The interaction of anthocyanins with bilitranslocase

Bilitranslocase (TC 2.A.65.1.1) is an organic anion membrane carrier expressed at the sinusoidal domain of the liver plasma membrane and in epithelial cells of the gastric mucosa. Its substrates are sulfobromophthalein, bilirubin, and nicotinic acid. This work reports on the identification of a new class of bilitranslocase substrates, i.e., anthocyanins. Seventeen out thes 20 compounds tested behaved as competitive inhibitors of bilitranslocase transport activity (K(I)=1.4-22 microM). Their structure-activity relationship reveals that mono- and di-glucosyl anthocyanins, the anthocyanin species occurring in food, are better ligands than the corresponding aglycones. Moreover, the first interaction of anthocyanins with the carrier occurs through hydrophilic moieties, such as the 3-glucosyl moiety and the B ring for the monoglucosides, through the 5-glucosyl moiety and the A ring for the diglucosides, and through either the B or the A ring for the aglycones. These findings suggest that bilitranslocase could play a role in the bioavailability of anthocyanins.     

A review of the role of reactive oxygen and nitrogen species in alcohol-induced mitochondrial dysfunction

Our understanding of the mechanisms involved in the development of alcohol-induced liver disease has increased substantially in recent years. Specifically, reactive oxygen and nitrogen species have been identified as key components in initiating and possibly sustaining the pathogenic pathways responsible for the progression from alcohol-induced fatty liver to alcoholic hepatitis and cirrhosis. Ethanol has been demonstrated to increase the production of reactive oxygen and nitrogen species and decrease several antioxidant mechanisms in liver. However, the relative contribution of the proposed sites of ethanol-induced reactive species production within the liver is still not clear. It has been proposed that chronic ethanol-elicited alterations in mitochondria structure and function might result in increased production of reactive species at the level of the mitochondrion in liver from ethanol consumers. This in turn might result in oxidative modification and inactivation of mitochondrial macromolecules, thereby contributing further to mitochondrial dysfunction and a loss in hepatic energy conservation. Moreover, ethanol-related increases in reactive species may shift the balance between pro- and anti-apoptotic factors such that there is activation of the mitochondrial permeability transition, which would lead to increased cell death in the liver after chronic alcohol consumption. This article will examine the critical role of these reactive species in ethanol-induced liver injury with specific emphasis on how chronic ethanol-associated alterations to mitochondria influence the production of reactive oxygen and nitrogen species and how their production may disrupt hepatic energy conservation in the chronic alcohol abuser.     

Influence of culture time on the expression of drug-metabolizing enzymes in primary human hepatocytes and hepatoma cell line HepG2

Primary cultures of human hepatocytes and hepatoma cell line HepG2 are frequently used to evaluate the hepatic disposition of drugs and other xenobiotics. To check the variability of the expression of drug-metabolizing enzymes in these in vitro models, expression of genes coding for several cytochrome P450 isoforms and phase II enzymes was quantified during culture time by real-time RT-PCR. Gene expression was determined daily for primary hepatocytes maintained in a sandwich culture over 1 week and for HepG2, during the first 10 passages. In primary hepatocytes characteristic expression trends were observed which could be abstracted into three major classes of time curves. Genes of the first and the second class had an expression maximum around day 6 and day 4 in culture, respectively. The third class of genes had two expression peaks: at day 1 and 5 in culture. Surprisingly, also the cell line HepG2 showed significant expression changes during passages. For example, gene expression of cytochrome 1A1 varied 8-fold, that of cytochrome 2B6 30-fold, and that of NADP-quinone reductase 1 more than 200-fold within the first 10 passages. In conclusion, neither primary hepatocytes nor HepG2 cell line display a model for constant expression of drug-metabolizing enzymes.     

Decrease of plasma taurine in Gaucher disease and its sustained correction during enzyme replacement therapy

Summary. Gaucher disease is caused by an autosomal-recessive deficiency of glucocerebrosidase. Cells of monocytic/macrophagic origin accumulate glucosylceramide. This leads to hepatosplenomegaly, bone destruction, thrombocytopenia and anemia. Enzyme replacement therapy (ERT) with macrophage-targeted glucocerebrosidase leads to normalization of these parameters. The way of macrophage activation in Gaucher disease is not known. Recently, the osmolytes taurine, betaine and inositol were identified as important regulators of macrophage function in liver. Therefore, the role of plasma taurine in Gaucher disease as a primarily macrophage-derived disease was studied. Fasting plasma levels were measured from blood samples of healthy control subjects (n = 29, m : f = 11 : 18, mean age 37 ± 3 years), from un-treated Gaucher patients (n = 16, m : f = 7 : 9, mean age 44 ± 4 years) and those treated for 37 ± 2 months (n = 54, m : f = 19 : 35, mean age 47 ± 2 years). Amino acid analysis was carried out in a BioChrom amino acid analyzer. In the untreated patients, plasma taurine was 45 ± 3 μM, as compared to the controls with a plasma taurine of 63 ± 4 μM (p < 0.01). The aver-age increase of plasma taurine during the first year of ERT was 18 ± 8 μM (n = 10). Patients treated for an average of 37 months (range 1–9 years of ERT) had a plasma taurine of 65 ± 4 μM (n = 54), which was not different from the controls. It is concluded that Gaucher patients show decreased plasma taurine levels and that therapy of Gaucher disease might correct this. It has to be established, whether decreased taurine availability is a cofactor of the permanent activation of glucosylceramide-storing monocytes/macrophages in this disease. Received January 25, 2000/Accepted January 31, 2000     

Tissue steroid sulfatase levels, testosterone and blood pressure

The objective of this study was to examine the response of tissue steroid sulfatase (STS) levels in hypertensive rat strains, when blood pressure (BP) was lowered by different techniques at an early age. A 4×3 factoral design was used, in which males (n=6–8) from four rat strains (WKY, SHR, SHR/a, SHR/y) at 4 weeks of age, were randomly assigned to one of three treatment groups: a hydralazine group, a castration group and a control group. BP was measured by the tail cuff technique and verified by tail catheter at the end of the experiment. BP was significantly reduced by both treatments in the hypertensive strains (SHR, SHR/a, SHR/y) compared to respective control groups. At 15–17 weeks of age, animals were euthanized and heart, kidney, adrenal glands and liver were assayed for STS levels. The major trend in tissue STS was that castration significantly lowered: adrenal, heart and liver STS in specific strains. In conclusion, castration and hydralazine significantly lowered the BP in the hypertensive rat strains, but only castration consistently lowered STS levels across strains implicating testosterone as a regulator of tissue STS.     

异种血清诱发肝纤维化模型中ECM变化的研究

为了观察异种血清诱发肝纤维化过程中细胞外基质（ECM）及其生成细胞的变化规律,经腹腔注射猪血清建立大鼠肝纤维化模型,以免疫组织化学方法显示肝内胶原（Col）Ⅰ、Ⅲ、Ⅳ、Ⅴ型和纤连蛋白（Fn）及其生成细胞。结果：①Fn最早在汇管区、小叶间隔内沉积,ColⅢ、Ⅳ、Ⅴ随之增多,停止血清注射后它们都明显减弱或消失;ColⅠ沉积出现最迟,但阳性迅速增强,停止注射后仍较强;②结蛋白（Dm）阳性的肝星状细胞（HSC,原称Ito细胞）其分布和数量变化与Fn和ColⅡ基本同步,而位于汇管区及细胞间隔的原始间叶细胞（PMC）Dm（-）。提示ECM生成细胞可能来自细胞骨架表型不同的PMC和HSC,本模型由于能清晰地显示ECM及其生成细胞而更适用于肝纤维化机制及防治的研究。     

肝源性肝细胞再生刺激因子(HSS)的制备及鉴定

本文采用新鲜乳牛肝脏作为原料。经酸处理、超滤、分离得到肝源性肝细胞再生刺激因子（ＨＳＳ）。ＳＤＳ－ＰＡＧＥ测得ＨＳＳ的蛋白质分子量为１２,０００～１５,０００Ｄａ。经动物试验和生物学活性鉴定,表明本法制备的ＨＳＳ符合《中国生物制品规程》（１９９５版）的有关要求,且工艺简便、有效     

Mutant p21ras in vinyl chloride exposed workers

The production of mutations in cellular oncogenes such as ras is involved in the development of many human cancers. These mutations result in the expression of mutant forms of the encoded p21 protein which can potentially serve as a biomarker for this carcinogenic process. Workers exposed to vinyl chloride (VC) who are at risk for the development of the sentinel neoplasm angiosarcoma of the liver (ASL) represent a model population for the study of such a mutant p21ras biomarker, since VC is known to cause a specific ras mutation in ASL. In order to determine the relationship between VC exposure and this biomarker, serum samples from a cohort of 225 French VC workers and 111 age-sex-race-smoking-drinking matched unexposed controls were examined for the presence of mutant p21ras by immunoblotting with a mouse monoclonal antibody specific for the mutant protein. Stratifying the exposed workers by degree of VC exposure in estimated ppm-years by quartiles yielded a statistically significant trend for increasing odds ratio for sero-positivity of the p21ras biomarker with increasing exposure. These results suggest that this serum biomarker is related to VC exposure and may be an early indicator of carcinogenic risk in exposed individuals.     

Molecular cloning of HCV and clinical application

Abstract: Fifty-five clones encoding epitopes of HCV were isolated from Japanese patients. Their amino acid homology (AAH) to the sequence of prototype (HCV-1) ranged from 47% to 94%. These sequences cover 60% of the HCV genome lacking M/E and NS2 regions suggesting a very low or lacking immunogenecity for these regions. Two test kits for detection of anti-HCV antibody were developed using a combination of a synthetic peptide (AR142) containing the epitope of N14 (QRKTKRSTNRR) having a homology to the core of HCV of | fr | sol 8/11AA and a non-fusion recombinant protein Y19 starting from amino acid number (AAN) 1380 to 1507 in the NS3 region showing a AAH to the HCV-1 of 90%, and a combination of a mixture of three synthetic peptides of S29 AAN of 1–30, 38–65 and 47–74 of the core and a non-fused recombinant protein S4 AAN of 1287–1506 having a 93% AAH of the NS3 region. They showed almost the same order of sensitivity and specificity of the second-generation kits when tested with serum from blood donors and patients with non-A, non-B hepatitis. It should also be stressed that in all of the complete responders of a recombinant α-interferon therapy, the antibody levels against AR142 gradually decreased during and after the treatment. In 1992, studies performed for 125 patients with hepatocellular carcinoma in our clinic shows that of these 16 patients might developed from either chronic non-B, non-C liver diseases or chronic liver diseases caused by mutant(s) of HCV as their serum were negative for HBsAg and second-generation of anti-HCV.