Relationship between microsatellite polymorphism in the haem oxygenase-1 gene promoter and longevity of the normal Japanese population

M Yamaya, K Nakayama, S Ebihara, H Hirai, S Higuchi, H Sasaki

JMedGenet2003;40:146–148

xidative stress is associated with the pathogenesis of O various diseases such as cancer, pulmonary disease, and cardiovascular and cerebrovascular disease.1–3 In addition to environmental factors, genetic factors may be involved in determining a person’s susceptibility to diseases induced by oxidative stress. Although it is not obvious that these genotypes might be more frequent in older people,as the majority of the elderly are subject to age related disorders, healthy elderly people with some protective genetic back- ground may escape from age related disorders and can reach an advanced age.

• We screened the frequencies of alleles with varying numbers of (GT) repeats in the HO-

gene in n healthy Japanese subjects. • The proportion of allelic frequencies in class L with a large size of (GT) repeat, as well as the genotypic n quencies in group I with class L alleles, was lower in the oldest male subjects than in the younger males. In contrast, in the oldest female subjects, the portion of allelic frequencies in class L and the frequencies in group I with class L alleles, did not from that in the other female subjects. • The present study suggests that a large (GT) repeat n the HO-

gene promoter may be a genetic factor that prevents the attainment of old age in male Japanese subjects.

Haem oxygenase (HO) oxidatively degrades haem to biliverdin, which is subsequently reduced to bilirubin, an effi- cient scavenger of reactive oxygen species, by biliverdin reductase.4 Because not only HO-1, an inducible form of HO, but also a constitutive form of HO, including HO-2, provide cellular protection against haem and non-haem mediated oxi- dant injury,4 HO is suggested to have an important role in pro- tection against various reactive oxygen species.

A (GT)n repeat of the rat prolactin gene is polymorphic,and this purine-pyrimidine alternating sequence, with Z confor- mation potential, negatively affects transcriptional activity in the rat prolactin gene.5 A (GT) repeat of the human HO-1 n gene is also polymorphic,6 and modulates human HO-1 gene transcription.7 We have shown that the large size (33< repeat) of a (GT) repeat in the HO-1 gene (X14782) is associated with n susceptibility to chronic pulmonary emphysema in Japanese populations and that a large (GT) repeat categorised as a class L allele inhibits HO-1 gene transcn ription induced by hydrogen peroxide in cultured cell lines.8 Based on these findings, we hypothesised that the number of (GT) repeats in the HO-1 n gene is associated with susceptibility to the development of diseases induced by oxidative stress.

oldest (>75 years, 108 male and 99 female). DNA was extracted from peripheral blood leucocytes obtained from each subject, and screening of the length of the (GT) repeats n in the HO-1 gene promoter was performed with methods described previously.8

Associations between groups and specific classes of allele,as well as age groups and genotype groups, were analysed for significance by the two tailed c 2 test.8 Probability values were corrected by Bonferroni’s method. Odds ratios and 95% confi- dence intervals (CIs) were calculated to assess the allelic fre- quencies with class L and the genotypic frequencies in group I with the class L allele. All subjects were Japanese and represented an ethnic isolate. Therefore, statistical artefacts caused by population stratification could be ruled out, as described by Reich and Goldstein.10 These parameters were chosen before the analysis and the present study was performed as a planned test. The HO-1 genotype distributions were in Hardy-Weinberg equilibrium.

To test this hypothesis, we screened allelic frequencies of the (GT) repeats in the HO-1 gene promoter in 512 healthy n Japanese subjects (20-94 years old).

SUBJECTS AND METHODS

We recruited the subjects at a health screening. In order to be defined as healthy, all the subjects were interviewed on past history and present illness and their daily life style was evalu- ated. Subjects who had obvious symptoms or disabilities were excluded. Subjects with good control in response to treatment for chronic diseases such as hypertension, diabetes mellitus, and hyperlipidaemia were included. All the subjects were Japanese and were recruited from a single area.

RESULTS

Table 1 shows the average age and the frequencies of hypertension, diabetes mellitus, or hyperlipidaemia, risk factors for cardiovascular and cerebrovascular disease, in each group. The frequencies of hypertension, hyperlipidaemia, or diabetes mellitus were not different between subjects with or without the class L allele, except that the frequencies of diabetes mellitus were higher in older male and female subjects in group I than those in group II.

The number of patients with cardiovascular and cerebrovas- cular diseases under medical supervision has increased in the Japanese population over 60 years old.9 The average life expectancy in Japan in 2000 was 77.6 years for men and 84.6 years for women. Therefore, we stratified the subjects into three age groups:younger (<60 years,59 male and 45 female), older (60 years < age <75 years,95 male and 106 female),and

The distribution of the numbers of (GT)n repeats was trimodal as previously described in Japanese smokers with and without CPE,8 with one peak at 23 GT repeats and the other two peaks located close together, at 30 and 33 GT

1 Average age and the number and ratio of subjects with hypertension, dialetes mellitus, or hyperlipidaemia