Modification of high saturated fat diet with n-3 polyunsaturated fat improves glucose intolerance and vascular dysfunction.

AIMS: The ability of dietary enrichment with monounsaturated fatty acid (MUFA), n-3 or n-6 polyunsaturated fatty acids (PUFAs) to reverse glucose intolerance and vascular dysfunction resulting from excessive dietary saturated fatty acids is not resolved. We hypothesized that partial replacement of dietary saturated fats with n-3 PUFA-enriched menhaden oil (MO) would provide greater improvement in glucose tolerance and vascular function compared to n-6 enriched safflower oil (SO) or MUFA-enriched olive oil (OO). METHODS: We fed mice a high saturated fat diet (HF) (60% kcal from lard) for 12 weeks before substituting half the lard with MO, SO or OO for an additional 4 weeks. At the end of 4 weeks, we assessed glucose tolerance, insulin signalling and reactivity of isolated pressurized gracilis arteries. RESULTS: After 12 weeks of saturated fat diet, body weights were elevated and glucose tolerance was abnormal compared to mice on control diet (13% kcal lard). Diet substituted with MO restored basal glucose levels, glucose tolerance and indices of insulin signalling (phosphorylated Akt) to normal, whereas restoration was limited for SO and OO substitutions. Although dilation to acetylcholine was reduced in arteries from mice on HF, OO and SO diets compared to normal diet, dilation to acetylcholine was fully restored and constriction to phenylephrine was reduced in MO-fed mice compared to normal. CONCLUSION: We conclude that short-term enrichment of an ongoing high fat diet with n-3 PUFA rich MO, but not MUFA rich OO or n-6 PUFA rich SO, reverses glucose tolerance, insulin signalling and vascular dysfunction.

Treatment of Zucker diabetic fatty rats with AVE7688 improves vascular and neural dysfunction.

AIM: Vasopeptidase inhibitors are drugs that inhibit angiotensin-converting enzyme and neutral endopeptidase (NEP). The latter is a protease that degrades vasoactive peptides and is increased in diabetes. We have previously shown that treating streptozotocin-induced diabetic rats, an animal model of type 1 diabetes, with AVE7688, a vasopeptidase inhibitor, improves neurovascular and neural function. In this study, we determined the effect of treating Zucker diabetic fatty (ZDF) rats, an animal model of type 2 diabetes, with AVE7688 on vascular and neural function. METHODS: ZDF rats at 12 weeks of age were treated for 12 weeks with AVE7688 (500 mg/kg diet). Afterwards, vascular reactivity of epineurial arterioles of the sciatic nerve and nerve conduction velocity and blood flow was determined. RESULTS: Vascular and neural function was significantly impaired in ZDF rats compared with age-matched lean (control) rats. Treating ZDF rats with AVE7688 improved vascular relaxation to acetylcholine and calcitonin gene-related peptide in epineurial arterioles. Motor and sensory nerve conduction velocity, endoneurial blood flow and thermal nociception end-points were also improved by treatment compared with untreated ZDF rats. Superoxide and expression of NEP were increased in epineurial arterioles from ZDF rats and attenuated by treatment with AVE7688. CONCLUSIONS: AVE7688 is an effective treatment for microvascular and neural disease in ZDF rats. Thus, vasopeptidase inhibitors may be an effective treatment for diabetic microvascular and neural complication in type 2 diabetes.

Vascular and neural dysfunction in Zucker diabetic fatty rats: a difficult condition to reverse.

AIM: We had previously demonstrated that vascular and neural dysfunction in Zucker diabetic fatty (ZDF) rats is progressive. In this study, we sought to determine whether monotherapy of ZDF rats can reverse the vascular and nerve defects. METHODS: ZDF rats at 16 weeks of age were treated for 12 weeks with the angiotensin-converting enzyme inhibitor enalapril, the antioxidant alpha-lipoic acid, the HMG-CoA reductase inhibitor rosuvastatin or the PPARgamma agonist rosiglitazone. Vasodilation of epineurial arterioles was measured by videomicroscopy. Endoneurial blood flow (EBF) was measured by hydrogen clearance, and nerve conduction velocity was measured following electrical stimulation of motor or sensory nerves. RESULTS: Motor nerve conduction velocity (MNCV), sensory nerve conduction velocity (SNCV) (70 and 77% of control, respectively), EBF (64% of control), and vascular relaxation in response to acetylcholine (50% of control) and calcitonin gene-related peptide (CGRP; 73% of control) are impaired in ZDF rats at 28 weeks of age compared with lean littermate controls. Treatment with enalapril and alpha-lipoic acid attenuated the decrease in MNCV and SNCV. Enalapril, alpha-lipoic acid and rosiglitazone treatment of ZDF rats were partially effective in improving endothelium-dependent vascular dysfunction as measured by vascular relaxation in response to acetylcholine. The same drugs also attenuated the decrease in EBF. However, impairment in vascular relaxation in response to CGRP was improved with only alpha-lipoic acid or rosuvastatin treatment. The increase in superoxide and nitrotyrosine levels in vascular tissue was attenuated by all treatments. CONCLUSIONS: The efficacy of monotherapy treatment of ZDF rats using different classes of drugs for vascular and neural dysfunction once complications have developed did not achieve expected levels. This could be because of the complex aetiology of vascular and neural disease in type 2 diabetes.

Effect of antioxidant treatment of streptozotocin-induced diabetic rats on endoneurial blood flow, motor nerve conduction velocity, and vascular reactivity of epineurial arterioles of the sciatic nerve.

We have shown that diabetes-induced reduction in endoneurial blood flow (EBF) and impaired endothelium-dependent vascular relaxation precede slowing of motor nerve conduction velocity (MNCV) and decreased sciatic nerve Na(+)/K(+) ATPase activity. Furthermore, vascular dysfunction was accompanied by an accumulation of superoxide in arterioles that provide circulation to the sciatic nerve. In the present study, we examined the effect that treatment of streptozotocin-induced diabetic rats with antioxidants has on vascular and neural function. Diabetic rats were treated with 0.5% alpha-lipoic acid as a diet supplement or with hydroxyethyl starch deferoxamine (HES-DFO) by weekly intravenous injections at a dose of 75 mg/kg. The treatments significantly improved diabetes-induced decrease in EBF, acetylcholine-mediated vascular relaxation in arterioles that provide circulation to the region of the sciatic nerve, and MNCV. The treatments also reduced the production of superoxide by the aorta and superoxide and peroxynitrite by arterioles that provide circulation to the region of the sciatic nerve. Treating diabetic rats with alpha-lipoic acid prevented the diabetes-induced increase in thiobarbituric acid-reactive substances in serum and significantly improved lens glutathione levels. In contrast, treating diabetic rats with HES-DFO did not prevent diabetes-induced changes of either of these markers of oxidative stress. Diabetes-induced increase in sciatic nerve conjugated diene levels was not improved by treatment with either alpha-lipoic acid or HES-DFO. Treating diabetic rats with alpha-lipoic acid but not HES-DFO partially improved sciatic nerve Na(+)/K(+) ATPase activity and myo-inositol content. The increase in sciatic nerve sorbitol levels in diabetic rats was unchanged by either treatment. These studies suggest that diabetes-induced oxidative stress and the generation of superoxide may be partially responsible for the development of diabetic vascular and neural complications.

Acetylcholine-induced arteriolar dilation is reduced in streptozotocin-induced diabetic rats with motor nerve dysfunction.

1. Diabetes mellitus produces marked abnormalities in motor nerve conduction, but the mechanism is not clear. In the present study we hypothesized that in the streptozotocin (STZ)-induced diabetic rat impaired vasodilator function is associated with reduced endoneural blood flow (EBF) which may contribute to nerve dysfunction. 2. We examined whether diabetes-induced reductions in sciatic nerve conduction velocity and EBF were associated with impaired endothelium-dependent dilation in adjacent arterioles. We measured motor nerve conduction velocity (MNCV) in the sciatic nerve using a non-invasive procedure, and sciatic nerve nutritive blood flow using microelectrode polarography and hydrogen clearance. In vitro videomicroscopy was used to quantify arteriolar diameter responses to dilator agonists in arterioles overlying the sciatic nerve. 3. MNCV and EBF in 4-week-STZ-induced diabetic rats were decreased by 22% and 49% respectively. Arterioles were constricted with U46619 and dilation to acetylcholine (ACh), aprikalim, or sodium nitroprusside (SNP) examined. All agonists elicited dose-dependent dilation in control and diabetic rats, although ACh-induced dilation was significantly reduced in diabetic rats. Treating vessels from normal or diabetic rats with indomethacin (INDO) alone did not significantly affect ACh-induced relaxation. However, ACh-induced vasodilation was significantly reduced by treatment with KCl or Nomega-nitro-L-arginine (LNNA) alone. Combining LNNA and KCl further reduced ACh-induced dilation in these vessels. 4. Diabetes causes vasodilator dysfunction in a microvascular bed that provides circulation to the sciatic nerve. These studies imply that ACh-induced dilation in these vessels is mediated by multiple mechanisms that may include the endothelial-dependent production of nitric oxide and endothelial-derived hyperpolarizing factor. This impaired vascular response is associated with neural dysfunction.

Effect of M40403 treatment of diabetic rats on endoneurial blood flow, motor nerve conduction velocity and vascular function of epineurial arterioles of the sciatic nerve.

1. To further explore the effect of antioxidants in preventing diabetes-induced vascular and neural dysfunction we treated streptozotocin-induced diabetic rats daily with subcutaneous injections of 10 mg kg(-1) of M40403 (n=11) and compared the results obtained from 17 control rats and 14 untreated diabetic rats. M40403 is a manganese(II) complex with a bis(cyclo-hexylpyridine)-substituted macrocyclic ligand that was designed to be a selective functional mimetic of superoxide dismutase. Thus, M40403 provides a useful tool to evaluate the roles of superoxide in disease states. 2. Treatment with M40403 significantly improved diabetes-induced decrease in endoneurial blood flow, acetylcholine-mediated vascular relaxation in arterioles that provide circulation to the region of the sciatic nerve, and motor nerve conduction velocity (P<0.05). M40403 treatment also reduced the appearance of superoxide in the aorta and epineurial vessels and peroxynitrite in epineurial vessels. Treating diabetic rats with M40403 reduced the diabetes-induced increase in thiobarbituric acid reactive substances in serum but did not prevent the decrease in lens glutathione level. Treating diabetic rats with M40403 did not improve sciatic nerve Na(+)/K(+) ATPase activity or the sorbitol, fructose or myo-inositol content of the sciatic nerve. 3. These studies provide additional evidence that diabetes-induced oxidative stress and the generation of superoxide and perhaps peroxynitrite may be partially responsible for the development of diabetic vascular and neural complications.

Sensory nerve innervation of epineurial arterioles of the sciatic nerve containing calcitonin gene-related peptide: effect of streptozotocin-induced diabetes.

The authors have determined that epineurial arterioles of the sciatic nerve are innervated by nonadrenergic, noncholinergic nerves that contribute to the regulation of vasodilation. Using immunohistochemistry, the authors determined that nerves innervating epineurial arterioles contain the neuropeptide calcitonin gene-related peptide (CGRP). Using streptozotocin-induced diabetic rats, the authors demonstrated that CGRP content in sensory nerves innervating epineurial arterioles and vasodilation in response to exogenous CGRP was decreased. In summary, epineurial arterioles of the sciatic nerve are innervated by sensory nerves containing the neuropeptide CGRP. The diabetes-like condition induced by streptozotocin reduces the content of CGRP in these nerves and exogenous CGRP-mediated vasodilation. CGRP is likely an important regulator of vascular tone and compromising its function could contribute to nerve ischemia and diabetic neuropathy.

Slowing of motor nerve conduction velocity in streptozotocin-induced diabetic rats is preceded by impaired vasodilation in arterioles that overlie the sciatic nerve.

Diabetes mellitus produces marked abnormalities in motor nerve conduction, but the mechanism is not clear. In the present study we hypothesized that in the streptozotocin (STZ)-induced diabetic rat impaired vasodilator function in arterioles that provide circulation to the region of the sciatic nerve is associated with reduced endoneural blood flow (EBF) and that these defects precede slowing of motor nerve conduction velocity, and thereby may contribute to nerve dysfunction. As early as three days after the induction of diabetes endoneural blood flow was reduced in the STZ-induced diabetic rat. Furthermore, after 1 week of diabetes acetylcholine-induced vasodilation was found to be impaired. This was accompanied by an increase in the superoxide level in arterioles that provide circulation to the region of the sciatic nerve as well as changes in the level of other markers of oxidative stress including an increase in serum levels of thiobarbituric acid reactive substances and a decrease in lens glutathione level. In contrast to the vascular related changes that occur within 1 week of diabetes, motor nerve conduction velocity and sciatic nerve Na+/K+ ATPase activity were significantly reduced following 2 and 4 weeks of diabetes, respectively. These studies demonstrate that changes in vascular function in the STZ-induced diabetic rat precede the slowing of motor nerve conduction velocity (MNCV) and are accompanied by an increase in superoxide levels in arterioles that provide circulation to the region of the sciatic nerve.

Effect of fidarestat and alpha-lipoic acid on diabetes-induced epineurial arteriole vascular dysfunction.

In the present study, the authors examined whether treating streptozotocin-induced diabetic rats with the combination of alpha-lipoic acid and fidarestat, an aldose reductase inhibitor, can promote the formation of dihydrolipoic acid in diabetic animals and thereby enhance the efficacy of alpha-lipoic acid as monotherapy toward preventing diabetic vascular and neural dysfunction. Treating diabetic rats with the combination of 0.25% alpha-lipoic acid (in the diet) and fidarestat (3 mg/kg body weight) prevented the diabetes-induced slowing of motor nerve conduction velocity and endoneurial blood flow. This therapy also significantly improved acetylcholine-mediated vasodilation in epineurial arterioles of the sciatic nerve compared to nontreated diabetic rats. Treating diabetic rats with 0.25% alpha-lipoic acid and fidarestat (3 mg/kg body weight) was equally or more effective in preventing vascular and neural dysfunction than was monotherapy of diabetic rats with higher doses of alpha-lipoic acid or fidarestat. Treating diabetic rats with the combination of 0.25% alpha-lipoic acid and fidarestat (3 mg/kg body weight) significantly improved several markers of oxidative stress and increased the serum levels of both alpha-lipoic acid and dihydrolipoic acid. These studies suggest that combination therapy consisting of alpha-lipoic acid and fidarestat may be more efficacious in preventing diabetes-induced vascular and neural dysfunction in peripheral tissue compared to monotherapy, which requires higher doses to be equally effective. The effect of this combination therapy may in part be due to the increased production and/or level of dihydrolipoic acid.

Role of the carrier solution in cyclosporine pharmacokinetics in the baboon.

Although numerous investigational models have demonstrated the potent immunosuppressive properties of cyclosporine, the effectiveness of any given dosage may vary with the metabolism of the animal, the route of administration, and the carrier solution of the drug. We investigated the pharmacokinetics of intramuscular cyclosporine administration in the baboon using three carriers: polyoxethylated castor oil (Cremophor), a mixture of octanoic and decanoic acids (Miglyol), and olive oil. Cyclosporine prepared in Cremophor, Miglyol, or olive oil was injected intramuscularly into the hindlegs of baboons. Specimens for cyclosporine assay were obtained 2, 4, 6, 12, 18, and 24 hours after single intramuscular injection of 10 mg/kg or 15 mg/kg. In addition, weekly, then monthly, levels were obtained on animals receiving daily intramuscular injections following heterotopic heart xenografts. Attempts at oral administration proved unreliable and were discontinued. Cyclosporine assay was performed on stored serum using the RIA-KIT (Sandoz Pharmaceuticals Corporation, East Hanover, N.J.). Cremophor provides a more bioavailable form of cyclosporine than Miglyol when administered intramuscularly. (Area under curve = 7776 +/- 1437 for Cremophor 15 mg/kg vs 1837 +/- 726 for Miglyol 15 mg/kg; 2579 +/- 694 for Cremophor 10 vs 1123 +/- 393 for Miglyol 10.) Long-term daily intramuscular administration of Cremophor provides a sustained drug serum trough level with wide variability between individual animals (80 to 825 ng/ml). Toxicity was limited to injection site inflammation. There was no biochemical evidence of renal toxicity; however, some animals did demonstrate early histologic changes of cyclosporine effect.(ABSTRACT TRUNCATED AT 250 WORDS)