2. Materials and methods

2.1. Subjects

One hundred control samples (59% female, 41% male with an age-range of 19-45 years) were chosen at random from the archival DNA bank of normal healthy Caucasian individuals from the Northern Ireland population. These samples were stored within the Northern Ireland Histocompatibility and Immunogenetics Laboratory (NIHIL) of Belfast City Hospital. The study involved 129 aged consecutive samples (70% female, 30% male with an age-range of 80-97 years) from the Belfast Elderly Longitudinal Free-living Ageing Study (Rea et al., 2000). All samples were from healthy Caucasian individuals from the Northern Ireland population displaying no signs of age-related disorders, still living independently and approximate SENIEUR status (Ligthart et al., found to be more frequent in the aged population compared to the control group. The frequency of the heterozygote TC genotype was significantly decreased in the aged group, and the TT genotype was significantly increased. These differences were observed independent of gender. The results suggest that increased expression or functionality of the HSP 70-Hom protein may provide better protection against oxidative stress and prevent accumulation of abnormal proteins within cells, thus potentially affecting lifespan. The distribution of T and C alleles in the normal population is highly skewed towards T, suggesting that homozygous CC is almost incompatible with life. The presence of the Met residue may result in individuals who are less susceptible to disease and more resistant to cellular stresses and aging. The study investigated a polymorphism of the HSP 70-Hom gene located in the class III region of the HLA complex, which displays strong linkage disequilibrium with HLA class I and II genes. Any polymorphic association with aging observed within the HSP genes may be a result of closely linked polymorphism elsewhere within the HLA region.The frequency of a variant HSP 70-Hom gene/protein was observed to be decreased by 77% in the control group and 83% in the cohort of aged subjects, although the difference was not significant. This suggests that an increase in the frequency of this variant is associated with the aged population, supporting the theory that the Thr residue (C allele) may affect the efficiency of the HSP 70-Hom protein by lowering the strength of hydrophobic interactions between the chaperone and target protein. In a proposed structural model of the peptide-binding, based on that of the HLA class I molecule, the altered amino acid at position 493 (Met to Thr) is on one of the β-sheets, which forms the floor of the peptide-binding groove affecting specificity (Pociot et al., 1993). 

Polymorphism of the HLA complex and immune system has been extensively studied in aging associations (Reviewed by Caruso et al., 2000; Caruso et al., 2001). However, one limitation in most studies is a lack of aged individuals available from both genders and complete background histories on health or ethnicity for study participants. This is especially important considering that it is postulated that both sexes follow different pathways to longevity (Franceschi et al., 2000). The relatively low numbers used in this study may be one possible factor in the lack of significant differences in allele frequencies between aged and control populations, although there is a recognizable tendency for an increase in T allele frequency in Irish aged populations.

Our results suggest that further examination of polymorphism within HSPs and other HLA-linked genes may provide interesting and informative data. 

Acknowledgments: 
The authors wish to acknowledge: The European Union for support under the aegis of European Union Concerted Action on the Molecular Biology of Immunosenescence (EUCAMBIS; Biomed 1 contract CT94-1209); and Immunology and Ageing in Europe (IMAGINE; QLK6-CT-1999-O2031). OA Ross was a recipient of a CAST Award jointly funded by the Department of Education and Learning (Northern Ireland) and the Northern Ireland Histocompatibility and Immunogenetics Charitable Trust Fund. For Josie O'Malley (1919-2002)

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