Mika Yamaoka

Decreases in percentages of naïve CD4 and CD8 T cells and increases in percentages of memory CD8 T-cell subsets in the peripheral blood lymphocyte populations of A-bomb survivors

Abstract Yamaoka, M., Kusunoki, Y., Kasagi, F., Hayashi, T., Nakachi, K. and Kyoizumi, S. Decreases in Percentages of Naïve CD4 and CD8 T Cells and Increases in Percentages of Memory CD8 T-Cell Subsets in the Peripheral Blood Lymphocyte Populations of A-Bomb Survivors. Radiat. Res. 161, 290–298 (2004). Our previous studies have revealed a clear dose-dependent decrease in the percentage of naïve CD4 T cells that are phenotypically CD45RA+in PBL among A-bomb survivors. However, whether there is a similar radiation effect on CD8 T cells has remained undetermined because of the unreliability of CD45 isoforms as markers of naïve and memory subsets among the CD8 T-cell population. In the present study, we used double labeling with CD45RO and CD62L for reliable identification of naïve and memory cell subsets in both CD4 and CD8 T-cell populations among 533 Hiroshima A-bomb survivors. Statistically significant dose-dependent decreases in the percentages of CD45RO−/CD62L+naïve cells were found in the CD8 T-cell population as well as in the CD4 T-cell population. Furthermore, the percentages of CD45RO+/ CD62L+and CD45RO+/CD62L−memory T cells were found to increase significantly with increasing radiation dose in the CD8 T-cell population but not in the CD4 T-cell population. These results suggest that the prior A-bomb exposure has induced long-lasting deficits in both naïve CD4 and CD8 T- cell populations along with increased proportions of these particular subsets of the memory CD8 T-cell population.

T-Cell Immunosenescence and Inflammatory Response in Atomic Bomb Survivors

Abstract In this paper we summarize the long-term effects of A-bomb radiation on the T-cell system and discuss the possible involvement of attenuated T-cell immunity in the disease development observed in A-bomb survivors. Our previous observations on such effects include impaired mitogen-dependent proliferation and IL-2 production, decreases in naive T-cell populations, and increased proportions of anergic and functionally weak memory CD4 T-cell subsets. In addition, we recently found a radiation dose-dependent increase in the percentages of CD25+/CD127− regulatory T cells in the CD4 T-cell population of the survivors. All these effects of radiation on T-cell immunity resemble effects of aging on the immune system, suggesting that ionizing radiation might direct the T-cell system toward a compromised phenotype and thereby might contribute to an enhanced immunosenescence. Furthermore, there are inverse, significant associations between plasma levels of inflammatory cytokines and the relative number of naïve CD4 T cells, also suggesting that the elevated levels of inflammatory markers found in A-bomb survivors can be ascribed in part to T-cell immunosenescence. We suggest that radiation-induced T-cell immunosenescence may result in activation of inflammatory responses and may be partly involved in the development of aging-associated and inflammation-related diseases frequently observed in A-bomb survivors.

Decreased Proportion of CD4 T Cells in the Blood of Atomic Bomb Survivors with Myocardial Infarction

Epidemiological studies of the atomic bomb survivors have suggested dose-related increases in mortality from diseases other than cancer. Cardiovascular disease is one such noncancer disease for which increases in both mortality and incidence have been found to be associated with radiation dose. Immunological studies have revealed long-term impairment of T-cell-mediated immunity, especially involving deficiencies of CD4 helper T cells, in atomic bomb survivors. In the present study, we investigated whether decreases in CD4 T cells were associated with myocardial infarction in atomic bomb survivors. Of 1,006 survivors examined to determine the proportion of CD4 T cells in peripheral blood lymphocytes, 18 persons had a history of myocardial infarction. The proportion of CD4 T cells was significantly decreased with increased radiation dose [corrected]. Further, the prevalence of myocardial infarction was significantly greater in individuals with a lower proportion of CD4 T cells. These results suggest that myocardial infarction in atomic bomb survivors may be associated with defects in CD4 helper T cells.

T-Cell Responses to Mitogens in Atomic Bomb Survivors: A Decreased Capacity to Produce Interleukin 2 Characterizes the T Cells of Heavily Irradiated Individuals

Abstract Kusunoki, Y., Hayashi, T., Morishita, Y., Yamaoka, M., Maki, M., Hakoda, M., Kodama, K., Bean, M. A. and Kyoizumi, S. T-Cell Responses to Mitogens in Atomic Bomb Survivors: A Decreased Capacity to Produce Interleukin 2 Characterizes the T Cells of Heavily Irradiated Individuals. Significant decreases in the fraction of lymphocytes that are CD4+ and increases in serum levels of some classes of immunoglobulin have been reported to occur in atomic bomb (A-bomb) survivors and in victims of the Chernobyl nuclear plant accident. To investigate the long-term effects of nuclear radiation on cellular immunity in more detail, we used limiting dilution assays with peripheral blood mononuclear cell preparations to analyze the T-cell responses of 251 A-bomb survivors exposed to less than 0.005 Gy and 159 survivors exposed to more than 1.5 Gy. The percentages of CD2-positive cells that were capable of proliferating in response to phytohemagglutinin (PHA) in the presence of exogenous interleukin 2 (IL2) did not differ substantially between distally exposed and more heavily exposed survivors. The heavily exposed survivors appeared to possess fewer T cells that were capable of proliferating in response to concanavalin A (Con A) or of producing interleukin 2. Assuming that CD4 T cells were the ones primarily responsible for producing IL2 in response to Con A, we were able to estimate how many cells in any given CD4 T-cell population were actually producing IL2. The results indicated that peripheral blood samples from heavily exposed survivors contained significantly fewer IL2-producing CD4 T cells than did similar samples from distally exposed survivors, indicating that significant exposure to A-bomb radiation may have a long-lasting negative effect on the capacity of CD4 T-cell populations to produce IL2.

T cells of atomic bomb survivors respond poorly to stimulation by Staphylococcus aureus toxins in vitro: Does this stem from their peripheral lymphocyte populations having a diminished naïve CD4 T-cell content?

Abstract Kusunoki, Y., Yamaoka, M., Kasagi, F., Hayashi, T., Koyama, K., Kodama, K., MacPhee, D. G. and Kyoizumi, S. T Cells of Atomic Bomb Survivors Respond Poorly to Stimulation by Staphylococcus aureus Toxins In Vitro: Does This Stem from their Peripheral Lymphocyte Populations Having a Diminished Naïve CD4 T-Cell Content? Radiat. Res. 158, 715–724 (2002). We found previously that the peripheral CD4 T-cell populations of heavily exposed A-bomb survivors contained fewer naïve T cells than we detected in the corresponding unexposed controls. To determine whether this demonstrable impairment of the CD4 T-cell immunity of A-bomb survivors was likely to affect the responsiveness of their immune systems to infection by common pathogens, we tested the T cells of 723 survivors for their ability to proliferate in vitro after a challenge by each of the Staphylococcus aureus toxins SEB, SEC-2, SEC-3, SEE and TSST-1. The results presented here reveal that the proliferative responses of T cells of A-bomb survivors became progressively weaker as the radiation dose increased and did so in a manner that correlated well with the decreasing CD45RA-positive (naïve) [but not CD45RA-negative (memory)] CD4 T-cell percentages that we found in their peripheral blood lymphocyte (PBL) populations. We also noted that the T cells of survivors with a history of myocardial infarction tended to respond poorly to several (or even all) of the S. aureus toxins, and that these same individuals had proportionally fewer CD45RA-positive (naïve) CD4 T cells in their PBL populations than we detected in survivors with no myocardial infarction in their history. Taken together, these results clearly indicate that A-bomb irradiation led to an impairment of the ability of exposed individuals to maintain their naïve T-cell pools. This may explain why A-bomb survivors tend to respond poorly to toxins encoded by the common pathogenic bacterium S. aureus.

Long-lasting changes in the T-cell receptor V beta repertoires of CD4 memory T-cell populations in the peripheral blood of radiation-exposed people

Summary. To study the long‐term effects of radiation‐induced T‐cell depletion on the T‐cell receptor (TCR) Vβ repertoires of human peripheral CD4 T‐cell populations, we measured the percentages of CD4 T cells representing each of the full range of possible TCR Vβ families in a cohort of atomic bomb survivors. We then estimated the extent to which the expression levels for individual TCR Vβ families differed from the average expression level for that particular TCR Vβ family across the entire cohort. We found no evidence of a systematic change in the TCR Vβ repertoires of the naïve CD4 T‐cell populations, but memory CD4 T‐cell TCR Vβ family expression levels diverged significantly from the population average for counterpart families, especially in individuals who had been exposed to higher doses and were at least 20 years of age at the time of the bombing. Comparisons of the TCR Vβ family expression profiles in the naïve and memory CD4 T‐cell pools of the same group of adult survivors revealed that differences in the TCR Vβ repertoires of these two types of CD4 T‐cell pool were larger in more heavily exposed survivors than in unexposed controls. These findings suggest that the memory CD4 T‐cell pools of individuals who received significant radiation doses in adulthood may well have become (and could still be) dependent upon a much less diverse complement of TCR Vβ families than would otherwise have been the case.

Inverse Associations between Obesity Indicators and Thymic T-Cell Production Levels in Aging Atomic-Bomb Survivors

Reduction of the naive T-cell population represents a deteriorating state in the immune system that occurs with advancing age. In animal model studies, obesity compromises the T-cell immune system as a result of enhanced adipogenesis in primary lymphoid organs and systemic inflammation. In this study, to test the hypothesis that obesity may contribute to the aging of human T-cell immunity, a thousand atomic-bomb survivors were examined for obesity status and ability to produce naive T cells, i.e., T-cell receptor excision circle (TREC) numbers in CD4 and CD8 T cells. The number of TRECs showed a strong positive correlation with naive T cell numbers, and lower TREC numbers were associated with higher age. We found that the TREC number was inversely associated with levels of obesity indicators (BMI, hemoglobin A1c) and serum CRP levels. Development of type-2 diabetes and fatty liver was also associated with lower TREC numbers. This population study suggests that obesity with enhanced inflammation is involved in aging of the human T-cell immune system. Given the fact that obesity increases the risk of numerous age-related diseases, attenuated immune competence is a possible mechanistic link between obesity and disease development among the elderly.

Aging-related changes in human T-cell repertoire over 20 years delineated by deep sequencing of peripheral T-cell receptors

Abstract Recent deep sequencing studies on T‐cell receptor (TCR) repertoire have provided robust data to characterize diversity of T‐cell immune responsiveness to a wide variety of peptide antigens, including viral and tumor antigens. The human TCR repertoire declines with age, but this decline has not been fully investigated longitudinally in individuals. Using a deep sequencing approach, we analyzed TCR&bgr; repertoires longitudinally over approximately 20 years, with ages ranging from 23 to 50 years at the start (23 to 65 years overall), in peripheral‐blood CD4 and CD8 T‐cell populations that were collected and cryopreserved 3 times at intervals of approximately 10 years from each of 6 healthy adults (3 men and 3 women). Sequence data at the hypervariable complementarity determining region 3 (CDR3) in the TCRB gene locus were evaluated by applying a random‐coefficient statistical regression model. Two outcomes were analyzed: total number of distinct TCRB CDR3 sequences as a TCR diversity metric, and clonality of the T‐cell populations. TCR repertoire diversity decreased (p < 0.001) and frequencies of clonal populations increased (p = 0.003) with age in CD8 T cells, whereas CD4 T cells retained fairly diverse TCR repertoires along with relatively low clonality. We also found that approximately 10–30% and 30–80% of read sequences in CD4 and CD8 T cells, respectively, overlapped at different ages within each individual, indicating long‐term stable maintenance of T‐cell clonal composition. Moreover, many of the most frequent TCRB CDR3 sequences (i.e., top T‐cell clones) persisted over 20 years, and some of them expanded and exerted a dominating influence on clonality of peripheral T‐cell populations. It is thus possible that persistence or expansion of top T‐cell clones is a driver of T‐cell immunity aging, and therefore represents a potential interventional target. HighlightsAging‐related changes in TCR&bgr; repertoire were delineated by deep sequencing combined with a longitudinal study design.In particular, TCR diversity of CD8 T cells decreased at −0.99 % per year during aging from 24 to 65 years.Many of the most frequent T‐cell clones (based on the CDR3 sequences) persisted, and some of them expanded over 20 years.

Memory CD4 T-cell subsets discriminated by CD43 expression level in A-bomb survivors

Purpose: Our previous study showed that radiation exposure reduced the diversity of repertoires of memory thymus-derived cells (T cells) with cluster of differentiation (CD)- 4 among atomic-bomb (A-bomb) survivors. To evaluate the maintenance of T-cell memory within A-bomb survivors 60 years after radiation exposure, we examined functionally distinct memory CD4 T-cell subsets in the peripheral blood lymphocytes of the survivors. Methods: Three functionally different subsets of memory CD4 T cells were identified by differential CD43 expression levels and measured using flow cytometry. These subsets consist of functionally mature memory cells, cells weakly responsive to antigenic stimulation, and those cells functionally anergic and prone to spontaneous apoptosis. Results: The percentages of these subsets within the peripheral blood CD4 T-cell pool all significantly increased with age. Percentages of functionally weak and anergic subsets were also found to increase with radiation dose, fitting to a log linear model. Within the memory CD4 T-cell pool, however, there was an inverse association between radiation dose and the percentage of functionally mature memory cells. Conclusion: These results suggest that the steady state of T cell memory, which is regulated by cell activation and/or cell survival processes in subsets, may have been perturbed by prior radiation exposure among A-bomb survivors.

Effects of NKG2D haplotypes on the cell-surface expression of NKG2D protein on natural killer and CD8 T cells of peripheral blood among atomic-bomb survivors.

NKG2D is a primary activating receptor that triggers cell-mediated cytotoxicity in NK cells against tumor and virus-infected cells. We previously identified the NKG2D haplotypes in the natural killer gene complex region on chromosome 12p. Two major haplotype alleles, LNK1 and HNK1, were closely related to low and high natural cytotoxic activity phenotypes, respectively. Furthermore, the haplotype of HNK1/HNK1 has revealed a decreased risk of cancer compared with LNK1/LNK1. In the present study, using flow cytometry, we evaluated the functional effects of NKG2D haplotypes and five htSNPs in terms of the cell-surface expression of NKG2D protein on NK and CD8 T cells of peripheral blood among 732 atomic-bomb survivors. NKG2D expression on NK cells showed significant increases, in the order of LNK1/LNK1, LNK1/HNK1 and HNK1/HNK1 haplotypes (p for trend=0.003), or with major homozygous, heterozygous, and minor homozygous genotypes for individual htSNPs (p for trend=0.02-0.003). The same trend was observed for NKG2D expression on CD8 T cells. Our findings indicate that the NKG2D haplotypes are associated with the expression levels of NKG2D protein on NK and CD8 T cells, resulting in inter-individual variations in human cytotoxic response.