Effects of NOS3 Glu298Asp Polymorphism on HemodynamicReactivity to Stress: Influences of Ethnicity and ObesitySurender Malhotra, Joseph Poole, Harry Davis, Yanbin Dong, Jennifer Pollock,Harold Snieder, Frank TreiberAbstract—Studies on the associations between the nitric oxide synthase gene (NOS3) Glu298Asp polymorphism andhypertension status or blood pressure (BP) levels have had inconsistent results. Potential moderating influences ofethnicity, sex, and obesity on the effects of the NOS3 polymorphism have not been examined. We evaluated theinfluence of these factors on associations between the NOS3 polymorphism, nitric oxide metabolites (NOx), andhemodynamics at rest and during stress. Subjects were 235 African American (AA) and 262 European American (EA)young adults (18.5⫾2.6 years). Hemodynamic measurements and blood samples for NOx assays were taken before andafter a competitive video game challenge. Glu298Asp polymorphism was detected by polymerase chain reaction–restriction enzyme digestion assay. A regression model was built using genotypes, ethnicity, sex, and obesity (body massindex ⬎85th percentile) and their interactions controlling for age; 20.1% of AAs and 49.8% of EAs were carriers of theAsp allele. AAs, regardless of obesity status, exhibited high diastolic blood pressure (DBP) reactivity unless they werenonobese and noncarriers of the Asp allele. EAs exhibited lower DBP reactivity unless they were obese Asp allelecarriers. AA nonobese carriers exhibited the greatest total peripheral resistance reactivity. Obese Asp allele carriersexhibited the greatest increases in cardiac output and the greatest decrease in NOx to the stressor. Results indicate theimportance of examining impact of BP control-related genetic polymorphisms within the context of moderating factorssuch as adiposity and ethnicity. (Hypertension. 2004;44:866-871.)Key Words: blood pressure䡲stress䡲nitric oxide䡲nitric oxide synthase䡲polymorphismNitric oxide (NO) is a potent vasodilator that plays asignificant role in vasomotor tone and in blood pressure(BP) control. Abrupt interruption of NO synthesis via phar-macological blockade markedly elevates BP in animals andhumans.1,2Individuals with essential hypertension have eitherdiminished whole-body NO production or increased inacti-vation leading to lower plasma levels.3,4NO is produced from L-arginine by nitric oxide synthase(NOS), which is a product of the NOS3 gene. The NOS3 geneis located on chromosome 7, spans 21 kb, and contains 26exons. Several studies have found an Asp for Glu substitutionin exon 7 at amino acid residue 298 (Glu298Asp, also calledG894T) associated with coronary spasm,5myocardial infarc-tion,6coronary artery disease,7and vascular responsiveness tophenylephrine.8The Asp variant of this polymorphism isbelieved to render the enzyme more susceptible to proteolyticcleavage.9There are contradictory results with regard to itsassociation with BP status. Some studies have found Aspallele carrier status to be associated with higher BP levels andincreased hypertension prevalence,10others have observedjust the opposite,11and still others have observed no signif-icant differences based on carrier status.12This inconsistencyacross studies has been partly attributed to lack of evaluationof possible modulating influences of subject characteristicsknown to differentially affect BP such as ethnicity, sex, age,and obesity.11Exaggerated BP reactivity to stress has been prospectivelylinked to BP levels and essential hypertension.13Males,African Americans (AAs), and obese individuals typicallyshow greater BP reactivity as opposed to females, EuropeanAmericans (EAs), and nonobese individuals, respective-ly.14 –16However, relatively little attention has been given tothe possible association of the Glu298Asp polymorphismwith BP reactivity to stress. Rankinen et al17showed thatamong participants in an exercise training study, carriers ofthe Asp allele showed a less beneficial training effect in BPreactivity to a submaximal exercise test compared withnoncarriers. The association between Asp carrier status andBP reactivity to behavioral stress has not been examined.A few studies have examined the relationship between theGlu298Asp polymorphism and plasma nitrite/nitrate (NOx)levels, metabolites of NO, and their findings have beenmixed. Asp carrier status has been associated with lowerbasal NOx levels in one study,18higher levels in a secondReceived May 7, 2004; first decision May 18, 2004; revision accepted October 1, 2004.From the Georgia Prevention Institute (S.M., J.P., H.D., Y.D., H.S., F.T.), Department of Pediatrics, Vascular Biology Center (J.S.P.), Medical Collegeof Georgia, Augusta, Ga; and the Twin Research and Genetic Epidemiology Unit (H.S.), St. Thomas’ Hospital, London, UK.Correspondence to Frank A. Treiber, Medical College of Georgia, Georgia Prevention Institute, Building HS1640, Augusta, GA 30912-3710. [email protected]© 2004 American Heart Association, Inc.Hypertension is available at http://www.hypertensionaha.org DOI: 10.1161/01.HYP.0000147578.84729.ac866study,19and a third study found no association.20To ourknowledge, associations between the Glu298Asp polymor-phism and behavioral stress induced changes in hemodynam-ic function and NOx levels have not been assessed.The purpose of the present study was to test the hypothesisthat carriers of the Asp allele, particularly those who weremales, AAs, or obese would show the highest levels of BPand lowest NOx levels at rest and in response to acutebehavioral stress. Because carriers of the Asp allele exhibittheir physiological effects through changes in production of apotent vasoactive substance, we hypothesized that the effectsof Asp carrier status would be mediated through changes invascular tone (ie, increased total peripheral resistance [TPR]).MethodsStudy PopulationA total of 497 young adults (235 AAs, 262 EAs; average age,18.5⫾2.6 years) participated in the study. Subjects are among theparticipants in a longitudinal study of the development of biobehav-ioral risk factors for cardiovascular disease.21All subjects have averified family history of cardiovascular disease (essential hyperten-sion and/or premature myocardial infarction).22All were normoten-sive and free of any chronic diseases.ProtocolThe study was approved by the institutional review board. Afterobtaining informed consent, subjects underwent a battery of anthro-pometric evaluations including