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Regulation of glucose transporters in human peritoneal mesothelial cells
Michael Fischereder1, Bernd Schröppel, Patrick Wiese
1Medical Policlinic, Ludwig-Maximilians-University, Munich, Germany. michael.fischereder@klinik.uni-regensburg.de
Background:
Risk factors for peritoneal fibrosis and mesothelial cell (MsC) injury in CAPD are infections and bioincompatibility of the dialysate, including high glucose concentrations. To study a potential link between dialysate and glucose toxicity in MsC, we investigated the expression of facilitative glucose transporters (GLUT), which could contribute to glucose toxicity.
Methods:
After induction of cell differentiation, MsC were incubated in regular medium or medium with 60 mM D-glucose, 30 mM glucose plus 30 mM mannitol, 60 mM mannitol, PD effluent, or with a cytokine mix. Expression of GLUT1, GLUT3, SGLT and GAPDH/L32 was studied by RNase protection assay. MsC were incubated under identical conditions with 14C-fluoro-deoxy-glucose for 30 minutes and glucose uptake was measured. To estimate Vmax and Km, 14C-fluoro-deoxy-glucose uptake rates were determined over a range of 0.6 to 10 mM unlabeled glucose.
Results:
The cytokine mix significantly stimulated GLUT1 expression (3-fold) and GLUT3 (1.7-fold). There was a 1.4-fold increase in GLUT1 (p<0.05) and a 1.7-fold increase in GLUT3 (p<0.05) after incubation in high glucose but not in mannitol or PD-effluent controls. Glucose uptake studies confirmed this increase after incubation in 30 mM (p<0.05) and 60 mM glucose solutions. Kinetic studies showed the Km was approximately 3.7 mM for this transport.
Conclusions:
GLUT mRNA expression and glucose uptake are induced by high ambient glucose concentrations and cytokines. Unlike many other cells, MsC are not able to protect themselves from increased glucose concentrations by downregulation of GLUTs. The intracellular glucose concentration may therefore increase during CAPD, affecting growth factor expression and glycosylation, and contributing to glucose toxicity.
Insights
High glucose and cytokines increase glucose transporters (GLUTs) in mesothelial cells (MsC), potentially causing glucose toxicity during peritoneal dialysis. MsC cannot downregulate GLUTs, leading to higher intracellular glucose levels.
Area of Science:
- Cell Biology
- Nephrology
- Biochemistry
Background:
- Peritoneal fibrosis and mesothelial cell (MsC) injury in continuous ambulatory peritoneal dialysis (CAPD) are linked to infections and dialysate bioincompatibility, including high glucose.
- Investigating the role of facilitative glucose transporters (GLUTs) in MsC is crucial for understanding glucose toxicity.
Purpose of the Study:
- To examine the expression of GLUTs in MsC under conditions mimicking CAPD.
- To determine if high glucose concentrations in dialysate affect glucose uptake and transporter expression in MsC.
Main Methods:
- MsC were incubated with high glucose (60 mM), glucose/mannitol mixtures, PD effluent, or cytokine mix.
- GLUT1, GLUT3, and SGLT expression was analyzed using RNase protection assay.
- 14C-fluoro-deoxy-glucose uptake and kinetic studies (Vmax, Km) were performed to measure glucose transport.
Main Results:
- High glucose significantly increased GLUT1 (1.4-fold) and GLUT3 (1.7-fold) mRNA expression in MsC.
- Cytokine mix also stimulated GLUT1 (3-fold) and GLUT3 (1.7-fold) expression.
- Increased glucose uptake was observed in 30 mM and 60 mM glucose, with a Km of approximately 3.7 mM.
Conclusions:
- High ambient glucose concentrations and cytokines induce GLUT mRNA expression and glucose uptake in MsC.
- MsC exhibit an inability to downregulate GLUTs, unlike other cells, potentially leading to increased intracellular glucose.
- Elevated intracellular glucose during CAPD may contribute to MsC dysfunction and glucose toxicity.