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Evidence against impaired brain microtubule protein polymerization at high glucose concentrations or during diabetes
E Y Eaker1, J M Angelastro, D L Purich
1Department of Medicine, University of Florida College of Medicine, Gainesville 32610.
Abstract:
Previous studies suggest that brain microtubule protein exposed to high glucose levels or isolated from diabetic rats can become glucosylated and that this impairs GTP-induced microtubule polymerization. We set out to extend that investigation to define the mechanistic basis for inhibition of microtubule assembly during diabetes or on incubation at high glucose levels. Rat and bovine brain microtubule protein was purified by cycles of polymerization/depolymerization. When microtubules were incubated for 1 h in either buffer or buffer containing glucose (up to 165 mM), there was no difference in polymerization, a finding contrary to the earlier study. Other rats were injected with vehicle or streptozotocin (90 mg/kg) to induce diabetes as evidenced by serum glucose in excess of 300 mg%, and at 4 weeks, brain microtubule protein was isolated by the polymerization cycling method. Again, there was no difference in the amount or purity of isolated microtubule protein between control or diabetic rats. We also observed no increase in microtubule glucosylation, and GTP-induced polymerization in vitro was indistinguishable for protein derived from brains of normal rats and rats with diabetes as measured by turbidity or electron microscopy. Our results suggest that in vitro incubation with glucose or in vivo elevation of glucose during diabetes fails to impair microtubule polymerization, pointing to other mechanisms for the neuropathy associated with diabetes.
Insights
High glucose levels and diabetes do not impair brain microtubule polymerization. This study found no evidence of glucosylation affecting microtubule assembly, suggesting other mechanisms for diabetic neuropathy.
Area of Science:
- Neuroscience
- Biochemistry
- Cell Biology
Background:
- Previous research indicated high glucose and diabetes impair brain microtubule polymerization via glucosylation.
- The mechanistic basis for this proposed impairment remained unclear.
Purpose of the Study:
- To investigate the mechanistic basis for inhibition of microtubule assembly during diabetes or high glucose incubation.
- To determine if glucose exposure or diabetes affects microtubule protein glucosylation and polymerization.
Main Methods:
- Purification of rat and bovine brain microtubule protein using polymerization/depolymerization cycles.
- In vitro incubation of microtubules with varying glucose concentrations.
- Induction of diabetes in rats using streptozotocin and subsequent isolation of brain microtubule protein.
- Assessment of microtubule glucosylation and GTP-induced polymerization via turbidity and electron microscopy.
Main Results:
- In vitro incubation with glucose did not alter microtubule polymerization.
- Microtubule protein isolated from diabetic rats showed no difference in amount, purity, or glucosylation compared to controls.
- GTP-induced polymerization of microtubule protein from normal and diabetic rats was indistinguishable.
- No increased microtubule glucosylation was observed in diabetic rats.
Conclusions:
- In vitro glucose incubation and in vivo diabetes do not impair brain microtubule polymerization.
- The proposed mechanism of glucosylation affecting microtubule assembly in diabetes is not supported by these findings.
- Alternative mechanisms likely underlie the development of diabetic neuropathy.