Amino acid sequence and the cellular location of the Na(+)-dependent D-glucose symporters (SGLT1) in the ovine enterocyte and the parotid acinar cell.
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ABSTRACT:
SUBMITTER: Tarpey PS
PROVIDER: S-EPMC1136258 | biostudies-other | 1995 Nov
REPOSITORIES: biostudies-other
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Project description:We have investigated the mechanisms of regulation of the Na+/glucose co-transporter (SGLT1) in a ruminant animal, which is an exceptional model system for studying intestinal glucose transport. Pre-ruminant lambs absorb glucose, produced by hydrolysis of the milk sugar lactose, in the intestine via apical SGLT1 and basolateral facilitative glucose transporters (GLUT2). Weaning coincides with the development of the rumen, and consequently the amount of hexoses reaching the small intestine of the ruminant sheep is undetectable. During development, SGLT1 activity and abundance in intestinal brush-border membranes decreased by over 200-fold, and either maintaining lambs on a milk replacer diet or infusing sheep intestine with D-glucose restored co-transporter activity and expression. We have measured ovine intestinal SGLT1 mRNA levels during development, with changes in diet and after direct infusion of D-glucose or methyl alpha-D-glucopyranoside into the intestinal lumen, in order to determine the level of regulation. During development, mRNA levels decreased only 4-fold. Lambs maintained on a milk replacer diet showed no change in mRNA levels relative to age-matched controls. Finally, upon infusion of the intestine of the ruminant sheep with sugars, D-glucose infusion increased SGLT1 mRNA, but only by 2-fold, compared with a 60-90-fold increase in co-transporter number and activity. Since the change in Na(+)-dependent glucose transport activity is correlated with SGLT1 protein abundance, and since changes in mRNA levels do not account for the dramatic changes in protein abundance, we conclude that the principal level of SGLT1 regulation by luminal sugar is translational or post-translational.
| S-EPMC1132544 | biostudies-other
Project description:It has been suggested that glucose absorption in the small intestine depends on both constitutively expressed SGLT1 and translocated GLUT2 in the brush border membrane, especially in the presence of high levels of luminal glucose. Here, we present a computational model of non-isotonic glucose uptake by small intestinal epithelial cells. The model incorporates apical uptake via SGLT1 and GLUT2, basolateral efflux into the blood via GLUT2, and cellular volume changes in response to non-isotonic conditions. The dependence of glucose absorption on luminal glucose, blood flow rate, and inlet blood glucose concentration is studied. Uptake via apical GLUT2 is found to be sensitive to all these factors. Under a range of conditions, the maximum apical GLUT2 flux is about half of the SGLT1 flux and is achieved at high luminal glucose (> 50 mM), high blood flow rates, and low inlet blood concentrations. In contrast, SGLT1 flux is less sensitive to these factors. When luminal glucose concentration is less than 10 mM, apical GLUT2 serves as an efflux pathway for glucose to move from the blood to the lumen. The model results indicate that translocation of GLUT2 from the basolateral to the apical membrane increases glucose uptake into the cell; however, the reduction of efflux capacity results in a decrease in net absorption. Recruitment of GLUT2 from a cytosolic pool elicits a 10-20% increase in absorption for luminal glucose levels in the a 20-100 mM range. Increased SGLT1 activity also leads to a roughly 20% increase in absorption. A concomitant increase in blood supply results in a larger increase in absorption. Increases in apical glucose transporter activity help to minimise cell volume changes by reducing the osmotic gradient between the cell and the lumen.
| S-EPMC8688934 | biostudies-literature
Project description:The stimulation of fluid and electrolyte secretion in salivary cells results in ionic changes that promote rapid increases in the activity of the Na,K-ATPase. In many cell systems, there are conflicting findings concerning the regulation of the phosphorylation of the Na,K-ATPase α subunit, which is the catalytic moiety. Initially, we investigated the phosphorylation sites on the α1 subunit in native rat parotid acinar cells using tandem mass spectrometry and identified two new phosphorylation sites (Ser(222), Ser(407)), three sites (Ser(217), Tyr(260), Ser(47)) previously found from large scale proteomic screens, and two sites (Ser(23), Ser(16)) known to be phosphorylated by PKC. Subsequently, we used phospho-specific antibodies to examine the regulation of phosphorylation on Ser(23) and Ser(16) and measured changes in ERK phosphorylation in parallel. The G-protein-coupled muscarinic receptor mimetic carbachol, the phorbol ester phorbol 12-myristate 13-acetate, the Ca(2+) ionophore ionomycin, and the serine/threonine phosphatase inhibitor calyculin A increased Ser(23) α1 phosphorylation. Inhibition of classical PKC proteins blocked carbachol-stimulated Ser(23) α1 subunit phosphorylation but not ERK phosphorylation, which was blocked by an inhibitor of novel PKC proteins. The carbachol-initiated phosphorylation of Ser(23) α1 subunit was not modified by ERK or PKA activity. The Na,K-ATPase inhibitor ouabain reduced and enhanced the carbachol-promoted phosphorylation of Ser(23) and Ser(16), respectively, the latter because ouabain itself increased Ser(16) phosphorylation; thus, both sites display conformational-dependent phosphorylation changes. Ouabain-initiated phosphorylation of Ser(16) α1 was not blocked by PKC inhibitors, unlike carbachol- or phorbol 12-myristate 13-acetate-initiated phosphorylations, suggesting that this site was also a substrate for a kinase other than PKC.
| S-EPMC2978561 | biostudies-literature
Project description:To clarify the physiological role of Na(+)-D-glucose cotransporter SGLT1 in small intestine and kidney, Sglt1(-/-) mice were generated and characterized phenotypically. After gavage of d-glucose, small intestinal glucose absorption across the brush-border membrane (BBM) via SGLT1 and GLUT2 were analyzed. Glucose-induced secretion of insulinotropic hormone (GIP) and glucagon-like peptide 1 (GLP-1) in wild-type and Sglt1(-/-) mice were compared. The impact of SGLT1 on renal glucose handling was investigated by micropuncture studies. It was observed that Sglt1(-/-) mice developed a glucose-galactose malabsorption syndrome but thrive normally when fed a glucose-galactose-free diet. In wild-type mice, passage of D-glucose across the intestinal BBM was predominantly mediated by SGLT1, independent the glucose load. High glucose concentrations increased the amounts of SGLT1 and GLUT2 in the BBM, and SGLT1 was required for upregulation of GLUT2. SGLT1 was located in luminal membranes of cells immunopositive for GIP and GLP-1, and Sglt1(-/-) mice exhibited reduced glucose-triggered GIP and GLP-1 levels. In the kidney, SGLT1 reabsorbed ∼3% of the filtered glucose under normoglycemic conditions. The data indicate that SGLT1 is 1) pivotal for intestinal mass absorption of d-glucose, 2) triggers the glucose-induced secretion of GIP and GLP-1, and 3) triggers the upregulation of GLUT2.
| S-EPMC3237647 | biostudies-literature
Project description:The expression of the Na+/glucose cotransporter (SGLT1) in response to thyroid hormone [3,5,3'-tri-iodo-l-thyronine (T3)] was investigated in the enterocytic model cell line Caco-2/TC7. In differentiated cells, T3 treatment induces an average 10-fold increase in glucose consumption as well as a T3 dose-dependent increase in SGLT1 mRNA abundance. Only cells grown on glucose-containing media, but not on the non-metabolizable glucose analogue alpha-methylglucose (AMG), could respond to T3-treatment. The Vmax parameter of AMG transport was enhanced 6-fold by T3 treatment, whereas the protein abundance of SGLT1 was unchanged. The role of Na+ recycling in the T3-related activation of SGLT1 activity was suggested by both the large increase in Na+/K+ATPase protein abundance and the inhibition, down to control levels, of AMG uptake in ouabain-treated cells. Further investigations aimed at identifying the presence of a second cotransporter that could be expressed erroneously in the colon cancer cell line were unsuccessful: T3-treatment did not modify the sugar-specificity profile of AMG transport and did not induce the expression of SGLT2 as assessed by reverse transcription-PCR. Our results show that T3 can stimulate the SGLT1 cotransport activity in Caco-2 cells. Both transcriptional and translational levels of regulation are involved. Finally, glucose metabolism is required for SGLT1 expression, a result that contrasts with the in vivo situation and may be related to the fetal phenotype of the cells.
| S-EPMC1219733 | biostudies-other
Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
2015-02-03 | E-GEOD-65587 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the SubSeries listed below.
2015-02-03 | GSE65587 | GEO
Project description:A correlation between gastrointestinal (GI) inflammation and gut hormones has reported that inflammatory stimuli including bacterial endotoxins, lipopolysaccharides (LPS), TNFα, IL-1β, and IL-6 induces high levels of incretin hormone leading to glucose dysregulation. Although incretin hormones are immediately secreted in response to environmental stimuli, such as nutrients, cytokines, and LPS, but studies of glucose-induced incretin secretion in an inflamed state are limited. We hypothesized that GI inflammatory conditions induce over-stimulated incretin secretion via an increase of glucose-sensing receptors. To confirm our hypothesis, we observed the alteration of glucose-induced incretin secretion and glucose-sensing receptors in a GI inflammatory mouse model, and we treated a conditioned media (Mϕ 30%) containing inflammatory cytokines in intestinal epithelium cells and enteroendocrine L-like NCI-H716 cells. In GI-inflamed mice, we observed that over-stimulated incretin secretion and insulin release in response to glucose and sodium glucose cotransporter (Sglt1) was increased. Incubation with Mϕ 30% increases Sglt1 and induces glucose-induced GLP-1 secretion with increasing intracellular calcium influx. Phloridzin, an sglt1 inhibitor, inhibits glucose-induced GLP-1 secretion, ERK activation, and calcium influx. These findings suggest that the abnormalities of incretin secretion leading to metabolic disturbances in GI inflammatory disease by an increase of Sglt1.
| S-EPMC6566487 | biostudies-literature
Project description:LEFTY2 (endometrial bleeding associated factor; EBAF or LEFTYA), a cytokine released shortly before menstrual bleeding, is a negative regulator of cell proliferation and tumour growth. LEFTY2 down-regulates Na+/H+ exchanger activity with subsequent inhibition of glycolytic flux and lactate production in endometrial cancer cells. Glucose can be utilized not only for glycolysis but also for glycogen formation. Both glycolysis and glycogen formation require cellular glucose uptake which could be accomplished by the Na+ coupled glucose transporter-1 (SGLT1; SLC5A1). The present study therefore explored whether LEFTY2 modifies endometrial SGLT1 expression and activity as well as glycogen formation. Ishikawa and HEC1a cells were exposed to LEFTY2, SGLT1 and glycogen synthase (GYS1) transcript levels determined by qRT-PCR. SGLT1, GYS1 and phospho-GYS1 protein abundance was quantified by western blotting, cellular glucose uptake from 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino)-2-Deoxyglucose (2-NBDG) uptake, and cellular glycogen content utilizing an enzymatic assay and subsequent colorimetry. As a result, a 48-hour treatment with LEFTY2 significantly increased SGLT1 and GYS1 transcript levels as well as SGLT1 and GYS1 protein abundance in both Ishikawa and HEC1a cells. 2-NBDG uptake and cellular glycogen content were upregulated significantly in Ishikawa (type 1) but not in type 2 endometrial HEC1a cells, although there was a tendency of increased 2-NBDG uptake. Further, none of the effects were seen in human benign endometrial cells (HESCs). Interestingly, in both Ishikawa and HEC1a cells, a co-treatment with TGF-? reduced SGLT1, GYS and phospho-GYS protein levels, and thus reduced glycogen levels and again HEC1a cells had no significant change. In conclusion, LEFTY2 up-regulates expression and activity of the Na+ coupled glucose transporter SGLT1 and glycogen synthase GYS1 in a cell line specific manner. We further show the treatment with LEFTY2 fosters cellular glucose uptake and glycogen formation and TGF-? can negate this effect in endometrial cancer cells.
| S-EPMC7112196 | biostudies-literature
Project description:Terminal differentiation in parotid acini relies on sustained changes in gene expression during the first few postnatal weeks. Little is known about what drives these changes. Expression measurements along with knowledgebased network analysis was used to develop a prospective gene regulatory network that drives differentiation. We used both microRNA and mRNA expression measurements along with knowledgebased network analysis was used to develop a prospective gene regulatory network that drives differentiation. laser capture microdissection was used to isolate acinar cells from the parotid at four timepoints in triplicate (E20, P5, P15, and P25). RNA was isolated, and used to measure microRNA expression.
2015-01-27 | E-GEOD-65324 | biostudies-arrayexpress