Active absorption of vitamin B12 and conjugated bile salts by guinea pig ileum occurs in villous and not crypt cells (original) (raw)
Intrinsic Factor-mediated Absorption of Cobalamin by Guinea Pig Ileal Cells
Journal of Clinical Investigation, 1983
A B S T R A C T To investigate the fate of intrinsic factor and cobalamin during cobalamin absorption, we incubated enterocytes isolated from guinea pig ileum for periods of up to 30 min with 57Co-labeled cyanocobalamin bound either to human intrinsic factor or to rabbit intrinsic factor biosynthetically labeled with [35S]methionine. When the labeled complex was incubated for 30 min with isolated ileal cells under conditions that block cellular metabolism, virtually all cellular radioactivity could be removed by washing the cell surface with EDTA or acid. In contrast, washing removed only half the radioactivity from cells incubated at 370C in 02 When residual cellular radioactivity was extracted and analyzed by gel filtration, 80-94% of both the 35S and 57Co radioactivity eluted in the same fractions as the original complex. The remaining 6-20% eluted as free [57Co]cobalamin or [35S]methionine. To examine events occurring after 30 min, we instilled into tied-off ileal loops of intact guinea pigs radiolabeled intrinsic factor-cobalamin complex and extracted nondissociable radioactivity 2-4.5 h later. The proportion of extracted 57Co eluting as free cobalamin increased to 39-46%, that eluting as intrinsic factor-cobalamin complex declined to 22-45%, and 9-34% now eluted as a macromolecule that reacted with antitranscobalamin II antibody but not antiintrinsic factor antibody. Extracted 35S radioactivity eluted in several peaks in addition to the intrinsic factor peak. These findings suggest that (a) after reversible attachment of intrinsic factor-cobalamin complex to its ileal surface receptor, an energy-dependent process prevents removal of the complex from the cell Parts of this research were reported at the Annual Meetings of the American
American Journal of Physiology-Gastrointestinal and Liver Physiology, 1999
This study was designed to identify the cellular component of the intestinal villus where transcobalamin II (TCII) is synthesized, because this protein provides an essential function in the intestinal absorption of vitamin B12 (cobalamin, Cbl). When a segment of proximal or distal small intestine of the guinea pig is cultured in medium containing [57Co]Cbl, TCII-[57Co]Cbl appears within 15 min. Northern blot analysis of RNA from both proximal and distal small intestine identified the TCII transcript. In situ hybridization of the distal ileum with35S-labeled TCII antisense transcript localized grains predominantly in crypts and in the lower third and central core of the villi. Grains were also evident at the base of the enterocytes in close apposition with the vascular network, whereas few grains appeared in the apical region of the columnar cells. This study provides evidence that TCII is constitutively expressed in the intestinal villi where vascular endothelium is abundant. In the...
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~
Transport of bile acids in a human intestinal epithelial cell line, Caco-2
Biochimica et Biophysica Acta (BBA) - General Subjects, 1990
The transport of tanrocholic acid (TA) across Caco-2 cell monolayers was dependent on time in culture and reached a plateau after 28 days, at which time the apical (AP)-to-basolateral (131,) transport was 10-times greater than BL-to-AP transport. The amounts of TA inside the cells following application of 10 nM [I4C]TA to tile AP or BL side of the monolayers (30 min) were approximately equal (54.4 + 2.7 and 64.6 ± 2.8 fmol/mg protein, respectively). AP-to-BL transport of TA was saturable and temperature-dependent. Vm~ , and K m for transport were 13.7 pmol / mg protein per min and 49.7 pM, respectively. The transport of TA had an activation energy of 13.2 kcal-mo1-1, required Na + and glucose. AP-to-BL transport of |14C]TA was inhibited by the co-administration (on the AP side) of either unlabeled TA or deoxycholate, but it was not reduced by the presence of unlabeled TA on the BL side.
Bile acid active and passive ileal transport in the rabbit: effect of luminal stirring
European Journal of Clinical Investigation, 1992
The intestinal absorption of bile acids (BA) with different chemical structure has been evaluated in the rabbit, after intestinal infusion of different concentrations (0.25-30 mM) of BA, by mesenteric blood sampling. Cholic (CA), chenodeoxycholic (CDCA), ursodeoxycholic (UDCA) acid, free and taurine (T-) conjugated, together with glycocholic (GCA) acid and deoxycholic acid (DCA) were studied. The apparent uptake parameters were calculated. All conjugated BA showed active transport (T max, nmol min-' cm-' int.), with Tmax values in the following order: TCA > TUDCA > TCDCA; unconjugated BA showed passive uptake, with values in the following order: DCA > CDCA > UDCA > CA. GCA and CA showed both passive uptake and active transport. For all BA studied the % uptake in the ileal segment considered was less than lo%, BA uptake being thus limited by transport and/or diffusion kinetics, rather than by flow velocity. The liquid resistance to BA radial diffusion inside the lumen was evaluated, and the infusate-to-blood uptake parameters corrected for it, in order to get the uptake parameters from the epithelium-to-liquid interface to mesenteric blood: the apparent Km decreased, passive uptake coefficient increased, while Tmax was unchanged. The passive component of the uptake, corrected for the luminal resistance, correlated with the BA hydrophobicity (r = 0.963; P < 0.01). These studies show that: (a) the active transport for BA in the rabbit ileum is mediated by a saturable, highefficiency, low-affinity carrier; (b) that passive transport is highly efficient for unconjugated BA, mainly for the most lipophilic ones; (c) that both systems are important in the intestinal absorption of BA.
Expression cloning and characterization of the hamster ileal sodium-dependent bile acid transporter
Journal of Biological Chemistry, 1994
Active uptake of bile acids from the lumen of the small intestine is mediated by an ileal Na+-dependent bile acid transport system. To identify components of this transport system, an expression cloning strategy was employed to isolate a hamster ileal cDNA that exhibits bile acid transport activity. By Northern blot analysis, mRNA for the cloned transporter was readily detected in ileum and kidney but was absent from liver and proximal small intestine. The transporter cDNA encoded a 348- amino acid protein with seven potential transmembrane domains and three possible N-linked glycosylation sites. The amino acid sequence was 35% identical and 63% similar to the rat liver Na+/bile acid cotransporter. After transfection into COS cells, the hamster cDNA transported taurocholate in a strict Na'dependent fashion with an apparent K , of 33 p ~. This taurocholate transport was inhibited by various bile acids but not by taurine or other organic anions. The Na+ dependence, saturability, and bile acid specificity of transport as well as the tissue specificity of mRNA expression strongly argue that the transporter cDNA characterized in this study is the Na+/bile acid cotransporter described previously in ileum. Bile acids are acidic sterols synthesized from cholesterol in the liver (Russell and . Following synthesis, the bile acids are secreted into bile and enter the lumen of the small intestine, where they facilitate absorption of fat-soluble vitamins and cholesterol. Bile acids are then absorbed from the small intestine, returned to the liver via the portal circulation, and resecreted into bile . In the small intestine, bile acids are absorbed by both passive and active mechanisms . The active absorption of bile acids, which was first described by Lack and Weiner (19611, has been shown in man and experimental animals to be restricted to the