Bile secretion results from coordinately regulated transport processes located at the sinusoid and bile canalicular domains of hepatocytes. Sodium-dependent taurocholate, and sodium-independent organic anion transporters as well as NaK-ATPase are localized to the sinusoidal domain. The long-range goal of this laboratory has been to identify and characterize the regulation of membrane transporters involved in bile formation and pathogenesis of cholestasis. Studies from our laboratory indicate that disorders of sinusoidal membrane transport processes may be the primary site for the pathogenesis of estrogen-induced cholestasis. The sinusoidal transporters are expressed in sexually dimorphic pattern, and decreased transporter expression by estrogens is mediated by the hypothalamic-pituitary-liveraxis. The first hypothesis we propose to test is: The sexually dimorphic expression of sodium-dependent taurocholate transporter is transcriptionally regulated by DNA elements in the 5'flanking region of the gene. Hormone replacement studies will be conducted in hypophysectomized rats and genetically deficient mice (lit/lit). We will also determine the specific hormone effects using cultured hepatocytes and unique cell lines which express the transporter. The promoter will be cloned and used for transient transfection assays and identification of DNase I sensitive sites, DNA-binding proteins, and the development of transgenic mice.
The second aim i s to examine the cell specific insulin regulation of hepatic NaK-ATPase, which plays a pivotal role in regulation of bile salt transport and bile flow. Preliminary data has shown that diabetes selectively increased NaK-ATPase beta-subunit. We propose to examine the hypothesis that beta-subunit, in part, regulates plasma membrane NaK-ATPase activity, and its expression is transcriptionally regulated by insulin. The effect of insulin deficiency on NaK-ATPase will be examined in diabetic rats, and transcriptional regulation of beta1 subunit by insulin will be determined using transient transfection assays in hepatoma cells. Taken together, these studies will provide an understanding of the hormonal control of the molecular events regulating sinusoidal membrane transporters which are involved in generation of bile flow.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK015851-26
Application #
2733961
Study Section
Special Emphasis Panel (ZRG2-NTN (03))
Project Start
1978-08-01
Project End
2000-06-30
Budget Start
1998-07-20
Budget End
1999-06-30
Support Year
26
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Colorado Denver
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
065391526
City
Aurora
State
CO
Country
United States
Zip Code
80045
Qadri, Ishtiaq; Hu, Ling-Jia; Iwahashi, Mieko et al. (2009) Interaction of hepatocyte nuclear factors in transcriptional regulation of tissue specific hormonal expression of human multidrug resistance-associated protein 2 (abcc2). Toxicol Appl Pharmacol 234:281-92
Qadri, Ishtiaq; Iwahashi, Mieko; Kullak-Ublick, Gerd A et al. (2006) Hepatocyte nuclear factor (HNF) 1 and HNF4 mediate hepatic multidrug resistance protein 2 up-regulation during hepatitis C virus gene expression. Mol Pharmacol 70:627-36
Simon, Francis R; Iwahashi, Mieko; Hu, Ling-Jia et al. (2006) Hormonal regulation of hepatic multidrug resistance-associated protein 2 (Abcc2) primarily involves the pattern of growth hormone secretion. Am J Physiol Gastrointest Liver Physiol 290:G595-608
Qadri, Ishtiaq; Iwahashi, Mieko; Capasso, Juan M et al. (2004) Induced oxidative stress and activated expression of manganese superoxide dismutase during hepatitis C virus replication: role of JNK, p38 MAPK and AP-1. Biochem J 378:919-28
Simon, Francis R; Fortune, John; Iwahashi, Mieko et al. (2004) Multihormonal regulation of hepatic sinusoidal Ntcp gene expression. Am J Physiol Gastrointest Liver Physiol 287:G782-94
Reichard, John F; Doorn, Jonathan A; Simon, Franz et al. (2003) Characterization of multidrug resistance-associated protein 2 in the hepatocellular disposition of 4-hydroxynonenal. Arch Biochem Biophys 411:243-50
Qadri, Ishtiaq; Iwahashi, Mieko; Simon, Francis (2002) Hepatitis C virus NS5A protein binds TBP and p53, inhibiting their DNA binding and p53 interactions with TBP and ERCC3. Biochim Biophys Acta 1592:193-204
Simon, Francis R; Fortune, John; Iwahashi, Mieko et al. (2002) Sexual dimorphic expression of ADH in rat liver: importance of the hypothalamic-pituitary-liver axis. Am J Physiol Gastrointest Liver Physiol 283:G646-55
Fiedler, J; Simon, F R; Iwahashi, M et al. (2001) Effect of peroxisome proliferator-activated receptor alpha activation on leukotriene B4 metabolism in isolated rat hepatocytes. J Pharmacol Exp Ther 299:691-7
Shiao, T; Iwahashi, M; Fortune, J et al. (2000) Structural and functional characterization of liver cell-specific activity of the human sodium/taurocholate cotransporter. Genomics 69:203-13

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