| Research Abstract |
We examined the effects of D-glucose on glucose transport activity and insulin-induced insulin resistance in vascular smooth muscle cells (VSMC) cultured in the absence and presence of various concentrations of glucose and insulin. Confluent cultures of VSMC were incubated with serum-free medium containing 0 to 25 mmol/L of D-glucose for 24 to 49 hours at 37ﾟC.After quiescent cells were washed and the glucose transport system was allowed to deactivate, we measured basal and maximally insulin-stimulated 2-deoxyglucose uptake rates. The rate of 2-deoxyglucose uptake decreased in association with increasing concentrations of D-glucose. The basal rates of radiolabeled 2-deoxyglucose uptake in cells pre-exposed to 0,5.5, and 25 mmol/L D-glucose for 5 minutes were 279,230, and 68 pmol/mg protein, respectively. Uptake of 2-deoxyglucose into cells was linear between 0 and 15 minutes of incubation at 37ﾟC.The effect of D-glucose was time-dependent and fully reversible. Insulin increased 2-deoxy
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glucose uptake in cells, but its effect dependent on the preincubation dose of D-glucose. Insulin-stimulated uptake was lower in cells pre-exposed to 25 mmol/L D-glucose than in cells pre-exposed to concentrations of D-glucose below 5.5 mmol/L.At 100 nmol/L insulin, the insulin-stimulated 2-deoxglucose uptake in cells pre-exposed to 0,5.5, and 25 mmol/L D-glucose was 779,514, and 99 pmol/mg protein/5 minutes, respectively. After long-term incubation with insulin insulin-stimulated glucose transport was inhibited. Recovery of glucose transport activity was assessed by incubating cells with D-glucose for 24 to 48 hours to induce desensitization. After 24-hour incubation with serum-free medium containing 5.5 or 25 mmol/L D-glucose, the glucose concentration decreased from 5.5 to 0.4 mmol/L and from 25 to 17.4 mmol/L,respectively. Uptake was lower in cells preincubated with high-dose glucose than cells preincubated with low-dose glucose. The effect of 24-hour glucose conditioning were fully reversible. Uptake of 2-deoxyglucose increased to 78 when cells preincubated in high-dose glucose were incubated further in 5.5 mmol/L glucose and decreased to 228 pmol/mg protein/5 minutes when cells preconditioned with low-dose glucose were incubated with 25 mmol/L glucose. When cells were incubated for 24 hours in serum-free medium containing 0 or 25 mmol/L D-glucose, the rates of 2-deoxyglucose uptake were 263 and 98 pmol/mg protein/5 minutes, respectively. When cells were then transferred to medium containing 5.5 mmol/L D-glucose and incubated for 0 to 24 hours, uptake increased, reaching 228 pmol/mg protein/5 minutes in both cultures. Reversal of the effect of high-dose glucose was rapid, but there was a lag period of 8 hours before 2-deoxyglucose uptake in cells preincubated with low-dose glucose decreased. Our results suggest that D-glucose regulates its own uptake independently of insulin and modulates the ability of insulin to induce insulin resistance in cultured VSMC.Gluc Less
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