Diabetes mellitus induces long lasting changes in the glucose transporter of rat heart endothelial cells.
The accumulation of glucose exerts various cytotoxic effects on endothelial and other vascular cells, and thereby contributes to the development of microvascular complications in diabetes. Since tissues, in which vascular complications typically occur, do not take up glucose in an insulin regulated manner, it is an important question to know whether other mechanisms exist in these cells to restrict the uptake and the accumulation of glucose. To study this question, we used microvascular endothelial cells isolated from rat heart endothelial cells (RHEC). In RHEC, the non-insulin regulated glucose transporter (Glut-1) was detected as a broad protein band of 50-65 kD. In contrast, the Glut-1 from rat brain, which was taken as reference, had a molecular weight of 45 kD. After treatment with endoglycosidase F, both proteins formed a band of approximately 40 kD on SDS-PAGE, demonstrating a more extensive glycosylation of Glut-1 in RHEC as compared to brain. Incubation of the cells in high glucose (22 mM, up to 10 days) did not down-regulate either Glut-1 protein or mRNA. In contrast to high glucose, deprivation of the cells from glucose led to an increase in Glut-1 mRNA and protein which is partly non-glycosylated. In cells from hearts of streptozotocin-diabetic rats (DRHEC), Glut-1 protein, but not Glut-1 mRNA, was reduced by about 40%. Additionally, a significant amount of glycosyl residues was resistant to the enzymatic treatment with N-endoglycosidase F. Both changes in Glut-1 were also observed when the cells were cultivated in low glucose (5.5 mM) for several passages indicating a long lasting, hardly reversible modification of Glut-1 by diabetes. These data indicate that Glut-1 is not down-regulated in RHEC by high glucose, and that this important mechanism to protect the endothelium against an intracellular accumulation of glucose is missing in RHEC. As a consequence, increases in blood glucose may lead to a glucose overload with the described deleterious effects on the structure and function of endothelium.
Hirsch
B
B
German Diabetes Research Institute, Department of Clinical Biochemistry, Düsseldorf.
Rösen
P
P
D000818:Animals; D001692:Biological Transport / Q000502:physiology; D015152:Blotting, Northern; D015153:Blotting, Western; D002478:Cells, Cultured; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism*; D004727:Endothelium / Q000166:cytology; D015870:Gene Expression / Q000502:physiology; D005947:Glucose / Q000378:metabolism / Q000493:pharmacokinetics; D051272:Glucose Transporter Type 1; D007700:Kinetics; D008297:Male; D009004:Monosaccharide Transport Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D018485:Muscle Fibers, Skeletal / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D009206:Myocardium / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D010766:Phosphorylation; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017208:Rats, Wistar
D051272:Glucose Transporter Type 1; D009004:Monosaccharide Transport Proteins; D012333:RNA, Messenger; C495679:Slc2a1 protein, rat; D005947:Glucose