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(Circulation. 1995;91:270-274.)
© 1995 American Heart Association, Inc.

Articles

A DNA Variant at the Angiotensin-Converting Enzyme Gene Locus Associates With Coronary Artery Disease in the Caerphilly Heart Study

Raj K. Mattu, MB, MRCP; Edward W. A. Needham, BSc; David J. Galton, DSc; Evanthia Frangos; Adrian J. L. Clark, MB, MRCP; Mark Caulfield, MB, MRCP

From the Medical Professorial Unit (R.K.M., E.W.A.N., D.J.G., E.F.), the Department of Clinical Pharmacology (M.C.), and the Department of Chemical Endocrinology (A.J.L.C.), St Bartholomew's Hospital, London, UK.

Correspondence to Dr R.K. Mattu, Department of Cardiological Sciences, St George's Hospital Medical School, Cranmer Terrace, London, UK, SW17 0RE.

	Abstract

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Background We analyzed an insertion/deletion (I/D) polymorphism of the angiotensin I–converting enzyme (ACE) gene in 1226 subjects from the Caerphilly Prospective Heart Disease Study. Amplification of genomic DNA using the polymerase chain reaction yielded the genotypes II, ID, and DD. Distribution of the polymorphism was analyzed among the whole group and within subgroups (specified following multiple risk factor analysis) for coronary artery disease (CAD) and against multiple risk factors.

Methods and Results Allele frequencies were I=0.413 and D=0.587. No association was observed between the polymorphism and CAD in the whole group. Among subjects defined at lower risk of CAD by total cholesterol/HDL cholesterol (TC/HDL) ratios, we found significant associations of the DD genotype with CAD (P<.0053, n=586 for TC/HDL <5.654 [median] and P<.009, n=385 for TC/HDL <5.0 [clinical threshold]). On further exclusion of subjects with blood pressures 140/90 or on hypotensive medications, the DD genotype still associated with CAD (P<.07, n=210, TC/HDL <5.654 and P<.016, n=135, TC/HDL <5.0). Further stratification of risk incorporating other risk factors, except body mass index, did not alter or enhance this association. Although similar association was observed when risk was specified by using HDL and apo B levels instead of TC/HDL, this association was lost when body mass index was included in the low-risk stratification.

Conclusions The DD genotype is a linkage marker for an etiologic mutation at or near the ACE gene that may confer risk of CAD detectable in subjects previously unidentifiable with "classic" risk factors. However, this risk may be quantitatively small among the general male population.

Key Words: genes • angiotensin • coronary disease • enzymes

	Introduction

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Coronary artery disease (CAD) arises from a genetic predisposition interacting with environmental risk factors^{1 2 3 4} and in 1990 accounted for 366 deaths per 100 000 men 45 to 64 years old in Wales.⁵ The genes conferring susceptibility to CAD are unknown for the majority of sufferers. However, mutations have been identified in the LDL receptor gene that determine the onset of premature CAD in familial hypercholesterolemia.⁶

The genes encoding components of the renin-angiotensin system (RAS) present attractive candidates for cardiovascular disease. The RAS is present in circulating and tissue-based forms^{7 8 9} and is involved in sodium homeostasis, cardiovascular remodeling, and maintenance of vascular tone. Angiotensin I–converting enzyme (ACE) is a key component within the RAS, where it hydrolyzes angiotensin I to generate angiotensin II (vasoconstrictor)¹⁰ and the kallikrein-kinin system, where it inactivates bradykinin (vasodilator).^{9 11} The observation that ACE inhibitors reduce atherosclerosis in cholesterol-fed rabbits supports the potential role for ACE or its substrates in the development of atheroma.¹²

The ACE gene has been mapped to chromosome 17q23,^{13 14} and an insertion/deletion (I/D) polymorphism, involving a 287-bp alu repeat sequence, has been located to intron 16.^{15 16 17} This deletion polymorphism was found to associate with survivors of a myocardial infarction (MI) among apparently low-risk subjects.¹⁸ In the present study, we investigate whether this polymorphism is a marker for CAD per se and not just the infarct-surviving subgroup and determine whether this polymorphism predicts CAD.

	Methods

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Subjects
Men (n=1226) were genotyped from the latter half of the Caerphilly Prospective Heart Disease Study. Details of this cohort and CAD status are given elsewhere.¹⁹ Briefly, Caerphilly is a medium-sized town in southern Wales with a relatively constant population just exceeding 40 000 people, predominantly of Celtic descent. All men between the ages of 45 and 59 years (inclusive) in the Caerphilly region were selected by use of the electoral roll and general practitioner records as the basic sampling frame; 89% collaborated. They were screened for CAD risk factors. CAD status was classified by use of a Rose questionnaire and ECGs (Minnesota coding) into three groups: subjects with CAD (n=297), possible CAD (n=107), and no CAD (control group, n=822).¹⁹ Use of the Rose questionnaire and ECGs to classify CAD subjects for a large population-based study is discussed elsewhere.^{20 21 22} Angiography was not routinely performed because of ethical considerations. The control subgroup represents asymptomatic and apparently clinically disease-free subjects. Subjects classified as having CAD and possible CAD had a similar incidence of coronary events at the 5-year follow-up, defined as fatal and nonfatal MI's, surgical intervention, and angina (personal communication, P. Sweetnam, 1994). Thus, analyses of CAD subjects were undertaken on these two groups both separately and in combination (n=404).

Using multiple logistic regression, we stratified the risk factors and determined that the best predictors of CAD were the lipids, in particular the total cholesterol/HDL cholesterol (TC/HDL) ratio (consistent with previous work²³ ), and blood pressure. Subsequently, we identified subjects at low risk of CAD by sequential addition of risk factors. Here, we present only the data defining low risk using lipids and blood pressure, since addition of further risk factors did not alter or enhance the associations (with the exception of body mass index [BMI]) but only served to reduce the sizes of the subgroups. We examined the clinical threshold above which drug treatment is indicated (TC/HDL <5, n=385) and the median TC/HDL (<5.654, n=586) among the population after excluding one subject on lipid-lowering drug therapy. In combination with these lipid criteria, normotensive subgroups were defined by the World Health Organization threshold blood pressure of <140/90 (n=135 and 210, respectively), which also excluded subjects on hypotensive medications. We investigated the possibility of a blood pressure threshold effect by increasing both systolic and diastolic thresholds by 10 mm Hg to <150/100 (n=200 and 307, respectively). An additional subgroup was also analyzed with the current British Hypertension Society's drug treatment threshold of <160/100²⁴ (n=257 and 386, respectively). We also defined low-risk individuals by using other cardiovascular risk factors in conjunction with these lipid criteria (ie, smoking, glucose, insulin, BMI, fibrinogen, and platelet count). High-risk subjects were defined as all subjects not in the low-risk group.

Laboratory Measurements
Measurements of biochemical and hematologic parameters are described elsewhere.²⁵

Isolation of DNA
DNA was isolated from frozen EDTA whole blood by a sucrose lysis procedure, as described elsewhere.²⁶ It was redissolved in Tris-EDTA buffer (10 mmol/L Tris, 1 mmol/L EDTA, pH 7.6) and stored at -20°C.

Oligonucleotides
Primers for polymerase chain reaction (PCR) were synthesized by standard ß-cyanoethyl phosphoramidite chemistry with a Biotech BT 8500 DNA synthesizer. They were purified on a Sephadex G-25 column. The primer sequences are described elsewhere.¹⁷

Amplification of Genomic DNA
Genomic DNA (0.2 to 0.5 µg) was amplified in a 25-µL reaction mixture containing 10 mmol/L Tris/HCl (pH 8.3); 50 mmol/L KCl; 200 µmol/L each of dATP, dCTP, dGTP, and dTTP; 2.5 mmol/L MgCl₂; 0.5 µmol/L each primer; and 0.5 U Taq DNA polymerase (Perkin Elmer Cetus). The mixture was overlaid with mineral oil. All tubes, pipette tips, and buffers were autoclaved, and the reaction mixture was UV irradiated before addition of genomic DNA and Taq DNA polymerase to minimize contamination.²⁷ Blank controls, containing no genomic DNA, were run with each set of amplifications. The amplification cycle was performed on a Perkin Elmer Cetus 480 Thermal Cycler and entailed 5 minutes of denaturation at 96°C, followed by 35 cycles of 1 minute at 94°C and 2 minutes at 68°C. This was followed by 10 minutes of extension at 72°C.

Genotyping
The samples were visualized after electrophoresis on a 2% agarose gel and stained with ethidium bromide (0.5 mg/mL). Genotyping was undertaken blind.

Statistical Analysis
Genotype distributions between the study groups were analyzed by construction of 2x2 and 3x2 contingency tables and ² analysis. Variations in the biochemical traits with respect to genotypes were analyzed by ANCOVA, adjusted for age and BMI. Triglycerides, Lp(a), and insulin were log₁₀ transformed before analysis. Odds ratios, which measure relative risk, were calculated with the method of Woolf.²⁸ Analyses were performed with the MINITAB, SPSS/PC+, and SAS statistical packages. All risk factors were subject to discriminant analysis to determine their predictive power for CAD, thereby generating low- and high-risk groups for further analysis. We analyzed multiple risk factors following multiple logistic regression to test the predictive power of the I/D polymorphism in CAD.

	Results

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PCR amplification of genomic DNA produced fragments of 490 and/or 190 bp, yielding the genotypes II, ID, and DD, respectively. The genotype distributions in cases and control subjects were in Hardy-Weinberg equilibrium and are shown with allelic frequencies for all subjects in Table 1. The allele frequencies for the whole group were D=0.587 and I=0.413. We found no significant differences in mean lipid and lipoprotein levels with ACE genotypes in the cohort (Table 2). The mean systolic and diastolic blood pressures, smoking, BMI, fasting blood glucose, platelet count, fibrinogen levels, and fasting median insulin levels also showed no significant differences with respect to genotype (Table 3). Similarly, these parameters did not vary significantly with genotypes in either the control or CAD subgroups.

View this table: [in this window] [in a new window]   Table 1. Distribution of ACE Genotypes and Allelic Frequencies Among Caerphilly Cohort

View this table: [in this window] [in a new window]   Table 2. Comparison of ACE I/D Polymorphism With Lipid and Lipoproteins Among Caerphilly Cohort (n=1226)

View this table: [in this window] [in a new window]   Table 3. Comparison of ACE I/D Polymorphism With Other Cardiovascular Risk Factors Among Caerphilly Cohort (n=1226)

Table 4 lists the genotypic distributions, allele frequencies, and odds ratios for subjects at lower risk of developing CAD. On analysis of normolipidemic subjects, defined by TC/HDL <5, the DD genotype was significantly more frequent among the CAD subjects compared with control subjects: DD versus II, P=.0195, and DD versus (II + ID), P<.009. Similarly, the D allele was significantly more frequent in these CAD subjects compared with control subjects, P<.01. We observed similar associations of the DD genotype with CAD when we defined the low-risk subjects using TC/HDL < 5.654: DD versus II, P=.029, and DD versus (II + ID), P=.005.

View this table: [in this window] [in a new window]   Table 4. Distribution of ACE Genotypes and Allele Frequencies Among Caerphilly Low-Risk Group

When we defined low-risk individuals using other cardiovascular risk factors in addition to lipids, the association of the DD genotype with CAD was still significant when dyslipidemia (TC/HDL <5 ) and hypertension (blood pressure <140/90) were excluded (Table 4): P=.03 for DD versus II and P=.016 for DD versus (II + ID). On alteration of the risk conferred with blood pressure to a threshold of <150/100, the association of the DD genotype was increased considerably and observed for both TC/HDL ratios (Table 5).

View this table: [in this window] [in a new window]   Table 5. Distribution of ACE Genotypes and Allele Frequencies Among Caerphilly Low-Risk Group With Blood Pressure <150/100

Interestingly, the likelihood of identifying CAD subjects with this marker shows a progressive increase in relative risk (odds ratio) with the lower TC/HDL ratio of <5 (1.79 to 2.22) and offers greater prediction of risk with the addition of blood pressure criteria to identify low risk (2.22 to 4.96). Conversely, we found no significant genotypic associations when we defined low-risk individuals using BMI with either dyslipidemia or hypertension. We also found no significant associations between genotypes and CAD among the higher-risk subjects. Analysis of multiple risk factors using multiple logistic regression provided further information only on the effect of age on genotypic associations.

The mean age of subjects with CAD (57.7 years; range, 49 to 67 years) was significantly higher than that of subjects without disease (56.8 years; range, 49 to 66 years), P=.001 (Table 1). Despite this, there were no significant age differences between the genotypes (Table 3) and similarly no significant differences in genotypic distribution between different age groups. Furthermore, multiple logistic regression showed that the associations of the DD genotype with CAD were not a consequence of age differences between the CAD and the disease-free groups.

	Discussion

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This study reveals association of the DD genotype and D allele with CAD in men apparently at low risk and offers a potential predictive marker for CAD among men previously unidentifiable by classic risk factors. This observation significantly extends the putative role of this genetic variant from the highly selected subgroup surviving a myocardial infarction¹⁸ to the wider CAD phenotype.

Association of the DD genotype with CAD shows a progressive increase in relative risk using lower TC/HDL ratios (<5.0) and rises further on exclusion of treatable hypertension. Although HDL and apo B were not the best discriminants for CAD, we also analyzed them in conjunction with other risk factors to define low-risk subjects, as a result of the recent report by Cambien et al.¹⁸ We used the control group's median apo B (<96 mg/dL) and HDL (>0.98 mmol/L) levels and noted that the frequency of the DD genotype was again significantly higher in the CAD subjects (n=345, P=.002; data not shown).

The striking increase in significance of association observed with the blood pressure threshold <150/100 (Table 5) compared with 140/90 (Table 4) probably arises from the larger numbers of subjects in the former cohort. However, the significance of the associations with the DD genotype is lost when a blood pressure threshold >160/100 (data not shown) is used. This suggests that the DD variant is predictive of CAD only in low-risk individuals because it does not identify CAD subjects possessing classic risk factors. This may result because the DD genotype confers risk that is quantitatively small compared with established risk factors, thereby accounting for the absence of a detectable significant association in our entire cohort and the high-risk group.

The ACE I/D polymorphism associates with CAD independently of mean systolic and diastolic blood pressure, fasting glucose, fasting insulin, fibrinogen, and platelet levels. Intriguingly, when individuals are stratified into those with BMI below the median, all association of the DD genotype disappears. This may reflect the weakness of BMI as a distinguishing risk factor among low-risk individuals, and a centripetal distribution may prove a better predictor of cardiovascular risk. This contrasts with the observation among survivors of MI¹⁸ and could represent differential influence of BMI on this complication of CAD.

ACE levels in healthy men can differ greatly (up to fivefold) between individuals but appear stable when repeatedly measured within a given individual^{29 30} and show familial resemblance.³¹ Environmental and hormonal determinants of ACE levels have not been found to account for this interindividual variability.³² Conversely, a major gene, in linkage disequilibrium with the ACE I/D polymorphism, accounts for up to 44% of this variability.^{31 33} Individuals with the DD genotype have levels approximately twice those of the II variant.¹⁵ The overall allele frequencies observed in our study (D=0.587 and I=0.413) are similar to those calculated from combined segregation and linkage analyses for a proposed etiological mutation at the ACE gene locus (having frequencies of 0.557 and 0.431) that accounts for variations in plasma ACE levels.³³ Therefore, it is possible that the association seen in our study arises from overactivity of the RAS or an alteration in the balance with the kallikrein-kinin system. ACE appears to influence the cardiovascular system at many sites and in multiple ways.^{34 35 36} Evidence for deleterious cardiovascular effects mediated through ACE emerges from several studies,^{37 38 39 40 41 42 43 44} although the mechanisms remain unclear. In particular, the Survival and Ventricular Enlargement Study (SAVE)³⁷ and Studies of Left Ventricular Dysfunction (SOLVD)⁴⁴ showed that postinfarction ACE inhibition not only reduced progression to heart failure but offered secondary protection against reinfarction. Recently, the DD genotype has been associated with dilated⁴⁵ and hypertrophic⁴⁶ cardiomyopathy (particularly sudden death), thus increasing the likelihood that ACE-dependent cardiovascular remodeling may be a key mechanism in these cardiovascular disorders. Because the genotypic effect is independent of hypertension in our study, a direct influence of ACE or its substrates on cardiovascular tissues, including atheroma, are plausible mechanisms. Supportive evidence for this view emerges from the effective reduction of atherosclerosis by ACE inhibitors in animal models, but not by an angiotensin type I receptor antagonist.¹² Although this observation does not exclude atherogenesis through other angiotensin receptors, it emphasizes the need to consider bradykinin.

Our study supports the hypothesis that the DD genotype of the ACE gene is a linkage marker for an underlying etiological mutation, which confers risk for development of CAD that is detectable in subjects previously unidentifiable by classic risk factors.

	Acknowledgments

 
We are grateful for support from the Joint Research Board of St Bartholomew's Hospital. Dr Mattu is supported by a grant from the McAlpine Trust. Dr Clark is supported by the Medical Research Council (MRC). We are extremely grateful to Dr P.C. Elwood and members of the MRC Epidemiology Unit, Cardiff, UK, who supplied blood samples and clinical data from subjects in the Caerphilly Prospective Heart Disease Study. We are also grateful for the advice and help provided by H. Watt and A. Hackshaw from the Statistics Department of the MRC Institute of Preventative Medicine, London.

Received May 23, 1994; accepted August 19, 1994.

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				H. Volzke, J. Engel, V. Kleine, C. Schwahn, J. B. Dahm, L. Eckel, and R. Rettig Angiotensin I-Converting Enzyme Insertion/Deletion Polymorphism and Cardiac Mortality and Morbidity After Coronary Artery Bypass Graft Surgery* Chest, July 1, 2002; 122(1): 31 - 36. [Abstract] [Full Text] [PDF]

				T. Isbir, B. Agachan, H. Yilmaz, M. Aydin, I. Kara, D. Eker, and E. Eker Interaction between apolipoprotein-E and angiotensin-converting enzyme genotype in Alzheimer's disease American Journal of Alzheimer's Disease and Other Dementias, July 1, 2001; 16(4): 205 - 210. [Abstract] [PDF]

				T. Mannami, T. Katsuya, S. Baba, N. Inamoto, K. Ishikawa, J. Higaki, T. Ogihara, and J. Ogata Low Potentiality of Angiotensin-Converting Enzyme Gene Insertion/Deletion Polymorphism as a Useful Predictive Marker for Carotid Atherogenesis in a Large General Population of a Japanese City : The Suita Study Stroke, June 1, 2001; 32(6): 1250 - 1256. [Abstract] [Full Text] [PDF]

				G. P. Rossi, S. Taddei, A. Virdis, L. Ghiadoni, G. Albertin, S. Favilla, I. Sudano, A. C. Pessina, and A. Salvetti Exclusion of the ACE D/I Gene Polymorphism as a Determinant of Endothelial Dysfunction Hypertension, February 1, 2001; 37(2): 293 - 300. [Abstract] [Full Text] [PDF]

				J. W. Knowles and N. Maeda Genetic Modifiers of Atherosclerosis in Mice Arterioscler. Thromb. Vasc. Biol., November 1, 2000; 20(11): 2336 - 2345. [Abstract] [Full Text] [PDF]

				A. Prasad, S. Narayanan, S. Husain, F. Padder, M. Waclawiw, N. Epstein, and A. A. Quyyumi Insertion-Deletion Polymorphism of the ACE Gene Modulates Reversibility of Endothelial Dysfunction With ACE Inhibition Circulation, July 4, 2000; 102(1): 35 - 41. [Abstract] [Full Text] [PDF]

				C Fatini, R Abbate, G Pepe, B Battaglini, F Gensini, G Ruggiano, G.F Gensini, and R Guazzelli Searching for a better assessment of the individual coronary risk profile. The role of angiotensin-converting enzyme, angiotensin II type 1 receptor and angiotensinogen gene polymorphisms Eur. Heart J., April 2, 2000; 21(8): 633 - 638. [Abstract] [PDF]

				B. Agerholm-Larsen, B. G. Nordestgaard, and A. Tybjarg-Hansen ACE Gene Polymorphism in Cardiovascular Disease : Meta-Analyses of Small and Large Studies in Whites Arterioscler. Thromb. Vasc. Biol., February 1, 2000; 20(2): 484 - 492. [Abstract] [Full Text] [PDF]

				B. T. Heijmans, R. G. J. Westendorp, D. L. Knook, C. Kluft, and P. E. Slagboom Angiotensin I-converting enzyme and plasminogen activator inhibitor-1 gene variants: risk of mortality and fatal cardiovascular disease in an elderly population-based cohort J. Am. Coll. Cardiol., October 1, 1999; 34(4): 1176 - 1183. [Abstract] [Full Text] [PDF]

				M. Pfohl, M. Koch, S. Prescod, K.K. Haase, H.U. Haring, and K.R. Karsch Angiotensin I-converting enzyme gene polymorphism, coronary artery disease and myocardial infarction. An angiographically controlled study Eur. Heart J., September 2, 1999; 20(18): 1318 - 1325. [Abstract] [PDF]

				S. T. Bogardus Jr, J. Concato, and A. R. Feinstein Clinical Epidemiological Quality in Molecular Genetic Research: The Need for Methodological Standards JAMA, May 26, 1999; 281(20): 1919 - 1926. [Abstract] [Full Text] [PDF]

				G. I. Rice, C. A. Foy, and P. J. Grant Angiotensin converting enzyme and angiotensin II type 1-receptor gene polymorphisms and risk of ischaemic heart disease Cardiovasc Res, March 1, 1999; 41(3): 746 - 753. [Abstract] [Full Text] [PDF]

				M. J. MALIARIK, B. A. RYBICKI, E. MALVITZ, R. G. SHEFFER, M. MAJOR, J. POPOVICH Jr., and M. C. IANNUZZI Angiotensin-converting Enzyme Gene Polymorphism and Risk of Sarcoidosis Am. J. Respir. Crit. Care Med., November 1, 1998; 158(5): 1566 - 1570. [Abstract] [Full Text] [PDF]

				C. J. O'Donnell, K. Lindpaintner, M. G. Larson, V. S. Rao, J. M. Ordovas, E. J. Schaefer, R. H. Myers, and D. Levy Evidence for Association and Genetic Linkage of the Angiotensin-Converting Enzyme Locus With Hypertension and Blood Pressure in Men but Not Women in the Framingham Heart Study Circulation, May 19, 1998; 97(18): 1766 - 1772. [Abstract] [Full Text] [PDF]

				D.-K. Kim, J.-W. Kim, S. Kim, H.-C. Gwon, J.-C. Ryu, J.-E. Huh, J.-A Choo, Y. Choi, C.-H. Rhee, and W.-R. Lee Polymorphism of Angiotensin Converting Enzyme Gene Is Associated With Circulating Levels of Plasminogen Activator Inhibitor-1 Arterioscler. Thromb. Vasc. Biol., November 1, 1997; 17(11): 3242 - 3247. [Abstract] [Full Text]

				M. Margaglione, E. Grandone, G. Vecchione, G. Cappucci, N. Giuliani, D. Colaizzo, E. Celentano, S. Panico, and G. Di Minno Plasminogen Activator Inhibitor-1 (PAI-1) Antigen Plasma Levels in Subjects Attending a Metabolic Ward: Relation to Polymorphisms of PAI-1 and Angiontensin Converting Enzyme (ACE) Genes Arterioscler. Thromb. Vasc. Biol., October 1, 1997; 17(10): 2082 - 2087. [Abstract] [Full Text]

				N. Iwai, S. Tamaki, N. Ohmichi, and M. Kinoshita The II Genotype of the Angiotensin-Converting Enzyme Gene Delays the Onset of Acute Coronary Syndromes Arterioscler. Thromb. Vasc. Biol., September 1, 1997; 17(9): 1730 - 1733. [Abstract] [Full Text]

				K. Hibi, T. Ishigami, K. Kimura, M. Nakao, T. Iwamoto, K. Tamura, T. Nemoto, T. Shimizu, Y. Mochida, H. Ochiai, et al. Angiotensin-Converting Enzyme Gene Polymorphism Adds Risk for the Severity of Coronary Atherosclerosis in Smokers Hypertension, September 1, 1997; 30(3): 574 - 579. [Abstract] [Full Text]

				B. Tian, Q. C. Meng, Y.-F. Chen, J. H. Krege, O. Smithies, and S. Oparil Blood Pressures and Cardiovascular Homeostasis in Mice Having Reduced or Absent Angiotensin-Converting Enzyme Gene Function Hypertension, July 1, 1997; 30(1): 128 - 133. [Abstract] [Full Text]

				B. Agerholm-Larsen, B. G. Nordestgaard, R. Steffensen, T. I.A. Sorensen, G. Jensen, and A. Tybjærg-Hansen ACE Gene Polymorphism: Ischemic Heart Disease and Longevity in 10 150 Individuals: A Case-Referent and Retrospective Cohort Study Based on the Copenhagen City Heart Study Circulation, May 20, 1997; 95(10): 2358 - 2367. [Abstract] [Full Text]

				J. H. Krege, H.-S. Kim, J. S. Moyer, J. C. Jennette, L. Peng, S. K. Hiller, and O. Smithies Angiotensin-Converting Enzyme Gene Mutations, Blood Pressures, and Cardiovascular Homeostasis Hypertension, January 1, 1997; 29(1): 150 - 157. [Abstract] [Full Text] [PDF]

				N. J. Samani, J. R. Thompson, L. O'Toole, K. Channer, and K. L. Woods A Meta-analysis of the Association of the Deletion Allele of the Angiotensin-Converting Enzyme Gene With Myocardial Infarction Circulation, August 15, 1996; 94(4): 708 - 712. [Abstract] [Full Text]

D. R.J. Singer, C. G. Missouris, and S. Jeffery Angiotensin-Converting Enzyme Gene Polymorphism: What to Do About All the Confusion? Circulation,