Cloning and molecular characterization of the ontogeny of a rat ileal sodium-dependent bile acid transporter (original) (raw)
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Increased expression of ileal apical sodium-dependent bile acid transporter in postpartum rats
American journal of physiology. Gastrointestinal and liver physiology, 2002
The expression and activity of the apical ileal sodium-dependent bile acid transporter (asbt) was examined in the small intestine of control, pregnant, and lactating postpartum rats 2, 12, and 21 days after delivery. Western blot analysis of brush border membrane vesicles (BBMV) prepared from different regions of the small intestine demonstrated that expression of asbt was maximal in the most distal segments for all experimental groups, was not substantially affected in pregnant and 2-day postpartum rats, and was significantly increased in 12- and 21-day postpartum rats. Analysis of mRNA suggested that asbt protein was regulated at the posttranscriptional level in postpartum rats. Increased expression of asbt protein postpartum was maximal (approximately 2-fold) in the proximal region of the ileum, consistent with a 60% increase in taurocholate (TC) transport in BBMV from the proximal ileum in 14- to 21-day postpartum rats relative to control rats. Absorption of TC, determined from ...
Development of active and passive transport of bile acids in rabbit intestine
Mechanisms of Ageing and Development, 1987
Previous studies have indicated that saturable, N absent in the ileum throughout most of the suckling t ileal bile acid uptake which occurs during weaning resl in functional bile acid carriers within the ileal brush 1 undertaken in weanling and adult rabbits to establish active ileal and passive jejunal and colonic uptake of 8 a range of concentrations of cholic (C), taurocholic (T(cholic (CDC), tauroehenodeoxycholic (TCDC), glychoc cholic (I)C) and taurodeoxycholic (TI)C) acid was detl TC, GC, DC and TI)C was greater in adult than in we~)rhea and mat me maturatac results primarily through an inc border membrane. This stud~ establish the effect of maturation ol bile acids. The in vitro upta (TC), glycocholic (GC), chenod¢ rchochenodeoxycholic (GCI)C), de determined. Active ileal uptake ruling animals, whereas uptaJ was similar in both groups. The relative permeability for p~ o the jejunum and colon was similar in young and adult rat area was similar in the two groups, but was greater in the i ,' weanling rabbits due to an increase in villus height, width serosal length. However, the age-associated differences in a not explained simply on the basis of these differences in • the concentration of bile acids in the intestinal lumen, q }it from weanling to adulthood does not influence the rel the jejunum or colon to bile acids, but does increase active td unconjugated cholic acid and deoxycholic acid, but not c[ 1; Aging; Bile acids; Colon; Ileum; Jejunum; Ontogeny; P~
Journal of Clinical Investigation, 1997
Although bile acid transport by bile duct epithelial cells, or cholangiocytes, has been postulated, the details of this process remain unclear. Thus, we performed transport studies with [ 3 H]taurocholate in confluent polarized monolayers of normal rat cholangiocytes (NRC). We observed unidirectional (i.e., apical to basolateral) Na ϩ -dependent transcellular transport of [ 3 H]taurocholate. Kinetic studies in purified vesicles derived from the apical domain of NRC disclosed saturable Na ϩ -dependent uptake of [ 3 H]taurocholate, with apparent K m and V max values of 209 Ϯ 45 M and 1.23 Ϯ 0.14 nmol/mg/10 s, respectively. Reverse transcriptase PCR (RT-PCR) using degenerate primers for both the rat liver Na ϩdependent taurocholate-cotransporting polypeptide and rat ileal apical Na ϩ -dependent bile acid transporter, designated Ntcp and ASBT, respectively, revealed a 206-bp product in NRC whose sequence was identical to the ASBT. Northern blot analysis demonstrated that the size of the ASBT transcript was identical in NRC, freshly isolated cholangiocytes, and terminal ileum. In situ RT-PCR on normal rat liver showed that the message for ASBT was present only in cholangiocytes. Immunoblots using a well-characterized antibody for the ASBT demonstrated a 48-kD protein present only in apical membranes. Indirect immunohistochemistry revealed apical localization of ASBT in cholangiocytes in normal rat liver. The data provide direct evidence that conjugated bile acids are taken up at the apical domain of cholangiocytes via the ASBT, and are consistent with the notion that cholangiocyte physiology may be directly influenced by bile acids. ( J. Clin. Invest. 1997. 100:2714-2721.) Key words: biliary epithelia • taurocholate • transport • liver • plasma membrane vesicles Preliminary portions of this work were presented at the 47th meeting of the American Association for the Study of Liver Diseases, and have been published in abstract form (1996. Hepatology. 94:897 a ).
Proceedings of The National Academy of Sciences, 1991
Liver parenchymal cells continuously extract high amounts of bile acids from portal blood plasma. This uptake process is mediated by a Na+/bile acid cotransport system. A cDNA encoding the rat liver bile acid uptake system has been isolated by expression cloning in Xenopus laevis oocytes. The cloned transporter is strictly sodium-dependent and can be inhibited by various non-bile-acid organic compounds. Sequence analysis of the cDNA revealed an open reading frame of 1086 nucleotides coding for a protein of 362 amino acids (calculated molecular mass 39 kDa) with five possible N-linked glycosylation sites and seven putative transmembrane domains. Translation experiments in vitro and in oocytes indicate that the transporter is indeed glycosylated and that its polypeptide backbone has an apparent molecular mass of 33-35 kDa. Northern blot analysis with the cloned probe revealed crossreactivity with mRNA species from rat kidney and intestine as well as from liver tissues of mouse, guinea pig, rabbit, and man.
Gastroenterology, 1996
The molecular regulation of headenosine triphosphate-dependent and potential-driven patic bile acid transporters during cholestasis is largely bile acid transporters, 4-9 although a recent study suggests unknown. Cloning of complementary DNAs for the sinuthat the latter system may be confined to the endoplasmic soidal sodium-dependent taurocholate cotransporting reticulum. 10 polypeptide (ntcp), the cytosolic bile acid-binding pro-Recently, complementary DNAs (cDNAs) encoding tein 3a-hydroxysteroid dehydrogenase (3a-HSD), and a for a sodium-dependent taurocholate cotransporting putative canalicular bile acid transporter Ca 2/ , Mg 2/polypeptide have been cloned from rat (ntcp) and human ecto-adenosine triphosphatase, now facilitates such liver. 11,12 Expression of the rat liver ntcp in oocytes, transtudies. Methods: Protein mass, steady-state messensiently transfected COS cells, or stably transfected CHO ger RNA (mRNA) levels, and gene transcription were cells resulted in a strictly sodium-dependent saturable assessed in rat livers after common bile duct ligation uptake of taurocholate with a similar Michaelis constant (CBDL) from 1-7 days, and taurocholate uptake was (K m ) of 30-40 mmol/L as previously determined in isodetermined in isolated hepatocytes. Results: After CBDL, Na / -dependent taurocholate uptake (V max ) delated hepatocytes and basolateral plasma membrane prepclined by 70%. The levels of ntcp protein were reduced Abbreviations used in this paper: CBDL, common bile duct ligation;
Cholesterol modulates human intestinal sodium-dependent bile acid transporter
AJP: Gastrointestinal and Liver Physiology, 2005
Bile acids are efficiently absorbed from the intestinal lumen via the ileal apical sodium-dependent bile acid transporter (ASBT). ASBT function is essential for maintenance of cholesterol homeostasis in the body. The molecular mechanisms of the direct effect of cholesterol on human ASBT function and expression are not entirely understood. The present studies were undertaken to establish a suitable in vitro experimental model to study human ASBT function and its regulation by cholesterol. Luminal membrane bile acid transport was evaluated by the measurement of sodium-dependent 3H-labeled taurocholic acid (3H-TC) uptake in human intestinal Caco-2 cell monolayers. The relative abundance of human ASBT (hASBT) mRNA was determined by real-time PCR. Transient transfection and luciferase assay techniques were employed to assess hASBT promoter activity. Caco-2 cell line was found to represent a suitable model to study hASBT function and regulation. 25-Hydroxycholesterol (25-HCH; 2.5 μg/ml fo...