HLA-A and HLA-B transcription decrease with ageing in peripheral blood leucocytes.

Immunosenescence involves modifications of humoral and cellular immunity. In a previous study, we have shown a locus-dependent reduction of HLA class-I cell surface expression on peripheral lymphocytes and monocytes with advancing age. Here we report the quantitative analysis of HLA-A and -B transcripts from PBL of 54 healthy subjects aged 21-90 years. Using a competitive RT-PCR method, we observed a significant decrease of HLA-A (P < 0.0001) and -B (P = 0.0025) mRNA contents with increasing age. Secondly, to investigate this locus-dependent alteration of HLA class-I transcription, we performed EMSA using nuclear extracts from PBL of five young (24-31-year-old) and 5 elderly (58-69 years old) donors with locus A and B sequences of the Enh-A as probes. No qualitative variation of EMSA profiles appeared between the two groups of donors with 6 and 4 bandshift for the locus A and B, respectively. Quantitatively, we observed a significant increase of B4 intensity in the elderly group compared to the young group (P < 0.05). These results suggest that the variation of DNA binding protein could contribute to the lower transcription of HLA-A and -B with ageing. These alterations of HLA class-I expression at the transcriptional level could lead to the unresponsiveness of CD8 T cells due to default of antigen presentation with ageing.

Localization of the 3' IgH locus elements that effect long-distance regulation of class switch recombination.

Four transcriptional enhancers lie downstream of the immunoglobulin heavy chain locus: Calpha3'/hs3a, hs1,2, hs3b, and hs4. Although individually weak, these elements have strong transcriptional synergies when combined and they altogether behave as a locus control region. Previous knockout experiments in the 3' region have shown that both hs3a and hs1,2 are dispensable for normal expression and rearrangement of the IgH locus but that their replacement with a transcribed neo gene severely affects class switch recombination. Here we show that even in the absence of a neo gene, joint deletion of the last two 3' enhancers, hs3b and hs4, severely impairs germline transcription and class switching to most isotypes and may in addition affect mu gene expression in resting B cells.

Polymorphism of the human alpha1 immunoglobulin gene 3' enhancer hs1,2 and its relation to gene expression.

We studied the hs1,2 transcriptional enhancer identified downstream of the human alpha1 gene of the immunoglobulin H (IgH) locus, for which two different allelic configurations (a and b) were previously reported by Southern blotting. By using a polymerase chain reaction (PCR) method we amplified minisatellites within the hs1,2 core enhancer, with variable numbers of tandem repeats (VNTR) defining three 'PCR alleles' alpha1A, alpha1B and alpha1C (including one, two and three repeats, respectively). Five different alpha1 h1,2 genotypes were encountered in a population of 513 donors, representing 13.8, 34.5, 49.7, 1.3 and 0.6% for the AA, BB, AB, AC and BC genotypes, respectively. Luciferase assays showed that increasing the number of minisatellites increased the transcriptional strength of the alpha1 hs1,2 enhancer. Simultaneous determination of Southern blot alleles and VNTR alleles only showed a partial linkage between both types of polymorphism, altogether defining at least six different allelic forms of the 3'alpha1 region. In conclusion, the present study further demonstrates the genetic instability of the 3'alpha region, for which multiple alleles have been generated through inversions and internal deletions and/or duplications. This study also strengthens the hypothesis that the polymorphism at the IgH 3' regulatory region of the alpha1 gene could play a role in the outcome of diseases involving immunoglobulin secretion.

An elevation in the concentration of HLA class I molecules in human blood due to ageing.

The quantification of plasmatic HLA class-I molecules (sHLA-I) was realized by a sandwich ELISA using a monomorphic mAb (W6/32). sHLA-I concentration is significantly increased in the elderly group (>50 years, 0.429+/-0.301 microg ml(-1)) as compared to the young group (<50 years, 0.126+/-0.085 microg ml(-1)). The variation of sHLA-I content could contribute to the unresponsiveness of ageing immune system.

Age-related alterations of somatic hypermutation and CDR3 lengths in human Vkappa4-expressing B lymphocytes.

The lower avidity and/or affinity of antibodies generated by an aged immune system could be attributed to two major changes in the antibody repertoire: a shift in germline gene usage and a decrease in the rate of immunoglobulin hypermutation. In an attempt to identify the mechanisms involved in the observed humoral immune deficiency in the elderly, we studied whether differences in the somatic diversity of a particular Vkappa region occurred with ageing. By using the polymerase chain reaction and sequencing, we analysed and compared Vkappa4-Jkappa rearrangements isolated from young (mean age 21 years) and aged (mean age 83 years) healthy adults. Mutations in the Vkappa4 gene compared with the germline sequence were determined as well as the length and structure of the CDR3 sequence. We analysed in detail various mechanisms contributing to CDR3 and Vkappa variability in rearrangements involving the Vkappa4 gene. Our data revealed that, despite strong individual variations, significantly lower levels of somatic mutation were found in the aged group, both for complementarity-determining regions (CDRs) and framework regions (FRs) encoding Vkappa4 sequences. This decrease mostly affected mutations responsible for replacements and thus resulted in a lowered somatic diversification of the encoded Vkappa4 proteins in aged individuals. Moreover, comparison of the CDR3 regions of the Vkappa4-Ckappa cDNA revealed changes in light-chain junctional diversity that correlated with age. Altogether these data suggest an impaired light-chain somatic diversity in connection with human senescence.

Modification of HLA expression on peripheral lymphocytes and monocytes during aging.

Immunosenescence involves modifications of humoral and cellular immunity. Here we report the analysis of human leukocyte antigen (HLA) expression on T lymphocytes, B lymphocytes and monocytes of 58 healthy subjects aged 23-95 years old. Using a double staining immunofluorescence and flow cytometry analysis, we have determined the percentages of cells expressing HLA class-I and HLA-DR antigens. The number of antigenic sites expressed per cell were evaluated for HLA-ABCw, HLA-A, HLA-B, HLA-DR locus with a flow cytometry quantification technique. With advancing age, we observed: (i) a significant decrease of the percentage of T cells and B cells expressing HLA-A products; (ii) a decrease of the number of HLA class-I antigenic sites expressed per cell on the three populations tested, predominantly on B cells and in a locus-dependent fashion; (iii) a decrease of the number of HLA-DR molecules expressed per T cell, although the percentage of T cells expressing DR products was increased; (iv) a significant diminution of the percentage of B cells expressing HLA-DR molecules, without changes of the number of HLA-DR antigenic sites per cells. These changes in HLA expression with increasing age could contribute to the decreased level of immunologic responsiveness observed with ageing and contribute to the modification of antigen recognition.

Differential effects of temperature on specific and nonspecific immune defences in fish.

The susceptibility of fish to disease is partly dependent on their environment, in particular on water temperature. It is generally accepted that lower temperatures adversely affect specific immune responses mediated by T helper cells. The probable mechanisms involved in such suppression in teleost fish are reviewed. Furthermore, the effects of temperature on nonspecific defences, such as phagocytosis and cytotoxicity, are described and total immune competence in teleosts at low environmental temperatures is discussed.

Effects and mechanisms of environmental temperature on carp (Cyprinus carpio) anti-DNP antibody response and non-specific cytotoxic cell activity: a kinetic study.

The effect of environmental temperatures on immune competence was investigated in carp which were subjected to changes in water temperature. The activity of non-specific cytotoxic cells (NCC) against P815 target cells, and the anti-DNP antibody response were evaluated until day 56 after transfer. Low environmental temperature (12 +/- 0.5 degrees C) enhanced NCC activity and decreased antibody production. In contrast a high environmental temperature (28 +/- 0.5 degrees C) was without effect on these parameters when compared to the standard temperature (20 +/- 0.5 degrees C). The results showed a maximum effect of low environmental temperature on day 28 and an adaptation in these immune responses 56 days following transfer. Collectively, the results indicated that non-specific immunity tends to offset specific immune suppression at low environmental temperatures. To determine the mechanism(s) by which environmental temperature affects cellular immune function, membrane fluidity measurements and sialic acid titration, as well as stress assessment by plasma cortisol measurement, were determined on day 28. Taken together, the results revealed a direct effect of temperature on cellular immune function which is modulated by membrane fluidity and sugar concentration and not by stress induction.