Exposure to Maternal Diabetes Is Associated With Early Abnormal Vascular Structure in Offspring

Aim/hypothesis: In utero exposure to maternal diabetes increases the risk of developing hypertension and cardiovascular disorders during adulthood. We have previously shown that this is associated with changes in vascular tone in favor of a vasoconstrictor profile, which is involved in the development of hypertension. This excessive constrictor tone has also a strong impact on vascular structure. Our objective was to study the impact of in utero exposure to maternal diabetes on vascular structure and remodeling induced by chronic changes in hemodynamic parameters. Methods and Results: We used an animal model of rats exposed in utero to maternal hyperglycemia (DMO), which developed hypertension at 6 months of age. At a pre-hypertensive stage (3 months of age), we observed deep structural modifications of the vascular wall without any hemodynamic perturbations. Indeed, in basal conditions, resistance arteries of DMO rats are smaller than those of control mother offspring (CMO) rats; in addition, large arteries like thoracic aorta of DMO rats have an increase of smooth muscle cell attachments to elastic lamellae. In an isolated perfused kidney, we also observed a leftward shift of the flow/pressure relationship, suggesting a rise in renal peripheral vascular resistance in DMO compared to CMO rats. In this context, we studied vascular remodeling in response to reduced blood flow by in vivo mesenteric arteries ligation. In DMO rats, inward remodeling induced by a chronic reduction in blood flow (1 or 3 weeks after ligation) did not occur by contrast to CMO rats in which arterial diameter decreased from 428 ± 17 μm to 331 ± 20 μm (at 125 mmHg, p = 0.001). In these animals, the transglutaminase 2 (TG2) pathway, essential for inward remodeling development in case of flow perturbations, was not activated in low-flow (LF) mesenteric arteries. Finally, in old hypertensive DMO rats (18 months of age), we were not able to detect a pressure-induced remodeling in thoracic aorta. Conclusions: Our results demonstrate for the first time that in utero exposure to maternal diabetes induces deep changes in the vascular structure. Indeed, the early narrowing of the microvasculature and the structural modifications of conductance arteries could be a pre-emptive adaptation to fetal programming of hypertension

Cyclooxygenase-2 preserves flow-mediated remodelling in old obese Zucker rat mesenteric arteries

AIMS: Resistance arteries have a key role in the control of local blood flow and pressure, and chronic increases in blood flow induce endothelium-dependent outward hypertrophic remodelling. The incidence of metabolic syndrome increases with age, and the combination of these two risk factors impairs endothelium integrity, in part through an inflammatory process. We hypothesized that cyclooxygenase-2 (COX2) would affect remodelling in 12-month-old obese rats compared with young rats. METHODS AND RESULTS: Mesenteric arteries of obese and lean Zucker rats were alternatively ligated to generate high flow (HF) in the median artery. After 21 days, arteries were isolated for in vitro analysis. After 21 days, outward hypertrophic remodelling occurred in HF arteries in obese (498 +/- 20 vs. 443 +/- 18 mum intraluminal diameter in normal flow (NF) arteries, P < 0.01), but not in lean rats (454 +/- 17 vs. 432 +/- 14, NS; n = 12 per group). Endothelium-dependent (acetylcholine)-mediated relaxation (AMR) was lower in obese than in lean rats. AMR was reduced by NO-synthase blockade in all groups, and eNOS expression was higher in HF than in NF arteries without difference between lean and obese rats. Indomethacin further reduced AMR in HF arteries from obese rats only. Obesity increased COX2 immunostaining in mesenteric arteries. Acute COX2 inhibition (NS398) significantly reduced AMR in HF arteries from obese rats only, suggesting production of vasodilator prostanoid(s). In obese rats chronically treated with the COX2 inhibitor celecoxib, outward remodelling did not occur in HF arteries and AMR was improved without reaching the level found in lean rats. CONCLUSION: COX2 preserved in part flow-mediated arterial remodelling in old obese rats. Nevertheless, this effect was not sufficient to keep endothelium-dependent relaxation to the level obtained in lean rats

The endothelial mineralocorticoid receptor regulates vasoconstrictor tone and blood pressure

Pathophysiological aldosterone (aldo)/mineralocorticoid receptor (MR) signaling has significant effects on the cardiovascular system, resulting in hypertension and cardiovascular remodeling; however, the specific contribution of the vascular MR to blood pressure regulation remains to be established. To address this question, we generated a mouse model with conditional overexpression of the MR in endothelial cells (MR-EC). In basal conditions, MR-EC mice developed moderate hypertension that could be reversed by canrenoate, a pharmacological MR antagonist. MR-EC mice presented increased contractile response of resistance arteries to vasoconstrictors (phenylephrine, thromboxane A(2) analog, angiotensin II, and endothelin 1) in the absence of vascular morphological alterations. The acute blood pressure response to angiotensin II or endothelin 1 infusion was increased in MR-EC mice compared with that in littermate controls. These observations demonstrate that enhanced MR activation in the endothelium generates an increase in blood pressure, independent of stimulation of renal tubular Na(+) transport by aldo/MR or direct activation of smooth muscle MR and establish one mechanism by which endothelial MR activation per se may contribute to impaired vascular reactivity

Heme oxygenase 1 is differentially involved in blood flow-dependent arterial remodeling: role of inflammation, oxidative stress, and nitric oxide

Heme oxygenase 1 is induced by hemodynamic forces in vascular smooth muscle and endothelial cells. We investigated the involvement of heme oxygenase 1 in flow (shear stress)-dependent remodeling. Two or 14 days after ligation of mesenteric resistance arteries, vessels were isolated. In rats, at 14 days, diameter increased by 23% in high-flow arteries and decreased by 22% in low-flow arteries compared with normal flow vessels. Heme oxygenase activity inhibition using Tin-protoporphyrin abolished diameter enlargement in high-flow arteries and accentuated arterial narrowing in low-flow arteries (32% diameter decrease versus 22% in control). Two days after ligation, heme oxygenase 1 expression increased in high-flow and low-flow vessels, in association with a reduced mitochondrial aconitase activity (marker of oxidative stress) in high-flow arteries only. Inhibition of macrophage infiltration (clodronate) decreased heme oxygenase 1 induction in low-flow but not in high-flow arteries. Similarly, inhibition of NADPH oxidase activity (apocynin) decreased heme oxygenase 1 induction in low-flow but not high-flow arteries. However, dihydroethidium staining was higher in high-flow and low-flow compared with normal flow arteries. In arteries cannulated in an arteriograph, heme oxygenase 1 mRNA increased in a flow-dependent manner and was abolished by N(G)-nitro-l-arginine methyl ester, catalase, or mitochondrial electron transport chain inhibition. Furthermore, heme oxygenase 1 induction using cobalt-protoporphyrin restored altered high-flow remodeling in endothelial NO synthase knockout mice. Thus, in high-flow remodeling, heme oxygenase 1 induction depends on shear stress-generated NO and mitochondria-derived hydrogen peroxide. In low-flow remodeling, heme oxygenase 1 induction requires macrophage infiltration and is mediated by NADPH oxidase-derived superoxide

Determinants of flow-mediated outward remodeling in female rodents: respective roles of age, estrogens, and timing

OBJECTIVE: Flow (shear stress)-mediated outward remodeling (FMR) of resistance arteries is a key adaptive process allowing collateral growth after arterial occlusion but declining with age. 17-beta-estradiol (E2) has a key role in this process through activation of estrogen receptor alpha (ERalpha). Thus, we investigated the impact of age and timing for estrogen efficacy on FMR. APPROACH AND RESULTS: Female rats, 3 to 18 months old, were submitted to surgery to increase blood flow locally in 1 mesenteric artery in vivo. High-flow and normal-flow arteries were collected 2 weeks later for in vitro analysis. Diameter increased by 27% in high-flow arteries compared with normal-flow arteries in 3-month-old rats. The amplitude of remodeling declined with age (12% in 18-month-old rats) in parallel with E2 blood level and E2 substitution failed restoring remodeling in 18-month-old rats. Ovariectomy of 3-, 9-, and 12-month-old rats abolished FMR, which was restored by immediate E2 replacement. Nevertheless, this effect of E2 was absent 9 months after ovariectomy. In this latter group, ERalpha and endothelial nitric oxide synthase expression were reduced by half compared with age-matched rats recently ovariectomized. FMR did not occur in ERalpha(-/-) mice, whereas it was decreased by 50% in ERalpha(+/-) mice, emphasizing the importance of gene dosage in high-flow remodeling. CONCLUSIONS: E2 deprivation, rather than age, leads to decline in FMR, which can be prevented by early exogenous E2. However, delayed E2 replacement was ineffective on FMR, underlining the importance of timing of this estrogen action

Diabetes mellitus abrogates erythropoietin-induced cardioprotection against ischemic-reperfusion injury by alteration of the RISK/GSK-3β signaling

Recent studies reported cardioprotective effects of erythropoietin (EPO) against ischemia–reperfusion (I/R) injury through activation of the reperfusion injury salvage kinase (RISK) pathway. As RISK has been reported to be impaired in diabetes and insulin resistance syndrome, we examined whether EPO-induced cardioprotection was maintained in rat models of type 1 diabetes and insulin resistance syndrome. Isolated hearts were obtained from three rat cohorts: healthy controls, streptozotocin (STZ)-induced diabetes, and high-fat diet (HFD)-induced insulin resistance syndrome. All hearts underwent 25 min ischemia and 30 min or 120 min reperfusion. They were assigned to receive either no intervention or a single dose of EPO at the onset of reperfusion. In hearts from healthy controls, EPO decreased infarct size (14.36 ± 0.60 and 36.22 ± 4.20% of left ventricle in EPO-treated and untreated hearts, respectively, p < 0.05) and increased phosphorylated forms of Akt, ERK1/2, and their downstream target GSK-3β. In hearts from STZ-induced diabetic rats, EPO did not decrease infarct size (32.05 ± 2.38 and 31.88 ± 1.87% in EPO-treated and untreated diabetic rat hearts, respectively, NS) nor did it increase phosphorylation of Akt, ERK1/2, and GSK-3β. In contrast, in hearts from HFD-induced insulin resistance rats, EPO decreased infarct size (18.66 ± 1.99 and 34.62 ± 3.41% in EPO-treated and untreated HFD rat hearts, respectively, p < 0.05) and increased phosphorylation of Akt, ERK1/2, and GSK-3β. Administration of GSK-3β inhibitor SB216763 was cardioprotective in healthy and diabetic hearts. STZ-induced diabetes abolished EPO-induced cardioprotection against I/R injury through a disruption of upstream signaling of GSK-3β. In conclusion, direct inhibition of GSK-3β may provide an alternative strategy to protect diabetic hearts against I/R injury

Involvement of angiotensin II in the remodeling induced by a chronic decrease in blood flow in rat mesenteric resistance arteries

Blood flow reduction induces inward remodeling of resistance arteries (RAs). This remodeling occurs in ischemic diseases, diabetes and hypertension. Nonetheless, the effect of flow reduction per se, independent of the effect of pressure or metabolic influences, is not well understood in RA. As angiotensin II is involved in the response to flow in RA, we hypothesized that angiotensin II may also be involved in the remodeling induced by a chronic flow reduction. We analyzed the effect of angiotensin I-converting enzyme inhibition (perindopril) and angiotensin II type 1 receptor blockade (candesartan) on inward remodeling induced by blood flow reduction in vivo in rat mesenteric RAs (low flow (LF) arteries). After 1 week, diameter reduction in LF arteries was associated with reduced endothelium-dependent relaxation and lower levels of eNOS expression. Superoxide production and extracellular signal-regulated kinases 1/2 (ERK1/2 phosphorylation were higher in LF than in normal flow arteries. Nevertheless, the absence of eNOS or superoxide level reduction (tempol or apocynin) did not prevent LF remodeling. Perindopril and candesartan prevented inward remodeling in LF arteries. Contractility to angiotensin II was reduced in LF vessels by perindopril, candesartan and the ERK1/2 blocker PD98059. ERK1/2 activation (ratio phospho-ERK/ERK) was higher in LF arteries, and this activation was prevented by perindopril and candesartan. ERK1/2 inhibition in vivo (U0126) prevented LF-induced diameter reduction. Thus, inward remodeling because of blood flow reduction in mesenteric RA depends on unopposed angiotensin II-induced contraction and ERK1/2 activation, independent of superoxide production. These findings might be of importance in the treatment of vascular disorders

Reactive oxygen species and cyclooxygenase 2-derived thromboxane A2 reduce angiotensin II type 2 receptor vasorelaxation in diabetic rat resistance arteries

Angiotensin II has a key role in the control of resistance artery tone and local blood flow. Angiotensin II possesses 2 main receptors. Although angiotensin II type 1 receptor is well known and is involved in the vasoconstrictor and growth properties of angiotensin II, the role of the angiotensin II type 2 receptor (AT2R) remains much less understood. Although AT2R stimulation induces vasodilatation in normotensive rats, it induces vasoconstriction in pathological conditions involving oxidative stress and cyclooxygenase 2 expression. Thus, we studied the influence of cyclooxygenase 2 on AT2R-dependent tone in diabetes mellitus. Mesenteric resistance arteries were isolated from Zucker diabetic fatty (ZDF) and lean Zucker rats and studied using in vitro using wire myography. In ZDF rats, AT2R-induced dilation was lower than in lean rats (11% versus 21% dilation). Dilation in ZDF rats returned to the control (lean rats) level after acute superoxide reduction (Tempol and apocynin), cyclooxygenase 2 inhibition (NS398), or thromboxane A(2) synthesis inhibition (furegrelate). Cyclooxygenase 2 expression and superoxide production were significantly increased in ZDF rat arteries compared with arteries of lean rats. After chronic treatment with Tempol, AT2R-dependent dilation was equivalent in ZDF and lean rats. Chronic treatment of ZDF rats with NS398 also restored AT2R-dependent dilation to the control (lean rats) level. Plasma thromboxane B(2) (thromboxane A(2) metabolite), initially high in ZDF rats, was decreased by chronic Tempol and by chronic NS398 to the level found in lean Zucker rats. Thus, in type 2 diabetic rats, superoxide and thromboxane A(2) reduced AT2R-induced dilation. These findings are important to take into consideration when choosing vasoactive drugs for diabetic patients

Microvascular vasodilator properties of the angiotensin II type 2 receptor in a mouse model of type 1 diabetes

Diabetes Mellitus is associated with severe cardiovascular disorders involving the renin-angiotensin system, mainly through activation of the angiotensin II type 1 receptor (AT1R). Although the type 2 receptor (AT2R) opposes the effects of AT1R, with vasodilator and anti-trophic properties, its role in diabetes is debatable. Thus we investigated AT2R-mediated dilatation in a model of type 1 diabetes induced by streptozotocin in 5-month-old male mice lacking AT2R (AT2R). Glucose tolerance was reduced and markers of inflammation and oxidative stress (cyclooxygenase-2, gp91phox p22phox and p67phox) were increased in AT2R mice compared to wild-type (WT) animals. Streptozotocin-induced hyperglycaemia was higher in AT2R than in WT mice. Arterial gp91phox and MnSOD expression levels in addition to blood 8-isoprostane and creatinine were further increased in diabetic AT2R mice compared to diabetic WT mice. AT2R-dependent dilatation in both isolated mesenteric resistance arteries and perfused kidneys was greater in diabetic mice than in non-diabetic animals. Thus, in type 1 diabetes, AT2R may reduce glycaemia and display anti-oxidant and/or anti-inflammatory properties in association with greater vasodilatation in mesenteric arteries and in the renal vasculature, a major target of diabetes. Therefore AT2R might represent a new therapeutic target in diabetes