Heart rate-associated mechanical stress impairs carotid but not cerebral artery compliance in dyslipidemic atherosclerotic mice

The cardiac cycle imposes a mechanical stress that dilates elastic carotid arteries, while shear stress largely contributes to the endothelium-dependent dilation of downstream cerebral arteries. In the presence of dyslipidemia, carotid arteries stiffen while the endothelial function declines. We reasoned that stiffening of carotid arteries would be prevented by reducing resting heart rate (HR), while improving the endothelial function would regulate cerebral artery compliance and function. Thus we treated or not 3-mo-old male atherosclerotic mice (ATX; LDLr(-/-):hApoB(+/+)) for 3 mo with the sinoatrial pacemaker current inhibitor ivabradine (IVA), the β-blocker metoprolol (METO), or subjected mice to voluntary physical training (PT). Arterial (carotid and cerebral artery) compliance and endothelium-dependent flow-mediated cerebral dilation were measured in isolated pressurized arteries. IVA and METO similarly reduced (P < 0.05) 24-h HR by ≈15%, while PT had no impact. As expected, carotid artery stiffness increased (P < 0.05) in ATX mice compared with wild-type mice, while cerebral artery stiffness decreased (P < 0.05); this paradoxical increase in cerebrovascular compliance was associated with endothelial dysfunction and an augmented metalloproteinase-9 (MMP-9) activity (P < 0.05), without changing the lipid composition of the wall. Reducing HR (IVA and METO) limited carotid artery stiffening, but plaque progression was prevented by IVA only. In contrast, IVA maintained and PT improved cerebral endothelial nitric oxide synthase-dependent flow-mediated dilation and wall compliance, and both interventions reduced MMP-9 activity (P < 0.05); METO worsened endothelial dysfunction and compliance and did not reduce MMP-9 activity. In conclusion, HR-dependent mechanical stress contributes to carotid artery wall stiffening in severely dyslipidemic mice while cerebrovascular compliance is mostly regulated by the endothelium.     

Selective cerebral vascular dysfunction in Mn-SOD-deficient mice.

We tested the hypothesis that the mitochondrial form of superoxide dismutase [manganese superoxide dismutase (Mn-SOD)] protects the cerebral vasculature. Basilar arteries (baseline diameter approximately 140 microm) from mice were isolated, cannulated, and pressurized to measure vessel diameter. In arteries from C57BL/6 mice preconstricted with U-46619, acetylcholine (ACh; an endothelium-dependent vasodilator) produced dilation that was similar in male and female mice and abolished by an inhibitor of nitric oxide synthase. Vasodilation to ACh was not altered in heterozygous male or female Mn-SOD-deficient (Mn-SOD+/-) mice compared with wild-type littermate controls (Mn-SOD+/+). Constriction of the basilar artery to arginine vasopressin, but not KCl or U-46619, was increased in Mn-SOD+/- mice (P<0.05), and this effect was prevented by tempol, a scavenger of superoxide. We also examined responses of cerebral (pial) arterioles (branches of the middle cerebral artery, control diameter approximately 30 microm) to ACh in anesthetized mice using a cranial window. Responses to ACh, but not nitroprusside (an endothelium-independent agonist), were reduced (P<0.05) in cerebral arterioles in Mn-SOD+/- mice, and this effect was prevented by tempol. Thus these are the first data on the role of Mn-SOD in cerebral circulation. In the basilar artery, ACh produced nitric oxide-mediated dilation that was similar in male and female mice. Under normal conditions in cerebral arteries, responses to ACh were not altered but constrictor responses were selectively enhanced in Mn-SOD+/- mice. In the cerebral microcirculation, there was superoxide-mediated impairment of responses to ACh.     

Effects of acute Rho kinase inhibition on chronic hypoxia-induced changes in proximal and distal pulmonary arterial structure and function

Hypoxic pulmonary hypertension (HPH) is initially a disease of the small pulmonary arteries. Its severity is usually quantified by pulmonary vascular resistance (PVR). Acute Rho kinase inhibition has been found to reduce PVR toward control values in animal models, suggesting that persistent pulmonary vasoconstriction is the dominant mechanism for increased PVR. However, HPH may also cause proximal arterial changes, which are relevant to right ventricular (RV) afterload. RV afterload can be quantified by pulmonary vascular impedance, which is obtained via spectral analysis of pulsatile pressure-flow relationships. To determine the effects of HPH independent of persistent pulmonary vasoconstriction in proximal and distal arteries, we quantified pulsatile pressure-flow relationships before and after acute Rho kinase inhibition and measured pulmonary arterial structure with microcomputed tomography. In control lungs, Rho kinase inhibition decreased 0 Hz impedance (Z?), which is equivalent to PVR, from 2.1 ± 0.4 to 1.5 ± 0.2 mmHg·min·ml?1 (P < 0.05) and tended to increase characteristic impedance (Z(C)) from 0.21 ± 0.01 to 0.22 ± 0.01 mmHg·min·ml?1. In HPH lungs, Rho kinase inhibition decreased Z? (P < 0.05) without affecting Z(C). Microcomputed tomography measurements performed on lungs after acute Rho kinase inhibition demonstrated that HPH significantly decreased the unstressed diameter of the main pulmonary artery (760 ± 60 vs. 650 ± 80 μm; P < 0.05), decreased right pulmonary artery compliance, and reduced the frequency of arteries of diameter 50-100 μm (both P < 0.05). These results demonstrate that acute Rho kinase inhibition reverses many but not all HPH-induced changes in distal pulmonary arteries but does not affect HPH-induced changes in the conduit arteries that impact RV afterload.     

Peroxisome proliferator-activated receptor-γ protects against vascular aging

Vascular disease occurs commonly during aging. Carotid artery and cerebrovascular disease are major causes of stroke and contributors to dementia. Recent evidence suggests that peroxisome proliferator-activated receptor-γ (PPARγ) may play a protective role in the vasculature, but the potential importance of PPARγ in vascular aging is unknown. To examine the hypothesis that PPARγ normally protects against vascular aging, we studied heterozygous knockin mice expressing a human dominant-negative mutation in PPARγ (P465L, designated L/+). Endothelial dysfunction, a major contributor to vascular disease, was studied using carotid arteries from adult (8 ± 1 mo) and old (24 ± 1 mo) L/+ mice and wild-type littermates. In arteries from wild-type mice, responses to the endothelium-dependent agonist ACh were similar in adult and old wild-type mice but were reduced by ～50% in old L/+ mice (n = 7-10, P < 0.05). Impaired responses in arteries from old L/+ mice were restored to normal by a scavenger of superoxide. Relaxation of arteries to nitroprusside (an NO donor) was similar in all groups. Contraction of arteries to U46619 was not affected by age or genotype, while maximal responses to endothelin-1 were reduced with age in both wild-type and L/+ mice. Vascular expression (mRNA) of the catalytic component of NADPH oxidase (Nox2) was not altered in wild-type mice but was increased significantly in old L/+ mice. These findings provide the first evidence that interference with PPARγ function accelerates vascular aging, suggesting a novel role for PPARγ in protecting against age-induced oxidative stress and endothelial dysfunction.     

Effect of chronic endothelin receptor antagonism on cerebrovascular function in type 2 diabetes

Diabetes increases the risk of stroke and contributes to poor clinical outcomes in this patient population. Myogenic tone of the cerebral vasculature, including basilar arteries, plays a key role in controlling cerebral blood flow. Increased myogenic tone is ameliorated with ET receptor antagonism in Type 1 diabetes. However, the role of endothelin-1 (ET-1) and its receptors in cerebrovascular dysfunction in Type 2 diabetes, a common comorbidity in stroke patients, remains poorly elucidated. Therefore, we hypothesized that 1) cerebrovascular dysfunction occurs in the Goto-Kakizaki (GK) model of Type 2 diabetes, and 2) pharmacological antagonism of ETA receptors ameliorates, while ETB receptor blockade augments vascular dysfunction. GK or control rats were treated with antagonists to either ETA (atrasentan, 5 mg.kg(-1).day(-1)) or ETB (A-192621, 15 or 30 mg.kg(-1).day(-1)) receptors for 4 wk and vascular function of basilar arteries was assessed using a wire myograph. GK rats exhibited increased sensitivity to ET-1. ET(A) receptor antagonism caused a rightward shift, indicating decreased sensitivity in diabetes, while it increased sensitivity to ET-1 in control rats. Endothelium-dependent relaxation was impaired in diabetes. ETA receptor blockade restored relaxation to control values in the GK animals with no significant effect in Wistar rats and ETB blockade with 30 mg.kg(-1).day(-1) A-192621 caused paradoxical constriction in diabetes. These studies demonstrate that cerebrovascular dysfunction occurs and may contribute to altered regulation of myogenic tone and cerebral blood flow in diabetes. While ETA receptors mediate vascular dysfunction, ETB receptors display differential effects. These results underscore the importance of ETA/ETB receptor balance and interactions in cerebrovascular dysfunction in diabetes.     

Nitrite supplementation reverses vascular endothelial dysfunction and large elastic artery stiffness with aging

We tested the hypothesis that short-term nitrite therapy reverses vascular endothelial dysfunction and large elastic artery stiffening with aging, and reduces arterial oxidative stress and inflammation. Nitrite concentrations were lower (P < 0.05) in arteries, heart, and plasma of old (26-28 month) male C57BL6 control mice, and 3 weeks of sodium nitrite (50 mg L(-1) in drinking water) restored nitrite levels to or above young (4-6 month) controls. Isolated carotid arteries of old control mice had lower acetylcholine (ACh)-induced endothelium-dependent dilation (EDD) (71.7 ± 6.1% vs. 93.0 ± 2.0%) mediated by reduced nitric oxide (NO) bioavailability (P < 0.05 vs. young), and sodium nitrite restored EDD (95.5 ± 1.6%) by increasing NO bioavailability. 4-Hydroxy-2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPOL), a superoxide dismutase (SOD) mimetic, apocynin, a nicotinamide adenine dinucleotide phosphate-oxidase (NADPH) inhibitor, and sepiapterin (exogenous tetrahydrobiopterin) each restored EDD to ACh in old control, but had no effect in old nitrite-supplemented mice. Old control mice had increased aortic pulse wave velocity (478 ± 16 vs. 332 ± 12 AU, P < 0.05 vs. young), which nitrite supplementation lowered (384 ± 27 AU). Nitrotyrosine, superoxide production, and expression of NADPH oxidase were ～100-300% greater and SOD activity was ～50% lower in old control mice (all P < 0.05 vs. young), but were ameliorated by sodium nitrite treatment. Inflammatory cytokines were markedly increased in old control mice (P < 0.05), but reduced to levels of young controls with nitrite supplementation. Short-term nitrite therapy reverses age-associated vascular endothelial dysfunction, large elastic artery stiffness, oxidative stress, and inflammation. Sodium nitrite may be a novel therapy for treating arterial aging in humans.     

Tumor necrosis factor-alpha impairs contraction but not relaxation in carotid arteries from iNOS-deficient mice

We used mice deficient in expression of inducible NO synthase (iNOS -/-) to directly examine the role of iNOS in impaired vasoconstrictor responses following tumor necrosis factor-alpha (TNF-alpha). In iNOS +/+ mice, contraction of carotid arteries in response to prostaglandin F(2alpha) (PGF(2alpha)) was impaired following TNF-alpha (100 microg/kg ip)(n = 10, P < 0.01). In contrast to responses in wild-type mice, contraction to low concentrations of PGF(2alpha) were normal, but maximum contraction to PGF(2alpha) was impaired in arteries from iNOS -/- mice treated with TNF-alpha [0.35 +/-.0.02 g (n = 8) following vehicle and 0.25 +/- 0.02 g (n = 7) following TNF-alpha (P < 0.05)]. Aminoguanidine, a relatively selective inhibitor of iNOS, partially restored contraction to PGF(2alpha) in vessels from iNOS +/+ mice but had no effect in iNOS -/- mice injected with TNF-alpha, suggesting that a mechanism(s) other than iNOS contributes to impaired responses. In contrast to contractile responses, relaxation of the carotid artery in response to acetylcholine and nitroprusside was not altered following TNF-alpha in iNOS +/+ or iNOS -/-mice. Responses of carotid arteries from iNOS -/- mice and effects of aminoguanidine suggest that both iNOS-dependent and iNOS-independent mechanisms contribute to impaired contractile responses following TNF-alpha.     

Statins and Selective Inhibition of Rho Kinase Protect Small Conductance Calcium-Activated Potassium Channel Function (KCa2.3) in Cerebral Arteries

Background

In rat middle cerebral and mesenteric arteries the K_Ca2.3 component of endothelium-dependent hyperpolarization (EDH) is lost following stimulation of thromboxane (TP) receptors, an effect that may contribute to the endothelial dysfunction associated with cardiovascular disease. In cerebral arteries, K_Ca2.3 loss is associated with NO synthase inhibition, but is restored if TP receptors are blocked. The Rho/Rho kinase pathway is central for TP signalling and statins indirectly inhibit this pathway. The possibility that Rho kinase inhibition and statins sustain K_Ca2.3 hyperpolarization was investigated in rat middle cerebral arteries (MCA).

Methods

MCAs were mounted in a wire myograph. The PAR2 agonist, SLIGRL was used to stimulate EDH responses, assessed by simultaneous measurement of smooth muscle membrane potential and tension. TP expression was assessed with rt-PCR and immunofluorescence.

Results

Immunofluorescence detected TP in the endothelial cell layer of MCA. Vasoconstriction to the TP agonist, U46619 was reduced by Rho kinase inhibition. TP receptor stimulation lead to loss of K_Ca2.3 mediated hyperpolarization, an effect that was reversed by Rho kinase inhibitors or simvastatin. K_Ca2.3 activity was lost in L-NAME-treated arteries, but was restored by Rho kinase inhibition or statin treatment. The restorative effect of simvastatin was blocked after incubation with geranylgeranyl-pyrophosphate to circumvent loss of isoprenylation.

Conclusions

Rho/Rho kinase signalling following TP stimulation and L-NAME regulates endothelial cell K_Ca2.3 function. The ability of statins to prevent isoprenylation and perhaps inhibit of Rho restores/protects the input of K_Ca2.3 to EDH in the MCA, and represents a beneficial pleiotropic effect of statin treatment.     

Protective Vascular and Cardiac Effects of Inducible Nitric Oxide Synthase in Mice with Hyperhomocysteinemia

Diet-induced hyperhomocysteinemia produces endothelial and cardiac dysfunction and promotes thrombosis through a mechanism proposed to involve oxidative stress. Inducible nitric oxide synthase (iNOS) is upregulated in hyperhomocysteinemia and can generate superoxide. We therefore tested the hypothesis that iNOS mediates the adverse oxidative, vascular, thrombotic, and cardiac effects of hyperhomocysteinemia. Mice deficient in iNOS (Nos2−/−) and their wild-type (Nos2+/+) littermates were fed a high methionine/low folate (HM/LF) diet to induce mild hyperhomocysteinemia, with a 2-fold increase in plasma total homocysteine (P<0.001 vs. control diet). Hyperhomocysteinemic Nos2+/+ mice exhibited endothelial dysfunction in cerebral arterioles, with impaired dilatation to acetylcholine but not nitroprusside, and enhanced susceptibility to carotid artery thrombosis, with shortened times to occlusion following photochemical injury (P<0.05 vs. control diet). Nos2−/− mice had decreased rather than increased dilatation responses to acetylcholine (P<0.05 vs. Nos2+/+ mice). Nos2−/− mice fed control diet also exhibited shortened times to thrombotic occlusion (P<0.05 vs. Nos2+/+ mice), and iNOS deficiency failed to protect from endothelial dysfunction or accelerated thrombosis in mice with hyperhomocysteinemia. Deficiency of iNOS did not alter myocardial infarct size in mice fed the control diet but significantly increased infarct size and cardiac superoxide production in mice fed the HM/LF diet (P<0.05 vs. Nos2+/+ mice). These findings suggest that endogenous iNOS protects from, rather than exacerbates, endothelial dysfunction, thrombosis, and hyperhomocysteinemia-associated myocardial ischemia-reperfusion injury. In the setting of mild hyperhomocysteinemia, iNOS functions to blunt cardiac oxidative stress rather than functioning as a source of superoxide.     

Endothelial dysfunction in Type 2 diabetes correlates with deregulated expression of the tail-anchored membrane protein SLMAP

The Type 2 diabetic db/db mouse experiences vascular dysfunction typified by changes in the contraction and relaxation profiles of small mesenteric arteries (SMAs). Contractions of SMAs from the db/db mouse to the alpha1-adrenoceptor agonist phenylephrine (PE) were significantly enhanced, and acetylcholine (ACh)-induced relaxations were significantly depressed. Drug treatment of db/db mice with a nonthiazolidinedione peroxisome prolifetor-activated receptor-gamma agonist and insulin sensitizing agent 2-[2-(4-phenoxy-2-propylphenoxy)ethyl]indole-5-acetic acid (COOH) completely prevented the changes in endothelium-dependent relaxation, but, with the discontinuation of therapy, endothelial dysfunction returned. Dysfunctional SMAs were found to specifically upregulate the expression of a 35-kDa isoform of sarcolemmal membrane-associated protein (SLMAP), which is a component of the excitation-contraction coupling apparatus and implicated in the regulation of membrane function in muscle cells. Real-time PCR revealed high SLMAP mRNA levels in the db/db microvasculature, which were markedly downregulated during COOH treatment but elevated again when drug therapy was discontinued. These data reveal that the microvasculature in db/db mice undergoes significant changes in vascular function with the endothelial component of vascular dysfunction specifically correlating with the overexpression of SLMAP. Thus changes in SLMAP expression may be an important indicator for microvascular disease associated with Type 2 diabetes.     

Impaired nitric oxide synthase-dependent dilatation of cerebral arterioles in type II diabetic rats

The goals of this study were to determine the effects of type II diabetes mellitus on nitric oxide synthase-dependent responses of cerebral arterioles and on endothelial nitric oxide synthase (eNOS) protein in cerebral arterioles. We examined dilatation of cerebral (pial) arterioles in 13-15 week old male lean and diabetic obese Zucker rats in response to nitric oxide synthase-dependent agonists (acetylcholine and adenosine diphosphate (ADP)) and a nitric oxide synthase-independent agonist (nitroglycerin). We found that acetylcholine (10 microM) increased cerebral arteriolar diameter by 10 +/- 3% (mean +/- SE) in lean Zucker rats, but by only 2 +/- 2% in diabetic obese Zucker rats (p<0.05). In addition, ADP (100 microM) increased cerebral arteriolar diameter by 20 +/- 2% in lean Zucker rats, but by only 8 +/- 2% in diabetic obese Zucker rats (p<0.05). In contrast, nitroglycerin produced similar vasodilatation in lean and diabetic obese Zucker rats. Thus, impaired dilatation of cerebral arterioles in diabetic obese Zucker rats is not related to non-specific impairment of vasodilatation. Following these functional studies, we harvested cerebral microvessels for Western blot analysis of eNOS protein. We found that eNOS protein was significantly higher in diabetic obese Zucker rats than in lean Zucker rats (p<0.05). Thus, type II diabetes mellitus impairs nitric oxide synthase-dependent responses of cerebral arterioles. In addition, eNOS protein from cerebral blood vessels is increased in diabetic obese Zucker rats.     

PPARgamma activation, by reducing oxidative stress, increases NO bioavailability in coronary arterioles of mice with Type 2 diabetes

We tested the hypothesis that short-term treatment of mice with Type 2 diabetes mellitus (DM) with rosiglitazone (ROSI), an agonist of peroxisome proliferator-activated receptor-gamma, ameliorates the impaired coronary arteriolar dilation by reducing oxidative stress via a mechanism unrelated to its effect on hyperglycemia and hyperinsulinemia. Control and Type 2 DM (db/db) mice were treated with ROSI (3 mg x kg(-1) x day(-1)) for 7 days, which did not significantly affect their serum concentration of glucose and insulin. Compared with controls, in db/db mice serum levels of 8-isoprostane and dihydroethydine-detectable superoxide production in carotid arteries were significantly elevated and were reduced by ROSI treatment. In coronary arterioles (diameter, approximately 80 microm) isolated from db/db mice, the reduced dilations to ACh, the nitric oxide (NO) donor NONOate, and increases in flow were significantly augmented either by in vitro administration of apocynin, an inhibitor of NAD(P)H-oxidase, or by in vivo ROSI treatment, responses that were then significantly reduced by the NO synthase inhibitor N(omega)-nitro-L-arginine methyl ester. In aortas of db/db mice, activity of SOD and catalase was reduced, whereas NAD(P)H oxidase activity was enhanced. ROSI treatment enhanced catalase and reduced NAD(P)H oxidase activity but did not affect the activity of SOD. These findings suggest that ROSI treatment enhances NO mediation of coronary arteriolar dilations due to the reduction of vascular NAD(P)H oxidase-derived superoxide production and enhancement of catalase activity. Thus, in addition to the previously revealed beneficial metabolic effects, the antioxidant action of rosiglitazone may protect coronary arteriolar function in Type 2 DM.     

H2O2 increases production of constrictor prostaglandins in smooth muscle leading to enhanced arteriolar tone in Type 2 diabetic mice

Our previous study showed that arteriolar tone is enhanced in Type 2 diabetes mellitus (T2-DM) due to an increased level of constrictor prostaglandins. We hypothesized that, in mice with T2-DM, hydrogen peroxide (H(2)O(2)) is involved in the increased synthesis of constrictor prostaglandins, hence enhanced basal tone in skeletal muscle arterioles. Isolated, pressurized gracilis muscle arterioles ( approximately 100 microm in diameter) of mice with T2-DM (C57BL/KsJ-db(-)/db(-)) exhibited greater basal tone to increases in intraluminal pressure (20-120 mmHg) than that of control vessels (at 80 mmHg, control: 25 +/- 5%; db/db: 34 +/- 4%, P < 0.05), which was reduced back to control level by catalase (db/db: 24 +/- 4%). Correspondingly, in carotid arteries of db/db mice, the level of dichlorofluorescein-detectable and catalase-sensitive H(2)O(2) was significantly greater. In control arterioles, exogenous H(2)O(2) (0.1-100 micromol/l) elicited dilations (maximum, 58 +/- 10%), whereas in arterioles of db/db mice H(2)O(2) caused constrictions (-28 +/- 8%), which were converted to dilations (maximum, 16 +/- 5%) by the thromboxane A(2)/prostaglandin H(2) (TP) receptor antagonist SQ-29548. In addition, arteriolar constrictions in response to the TP receptor agonist U-46619 were not different between the two groups of vessels. Endothelium denudation did not significantly affect basal tone and H(2)O(2)-induced arteriolar responses in either control or db/db mice. Also, in arterioles of db/db mice, but not in controls, 3-nitrotyrosine staining was detected in the endothelial layer of vessels. Thus we propose that, in mice with T2-DM, arteriolar production of H(2)O(2) is enhanced, which leads to increased synthesis of the constrictor prostaglandins thromboxane A(2)/prostaglandin H(2) in the smooth muscle cells, which enhance basal arteriolar tone. These alterations may contribute to disturbed regulation of skeletal muscle blood flow in Type 2 diabetes mellitus.     

Activated Platelets from Diabetic Rats Cause Endothelial Dysfunction by Decreasing Akt/Endothelial NO Synthase Signaling Pathway

Diabetes is associated with endothelial dysfunction and platelet activation, both of which may contribute to increased cardiovascular risk. The purpose of this study was to characterize circulating platelets in diabetes and clarify their effects on endothelial function. Male Wistar rats were injected with streptozotocin (STZ) to induce diabetes. Each experiment was performed by incubating carotid arterial rings with platelets (1.65×10⁷ cells/mL; 30 min) isolated from STZ or control rats. Thereafter, the vascular function was characterized in isolated carotid arterial rings in organ bath chambers, and each expression and activation of enzymes involved in nitric oxide and oxidative stress levels were analyzed. Endothelium-dependent relaxation induced by acetylcholine was significantly attenuated in carotid arteries treated with platelets isolated from STZ rats. Similarly, treatment with platelets isolated from STZ rats significantly reduced ACh-induced Akt/endothelial NO synthase signaling/NO production and enhanced TXB₂ (metabolite of TXA₂), while CD61 (platelet marker) and CD62P (activated platelet marker) were increased in carotid arteries treated with platelets isolated from STZ rats. Furthermore, the platelets isolated from STZ rats decreased total eNOS protein and eNOS dimerization, and increased oxidative stress. These data provide direct evidence that circulating platelets isolated from diabetic rats cause dysfunction of the endothelium by decreasing NO production (via Akt/endothelial NO synthase signaling pathway) and increasing TXA₂. Moreover, activated platelets disrupt the carotid artery by increasing oxidative stress.     

Vascular dysfunction in type 2 diabetic TallyHo mice: role for an increase in the contribution of PGH2/TxA2 receptor activation and cytochrome p450 products

In this study, we tested the hypothesis that spontaneously diabetic TallyHo (TH) mice, a novel polygenic model for type 2 diabetes, will exhibit endothelial dysfunction associated with an increased contribution from endothelium-derived contractile factors (EDCF). The cellular mechanisms underlying the increased contribution of EDCF were explored in 16 and 30-week-old male TH and age-matched male C57BL/6J mice (n=4-9). Blood glucose and serum lipid profiles were markedly increased in the TH mice. Superoxide generation, assessed with a lucigenin chemiluminescence assay, was markedly increased in the aortae of TH mice. Endothelium-dependent vascular relaxations and contractions to acetylcholine (ACh), but not endothelium-independent relaxations to sodium nitroprusside, were impaired and vascular contractions to phenylephrine were significantly enhanced in aortae from TH mice. Nomega-nitro-L-arginine methyl ester markedly increased the ACh-induced contractions in TH mice, whereas SQ29548, a thromboxane receptor antagonist, and cytochrome P450 (CYP) inhibitors 17-octadecynoic acid and sulfaphenazole, the latter being specific for CYP2C6 and 2C9, decreased and (or) normalized the contractile response to ACh in TH mice. The present study indicates that enhanced contribution of prostaglandin H2/thromboxane A2 receptor and CYP, likely CYP2C6 and 2C9, play a critical role in the pathogenesis of increased EDCF in the aortae of type 2 diabetic TH mice.     

Glucose intolerance in young TallyHo mice is induced by leptin-mediated inhibition of insulin secretion

The pathophysiology of TallyHo mouse, a recently established animal model for type 2 diabetes mellitus, was analyzed at prediabetic state to examine the inherent defects which contribute to the development of diabetes. At 4 weeks of age, the TallyHo mice already revealed glucose intolerance while their peripheral tissues exhibited normal insulin sensitivity. On the other hand, decreased plasma insulin concentration was observed with little differences in pancreatic insulin contents, indicating the impaired insulin secretion. Such defect, however, was not found in the isolated islets, which suggests a role of endocrine factor in impaired insulin secretion of TallyHo mice. Treatment of leptin inhibited the glucose-stimulated insulin secretion from the isolated islets of TallyHo mice, while in vivo administration of anti-leptin antibody lowered plasma glucose concentration with increased insulin level in TallyHo mice. Expression of glucokinase mRNA was decreased both in whole pancreas and leptin treated islets of TallyHo mice compared with whole pancreas in C57BL/6 mice and untreated islets of TallyHo mice, respectively. These results suggest that elevated plasma leptin can, through the inhibition of insulin secretion, induce glucose intolerance in TallyHo mice.     

Progression of coronary and mesenteric vascular dysfunction in Zucker obese and Zucker diabetic fatty rats

We investigated the progression of vascular dysfunction associated with the metabolic syndrome with and without hyperglycemia in lean, Zucker obese, and Zucker diabetic fatty (ZDF) rats. Responses of aorta and small coronary and mesenteric arteries were measured to endothelium-dependent and -independent vasodilators. Indices of oxidative stress were increased in serum from ZDF rats throughout the study, whereas values were increased in Zucker obese rats later in the study [thiobarbituric acid reactive substances: 0.45 +/- 0.02, 0.59 +/- 0.03 (P < 0.05), and 0.58 +/- 0.03 (P < 0.05) mug/ml in serum from 28- to 40-wk-old lean, Zucker obese, and ZDF rats, respectively]. Acetylcholine (ACh)-induced relaxation was not altered in vessels from lean animals from 8-40 wk. ACh-induced relaxation was nearly abolished in coronary arteries from 28- to 36-wk-old Zucker obese rats and by 16-36 wk in ZDF rats and was attenuated in aorta and mesenteric vessels from ZDF rats [%relaxation to 10 muM ACh: 72.2 +/- 7.1, 17.9 +/- 5.9 (P < 0.05), and 23.0 +/- 4.5 (P < 0.05) in coronary vessels; and 67.9 +/- 9.2, 50.1 +/- 5.5, and 42.3 +/- 4.7 (P < 0.05) in mesenteric vessels from 28- to 40-wk-old lean, Zucker obese, and ZDF rats, respectively]. The attenuated ACh-induced relaxation was improved when vessels were incubated with tiron, suggesting superoxide as a mechanism of endothelial dysfunction. Sodium nitroprusside-induced relaxation was not altered in aorta or coronary arteries and was potentiated in mesenteric arteries from Zucker obese rats. Our data suggest that diabetes enhances the progression of vascular dysfunction. Increases in indices of oxidative stress precede the development of dysfunction and may serve as a marker of endothelial damage.     

Gender differences in vascular reactivity to endothelin-1 (1-31) in mesenteric arteries from diabetic mice

Endothelin-1 (1-31) [ET-1 (1-31)], a novel member of the ET family, comprises 31 amino acids and is derived from the selective hydrolysis of big ET-1 by chymase. Although ET-1 (1-31) reportedly exerts biological effects by direct or indirect [via its conversion to ET-1 (1-21)] mechanisms, it is unclear whether in diabetes the vascular effects of ET-1 (1-31) display gender differences. We investigated this question by exposing mesenteric artery rings to ET-1 (1-31), using arteries from mice in the early or chronic phase of diabetes. In the early stage of diabetes, the ET-1 (1-31)-induced contraction was similar between age- and sex-matched control and streptozotocin (STZ)-induced diabetic mice. In the chronic stage of diabetes, the ET-1 (1-31)-induced contraction was enhanced in diabetic female mice, but not in diabetic male mice (vs. both age-matched control and early-stage diabetic mice). This enhancement was largely prevented by Y27632 (Rho kinase inhibitor), PD98059 [inhibitor of extracellular signal related kinases 1 and 2 (ERK1/2)], or SP600125 [C-jun terminal kinase (JNK) inhibitor]. These data indicate that the ET-1 (1-31)-induced vasoconstriction in the mesenteric artery may be specifically enhanced in established diabetic female mice, and that this enhancement may be due to alterations in the activities of Rho/Rho kinase or mitogen-activated protein kinase.     

Catechin prevents severe dyslipidemia-associated changes in wall biomechanics of cerebral arteries in LDLr-/-:hApoB+/+ mice and improves cerebral blood flow

Endothelial dysfunction and oxidative stress contribute to the atherosclerotic process that includes stiffening of large peripheral arteries. In contrast, our laboratory previously reported a paradoxical increase in cerebrovascular compliance in LDLr(-/-):hApoB(+/+) atherosclerotic (ATX) mice (7). We hypothesized that prevention of cerebral artery endothelial dysfunction with a chronic dietary antioxidant intake would normalize the changes in cerebral artery wall structure and biomechanics and prevent the decline in basal cerebral blood flow associated with atherosclerosis. Three-month-old ATX mice were treated, or not, for 3 mo with the polyphenol (+)-catechin (CAT; 30 mg·kg(-1)·day(-1)) and compared with wild-type controls. In isolated, pressurized cerebral arteries from ATX mice, CAT prevented endothelial dysfunction (deterioration of endothelium-dependent, flow-mediated dilations; P < 0.05), the inward hypertrophic structural remodeling (increase in the wall-to-lumen ratio; P < 0.05), and the rise in cerebrovascular compliance (rightward shift of the stress-strain curve measured in passive conditions, reflecting mechanical properties of the arterial wall; P < 0.05). Doppler optical coherence tomography imaging in vivo confirmed these findings, showing that cerebral compliance was higher in ATX mice and normalized by CAT (P < 0.05). CAT also prevented basal cerebral hypoperfusion in ATX mice (P < 0.05). Active remodeling of the cerebrovascular wall in ATX mice was further suggested by the increase (P < 0.05) in pro-metalloproteinase-9 activity, which was normalized by CAT. We conclude that, by preserving the endothelial function, a chronic treatment with CAT prevents the deleterious effect of severe dyslipidemia on cerebral artery wall structure and biomechanical properties, contributing to preserving resting cerebral blood flow.