Glycemia-lowering effect of cobalt chloride in the diabetic rat: increased GLUT1 mRNA expression.

We have recently shown that expression of the GLUT1 glucose transporter isoform is augmented in cells exposed to cobalt chloride [Co(II)], an agent that stimulates the expression of hypoxia-responsive genes (Behrooz, A., Ismail-Beigi, F., 1997. J. Biol. Chem. 272, 5555-5562.). Here, we examine the effect of Co(II) on glycemia and tissue GLUT1 mRNA content of normal and diabetic rats. The addition of 2 mM Co(II) in the drinking water reduced the glycemia of streptozotocin-induced diabetic rats by day 3 from 32.3 +/- 2.1 to 21.0 +/- 1.9 mM (non-fasting). Co(II) resulted in no change in serum insulin levels of normal or diabetic rats. Treatment with 4 mM Co(II) was more effective than 2 mM Co(II) in reducing the glycemia of diabetic rats, while 6 mM Co(II) was associated with severe toxicity. GLUT1 mRNA content increased significantly in ventricular myocardium, renal cortex, skeletal muscle, cerebrum and liver of normal and diabetic rats treated with 2 mM cobalt chloride (ranging from 1.3- to 2.9-fold in the different tissues). It is concluded that: (1) treatment with Co(II) decreases the glycemia of diabetic rats, and (2) the glycemia-lowering effect of Co(II) is associated with, and may be mediated by, enhanced expression of GLUT1 mRNA.

Mechanism of stimulation of glucose transport in response to inhibition of oxidative phosphorylation: analysis with myc-tagged Glut1.

To determine the role of 'translocation' vs. 'activation' of Glut1 in the stimulation of glucose transport in response to inhibition of oxidative phosphorylation, we measured the abundance of myc-tagged Glut1 in plasma membrane of stably transfected Clone 9 cells, a rat liver cell line expressing only the Glut1 isoform. The myc epitope-tag is located between Ile56 and Pro57 in the putative first extracellular loop of Glut1. Under basal conditions, transfected cells expressed approximately 3 fold higher levels of Glut1 and exhibited a approximately 3 fold higher rate of glucose transport than non-transfected cells. To delineate the mechanism mediating the stimulation of glucose transport by a azide we employed two strategies: (1) mild cell surface biotinylation followed by isolation of plasma membranes and quantitation of Glut1 sites in Western blots employing anti-Glut1 and anti-myc antibodies, and (2) quantitative immunofluorescence of myc epitopes in plasma membrane sheets. The rate of glucose transport increased 2.9 +/- 0.5 fold in transfected cells exposed to 5 mM azide for 1 h. Exposure to azide, however, resulted in no significant increase in Glut1 content of plasma membranes using anti-Glut1 or anti-myc antibodies in Western blots (1.0 +/- 0.1 and 0.9 +/- 0.2 fold, respectively; azide/control), and was associated with no detectable increase in immunofluorescence using either anti-Glut1 or anti-myc antibodies (p > 0.1 for both measurements). Treatment of cells with cobalt chloride (employed as a positive control) resulted in marked increases in glucose transport, cell and plasma membrane Glut1 content, and immunofluorescence of plasma membrane sheets (8-10 fold increase in each parameter). We conclude that the stimulation of glucose transport by azide results mainly from activation of Glut1 transporters pre-existing in the plasma membrane.

Effects of intra-articular vitamin E and corticosteroid injection in experimental hemarthrosis in rabbits.

The effects of intra-articular injections of vitamin E and corticosteroid were investigated in skeletally mature New Zealand white rabbits in which experimental hemarthrosis was induced for 14 days. The rabbits were divided into three groups composed of eight rabbits each: the first group comprised of animals with hemarthrosis, the second group animals with hemarthrosis and intra-articular injections of 20 mg vitamin E, and the third group animals with hemarthrosis and intra-articular injections of 10 mg of triamcinolone acetonide (TCA). Proteoglycan levels in the joint cartilage of the hemarthrosis group were found to be increased significantly compared with the controls (P < .01), whereas in the vitamin E-injected group they were significantly decreased (P < .05). In the TCA-injected group, proteoglycan levels were not found to be significantly different from those in the hemarthrosis group (P > .05). Histopathological evaluation showed that the cartilage structure in the joint of the control limbs was identical to that in the vitamin E- and TCA-injected limbs. In the hemarthrosis group, in comparison with the controls, the joint surface was roughened and fibrillated. In the superficial areas of the cartilage tissue, chondrocytes were decreased in number. These findings suggest that in this model, vitamin E and TCA may be helpful in preventing the joint cartilage changes seen in hemarthrosis.