Nondiabetic Hamsters Research Articles

The effect of vitamin E, probucol, and lovastatin on oxidative status and aortic fatty lesions in hyperlipidemic-diabetic hamsters

Diabetes mellitus is associated with an increased risk of premature atherosclerosis, which may be due in part to an increased rate of low density lipoprotein (LDL) oxidation. Previous studies have shown that vitamin E, probucol, and lovastatin can reduce the oxidative susceptibility of LDL in normoglycemic animal models; however, few studies have investigated this in conjunction with aortic fatty streak lesion formation in diabetic hyperlipidemic models. Forty-eight Syrian hamsters were made diabetic by intraperitoneal injection of low dose streptozotocin. Diabetic animals (12 animals/groups) received a high saturated fat and cholesterol diet for 12.5 weeks. At 2.5 week of dietary treatments, the diet was supplemented with either: (1) 500 IU/day vitamin E (D+E); (2) 1% probucol w/w of the diet (D+P); (3) 25 mg/kg lovastatin (D+L); or (4) diabetic control (D). An age-matched group of hamsters ( n=6) receiving the same diet but not made diabetic (ND) was used as control. At the end of the study, aortic arch foam cell-rich fatty streak lesion, plasma glucose, total cholesterol (TC), high density lipoprotein cholesterol (HDL-C), non-HDL-C, triglycerides (TG), phospholipids, α-tocopherol, plasma lipid peroxide and the susceptibility of LDL to copper-catalyzed oxidation were determined. Diabetes increased plasma glucose, and when combined with an atherogenic diet resulted in a further increase of plasma lipids. Vitamin E, probucol, and lovastatin significantly reduced plasma TG in the diabetic hamsters fed the atherogenic diet. Vitamin E treatment increased TC, probucol reduced HDL-C without affecting TC; whereas lovastatin reduced TC and selectively decreased non-HDL-C, and significantly reduced fatty streak lesion formation in the aortic arch. While vitamin E and probucol were effective in reducing several indices of oxidative stress including plasma lipid peroxides, cholesterol oxidation products and in vitro LDL oxidation, they had no effect on fatty streak lesion formation. Our results indicate that the LDL in diabetic animals is more susceptible to oxidation than in non-diabetic hamsters and that not only vitamin E and probucol but also lovastatin provide antioxidant protection. It appears that in this combined model of diabetes and hypercholesterolemia, progression of fatty streak lesion formation is mainly associated with changes in TC and non-HDL-C as affected by lovastatin, and is less dependent on the extent of LDL oxidation, changes in plasma TG level and oxidative stress status.

Aminoguanidine Suppresses Basal Macromolecular Extravasation during Diabetes Mellitus

The goal of this study was to determine the effect of aminoguanidine on basal macromolecular efflux from the microcirculation of the hamster cheek pouch during diabetes mellitus. We used intravital fluorescent microscopy and fluorescein isothiocyanate dextrans (FITC-dextran; mw = 70 and 20K) to examine basal macromolecular extravasation in nondiabetic hamsters, nondiabetic hamsters treated with a topical application of aminoguanidine (0.5 mM), diabetic hamsters (6–10 weeks after injection of streptozotocin), and diabetic hamsters treated with a topical application of aminoguanidine (0.5 mM). Increases in macromolecular efflux were quantitated by calculating the clearance (ml/s × 10−6) of FITC-dextran-70K and -20K. In nondiabetic hamsters, the clearance of FITC-dextran-70K and -20K remained relatively constant during the experimental period, although the clearance of FITC-dextran-70K was less than that for FITC-dextran-20K. Topical application of aminoguanidine did not alter basal permeability characteristics in nondiabetic hamsters. In diabetic hamsters, clearance of FITC-dextran-70K and -20K also remained relatively constant during the experimental period. However, the magnitude of clearance of FITC-dextran-70K and and -20K was significantly greater in diabetic compared to nondiabetic hamsters (P < 0.05). Topical application of aminoguanidine restored basal permeability characteristics of diabetic hamsters to that observed in nondiabetic hamsters. These findings suggest that acute treatment of the microcirculation in vivo with aminoguanidine ameliorates basal increases in extravasation of macromolecules during diabetes mellitus. We suggest that aminoguanidine suppresses basal macromolecular efflux in diabetic hamsters via inhibition of nitric oxide synthase. Thus, it appears that the use of aminoguanidine may be an important therapeutic approach for the treatment of diabetes-related vascular dysfunction.

Constrictor responses of resistance arterioles during diabetes mellitus

While diabetes mellitus appears to alter nitric oxide synthase-dependent vasodilatation, the effect of diabetes on constrictor responses of resistance arterioles is not clear. Our goal was to examine the effect of diabetes on constrictor responses of cheek pouch arterioles. In vivo diameter of arterioles (∼50 μm) was measured in response to norepinephrine, the thromboxane analogue (U-46619) and endothelin-1 in nondiabetic and diabetic hamsters (4–6 weeks post streptozotocin). Norepinephrine (1.0 and 10 nM) and U-46619 (0.1 and 1.0 nM) produced similar dose-related vasoconstriction in nondiabetic and diabetic hamsters ( P>0.05). In contrast, vasoconstriction to endothelin-1 (0.1 and 1.0 pM) was greater in diabetic than nondiabetic hamsters ( P<0.05). Next, we examined the role of nitric oxide in basal vascular tone and whether enhanced vasoconstriction in diabetic hamsters to endothelin-1 might be related to an alteration in the modulatory role of nitric oxide. N G-monomethyl- l-arginine ( l-NMMA) (1.0, 10 and 100 μM) produced dose-related vasoconstriction in nondiabetic, but not diabetic hamsters. Further, l-NMMA did not alter vasoconstriction in response to endothelin-1 in nondiabetic and diabetic hamsters. These findings suggest that diabetes alters constriction of cheek pouch resistance arterioles to endothelin-1 which appears to be independent of the synthesis/release of nitric oxide. In addition, based upon findings using l-NMMA, it appears that there is a reduced influence of nitric oxide on basal diameter of resistance arterioles during diabetes.

Superoxide Dismutase Restores Impaired Histamine‐Induced Increase in Venular Macromolecular Efflux During Diabetes Mellitus

The goal of this study was to determine the role of oxygen radicals in impaired histamine-induced increases in venular macromolecular efflux from the hamster cheek pouch. We used intravital fluorescent microscopy and fluorescein isothiocyanate dextran (FITC-dextran; MW = 70K) to examine macromolecular extravasation from post-capillary venules in nondiabetic and diabetic (2-4 weeks after injection of streptozotocin) hamsters in response to histamine. Increases in extravasation of macromolecules were quantitated by counting venular leaky sites and by calculating clearance (ml/s x 10(-6)) of FITC-dextran-70K. In nondiabetic hamsters, superfusion with histamine (1.0 and 5.0 microM) increased venular leaky sites from 0 +/- 0 to 17 +/- 6 and 35 +/- 6 per 0.11 cm2, respectively. In addition, clearance of FITC-dextran-70K increased during superfusion with histamine. In contrast, superfusion with histamine did not increase the formation of venular leaky sites (0 +/- 0) or clearance of FITC-dextran-70K in diabetic hamsters. Next, we examined whether alterations in histamine-induced increases in macromolecular efflux in diabetic hamsters may be related to the production of oxygen radicals. We examined whether exogenous application of superoxide dismutase (150 U/ml) could restore impaired histamine-induced increases in macromolecular extravasation in diabetic hamsters. Application of superoxide dismutase did not alter histamine-induced increases in venular leaky sites or clearance of FITC-dextran-70K in nondiabetic hamsters. However, application of superoxide dismutase restored histamine-induced increases in leaky site formation and clearance of FITC-dextran-70K in diabetic hamsters towards that observed in nondiabetic hamsters. These findings suggest that oxygen radical formation appears to contribute to impaired macromolecular efflux in response to histamine during short-term diabetes mellitus.

Superoxide Dismutase Restores Impaired Histamine-Induced Increase in Venular Macromolecular Efflux During Diabetes Mellitus

The goal of this study was to determine the role of oxygen radicals in impaired histamine-induced increases in venular macromolecular efflux from the hamster cheek pouch. We used intravital fluorescent microscopy and fluorescein isothio-cynate dextran (FITC-dextran; MW = 70 K) to examine macromolecular extravasation from post-capillary venules in nondiabetic and diabetic (2–4 weeks after injection of streptozoticin) hamsters in response to histamine. Increases in extravasation of macromolecules were quantitated by counting venular leaky sites and by calculating clearance (ml/s × 10−6) of FITC-dextran-70 K. In non-diabetic hamsters, superfusion with histamine (1.0 and 5.0 µM) increased venular leaky sites from 0 ± 0 to 17 ± 6 and 35 ± 6 per 0.11 cm2, respectively. In addition, clearance of FITC-dextran-70 K increased during superfusion with histamine. In contrast, superfusion with histamine did not increase the formation of venular leaky sites (0 ± 0) or clearance of FITC-dextran-70 K in diabetic hamsters. Next, we examined whether alterations in histamine-induced increases in macromolecular efflux in diabetic hamsters may be related to the production of oxygen radicals. We examined whether exogenous application of superoxide dismutase (150 U/ml) could restore impaired histamine-induced increases in macromolecular extravasation in diabetic hamsters. Application of superoxide dismutase did not alter histamine-induced increases in venular leaky sites or clearance of FITC-dextran-70 K in nondiabetic hamsters. However, application of superoxide dismutase restored histamine-induced increases in leaky site formation and clearance of FITC-dextran-70 K in diabetic hamsters towards that observed in nondiabetic hamsters. These findings suggest that oxygen radical formation appears to contribute to impaired macromolecular efflux in response to histamine during short-term diabetes mellitus.

L-arginine does not reverse impaired agonist-induced increases in macromolecular efflux during diabetes mellitus.

The first goal of this study was to determine the effect of diabetes mellitus on agonist-induced increases in venular macromolecular permeability of the hamster cheek pouch. The second goal was examine the role for an alteration in the availability of L-arginine to nitric oxide synthase in impaired agonist-induced increases in macromolecular permeability during diabetes. We used intravital fluorescent microscopy and fluorescein isothiocyanate dextran (FITC-dextran; mw = 70 K) to examine macromolecular extravasation from post-capillary venules in non-diabetic and diabetic (2 weeks after injection of streptozotocin) hamsters in response to histamine and substance P. Increases in extravasation of macromolecules were quantitated by counting the number of venular leaky sites and by calculating the clearance (ml/s x 10(-6)) of FITC-dextran-70 K. In non-diabetic hamsters, histamine (1.0 and 5.0 microM) and substance P (50 and 100 nM) increased permeability of the cheek pouch microcirculation to FITC-dextran-70 K. In contrast, histamine- and substance P-induced increases in macromolecular extravasation were markedly reduced in diabetic hamsters. Next, we investigated whether alterations in histamine- and substance P-induced changes in macromolecular extravasation in diabetic hamsters may be related to an alteration in the availability of L-arginine. We examined whether exogenous application of L-arginine (100 microM) could restore impaired histamine- and substance-P-induced increases in macromolecular extravasation in diabetic hamsters. We found that L-arginine potentiated agonist-induced increases in macromolecular extravasation in non-diabetic hamsters, but did not alter responses in diabetic hamsters. These findings suggest that short-term diabetes mellitus alters agonist-induced alterations in microvascular permeability. The mechanism of altered microvascular permeability during diabetes mellitus does not appear to be related to an impaired availability of L-arginine.

Effect of L-arginine on reactivity of hamster cheek pouch arterioles during diabetes mellitus.

The goal of this study was to determine whether exogenous application of L-arginine could restore impaired agonist-induced increases in arteriolar diameter during diabetes mellitus. We used intravital microscopy to examine reactivity of cheek pouch arterioles (50 microns in diameter) in nondiabetic and diabetic (2 weeks after injection of streptozotocin) hamsters in response to histamine and substance P. In nondiabetic hamsters histamine (1.0 and 5.0 microM) dilated cheek pouch arterioles by 15 +/- 1 and 22 +/- 1%, respectively, and substance P (50 and 100 nM) dilated arterioles by 14 +/- 3 and 21 +/- 4%, respectively. In addition, dilatation of arterioles in response to histamine and substance P in nondiabetic hamsters was abolished by application of an enzymatic inhibitor of nitric oxide synthase (L-NMMA). In contrast, histamine- and substance P-induced increases in arteriolar diameter were markedly reduced in diabetic hamsters. Histamine (1.0 and 5.0 microM) dilated arterioles by only 5 +/- 1 and 4 +/- 2%, respectively, and substance P (50 and 100 nM) dilated arterioles by only 6 +/- 2 and 5 +/- 3%, respectively (p < 0.05 vs. nondiabetic hamsters). Nitroglycerin produced similar vasodilatation in nondiabetic and diabetic hamsters. Next, we examined whether exogenous application of L-arginine (100 microM) could restore impaired histamine- and substance P-induced increases in arteriolar diameter in diabetic hamsters. We found that L-arginine did not restore altered nitric oxide synthase-dependent vasodilatation in diabetic hamsters. These findings suggest that short-term diabetes mellitus alters agonist-induced increases in arteriolar diameter. In addition, the mechanism of altered arteriolar reactivity during diabetes mellitus does not appear to be related to an impaired availability of L-arginine.

Loss of GLUT2 glucose transporter expression in pancreatic beta cells from diabetic Chinese hamsters

The diabetic Chinese hamster is a well-established animal model for NIDDM with a defective glucose-induced insulin secretory response. In the pancreas of nondiabetic hamsters, the GLUT2 glucose transporter was localized in the plasma membrane of insulin-positive beta cells. At variance with the rat, immunoreactivity was also detected in the cytoplasm. Other islet cell types were not GLUT2 positive. GLUT2 immunoreactivity was already significantly reduced in beta cells from mildly diabetic animals in spite of a normal insulin immunoreactivity. In severely diabetic animals the majority of the beta cells had lost GLUT2 immunostaining. This observation was confirmed in a Western blot analysis of the GLUT2 protein in isolated pancreatic islets. Only beta cells that were densely immunostained for insulin were still GLUT2 positive. However, around 40% of the beta cells devoid of GLUT2 immunoreactivity were still insulin immunoreactive. Thus, the loss of GLUT2 immunoreactivity, which is an important component of the glucose recognition apparatus of the pancreatic beta cell, is an early indicator of beta cell dysfunction before the development of degenerative lesions or the loss of insulin immunoreactivity. GLUT2 loss may be important in the deterioration of glucose-induced insulin secretion in the diabetic Chinese hamster.

Alterations in erythrocyte plasma membrane ATPase activity and adenine nucleotide content in a spontaneously diabetic subline of the Chinese hamster.

The CHIG/Han subline of the Chinese hamster develops noninsulin-dependent diabetes mellitus characterized by hyperinsulinemia and different degrees of glucose intolerance. To study whether these abnormalities could affect transmembrane cation transport activity, we determined membrane ATPase activity and ATP concentrations in red blood cells of diabetes-resistant CHIA and diabetes-susceptible CHIG sublines of the Chinese hamster. Mg(2+)-ATPase activity was increased in red blood cell membranes of diabetic hamsters compared with that of nondiabetic CHIG and the diabetes-resistant CHIA animals and correlated with plasma triglyceride and cholesterol levels. Ca(2+)-ATPase and Na+/K+ATPase activity were not significantly different between diabetic and nondiabetic hamsters, but for the Na+/K(+)-ATPase, Km was decreased and the Vmax value increased in membrane preparations from severely diabetic hamsters. Both ATP and ADP content were lower in erythrocytes from diabetic than nondiabetic hamsters. Independently of the levels of glycemia, AMP concentrations were higher in CHIG than in CHIA hamsters. While ATP/AMP ratios were found to be decreased in erythrocytes from diabetes-susceptible CHIG hamsters compared to the diabetes-resistant CHIA animals, they were significantly correlated with the levels of glycemia. Furthermore, the relationship between blood glucose levels and kidney weight in hamsters of the diabetes-susceptible CHIG subline was such, that severely hyperglycemic animals displayed the greatest increase in kidney wet weight. These results indicate that the progressive metabolic deterioration in the development of noninsulin-dependent diabetes is associated with significant changes in the activity and kinetic parameters of cellular ATPases which could probably indicate early membrane alterations which may eventually result in the late microangiopathic complications of diabetes.

Major plasma glycoproteins of diabetic and nondiabetic Chinese hamsters.

Differences were noted in the plasma glycoprotein electrophoretic profile of diabetic versus nondiabetic chinese hamster. The variant plasma proteins were identified by chromatographic and electrophoretic procedures as transferrin (Tf) and alpha 2 macroglobulin (alpha 2M). The transferrin variants were line dependent and not altered in streptozotesin induced diabetics. Alpha 2M patterns were changed to that of the diabetics after streptozotesin treatment, and therefore are metabolic variants.

Mutagenicity of oral hypoglycaemic agents.

Summary Oral hypoglycaemic agents, commonly used in the therapy of diabetes mellitus, weretested for mutagenicity in vivo and in vitro. All drugs were non-mutagenic in the Salmonella/microsome assay (with and without microsomal activation). Only chlorpropam.ide gave a positive reaction performing the sister chromatid exchange test in vivo. Analysing bone marrow chromosomes of diabetic and non-diabetic mice and hamsters it was possible to exclude the possibility that the diabetic state itself is the inducing factor.

The polyol pathway for glucose metabolism in tissues from normal, diabetic, and ketotic Chinese hamsters.

Increased activity of the polyol pathway for glucose metabolism and increased intracellular levels of glucose, polyol, and fructose have been implicated in the onset of chronic complications of diabetes in certain tissues where the transport of glucose is not insulin-dependent. Levels of all three carbohydrates were found to be significantly higher in sciatic nerves and lenses from diabetic and ketotic hamsters than in tissues from nondiabetic control animals (p<0.01). Glucose, polyol, and fructose levels were not significantly different in spinal cords or gastrocnemius muscles from diabetic and ketotic hamsters than in tissues from nondiabetic hamsters. There was a linear relationship between blood glucose levels and tissue levels of glucose (r=0.885), polyol (r= 0.797), fructose (r=0.886) and the total tissue concentration of all three substances (r=0.916) in hamster sciatic nerves. Similarly, a linear correlation between blood glucose levels and levels of glucose (r=0.934), polyol (r=0.784), fructose (r=0.866) and the total tissue concentration of the three carbohydrates (r= 0.915) in lenses was also established. The results of these studies provide evidence that glucose, polyol, and fructose levels are elevated in certain tissues from animals with spontaneous diabetes. The significance of these observations with respect to the pathogenesis of complications of diabetes remains to be determined.