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Apical and basolateral localisation of GLUT2 transporters in human lung epithelial cells.

Kalsi KK, Baker EH, Medina RA, Rice S, Wood DM, Ratoff JC, Philips BJ, Baines DL - Pflugers Arch. (2008)

Bottom Line: In non-polarised H441 cells, uptake of D: -glucose and deoxyglucose was similar.We could not conclusively demonstrate sodium/glucose transporter-mediated transport in non-polarised or polarised cells.We speculate that these transporters could contribute to glucose uptake/homeostasis in the human airway.

View Article: PubMed Central - PubMed

Affiliation: Centre for Ion Channel and Cell Signalling, Division of Basic Medical Sciences, St George's, University of London, Cranmer Terrace, London, SW17 0RE, UK.

ABSTRACT
Glucose concentrations of normal human airway surface liquid are approximately 12.5 times lower than blood glucose concentrations indicating that glucose uptake by epithelial cells may play a role in maintaining lung glucose homeostasis. We have therefore investigated potential glucose uptake mechanisms in non-polarised and polarised H441 human airway epithelial cells and bronchial biopsies. We detected mRNA and protein for glucose transporter type 2 (GLUT2) and glucose transporter type 4 (GLUT4) in non-polarised cells but GLUT4 was not detected in the plasma membrane. In polarised cells, GLUT2 protein was detected in both apical and basolateral membranes. Furthermore, GLUT2 protein was localised to epithelial cells of human bronchial mucosa biopsies. In non-polarised H441 cells, uptake of D: -glucose and deoxyglucose was similar. Uptake of both was inhibited by phloretin indicating that glucose uptake was via GLUT-mediated transport. Phloretin-sensitive transport remained the predominant route for glucose uptake across apical and basolateral membranes of polarised cells and was maximal at 5-10 mM glucose. We could not conclusively demonstrate sodium/glucose transporter-mediated transport in non-polarised or polarised cells. Our study provides the first evidence that glucose transport in human airway epithelial cells in vitro and in vivo utilises GLUT2 transporters. We speculate that these transporters could contribute to glucose uptake/homeostasis in the human airway.

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Western blot analysis of total protein and non-bound fraction (30 μg) or biotinylated apical and basolateral proteins isolated from 1 mg of total protein extracted from polarised H441 cells. a Immunostained proteins corresponding to GLUT2 (∼60 kDa) were present in the total, biotinylated apical and basolateral protein fractions. b Immunostained proteins corresponding to the α1 subunit of Na+K+ATPase (∼113 kDa) were present predominantly in the total protein and basolateral fractions. a and b Immunostained products corresponding to β-actin (∼42 kDa) were present in the total protein and non-bound fractions. c Effect of 1, 5 and 15 mM glucose on GLUT2 transporter abundance in biotinylated apical and basolateral proteins from polarised H441 cells
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Fig8: Western blot analysis of total protein and non-bound fraction (30 μg) or biotinylated apical and basolateral proteins isolated from 1 mg of total protein extracted from polarised H441 cells. a Immunostained proteins corresponding to GLUT2 (∼60 kDa) were present in the total, biotinylated apical and basolateral protein fractions. b Immunostained proteins corresponding to the α1 subunit of Na+K+ATPase (∼113 kDa) were present predominantly in the total protein and basolateral fractions. a and b Immunostained products corresponding to β-actin (∼42 kDa) were present in the total protein and non-bound fractions. c Effect of 1, 5 and 15 mM glucose on GLUT2 transporter abundance in biotinylated apical and basolateral proteins from polarised H441 cells

Mentions: GLUT2 (but not GLUT1, GLUT3 or GLUT4) proteins were detected in the membranes of polarised cells. GLUT2 was equally distributed in the biotinylated apical and basolateral membrane fractions but was not detected in the intracellular non-bound protein fraction (Fig. 8a). α1-Na+K+ATPase was predominantly located in the basolateral biotinylated fraction and the total protein lysate but not in the non-bound fraction consistent with its distribution in the basolateral membrane of these cells (Fig. 8b). β-actin was present at similar abundance in all total protein preparations but was not detected in apical and basolateral membrane preparations, indicating that these fractions were not contaminated with intracellular proteins (Fig. 8a,b).Fig. 8


Apical and basolateral localisation of GLUT2 transporters in human lung epithelial cells.

Kalsi KK, Baker EH, Medina RA, Rice S, Wood DM, Ratoff JC, Philips BJ, Baines DL - Pflugers Arch. (2008)

Western blot analysis of total protein and non-bound fraction (30 μg) or biotinylated apical and basolateral proteins isolated from 1 mg of total protein extracted from polarised H441 cells. a Immunostained proteins corresponding to GLUT2 (∼60 kDa) were present in the total, biotinylated apical and basolateral protein fractions. b Immunostained proteins corresponding to the α1 subunit of Na+K+ATPase (∼113 kDa) were present predominantly in the total protein and basolateral fractions. a and b Immunostained products corresponding to β-actin (∼42 kDa) were present in the total protein and non-bound fractions. c Effect of 1, 5 and 15 mM glucose on GLUT2 transporter abundance in biotinylated apical and basolateral proteins from polarised H441 cells
© Copyright Policy
Related In: Results  -  Collection

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Fig8: Western blot analysis of total protein and non-bound fraction (30 μg) or biotinylated apical and basolateral proteins isolated from 1 mg of total protein extracted from polarised H441 cells. a Immunostained proteins corresponding to GLUT2 (∼60 kDa) were present in the total, biotinylated apical and basolateral protein fractions. b Immunostained proteins corresponding to the α1 subunit of Na+K+ATPase (∼113 kDa) were present predominantly in the total protein and basolateral fractions. a and b Immunostained products corresponding to β-actin (∼42 kDa) were present in the total protein and non-bound fractions. c Effect of 1, 5 and 15 mM glucose on GLUT2 transporter abundance in biotinylated apical and basolateral proteins from polarised H441 cells
Mentions: GLUT2 (but not GLUT1, GLUT3 or GLUT4) proteins were detected in the membranes of polarised cells. GLUT2 was equally distributed in the biotinylated apical and basolateral membrane fractions but was not detected in the intracellular non-bound protein fraction (Fig. 8a). α1-Na+K+ATPase was predominantly located in the basolateral biotinylated fraction and the total protein lysate but not in the non-bound fraction consistent with its distribution in the basolateral membrane of these cells (Fig. 8b). β-actin was present at similar abundance in all total protein preparations but was not detected in apical and basolateral membrane preparations, indicating that these fractions were not contaminated with intracellular proteins (Fig. 8a,b).Fig. 8

Bottom Line: In non-polarised H441 cells, uptake of D: -glucose and deoxyglucose was similar.We could not conclusively demonstrate sodium/glucose transporter-mediated transport in non-polarised or polarised cells.We speculate that these transporters could contribute to glucose uptake/homeostasis in the human airway.

View Article: PubMed Central - PubMed

Affiliation: Centre for Ion Channel and Cell Signalling, Division of Basic Medical Sciences, St George's, University of London, Cranmer Terrace, London, SW17 0RE, UK.

ABSTRACT
Glucose concentrations of normal human airway surface liquid are approximately 12.5 times lower than blood glucose concentrations indicating that glucose uptake by epithelial cells may play a role in maintaining lung glucose homeostasis. We have therefore investigated potential glucose uptake mechanisms in non-polarised and polarised H441 human airway epithelial cells and bronchial biopsies. We detected mRNA and protein for glucose transporter type 2 (GLUT2) and glucose transporter type 4 (GLUT4) in non-polarised cells but GLUT4 was not detected in the plasma membrane. In polarised cells, GLUT2 protein was detected in both apical and basolateral membranes. Furthermore, GLUT2 protein was localised to epithelial cells of human bronchial mucosa biopsies. In non-polarised H441 cells, uptake of D: -glucose and deoxyglucose was similar. Uptake of both was inhibited by phloretin indicating that glucose uptake was via GLUT-mediated transport. Phloretin-sensitive transport remained the predominant route for glucose uptake across apical and basolateral membranes of polarised cells and was maximal at 5-10 mM glucose. We could not conclusively demonstrate sodium/glucose transporter-mediated transport in non-polarised or polarised cells. Our study provides the first evidence that glucose transport in human airway epithelial cells in vitro and in vivo utilises GLUT2 transporters. We speculate that these transporters could contribute to glucose uptake/homeostasis in the human airway.

Show MeSH