Hiromu Toma

Functional alteration of granulocytes, NK cells, and natural killer T cells in centenarians

The immune system in centenarians was characterized as elevated levels in the proportion and number of granulocytes, NK cells, and extrathymic T cells (including NKT cells) in the peripheral blood. Conventional T cells, abundant in youth, were decreased in proportion and number. In addition to this numerical change in centenarians, the function was significantly altered in comparison with that in middle-aged subjects. The phagocytic function and cytokine production of granulocytes in centenarians increased whereas the production of superoxides from granulocytes decreased. This tendency was almost the same in both healthy and unhealthy centenarians. IFN gamma production by NK and extrathymic T cells in centenarians seemed to be augmented and resulted in an elevated level of serum IFN gamma. Possibly due to the effect of this endogenous IFN gamma, the proportion of CD64(+) (Fc gamma RI) cells among granulocytes was elevated. The expansion of CD64 antigens on granulocytes is known to be regulated by IFN gamma and to be associated with their induction of phagocytosis. These results suggest that the immune system of centenarians is not merely impaired, but altered in terms of the number and functions of granulocytes, NK cells, NKT cells.

Numerical and functional characteristics of lymphocyte subsets in centenarians

The immune system in the aged is a very interesting subject for study. In this study, analysis was extended to extrathymic T cells as well as NK cells and “conventional” T cells (i.e., thymus-derived T cells) in terms of their constitution and function in both healthy and unhealthy centenarians. Middle-aged persons were used as controls. Healthy and unhealthy centenarians showed lower levels in the proportion and absolute number of lymphocytes. The major change in the constitution of lymphocyte subsets was increased levels in the proportion of NK cells (CD56+/CD57+) and extrathymic T cells (CD3+CD57+). Inversely, conventional T cells decreased in proportion and function (i.e., proliferative response to mitogen). Although NK cells increased in centenarians, NK activity by whole lymphocytes and the purified NK fraction decreased. The difference between healthy and unhealthy centenarians was small in all parameters, the only difference being a lower level of expression of CD56 antigens on CD57+ T cells in unhealthy centenarians. These results indicate that there is a major shift in lymphcyte population from conventional T cells to NK cells and extrathymic T cells with aging. Concerning the age-associated increases in CD56+ T and CD57+ T cells, these cells correspond to NK1+ T cells in mice.

Strongyloides venezuelensis infections in mice

The course and intensity of Strongyloides venezuelensis infection as compared with S. ratti infection were investigated in BALB/c mice. The mice were found to be much more susceptible to infection with S. venezuelensis than S. ratti. The majority of worms inoculated were recovered from the lungs and subsequently from the small intestines, suggesting that their migratory route via the lungs to the small intestine was comparable to that of S. stercoralis in humans. Spontaneous expulsion of worms occurred by about 10 days after infection, which was the same as that of S. ratti. Different infectivities, as assessed by faecal egg excretion, age, sex and strain of mouse were observed in mice infected with S. venezuelensis, as well as in those infected with S. ratti. A striking immunity was acquired following a primary exposure to S. venezuelensis. Mice infected with S. venezuelensis are considered to provide as useful a model as those infected with S. ratti for the study of human strongyloidiasis.

Involvement of IL-2/IL-2R system activation by parasite antigen in polyclonal expansion of CD4(+)25(+) HTLV-1-infected T-cells in human carriers of both HTLV-1 and S. stercoralis.

The intermediate state of HTLV-1 infection, often found in individuals dually infected with Strongyloides stercoralis (S. stercoralis) and HTLV-1, is assumed to be a preleukemic state of adult T-cell leukemia (ATL). To investigate the effects of S. stercoralis superinfection on the natural history of HTLV-1 infection, we characterized peripheral blood samples of these individuals in Okinawa, Japan, an endemic area for both HTLV-1 and S. stercoralis and we studied effects of the parasite antigen on T-cells. The dually infected individuals showed a significantly higher provirus load and an increase in CD4+25+ T cell population, with a significant, positive correlation. This increase was attributable to polyclonal expansion of HTLV-1-infected cells, as demonstrated by inverse-long PCR analysis of the integration sites. S. stercoralis antigen activated the IL-2 promoter in reporter gene assays, induced production of IL-2 by PBMC in vitro, and supported growth of IL-2 dependent cell lines immortalized by HTLV-1 infection or the transduction of Tax. Taken collectively, these results indicate that S. stercoralis infection induces polyclonal expansion of HTLV-1-infected cells by activating the IL-2/IL-2R system in dually infected carriers, an event which may be a precipitating factor for ATL and inflammatory diseases.

Application of enzyme immunoassay for postchemotherapy evaluation of human strongyloidiasis

Posttherapy evaluation of strongyloidiasis is frequently difficult because coprologic examination is not sensitive enough for diagnosis of chronic infection. In the present study, anti-Strongyloides enzyme-linked immunosorbent assay antibodies were monitored before and after treatment with thiabendazole and pyrvinium pamoate in 199 patients with chronic strongyloidiasis in Okinawa, Japan. A significant decrease in antibody levels was observed in patients who became negative for fecal larvae after the treatment, whereas the antibody levels did not show a significant change after the treatment in patients who were still harboring the parasite. In the group coprologically negative in the follow-up examination, however, many individuals did not show a significant fall in antibody titers after treatment, which suggests that these cases were equivocal for complete cure. By the subsequent fecal reexamination performed on the equivocal cases, approximately 20% were additionally found to be still harboring the parasite. These results indicate that serologic testing is useful to check whether a real cure has been achieved among the patients in whose fecal samples the presence of larvae has not been demonstrated after treatment.

Reduced efficacy of treatment of strongyloidiasis in HTLV‐I carriers related to enhanced expression of IFN‐γ and TGF‐β1

Strongyloidiasis, a human intestinal infection caused by Strongyloides stercoralis (S. stercoralis), is difficult to cure with drugs. In particular, a decrease of the efficacy of treatment has been reported in patients dually infected with S. stercoralis and human T‐cell leukaemia virus type I (HTLV‐I), both of which are endemic in Okinawa, Japan. However, the factors influencing this resistance remain unclear. In the present study, patients infected with S. stercoralis, with or without HTLV‐I infection, were treated with albendazole, followed up for one year and separated into two groups, cured and non‐cured. The cure rate of S. stercoralis was lower in HTLV‐I carriers (P < 0·05). Serum levels of S. stercoralis‐specific IgA, IgE, IgG, IgG1 and IgG4 antibodies were estimated, and a decrease of IgE (P < 0·05) and an increase of IgG4 (P < 0·05) were observed in the non‐cured group, especially in HTLV‐I carriers. RT‐PCR of cytokines using peripheral blood mononuclear cells revealed that S. stercoralis patients with HTLV‐I showed a high frequency of expression of IFN‐γ and TGF‐β1, whereas those without HTLV‐I showed no expression of these cytokines. IFN‐γ‐ and TGF‐β1‐positive HTLV‐I carriers showed a decrease of IgE (P < 0·05), an increase of IgG4 (P < 0·01) and a lower cure rate (P < 0·01) compared with those who were negative for both cytokines. These results suggest that persistent infection with HTLV‐I affected S. stercoralis‐specific immunity and reduced therapeutic efficacy.

Predictive markers for development of strongyloidiasis in patients infected with both Strongyloides stercoralis and HTLV-1

Severe strongyloidiasis has often been reported to occur in some patients infected with both Strongyloides stercoralis (S. stercoralis) and human T‐cell leukaemia virus type 1 (HTLV‐1); however, there are few useful predictive markers for the risk of development of strongyloidiasis in these patients. To search for such predictive markers, we examined peripheral blood and stool samples of individuals infected with both S. stercoralis and HTLV‐1 in Okinawa, Japan, an area in which both of these are endemic. The HTLV‐1 proviral load and antibody titre were examined in relation to the S. stercoralis load as measured by the direct faecal smear method in patients infected with both S. stercoralis and HTLV‐1. The Epstein‐Barr virus (EBV)‐associated nuclear antigen (EBNA) antibody titre was also measured in these patients in order to examine the relationship between host immunity and HTLV‐1 proviral load or antibody titre. The direct faecal smear‐positive group showed both a higher HTLV‐1 proviral load and HTLV‐1 antibody titre than the ‐negative group (P < 0·05). In contrast, inverse correlations of these parameters with the EBNA antibody titre were observed, especially for proviral load (ρ = −0·387, P <  0·05). These results suggest that HTLV‐1 proviral load and antibody titre influence the S. stercoralis load via disturbance of the host immunity, and that proviral load would be an especially useful predictive marker of the risk of development of strongyloidiasis in patients infected with both S. stercoralis and HTLV‐1.

Expansion of unconventional T cells with natural killer markers in malaria patients

Immunological states during human malarial infection were examined. In parallel with parasitemia and anemia, granulocytosis was induced in the blood of patients, especially those infected with Plasmodium (P.) falciparum. At that time, the level of lymphocytes remained unchanged or slightly increased in the blood. However, the distribution of lymphocyte subsets was modulated, showing that the proportion of CD56(+)T cells, CD57(+)T cells, and gammadeltaT cells (i.e. all unconventional T cells) had increased in patients infected with P. falciparum or P. vivax. This phenomenon occurred at the early phase of infection and disappeared in the course of recovery. The data from patients with multiple attacks of P. vivax infection showed that there was no augmentation of these responses. In adult cases, the increase in the proportion of unconventional T cells seemed to closely parallel disease severity. However, all these responses were weak in children, even those infected with P. falciparum. In conjunction with accumulating evidence from mouse malaria experiments, the present results suggest that the immunological state induced by malarial infection might mainly be an event of unconventional T cells and that the immunological memory might not be long-lasting, possibly due to the properties of unconventional T cells.

Effects of spleen cells and serum on transfer of immunity to Strongyloides venezuelensis infection in hypothymic (nude) mice

The course of Strongyloides venezuelensis infection in congenitally hypothymic (nu/nu) mice and their heterozygous thymus-bearing littermates (nu/+) was followed. Unlike the infected nu/+ mice, the nu/nu mice were unable to expel the worms until the end of the observation period (98 days post-infection). In addition, about three times as many eggs were counted at the peak level of infection in faeces of the infected nu/nu mice in comparison with the nu/+ mice. No acquired resistance to rechallenge was observed among the nu/nu mice. Auto-reinfection within the infected nu/nu mice could not be supposed in the present study. The worm expulsion mechanism was generated by nu/nu mice which had been given syngeneic spleen cells from intact +/+ mice. The expulsion of adult worms, as well as the protection against migrating larvae, occurred anamnestically when spleen cells from immune +/+ mice were transferred. The serum transfer, however, only caused a retardation of larval migration. The results support the hypothesis that direct worm immunity and worm expulsion are a T cell-dependent phenomenon.

Application of enzyme-linked immunosorbent assay for mass examination of strongyloidiasis in okinawa, Japan

The enzyme-linked immunosorbent assay was tested to evaluate whether it could be applicable in screening for mass examination of strongyloidiasis. A total of 2906 inhabitants in three areas (858 in Gushikawa Village, 849 in Nakazato Village and 1199 in Sashiki Town) were screened by the enzymatic assay and approximately 11-30% (11.8% in Gushikawa, 17.0% in Nakazato and 27.7% in Sashiki) were considered to be antibody positive. In the parasitological follow-up examinations of those who were antibody positive, actual infection was found in more than half (51%) the subjects. The overall infection rates estimated from the results reached 5.8% in Gushikawa, 9.1% in Nakazato and 14.0% in Sashiki (mean = 10.4%). The infection rates were significantly higher than those in previous surveys conducted in the same areas. The ELISA technique was found to be useful for strongyloidiasis screening and for seroepidemiological purposes in Okinawa.