301 Sustained Hypertension Despite Endothelium-Specific Reintroduction of Functionally Active eNOS in eNOS-Deficient Mice
T Suvorava, VT Dao, M Oppermann, G Kojda
Institute of Pharmacology and Clinical Pharmacology, University Hospital, Duesseldorf, Germany
Topic: Hypertension
Purpose: Several studies have shown that eNOS-deficient mice (eNOS−/−) have higher blood pressure than wild-type mice. It is frequently assumed that hypertension in eNOS−/− is caused by the lack of endothelium-derived nitric oxide and the resulting increase in arterial tone and peripheral resistance. We sought to investigate whether endothelial-specific targeting of eNOS introduced in eNOS−/− can normalize aortic reactivity and blood pressure.
Methods: Transgenic mice carrying bovine eNOS (eNOSbov+) have been generated on C57Bl/6 background using the endothelium-specific Tie-2 promotor. By breeding these mice with eNOS knockouts (eNOS−/−), mice that only express eNOSbov (eNOS−/−/eNOSbov+) were obtained.
Results: Western blot analysis confirmed eNOS protein expression in aorta (67.7±12.6), myocardium (49.9±5.4), lung (124.5±2.5) and skeletal muscle (87.0±25.1) of eNOS−/−/ eNOSbov+ as compared to C57Bl/6 (100%, n=3–6). Aortas of eNOS−/−/eNOSbov+ showed complete restoration of endothelium-dependent relaxation to acetylcholine. the dose-response-patterns to acetylcholine did not differ significantly (P=0.562, n=7–11, two-way ANOVA), and the maximal relaxations were similar in eNOS−/−/eNOSbov+ (98.3±2.14 %, n=7) and C57Bl/6 mice (92.4±3.6 %, n=11) while no relaxation was observed in eNOS−/− (137.5±12.1, n=11). Hypersensitivity to phenylephrine observed in eNOS−/− mice (maximal contraction 15.2±0.7 mN, n=6, P>0.0001) vs C57Bl/6 (4.6±1.1mN, n=5) was blunted by endothelium-targeted reintroduction of eNOS (2.9±0.8, mN, n=6). Likewise, there was a significant increase in aortic sensitivity to NO-donors S-nitroso-N-acetyl-penicillamine and diethylamine/NO in eNOS−/− as compared to C57Bl/6 (n=4–5, P>0.05) and this was completely abolished in eNOS−/−/eNOSbov+ (n=4–5, P= 0.0278). the expression of both sGC-alpha1 and sGC-beta1 did not reveal any significant difference between eNOS−/−, eNOS−/−/eNOSbov+ and C57Bl/6 (n=4, each P>0.05, ANOVA). Despite complete restoration of aortic reactivity, eNOS−/−/eNOSbov+ mice have strongly elevated systolic blood pressure (n=137.1± 2.26 mmHg, n=8) as compared to C57Bl/6 (118.4±3.1 mmHg, n=6, P>0.05), but not to eNOS−/− (135.9±2.07, n=8, P=0.7).
Conclusions: Endothelium-specific reintroduction of functionally active eNOS in eNOS-deficient mice resulted in complete normalization of aortic reactivity but not blood pressure. these data suggest that eNOS appears to have limited effect on systemic blood pressure.
302: Regular exercise prevents calcific aortic valve disease in mice
Y Matsumoto, V Adams, S Jacob, N Mangner, G Schuler, A Linke
University of Leipzig, Heart Center, Leipzig, Germany
Topic: Exercise physiology, testing and training
Background: Currently no effective therapy to prevent calcific aortic valve (AV) disease is established. Hence, novel therapies and the optimum timing are active areas of investigation. Calcific AV disease confers significant morbidity and mortality as the severity of disease progresses. thus, as suggested by recent reports, adverse events can be avoided or delayed if it is possible to prevent the progression of AV disease at an earlier time point before presenting severe valve calcification (eg, AV sclerosis). Regular exercise training (ET) slows the progression of atherosclerotic lesions, reduces oxidative stress and increases nitric oxide bioavailability, all of which may be expected to improve degenerative AV disease. this study was designed to examine if ET prevents AV sclerosis, and if so, what cellular and molecular mechanisms are involved, using a well-established animal model revealing calcific AV disease.
Methods and Results: Four-week-old LDL receptor deficient (LDLR−/−) mice (n=94) were randomly divided into four groups: Group 1 (N), normal diet plus sedentary; group 2 (Chol), cholesterol(chol)-diet plus sedentary; group 3 (Reg), chol-diet plus regular ET for 16 wks (Treadmil, 60min/d, 5d/wk); group 4 (Occa), chol-diet plus occasional ET for 16wks (1d/wk). Histological analysis at 20-wk-old showed that AV thickness increased significantly in chol-group compared to N-group. Importantly, regular ET but not occasional ET significantly reduced AV thickness as compared tochol-group (N;31.3±3.0, Chol; 50.1±3.4, Reg;30.4±1.2, Occa; 48.9±3.2çm, P>0.001). Immunohistochemistory revealed that chol-diet disrupted and regular ET preserved endothelial integrity on the AV surface (N; 95.7±1.0, Chol; 73.7±2.2, Reg; 95.8±2.3%, P>0.001). Exercise did not affect cholesterol levels. However, plasma myeloperoxidase, accumulation of macrophages and oxidized-LDL, in-situ superoxide, myofibroblasts/osteoblasts phenotypes and mineralization at the AVs were markedly increased in chol-group and significantly decreased by regular ET. PCR revealed mRNA for Runx2 (osteogenic transcription factor) was increased in chol-group and significantly diminished by regular ET (P>0.05).
Conclusions: In the LDLR−/− mice, regular ET prevents calcific AV sclerosis by numerous mechanisms including preservation of valvular endothelial integrity, reduced inflammation or oxidative stress and finally decrease in mineralization through the inhibition of osteogenic pathway. Based on our new insights, regular ET may be useful for preventing calcific AV disease particularly at an early stage of disease.
303: the ER22/23EK polymorphism in the glucocorticoid receptor gene is associated with changes in aerobic capacity after training in CAD patients
T thomaes, MA thomis, S Onkelinx, L Vanhees
Catholic University of Leuven, Leuven, Belgium
Topic: Cardiovascular rehabilitation
Background and Objectives: Previous studies reported associations between polymorphisms of different genes with muscular strength, anaerobic phenotypes and body composition. Because of the correlation of these parameters with aerobic capacity, the aim of this study was to investigate the influence of polymorphismsassociated with muscular function on the aerobic capacity at baseline and after 3 months of physical training in CAD patients.
Methods: 935 CAD patients who performed an incremental exercise test until exhaustion were included in this study. Polymorphisms of Alpha-actinin 3 (Q523R, R577X), Myosine light chain kinase (C49T, C37885A), Activin-type II receptor B (Rs2268757), Follistatin (rs12152850, rs12153205, rs3756498), IGF-II (ApaI), AKT1 (G205T), Glucocorticoid receptor (ER22, 23EK, N363S, BcII), IL-15 Receptor a (PstI, BstNI, HpaII), AMP-deaminase (C34T) and Vitamin D receptor (BsmI, TaqI, ApaI) were detected using the invader assay. Genotype-phenotype association analyses were performed using ANCOVA. Haplotype analysis was performed for the different gene polymorphisms using PHASE software (>2% frequency).
Results: For the ER22/23EK polymorphisms of the glucocorticoid receptor gene, carriers of the mutant allele had a significantly greater absolute and relative increase in VO2peak after 3 months of training (Table 1). Significant differences were observed for the haplotypes in the Myosine light chain kinase gene, the Follistatin gene and the Glucocorticoid receptor gene. No significant differences were observed in the other polymorphisms and haplotype combinations.
Conclusions: the ER22/23EK polymorphism of the glucocorticoid receptor gene is associated with a 5% additional change in aerobic capacity after a three month training program in CAD patients. this supplementary improvement of themutant allele-carriers might be due to an improved muscular function as an endophenotype of aerobic capacity.
Results of the ER22/23EK polymorphism
Allele
VO2 baseline ± SE
Δ VO2 ± SE
Δ VO2 ± SE (%)
combination
(ml/min)
(ml/min)
E22E
G(856)
1712.7 ± 12.0
382.9 ± 8.5
23.9 ± 0.6
GA+A (60)
1782.4 ± 45.3
461.3 ± 32.0 ∗
28.2 ± 2.1 ∗
R23K
G (857)
1715.92± 12.0
382.8 ± 8.5
23.8 ± 0.5
GA+A (55)
1789.8 ± 47.6
461.7 ± 33.6∗
28.2 ± 2.2∗
∗p < 0.05 (corrected for gender, age, heigth, weigth and baseline VO2peak), n: number of patients.
304: Skeletal muscle metabolic recovery following submaximal exercise in chronic heart failure is limited more by O2 delivery than O2 utilization
HMC Hareld Kemps1, JJ Prompers2, B Wessels2, WR De Vries3, ML Zonderland3, K Nicolay2, G Schep1, PAFM Doevendans3
1Maxima Medical Centre, Veldhoven, Netherlands, 2Eindhoven University of Technology, Eindhoven, Netherlands, 3University Medical Center Utrecht, Utrecht, Netherlands
Topic: Exercise physiology, testing and training
Purpose: Chronic heart failure (CHF) is associated with a prolonged recovery of skeletal muscle energy stores following submaximal exercise, limiting the ability to perform repetitive daily activities. the purpose of this study was to investigate whether muscle metabolic recovery following submaximal exercise in patients with chronic heart failure (CHF) is limited by O2 delivery or O2 utilization.
Methods: thirteen stable CHF patients (New York Heart Association class II-III) and 8 healthy subjects, matched for age and body mass index were included. All subjects performed repetitive submaximal dynamic single-leg extensions in the supine position. Post-exercise phosphocreatine (PCr) resynthesis was assessed by 31P magnetic resonance spectroscopy, using the time constant of the PCr increase as an index of muscle metabolic recovery (Tau-PCr). Near-infrared spectroscopy was applied simultaneously, using the rate of decrease in deoxygenated hemoglobin (HHb) as an index of post-exercise muscle re-oxygenation (MRT-HHb).
Results: As expected, PCr recovery was slower in CHF patients than in control subjects (Tau-PCr: 47 ±10 sec versus 35 ±12 sec, p=0.04). HHb recovery kinetics were also prolonged in CHF patients (MRT-HHb: 74±41 sec versus 44±17 sec, p=0.04). In the patient group, HHb recovery kinetics were slower than PCr recovery kinetics (p=0.02), whereas no difference existed in the control group (p=0.32).
Conclusion: the prolonged metabolic recovery in CHF patients is associated with an even slower muscle tissue re-oxygenation, indicating a lower O2 delivery relative to metabolic demands. therefore we postulate that the impaired ability to perform repetitive daily activities in these patients depends more on a reduced muscle blood flow than on limitations in O2 utilization.
Kinetic parameters (∗ p < 0.05)
305: Intensive exercise training leads to improved endothelial function in school children
C Walther, V Adams, K Machalica, U Mueller, S Erbs, M Sonnabend, L Gaede, G Schuler
University of Leipzig, Heart Center, Leipzig, Germany
Topic: Physical activity
Background: Vascular and endothelial markers are known to have a long lasting impact on cardiovascular risk. Physical exercise has the potential to positively influence vascular and endothelial markers in healthy individuals and in patients with coronary artery disease. Aim of this study was to investigate the effect of intensive exercise on these markers in children and adolescents.
Methods: Vascular and endothelial markers (Granulocyte-colony stimulating factor (GCSF), monocyte chemotactic protein-1 (MCP1a), vascular endothelial growth factor (VEGF)), inflammatory markers (Interleukin –6(IL-6), CRP) and an anti-inflammatory marker (Interleukin –10 (IL-10)) were investigated in 36 students (mean age 12.1 ±0.1 years) with a high level of physical activity (PA) (> 15 hours/week) and in 77 students (mean age 11.8 ±0.1 years) with regular exercise lessons (CG) (2–3 hours/week). In addition lipid profile and anthropometric data were assessed and a treadmill exercise test with spirometry was performed. Endothelial function was assessed analysing the reactive hyperemic index (RHI) via a finger-tonometry.
Results: there was no significant difference regarding mean BMI and the lipid profile between both groups. Students of PA were significantly taller and had a significantly better physical fitness (VO2max) compared to children of CG (60.6 ±1.2 ml/kg∗min versus 47.3 ±0.6 ml/kg∗min, p>0.001). PA was associated with significantly higher levels of GCSF (17.9 ±4.1 in PA vs. 11.6 ±1.0 pg/ml in CG, p< 0.001), MCP1 (16.2 ±3.5 in PA vs.12.3 ±4.7 pg/ml in CG, p>0.01) and IL-10 (8.6 ±2.2 in PA vs. 4.7 ±0.6 pg/ml in CG, p>0.01). Furthermore children of PA had a significantly better endothelial function (RHI: 1.7 ±0.6) compared to the other students (RHI 1.47 ±0.8), p>0.05.
Conclusion: the amount of physical activity has a significant impact on markers of vascular function already in childhood. Primary prevention by means of increased physical activity during childhood is an important factor to improve the cardiovascular risk profile.
306: Effects of high intensity interval training versus moderate exercise on hemodynamic, metabolic, and neuro-humoral abnormalities of young normotensive Women at high familial risk for hypertension
EG Ciolac1, EA Bocchi2, LA Bortolotto2, VO Carvalho3, JM Greve1, GV Guimaraes2
1Laboratory of Kinesiology, Institute of Orthopedics and Traumatology do Hospital das Clinicas da Fac, Sao Paulo, Brazil, 2Heart Institute do Hospital das Clinicas da Faculdade de Medicina da Universidade de Sao Paulo, Sao Paulo, Brazil, 3Instituto da Crianoa do Hospital das Clinicas da Faculdade de Medicina da Universidade de Sao Paulo, Sao Paulo, Brazil
Topic: Hypertension
Background: Family history of hypertension is associated with higher risk of hypertension and subsequent cardiovascular disease. Moreover, off spring of essential hypertensive parents (FH+) have shown early metabolic and hemodynamic abnormalities associated to hypertension. Exercise training has an important role in the prevention and treatment of hypertension, but its effects on these early abnormalities observed in FH+ has not been studied. Our aim was to compare high-intensity interval (AIT) and moderate-intensity continuous exercise training (CMT) with regard hemodynamic, metabolic and hormonal variables in FH+.
Methods: Forty-four healthy women FH+ (25.0 ± 4.4 years), randomized to control (ConFH+) or to a 3 times-a-week equal volume AIT (80–90% of VO2PEAK) or CMT (50–60% of VO2PEAK), and 15 healthy women with normotensive parents (ConFH-; 25.3 ±3.1 years) had their ambulatorial blood pressure (ABP), carotid-femoral pulse wave velocity (PWV), and biochemistry (total cholesterol and fractions, triglycerides, glucose, insulin and insulin sensitivity) analyzed before and after a 16-week follow-up. BP, nor-epinephrine (NE), endothelin-1 (ET-1) and nitrite/nitrate (NOx) levels were also analyzed during a graded exercise test (GXT).
Results: ABP, glucose and cholesterol levels were similar among all groups, but FH+ groups displayed higher insulin, insulin sensitivity, PWV, NE and ET-1 levels, and lower NOx levels than ConFH-. AIT and CMT were equally effective in improving ABP (P < 0.05), insulin and insulin sensitivity (P < 0.001); however, AIT was superior in improving cardiorespiratory fitness (15% versus 8%; P < 0.05) and PWV (P < 0.01). BP, norepinephrine, ET-1 and NOx response to exercise also improved more after AIT than CMT (P < 0.05). All the variables did not change in ConFH+ and ConFH- after the follow-up. With the improvements, AIT group displayed insulin and insulin sensitivity, PWV, NE and ET-1 levels lower than ConFH+ and silimar to ConFH-, and NOx levels higher than ConFH+ and similar to ConFH-. In the CMT group, only ET-1 level were lower when compared to ConFH+.
Conclusion: Exercise intensity was an important factor for improving cardiorespiratory fitness and reversing hemodynamic, metabolic and hormonal alterations involved in the pathophysiology of HPT. these findings may have important implications for exercise training programs for the prevention of an inherited hypertensive disorder.
