Signal Transduction; Sphingolipids

Men with non-alcoholic fatty liver disease (NAFLD) are more likely to progress to non-alcoholic steatohepatitis (NASH) and liver fibrosis than women. However, the underlying molecular mechanisms of this dimorphism is unclear. We have previously shown that mice with global deletion of SphK1, the enzyme that produces the bioactive sphingolipid metabolite sphingosine 1-phosphate (S1P), were protected from development of NASH. The aim of this study was to elucidate the role of hepatocyte-specific SphK1 in development of NASH and to compare its contribution to hepatosteatosis in male and female mice.. We assessed mouse livers in early-stage fibrosis induced by high fat feeding, using single harmonic generation microscopy, LC-MS/MS analysis of hydroxyproline levels, and expression of fibrosis markers. We identified an antifibrotic intercellular signaling mechanism by culturing primary mouse hepatocytes alongside, and in co-culture with, LX2 hepatic stellate cells.. We generated hepatocyte-specific SphK1 knockout mice (SphK1-hKO). Unlike the global knockout, SphK1-hKO male mice were not protected from diet-induced steatosis, inflammation, or fibrogenesis. In contrast, female SphK1-hKO mice were protected from inflammation. Surprisingly, however, in these female mice, there was a ∼10-fold increase in the fibrosis markers Col1α1 and 2-3 fold induction of alpha smooth muscle actin and the pro-fibrotic chemokine CCL5. Because increased fibrosis in female SphK1-hKO mice occurred despite an attenuated inflammatory response, we investigated the crosstalk between hepatocytes and hepatic stellate cells, central players in fibrosis. We found that estrogen stimulated release of S1P from female hepatocytes preventing TGFβ-induced expression of Col1α1 in HSCs via S1PR3.. The results revealed a novel pathway of estrogen-mediated cross-talk between hepatocytes and HSCs that may contribute to sex differences in NAFLD through an anti-fibrogenic function of the S1P/S1PR3 axis. This pathway is susceptible to pharmacologic manipulation, which may lead to novel therapeutic strategies.

Topics: Animals; Chromatography, Liquid; Disease Models, Animal; Estrogens; Female; Humans; Liver Cirrhosis; Male; Mice; Mice, Knockout; Non-alcoholic Fatty Liver Disease; Phosphotransferases (Alcohol Group Acceptor); Sex Characteristics; Tandem Mass Spectrometry

Sphingosine kinase 1 (SphK1) plays critical roles in the activation of hepatic stellate cells (HSCs) and liver fibrosis. Our previous study found that polydatin ameliorates chronic liver injury and fibrosis by inhibiting oxidative stress. However, whether polydatin exerts an anti-fibrotic effect on liver fibrosis dependent on SphK1 signaling is unknown. We aimed to investigate the role of polydatin in SphK1, which mediates HSC activation and liver fibrosis. C57BL/6 mice were induced using CCl

Topics: Animals; Apoptosis; Biomarkers; Cell Line, Transformed; Cell Proliferation; Disease Models, Animal; Enzyme Inhibitors; Glucosides; Hepatic Stellate Cells; Humans; Liver Cirrhosis; Mice; Phosphotransferases (Alcohol Group Acceptor); Signal Transduction; Stilbenes

Chronic liver disease mediated by activation of hepatic stellate cells (HSCs) and Kupffer cells (KCs) leads to liver fibrosis. Here, we aimed to investigate the molecular mechanism and define the cell type involved in mediating the sphingosine kinase (SphK)1-dependent effect on liver fibrosis. The levels of expression and activity of SphK1 were significantly increased in fibrotic livers compared with the normal livers in human. SphK1 was coexpressed with a range of HSC/KC markers including desmin, α-smooth muscle actin (α-SMA) and F4/80 in fibrotic liver. Deficiency of SphK1 (SphK1. SphK1 has distinct roles in the activation of KCs and HSCs in liver fibrosis. Mechanistically, SphK1 in KCs mediates CCL2 secretion, and SphK1 in HSCs upregulates CCR2 by downregulation of miR-19b-3p. (Hepatology 2018).

Topics: Animals; Bone Marrow Transplantation; Chemokine CCL2; Hepatic Stellate Cells; Humans; Kupffer Cells; Liver Cirrhosis; Mice, Inbred C57BL; Mice, Knockout; MicroRNAs; Phosphotransferases (Alcohol Group Acceptor); Receptors, CCR2

There is growing interest in the use of bone marrow cells to treat liver fibrosis, however, little is known about their antifibrotic efficacy or the identity of their effector cell(s). Sphingosine-1-phosphate (S1P) mediates egress of immune cells from the lymphoid organs into the lymphatic vessels; we investigated its role in the response of hematopoietic stem cells (HSCs) to liver fibrosis in mice.. Purified (c-kit+/sca1+/lin-) HSCs were infused repeatedly into mice undergoing fibrotic liver injury. Chronic liver injury was induced in BoyJ mice by injection of carbon tetrachloride (CCl. Infusions of HSCs into mice with liver injury reduced liver scarring based on picrosirius red staining (49.7% reduction in mice given HSCs vs control mice; P < .001), and hepatic hydroxyproline content (328 mg/g in mice given HSCs vs 428 mg/g in control mice; P < .01). HSC infusion also reduced hepatic expression of α-smooth muscle actin (0.19 ± 0.007-fold compared with controls; P < .0001) and collagen type I α 1 chain (0.29 ± 0.17-fold compared with controls; P < .0001). These antifibrotic effects were maintained with infusion of lymphoid progenitors that lack myeloid potential and were associated with increased numbers of recipient neutrophils and macrophages in liver. In studies of HSC cell lines, we found HSCs to recruit monocytes, and this process to require C-C motif chemokine receptor 2. In fibrotic liver tissue from mice and patients, hepatic S1P levels increased owing to increased hepatic sphingosine kinase-1 expression, which contributed to a reduced liver:lymph S1P gradient and limited HSC egress from the liver. Mice given the S1P antagonist (FTY720) with HSCs had increased hepatic retention of HSCs (1697 ± 247 cells in mice given FTY720 vs 982 ± 110 cells in controls; P < .05), and further reductions in fibrosis.. In studies of mice with chronic liver injury, we showed the antifibrotic effects of repeated infusions of purified HSCs. We found that HSCs promote recruitment of endogenous macrophages and neutrophils. Strategies to reduce SIP signaling and increase retention of HSCs in the liver could increase their antifibrotic activities and be developed for treatment of patients with liver fibrosis.

Topics: Actins; Aldehyde-Lyases; Animals; Cell Line; Cell Movement; Chemical and Drug Induced Liver Injury, Chronic; Collagen Type I; Collagen Type I, alpha 1 Chain; Female; Fingolimod Hydrochloride; Gene Expression; Hematopoietic Stem Cell Transplantation; Hematopoietic Stem Cells; Humans; Immunosuppressive Agents; Liver; Liver Cirrhosis; Lymph; Lysophospholipids; Macrophages; Male; Membrane Proteins; Mice; Monocytes; Neutrophils; Phosphoric Monoester Hydrolases; Phosphotransferases (Alcohol Group Acceptor); Sphingosine

Sphingosine-1-phospate is a potent bioactive lipid metabolite that regulates cancer progression. Because sphingosine kinase 1 and sphingosine kinase 2 (SPHK 1/2) are both essential for sphingosine-1-phospate production, they could be a therapeutic target in various cancers. Peretinoin, an acyclic retinoid, inhibits post-therapeutic recurrence of hepatocellular carcinoma via unclear mechanisms. In this study, we assessed effects of peretinoin on SPHK expression and liver cancer development in vitro and in vivo. We examined effects of peretinoin on expression, enzymatic and promoter activity of SPHK1 in a human hepatoma cell line, Huh-7. We also investigated effects of SPHK1 on hepatocarcinogenesis induced by diethylnitrosamine using SPHK1 knockout mice. Peretinoin treatment of Huh-7 cells reduced mRNA levels, protein expression and enzymatic activity of SPHK1. Peretinoin reduced SPHK1 promoter activity; this effect of peretinoin was blocked by overexpression of Sp1, a transcription factor. Deletion of all Sp1 binding sites within the SPHK1 promoter region abolished SPHK1 promoter activity, suggesting that peretinoin reduced mRNA levels of SPHK1 via Sp1. Additionally, diethylnitrosamine-induced hepatoma was fewer and less frequent in SPHK1 knockout compared to wild-type mice. Our data showed crucial roles of SPHK1 in hepatocarcinogenesis and suggests that peretinoin prevents hepatocarcinogenesis by suppressing mRNA levels of SPHK1.

Topics: Animals; Antineoplastic Agents; Carcinoma, Hepatocellular; Cell Line, Tumor; Diethylnitrosamine; Gene Expression Regulation, Enzymologic; Hepatitis C; Humans; Liver; Liver Cirrhosis; Liver Neoplasms; Liver Neoplasms, Experimental; Mice, Knockout; Mice, Transgenic; Phosphotransferases (Alcohol Group Acceptor); Retinoids; Sphingolipids

Topics: Animals; Carbon Tetrachloride; Cell Line; Gene Expression Regulation; Hepatic Stellate Cells; Humans; Indoles; Lipid Metabolism; Liver Cirrhosis; Lysophospholipids; Male; Melatonin; Mice; Phosphotransferases (Alcohol Group Acceptor); Receptors, Lysosphingolipid; Signal Transduction; Sphingosine; Sphingosine-1-Phosphate Receptors

RNA-binding protein HuR mediates transforming growth factor (TGF)-β1-induced profibrogenic actions. Up-regulation of Sphingosine kinase 1 (SphK1) is involved in TGF-β1-induced activation of hepatic stellate cells (HSCs) in liver fibrogenesis. However, the molecular mechanism of TGF-β1 regulates SphK1 remains unclear. This study was designed to investigate the role of HuR in TGF-β1-induced SphK1 expression and identify a new molecular mechanism in liver fibrogenensis. In vivo, HuR expression was increased, translocated to cytoplasm, and bound to SphK1 mRNA in carbon tetrachloride- and bile duct ligation-induced mouse fibrotic liver. HuR mRNA expression had a positive correlation with mRNA expressions of SphK1 and fibrotic markers, α-smooth muscle actin (α-SMA) and Collagen α1(I), respectively. In vitro, up-regulation of SphK1 and activation of HSCs stimulated by TGF-β1 depended on HuR cytoplasmic accumulation. The effects of TGF-β1 were diminished when HuR was silenced or HuR cytoplasmic translocation was blocked. Meanwhile, overexpression of HuR mimicked the effects of TGF-β1. Furthermore, TGF-β1 prolonged half-life of SphK1 mRNA by promoting its binding to HuR. Pharmacological or siRNA-induced SphK1 inhibition abrogated HuR-mediated HSC activation. In conclusion, our data suggested that HuR bound to SphK1 mRNA and played a crucial role in TGF-β1-induced HSC activation.

Topics: Adult; Aged; Animals; ELAV-Like Protein 1; Female; Hepatic Stellate Cells; Humans; Liver Cirrhosis; Male; Mice, Inbred ICR; Middle Aged; Phosphotransferases (Alcohol Group Acceptor); RNA-Binding Proteins; Transforming Growth Factor beta1

The role of sphingosine 1-phosphate (S1P) in liver fibrosis or inflammation was not fully examined in human. Controversy exists which S1P receptors, S1P1 and S1P3 vs S1P2, would be importantly involved in its mechanism. To clarify these matters, 80 patients who received liver resection for hepatocellular carcinoma and 9 patients for metastatic liver tumor were enrolled. S1P metabolism was analyzed in background, non-tumorous liver tissue. mRNA levels of sphingosine kinase 1 (SK1) but not SK2 were increased in livers with fibrosis stages 3-4 compared to those with 0-2 and to normal liver. However, S1P was not increased in advanced fibrotic liver, where mRNA levels of S1P transporter spinster homolog 2 (SPNS2) but not S1P-degrading enzymes were enhanced. Furthermore, mRNA levels of S1P2 but not S1P1 or S1P3 were increased in advanced fibrotic liver. These increased mRNA levels of SK1, SPNS2 and S1P2 in fibrotic liver were correlated with α-smooth muscle actin mRNA levels in liver, and with serum ALT levels. In conclusion, S1P may be actively generated, transported to outside the cells, and bind to its specific receptor in human liver to play a role in fibrosis or inflammation. Altered S1P metabolism in fibrotic liver may be their therapeutic target.

Topics: Aged; Anion Transport Proteins; Female; Gene Expression Regulation; Humans; Liver; Liver Cirrhosis; Male; Phosphotransferases (Alcohol Group Acceptor); Receptors, Lysosphingolipid; RNA, Messenger; Sphingosine-1-Phosphate Receptors

Sphingosine 1-phosphate (S1P) is involved in multiple pathological processes, including fibrogenesis. S1P participates in mouse liver fibrogenesis via a paracrine manner. Herein, we investigated the involvement of S1P in human liver fibrosis. Human fibrotic samples were obtained from livers of patients undergoing liver transplantation. Expression of sphingosine kinase (SphK1), collagen (Col) α1(I), Col α1(III), α-smooth muscle actin, and p-Smad2/3 was characterized by immunofluorescence, real-time RT-PCR, high-content analysis, or Western blot analysis in the fibrotic liver, human bone marrow-derived mesenchymal stem cells, and human hepatogenic profibrotic cells. The effect of SphK1 was assessed using siSphK1 or SphK-specific inhibitor. SphK1, which was expressed in human fibrotic liver myofibroblasts, could be detected in human bone marrow-derived mesenchymal stem cells or human hepatogenic profibrotic cells activated by transforming growth factor β1 (TGF-β1). TGF-β1 evoked the activation of SphK1, increased intracellular S1P, and up-regulated expression of SphK1, Col α1(I), and Col α1(III) in a TGF-β receptor-dependent manner. TGF-β1 induced expression of Col α1(I) and Col α1(III) via SphK1, which was mediated by intracellular S1P, independent of S1P receptors. TGF-β1 evoked nuclear translocation of p-Smad2 and p-Smad3 in TGF-β receptor-dependent, but SphK1-independent, manner. In conclusion, intracellular S1P plays a crucial role in the TGF-β1-induced expression of Col α1(I) and Col α1(III), which is required for human fibrosis development. S1P exerts its effects in S1P receptor-independent manner.

Topics: Adult; Aged; Animals; Collagen Type I; Collagen Type I, alpha 1 Chain; Collagen Type III; Female; Gene Expression Regulation; Humans; Liver Cirrhosis; Lysophospholipids; Male; Mice; Middle Aged; Myofibroblasts; Phosphotransferases (Alcohol Group Acceptor); Smad2 Protein; Smad3 Protein; Sphingosine; Transforming Growth Factor beta1

Sphingosine kinase (SphK)/sphingosine 1-phosphate (S1P)/S1P receptor (S1PR) axis is involved in multiple biological processes, including liver fibrosis. Angiogenesis is an important pathophysiological process closely associated with liver fibrosis; however, the functional role of SphK/S1P/S1PR in this process remains incompletely defined.. Bile duct ligation or carbon tetrachloride was used to induce liver fibrosis in mice. Human fibrotic samples were obtained from livers of patients undergoing liver transplantation. S1P levels in the liver were examined by HPLC. Expression of angiogenic markers, including angiopoietin 1, CD31, vascular cell adhesion molecule-1, and von Willebrand factor, was characterized by immunofluorescence, real-time RT-PCR, and Western blot in the fibrotic liver and primary mouse hepatic stellate cells (HSCs). SphK inhibitor (SKI) or S1PR antagonists were administered intraperitoneally in mice.. S1P levels in the liver were closely correlated with mRNA expression of angiogenic markers. Ang1 is expressed in activated HSCs of the fibrotic liver and in primary HSCs. In HSCs, by using specific antagonists or siRNAs, we demonstrated S1P stimulation induced Ang1 expression via S1PR1 and S1PR3. In vivo, S1P reduction by SKI inhibited angiogenesis in fibrotic mice. Furthermore, S1PR1/3 antagonist significantly blocked upregulation of angiogenic markers in the injured liver, and attenuated the extent of liver fibrosis, while S1PR2 antagonist had no effect on angiogenesis, supporting the key role of S1PR1 and S1PR3 in angiogenesis underlying liver fibrosis process.. SphK1/S1P/S1PR1/3 axis plays a crucial role in the angiogenic process required for fibrosis development, which may represent an effective therapeutic strategy for liver fibrosis.

Topics: Adult; Aged; Angiopoietin-1; Animals; Female; Hepatic Stellate Cells; Humans; Liver; Liver Cirrhosis; Liver Cirrhosis, Experimental; Lysophospholipids; Male; Mice; Mice, Inbred ICR; Middle Aged; Neovascularization, Pathologic; Phosphotransferases (Alcohol Group Acceptor); Receptors, Lysosphingolipid; Signal Transduction; Sphingosine

Sphingosine kinase (SphK) is involved in numerous biological processes, including cell growth, proliferation, and differentiation. However, whether SphK participates in the differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) to myofibroblasts has been unknown. In a carbon tetrachloride-treated mouse model, SphK1 was expressed in BMSCs in damaged liver. Furthermore, mRNA expression of both SphK1 and transforming growth factor β1 (TGF-β1) was significantly increased after liver injury, with a positive correlation between them. The SphK inhibitor SKI significantly blocked BMSC differentiation to myofibroblasts during liver injury (the proportion of BMSC-derived myofibroblasts decreased markedly, compared with no SKI treatment) and attenuated the extent of liver fibrosis. Using primary mouse BMSCs, we demonstrated that TGF-β1 induced BMSC differentiation to myofibroblasts, accompanied by the up-regulation of SphK1 and modulation of sphingosine 1-phosphate (S1P) receptor (S1PR) expression. Notably, pharmacological or siRNA-mediated inhibition of SphK1 abrogated the prodifferentiating effect of TGF-β1. Moreover, using either S1PR subtype-specific antagonists or specific siRNAs, we found that the prodifferentiating effect of TGF-β1 was mediated by S1PR(1) and S1PR(3). These data suggest that SphK1 activation by TGF-β1 leads to differentiation of BMSCs to myofibroblasts mediated by S1PR(1) and S1PR(3) up-regulation, thus providing new information on the mechanisms by which TGF-β1 gives rise to fibrosis and opening new perspectives for pharmacological treatment of liver fibrosis.

Topics: Animals; Bone Marrow; Carbon Tetrachloride; Cell Differentiation; Cell Proliferation; Cells, Cultured; Enzyme Inhibitors; G(M1) Ganglioside; Gene Knockdown Techniques; Liver Cirrhosis; Mesenchymal Stem Cells; Mice; Mice, Inbred ICR; Myofibroblasts; Phosphotransferases (Alcohol Group Acceptor); Receptors, Lysosphingolipid; RNA, Small Interfering; Transforming Growth Factor beta1; Up-Regulation

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.. S1P 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.. S1P 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.. 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.

Topics: Base Sequence; Cell Movement; Cells, Cultured; Humans; Liver Cirrhosis; Lysophospholipids; Myofibroblasts; Oxadiazoles; Phosphotransferases (Alcohol Group Acceptor); Receptors, Lysosphingolipid; RNA, Messenger; RNA, Small Interfering; Signal Transduction; Sphingosine; Sphingosine-1-Phosphate Receptors; Suramin; Thiophenes

Myofibroblasts play a central role in the pathogenesis of liver fibrosis. Myofibroblasts of bone marrow (BM) origin have recently been identified in fibrotic liver. However, little is known about the mechanism that controls their mobilization in vivo. Here we confirmed that BM mesenchymal stem cells (BMSCs) can migrate to the damaged liver and differentiate into myofibroblasts. We also investigated the mechanism underlying the homing of BMSCs after liver injury.. ICR mice were lethally irradiated and received BM transplants from enhanced green fluorescent protein transgenic mice. Carbon tetrachloride or bile duct ligation was used to induce liver fibrosis. The fibrotic liver tissue was examined by immunofluorescent staining to identify BM-derived myofibroblasts.. BMSCs contributed significantly to myofibroblast population in fibrotic liver. Moreover, analysis in vivo and in vitro suggested that homing of BMSCs to the damaged liver was in response to sphingosine 1-phosphate (S1P) gradient between liver and BM. Furthermore, S1P receptor type 3 (S1P3) was required for migration of BMSCs triggered by S1P.. S1P mediates liver fibrogenesis through homing of BMSCs via S1P3 receptor, which may represent a novel therapeutic target in liver fibrosis through inhibiting S1P formation and/or receptor activation.

Topics: Animals; Base Sequence; Bone Marrow Cells; Bone Marrow Transplantation; Cell Differentiation; Cell Movement; DNA Primers; Green Fluorescent Proteins; In Vitro Techniques; Liver Cirrhosis; Lysophospholipids; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Inbred ICR; Mice, Transgenic; Phosphotransferases (Alcohol Group Acceptor); Receptors, Lysosphingolipid; Recombinant Proteins; Sphingosine; Suramin; Transforming Growth Factor beta1