Ultrasound Screening of Men with Coronary Artery Disease for Abdominal Aortic Aneurysms: A Prospective Dual Center Study.

BACKGROUND AND AIMS: According to the heterogeneous results of previous studies, the prevalence of abdominal aortic aneurysm seems high among men with coronary artery disease. The associating risk factors for abdominal aortic aneurysm in this population require clarification. Our objective was to assess the prevalence of non-diagnosed abdominal aortic aneurysms in men with angiographically verified coronary artery disease and to document the associated co-morbidities and risk factors. MATERIAL AND METHODS: Altogether, 407 men with coronary artery disease were screened after invasive coronary angiography in two series at independent centers. Risk factor data were recorded and analyzed. RESULTS AND CONCLUSION: The mean age of the study cohort was 70.0 years (standard deviation: 11.0). The prevalence of previously undiagnosed abdominal aortic aneurysms in the whole screened population of 407 men was 6.1% (n = 25/407). In a multivariate analysis of the whole study population, the only significant risk factors for abdominal aortic aneurysm were age (odds ratio: 1.04, 95% confidence interval: 1.00-1.09) and history of smoking (odds ratio: 3.13, 95% confidence interval: 1.26-7.80). Non-smokers with abdominal aortic aneurysm were significantly older than smokers (mean age: 80.7 (standard deviation: 8.0) vs 68.0 (standard deviation: 11.1), p = 0.003), and age was a significant risk factor only among non-smokers (p = 0.011; p = 0.018 for interaction). Among smokers, the prevalence of abdominal aortic aneurysm was 8.8%, and 72% (n = 18/25) of all diagnosed abdominal aortic aneurysm patients were smokers. Prevalence of undiagnosed abdominal aortic aneurysms among patients with coronary artery disease is high, and history of smoking is the most significant risk factor for abdominal aortic aneurysm. Effectiveness of selective screening of abdominal aortic aneurysm in male patients with coronary artery disease warrants further studies.

Improvement of myocardial blood flow by lipid-lowering therapy with pravastatin is modulated by apolipoprotein E genotype.

OBJECTIVE: Apolipoprotein E (apoE) polymorphism affects the risk of advanced coronary artery disease, but its role in early atherosclerosis remains unknown. We used positron emission tomography (PET) to study whether coronary reactivity or its response to pravastatin is related to the apoE genotype. MATERIAL AND METHODS: Samples from 44 mildly hypercholesterolaemic men (aged 35 +/- 4 years) of an earlier trial were re-analysed according to apoE genotype. Subjects were randomized to receive either 40 mg/day pravastatin or placebo for 6 months. To assess coronary reactivity, myocardial blood flow was measured by PET at rest and during adenosine infusion. PET studies and lipid analyses were done at baseline and after 6 months of therapy. RESULTS: There were no differences between apoE epsilon3/3 and epsilon4/3 genotypes in basal or adenosine-stimulated flow or in coronary flow reserve (CFR) at baseline. There was a significant apoE genotype-by-treatment group interaction regarding the change in adenosine-stimulated flow (ANCOVA; p = 0.018) and CFR (p = 0.020) at the end of the study. In the pravastatin group, the adenosine-stimulated flow increased by 32.5 % in subjects with epsilon3/3 (n = 9), but decreased non-significantly (-14.4 %) in subjects with epsilon4/3 (n = 9) (p = 0.0009). The corresponding changes in CFR were +17.8 % for epsilon3/3 and (-11.9 % for epsilon4/3 (p = 0.05). There were no significant changes from the baseline values in placebo recipients. After pravastatin treatment, both genotype groups showed a similar decrease in serum total and low-density lipoprotein cholesterol (p<0.0001 for both). CONCLUSIONS: Coronary function improves by 6 months of pravastatin in subjects with the apoE epsilon3/3 genotype, but not in those with the epsilon4/3.

The hepatic lipase gene C-480T polymorphism in the development of early coronary atherosclerosis: the Helsinki Sudden Death Study.

BACKGROUND: The T allele of the hepatic lipase (HL) C-480T polymorphism was previously found to be associated with lower post-heparin plasma HL activity, atherosclerosis and risk of coronary artery disease. We studied the association of HL C-480T polymorphism with the extent of atherosclerosis at vessel-wall level in an autopsy series of middle-aged men. MATERIALS AND METHODS: An autopsy cohort of 700 Caucasian Finnish men aged 33-70 years (mean 53 years), which comprised two autopsy series, collected 10 years apart during 1981-82 and 1991-92, were analysed. Areas of coronary wall covered with fatty streaks and fibrotic and complicated lesions were measured using computer-assisted planimetry and related to HL C-480T genotypes (CC, CT, and TT). RESULTS: There was a significant age-by-genotype interaction on the mean percentage area of fatty streaks (P = 0.01). The HL C-480T polymorphism was a significant explanatory factor for fatty streak area in men under 53 years of age with or without age, body mass index, hypertension, diabetes, smoking, alcohol consumption, apolipoprotein E genotype, and series number as covariates. Men carrying the TT genotype had two times larger areas of fatty streaks compared to the CC carriers (8.8% vs. 4.3%, P = 0.009). However, this association disappeared in men over 53 years. The areas of more advanced atherosclerotic lesions did not vary significantly among the genotype groups. CONCLUSIONS: Our results suggest that the HL C-480T polymorphism affects the formation of early coronary atherosclerotic lesions in men in their early middle age.

Toll-like receptor 4 polymorphism is associated with coronary stenosis but not with the occurrence of acute or old myocardial infarctions.

OBJECTIVE: Atherosclerosis is considered to be a chronic inflammatory disease. Toll-like receptor 4 (TLR-4), a key mediator in activating inflammatory cascade, has an A-to-G functional polymorphism that changes aspartic acid to glycine at position 299. TLR-4 is activated by, for example, lipopolysaccharides. The purpose of this study was to investigate the role of a common Asp299Gly polymorphism of the TLR-4 gene in atherosclerosis. MATERIAL AND METHODS: The study comprised autopsy material from 657 men (the Helsinki Sudden Death Study; mean age 53, range 33-70 years). RESULTS: Fewer G-allele carriers had 3-vessel coronary artery disease compared with AA homozygotes (OR 0.32; 95 % CI, 0.12-0.88, p = 0.027), and they also had a lower mean value for maximal coronary stenosis (p = 0.019). TLR-4 polymorphism was not significantly associated with the occurrence of acute or old myocardial infarction (MI). CONCLUSIONS: The G allele of the TLR-4 gene, which is associated with a lower inflammation response, was associated with a lower risk of coronary stenosis but not with the occurrence of MI and hence is not a major factor in the development of coronary atherosclerosis.

Radiographic assessment of dental health in middle-aged men following sudden cardiac death.

Poor oral health has been suggested to be a risk factor for myocardial infarction. To study if dental pathology might predispose to pre-hospital sudden cardiac death, and using a sum index of panoramic tomography findings, we compared the oral health of middle-aged (33-69 yrs) male victims (Helsinki Sudden Death Study) of sudden cardiac death (n = 117) with that of controls, who died of non-cardiac diseases (n = 63) or suffered unnatural sudden death (n = 120). The mean number of teeth was 15.2, and 17.4% of the men were edentulous. Frequent age-associated findings in dentate victims were fillings (79.9%), horizontal bone loss (72.1%), periapical lesions (45.6%), residual roots (38.2%), and vertical pockets (30.9%). In multivariate analysis with coronary heart disease risk factors and number of teeth as covariates, poor oral health was associated (p = 0.053) with the risk of sudden cardiac death along with age, smoking, and body mass index. This association was especially strong (p = 0.009) among victims < 50 yrs.

A functional variant of the iNOS gene flanking region is associated with LAD coronary artery disease: an autopsy study.

BACKGROUND: Recent studies using reporter gene constructs have indicated significant differences in the promoter activity of inducible nitric oxide synthase (iNOS) gene variants. Although the exact role of iNOS in atherogenesis is unclear, it is possible that this variation site may influence the extent of coronary artery disease (CAD). METHODS: We amplified these (AAAT) repeat variants from the NOS2A gene (denoted iNOS R4 and iNOS R5) from 325 Finnish men included in the Helsinki Sudden Death Study, and studied their association with indices of stenosis and atherosclerosis of the left anterior descending artery (LAD), right coronary artery (RCA) and left circumflex artery (LCX). In order to understand the effect of iNOS genotype on different stages of CAD, our study population was divided into age groups. RESULTS: In the LAD, the progression of atherosclerosis seemed to be more pronounced in the 4/5 genotype carriers than in those with the 4/4 genotype when the different age groups were compared. More specifically, statistically significant differences between the genotypes were found in the subgroup of men aged > 55 years. In this group, men carrying the rare R4/5 genotype presented higher mean values of stenosis percentages (55% vs. 42%, P = 0.008), larger areas of fatty streaks (10.4% vs. 5.9%; P = 0.01) and complicated lesions (3.5% vs. 1.3%; P = 0.001) compared with the R4/4 carriers. No significant association of iNOS genotypes with stenosis and atherosclerosis of RCA and LCX was found. CONCLUSIONS: It appears unlikely the R4/5 genotype plays a major role in the pathogenesis of CAD, as it was not associated with stenosis and atherosclerosis in RCA and LCX. However this genotype may have some role in more pronounced CAD, as seen in the LAD.

Fibrinogen gene promoter -455 A allele as a risk factor for lacunar stroke.

BACKGROUND AND PURPOSE: Elevated fibrinogen levels are suggested to increase the risk of myocardial infarction and stroke. Carriers of the A allele of the fibrinogen -455G/A polymorphism have increased plasma fibrinogen levels. We studied the association of this polymorphism with stroke subtype in the Stroke Aging Memory (SAM) cohort. METHODS: The SAM cohort comprises 486 consecutive patients 55 to 85 years of age who, 3 months after ischemic stroke, completed a detailed stroke assessment. Stroke subtypes were examined with MRI. -455G/A genotype was determined by polymerase chain reaction. MRI and genotype data were available for the 299 patients who constitute the present study population. RESULTS: Genotype distributions were 64.9% (GG), 31.8% (GA), and 3.3% (AA). In a logistic regression model with age, sex, hypertension, diabetes, hypercholesterolemia, hypertriglyceridemia, myocardial infarction, arrhythmia, atrial fibrillation, peripheral arterial disease, and smoking as possible confounders, there was a significant association between A+ genotype and >or=3 lacunar infarcts (odds ratio [OR], 2.57; 95% CI, 1.23 to 5.36; P=0.01). Hypertensive patients carrying the A allele had increased risk (OR, 4.24; 95% CI, 1.29 to 13.99; P=0.02) for >or=3 lacunar infarcts. A similar increase in risk was observed among smokers with the A+ genotype (OR, 2.67; 95% CI, 0.92 to 7.77; P=0.07). CONCLUSIONS: Stroke patients carrying the A allele of the Bbeta-fibrinogen -455G/A polymorphism frequently presented with multiple lacunar infarcts. This association was stronger among hypertensives and smokers. These associations suggest that the A allele may predispose to atherothrombotic events in cerebrovascular circulation.

Methylenetetrahydrofolate reductase gene C677T mutation is related to the defects in the internal elastic lamina of the artery wall.

BACKGROUND: The C677T mutation of the methylenetetrahydrofolate reductase (MTHFR) gene leads to C/C, C/T and T/T genotypes, which affect the plasma homocysteine concentration in humans. In mini-pigs, high serum homocysteine levels are associated with defects in the internal elastic lamina (IEL) of the artery wall, which are apparently related to the migration of smooth muscle cells into the intima during atherogenesis. We studied the association between the MTHFR genotypes and the number of gaps in the IEL in the wall of the five major abdominal arteries. MATERIALS AND METHODS: The autopsy study included 123 subjects (90 males and 33 females) aged 18-93. For the light microscopy, a 0.5 cm circular segment of the coeliac, the superior mesenteric, the inferior mesenteric and the renal arteries were cut and embedded in paraffin blocks. The circumference of the IEL, the thickness of the intima and the number of the gaps per millimetre in the IEL were measured by MOP 3 image analysis. RESULTS: The T-allele carriers (C/T and T/T) of the MTHFR gene had significantly less gaps in the IEL than the subjects with the C/C genotype in the superior mesenteric and in the left renal arteries (2.02 +/- 2.25 vs. 2.53 +/- 1.89, P < 0.04 and 0.56 +/- 1.09 vs. 1.82 +/- 2.66, P < 0.02, respectively). The trend was similar for the coeliac and the right renal arteries. CONCLUSIONS: Our result suggests that MTHFR polymorphism may be involved in the fragmentation of the IEL.

Association of neuropeptide y polymorphism with the occurrence of type 1 and type 2 alcoholism.

BACKGROUND: The susceptibility to alcoholism can be explained partially by genetic factors. Neuropeptide Y (NPY) has emerged as one potential factor contributing the development of alcoholism. A recent study indicated that the NPY gene variant producing a leucine-to-proline substitution (T to C at position 1128) was associated with 34% higher average alcohol consumption. METHODS: The subjects consisted of 122 alcoholics classified as type 1 and type 2 subtypes by psychiatric evaluation. A random sample of 59 social drinkers was used as a control group to compare the distribution of NPY genotypes with those of alcoholics. RESULTS: In a logistical regression model, there was a significantly lower frequency of the leucine(7)/proline(7) heterozygotes among well characterized type 2 alcoholics, compared with the controls (10.8 vs. 24.1%, p = 0.028). CONCLUSIONS: We speculate that the genetic polymorphism producing the proline(7) substitution of NPY might not predispose to alcoholism, but indeed retard the transition to alcoholism.

Association of apolipoprotein E polymorphism with outcome after aneurysmal subarachnoid hemorrhage: a preliminary study.

BACKGROUND AND PURPOSE: Variation in the outcome after aneurysmal subarachnoid hemorrhage (SAH) is not fully explained by known prognostic factors. APOE genotype is the most important genetic determinant of susceptibility to Alzheimer's disease, and it is also shown to be associated with the outcome after traumatic brain injury. We studied the association of apolipoprotein E polymorphism with the outcome after aneurysmal SAH. METHODS: A total of 160 consecutive patients were admitted after SAH to a neurosurgical unit. The clinical assessment after the SAH was performed with the Hunt and Hess grading scale. The severity of the bleeding as visualized on CT was assessed by Fisher's grading system. Outcome was assessed with the Glasgow Outcome SCALE: APOE genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism. RESULTS: 126 patients had aneurysmatic SAH, and detailed information on outcome and APOE genotype was available for 108 patients (86%). Sixteen (40%) of 40 patients with APOE epsilon4 had an unfavorable outcome compared with 13 (19%) of 68 without the APOE epsilon4 allele (OR 2.8, 95% CI 1.18 to 6.77). Association was more significant after adjustment for age, rebleeding, clinical status on admission, and CT scan findings (OR 7.1, 95% CI 1.9 to 26.3; P=0.0035). CONCLUSIONS: Our findings show a significant genetic association of APOE polymorphism with outcome after spontaneous aneurysmal SAH. Genetic factors thus seem to explain a part of individual differences in the recovery of SAH.

Polymorphisms within the tumor necrosis factor locus and prevalence of coronary artery disease in middle-aged men.

Tumor necrosis factor (TNF) is an important cytokine in the inflammation process of atherosclerosis and is also involved in lipid metabolism. Two biallelic polymorphisms within TNF gene locus-TNFA at the position -308 in the promoter region of the TNF gene and TNFB in the first intron of the lymphotoxin-alpha (LT-alpha) have been reported to be associated with TNF production and with susceptibility to inflammatory diseases. We studied the association of these polymorphisms within the major histocompatibility complex (MHC) III region with coronary atherosclerosis and its manifestations. The autopsy series comprised 700 Caucasian Finnish men, aged 33-70 years (The Helsinki Sudden Death Study). Coronary stenosis and surface area of atherosclerotic changes (fatty streaks, fibrous plaques, complicated lesions and calcification) were measured and the presence of myocardial infarction and coronary thrombosis recorded. TNFA and TNFB genotypes were determined by the PCR-RFLP technique. The allele frequencies were TNFA1/TNFA2=0.88/0.12 and TNFB1/TNFB2=0.30/0.70. There was a strong linkage disequilibrium between the two polymorphisms. There were no differences in coronary stenosis and in the frequency of old or recent myocardial infarction or coronary thrombosis between men with different genotype status in either locus. Men with TNFA22 or TNFB11 genotype tended to have more fibrous lesions and calcification in their coronary arteries. TNFA and TNFB polymorphisms are unlikely to contribute to progression of atherosclerosis in a way clinically important.

Apolipoprotein E polymorphism and carotid artery intima-media thickness in a random sample of middle-aged men.

Genetic polymorphism of apolipoprotein E (apoE) is an important factor in the development of coronary artery disease but the results concerning apoE genotype and carotid artery atherosclerosis remain controversial. We investigated a random sample of 189 Finnish middle aged men (mean age 54 years, range 50-59) to assess the role of apoE in the process of carotid atherosclerosis. Intima-media thickness (IMT) of the carotid artery wall was measured at three standardised segments (common carotid artery, bifurcation and internal carotid artery) by B-mode ultrasonography. Overall mean IMT value was also calculated. The carriers of E3/2 (n=20) genotype had significantly lower (P<0.01) total cholesterol and LDL cholesterol concentrations than carriers of E3/3 genotype (n=109) or the E4 allele (n=60). ApoE polymorphism was associated with common carotid artery IMT (P=0.034) when adjusted for age and body-mass index (model 1). The carriers of E3/2 had on average 9% (95% CI 0.8-16%, P=0.028) lower common carotid IMT values than the carriers of E3/3. After further adjustment with LDL and HDL cholesterol, systolic blood pressure, lipoprotein (a), apolipoprotein B and pack-years of smoking (model 2) the association was not statistically significant. The overall mean IMT varied significantly with apoE genotype (P=0.03 for model 1 and P=0.07 for model 2), and it was also lowest in the carriers of E3/2 genotype. This suggests that apoE E3/2 genotype is a protective factor in the development of carotid artery atherosclerosis in randomly selected middle-aged men. The favourable effect might be mediated at least partly by the lowering effect of E3/2 genotype on serum cholesterol.

Apolipoprotein E polymorphism and atherosclerosis: association of the epsilon4 allele with defects in the internal elastic lamina.

The defects in the internal elastic lamina (IEL) have been proposed to be important for the migration of smooth muscle cells into the intima during atherosclerosis. We investigated the association of a genetic factor--apolipoprotein E (apoE) genotype--with the number of gaps in the IEL of the artery wall in 123 consecutive autopsy cases (90 male, 33 female) aged 18-93. At autopsy, the circumference of the IEL and the number of gaps in the IEL were measured in circular samples of the coeliac; (CA), superior mesenteric (SMA) and inferior mesenteric (IMA) arteries. In the series, the number of gaps per millimetre in the IEL of CA, SMA and IMA were associated with intimal thickening (P<0.0001, P=0.01 and P=0.005, respectively). In men, apoE genotype was significantly associated with the number of gaps in the IEL of the CA and IMA (P=0.033 and P=0.041, respectively). The carriers of epsilon4/3 or epsilon4/4 genotype had higher number of gaps in CA than the carriers of epsilon3/3 genotype (2.30+/-2.63 vs. 1.38+/-1.83 gaps/mm, P=0.035) and also higher number of gaps in IMA than the carriers of epsilon3/2 (2.18+/-1.71 vs. 0.66+/-0.60 gaps/mm, P=0.041). The results suggest that the apoE varepsilon4 allele may be involved with IEL fragmentation in men. This may be mediated through higher serum cholesterol associated with the varepsilon4 allele.

Apolipoprotein E genotype is not linked to locally recurrent hormone-refractory prostate cancer.

Mechanisms of prostate cancer recurrence during androgen deprivation are poorly understood. Recently, the putative role of apolipoprotein E varepsilon 4 allele in the aetiology of prostate cancer was raised. To investigate the hypothesis that varepsilon 4 allele of apolipoprotein E gene predisposes to prostate cancer and is involved in the relapse of hormonal therapy response, 38 hormone-refractory locally recurrent carcinoma samples from 38 prostate cancer patients were screened for apolipoprotein E genotype. The frequency distribution of apolipoprotein E genotypes among tumours did not differ significantly from that among controls. The allele frequency of varepsilon 4 was 19.7% and 19.3% in tumours and controls, respectively. The results suggest that clinical progression of prostate cancer during androgen withdrawal therapy is not associated with apolipoprotein E genotype. Prostate Cancer and Prostatic Diseases (2000) 3, 107-109

Age-dependent association of apolipoprotein E genotype with coronary and aortic atherosclerosis in middle-aged men: an autopsy study.

BACKGROUND: Apolipoprotein E (apoE) polymorphism is one of the genetic determinants of serum cholesterol values. The apoE epsilon4 allele has been associated with advanced coronary heart disease (CHD) diagnosed by angiography, but the role of the apoE genotype in atherosclerosis has not been confirmed at vessel-wall level, nor is any age-dependent effect of the apoE genotype on the development of CHD known. METHODS AND RESULTS: The right and left anterior descending coronary arteries (RCA and LAD) and the aorta from 700 male autopsy cases (Helsinki Sudden Death Study) in 1981-1982 and 1991-1992 (average age 53 years, range 33 to 70 years) were stained for fat, and all areas covered with fatty streaks, fibrotic plaques, and complicated lesions were measured. In the RCA and LAD, the apoE genotype was significantly associated with the area of total atherosclerotic lesions in men <53 years old but not with that in older men (P=0.0085 and P=0.041, respectively, for age-by-genotype interaction). Men <53 years old with the epsilon4/3 genotype showed 61% larger total atherosclerotic lesion area in the RCA (P=0.0027) and 26% larger area in the LAD (P=0.12) than did men with the epsilon3/3. The apoE epsilon4/3 was also associated with atherosclerotic lesions in the abdominal (P=0.014) and thoracic (P=0.12) aorta, but this effect, unlike that of the coronary arteries, was not age-related. CONCLUSIONS: In men, the apoE epsilon4 allele is a significant genetic risk factor for coronary atherosclerosis in early middle age. This suggests that at older age, other known risk factors of CHD play a more important role in the atherosclerotic process than apoE polymorphisms.

Apolipoprotein E epsilon4 allele frequency is increased in Parkinson's disease only with co-existing Alzheimer pathology.

We determined the apolipoprotein E (apoE) genotype in clinically diagnosed and neuropathologically verified cases of Parkinson's disease (PD) (n = 45), with or without Alzheimer (AD)-type changes, and compared the apoE genotype with that in healthy age-matched controls (n = 59). The PD cases were divided into two groups according to the CERAD criteria: "O + A", with no or only uncertain histological findings of AD, and "B + C" with histological findings suggestive or indicative of AD. DNA was isolated from frozen brain samples, and the apoE genotypes were determined using polymerase chain reaction amplification and subsequent restriction analysis by HhaI enzyme. The frequency of the apo epsilon4 allele (29.4%) was significantly increased in the B + C group. The odds ratio for an apo epsilon4 allele in the B + C group was 2.5 as compared to controls (95% confidence interval, 1.2-5.2). In the O + A group, the frequency of apo epsilon4 allele (13.6%) was similar to that in controls (14.4%) and the risk of an apo epsilon4 allele was not increased (odds ratio 0.94). The PD cases with an apo epsilon4 allele had a greater number of cortical (P = 0.02) but not nigral Lewy bodies than those without an apo epsilon4 allele (P = 0.57). The results show that neuropathologically verified PD as such is not associated with increased apo epsilon4 allele frequency.

Rapid apolipoprotein E genotyping from mailed buccal swabs.

Genetic polymorphism of apolipoprotein E (apoE) is linked with the risk of cardiovascular and Alzheimer's disease (AD). We determined apoE genotypes from mailed buccal cell swabs in 36 healthy individuals and 18 demented patients suffering from AD. DNA was released from the swabs by a rapid lysis technique. Polymerase chain reaction using 'hot start' and restriction enzyme digestion by HhaI was used for genotyping of apoE alleles, which were detected by polyacrylamide or MetaPhor agarose gel. ApoE genotypes determined from buccal cells or from stored leucocytes were identical to previously done phenotyping. An advantage of the present simplified buccal cell collection procedure is the possibility to collect DNA samples for apoE genotyping rapidly by mail from patients visiting their physician geographically distant from research laboratory. Moreover, venepuncture and storing of the samples in refrigerators are avoided and costs are reduced. This non-invasive and painless method is also applicable to other genetic screening tests in risk populations.

Fast 5-hour determination of angiotensin-converting enzyme genotypes from blood by SDS-PAGE using PhastSystem.

Polymorphism in the angiotensin-converting enzyme (ACE) gene has been shown to correlate with circulating ACE concentrations in plasma, and also to be an independent risk factor in the development of many cardiovascular diseases. However, methods available today for ACE genotyping are still laborious and time-consuming. Here we report a sensitive, simple and non-isotopic procedure with commercially available gels for the identification of ACE insertion/ deletion polymorphism. This technique includes rapid extraction of the DNA by the QIAamp Blood Kit prior to polymerase chain reaction, followed by sodium dodecylsulphate polyacrylamide gel electrophoresis using the PhastSystem (Pharmacia). The procedure can be accomplished in five-hours from drawing the blood samples to the final result.