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H M Said

Publications and source records attributed to H M Said.

At least 73 records · Page 4Linked to original sources
PubMed1991

Carrier-mediated mechanism for biotin transport in rabbit intestine: studies with brush-border membrane vesicles.

Simple diffusion has been reported as the mechanism of biotin transport in rabbit intestine. In this study, we reevaluated this concept by examining biotin transport in rabbit intestine using optimal experimental conditions and a well-established brush-border membrane vesicles (BBMV) technique. Uptake of biotin by rabbit intestinal BBMV was found by an osmolarity study to be mostly the result of transport of the vitamin into an osmotically sensitive intravesicular space with little binding to membrane surfaces. Biotin transport in rabbit intestinal BBMV was 1) Na+ gradient dependent (out greater than in) with a clear "overshoot" phenomenon, indicating the accumulation of the substrate against a concentration gradient; 2) initial rate of biotin transport by the Na+ gradient-dependent component was saturable as a function of substrate concentration with apparent Km and maximum velocity (Vmax) values of 6.7 microM and 10.7 pmol.mg protein-1 x 10 s-1, respectively; 3) inhibited by high concentrations of unlabeled biotin and its related compounds desthiobiotin and thioctic acid in the presence, but not absence, of a Na+ gradient; and 4) not affected by inducing a relatively positive or negative intravesicular space with the use of valinomycin-induced K+ diffusion potential. These findings indicate that the biotin transport mechanism in rabbit intestine is carrier mediated in nature. Furthermore, this mechanism is Na+ gradient dependent, capable of accumulating the substrate against a concentration gradient and transport the vitamin via an electroneutral process.

Animals↗
PubMed1990

Effect of chemically induced diabetes mellitus on glutamine transport in rat intestine.

The effect of chemically induced diabetes mellitus on the intestinal transport of glutamine was examined using a brush-border membrane vesicle technique. Diabetes was induced by a single intraperitoneal injection of streptozotocin (100 mg/kg body weight). Control and diabetic rats were studied 5 days following the induction of diabetes. Na(+)-dependent and Na(+)-independent glutamine (0.5 mM) transport was found to be significantly higher in the diabetic rats than in the control rats. This increase was found to be caused by a significant increase in the Vmax of the Na(+)-dependent and the Na(+)-independent glutamine transport processes in the diabetic rats (Vmax of 3742 +/- 487 and 2055 +/- 279 pmol/mg protein per 7 s, respectively) compared with that of the control rats (2183 +/- 75 and 1271 +/- 83 pmol/mg protein per 7 s, respectively). The apparent Km values of glutamine transport systems, on the other hand, were similar in the two rat groups. Insulin treatment of the diabetic rats significantly reduced the Vmax of glutamine transport by both the Na(+)-dependent and the Na(+)-independent processes to a level similar to that of the control rats (Vmax in the insulin-treated diabetic rats of 2036 +/- 123 and 1247 +/- 105 pmol/mg protein per 7 s, respectively). This study demonstrates that chemically induced diabetes mellitus is associated with an increase in intestinal glutamine transport. This increase is the result of the diabetic condition itself and appears to be mediated through an increase in the number of the transport carriers of glutamine.

Animals↗
PubMed1990

Effect of aging on intestinal biotin transport in the rat.

The influence of aging of biotin intestinal transport was examined in Fisher 344 rats (3- 12- and 24-month old) using a brush border membrane vesicle (BBMV) technique. In all age groups examined, transport of biotin as a function of concentration was saturable in the presence of a Na+ gradient (out greater than in), but was lower and linear in the presence of a choline gradient (out greater than in.). The Vmax of the biotin transport process was found to be significantly (p less than 0.01) higher in 24-month-old rats compared to 3- and 12-month old rats. On the other hand, the apparent Km of the biotin transport system was similar in all age groups. We also measured plasma levels of biotin in the different age groups. The mean plasma levels of biotin in 24-month-old rats was found to be significantly higher than that of 3-month (p less than 0.005) and 12-month (p less than 0.025) old rats. These results demonstrate that aging is associated with an increase in biotin intestinal transport. This increase appears to be due to changes in the activity (and/or number) but not the affinity of the biotin transport system. Furthermore, an increase in plasma biotin levels was observed with aging, which might be a consequence to the increase in the vitamin's intestinal transport.

Aging↗
PubMed1990

Chronic ethanol feeding and acute ethanol exposure in vitro: effect on intestinal transport of biotin.

This study examined the effects of chronic ethanol feeding and acute ethanol exposure in vitro on biotin transport in rat intestine. Transport studies were performed with intestinal everted sacs. Ethanol was fed to rats for 6-7 wk. Compared with pair-fed controls, ethanol feeding significantly decreased plasma biotin concentrations and transport at physiological concentrations (0.01, 0.1, and 0.3 mumol/L) but not at pharmacological concentration (100 mumol/L). When added to the incubation medium of everted jejunal sacs from dry-food-fed rats, ethanol (2% vol:vol) significantly inhibited the transport of biotin at a physiological concentration (0.1 mumol/L) but not at a pharmacological concentration (100 mumol/L). The inhibitory effect of ethanol on the transport of 0.1 mumol biotin/L increased with increasing concentration of ethanol in the incubation medium [0.5% to 5% (vol:vol)]. Acetaldehyde, the major ethanol metabolite, also significantly inhibited biotin transport at 0.2% (vol:vol). These data demonstrate that chronic ethanol feeding and acute ethanol exposure in vitro inhibit the intestinal transport of biotin by the carrier-mediated process. Chronic ethanol feeding is also associated with a significant decrease in plasma biotin concentrations. The ethanol-induced inhibition in intestinal transport of biotin may be a contributing factor in reducing plasma biotin concentrations.

Alcoholism↗
PubMed1990

Uptake of biotin by native Xenopus laevis oocytes.

The present study examined biotin uptake by Xenopus laevis oocytes in vitro. Uptake of low (0.03 microM) and high (10 microM) concentrations of biotin was linear with time for up to 4 h of incubation and occurred with little initial binding to oocytes. Uptake of biotin was dependent on extracellular Na+ concentration [Na+]o and was severely inhibited when Na+ was replaced by other monovalent cations [choline, tetraethylammonia, Li+, and tris(hydroxymethyl)aminomethane]. The initial rate of biotin uptake was saturable as a function of concentration with an apparent Michaelis constant of 3.9 +/- 0.5 microM and maximum velocity of 1,559 +/- 70 fmol.oocyte-1.h-1. Addition to the incubation medium of biotin structural analogues desthiobiotin and thioctic acid caused significant and concentration-dependent inhibition in the uptake of [3H]biotin. This inhibition was found to be competitive in nature with inhibition constant values of 9 and 17.5 microM. In contrast, neither the structural analogue biocytin nor biotin methyl ester (compounds in which the carboxyl group of the valeric acid moiety is blocked) showed any effect on the uptake of [3H]biotin. Biotin uptake was significantly blocked by the metabolic inhibitors dinitrophenol, cyanide, and azide and by incubation at 4 degrees C. Also, the sulfhydryl group blocker p-(chloromercuri)phenylsulfonate caused significant inhibition in biotin uptake. These results demonstrate that Xenopus oocytes possess an uptake system for biotin in its cell membrane that is Na+, energy, and temperature dependent. These characteristics of biotin uptake are similar to those reported in mammalian cells. It is suggested that Xenopus oocytes might be a useful in vitro model system to study the details of the mechanisms and regulation of biotin movement across biological membranes.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Transport of biotin in basolateral membrane vesicles of rat liver.

We examined biotin transport across the basolateral membrane (BLM) of rat liver using BLM vesicles (BLMV) technique. The purity and suitability for transport studies of liver BLMV were demonstrated by morphological (electron microscopy), enzymatic, and functional criteria. Orientation of liver BLMV was determined by freeze-fracture electron microscopy and by [3H]ouabain binding methodology and was found to be 65.3-69.7% in the right-side-out orientation. Uptake of biotin by liver BLMV was found by osmolarity and temperature studies to be mostly the result of transport of the substrate into an active intravesicular space with little binding to membrane surfaces. Transport of biotin was found to be Na+ gradient dependent with a distinct "over-shoot" phenomenon. Initial rate of transport of biotin as a function of concentration was found to include a saturable component in the presence of a Na+ gradient (out greater than in) but was linear and lower in the presence of a K+ gradient (no Na+). Kinetic parameters of the saturable Na+ gradient-dependent transport process were 0.39 microM and 1,807 fmol.mg protein-1.20 s-1 for the apparent Km and Vmax, respectively. In the presence, but not the absence, of a Na+ gradient (out greater than in), the addition of structural analogues to the incubation medium caused significant inhibition in the transport of 0.079 microM [3H]biotin. Induction, with the use of valinomycin and an inwardly directed K+ gradient, of a relatively positive intravesicular space caused significant inhibition in the initial rate of biotin transport.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Transport of biotin in the ileum of suckling rats: characteristics and ontogeny.

Unlike adult rats, which transport biotin preferentially in the jejunum of the small intestine, suckling rats transport biotin preferentially in the ileum. The characteristics and kinetics of the transport process of biotin in the ileum of developing suckling rats and the subsequent maturation of the process in weanling and adult rats are not known and, therefore, are examined in this study. Transport studies were performed in suckling (16-d-old), weanling (25- to 27-d-old), and adult (90-d-old) rats using the everted sac technique. Transport of biotin in the ileum of suckling rats was significantly higher (p less than 0.01) than that in weanling rats, and transport in weanling rats was in turn significantly higher (p less than 0.01) than that in adult rats. In all age groups examined, transport of biotin was found to be Na(+)-dependent and saturable as a function of concentration within the physiologic range (less than 10 microM), but linear at high concentrations. However, the Vmax of the carrier-mediated transport system of biotin showed progressive decrease with maturation (3290 +/- 363, 829 +/- 47, and 463 +/- 19 pmol/g tissue wet wt/15 min in suckling, weanling, and adult rats, respectively), whereas no significant changes were observed in the apparent Km (2.8 +/- 0.6, 2.6 +/- 0.4, and 3.4 +/- 0.4 microM in suckling, weanling, and adult rats, respectively). These findings suggest that maturation is associated with a decrease in the number (and/or activity) but not the affinity of the ileal biotin transport carriers. Transport of biotin by the diffusion process was also found to decrease with maturation (diffusion rates of 144, 86.5, and 40 pmol/g tissue wt/15 min were found for suckling, weanling, and adult rats, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Transport of glutamine in rat intestinal brush-border membrane vesicles.

Transport of glutamine across the brush-border membrane of the rat intestine was examined using brush-border membrane vesicle (BBMV) technique. Osmolarity and temperature studies indicated that the uptake of glutamine by BBMV is mostly the result of transport of the substrate into the intravesicular space. Transport of glutamine was Na+-gradient dependent (out greater than in) with a distinct 'overshoot' phenomenon. Initial rate of transport of glutamine as a function of concentration was saturable both in the presence and absence of a Na+ gradient (out greater than in). Apparent Km of 3.50 and 3.34 mM and Vmax of 707 and 282 pmol/mg protein per 7 s, were calculated for the Na+-dependent and the Na+-independent transport processes of glutamine. The transport of [3H]glutamine by the Na+-dependent and the Na+-independent processes was severely inhibited by the addition to the incubation medium of other amino acids and unlabelled glutamine. Inducing a relatively negative intravesicular compartment with the use of valinomycin and an outwardly directed K+ gradient stimulated glutamine transport. This indicates that transport of the substrate by the Na+-dependent process is electrogenic in nature. Transport of glutamine by the Na+-independent process, however, appeared to be electroneutral in nature. These results demonstrate the existence of two carrier-mediated transport processes for glutamine in the rat intestinal BBMV, one is Na+-dependent and the other is Na+-independent. Furthermore, the results suggest that glutamine transport by the Na+-dependent process probably occurs by a glutamine/Na+ cotransport mechanism.

Animals↗
PubMed1989

Metabolism of 3H- and 14C-labeled glutamate, proline, and alanine in normal and adrenalectomized rats using different sites of tracer administration and sampling.

Alanine, glutamate and proline labeled with 14C and 3H were infused into fasted normal and adrenalectomized rats. Alanine was administered by the A-V mode (arterial administration-venous sampling), and glutamate and proline by both the A-V and V-A (venous administration-arterial sampling) modes. The kinetics of 14C alanine and 14C glutamate differed markedly from those of the tritium-labeled compounds, but there was little difference in the kinetics of 3H and 14C proline. The replacement rate calculated from the A-V mode for glutamate was about half that obtained in the V-A mode, but there was little difference with proline. The masses of the amino acids (total content of amino acids in the body) were calculated from the washout curves of the tritium-labeled compounds after the infusion of tracer was terminated. The masses for the normal rats were 407 mumol/kg for alanine, 578 mumol/kg for glutamate and 296 mumol/kg for proline. The so-called distribution spaces calculated conventionally from total masses and the amino acid concentrations in plasma are much greater than the volume of the body, reflecting the fact that amino acid concentrations in tissues greatly exceed those in plasma. Adrenalectomy markedly affected the kinetics of the three amino acids, and their replacement rates were greatly reduced. The proline and glutamate masses were reduced by at least one half, while that of alanine was unchanged. Adrenalectomy markedly reduced the conversion of proline to glutamate. The hydrocortisone regimen used in this study restored the metabolism of alanine and glutamate to normal, but had no effect on that of proline.

Adrenalectomy↗
PubMed1989

Biotin transport in the human intestine: inhibition by anticonvulsant drugs.

The effect of the anticonvulsant drugs carbamazepine and primidone on the transport of biotin in the human intestine was examined with purified brush border membrane vesicles (BBMVs) and basolateral membrane vesicles (BLMVs). Both agents inhibited biotin transport in BBMV in a concentration-dependent manner. The inhibition by both carbamazepine and primidone was competitive (inhibition constant [Ki] of 4.70 and 2.25 mmol/L, respectively) and appeared to be specific because the transport of D-glucose was not affected by different concentrations of these pharmacologic agents. The transport of biotin in BLMV was not affected by carbamazepine or primidone. These results demonstrate that carbamazepine and primidone are competitive inhibitors of biotin transport in the human intestine and that the inhibitory effect is directed toward the substrate transport system at the brush border membrane of the enterocyte. These findings may relate to possible impairment of biotin status in patients on long-term therapy with anticonvulsant agents.

Anticonvulsants↗
PubMed1989

Intestinal uptake of retinol: enhancement by bovine milk beta-lactoglobulin.

The effect of bovine milk beta-lactoglobulin (BLG) on intestinal uptake of retinol was examined in suckling rats with the everted sac technique. Uptake of 0.06 mumol retinol/L bound to BLG (BLG-retinol) was significantly (p less than 0.01) higher than that of 0.06 mumol free retinol/L both in the jejunum and the ileum. The enhancing effect of BLG on retinol uptake was specific because equimolar concentrations of bovine serum albumin and lactoferrin had no effect on retinol uptake. However, serum retinol-binding protein (RBP), which shares structural and conformational similarities with BLG, also enhanced retinol uptake. BLG, BLG-retinol, and RBP-retinol all inhibited the uptake of retinol from BLG-[3H]retinol in a concentration-dependent manner. Uptake of retinol from BLG-retinol was saturable (apparent Km = 5.6 mumol/L, Vmax = 22.7 nmol.g-1.5 min-1), not affected by metabolic inhibitors, and partially temperature dependent (Q10 = 2.77). BLG also significantly (p less than 0.01) enhanced retinol uptake in the intestine of adult rats. These results demonstrate that BLG specifically enhances intestinal uptake of retinol and suggest the possibility of a receptor for BLG-like proteins at the brush border membrane of the enterocyte.

Animals↗
PubMed1989

Aging: effect on hepatic metabolism and transport of folate in the rat.

Effects of aging on hepatic folate metabolism and transport were assessed in male Fisher 344 rats. Total serum and hepatic folate levels were measured. Hepatic folates were measured by high-performance liquid chromatography and by Lactobacillus casei assay. Transport of 5-methyltetrahydrofolate (5-CH3-H4PteGlu) was measured in isolated hepatocytes. Serum folate declined with aging; however, neither the total folate level nor the distribution of hepatic folate coenzymes was affected by the aging process. The level of liver folate monoglutamates was not significantly different in any group. The initial rate of uptake of 5-CH3-H4PteGlu was significantly decreased in hepatocytes from the 24-mo-old rats, as was the ability to concentrate this folate from the medium. Aged rats maintain apparently normal levels of hepatic folates despite decreased serum levels and decreased ability to take up folates, suggesting that membrane transport of folates may not be a limiting factor in hepatic folate assimilation.

Aging↗
PubMed1989

Transport characteristics of glutamine in human intestinal brush-border membrane vesicles.

Transport of glutamine (Gln) across the brush-border membrane of the human intestine was examined using the brush-border membrane vesicle (BBMV) technique. Osmolarity and temperature studies indicate that the uptake of Gln by BBMV is mostly the result of transport of the substrate into the intravesicular space. Transport of Gln was Na+ gradient dependent (out greater than in) with a distinct "overshoot" phenomenon. Initial rate of transport of Gln as a function of concentration was saturable both in the presence and absence of a Na+ gradient (out greater than in). Apparent Km of 1.86 and 1.36 mM and Vmax of 1,906 and 637 pmol.mg protein-1 . 7 s-1 were calculated for the Na+-dependent and the Na+-independent transport processes of Gln, respectively. The transport of [3H]Gln (0.58 mM) by the Na+-dependent and the Na+-independent processes was severely inhibited by the addition to the incubation medium of serine, asparagine, and unlabeled Gln. Inducing a relatively negative intravesicular compartment with the use of valinomycin and an outwardly directed K+ gradient or with the use of anions of different lipid permeabilities indicates that Gln transport by the Na+-dependent process is electrogenic in nature. Transport of Gln by the Na+-independent process, however, appeared to be electroneutral in nature. These results demonstrate the existence of two carrier-mediated transport processes for Gln in the human intestinal BBMV, one is Na+ dependent and the other is Na+ independent. Furthermore, the results suggest that Gln transport by the Na+-dependent process probably occurs by a Gln-Na+ cotransport mechanism.

Asparagine↗
PubMed1989

Regulation of intestinal biotin transport in the rat: effect of biotin deficiency and supplementation.

The effect of biotin deficiency and supplementation at pharmacological doses on the intestinal transport of the vitamin was examined in the rat using a brush-border membrane vesicle (BBMV) technique. Transport of biotin in both jejunal and ileal BBMV was significantly (P less than 0.05-0.01) higher in biotin-deficient rats compared with control (pair-fed) rats. This increase in biotin transport appeared to be specific because transport of D-glucose was similar in the two rat groups. The increase in biotin transport in the deficient rats appeared to be mediated through a marked increase (146-230%) in the Vmax of the biotin transport process (with minimal change in the apparent Km), suggesting an increase in the number of the transport carriers. In contrast, supplementation at pharmacological doses of biotin caused significant (P less than 0.05-0.01) and specific decrease (suppression) in biotin transport compared with (unsupplemented) controls. The suppression of biotin transport in the supplemented rats appeared to be mediated through a marked decrease (58%) in the Vmax of the biotin transport process (with minimal change in the apparent Km), suggesting a decrease in the number of the transport carriers. These results provide evidence that biotin transport in the intestine is regulated by the level of the vitamin in the diet (and/or body stores). Furthermore, the results demonstrate the ability of the small intestine to adapt to the challenge of deficiency of an essential nutrient, a capability that may be crucial for the survival of the animal.

Animals↗
PubMed1988

Biotin transport in rat intestinal brush-border membrane vesicles.

Transport of biotin across rat intestinal brush-border membrane was examined using the brush-border membrane vesicle (BBMV) technique. Uptake of biotin by BBMV is the result of transport of the substrate into the intravesicular space with negligible binding to membrane surfaces. In the presence of a Na+ gradient (out greater than in), transport of biotin was higher with a transient 'overshoot' phenomenon. In comparison, transport of biotin in the presence of a choline gradient (out greater than in) was lower with no 'overshoot' phenomenon. In both jejunal and ileal BBMV, the transport of biotin as a function of concentration was saturable in the presence of a Na+ gradient (out greater than in) but was linear in the presence of a choline gradient (out greater than in). Vmax of the Na+-dependent transport system was 0.88 and 0.37 pmol/mg protein per s and apparent Kt was 7.57 and 7.85 microM in jejunal and ileal BBMV, respectively. Structural analogues inhibited the transport process of biotin. Unlike the electrogenic transport of D-glucose, the transport of the anionic biotin was not affected by imposing a relatively positive intravesicular potential with the use of valinomycin and an inwardly-directed K+ gradient, suggesting that biotin transport is most probably an electroneutral process. This suggestion was further supported by studies on biotin transport in the presence of anions of different lipid permeability. The results of this study demonstrate that biotin transport across rat intestinal brush-border membrane is by a carrier-mediated, Na+-dependent and electroneutral process. Furthermore, transport of biotin is higher in the jejunum than the ileum.

Animals↗
PubMed1988

Transport of glycyl-L-proline in intestinal brush-border membrane vesicles of the suckling rat: characteristics and maturation.

Transport of the dipeptide glycine-L-proline (Gly-L-Pro) in the developing intestine of suckling rats and its subsequent maturation in adult rats was examined using the brush-border membrane vesicles (BBMV) technique. Uptake of Gly-L-Pro by BBMV was mainly the result of transport into the intravesicular space with little binding to membrane surfaces. Transport of Gly-L-Pro in BBMV of suckling rats was: (1) Na+ independent; (2) pH dependent with maximum uptake at an incubation buffer pH of 5.0; (3) saturable as a function of concentration (apparent Km = 21.5 +/- 7.9 mM, Vmax = 8.6 +/- 1.5 nmol/mg protein per 10 s); (4) inhibited by other di- and tripeptides; and (5) stimulated and inhibited by inducing a negative and positive intravesicular membrane electrical potential, respectively. Similarly, transport of Gly-L-Pro in intestinal BBMV of adult rats was saturable as a function of concentration (apparent Km = 17.4 +/- 8.6 mM, Vmax = 9.1 +/- 2.1 nmol/mg protein per 10 s) and was stimulated and inhibited by inducing a relatively negative and positive intravesicular membrane potential, respectively. No difference in the transport kinetic parameters of Gly-L-Pro was observed in suckling and adult rats, indicating a similar activity (and/or number) and affinity of the transport carrier in the two age groups. These results demonstrate that the transport of Gly-L-Pro is by a carrier-mediated process which is fully developed at the suckling period. Furthermore, the process is H+-dependent but not Na+-dependent, electrogenic and most probably occurs by a Gly-L-Pro/H+ cotransport mechanism.

Animals↗
PubMed1988

Biotin transport in basolateral membrane vesicles of human intestine.

The characteristics of the exit process of biotin from the enterocyte, i.e., transport across the basolateral membrane, was determined using an enriched basolateral membrane vesicle preparation of human intestine. Purity and suitability of basolateral membrane vesicles for transport studies was confirmed by enzymatic and functional criteria. Orientation of human basolateral membrane vesicles was determined by [3H]ouabain binding studies and was found to be 64% inside-out vesicles and the rest right-side-out vesicles and membrane sheets. Osmolarity studies indicated that the uptake of biotin by these vesicles represents transport into the intravesicular compartment, with little binding to membrane surfaces. The rate of biotin transport was linear for approximately 40 s but decreased thereafter. Transport of biotin was (a) Na+-independent, (b) saturable as a function of concentration, with an apparent KM of 1.1 microM and Vmax of 0.9 pmol/mg protein.15 s, (c) inhibited by structural analogues (desthiobiotin and biotin methyl ester) and related compounds (thioctic acid and thioctic amide), and (d) stimulated by inducing a positive intravesicular electrical potential. These studies are the first to demonstrate the existence of a carrier-mediated transport system for biotin in the basolateral membrane of human intestine.

Basement Membrane↗