Remodeling Of Resistance Arteries Research Articles

Abstract 024: The Direct Renin Inhibitor Aliskiren Improves Vasodilation and Endothelial Function in Resistance Arteries from Diabetic and Hypertensive Patients

We previously demonstrated that the direct renin inhibitor aliskiren (ALK) significantly reduced the remodeling of subcutaneous resistance arteries of hypertensive patients as compared to the angiotensin-converting enzyme inhibitor ramipril (RAM). Here we questioned whether endothelial function of resistance arteries would improve after 1 year of blood pressure (BP) control with ALK or RAM. Sixteen diabetic patients with mild essential hypertension were randomized to ALK (150-300 mg once daily, n=9) or RAM (5-10 mg once daily, n=7). Subcutaneous resistance arteries were dissected from gluteal biopsy and mounted on a pressurized micromyograph. Endothelium-dependent and -independent relaxations were assessed by concentration-response curves to acetylcholine (1 nM to 100 μM) and sodium nitroprusside (10 nM to 1 mM) respectively, in arteries pre-contracted with norepinephrine (10 μM). Carotid-femoral pulse wave velocity (PWV) was assessed by applantion tonometry. Forearm flow mediated dilation (FMD) was assessed by ultrasounds. The expression of P-eNOS/e-NOS and the markers of oxidative stress nitrotyrosine and LOX-1 were assessed by immunohistochemistry in resistance arteries. Patients were similar for age, sex, BMI and glycemic and metabolic control. Systolic BP was significantly and equally reduced by both ALK and RAM (153±8.9 mmHg reduced to 128±7.3 mmHg and 151±10.6 mmHg reduced to 121±12.1, respectively, P<0.01) whereas diastolic BP was significantly reduced only in ALK-treated but not in RAM-treated patients (94.2±7.17 mmHg reduced to 81.4±6.31 mmHg, P<0.01; and 84.7±12.22 reduced to 78.6±7.48, NS, respectively). PWV and FMD were similar in both groups before and after treatment. Endothelium-dependent vasodilation was improved only by ALK (max dilation 92.5±3.8% vs 50.5±14%, P<0.05) but not by RAM (max dilation 72.9±5.6% vs 61.8±14.5%, NS). Endothelium-independent vasodilation was similar in all the groups. Only ALK increased P-eNOS/eNOS expression (+44% vs before treatment, p<0.05). The markers of oxidative stress were similar in both groups before and after treatment. In conclusion ALK improved endothelial function and induced vasodilation in resistance arteries from diabetic and hypertensive patients.

Abstract P252: Effect of Maternal Diabetes on Fetal Programming of Vascular Remodeling Mechanisms in Adult Rats

Modifications of the intra-uterine environment are now recognized as an important cause of fetal stress. Leading to several responses such as loss of structure/function and pre-emptive adaptations to an adverse post-natal environment, and finally to adult diseases. It has been demonstrated that children of diabetic mothers have an increased risk of developing cardiovascular diseases (i.e. hypertension) during adulthood. Blood vessels are able to reorganize their structure in response to physiological alteration of blood flow or pathological stimuli. Constrictive vascular remodeling is currently associated with the occurrence of cardiovascular diseases. Thus, our objective was to study vascular remodeling in case of in utero exposure to maternal diabetes. This is in order to investigate the effect of intra-uterine environment perturbations on vascular fetal programming. We have developed an animal model of rats exposed in utero to moderated maternal hyperglycemia (DMO), which developed an hypertension around 6 months of age. We analyzed structure of elastic and resistance arteries (internal diameter, MCSA, intima-media thickness and remodeling parameters) in absence or presence of established hypertension in male DMO compared to controls (CMO) (at 3 and 18 months of age). Moreover, by in vivo sequential ligation of mesenteric arteries, we have studied vascular remodeling of resistance arteries in response to low (LF), normal (NF) or high flow (HF). In old DMO, we did not observe any vascular remodeling induced by hypertension neither in aorta nor in mesenteric arteries. This could be related to the increase of smooth muscle cell attachments that we observed. Moreover, after 1 or 3 weeks of mesenteric arteries ligation, DMO did not exhibit any constrictive remodeling in LF although expansive remodeling in HF is maintained. Interestingly, we measured an increase of eNOS activity and GP91 in LF arteries of theses animals although transglutaminase-2 protein expression was not modified. Our results demonstrate that in utero exposure to maternal diabetes induce modification of vascular remodeling mechanisms in adulthood. Absence of vascular response could be a pre-emptive adaptation to fetal programmed-hypertension in these animals.

Association of insulin resistance and coronary artery remodeling: an intravascular ultrasound study.

BackgroundThere are few studies that investigated the correlation between insulin resistance (IR) and the coronary artery remodeling. The aim of the study is to investigate the association of IR measured by homeostasis model assessment of insulin resistance (HOMA-IR) and coronary artery remodeling evaluated by intravascular ultrasound (IVUS).MethodsA total of 298 consecutive patients who received percutaneous coronary interventions under IVUS guidance were retrospectively enrolled. The value of HOMA-IR more than 2.5 was considered as IR positive. Metabolic syndrome was classified according to NCEP ATP III guidelines. The remodeling index was defined as the ratio of the external elastic membrane (EEM) area at the lesion site to the EEM area at the proximal reference site.ResultsA total of 369 lesions were analyzed (161 lesions in HOMA-IR positive and 208 lesions in HOMA-IR negative). Remodeling index was significantly higher in the HOMA-IR positive group compared with the negative group (HOMA-IR positive vs. negative: 1.074 ± 0.109 vs. 1.042 ± 0.131, p = 0.013). There was a significant positive correlation between remodeling index and HOMA-IR (p = 0.010). Analysis of HOMA-IR according to remodeling groups showed increasing tendency of HOMA-IR, and it was statistically significant (p = 0.045). Multivariate analysis revealed that only HOMA-IR was an independent predictor of remodeling index (r = 0.166, p = 0.018).ConclusionIncreased IR estimated by HOMA-IR was significantly associated with a higher remodeling index and positive coronary artery remodeling.

Mast cell stabilization with sodium cromoglycate modulates pulmonary vessel wall remodeling during four-day hypoxia in rats

ABSTRACTAim of the study: In rats, the environment with low content of oxygen induces hypoxic pulmonary hypertension. Remodeling of pulmonary resistance arteries is particularly triggered by the mast cell degranulation products, e.g., rodent-like interstitial collagenase (matrix metalloproteinase 13). Administration of sodium cromoglycate leads to stabilization of mast cell granules, and thus to the modified remodeling process. Materials and Methods: During four-day hypoxia, we treated rats with sodium cromoglycate. Pulmonary vascular remodeling was assessed as well as counts of periarterial pulmonary mast cells, both total and matrix metalloproteinase 13-positive ones. Results: Four-day hypoxia induced remodeling of both resistance arteries and large conduit arteries. We have found increase in the tunica media thickness of resistance arteries. Tunica adventitia thickness of both resistance arteries and large conduit arteries with a diameter of over 300 μm increased as well; the latter ones revealed increase in the number of vasa vasorum in their walls. Mast cell stabilization suppressed hypoxic pulmonary vascular remodeling in resistance pulmonary arteries. Four-day hypoxia led to changes in distribution of toluidine blue-detected and MMP-13 positive periarterial mast cells; this redistribution was also influenced by the administration of sodium cromoglycate. Conclusions: The number of pulmonary periarterial mast cells seemingly decreases during hypoxia due to their degranulation, which disables their identification. Large conduit arteries do not affect final blood pressure in the pulmonary vascular bed; however, their structure changes substantially under hypoxia. Such remodeling changes are not mediated by mast cell products only since they have occurred in spite of stabilization of mast cell granules.

Microvessel mechanobiology in pulmonary arterial hypertension: cause and effect.

Pulmonary arterial hypertension (PAH, or group 1 pulmonary hypertension) is a progressive, insidious, and fatal illness of the pulmonary microvasculature. Proliferative remodeling of the small resistance arteries results in an elevation of mean pulmonary arterial pressure (mPAP) and total pulmonary vascular resistance (PVR), increasing the workload on the right ventricle (RV) until it eventually fails.1 Although these hemodynamic parameters are useful for clinical diagnosis, alone they have limited prognostic value; furthermore, vasodilator therapy aimed at reduction of PVR has achieved only modest success, and increases in patient survival are attributed primarily to earlier diagnosis and intervention.2 These limitations have motivated research focused on identifying measurable physiological parameters that could both predict mortality and serve as measures for therapeutic efficacy. Decreased pulmonary arteriolar compliance (PAC) is a major factor contributing to the increased RV workload and failure in PAH.3–5 PAC measures a vessel’s ability to deform under loading, and as a blood vessel stiffens its compliance decreases. Total vessel compliance is estimated as stroke volume divided by pulse pressure (PP). This estimation alone is a strong predictor of survival in idiopathic and familial PAH.5 Although proximal artery stiffness has received a great deal of attention in hypoxic pulmonary hypertension and is important in increasing RV workload, changes in PAC affect the entire pulmonary vasculature with the largest portion of that change occurring in vessels distal to the lung hilum.6 Understanding how vascular remodeling changes PAC, especially in the distal vasculature where much of our knowledge on vessel mechanics is currently lacking, will allow us to link the cellular and molecular mediators commonly studied in PAH with the biomechanical changes that cause elevated pressures and RV failure. Pulmonary arterial smooth muscle cells (PASMCs) and adventitial fibroblasts decrease PAC by altering the composition, amount, and organization …

Abstract MP10: Early Functional and Structural Alterations of Resistance Arteries in Mice Treated with an Inhibitor of the Vascular Endothelial Growth Factor Receptor

Background: Inhibition of tyrosine kinases receptors such as vascular endothelial growth factor receptor (VEGFR) and epidermal growth factor receptor (EGFR) improves outcomes in patients with cancers. Only VEGFR inhibitors, however, induce severe hypertension whose mechanisms remain unclear. We hypothesized that VEGFR inhibitors may induce early vascular functional and structural alterations, that may precede the development of hypertension. Methods and results: Normotensive SV-129 mice (8 weeks old, 5 for each group) were treated or not with the VEGFR inhibitor Vatalanib (VAT, 100 mg/Kg/day) or the EGFR inhibitor Gefitinib (GEF, 100 mg/Kg/day). Vehicle-treated control mice were also studied. Blood pressure (BP) was measured by tail-cuff method. Endothelium-dependent and -independent relaxations were assessed by concentration-response curves to acetylcholine (1 nM to 100 μM) ± L-NAME (100 μM) and sodium nitroprusside (10 nM to 1 mM) respectively, in mesenteric arteries pre-contracted with norepinephrine (10 μM). Media-to-lumen ratio (M/L, an index of early vascular remodeling), and cross sectional area (CSA) were evaluated on pressurized preparations. After two weeks, BP was similarly preserved in both VAT- and GEF-treated mice as compared to vehicle-treated mice (89.8±1.5 mmHg and 87.2±2.8 mmHg vs 92.2±2.2 mmHg, respectively, NS). Endothelium-dependent relaxation was similarly preserved in vehicle-treated and GEF-treated mice, whereas it was reduced in VAT-treated mice (-17% vs vehicle-treated mice, P<0.05). L-NAME blunted acetylcholine-induced relaxation in all groups except in VAT-treated mice, suggesting an impairment of NO production only in this group. Endothelium-independent relaxation was similar in all groups. Only VAT-treated mice presented increased M/L as compared to vehicle-treated mice (6.3±0.1% vs 5.4±0.1%, P<0.05). M/L resulted similar in GEF-treated and vehicle-treated mice. CSA was similar in all groups. Conclusion: In normotensive mice, only VAT promoted early vascular alterations such as endothelial dysfunction and vascular remodeling in resistance arteries. Those changes in the vasculature are distinctive of hypertension and might precede and sustain the development of the hypertensive disease.

Abstract 346: Subcutaneous Resistance Artery Remodeling And Function In Chronic Kidney Disease Patients

Background: Chronic kidney disease (CKD) is associated with cardiovascular (CV) complications. However, interventional trials targeting classical CV risks factors have been often unsuccessful in advanced stage CKD, which emphasizes the need to better understand CKD-associated vascular disorders. Resistance arteries are a key determinant of blood pressure (BP) and their changes in different CV conditions contribute to target organ damage. The aim of the present study was to characterize resistance artery remodeling and function in CKD patients, compared to vessels from hypertensive (HTN) subjects. Method: Twenty-two stage 4 CKD patients (aged 63.6±3.1 years) and 16 HTN subjects (45.6±16.1 years) were included in the present study. They all underwent a subcutaneous biopsy under local anaesthesia. Small artery remodeling and function were studied on a pressurized myograph, and subcutaneous fat CD3 infiltration and media fibronectin expression by immunostaining. Vascular smooth muscle cells (VSMCs) were counted after hematoxilin-eosin staining. Results: CKD systolic BP was similar to HTN (133±18 vs. 143±10 mmHg, respectively). Vasodilatory responses to acetylcholine were lower in CKD compared to HTN (maximal relaxation (%), 74.3±3.4 vs. 87.5±2.7, P<0.05). Media/lumen at 60 mmHg was lower in CKD than in HTN (6.7±0.5 vs 8.8±0.7, P<0.05). Resistance artery stiffness was lower in CKD compared to HTN (strain at 120 mmHg, 0.845±0.126 vs 0.585±0.099, P<0.05). Fibronectin staining in resistance arteries was lower in CKD than HTN (8.2±0.8 vs 23.3±1.7 RFU/μm2, P<0.001). Less VSMCs were present in the arterial wall of CKD compared to HTN (5.4±0.4 vs 7.2±0.5 cells/μm2, P<0.05). Subcutaneous fat presented fewer CD3+ cells in CKD than HTN (12.8±4.1 vs 23.7±12.8 cells/mm2, P<0.05). Conclusion: Despite higher levels of BP, resistance arteries isolated from CKD patients exhibited no vascular remodeling and lower arterial stiffness compared with HTN patients. These results are in line with the maladaptive hypotrophic remodeling observed in large vessels in CKD, suggesting a generalized vascular defect in mechanotransduction in CKD.

Role of estrogens and age in flow-mediated outward remodeling of rat mesenteric resistance arteries.

In resistance arteries, a chronic increase in blood flow induces hypertrophic outward remodeling. This flow-mediated remodeling (FMR) is absent in male rats aged 10 mo and more. As FMR depends on estrogens in 3-mo-old female rats, we hypothesized that it might be preserved in 12-mo-old female rats. Blood flow was increased in vivo in mesenteric resistance arteries after ligation of the side arteries in 3- and 12-mo-old male and female rats. After 2 wk, high-flow (HF) and normal-flow (NF) arteries were isolated for in vitro analysis. Arterial diameter and cross-sectional area increased in HF arteries compared with NF arteries in 3-mo-old male and female rats. In 12-mo-old rats, diameter increased only in female rats. Endothelial nitric oxide synthase expression and endothelium-mediated relaxation were higher in HF arteries than in NF arteries in all groups. ERK1/2 phosphorylation, NADPH oxidase subunit expression levels, and arterial contractility to KCl and to phenylephrine were greater in HF vessels than in NF vessels in 12-mo-old male rats only. Ovariectomy in 12-mo-old female rats induced a similar pattern with an increased contractility without diameter increase in HF arteries. Treatment of 12-mo-old male rats and ovariectomized female rats with hydralazine, the antioxidant tempol, or the angiotensin II type 1 receptor blocker candesartan restored HF remodeling and normalized arterial contractility in HF vessels. Thus, we found that FMR of resistance arteries remains efficient in 12-mo-old female rats compared with age-matched male rats. A balance between estrogens and vascular contractility might preserve FMR in mature female rats.

Structural remodeling of coronary resistance arteries: effects of age and exercise training.

Age is known to induce remodeling and stiffening of large-conduit arteries; however, little is known of the effects of age on remodeling and mechanical properties of coronary resistance arteries. We employed a rat model of aging to investigate whether 1) age increases wall thickness and stiffness of coronary resistance arteries, and 2) exercise training reverses putative age-induced increases in wall thickness and stiffness of coronary resistance arteries. Young (4 mo) and old (21 mo) Fischer 344 rats remained sedentary or underwent 10 wk of treadmill exercise training. Coronary resistance arteries were isolated for determination of wall-to-lumen ratio, effective elastic modulus, and active and passive responses to changes in intraluminal pressure. Elastin and collagen content of the vascular wall were assessed histologically. Wall-to-lumen ratio increased with age, but this increase was reversed by exercise training. In contrast, age reduced stiffness, and exercise training increased stiffness in coronary resistance arteries from old rats. Myogenic responsiveness was reduced with age and restored by exercise training. Collagen-to-elastin ratio (C/E) of the wall did not change with age and was reduced with exercise training in arteries from old rats. Thus age induces hypertrophic remodeling of the vessel wall and reduces the stiffness and myogenic function of coronary resistance arteries. Exercise training reduces wall-to-lumen ratio, increases wall stiffness, and restores myogenic function in aged coronary resistance arteries. The restorative effect of exercise training on myogenic function of coronary resistance arteries may be due to both changes in vascular smooth muscle phenotype and expression of extracellular matrix proteins.

Determinants of Flow-Mediated Outward Remodeling in Female Rodents

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-β-estradiol (E2) has a key role in this process through activation of estrogen receptor α (ERα). 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, ERα and endothelial nitric oxide synthase expression were reduced by half compared with age-matched rats recently ovariectomized. FMR did not occur in ERα −/− mice, whereas it was decreased by 50% in ERα +/− 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.

0362: Interleukin 17 production induced by angiotensin II type 2 receptor activation in T cells drives flow-mediated outward remodeling of mouse resistance arteries

Flow (shear stress)-mediated outward remodeling of resistance arteries is involved in collateral growth during postischemic revascularization. Besides the NO pathway the involvement of inflammatory factors and reactive oxygen species has been suggested in this remodeling although the mechanism and the sequence remains unknown. As T cells drive the inflammatory response, we investigated their role in flow-mediated remodeling. Mouse mesenteric resistance arteries (250μm internal diameter) were submitted in vivo to a chronic increase (144±18 to 239±25 μl/min) in blood flow. Arteries were collected for in vitro analysis after 1 to 7 days. After 1 week, remodeling occurred in high flow (HF) arteries (diameter increased from 242±21μm to 324±20μm, n=xx per group, P<00.1) in control mice. No remodeling occurred in nude mice and in mice treated with anti-CD3 antibodies. In control arteries lymphocyte T cells accumulate around HF arteries at day 1 (shown using both immunohistology and flow cytometry) and they expressed angiotensin II type 2 receptor (AT2R). In isolated T cells AT2R stimulation induced preferentially IL-17 production (20±6pg/ml to 50±5pg/ml, n=xx per group, P<00.1). We then tested the occurrence of remodeling in mice lacking AT2R or IL-17. In both AT2R-/- and IL-17-/- mice diameter expansion did not occur whereas T cells and myeloid cells accumulation around the HF artery was not affected. IL-17 infusion (5ng/h, Alzet osmotic minipump) to nude mice and to AT2R-/- mice restored HF remodeling to control level. In order to confirm the role of AT2R in flow-mediated remodeling in another vascular territory, neovascularization was examined using an ischemic skin flap model in AT2R-/- mice. As in mesenteric arteries, the absence of AT2R prevented diameter expansion in the arteries feeding the skin flap. we demonstrate for the first time the involvement of lymphocytes T cells polarization into TH17 by angiotensin II type 2 receptor in flow-mediated outward remodeling of resistance arteries.

Chronic intermittent hypoxia alters blood pressure and mesenteric small resistance arterial responsiveness in mice (710.9)

Blood pressure (BP) is elevated in rats and mice exposed to chronic intermittent hypoxia (CIH). It is unclear if altered vascular function, such as myogenic reactivity (MR), contributes to BP elevation in CIH. We aimed to characterize the vascular alterations in small resistance arteries from CIH‐exposed mice. C57BL/6J mice were exposed to either CIH (1 min alterations of 5% and 21% O2) or normoxia (sham control, constant 21% O2), 8 hrs/day for 5 weeks. Compared to sham controls, CIH‐exposed mice had significantly higher BP, measured by radio telemetry. Mesenteric small resistance arteries were isolated and pressurized for diameter and wall thickness measurement. Myogenic constriction in response to intraluminal pressure increase (i.e., MR) was significantly attenuated, especially at pressures 蠅 70 mm Hg. At 70 mm Hg, vasodilations to acetylcholine and sodium nitroprusside, but not to external Ca2+ removal, were significantly impaired in CIH, whereas vasoconstrictions to phenylephrine (PE), endothelin‐1, U46619, and high [K+]o were similar between sham and CIH arteries. When intraluminal pressure was increased to 110 mm Hg, however, PE‐induced vasoconstriction was significantly reduced, and nitric oxide‐dependent vasodilation was further attenuated in CIH arteries. Arterial remodeling was not obvious since wall thickness, wall to lumen ratio, and passive diameters in Ca2+‐free solution were similar between two groups. Thus, CIH 1) elevates BP; 2) reduces MR; 3) reduces arterial responsiveness to vasodilators, and, at higher intraluminal pressure, to vasoconstrictors; 4) has no effect on arterial remodeling in mouse mesenteric small resistance arteries. It is likely that the altered arterial responsiveness is involved in the regulation of BP in CIH.Grant Funding Source: NIH/NHLBI R01 HL107654

Structural remodeling of coronary resistance arteries: effects of age and exercise training (1079.24)

Age induces medial thickening and stiffening of conduit arteries; however, little is known of the effects of age on remodeling and mechanical properties of coronary resistance arteries. We determined 1) whether age increases wall thickness and stiffness of coronary resistance arteries, and 2) whether exercise training reverses age‐induced increases in wall thickness and stiffness. Young and old rats remained sedentary or underwent 10 wks of exercise training. Coronary resistance arteries were isolated for determination of wall/lumen ratio, wall stiffness, and myogenic contractile responses to intraluminal pressure changes. Collagen and elastin content of the vessel wall were assessed histologically. Age increased wall/lumen ratio, but reduced wall stiffness. Collagen/elastin ratio did not change with age. Exercise training reduced wall/lumen ratio and collagen/elastin ratio, but increased stiffness in old arteries. Myogenic responses were impaired with age and restored by exercise training. Thus, age induces inward remodeling and reduces wall stiffness of coronary resistance arteries, leading to loss of myogenic responsiveness. Exercise training induces outward remodeling and increases wall stiffness of aged coronary resistance arteries, thus restoring myogenic function. The restorative effects of exercise training may be due to changes in smooth muscle and extracellular matrix proteins.Grant Funding Source: NIH R01 HL 90937

AGEs breaking and antioxidant treatment improves endothelium-dependent dilation without effect on flow-mediated remodeling of resistance arteries in old Zucker diabetic rats

BackgroundA chronic increase in blood flow in resistance arteries is associated with increased lumen diameter (outward remodeling) and improved endothelium (NO)-mediated relaxation. Flow-mediated remodeling of resistance arteries is essential for revascularization in ischemic diseases. Nevertheless, it is impaired in 12 to 24-month old rats and in young Zucker Diabetic Fatty (ZDF) rats due to advanced glycation end products (AGEs) and oxidative stress. As type 2 diabetes occurs preferentially in older subjects we investigated flow-mediated remodeling and the effect of the AGEs breaker ALT-711 associated or not to the antioxidant TEMPOL in one-year old lean (LZ) and ZDF rats.MethodsMesenteric resistance arteries were exposed to high (HF) or normal blood flow (NF) in vivo. They were collected after 2 weeks for in vitro analysis.ResultsIn LZ rats, diameter expansion did not occur despite a significant increase in blood flow in HF arteries. Nevertheless, endothelium-mediated relaxation was higher in HF than in NF arteries. ALT-711, alone or in combination with TEMPOL, restored outward remodeling in HF arteries in association with AGEs reduction. TEMPOL alone had no effect. ALT-711, TEMPOL or the combination of the 2 drugs did not significantly affect endothelium-mediated relaxation in HF and NF arteries.In ZDF rats, diameter did not increase despite the increase in blood flow and endothelium-mediated relaxation was further decreased in HF arteries in association with AGEs accumulation and excessive oxidative stress. In both NF and HF arteries, endothelium-mediated relaxation was lower in ZDF than in LZ rats. ALT-711, TEMPOL or their combination did not improve remodeling (diameter equivalent in HF and NF arteries). In parallel, they did not reduce AGEs level and did not improve MMPs activity. Nevertheless, ALT-711 and TEMPOL partly improved endothelium-mediated relaxation through a reduction of oxidative stress and the association of ALT-711 and TEMPOL fully restored relaxation to the level found in LZ rats.ConclusionsALT-711 did not improve outward remodeling in mature ZDF rats but it reduced oxidative stress and consequently improved endothelium-dependent relaxation. In mature LZ rats, ALT-711 improved outward remodeling and reduced AGEs level. Consequently, AGEs breaking is differently useful in ageing whether it is associated with diabetes or not.

Therapeutic hypercapnia prevents inhaled nitric oxide-induced right-ventricular systolic dysfunction in juvenile rats

Chronic pulmonary hypertension in the neonate and infant frequently presents with right-ventricular (RV) failure. Current clinical management may include protracted treatment with inhaled nitric oxide (iNO), with the goal of reducing RV afterload. We have previously reported that prolonged exposure to iNO causes RV systolic dysfunction in the chronic hypoxia-exposed juvenile rat, which was prevented by a peroxynitrite decomposition catalyst. Given that inhalation of CO2 (therapeutic hypercapnia) may limit oxidative stress and upregulated cytokine expression in the lung and other organs, we hypothesized that therapeutic hypercapnia would attenuate cytokine-mediated nitric oxide synthase (NOS) upregulation, thus limiting peroxynitrite generation. Sprague–Dawley rat pups were exposed to chronic hypoxia (13% O2) from postnatal day 1 to 21, while receiving iNO (20ppm) from day 14 to 21, with or without therapeutic hypercapnia (10% CO2). Therapeutic hypercapnia completely normalized RV systolic function, RV hypertrophy, and remodeling of pulmonary resistance arteries in animals exposed to iNO. Inhaled nitric oxide-mediated increases in RV peroxynitrite, apoptosis, and contents of tumor necrosis factor (TNF)-α, interleukin (IL)-1α, and NOS-2 were all attenuated by therapeutic hypercapnia. Inhibition of NOS-2 activity with 1400W (1mg/kg/day) prevented iNO-mediated upregulation of peroxynitrite and led to improved RV systolic function. Blockade of IL-1 receptor signaling with anakinra (500mg/kg/day) decreased NOS-2 content and had similar effects compared to NOS-2 inhibition on iNO-mediated effects, whereas blockade of TNF-α signaling with etanercept (0.4mg/kg on alternate days) had no effects on these parameters. We conclude that therapeutic hypercapnia prevents the adverse effects of sustained exposure to iNO on RV systolic function by limiting IL-1-mediated NOS-2 upregulation and consequent nitration. Therapeutic hypercapnia also acts synergistically with iNO in normalizing RV hypertrophy, vascular remodeling, and raised pulmonary vascular resistance secondary to chronic hypoxia.

Time-related alteration in flow- (shear stress-) mediated remodeling in resistance arteries from spontaneously hypertensive rats.

Hypertension is a major risk factor for cardiovascular disorders. As flow-mediated outward remodeling has a key role in postischemic revascularization, we investigated this remodeling in mesenteric resistance arteries of normotensive (WKY) and spontaneously hypertensive rats (SHRs) aged 3 to 9 months. Sequential ligation of mesenteric resistance arteries allowed modifying blood flow in vivo, thus exposing arteries to low, normal, or high flow. After 1, 3, 8, or 24 weeks, arteries were isolated for in vitro study. High flow (HF) induced outward hypertrophic remodeling in WKY rats after 1 week and persisted until 24 weeks without change in wall to lumen ratio. In SHRs, diameter increase was delayed, occurring only after 3 weeks. Nevertheless, it was reduced at 8 weeks and no longer significant after 24 weeks. In parallel, media cross-section area increased more with time in SHRs than in WKY rats and this was associated with increased contractility and oxidative stress with decreased NO-dependent relaxation. Low flow induced progressive inward remodeling until 24 weeks in both strains with excessive hypertrophy in SHRs. Thus, a chronic increase in flow induced transitory diameter expansion and long-lasting hypertrophy in SHRs. This could contribute to the higher susceptibility of hypertensive subjects to ischemic diseases.

Abstract 159: Inhibition of Ace Does Not Prevent 20-hete-mediated Vascular Injury in Cyp4a12tg Mice

20-hydroxyeicosatetraenoic acid (20-HETE), a cytochrome P450-arachidonic acid metabolite in the microcirculation, increases smooth muscle contraction and proliferation, stimulates endothelial dysfunction and activation and promotes hypertension that is dependent, in part, on the renin angiotensin system (RAS). We showed that in cultured human microvascular endothelial cells, 20-HETE increases angiotensin converting enzyme (ACE) mRNA by 3.1-fold (± 0.16; p&lt;0.05), ACE protein by 4.2-fold (±0.99; p&lt;0.05) and cellular and extracellular ACE activity by 2.2-fold (±0.09; p&lt;0.05) and 1.96-fold (±0.16; p&lt;0.05), respectively. The contribution of ACE to vascular 20-HETE-mediated microvascular remodeling in hypertension was assessed using the Cyp4a12 transgenic mouse in which the expression of the Cyp4a12-20-HETE synthase is driven by a tetracyclin (doxycycline, DOX)-sensitive promoter. Treatment of Cyp4a12tg mice with DOX increased blood pressure (133±6 vs. 107±3 mmHg; p&lt;0.05) and remodeling of renal resistance arteries (M/L, 0.15±0.01 vs. 0.07±0.01; p&lt;0.05); these increases were abrogated by co-treatment with the 20-HETE antagonist, 20-HEDGE. Renal ACE protein expression was significantly higher in DOX-treated (1.6 ± 0.17 fold; p&lt;0.05) as compared to untreated Cyp4a12tg mice; co-treatment with 20-HEDGE prevented the increase in ACE expression. Lisinopril treatment of Cyp4a12tg mice concurrently receiving DOX prevented the blood pressure increase (133±6 vs. 104±2 mmHg; p&lt;0.05) but did not interfere with renal microvascular 20-HETE production (1.05±0.18 vs 1.09±0.30 ng/mg). Co-treatment with lisinopril and DOX attenuated but did not prevent remodeling of renal microvessels (M/L, 0.07±0.0 vs. 0.15±0.01 vs. 0.11±0.01; p&lt;0.05 in water, DOX and DOX+Lisinopril, respectively). In contrast, co-treatment with 20-HEDGE prevented DOX-induced remodeling of resistance arteries of Cyp4a12tg mice. This study demonstrates that 20-HETE is a key determinant of microvascular remodeling in DOX-induced Cyp4a12tg hypertensive mice. 20-HETE’s mechanism only partially relies on activation of the RAS. Thus, the 20-HETE-driven microvascular remodeling does not fully depend on the upregulation of ACE and elevation of blood pressure.

Abstract 238: Interleukin 17 Production Induced by Angiotensin Ii Type 2 Receptor Activation in T Cells Drives Flow-mediated Outward Remodeling of Mouse Resistance Arteries

Objective: Flow (shear stress)-mediated outward remodeling of resistance arteries is involved in collateral growth during postischemic revascularization. Besides the NO pathway the involvement of inflammatory factors and reactive oxygen species has been suggested in this remodeling although the mechanism and the sequence remains unknown. As T cells drive the inflammatory response, we investigated their role in flow-mediated remodeling. Mouse mesenteric resistance arteries (250μm internal diameter) were submitted in vivo to a chronic increase (144±18 to 239±25 μl/min) in blood flow. Arteries were collected for in vitro analysis after 1 to 7 days. Results: After 1 week, remodeling occurred in high flow (HF) arteries (diameter increased from 242±21μm to 324±20μm, n=9 per group, P&lt;00.1) in control mice. No remodeling occurred in nude mice and in mice treated with anti-CD3 antibodies. In control arteries lymphocyte T cells accumulate around HF arteries at day 1 (shown using both immunohistology and flow cytometry) and they expressed angiotensin II type 2 receptor (AT2R). In isolated T cells AT2R stimulation induced preferentially IL-17 production (20±6pg/ml to 50±5pg/ml, n=7 per group, P&lt;00.1). We then tested the occurrence of remodeling in mice lacking AT2R or IL-17. In both AT2R-/- and IL-17-/- mice diameter expansion did not occur whereas T cells and myeloid cells accumulation around the HF artery was not affected. IL-17 infusion (5ng/h, Alzet osmotic minipump) to nude mice and to AT2R-/- mice restored HF remodeling to control level. In order to confirm the role of AT2R in flow-mediated remodeling in another vascular territory, neovascularization was examined using an ischemic skin flap model in AT2R-/- mice. As in mesenteric arteries, the absence of AT2R prevented diameter expansion in the arteries feeding the skin flap. Conclusion: we demonstrate for the first time the involvement of lymphocytes T cells polarization into TH17 by angiotensin II type 2 receptor in flow-mediated outward remodeling of resistance arteries.

Rho-Kinase Inhibitor Prevents Bleomycin-Induced Injury in Neonatal Rats Independent of Effects on Lung Inflammation

Bleomycin-induced lung injury is characterized in the neonatal rat by inflammation dominated by neutrophils and macrophages, inhibited distal airway and vascular development, and pulmonary hypertension, similar to human infants with severe bronchopulmonary dysplasia. Rho-kinase (ROCK) is known to mediate lung injury in adult animals via stimulatory effects on inflammation. We therefore hypothesized that inhibition of ROCK may ameliorate bleomycin-induced lung injury in the neonatal rat. Pups received daily intraperitoneal bleomycin or saline from Postnatal Days 1 through 14 with or without Y-27632, a ROCK inhibitor. Treatment with Y-27632 prevented bleomycin-induced pulmonary hypertension, as evidenced by normalized pulmonary vascular resistance, decreased right-ventricular hypertrophy, and attenuated remodeling of pulmonary resistance arteries. Bleomycin-induced changes in distal lung architecture, including septal thinning, inhibited alveolarization, and decreased numbers of peripheral arteries and capillaries, were partially or completely normalized by Y-27632. Treatment with Y-27632 or a CXCR2 antagonist, SB265610, also abrogated tissue neutrophil influx, while having no effect on macrophages. However, treatment with SB265610 did not prevent bleomycin-induced lung injury. Lung content of angiostatic thrombospondin-1 (TSP1) was increased significantly in the lungs of bleomycin-exposed animals, and was completely attenuated by treatment with Y-27632. Thrombin-stimulated TSP1 production by primary cultured rat pulmonary artery endothelial cells was also attenuated by Y-27632. Taken together, our findings suggest a preventive effect of Y-27632 on bleomycin-mediated injury by a mechanism unrelated to inflammatory cells. Our data suggest that improvements in lung morphology may have been related to indirect stimulatory effects on angiogenesis via down-regulation of TSP1.

20-HETE induces remodeling of renal resistance arteries independent of blood pressure elevation in hypertension

20-Hydroxyeicosatetraenoic acid (20-HETE) is a cytochrome P-450 (Cyp)-derived arachidonic acid metabolite that has been shown to increase smooth muscle contractions and proliferation, stimulate endothelial dysfunction and activation, and promote hypertension. We examined if 20-HETE contributes to microvascular remodeling in hypertension. In Sprague-Dawley rats, administration of the 20-HETE biosynthesis inhibitor HET0016 or the 20-HETE antagonist N-20-hydroxyeicosa-6(Z),15(Z)-dienoic acid (20-HEDE) prevented 5α-dihydrotestosterone (DHT)-induced increases in blood pressure as well as abrogated DHT-induced increases in the media-to-lumen ratio (M/L), media thickness, and collagen IV deposition in renal interlobar arteries. Reserpine prevented blood pressure elevation in DHT-treated rats but did not affect microvascular remodeling (M/L, media thickness, and collagen deposition); under these conditions, treatment with the 20-HETE antagonist attenuated microvascular remodeling, suggesting that 20-HETE contributes to DHT-induced vascular remodeling independent of blood pressure elevation. In Cyp4a14(-/-) mice, which display androgen-driven and 20-HETE-dependent hypertension, treatment with the 20-HETE antagonist abolished remodeling of renal resistance arteries measured as media thickness (24 ± 1 vs. 15 ± 1 μm) and M/L (0.29 ± 0.03 vs. 0.17 ± 0.01). Moreover, in Cyp4a12 transgenic mice in which the expression of Cyp4a12-20-HETE synthase is driven by a tetracycline-sensitive promoter, treatment with doxycycline resulted in blood pressure elevation (140 ± 4 vs. 92 ± 5 mmHg) and a significant increase in remodeling of renal resistance arteries (media thickness: 23 ± 1 vs. 16 ± 1 μm; M/L: 0.39 ± 0.04 vs. 0.23 ± 0.02); these increases were abrogated by cotreatment with 20-HEDE. This study demonstrated that 20-HETE is a key regulator of microvascular remodeling in hypertension; its effect is independent of blood pressure elevation and androgen levels.