Identification of persistent defects in insulin receptor structure and function capillary endothelial cells from diabetic rats.
Insulin actions and receptors were studied in capillary endothelial cells cultured from diabetic BB rats and their nondiabetic colony mates. The endothelial cells from diabetic rats of 2 mo duration had persistent biological and biochemical defects in culture. Compared with normal rats, endothelial cells from diabetic rats grew 44% more slowly. Binding studies of insulin and insulin-like growth factor I (IGF-I) showed that cells from diabetic rats had 50% decrease of insulin receptor binding (nondiabetic: 4.6 +/- 0.7; diabetic: 2.6 +/- 0.4% per milligram protein, P less than 0.01), which was caused by a 50% decrease in the number of binding sites per milligram protein, whereas IGF-I binding was not changed. Insulin stimulation of 2-deoxy-glucose uptake and alpha-aminoisobutyric acid uptake were also severely impaired with a 80-90% decrease in maximal stimulation, in parallel with a 62% decrease in insulin-stimulated autophosphorylation (P less than 0.05). 125I-insulin cross-linking revealed an 140-kD alpha subunit of the insulin receptor similar to that in cells from nondiabetic rats, although bands at greater than 200 kD were also detected. The molecular weight of the insulin receptor beta subunit (by SDS-PAGE) was smaller in cells from diabetic than from normal rats (88-90 vs. 95 kD). Neuraminadase treatment of the partially purified insulin receptors decreased the molecular weight of the insulin receptors from nondiabetic rats to a greater degree than its diabetic counterpart. In contrast, Northern blot analysis of insulin receptor mRNAs using human cDNA probes revealed two species of 9.4 and 7.2 kb with no difference in mRNA abundance between cells from diabetic and nondiabetic rats. We conclude that the exposure of capillary endothelial cells to a diabetic milieu in vivo can cause specific and persistent changes in the insulin receptor and insulin action.
Kwok
C F
CF
Research Division, Joslin Diabetes Center, Boston, Massachusetts 02215.
Goldstein
B J
BJ
Muller-Wieland
D
D
Lee
T S
TS
Kahn
C R
CR
King
G L
GL
D000818:Animals; D001667:Binding, Competitive; D002196:Capillaries / Q000032:analysis; D004247:DNA / Q000378:metabolism; D003921:Diabetes Mellitus, Experimental / Q000473:pathology*; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000032:analysis*; D006943:Hyperglycemia / Q000378:metabolism; D009439:Neuraminidase / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D011972:Receptor, Insulin / Q000032:analysis*; D013936:Thymidine / Q000378:metabolism
D004247:DNA; D011972:Receptor, Insulin; D009439:Neuraminidase; D013936:Thymidine