Dexamethasone up-regulates the inhibitory adaptor protein Dok-1 and suppresses downstream activation of the mitogen-activated protein kinase pathway in antigen-stimulated RBL-2H3 mast cells

The glucocorticoid dexamethasone suppresses antigen-induced degranulation, cytokine production, and intermediate signaling events in RBL-2H3 mast cells, although the exact mechanisms are uncertain. By microarray analysis, we discovered that expression of the inhibitory adaptor protein, downstream of tyrosine kinase (Dok)-1, was up-regulated 4-fold in dexamethasone-treated RBL-2H3 cells. The up-regulation was apparent with as little as 1 to 10 nM dexamethasone. Treatment with dexamethasone also enhanced tyrosine phosphorylation of Dok-1, augmented recruitment of Ras GTPase-activating protein (RasGAP) by Dok-1, and inhibited activation of the mitogen-activated protein (MAP) kinase pathway in antigen-stimulated cells. The same effects were obtained by transient overexpression of Dok-1 but not by overexpression of Dok-1 that was mutated in RasGAP-binding domain. The negative regulatory role of Dok-1 was further validated by the expression of small interfering RNA directed against Dok-1, which enhanced activation of MAP kinase and subsequent release of arachidonic acid and tumor necrosis factor-alpha. These findings identify Dok-1 as mediator of the antiallergic actions of dexamethasone and as a negative regulator of the MAP kinase pathway and downstream release of inflammatory mediators.     

Infection of a Japanese patient by genotype 4 hepatitis e virus while traveling in Vietnam

Cases of imported hepatitis E in industrialized countries infected with a genotype 1 hepatitis E virus (HEV) have been identified. We report a 56-year-old Japanese man who acquired infection with a genotype 4 HEV with 98.8% identity to a Vietnamese isolate after ingestion of uncooked shellfish while traveling in Vietnam.     

Immunohistochemical expression of 14-3-3 sigma protein in various histological subtypes of uterine cervical cancers

14-3-3 sigma (σ) has been a major G2/M checkpoint control gene and has demonstrated that its inactivation in various cancers occurs mostly by epigenetic hypermethylation, not by genetic change. In order to confirm 14-3-3σ protein expression together with p16 and p53 in cervical cancers, immunohistochemistry was performed using various histological subtypes of cervical cancers and dysplasia. Strong and diffuse immunoreactivity for 14-3-3σ was uniformly observed in all the cervical dysplasia (17/17) and squamous cell carcinomas (29/29) including human papillomavirus (HPV)-negative cases. Even in adenosquamous carcinomas and adenocarcinomas of the cervix, immunohistochemical expression of 14-3-3σ was shown with relatively high frequency (13/15, 87% and 22/27, 81%). In the in situ hybridization study, mRNA of 14-3-3σ was expressed in six of eight immunohistochemical-negative cases. Therefore, the undetectable expression of 14-3-3σ protein in cervical cancers might, at least in part, be due to a proteolysis not epigenetic hypermethylation. It is of interest that cancers without 14-3-3σ expression were predominantly those lacking HPV DNA, and that there were no cases with concomitant inactivation of 14-3-3σ and p16 in the present study. These observations are consistent with the hypothesis that inactivation of either 14-3-3σ or p16 has an effect equivalent to the expression of E6 and E7 oncoproteins of HPV.     

Involvement of multiple transporters in the efflux of 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors across the blood-brain barrier

Statins, 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors, are frequently used for the treatment of hypercholesterolemia. The present study aimed to examine the involvement of organic anion transporters in the efflux transport of pravastatin and pitavastatin across the blood-brain barrier (BBB). Transport studies using cDNA-transfected cells revealed that these statins are substrates of multispecific organic anion transporters expressed at the BBB (rOat3:Slc22a8 and rOatp2:Slco1a4). The efflux of these statins across the BBB was characterized using the brain efflux index method. The efflux clearance of pitavastatin across the BBB, obtained from the elimination rate constant and the distribution volume in the brain, was greater than that of pravastatin (364 versus 59 microl/min/g brain). The efflux of pravastatin and pitavastatin was saturable (apparent Km values: 18 and 5 muM, respectively) and inhibited by probenecid but unaffected by tetraethylammonium. Furthermore, an inhibitor of the efflux pathway for hydrophilic organic anions across the BBB (p-aminohippurate), and inhibitors of the efflux pathway for amphipathic organic anions (taurocholate and digoxin) inhibited the efflux of both statins. The degree of inhibition by p-aminohippurate was similar and partial for the efflux of pravastatin and pitavastatin. Taurocholate and digoxin completely inhibited the efflux of pitavastatin, whereas their effect was partial for the efflux of pravastatin. The results of the present study suggest the involvement of multiple transporters, including rOat3 and rOatp2, in the efflux transport of pravastatin and pitavastatin across the BBB, each making a different contribution.