In vivo effect of calcium (Ca) on brush border intestinal alkaline phosphatase (BBIAP) in the rat

Bone ◽  
2006 ◽  
Vol 38 (5) ◽  
pp. S16-S17 ◽  
Author(s):  
M.L. Brance ◽  
R.M. Brun ◽  
A. Rigalli ◽  
L. De Candia ◽  
R.C. Puche
Keyword(s):  
Alkaline Phosphatase ◽  
Brush Border ◽  
Intestinal Alkaline Phosphatase

scholarly journals Independent biosynthesis of soluble and membrane-bound alkaline phosphatases in the suckling rat ileum

1981 ◽  
Vol 200 (3) ◽  
pp. 645-654 ◽  
Author(s):  
Graeme P. Young ◽  
Steven T. Yedlin ◽  
David H. Alpers
Keyword(s):  
Alkaline Phosphatase ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Double Diffusion ◽  
Soluble Form ◽  
Soluble Enzyme ◽  
Intestinal Alkaline Phosphatase ◽  
Absolute Increase ◽  
Membrane Bound

Enzymically active intestinal alkaline phosphatase exists in both soluble and membrane-bound forms in the suckling rat. Antiserum prepared against purified soluble alkaline phosphatase (anti-AlP) was shown to be monospecific when assessed by Ouchterlony double-diffusion analysis and immunoelectrophoresis. The two forms of alkaline phosphatase were antigenically identical and possessed similar affinities for anti-AlP. To study the biosynthesis of the two forms, 14-day-old rats were injected intraperitoneally with [3H]leucine. The labelling kinetics of alkaline phosphatase, extracted from supernatant and brush-border membrane fractions with anti-AlP, was followed over 20h. Incorporation of [3H]leucine into membrane-bound alkaline phosphatase was rapid, reaching a plateau at 6h. The soluble enzyme showed slower incorporation of label and maximal radioactivity was not reached until 12h after labelling, a lag of 6h behind the membrane-bound enzyme. Soluble alkaline phosphatase could not have been a precursor of the membrane form, as there was no early peak of radioactivity in the soluble form. To determine if the soluble enzyme was irreversibly derived from the membrane enzyme, a newly developed technique of labelling brush-border membrane proteins in vivo by intraluminal injection of diazotized [125I]iodosulphanilic acid was used. The appearance of 125I in soluble and membrane alkaline phosphatase was then monitored over a 7h period, encompassing the lag between maximal leucine labelling of the two forms. The results failed to show either a proportional transfer of radioactivity from membrane to soluble alkaline phosphatase or an absolute increase in radioactivity of the soluble form during degradation of brush-border alkaline phosphatase. Therefore there does not appear to be a serial precursor/product relationship between the soluble and membrane-bound forms of suckling-rat intestinal alkaline phosphatase.

Duodenal brush border intestinal alkaline phosphatase activity affects bicarbonate secretion in rats

2007 ◽  
Vol 293 (6) ◽  
pp. G1223-G1233 ◽  
Author(s):  
Yasutada Akiba ◽  
Misa Mizumori ◽  
Paul H. Guth ◽  
Eli Engel ◽  
Jonathan D. Kaunitz
Keyword(s):  
Alkaline Phosphatase ◽  
Brush Border ◽  
Intestinal Alkaline Phosphatase ◽  
Glycerol Phosphate ◽  
Secretory Rate ◽  
Rat Duodenum ◽  
Ph Stat ◽  
Cystic Fibrosis Transmembrane

We hypothesized that duodenal HCO3− secretion alkalinizes the microclimate surrounding intestinal alkaline phosphatase (IAP), increasing its activity. We measured AP activity in rat duodenum in situ in frozen sections with the fluorogenic substrate ELF-97 phosphate and measured duodenal HCO3− secretion with a pH-stat in perfused duodenal loops. We examined the effects of the IAP inhibitors l-cysteine or l-phenylalanine (0.1–10 mM) or the tissue nonspecific AP inhibitor levamisole (0.1–10 mM) on AP activity in vitro and on acid-induced duodenal HCO3− secretion in vivo. AP activity was the highest in the duodenal brush border, decreasing longitudinally to the large intestine with no activity in stomach. Villous surface AP activity measured in vivo was enhanced by PGE2 intravenously and inhibited by luminal l-cysteine. Furthermore, incubation with a pH 2.2 solution reduced AP activity in vivo, whereas pretreatment with the cystic fibrosis transmembrane regulator (CFTR) inhibitor CFTRinh-172 abolished AP activity at pH 2.2. l-Cysteine and l-phenylalanine enhanced acid-augmented duodenal HCO3− secretion. The nonselective P2 receptor antagonist suramin (1 mM) reduced acid-induced HCO3− secretion. Moreover, l-cysteine or the competitive AP inhibitor glycerol phosphate (10 mM) increased HCO3− secretion, inhibited by suramin. In conclusion, enhancement of the duodenal HCO3− secretory rate increased AP activity, whereas inhibition of AP activity increased the HCO3− secretory rate. These data support our hypothesis that HCO3− secretion increases AP activity by increasing local pH at its catalytic site and that AP hydrolyzes endogenous luminal phosphates, presumably ATP, which increases HCO3− secretion via activation of P2 receptors.

The two isozymes of rat intestinal alkaline phosphatase are products of two distinct genes

2000 ◽  
Vol 3 (1) ◽  
pp. 1-8 ◽  
Author(s):  
Q. XIE ◽  
D. H. ALPERS
Keyword(s):  
Alkaline Phosphatase ◽  
Transcriptional Start Site ◽  
In Vivo Studies ◽  
Peroxisome Proliferator ◽  
Start Site ◽  
Intestinal Alkaline Phosphatase ◽  
Promoter Regions ◽  
Flanking Regions

Xie, Q., and D. H. Alpers. The two isozymes of rat intestinal alkaline phosphatase are products of two distinct genes. Physiol Genomics 3: 1–8, 2000.—Rat intestinal alkaline phosphatases (IAP-I and -II) differ in primary structure, substrate specificity, tissue localization, and response to fat feeding. This study identifies two distinct genes (∼5–6 kb) corresponding to each isozyme and containing 11 exons of nearly identical size. The exon-intron junctions are identical with those found in IAP genes from other species. The 1.7 and 1.2 bp of 5′ flanking regions isolated from each gene, respectively, contain Sp1 and gut-enriched Kruppel-like factor (GKLF) binding sites, but otherwise show little identity. There is a potential CAAT-box 14 bp 5′ to the transcriptional start site, 36 bp upstream from IAP-I, and a TATA-box 31 bp 5′ to the transcriptional start site, 55 bp upstream from IAP-II. Transfection of these promoter regions (linked to luciferase as a reporter gene) into a kidney cell line, COS-7, produced the differential response to oleic acid expected from in vivo studies, i.e., threefold increase using the 5′ flanking region of IAP-II, but not IAP-I. This response was not reproduced by 5,8,11,14-eicosatetraynoic acid (ETYA) or clofibrate, suggesting that peroxisome proliferator response elements are not involved. Isolation of the IAP-II gene will allow determination of the sequences responsible for dietary fat response in the enterocyte.

Anion-stimulated ATPase activity of brush border from rat small intestine.

1979 ◽  
Vol 236 (1) ◽  
pp. E70 ◽  
Author(s):  
M H Humphreys ◽  
L Y Chou
Keyword(s):  
Alkaline Phosphatase ◽  
Atpase Activity ◽  
Brush Border ◽  
Atp Hydrolysis ◽  
Intestinal Transport ◽  
Mitochondrial Enzymes ◽  
Intestinal Alkaline Phosphatase ◽  
Ph Optimum ◽  
Low Concentrations ◽  
Small Intestinal

Differential centrifugation of rat small intestinal homogenates produced a crude brush border (BB) fraction that was enriched 15-fold for the marker enzymes, alkaline phosphatase and sucrase; contamination with mitochondrial enzymes, monoamine oxidase and succinate dehydrogenase, was minimal. ATP hydrolysis by this BB fraction was stimulated by addition of several anions to the incubation medium: HCO3 and Cl were equally effective in this regard, with NO3, NO2, SO4, and acetate being less stimulatory. SCN and CNO inhibited ATPase activity, whereas the divalent anion SO3 was stimulatory at low concentrations (less than 25 mM) but inhibitory at 100 mM. Maximum anion stimulation was observed at a Mg concentration of 0.5 mM, and pH optimum was 8.5. Kinetic analysis showed that HCO3 increased the Vmax without altering the Km for ATP; the Ka for this effect of HCO3 was 35 mM. This enzyme activity was completely inhibited by 20 mM L-phenylalanine, 10 mM L-cysteine, and 3 mM EDTA, compounds that also inhibited intestinal alkaline phosphatase. These results demonstrate the presence of anion-stimulated ATPase activity in rat small intestinal brush border and suggest that this activity may be related to intestinal alkaline phosphatase. The role of this enzyme in intestinal transport is not known, but could relate to the regulation of intestinal absorption and secretion.

The similarity between alkaline phosphatase (EC 3.1.3.1) and phytase (EC 3.1.3.8) activities in rat intestine and their importance in phytate-induced zinc deficiency

1978 ◽  
Vol 39 (2) ◽  
pp. 307-316 ◽  
Author(s):  
N. T. Davies ◽  
A. A. Flett
Keyword(s):  
Alkaline Phosphatase ◽  
Brush Border ◽  
Regional Differences ◽  
Maximal Activity ◽  
Duodenal Mucosa ◽  
Zn Deficiency ◽  
Intestinal Segments ◽  
Phytate Hydrolysis ◽  
Feeding Diets

1. The activities of alkaline phosphatase (EC 3.1.3.1) and phytase (EC3.1.3.8) were similarly distributed in the small intestine of rats. Regional differences in activity were reflected by similar differences in the capacity of ligated intestinal segments to hydrolyse phytate in vivo. Activities were greatest in the duodenum and lowest in the terminal ileum.2. Specific activities of both enzymes were tenfold greater in the brush border fraction of duodenal mucosa compared with entire mucosal homogenates.3. Brush-border alkaline phosphatase and phytase activities required both magnesium and zinc ions for maximal activity.4. Zn deficiency induced by feeding a diet low in Zn (0.5 mg Zn/kg) caused similar reductions in activity of both enzymes.5. Zn deficiency induced by feeding diets marginally adequate in Zn (12 mg/kg) and phytate (10g/kg) caused reductions in alkaline phosphatase, phytase activities and phytate hydrolysis in vivo.6. It is suggested that phytase activity is a manifestation of alkaline phosphatase and the significance of this in relation to phytate-induced Zn deficiency is discussed.

Pattern of rat intestinal brush-border enzyme gene expression changes with epithelial growth state

1995 ◽  
Vol 269 (2) ◽  
pp. C385-C391 ◽  
Author(s):  
R. A. Hodin ◽  
S. M. Chamberlain ◽  
S. Meng
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Brush Border ◽  
Longitudinal Axis ◽  
Mrna Levels ◽  
Intestinal Alkaline Phosphatase ◽  
Enzyme Gene ◽  
Growth State ◽  
Brush Border Enzyme ◽  
Epithelial Growth

Enterocyte growth and differentiation occur simultaneously within the epithelium, but little is known regarding any relationship between these two processes. Four rat models of small intestinal epithelial hypo- and hyperplasia (neonatal ontogeny, fasting/refeeding, hypo-/hyperthyroidism, and bombesin treatment) were used to study the regulation of enterocyte gene expression in relation to epithelial growth state. Mucosal scrapings, as well as crypt and villus cell populations, were subjected to Northern blot analyses using radiolabeled cDNA probes corresponding to lactase, intestinal alkaline phosphatase, villin, ornithine decarboxylase (ODC), and the actin control. In all four models, the hypoplastic (atrophic) condition is characterized by high levels of lactase and low levels of the 3.0-kb intestinal alkaline phosphatase mRNA, whereas under hyperplastic conditions this pattern is reversed. The changes in intestinal alkaline phosphatase and lactase are qualitatively similar along the longitudinal axis of the intestine and are proportional to the degree of hyperplasia, as verified by ODC mRNA levels. Furthermore, the crypt-villus axis of differentiation is maintained regardless of epithelial growth state. In conclusion, the pattern of brush-border enzyme gene expression changes as a function of epithelial growth state, indicating a previously unrecognized degree of plasticity to the state of enterocyte differentiation.

scholarly journals The enterocyte microvillus is a vesicle-generating organelle

2009 ◽  
Vol 185 (7) ◽  
pp. 1285-1298 ◽  
Author(s):  
Russell E. McConnell ◽  
James N. Higginbotham ◽  
David A. Shifrin ◽  
David L. Tabb ◽  
Robert J. Coffey ◽  
...  
Keyword(s):  
Brush Border ◽  
Membrane Surface ◽  
Intestinal Lumen ◽  
Intestinal Alkaline Phosphatase ◽  
Brush Border Enzymes ◽  
Membrane Surface Area ◽  
Catalytically Active ◽  
The Right

For decades, enterocyte brush border microvilli have been viewed as passive cytoskeletal scaffolds that serve to increase apical membrane surface area. However, recent studies revealed that in the in vitro context of isolated brush borders, myosin-1a (myo1a) powers the sliding of microvillar membrane along core actin bundles. This activity also leads to the shedding of small vesicles from microvillar tips, suggesting that microvilli may function as vesicle-generating organelles in vivo. In this study, we present data in support of this hypothesis, showing that enterocyte microvilli release unilamellar vesicles into the intestinal lumen; these vesicles retain the right side out orientation of microvillar membrane, contain catalytically active brush border enzymes, and are specifically enriched in intestinal alkaline phosphatase. Moreover, myo1a knockout mice demonstrate striking perturbations in vesicle production, clearly implicating this motor in the in vivo regulation of this novel activity. In combination, these data show that microvilli function as vesicle-generating organelles, which enable enterocytes to deploy catalytic activities into the intestinal lumen.

Intestinal alkaline phosphatase: selective endocytosis from the enterocyte brush border during fat absorption

2007 ◽  
Vol 293 (6) ◽  
pp. G1325-G1332 ◽  
Author(s):  
Gert H. Hansen ◽  
Lise-Lotte Niels-Christiansen ◽  
Lissi Immerdal ◽  
Birthe T. Nystrøm ◽  
E. Michael Danielsen
Keyword(s):  
Alkaline Phosphatase ◽  
Brush Border ◽  
Immunofluorescence Microscopy ◽  
Lipid Analysis ◽  
Marked Change ◽  
Fat Absorption ◽  
Intestinal Alkaline Phosphatase ◽  
Coated Pits ◽  
Brush Border Enzymes ◽  
Detergent Resistant Membranes

Absorption of dietary fat in the small intestine is accompanied by a rise of intestinal alkaline phosphatase (IAP) in the serum and of secretion of IAP-containing surfactant-like particles from the enterocytes. In the present work, fat absorption was studied in organ cultured mouse intestinal explants. By immunofluorescence microscopy, fat absorption caused a translocation of IAP from the enterocyte brush border to the interior of the cell, whereas other brush-border enzymes were unaffected. By electron microscopy, the translocation occurred by a rapid (5 min) induction of endocytosis via clathrin-coated pits. By 60 min, IAP was seen in subapical endosomes and along membranes surrounding fat droplets. IAP is a well-known lipid raft-associated protein, and fat absorption was accompanied by a marked change in the density and morphology of the detergent-resistant membranes harboring IAP. A lipid analysis revealed that fat absorption caused a marked increase in the microvillar membrane contents of free fatty acids. In conclusion, fat absorption rapidly induces a transient clathrin-dependent endocytosis via coated pits from the enterocyte brush border. The process selectively internalizes IAP and may contribute to the appearance of the enzyme in serum and surfactant-like particles.

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