• angiotensin I converting enzyme
• genetics
• Alzheimer’s disease

Early studies suggested that angiotensin I converting enzyme (peptidyl-dipeptidase A) 1 (ACE) gene polymorphism is associated with an increased risk of coronary artery disease and, more recently, with sporadic late onset Alzheimer’s disease.¹ Studies conducted in northern European populations have considered the ACE*I allele to be a risk factor for various types of cognitive decline.^1,2 One such study in a French population found an association between the ACE*D allele and dementia,³ while other studies in southern European populations found either a slight but significantly increased frequency of ACE*I in Alzheimer’s disease patients⁴ or did not detect any effect of ACE polymorphism.⁵

Our group recently reported the novel finding that apolipoprotein E (APOE) ε4 allele shows a geographical trend, decreasing in frequency from northern to southern Europe.⁶ We hypothesised that the variability in the strength of evidence for an association between ACE polymorphism and Alzheimer’s disease was related to similar geographical variations in ACE*I frequency. We investigated whether there was evidence in southern Italy of an association between the ACE polymorphism and increased risk of Alzheimer’s disease. Secondly, we compared our results with the findings from published studies on other European populations.^1,2,4

Between June 1998 and October 2001, we consecutively examined in our centre 141 patients with Alzheimer’s disease (51 men, 90 women; mean (SD) age at onset, 71 (8.5) years), and 268 unrelated caregivers, spouses, friends, neighbours, or volunteers (118 men, 150 women; mean age at collection, 72 (7.1) years). A clinical diagnosis of probable Alzheimer’s disease was made according to the criteria of the National Institute for Neurological and Communicative Disorders and Stroke/Alzheimer’s Disease and Related Disorders Association, and the group of non-demented elderly control subjects was sex and age matched. The ascertainment, diagnosis, and collection of cases and controls are described in detail elsewhere.⁶ The age at onset of Alzheimer’s disease symptoms was estimated from semistructured interviews with the patients’ caregivers. The study protocol was approved by the ethics committee of the University of Bari. After a complete explanation of the study, written informed consent was obtained from all the subjects or their relatives.

APOE genotypes were determined as detailed elsewhere.⁶ ACE genotypes were produced using established methods, followed by a quality control amplification step necessary in detecting underamplified ACE*I alleles.¹

The statistical analysis was performed by Pearson χ² test to make genotype and allele comparisons as well as test for agreement of data with Hardy-Weinberg principles. Allele frequencies were determined by allele counting. To express variances of the allele frequencies, we used 95% CIs, calculated by Wilson’s formulas. The differences among age at onset of Alzheimer’s disease symptoms in relation to different ACE genotypes were calculated with Mann-Whitney test. To evaluate whether the association between Alzheimer’s disease and ACE genotypes were homogeneous in all APOE strata we used a permutation based exact logistic model by LogXact procedure implemented in the SAS system (Proc-LogXact 4; Copyright 2001 by CYTEL Software Corporation, Cambridge, MA 021139). In order to correct for multiple statistical testing, the results were adjusted according to Bonferroni inequality. The Cochran-Armitage trend test was carried out to evaluate the geographical trend among ACE allele and genotype frequencies in Alzheimer’s disease patients and controls of three European countries (Italy, Spain, and United Kingdom), from published studies.^1,2,4 The data were analysed by SAS FREQ procedure (version 8.2).

Table 1 shows ACE allele and genotypes frequencies in Alzheimer’s disease patients and controls in southern Italy. The frequencies of the different ACE genotypes in our population were in Hardy–Weinberg equilibrium (HWE) (cases: Pearson χ² = 2.09, p = 0.15; controls: χ² = 2.49, p = 0.11). Moreover, there was no evidence that the genotypic counts of Alzheimer’s disease patients and controls were not under HWE (Alzheimer’s disease patients under HWE, given the controls were under HWE: likelihood ratio, χ² = 2.18, p = 0.34). No significant differences were found in ACE genotype frequencies between patients with Alzheimer’s disease and controls in this southern Italian population.

We did not find any statistically significant differences in rates between ACE alleles and Alzheimer’s disease among APOE allele strata, nor did we observe any lack of homogeneity among response differences (data not shown). Interestingly, Alzheimer patients with the ACE*I/*I genotype were on average 3.6 years younger at onset than those with the ACE*D/*D genotype (mean (SD) age at onset: ACE*D/*D, 72.1 (6.8) years; ACE*I/*D, 70.3 (8.1) years; ACE*I/*I, 68.5 (7.3) years). However, this difference did not reach statistical significance (z = 1.49; Bonferroni p > 0.05).

The ACE*I allele frequency in Alzheimer’s disease patients and controls showed a statistically significant decreasing trend from northern to southern regions of Europe (z = 5.36 p <0.001; z = 4.33 p <0.001, respectively), while there was a concomitant increase in ACE*D allele frequency (table 1). This was reflected by genotype data whereby a decreasing geographical trend from north to south was found for ACE*I/*I genotype (cases, z = 3.92 p <0.01; controls, z = 4.15 p <0.001) and an inverse trend for ACE*D/*D genotype (cases, z = 3.29 p <0.001; controls, z = 3.46 p<0.001). Interestingly, we found a statistically significant decreasing trend from northern to southern regions for the ACE*I/*D genotype, but this was observed only in Alzheimer’s disease patients (z = 3.12 p <0.01).