No abstract

Sphingosine 1-phosphate (S1P), a bioactive lipid mediator, stimulates proliferation and contractility in hepatic stellate cells, the principal matrix-producing cells in the liver, and inhibits proliferation via S1P receptor 2 (S1P 2 ) in hepatocytes in rats in vitro. A potential role of S1P and S1P 2 in liver regeneration and fibrosis was examined in S1P 2 -deficient mice. Nuclear 5-bromo-2′-deoxy-uridine labeling, proliferating cell nuclear antigen (PCNA) staining in hepatocytes, and the ratio of liver weight to body weight were enhanced at 48 h in S1P 2 -deficient mice after a single carbon tetrachloride (CCl 4 ) injection. After dimethylnitrosamine (DMN) administration with a lethal dose, PCNA staining in hepatocytes was enhanced at 48 h and survival rate was higher in S1P 2 -deficient mice. Serum aminotransferase level was unaltered in those mice compared with wild-type mice in both CCl 4 -and DMN-induced liver injury, suggesting that S1P 2 inactivation accelerated regeneration not as a response to enhanced liver damage. After chronic CCl 4 administration, fibrosis was less apparent, with reduced expression of smooth-muscle a-actin-positive cells in the livers of S1P 2 -deficient mice, suggesting that S1P 2 inactivation ameliorated CCl 4 -induced fibrosis due to the decreased accumulation of hepatic stellate cells. Thus, S1P plays a significant role in regeneration and fibrosis after liver injury via S1P 2 . Sphingosine 1-phosphate (S1P), which elicits a wide variety of cell responses (1), has emerged as a novel lipid intracellular mediator. S1P was shown to act as an intracellular second messenger of platelet-derived growth factor and serum in their mitogenic actions in cultured fibroblasts (2, 3), and furthermore, intracellular levels of S1P and ceramide were reported to determine cell survival or death (4, 5). However, evidence indicating that S1P also acts as an extracellular mediator has been reported; some of the diverse effects of S1P, such as stimulation of cell proliferation or contractility, are known to be sensitive to pertussis toxin (6) or ADP-ribosyltransferase C3 from Clostridium botulinum (7, 8), suggesting that S1P may activate a receptor coupled to G protein(s). Indeed, recent investigation has revealed that S1P acts through at least five high-affinity G proteincoupled receptors referred to as S1P 1-5 (9, 10). Regarding the source of S1P in vivo, it is shown to be stored in platelets (11), and recent data using conditional knockouts of sphingosine kinases support release of S1P from erythrocytes (12, 13). These findings suggest that S1P has normal in vivo roles as well as potentially pathophysiological roles as a circulating paracrine mediator, a view further supported by the phenotypes of S1P receptor mutants (10,14,15). S1P receptors are also expressed in the liver (14). To investigate the function of S1P in liver pathophysiology, we have determined the effect of S1P on liver cells in culture. We first demonstrated that S1P stimulates proliferation and contractility in rat hepati...

The clinical superiority of REG over SOR is partially attributable to reduced MICA shedding via transcriptional suppression of ADAM9 and ADAM10.

Sinusoidal vasoconstriction, in which hepatic stellate cells operate as contractile machinery, has been suggested to play a pivotal role in the pathophysiology of portal hypertension. We investigated whether sphingosine 1-phosphate (S1P) stimulates contractility of those cells and enhances portal vein pressure in isolated perfused rat livers with Rho activation by way of S1P receptor 2 (S1P 2 ). Rho and its effector, Rho kinase, reportedly contribute to the pathophysiology of portal hypertension. Thus, a potential effect of S1P 2 antagonism on portal hypertension was examined. Intravenous infusion of the S1P 2 antagonist, JTE-013, at 1 mg/kg body weight reduced portal vein pressure by 24% without affecting mean arterial pressure in cirrhotic rats induced by bile duct ligation at 4 weeks after the operation, whereas the same amount of S1P 2 antagonist did not alter portal vein pressure and mean arterial pressure in control sham-operated rats. Rho kinase activity in the livers was enhanced in bile duct-ligated rats compared to sham-operated rats, and this enhanced Rho kinase activity in bile duct-ligated livers was reduced after infusion of the S1P 2 antagonist. S1P 2 messenger RNA (mRNA) expression, but not S1P 1 or S1P 3 , was increased in bile ductligated livers of rats and mice and also in culture-activated rat hepatic stellate cells. S1P 2 expression, determined in S1P LacZ =þ 2 mice, was highly increased in hepatic stellate cells of bile duct-ligated livers. Furthermore, the increase of Rho kinase activity in bile duct-ligated livers was observed as early as 7 days after the operation in wildtype mice, but was less in S1P À=À 2 mice. Conclusion: S1P may play an important role in the pathophysiology of portal hypertension with Rho kinase activation by way of S1P 2 . The S1P 2 antagonist merits consideration as a novel therapeutic agent for portal hypertension.

In our previous study on hepatocellular carcinoma (HCC) susceptibility genes in chronic hepatitis patients, we identified the MHC class I polypeptide-related sequence A (MICA). Natural killer cells eliminate various cancer cells, including HCC, by suppressing MICA shedding. Therefore, we investigated MICA sheddases and inhibitors for HCC immunotherapy. In this study, HepG2, PLC/PRF/5, and Hep3B were treated with the siRNA of a disintegrin and metalloproteases (ADAMs) and matrix metalloproteases to measure the concentration of soluble MICA (sMICA) by ELISA to detect the therapeutic target. Furthermore, an FDA-approved drug library was tested for the enzymatic inhibition of the targeted enzyme in an in vitro drug screening assay system. ADAM17 knockdown reduced sMICA levels and increased membrane-bound MICA (mMICA) expression in HCC cells. In an in vitro drug screen using an FDA-approved drug library, lomofungin, an antifungal drug, was found to strongly decrease ADAM17 activity. In HCC cells, mMICA expression was induced and sMICA production was inhibited in a dose-dependent manner. These effects were cancelled upon ADAM17 knockdown, suggesting that lomofungin targeted ADAM17. Analysis of lomofungin analogs revealed the responsible functional groups. In summary, we suggest lomofungin to be an attractive agent for the immunological control of HCC, via the suppression of ADAM17.