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The objective of this study was to evaluate effects of whole body radiation exposure early in life on influenza vaccination immune responses much later in life. A total of 292 volunteers recruited from the cohort members of ongoing Adult Health Study (AHS) of Japanese atomic bomb (A-bomb) survivors completed this observational study spanning two influenza seasons (2011-2012 and 2012-2013). Peripheral blood samples were collected prior to and three weeks after vaccination. Serum hemagglutination inhibition (HAI) antibody titers were measured as well as concentrations of 25 cytokines and chemokines in culture supernatant from peripheral blood mononuclear cells, with and without in vitro stimulation with influenza vaccine. We found that influenza vaccination modestly enhanced serum HAI titers in this unique cohort of elderly subjects, with seroprotection ranging from 18 to 48% for specific antigen/season combinations. Twelve percent of subjects were seroprotected against all three vaccine antigens post-vaccination. Males were generally more likely to be seroprotected for one or more antigens post-vaccination, with no differences in vaccine responses based on age at vaccination or radiation exposure in early life. These results show that early life exposure to ionizing radiation does not prevent responses of elderly A-bomb survivors to seasonal influenza vaccine.

Summary Aging‐related decline in immunity is believed to be the main driver behind decreased vaccine efficacy and reduced resistance to infections in older adults. Unrepaired DNA damage is known to precipitate cellular senescence, which was hypothesized to be the underlying cause of certain age‐related phenotypes. Consistent with this, some hallmarks of immune aging were more prevalent in individuals exposed to whole‐body irradiation (WBI), which leaves no anatomical repository of undamaged hematopoietic cells. To decisively test whether and to what extent WBI in youth will leave a mark on the immune system as it ages, we exposed young male C57BL/6 mice to sublethal WBI (0.5–4 Gy), mimicking human survivor exposure during nuclear catastrophe. We followed lymphocyte homeostasis thorough the lifespan, response to vaccination, and ability to resist lethal viral challenge in the old age. None of the irradiated groups showed significant differences compared with mock‐irradiated (0 Gy) animals for the parameters measured. Even the mice that received the highest dose of sublethal WBI in youth (4 Gy) exhibited equilibrated lymphocyte homeostasis, robust T‐ and B‐cell responses to live attenuated West Nile virus (WNV) vaccine and full survival following vaccination upon lethal WNV challenge. Therefore, a single dose of nonlethal WBI in youth, resulting in widespread DNA damage and repopulation stress in hematopoietic cells, leaves no significant trace of increased immune aging in a lethal vaccine challenge model.

Previous immunological studies in atomic bomb survivors have suggested that radiation exposure leads to long-lasting changes, similar to immunological aging observed in T-cell-adaptive immunity. However, to our knowledge, late effects of radiation on dendritic cells (DCs), the key coordinators for activation and differentiation of T cells, have not yet been investigated in humans. In the current study, we hypothesized that numerical and functional decreases would be observed in relationship to radiation dose in circulating conventional DCs (cDCs) and plasmacytoid DCs (pDCs) among 229 Japanese A-bomb survivors. Overall, the evidence did not support this hypothesis, with no overall changes in DCs or functional changes observed with radiation dose. Multivariable regression analysis for radiation dose, age and gender effects revealed that total DC counts as well as subpopulation counts decreased in relationship to increasing age. Further analyses revealed that in women, absolute numbers of pDCs showed significant decreases with radiation dose. A hierarchical clustering analysis of gene expression profiles in DCs after Toll-like receptor stimulation in vitro identified two clusters of participants that differed in age-associated expression levels of genes involved in antigen presentation and cytokine/chemokine production in cDCs. These results suggest that DC counts decrease and expression levels of gene clusters change with age. More than 60 years after radiation exposure, we also observed changes in pDC counts associated with radiation, but only among women.

The relationships between commitments of dendritic cells (DCs) and T cells in human hematopoietic stem cells are not well understood. In this study, we enumerate and characterize conventional DC and plasmacytoid DC precursors in association with T cell and thymus-derived types of NK cell precursors among CD34+ hematopoietic progenitor cells (HPCs) circulating in human peripheral blood. By limiting-dilution analyses using coculture with stroma cells expressing Notch1 ligand, the precursor frequencies (PFs) of DCs in HPCs were found to significantly correlate with T cell PFs, but not with NK cell PFs, among healthy donors. Clonal analyses showed that the majority of T/NK dual- and T single-lineage precursors—but only a minority of NK single-lineage precursors—were associated with the generation of DC progenies. All clones producing both DC and T cell progenies were found with monocyte and/or granulocyte progenies, suggesting DC differentiation via myeloid DC pathways. Analyses of peripheral blood HPC subpopulations revealed that the lineage split between DC and T/NK cell progenitor occurs at the stage prior to bifurcation into T and NK cell lineages. The findings suggest a strong linkage between DC and T cell commitments, which may be imprinted in circulating lymphoid-primed multipotent progenitors or in more upstream HPCs.

Gastric cancer (GC) is one of the cancers that reveal increased risk of mortality and incidence in atomic bomb survivors. The incidence of gastric cancer in the Life Span Study cohort of the Radiation Effects Research Foundation (RERF) increased with radiation dose (gender-averaged excess relative risk per Gy = 0.28) and remains high more than 65 years after exposure. To assess a possible role of gene-environment interaction, we examined the dose response for gastric cancer incidence based on immunosuppression-related IL-10 genotype, in a cohort study with 200 cancer cases (93 intestinal, 96 diffuse and 11 other types) among 4,690 atomic bomb survivors participating in an immunological substudy. Using a single haplotype block composed of four haplotype-tagging SNPs (comprising the major haplotype allele IL-10-ATTA and the minor haplotype allele IL-10-GGCG, which are categorized by IL-10 polymorphisms at -819AG and -592TG, +1177TC and +1589AG), multiplicative and additive models for joint effects of radiation and this IL-10 haplotyping were examined. The IL-10 minor haplotype allele(s) was a risk factor for intestinal type gastric cancer but not for diffuse type gastric cancer. Radiation was not associated with intestinal type gastric cancer. In diffuse type gastric cancer, the haplotype-specific excess relative risk (ERR) for radiation was statistically significant only in the major homozygote category of IL-10 (ERR = 0.46/Gy, P = 0.037), whereas estimated ERR for radiation with the minor IL-10 homozygotes was close to 0 and nonsignificant. Thus, the minor IL-10 haplotype might act to reduce the radiation related risk of diffuse-type gastric cancer. The results suggest that this IL-10 haplotyping might be involved in development of radiation-associated gastric cancer of the diffuse type, and that IL-10 haplotypes may explain individual differences in the radiation-related risk of gastric cancer.

Ionizing radiation (IR) is a major source of cellular damage and the immediate cellular response to IR has been well characterized. But the long-term impact of IR on cell function and its relationship with aging are not known. Here, we examined the IR effects on telomere length and other biomarkers 50 to 68 years post-exposure (two time points per person) in survivors of the atomic bombing at Hiroshima during WWII. We found that telomere length of leukocytes was inversely correlated with the dose of IR (p=0.008), and this effect was primarily found in survivors who were exposed at younger ages; specifically those

Past exposure to atomic bomb (A-bomb) radiation has exerted various long-lasting deleterious effects on the health of survivors. Some of these effects are seen even after >60 yr. In this study, we evaluated the subclinical inflammatory status of 442 A-bomb survivors, in terms of 8 inflammation-related cytokines or markers, comprised of plasma levels of reactive oxygen species (ROS), interleukin (IL)-6, tumor necrosis factor α (TNF-α), C-reactive protein (CRP), IL-4, IL-10, and immunoglobulins, and erythrocyte sedimentation rate (ESR). The effects of past radiation exposure and natural aging on these markers were individually assessed and compared. Next, to assess the biologically significant relationship between inflammation and radiation exposure or aging, which was masked by the interrelationship of those cytokines/markers, we used multivariate statistical analyses and evaluated the systemic markers of inflammation as scores being calculated by linear combinations of selected cytokines and markers. Our results indicate that a linear combination of ROS, IL-6, CRP, and ESR generated a score that was the most indicative of inflammation and revealed clear dependences on radiation dose and aging that were found to be statistically significant. The results suggest that collectively, radiation exposure, in conjunction with natural aging, may enhance the persistent inflammatory status of A-bomb survivors.—Hayashi, T., Morishita, Y., Khattree, R., Misumi, M., Sasaki, K., Hayashi, I., Yoshida, K., Kajimura, J., Kyoizumi, S., Imai, K., Kusunoki, Y., Nakachi, K. Evaluation of systemic markers of inflammation in atomic-bomb survivors with special reference to radiation and age effects.

It is not yet known whether hematopoietic stem and progenitor cells (HSPCs) are compromised in the aging population of atomic bomb (A-bomb) survivors after their exposure nearly 70 years ago. To address this, we evaluated age- and radiation-related changes in different subtypes of circulating HSPCs among the CD34-positive/lineage marker-negative (CD34(+)Lin(-)) cell population in 231 Hiroshima A-bomb survivors. We enumerated functional HSPC subtypes, including: cobblestone area-forming cells; long-term culture-initiating cells; erythroid burst-forming units; granulocyte and macrophage colony-forming units; and T-cell and natural killer cell progenitors using cell culture. We obtained the count of each HSPC subtype per unit volume of blood and the proportion of each HSPC subtype in CD34(+)Lin(-) cells to represent the lineage commitment trend. Multivariate analyses, using sex, age and radiation dose as variables, showed significantly decreased counts with age in the total CD34(+)Lin(-) cell population and all HSPC subtypes. As for the proportion, only T-cell progenitors decreased significantly with age, suggesting that the commitment to the T-cell lineage in HSPCs continuously declines with age throughout the lifetime. However, neither the CD34(+)Lin(-) cell population, nor HSPC subtypes showed significant radiation-induced dose-dependent changes in counts or proportions. Moreover, the correlations of the proportions among HSPC subtypes in the survivors properly revealed the hierarchy of lineage commitments. Taken together, our findings suggest that many years after exposure to radiation and with advancing age, the number and function of HSPCs in living survivors as a whole may have recovered to normal levels.

This study aims to deepen understanding of lymphocyte phenotypes related to the course of hepatitis C virus (HCV) infection and progression of liver fibrosis, in a cohort of atomic-bomb survivors. The study subjects comprise three groups: 162 HCV persistently infected, 145 spontaneously cleared, and 3511 uninfected individuals. We found increased percentages of peripheral blood TH1 and total CD8 T cells and decreased percentages of NK cells in the HCV persistence group, compared with the other two groups, after adjustment for age, gender, and radiation exposure dose. Subsequently, we found that increased TH1 cell percentages in the HCV persistence group were significantly associated with an accelerated time-course reduction in platelet counts―accelerated progression of liver fibrosis―while TC1 and NK cell percentages were inversely associated with the progression. This study suggests that TH1 immunity is enhanced by persistent HCV infection, and that percentages of peripheral TH1, TC1, and NK cells may help predict progression of liver fibrosis., This research was based on RERF Research Protocols 3-09, 4-02, 2-00, 9-92, and was supported in part by the U.S. National Institute of Allergy and Infectious Diseases (NIAID Contract HHSN272200900059C).

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.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.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.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.

Lung cancer is a leading cause of cancer death worldwide. Prevention could be improved by identifying susceptible individuals as well as improving understanding of interactions between genes and etiological environmental agents, including radiation exposure. The epidermal growth factor receptor (EGFR)-signaling pathway, regulating cellular radiation sensitivity, is an oncogenic cascade involved in lung cancer, especially adenocarcinoma. The cytosine adenine (CA) repeat number polymorphism in the first intron of EGFR has been shown to be inversely correlated with EGFR production. It is hypothesized that CA repeat number may modulate individual susceptibility to lung cancer. Thus, we carried out a case-cohort study within the Japanese atomic bomb (A-bomb) survivor cohort to evaluate a possible association of CA repeat polymorphism with lung cancer risk in radiation-exposed or negligibly exposed (5 mGy) A-bomb survivors. First, by dividing study subjects into Short and Long genotypes, defined as the summed CA repeat number of two allelesor = 37 andor = 38, respectively, we found that the Short genotype was significantly associated with an increased risk of lung cancer, specifically adenocarcinoma, among negligibly exposed subjects. Next, we found that prior radiation exposure significantly enhanced lung cancer risk of survivors with the Long genotype, whereas the risk for the Short genotype did not show any significant increase with radiation dose, resulting in indistinguishable risks between these genotypes at a high radiation dose. Our findings imply that the EGFR pathway plays a crucial role in assessing individual susceptibility to lung adenocarcinoma in relation to radiation exposure.

To elucidate the mechanism of radiation-induced cancers, molecular analysis of cancers in atomic-bomb survivors is important. In our study, we developed a custom oligonucleotide array of 208 genes. We analyzed gene expression profiles of gastric cancers (GCs) from atomic-bomb survivors and identified 9 genes with significantly lower expression in GCs from exposed patients than in GCs from nonexposed patients. Among these 9 genes, expression of versican and osteonectin was investigated in greater detail using immunohistochemistry in 116 GCs from 64 exposed and 52 nonexposed patients who developed GC after the bombing. In the Stage I/II GCs, the clinicopathologic, phenotypic and proliferative characteristics of GCs from exposed and nonexposed patients did not differ significantly; however, versican and osteonectin were expressed at much lower levels in the area of tumor-associated stroma of exposed patients than in nonexposed patients (p = 0.026 and p = 0.024, respectively). These results suggest that the characteristics of tumor-associated stromal cells differ between GCs from exposed and nonexposed patients. © 2008 Wiley-Liss, Inc.

To assess oxidative stress (OS) induced by endurance exercise, concentrations of serum reactive oxygen species (ROS) were determined in 70 Japanese male amateur runners completing a two-day ultra-marathon race. Serum ROS levels were analyzed at three time points: before the race (baseline), after the 1st day race (mid-race), and after the 2nd day race (goal) (post-race). The means (SE) of ROS were 151.4(3.7) (U. CARR.), 168.7(4.4), and 156.8(4.4), respectively. Significant positive trends were noted between age and serum ROS concentrations at the three race points (p

Epigenetic silencing of genes by aberrant DNA methylation is recognized as a crucial component of the mechanism underlying tumorigenesis. However, the relationship between DNA methylation and the past lifestyle in cancer patients remains largely unknown. We examined the methylation statuses of 6 tumor-related genes, CDX2 (homeobox transcription factor), BMP-2 (bone morphogenetic protein 2), p16 (INK4A), CACNA2D3 (calcium channel-related), GATA-5 (transcription factor) and ER (estrogen receptor), in 106 primary gastric carcinomas by methylation-specific PCR and compared them with the past lifestyles of the patients. The methylation frequencies of the genes were 23.6, 21.7, 9.4, 32.4, 40.8 and 59.1%, respectively. Significant association was found between a decreased intake of green tea and methylation of CDX2 and BMP-2. More physical activity was correlated with a lower methylation frequency of CACNA2D3. Of these 6 genes, the methylation statuses of CDX2, BMP-2 and p16 revealed a significant interrelationship and those of CACNA2D3, GATA-5 and ER did likewise. Thus, some epidemiological factors, such as green tea intake, could be important as to determination of the methylation statuses of selected genes and may influence the development of cancer, including that of the stomach.

Epidermal growth factor receptor (EGFR) mutations in lung cancer enhance tyrosine kinase activity and increase sensitivity to the EGFR tyrosine kinase inhibitor, gefitinib. Mutation analysis of the EGFR gene is therefore indispensable for predicting gefitinib response. We investigated a CA-repeat polymorphism in the EGFR gene related to EGFR mutations. Because an increasing number of CA-repeats at intron 1 of the EGFR gene has been reported to reduce transcription activity, we examined the relationship between EGFR mutations and this CA-repeat polymorphism. EGFR mutations at exon 19 were closely associated with shorter CA-repeat length in the shorter allele, but this was not the case for EGFR mutations at exons 18 or 21. Increased intrinsic EGFR mRNA expression in non-cancerous lung tissues from lung adenocarcinoma patients was also significantly associated with shorter CA-repeat length. A higher frequency of EGFR mutations at exon 19 was associated with shorter CA-repeat length only in patients with high levels of EGFR mRNA expression. To determine the phenotypes of cells possessing shorter CA-repeats, an in vitro study using human bronchial epithelial cells with different CA-repeat lengths was performed; more rapid cell growth and activated EGF/EGFR signaling were found more often in the cells having both shorter CA-repeats and increased EGFR mRNA expression. These results suggest that CA-repeat length in the EGFR gene may be a genetic factor related to cancer in the case of EGFR mutations at exon 19. The mechanism likely involves enhanced intrinsic expression of EGFR mRNA and activated EGF/EGFR signaling that accompany shorter CA-repeats.

The derivatives of reactive oxygen metabolites (D-ROM) test has been developed to determine the amount of oxygen-centered free radicals in a blood sample as a marker of oxidative stress. This study aims to improve the D-ROM test and develop an automated assay system by use of a clinical chemistry analyzer. Five microliters of serum was added to 1 well of a 96-well microtiter plate for a total 240microl of reaction solution containing alkylamine and metals. This was followed by automatic mixing, incubation and measurement of reactive oxygen species (ROS) levels as a color development at 505nm using a spectrophotometer with catalytic capability for transition metals. This assay system was used to measure serum levels of ROS in cigarette smokers and never-smokers, by way of example. The levels of serum ROS determined by this system correlate with the amounts of free radicals and peroxides, which reacted with various molecules in the body and formed stable metabolites. This test can use frozen sera as well as fresh ones. The inter- and intra-deviation of this system was within 5% and showed consistent linearity in the range between 4 and 500mg/l of hydrogen peroxides. Serum ROS levels among smokers increased with the number of cigarettes smoked per day (36.5% increment per pack per day; P0.0001). This assay system will be a simple, inexpensive, and reliable tool for assessing oxidative stress in human populations. Our preliminary results on cigarette smoking imply that this assay system has potential for application in various epidemiological and clinical settings.

In the normal human life span, there occur lifestyle-related diseases that may be preventable with nontoxic agents. This paper deals with the preventive activity of green tea in some lifestyle-related diseases. Green tea is one of the most practical cancer preventives, as we have shown in various in vitro and in vivo experiments, along with epidemiological studies. Among various biological effects of green tea, we have focused on its inhibitory effect on TNF-alpha gene expression mediated through inhibition of NF-kappaB and AP-1 activation. Based on our recent results with TNF-alpha-deficient mice, TNF-alpha is an endogenous tumor promoter. TNF-alpha is also known to be a central mediator in chronic inflammatory diseases such as rheumatoid arthritis and multiple sclerosis. We therefore hypothesized that green tea might be a preventive agent for chronic inflammatory diseases. To test this hypothesis, TNF-alpha transgenic mice, which overexpress TNF-alpha only in the lungs, were examined. The TNF-alpha transgenic mouse is an animal model of human idiopathic pulmonary fibrosis which also frequently develops lung cancer. Expressions of TNF-alpha and IL-6 were inhibited in the lungs of these mice after treatment with green tea in drinking water for 4 months. In addition, judging from the results of a prospective cohort study in Saitama Prefecture, Japan, green tea helps to prevent cardiovascular disease. In this study, a decreased relative risk of death from cardiovascular disease was found for people consuming over 10 cups of green tea a day, and green tea also had life-prolonging effects on cumulative survival. These data suggest that green tea has preventive effects on both chronic inflammatory diseases and lifestyle-related diseases (including cardiovascular disease and cancer), resulting in prolongation of life span.

We have previously shown that natural cytotoxic activity of peripheral blood lymphocytes was inversely related to cancer development based on a prospective cohort study. The genetic fraction of cytotoxic activity needs to be clarified, identifying individuals immunogenetically susceptible to cancer. A case-control study within the cohort members was designed: 102 cancer cases with peripheral lymphocyte DNA available and three control groups, each of which consisted of 204 subjects with each tertile level of cytotoxic activity. We first compared two control groups with high and low cytotoxic activity in terms of the single nucleotide polymorphisms in the natural killer complex gene region on chromosome 12p, identifying the haplotype alleles that were associated with the activity. Next, cancer risks were assessed for these haplotypes. We found two haplotype blocks, each of which generated two major haplotype alleles: low-activity-related LNK1 (frequency 0.478 and 0.615 in groups with high and low activity, respectively; P < 0.00008) and high-activity-related HNK1 (0.480 and 0.348; P < 0.0001), LNK2 (0.711 and 0.821; P < 0.0002), and HNK2 (0.272 and 0.174; P < 0.0008). These NKG2D haplotype alleles showed a significant difference between cases (0.632 for LNK1 and 0.333 for HNK1) and controls (0.554 for LNK1 and 0.406 for HNK1). The haplotype HNK1/HNK1 revealed a decreased risk of cancer (odds ratio, 0.471; 95% confidence interval, 0.233-0.952) compared with LNK1/LNK1. Individuals who are genetically predisposed to have low or high natural cytotoxic activity can in part be determined by NKG2D haplotyping, which in turn reveals an increased or decreased risk of cancer development. (Cancer Res 2006; 66(1): 563-70)

It has previously been reported that hemizygous mutant fraction (Mf) at the glycophorin A (GPA) locus in erythrocytes increased with radiation dose in heterozygotes among Hiroshima and Nagasaki atomic bomb survivors. In the present study, we analyzed the relationship between GPA Mf and cancer risk using newly developed cancers among previously cancer-free subjects whose GPA Mf had been measured between 1988 and 1996. Among 1,723 survivors (1,117 in Hiroshima and 606 in Nagasaki), we identified 186 subjects who developed a first cancer by the end of 2000. We compared the radiation dose responses of GPA Mf between cancer and cancer-free groups using a linear-quadratic model fit by multiple regression analysis in combination with age, sex, and city. The slope of the GPA Mf dose-response curve was significantly higher in the cancer group than in the cancer-free group among Hiroshima subjects. Moreover, no significant difference of GPA Mf between cancer and cancer-free groups was found in unexposed controls in the two cities. The same conclusions were obtained using a linear dose-response model and by further analysis using Cox regression of cancer incidence. These findings suggest that there might be interindividual variation in mutability of somatic genes and that Hiroshima survivors who have higher mutability in response to radiation exposure would be expected to have a higher probability of suffering radiation-related cancer.

BACKGROUND: Subjects of the Japan Collaborate Cohort Study (JACC Study) gave peripheral blood samples collected between 1988 and 1990. We conducted to investigate whether levels of serum components measured after 9 years of frozen storage are stable or not.METHODS: To assess the degradation of frozen serum components in the JACC Study, we compared levels of various components (IGF-I, IGF-II, IGFBP-3, TGF-β1, sFas, and total SOD activity) between fresh and stored sera collected from other inhabitants. Serum levels of constituents were measured by immunoradiometric assay (IGF-I, IGF-II and IGFBP-3), quantitative enzyme immunoassay (TGF-β1), enzyme-linked immuno-adsorbent assay (sFas), and an improved nitrite method (SOD activity). RESULTS: The coefficients of variation for intra- and inter-assay precisions of the measurements were less than 9%. Levels of IGF-I, IGF-II, IGFBP-3, TGF-β1 and sFas in sera after storage for 9 years at - 80°C were similar to those of fresh sera newly collected from inhabitants. The distributions of serum IGF-I, IGF-II, IGFBP-3, TGF-β1, sFas and SOD activity for specimens collected from different individuals tended to be similar to those of serum levels for frozen specimens collected from different individuals and stored for 9 years.CONCLUSIONS: There was no statistically significant difference in distribution of measured values of IGF-I, IGF-II, IGFBP-3, TGF-β1, and sFas between newly collected sera and frozen specimens stored for 9 years. Thus, measurements of these serum constituents of specimens stored for the JACC Study can be reliably used in nested case-control study.J Epidemiol 2005; 15: S67-S73.

Objective: Epidermal growth factor (EGF) has many biological functions and plays an important role in the progression of various tumors including gastric cancer. An A-G single nucleotide polymorphism (SNP) at position 61 in the 5′-untranslated region (UTR) of the EGF gene has recently been reported to be associated with different levels of EGF production. We examined whether this polymorphism is correlated with the development and malignant phenotypes of gastric cancer. Methods: The study population included 200 gastric cancer patients and 230 healthy control subjects. The SNP in the 5′-UTR of the EGF gene was analyzed by polymerase chain reaction-restriction fragment length polymorphism. Results: The A allele was significantly less frequent in patients than in controls (p = 0.01). Individuals with the A/A or A/G genotype showed a significantly lower risk of gastric cancer than those with the G/G genotype [adjusted odds ratio (OR) = 0.56], whereas the same genotypes were associated with malignant progression of this cancer, e.g. deeper tumor invasion, increased lymph node metastasis and advanced clinical stage, and histological classification in gastric cancer patients (adjusted OR = 1.80, 1.98, 2.26 and 1.89, respectively). Conclusions: Our findings suggest that the A-G polymorphism of EGF is involved not only in the occurrence but also in the malignant progression of gastric cancer.

Objective: A population-based cohort study was conducted to examine concordance of cancer incidence and lifestyle in married couples. Methods: Cancer incidence from 1 September 1986 to 30 June 2000 was followed for a baseline cohort of 2601 community-living married couples aged 40-84 years who were cancer-free and had completed an epidemiologic questionnaire in 1986 in Saitama, Japan. We computed age-adjusted cross-ratios: spouse-with/spouse-without rate ratios as a measure of association for paired rate data, using a bivariate survival analysis method. Results: Concordance of lifestyles for couples was high for dietary habits, and low for smoking and alcohol habits. During the 14 year follow-up, 464 cancer cases occurred (husbands only: 279, wives only: 119, both: 33 couples). Wives whose husbands developed cancer were at an increased risk of developing cancer themselves, compared with those whose husbands had not had cancer (CR: 1.70, 95% CI: 1.12-2.58). Mean age at diagnosis was 68.2 years for husbands and 66.0 years for wives. And correlation of cancer incidence was further seen among couples who shared habits of smoking and/or drinking. Conclusions: Our data suggest that shared lifestyles including smoking and drinking habits are associated with an excess concordance of cancer incidence among married couples.

Matrix metalloproteinase-1 (MMP-1) plays a key role in cancer invasion and metastasis by degradation of extracellular matrix (ECM) and basement membrane barriers. The 1G/2G single nucleotide polymorphism (SNP) in the MMP-1 promoter at position –1607 bp has been reported to affect the transcriptional activity. In the light of these findings, we investigated whether this SNP in the MMP-1 promoter is associated with the development, differentiation, and progression of gastric cancer. The 215 gastric cancer patients and 166 controls were used in this study. The SNP of the MMP-1 promoter was analyzed by PCR-RFLP and sequencing. The genotype frequency was compared between cases and controls, and the association with clinicopathological parameters among cases was studied. The frequency of 1G/2G genotypes in gastric cancer patients was similar to those in controls (p=0.57). The degree of tumor invasion, the presence of lymph node metastasis, and clinical stage showed no significant association with the SNP. On the other hand, we found a significant association with histological differentiation and gender among gastric cancer patients (p

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.

We investigated progressive telomere shortening in normal human epidermis and lingual epithelium during aging, and attempted, in particular, to ascertain whether the telomere shortening that accompanies aging occurs at the same rate in different tissues. We studied telomeric DNA integrity, and estimated annual telomere loss, in 52 specimens of epidermis and 48 specimens of lingual epithelium collected at autopsy from subjects who had died at ages between 0 and 101 y. Most of the DNA samples were measured twice by southern blot hybridization. In addition, the correlation between telomere lengths in the two types of tissues was examined. The telomere reduction rates in epidermis and lingual epithelium were 36 bp and 30 bp per y, respectively, and these were significantly different. The rates obtained by the second measurements in epidermis and lingual epithelium were 39 and 32 bp per y, respectively, and these were also significantly different. The mean telomere lengths in the epidermis of eight neonates and the lingual epithelium of seven neonates were 13.2+/-1.0 and 13.8+/-1.0 kb, respectively. Comparison of telomere lengths in the two tissues for 41 paired samples showed that the mean telomere length in the epidermis (10.7+/-2.3 kb) was less than that in the lingual epithelium (12.4+/-2.5 kb); however, statistical analysis revealed a very significant relationship between epidermal and lingual epithelial telomere length (r=0.842, p0.0001). These results indicate that the telomeres in epidermis and lingual epithelium are characterized by tissue-specific loss rates.

The number of murine mature blood cells recovered within 6 weeks after 2-Gy whole-body irradiation at 6 weeks of age, whereas in the case of the undifferentiated hematopoietic stem/progenitor cell (HSC/HPC) compartment [cells in the lineage-negative, c-kit-positive and stem-cell-antigen-1-positive (LKS) fraction], the numerical differences between mice with and without irradiation remained more than a year, but conclusively the cells showed numerical recovery. When mice were exposed to radiation at 6 months of age, acute damages of mature blood cells were rather milder probably because of their maturation with age; but again, cells in the LKS fraction were specifically damaged, and their numerical recovery was significantly delayed probably as a result of LKS-specific cellular damages. Interestingly, in contrast to the recovery of the number of cells in the LKS fraction, their quality was not recovered, which was quantitatively assessed on the basis of oxidative-stress-related fluorescence intensity. To investigate why the recovery in the number of cells in the LKS fraction was delayed, expression levels of genes related to cellular proliferation and apoptosis of cells in the bone marrow and LKS fraction were analyzed by real-time polymerase chain reaction (RT-PCR). In the case of 21-month-old mice after radiation exposure, Ccnd1, PiK3r1 and Fyn were overexpressed solely in cells in the LKS fraction. Because Ccnd1and PiK3r1 upregulated by aging were further upregulated by radiation, single-dose radiation seemed to induce the acceleration of aging, which is related to the essential biological responses during aging based on a lifetime-dependent relationship between a living creature and xenobiotic materials.

The study of green tea polyphenols as a cancer preventative is approaching a new era, with significant results accumulating rapidly. This paper briefly reviews four topics related to mechanisms of action of tea polyphenols: (I) identification of the genes commonly affected by EGCG, as demonstrated by Clontech's Atlas cDNA Expression Array; (II) the significance of heterogeneous nuclear ribonucleoprotein B1 (hnRNP B1) as a new biomarker for early detection of lung cancer, and inhibition of its expression by EGCG; (III) the synergistic or additive effects of EGCG with the cancer preventive agents, sulindac and tamoxifen, on induction of apoptosis in PC-9 cells and on inhibition of intestinal tumor development in multiple intestinal neoplasia (Min) mice; (IV) the results of a 10 year prospective cohort study demonstrating the effectiveness of daily consumption of green tea in preventing cancer, and a prototype study for developing green tea beverage as cancer preventive.

We examined the effect of smoking cessation on natural killer (NK) activity of peripheral blood lymphocytes in terms of a prospective study of 27 Japanese subjects who participated in a smoking cessation intervention program. This program was delivered by means of groupcounseling offering 7 sessions of about 2 hours over 6 months to help smokers to discontinue the habit. Thirteen subjects ceased smoking (quitters), while 14 continued to smoke (cigarette smokers). NK activity before the intervention was correlated positively with age (correlation coefficient=0.46, P

p73 gene, a new p53 homologue, has been identified: it supposedly acts as tumour suppressor gene in neuroblastoma. To clarify whether p73 might be involved in lung carcinogenesis, we examined p73 expression in resected lung cancer and paired normal lung in 60 cases using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). We also examined p73 gene status in three representative cases using Southern blot, and p53 gene alteration in 49 cases using PCR-single-strand conformation polymorphism (PCR-SSCP) and direct sequence. In 87% of the cases (52/60) p73 expression in tumour was more than twice as high as that in paired normal lung tissues, and the difference between p73 expression in tumour and normal lung tissue was significant (P < 0.0001). However, Southern blot analysis revealed that none of the cases showed p73 gene amplification. Compared with clinicopathological characteristics, p73 expression correlates significantly with histological differences and age of patient, independently (P < 0.05). Concerning p53 gene status, 43% (21/49) showed p53 gene alteration, but there was no correlation between p73 overexpression and p53 gene alteration. Our results suggest that need for further functional analysis of the role of p73 in lung carcinogenesis. © 1999 Cancer Research Campaign

Background.Laboratory studies have revealed the cancer preventive effects of green tea, so the association between green tea consumption and cancer was examined in a human population. Methods.The association between green tea consumption and cancer incidence was studied in our prospective cohort study of a Japanese population. We surveyed 8,552 individuals over 40 years of age living in a town in Saitama prefecture on their living habits, including daily consumption of green tea. During the 9 years of follow-up study (71,248.5 person-years), we identified a total of 384 cases of cancer in all sites. Results.We found a negative association between green tea consumption and cancer incidence, especially among females drinking more than 10 cups a day. The slowdown in increase of cancer incidence with age observed among females who consumed more than 10 cups a day is consistent with the finding that increased consumption of green tea is associated with later onset of cancer. Age-standardized average annual incidence rate was significantly lower among females who consumed large amounts of green tea. Relative risk (RR) of cancer incidence was also lower among both females (RR = 0.57, 95% CI = 0.33–0.98) and males (RR = 0.68, 95% CI = 0.39–1.21) in groups with the highest consumption, although the preventive effects did not achieve statistical significance among males, even when stratified by smoking and adjusted for alcohol and dietary variables. Conclusion.Our epidemiological study showed that green tea has a potentially preventive effect against cancer among humans.

We have previously reported on an association of genetic susceptibility to squamous cell carcinoma of the lung with two polymorphisms of the CYP1A1 gene, an MspI polymorphism and an isoleucine-valine (Ile-Val) polymorphism. We report here that the two CYP1A1 polymorphisms were associated with current or ex-smokers with adenocarcinoma of the lung. We first compared smoking status of the patients by differentiation grades of adenocarcinoma. Proportion of current or ex-smokers and their cigarette consumption was the highest among the poorly differentiated. Second, the two polymorphisms of CYP1A1 were examined among current or ex-smokers with three differentiation grades, and we found that poorly differentiated adenocarcinoma showed significantly high frequencies of genotypes C and Val/Val, which have been found to be 'susceptible' in squamous cell carcinoma of the lung. Third, a case-control study was carried out to estimate the genetic risk for CYP1A1 genotypes to differentiation grades, selecting only current or ex-smokers in patients and controls. Current or ex-smokers with 'susceptible' genotype C or Val/Val were at significant risk, with an odds ratio of 3.25 or 3.22 (95% confidence interval (CI), 1.40-7.56 or 1.00-10.32) for adenocarcinoma as a whole, respectively. The odds ratio increased to 4.51 or 4.09 (95% CI, 1.73-11.78 or 1.12-14.91) for poorly differentiated, while the odds ratios for other differentiated grades were not significant. Finally, a relation of the genetic risk to cigarette dose levels was evaluated.

Drinking green tea daily is part of Japanese culture, and various studies have revealed that green tea is a cancer preventive. We here review our progress in cancer prevention with green tea on 12 main topics, from basic to clinical level. TOPICS AND METHODS: Biochemical and biological studies of green tea catechins, a prospective cohort study, preclinical safety trials with tablets of green tea extract, double-blind randomized clinical phase II prevention trial for recurrence of colorectal adenomas, and synergistically enhanced inhibition by the combination of green tea catechins and anticancer drugs. All results were significant, including human studies with informed consent.Drinking 10 Japanese-size cups of green tea per day delayed the cancer onset of humans 7 years for females. For tertiary cancer prevention, consuming 10 cups of green tea per day fortified by green tea tablets, 50 %, significantly prevented the recurrence of colorectal adenomas. A minimum effective amount of green tea catechins for cancer prevention was found in humans. In addition, the combination of green tea catechins and anticancer drugs engendered a new cancer therapeutic strategy.The consumption of 10 Japanese-size cups of green tea per day is a significant factor in primary cancer prevention for the general population, and the preventive effect on recurrence of colorectal adenomas in patients is vital evidence in tertiary cancer prevention.

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.

Tumor protein p53 (TP53) is the best-known tumor suppressor gene and plays a crucial role in carcinogenesis. The TP53 Arg 72 Pro polymorphism has been reported to be a risk factor for several types of cancer, but its association with pancreatic cancer has not been fully evaluated. Therefore, we investigated the effects of this polymorphism on pancreatic cancer in relation to smoking and drinking habits by examining the distribution of the SNP genotypes in 226 pancreatic cancer patients and 448 healthy controls. The frequencies of Arg/Arg, Arg/Pro and Pro/Pro were found to be 37, 49 and 15% in the pancreatic cancer cases and 44, 46 and 10% in the controls, respectively. Compared to the controls with the Arg/Arg genotype, cases with Pro/Pro homozygosity exhibited a significantly increased risk [adjusted odds ratio (OR)=1.70; 95% confidence interval (CI) 1.01-2.88]. In stratified studies, the association was particularly strong in males (OR=2.62; 95% CI 1.32-5.23), particularly in those smoking in excess of 20 pack-years and drinking in excess of 23 g ethanol/day (OR=5.02; 95% CI 1.12-22.51). We found that the TP53 Pro/Pro genotype compared to the Arg/Arg genotype had a profound effect on pancreatic cancer risk among males, particularly among heavy smokers and excessive alcohol drinkers.

The p53 tumor suppressor gene is mutated in diverse types of human cancer, and the normal allele encodes a nuclear protein that regulates expression of cell cycle-related genes as a transcription factor. The wild-type of p53 protein exists as at least two forms of variants among human populations, ascribed to amino acid replacement at codon 72 of Arg by Pro. In this study, we show that this germ line Arg-Pro polymorphism at codon 72 of the p53 gene is associated with genetically determined susceptibility to smoking-induced lung cancer; a susceptible genotype Pro/Pro has a 1.7-fold higher risk of this cancer compared with other genotypes. This p53 polymorphism modulates risk to smoking-induced lung cancer independently of other genetic risk factors such as germ line polymorphism of CYP1A1 or GST1 genes.

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.

Metabolic risk factors are known to cause atherosclerosis through inflammation. In the process of inflammation, soluble Fas (sFas) may interfere with the apoptotic pathway and contribute to dysregulated inflammation. Recent studies suggest sFas as a marker of inflammation in patients with cardiovascular diseases. However, whether a relationship exists between sFas levels and metabolic risk factors among healthy subjects remains unclear.We measured the serum sFas levels of 876 subjects selected as controls for a nested case-control study within the JACC Study. The adjusted means of the sFas levels were compared according to the presence of overweight/obesity, hypertension, hyperlipidaemia, diabetes and their clusters.sFas level was significantly associated with overweight/obesity (2.42 ng mL(-1) in overweight/obese men and 2.19 in others) and hyperlipidaemia (2.34 ng mL(-1) in men with hyperlipidaemia and 2.19 in others) among men, though not with hypertension or diabetes. Moreover, a clear association between sFas levels and the cluster number of metabolic risk factors was observed independently with age, smoking and drinking(2.39, 2.28, 2.24 and 2.11 ng dL(-1) in men with three to four, two, one and none of the four metabolic risk factors respectively). However, among women, clear associations were not observed between sFas levels and the four metabolic risk factors or their clustering.Serum sFas levels appear to be associated with overweight/obesity, hyperlipidaemia and clusters of metabolic risk factors among men, suggesting that sFas may elevate to down-regulate increased apoptosis in atherogenesis processes.

Recent studies have linked elevated serum sFas levels to atherosclerotic disease among patients. Confirming an association between obesity and serum sFas levels in healthy subjects would facilitate our understanding of obesity and its related disorders. We therefore analyzed serum sFas levels of 8,541 subjects selected as controls for a nested case-control study within the JACC Study. Body mass index (BMI) was calculated as the indicator of obesity based on self-reported height and weight. We found a statistically significant positive association between serum sFas levels and BMI among our apparently healthy subjects. Our result suggests that serum sFas rises to down-regulate increased apoptosis in atherogenesis processes caused by obesity.

Superoxide dismutase (SOD) is an antioxidant enzyme that acts to degrade superoxide, a major causitive factor for oxidative stress associated with cancer, cardiovascular disease, and various other ailments. Here, to assess an association between antioxidants and lifestyle factors related to cancer risk, we analyzed serum SOD activity among the subjects within a large-scale cohort study in Japan. As results, significant differences in serum SOD activity were found between the sexes (lower in males), among female age groups (lower in younger individuals), and in males with the BMI (lower in those with a high BMI). Linear increase in serum SOD activity with aging and decrease with BMI were observed in females. Significantly low SOD activity was evident in male heavy smokers. In contrast, elevation was noted in female frequent drinkers. In conclusion, our findings do suggest associations between serum SOD activity and lifestyle factors. However, for further study, establishment of a standard measurement method for SOD activity should be a high priority.

Insulin-like growth factor (IGF)-I and its main binding protein, IGFBP-3, modulate cell growth and survival, and thus are thought to be important for tumor development. Carotenoids and retinol have been linked to the prevention of several cancers. We here evaluated associations of serum levels of carotenoids and retinol with IGF-I, IGF-II, and IGFBP-3 within the context of the JACC Study. The study subjects were 924 controls (578 men and 346 women) of a nested case-control study of lung and colorectal cancer risk. Using frozen-stored sera, serum levels of a-carotene, b-carotene, lycopene, b-cryptoxanthin, zeaxanthin/lutein, and retinol were separately determined using high-performance liquid chromatography. Serum levels of IGF-I, IGF-II, and IGFBP-3 were measured by immuno-radiometric assay. Confounding factors-adjusted least squares mean levels of serum IGF-I, IGF-II, and IGFBP-3 for each quartile of serum levels of carotenoids and retinol were estimated. Serum IGF-I, IGF-II, and IGFBP-3 levels increased with increasing serum retinol levels. Moreover, serum IGF-I levels were significantly higher in highest quartile of serum provitamin A, such as a-carotene, b-carotene, and b-cryptoxanthin, among women. Serum IGFBP-3 levels decreased with increasing serum lycopene levels in women and with increasing serum zeaxanthin/lutein levels in men. The current study indicates that positive associations exist for serum retinol levels with serum levels of IGF-I, IGF-II, and IGFBP-3 independent of age, BMI, smoking habits, drinking habits, and intake of energy and protein among Japanese healthy men and women.

The preventive effects of physical activity against cancer may partly be ascribable to a possible decrease in insulin-like growth factor-I (IGF-I) induced by the activity. To examine the association of physical activity with IGF-I and IGF-binding protein-3 (IGFBP-3), we analyzed the data for control participants in a case-control study nested in the Japan Collaborative Cohort Study. A total of 3,598 men and 3,359 women throughout Japan, aged 40 to 79 years, were administered a lifestyle questionnaire and provided serum samples. The age- and body mass index (BMI)- adjusted serum levels of IGF-I and IGFBP-3 were lower among those who walked for longer times with a significant trend in both sexes (trend P0.01). Among participants who walked one hour or more per day, the mean levels of serum IGF-I and IGFBP-3 were 138.7 ng/ml and 2.87 microg/ml in men and 125.7 ng/ml and 3.14 microg/ml in women, respectively. The corresponding figures among those rarely walked were 147.4 ng/ml and 2.99 microg/ml in men and 132.3 ng/ml and 3.21 microg/ml in women. For IGF-I, adjustment for serum IGFBP-3 did not essentially alter such associations although the trend in women did not reach statistical significance. A decreased level of serum IGF-I was associated with longer exercise time in men even after adjustment for serum IGFBP-3 (trend P = 0.033 after adjustment for age, BMI, and serum IGFBP-3), whereas the time was positively correlated with serum IGF-I in women (trend P = 0.048). Our findings may partly explain the protective effects of physical activity against several sites of cancer.

Insulin-like growth factor (IGF)-I and IGF-II are important mitogen and IGF binding protein-3 (IGFBP-3) exerts opposite effects. However, the results of epidemiological studies on cancer influence are somewhat controversial, and mainly from Western countries. In the present study, we therefore examined associations of serum IGF-I, IGF-II and IGFBP-3 with colorectal cancer risk among participants in the JACC Study in Japan. After matching 3 controls to cases by sex, age, and study area, a total 101 risk sets were examined using a conditional logistic regression model adjusted for body mass index, smoking habit, alcohol consumption and family history of colorectal cancer. The odds ratios (and 95% CIs) for colorectal cancer mortality among the highest tertiles of IGF-I, IGF-II, and IGFBP-3, compared to the lowest tertiles were 1.01 (0.49-2.10), 1.02 (0.55- 1.91), and 1.22 (0.63-2.38), respectively. No linear trends were observed. The lack of any association was not altered after additional adjustment for mutual markers of IGF-I/IGF-II or IGFBP-3, 0.76 (0.34-1.71) for IGF-I, 0.66 (0.30-1.45) for IGF-II, and 1.11 (0.47-2.66) for IGFBP-3. Our prospective data thus indicated that there is no association of IGF-I, IGF-II, and IGFBP-3 with colorectal cancer risk in the Japanese population. Although these markers might be etiologically significant in relation to colorectal cancer, we did not obtain evidence supporting this hypothesis.

No observational study has examined whether cancer-related biomarkers are associated with diet in Japanese. We therefore assessed sex-specific food and nutrient intakes according to serum IGF-I, IGF-II, IGFBP-3, TGF-b1, total SOD activity and sFas levels, under a cross-sectional study of 10,350 control subjects who answered the food frequency questionnaire in the first-wave nested case-control study within the Japan Collaborative Cohort Study. For both men and women, IGF-I levels were associated with higher intakes of milk, fruits, green tea, calcium and vitamin C. IGF-II levels were associated with higher intakes of milk, yogurt, fruits and miso soup, and lower intakes of rice, coffee and carbohydrate. IGFBP-3 levels were associated with higher intakes of milk, yogurt, fruits and vitamin C, and lower intakes of rice, energy, protein, carbohydrate, sodium and polyunsaturated fatty acids. TGF-b1 levels were associated with lower intakes of coffee intakes, and higher intakes of miso soup and sodium. Total SOD activity levels were associated with lower intakes of most nutrients other than energy, carbohydrate, iron, copper, manganese, retinol equivalents, vitamin A, B2, B12, niacin, folic acid, vitamin C and fish fat. sFas levels were associated with higher intakes of manganese and folic acids. The results of the present study should help to account for findings on those biomarkers regarding risks of cancer and other lifestyle-related diseases in terms of dietary confounding as causality.

Biliary tract cancer, encompassing gallbladder and bile duct cancers, has a poor prognosis, but little is known of the etiology. A nested case-control study was here conducted to evaluate the association between serum levels of IGF-I, IGF-II and IGFBP-3 and death from biliary tract cancer. In a large scale cohort study, 35 gallbladder and 42 bile duct cancers were observed during the follow-up. For each subject in the case group, 1-3 control subjects (228 in total) were selected randomly, matched for sex, age (as near as possible) and residential area. The subjects were divided into tertiles by circulating levels of IGF-I, IGF-II or IGFBP-3. Using conditional logistic regression, risks among the tertiles were compared adjusted for defecation, smoking and drinking habits. No remarkable differences in risks of gallbladder or bile duct cancer were observed among tertiles of IGF-I or IGF-II, and no remarkable trend was observed. Circulating IGFBP-3 showed an inverse U-shape association with gallbladder cancer and a U-shaped one with bile duct cancer. Associations between IGF-I or IGF-II and gallbladder or bile duct cancer thus were lacking or very weak. The observed U- and inverse U-shaped association of IGFBP-3 with the cancers is not suggestive of any meaningful relationships.

The expression of superoxide dismutases (SODs) has been shown to differ between lung tumor and tumor-free tissues. In the present study, we investigated the association between serum SOD activity and the risk of lung cancer mortality, based on a nested case-control design study within the Japan Collaborative Cohort Study, with a sample of 193 lung cancer patients and 573 matched controls. Blood samples were obtained at the baseline and stored at -80 degrees C until analysis for SOD levels. Serum levels of SODs were divided into quartiles, with the first quartile used as the reference. A conditional logistic model was used to estimate odds ratios (ORs) for lung cancer mortality associated with serum SOD quartile levels. The adjusted ORs and 95% CIs for the second, third; and fourth SOD quartiles were 0.80 (95%CI: 0.49-1.29), 1.32 (0.78-2.25), and 1.07 (0.60-1.89), respectively. In analyses stratified by observation period, the adjusted ORs of the respective quartiles were 0.56 (95%CI: 0.30-1.07), 1.16 ( 0.57-2.37), and 1.11 (0.52-2.35) for the period from the baseline to 1994; and the adjusted ORs of 1.36 (95%CI: 0.65-2.85), 1.71 (0.75-3.87), and 1.06 (0.44-2.53) for the period after 1994. To conclude, we found no significant association between serum SOD level and the risk of deaths from lung cancer in the present study.

Superoxide dismutases (SODs) are antioxidant enzymes that play a role in the defense system of the body. They may be involved in protection against carcinogenesis processes. In the present study, we investigate the association between serum SOD activity and the risk of deaths due to all cancers combined, based on a nested case-control study within the Japan Collaborative Cohort Study of 914 cancer deaths and 2,739 matched controls. Blood samples were obtained at the baseline and stored at -80 degrees C until analysis for SOD levels. Serum levels of SODs were divided into quartiles, with the first quartile used as the reference. A conditional logistic model was used to estimate odds ratios (ORs) for total cancer mortality associated with serum SOD quartile levels. The adjusted ORs and 95% confidence intervals (CIs) for the second, third and fourth SOD quartiles were 0.96 (95%CI: 0.77-1.19), 1.18 (0.92-1.51), and 1.32 (1.04-1.69), respectively. In analyses stratified by observation period, the adjusted ORs of the respective quartiles were 0.81 (95%CI: 0.60-1.08), 0.98 (0.70-1.37), and 1.28 (0.92-1.79) for the period from the baseline to 1994; and the adjusted ORs were 1.18 (95%CI: 0.85-1.63), 1.47 (1.04-2.10), and 1.41 (1.00-2.04) for the period after 1994. To conclude, we found a slightly positive association between serum SOD level and the risk of all cancer mortality in the present study. Elevated serum SOD levels might reflect a response to oxidative stress, and then may predict a state of excess reactive oxygen species in the carcinogenesis process. Detailed studies of associations between serum SOD levels and cancers in specific sites should now be performed, with attention to particular tumour types.

High levels of insulin-like growth factor (IGF)-I are reported to be associated with an increased risk of prostate cancer. On the other hand, the insulin-like growth factor binding protein-3 (IGFBP-3) may decrease the risk. We therefore investigated the influence of serum IGF-I and IGFBP-3 on prostate cancer risk in a case-control study nested within a large-scale cohort in Japan (the Japan Collaborative Cohort Study). Information on lifestyle and sera of the subjects were collected in 1988-90. Serum IGF-I, IGF-II and IGFBP-3 were measured in sera stored at -80 degrees C by immuno-radiometric assay. In 13,508 male subjects of the cohort who donated sera, 40 cases and 120 controls (1:3 matched with age and survey area) were identified. Ages of the cases ranged from 59 to 79 years, with a mean of 69.8. Odds ratios (ORs) and 95% confidence intervals (CIs) were estimated for the highest and middle tertiles compared with the lowest in controls using a conditional logistic model. Non-adjusted ORs for the highest tertiles were 0.99 (95% CI, 0.34-2.91) for IGF-I (trend-P = 0.60), 1.91 (95% CI, 0.68-5.38) for IGFBP-3 (trend-P = 0.23), 1.73 (95% CI, 0.69-3.47) for IGF-II (trend-P = 0.23), and 0.67 (95% CI, 0.26-1.76) for the IGF-I/IGBP-3 ratio (trend-P = 0.83). Serum levels of IGF-I, IGF-II, IGFBP-3, or IGF-I/IGFBP-3 ratio were thus not thought to be associated with risk of prostate cancer.

In a search for novel circulating biomarkers for pancreatic cancer, we examined the association between serum soluble Fas (sFas) levels and superoxide dismutase (SOD) activity and the risk of death from pancreatic cancer in a nested case-control study within the Japanese Collaborative Cohort Study. Case subjects were 68 persons who were free of morbidity, had provided a blood sample at baseline (1988-1990), and subsequently died from pancreatic cancer before December 31, 1997. Control subjects were 199 matched persons who were selected from the remaining participants in the cohort. Conditional logistic regression models were used to estimate age-adjusted and multivariate-adjusted odds ratios (ORs) and their 95% confidence intervals (CIs). No statistically significant differences were noted in mean sFas levels (p=0.11) and SOD activity (p=0.42) between cases and controls. Overall, neither serum sFas levels nor SOD activity were associated with the risk of pancreatic cancer deaths, after adjustment for area, BMI, cigarette smoking, and history of diabetes. Furthermore, no significant risk trends were noted. Our results do not support the hypothesis that serum sFas levels and SOD activity are associated with pancreatic cancer risk.

We examined the association of natural killer (NK) cell activity with a number of life style factors by a cross-sectional analysis on 2892 Japanese individuals. The following habits were found to be associated with increased NK activity: 1) alcohol drinking (males, P less than 0.05), 2) not smoking cigarettes (males and females, P less than 0.001), 3) increased intake of green vegetables (females, P less than 0.001), 4) increased intakes of meat, milk, dairy products, and soybean products (males, P less than 0.01), 5) daily workload of less than 3 hours per day (females, P less than 0.05), 6) regular meals (females, P less than 0.05), 7) regular sleep of more than 7 hours (females, P less than 0.05), 8) proper body weight. In addition, systolic blood pressure showed a positive correlation with NK activity (P less than 0.001), while proportion of helper/inducer (OKT4+) T cells and fraction of beta-globulin showed negative correlations (P less than 0.001). We thus found that the living habits associated with increased NK activity were consistent with generally accepted good health practices, except alcohol drinking.

Dietary, smoking, and drinking habits, as well as sociopsychological factors and familial history, were investigated in a case-control study on the etiology of esophageal cancer (EC) in two areas of Shanxi (Yangcheng and Linfen), north central China. Data were analyzed from 326 cases and 396 controls. We identified several factors associated with high or low risk; some were common across the areas and others were area-specific. Consumption of millet gruel was associated positively with EC, in a dose-response relationship. An increase in EC risk was seen for consumption of millet soup with noodles, and also with certain sociopsychological factors, in both areas. A large increase in risk was found with consumption of boiled vegetables in Linfen, with a dose-response relationship. EC risk tended to become greater with the increasing intake of moldy foods and of pickled vegetable juice. A positive association between EC risk and family history of EC was observed only in Yangcheng. Soybean consumption was found to be associated with reduced risk. Dental hygiene (brushing teeth) was associated with reduced risk in Linfen. There was a suggestion of increased risk associated with heavy tobacco smoking, but it was not significant in either area. Alcohol consumption had a marginally significant association with risk in the high risk area, but not in Linfen.