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Changes in Isoprenaline-Induced Endothelium-Dependent and -Independent Relaxations of Aorta in Long-Term STZ-Diabetic Rats: Reversal Effect of Dietary Vitamin E

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Abstract

  • 1.

    The present study concerns in vitro isoprenaline (ISO)-induced relaxation of aortic rings of long-term streptozotocin (STZ)-diabetic and nondiabetic rats, both with and without dietary vitamin E supplementation.

  • 2.

    Incubation with propranolol, NG-nitro-L-arginine methyl ester and methylene blue, as well as absence of endothelium, all negatively affect the ISO-induced relaxations.

  • 3.

    Thiobarbituric acid reactivity levels used as an index of lipid peroxidation are elevated in the aorta by diabetes. Four months of STZ-diabetes results in a significant increase in the ISO-induced relaxations together with endothelial dysfunction in the rat aorta. Diabetes also causes the loss of vascular integrity.

  • 4.

    Dietary vitamin E supplementation during the last 2 months of diabetes allows normalization of the levels of lipid peroxides. This vitamin also completely reverses the increased sensitivity (pD2 value) of the aorta to ISO, whereas the maximum ISO-induced relaxations are partially restored after the treatment in diabetic rats.

  • 5.

    The results suggest that ISO-induced relaxation in the aorta partially depends on the intact endothelium and that the endothelium-dependent relaxant effect of ISO is mediated by endothelium-derived relaxing factor. Results also indicate that abnormal vascular reactivity and structure of the diabetic rat aorta may be related to the increased lipid peroxidation. In conclusion, vitamin E can protect the arterial wall from oxidative stress–induced injury associated with chronic STZ-diabetes and allows normalization of the response to ISO and the structure of the aorta in diabetic rats.

Introduction

Relaxation responses of the blood vessels to isoprenaline (ISO) are mediated by β-adrenoceptors on the smooth muscle and are conventionally endothelium independent (Heeson and De Mey 1990; Kukovetz et al., 1981). In this regard, Moncada et al. 1991found that ISO-induced relaxation in the rat aorta is neither endothelium dependent nor affected by inhibitors of NO synthase. Contrary to the expectation, some reports suggested that endothelium may play a facilitatory role in the relaxation response to β-adrenoceptor agonists. These include the demonstration that removal of the endothelium from the rat aorta (Kamata et al., 1989) and canine coronary arteries (Rubanyi and Vanhoutte 1985) attenuates ISO-induced relaxation. Similarly, it was demonstrated in the rat aorta that ISO-induced relaxations are inhibited by methylene blue (MB) or by hemoglobin (Grace et al., 1988), agents known to affect the action of NO (Martin et al., 1985). Other studies showed that ISO-induced relaxations are attenuated by NO synthase inhibitors in the rat aorta (Gray and Marshall 1991) and in rat mesenteric resistance arteries (Graves and Poston 1993). Interestingly, Gray and Marshall 1992showed that ISO-induced relaxations totally depend on the presence of intact endothelium in the rat thoracic aorta. Owing to the discrepancies among the previous reports, the first aim of the present study was to investigate whether intact endothelium does in fact mediate relaxations induced by ISO in the rat thoracic aorta.

Diabetes mellitus is known to produce alterations in the responsiveness of vascular smooth muscle to various vasoactive agents (Karasu and Altan 1993). This is an important reason for the development of cardiovascular disease associated with diabetes (Karasu and Altan 1993; MacLeod and McNeill 1985). Accordingly, most investigators have demonstrated that endothelial dysfunction accompanies the increased responsiveness of vessels to vasoconstrictor agents in experimental diabetic rats (Abiru et al., 1990; Karasu and Altan 1994). To our knowledge, there is one study, reported by Kamata et al., 1989, investigating the effects of chemically induced diabetes on the response of the aorta to β-adrenoceptor agonists. The second aim of the present study, therefore, was to determine the effects of long-term STZ-diabetes on ISO-induced relaxations.

On the other hand, the exaggerated free radical activity and increased lipid peroxidation with a reduction in plasma vitamin E content have been well documented in diabetes (Chisolm et al., 1992; Kunisaki et al., 1990). These mechanisms are central to the development of the diabetes-induced micro- and macroangiophatic diseases (Giugliano et al., 1995). We have consistently found that vitamin E treatment is beneficial in the protection of vascular (Karasu et al., 1997) and neuronal (Karasu et al., 1995) functions against streptozotocin (STZ)-diabetes–induced injury. However, the effectiveness of vitamin E supplementation on ISO-induced relaxations in diabetic rats has not been questioned. For this reason, a third aim of the present study was to investigate whether the long-term diabetes-caused changes in ISO-induced relaxations in the rat aorta can be reversed by the dietary supplementation of vitamin E as a strong inhibitor of lipid peroxidation (Esterbauer et al., 1991).

Section snippets

Experimental organization

The experiments were carried out on male albino rats with a starting weight of 200–250 g and an age range of 8–10 weeks. In the experiments, the animals were randomly assigned to four experimental groups. Age-matched nondiabetic rats were employed as onset control group. Nondiabetic control rats were fed a standard rat diet or a diet supplemented with vitamin E (dl-α-tocopheryl acetate 0.5% w/w) for 2 months. Diabetes was induced by a single intraperitoneal injection of STZ (60 mg/kg, freshly

Body weights and plasma glucose concentrations of rats, and TBARS levels of aortae

The general characteristics of rats are shown in Table 1. Diabetic rats gained no weight during the experiments, whereas nondiabetic control rats almost doubled in weight; vitamin E treatment had no effect on the body weight of diabetic rats. Four-month diabetes caused hyperglycemia. Vitamin E treatment of diabetic rats had no significant effects on the increased plasma glucose.

Thiobarbituric acid reactive substances (TBARS) levels, which were used as an index of lipid peroxidation, were

Discussion

The present study demonstrates that ISO causes relaxations in the rat aorta that partially depend on the presence of intact endothelium. These findings confirm the results of previous studies that ISO-induced relaxations are inhibited by mechanical removal of the endothelium (Grace et al., 1988; Kamata et al., 1989) or by pretreatment with either MB (Grace et al., 1988) or L-NAME (Gray and Marshall 1991) in the rat aorta. The magnitudes of the inhibitory effects of L-NAME, MB and absence of

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