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Introduction: Eosinophilic chronic rhinosinusitis (ECRS) is a refractory chronic disease defined by recurrent nasal polyps with severe eosinophilic infiltration. This is mainly due to enhanced type 2-dominant immune responses, but the underlying mechanism is still not fully understood., Objective and Methods: In the present study, we aimed to determine the characteristics of dendritic cells (DCs) and cytokine profiles of T cells in the peripheral blood of individuals with ECRS and age- and sex-matched healthy controls (HC)., Results and Conclusion: The ratios of myeloid (m)DC1s to DCs and PD-L1+ mDC1s to mDC1s were higher in ECRS patients than in HC. The proportions of plasmacytoid (p)DCs in DCs, and human leukocyte antigen-DR+ pDCs and ILT3+ pDCs in pDCs were lower in ECRS patients than in HC. In a characterization of T cells, IL-4+CD4+, IFN-γ+CD4+, IL-4+IFN-γ+CD4+, IL-4+Foxp3+CD4+, IFN-γ+Foxp3+CD4+, IFN-γ+IL-4-Foxp3-CD4+, IL-4+CD8+, IL-4+IFN-γ+CD8+, and IL-4+Foxp3+CD8+ T-cell populations were significantly higher in ECRS patients than in HC. These results suggest that the enhanced immune regulation of mDC1, diminished capacity of pDCs, and increased proportion of the T-cell phenotypes in peripheral blood might be factors in ECRS pathogenesis., (© 2021 S. Karger AG, Basel.)

Although gemcitabine is an effective chemotherapeutic for pancreatic cancer, severe side effects often accompany its use. Since we have discovered that locally administered C1B domain peptides effectively control tumor growth without any side effects, the efficacy of co-treatment with this peptide and a low dose of gemcitabine on the growth of pancreatic cancer was examined. Two- and three-dimensional cell culture studies clarified that a co-treatment with C1B5 peptide and gemcitabine significantly attenuated growth of PAN02 mouse and PANC-1 human pancreatic cancer cells in 2D and 3D cultures. Although treatment with the low dose of gemcitabine alone (76%) or the C1B5 peptide alone (39%) inhibited tumor growth moderately, a co-treatment with C1B5 peptide and a low dose of gemcitabine markedly inhibited the growth of PAN02 autografts in the mouse peritoneal cavity (94% inhibition) without any noticeable adverse effect. The number of peritoneal cavity-infiltrating neutrophils and granzyme B + lymphocytes was significantly higher in the co-treatment group than in the control group. A significant increase of granzyme B mRNA expression was also detected in human T cells by the co-treatment. Taken together, the current study suggests that C1B5 peptide offers a remarkably effective combination treatment strategy to reduce side effects associated with gemcitabine, without losing its tumoricidal effect., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Calpain is a calcium-dependent cystein protease, and the homologues of schistosome are known as one of vaccine candidate molecules against schistosomiasis. Here, we established two IL-2 producing T cell hybridoma cell lines specific for Schistosoma japonicum calpain, to identify T cell epitope(s) on the molecule. Overlapping 15mer oligopeptides of calpain were synthesized and tested for their stimulatory abilities to the hybridomas. As a result, epitopes recognized by the two hybridoma lines were the same: EQLKIYAQRC. Spleen cells from calpain multiple antigenic peptide (MAP)-immunized BALB/c mice produced IFNgamma upon stimulation with MAP or soluble worm antigen preparation (SWAP). The identification of the T cell epitope to stimulate Th1 response will contribute to the proper design of synthetic vaccines, evaluation of their protective potentials and elucidation of protective mechanisms in murine experimental schistosomiasis.

A Th1/Th2 cytokine imbalance with a predominance of Th1 cytokines has been suggested to be of pathogenic importance in Wegener's granulomatosis. To evaluate the role of Th1/Th2 cytokines in Wegener's granulomatosis, the subsets of Th1, Th2, Tc1 and Tc2 cells from patients with active Wegener's granulomatosis were examined by intracellular cytokine flow cytometry. The population of Tc1 cells (72.0 +/- 14.4 per cent) in Wegener's granulomatosis was significantly increased compared with Tc1 cells (37.3 +/- 14.6 per cent) in control (p<0.05). Th1, Th2 and Tc2 cells in Wegener's granulomatosis were not significantly increased compared with the control cells. These results indicate that the predominance of Tc1 cells might contribute to the mechanism of the pathogenesis of Wegener's granulomatosis.

T-cell mediated immune response, toward the house dust mite (HDM) antigens in particular, has been reported to be involved in the pathogenesis of atopic dermatitis (AD). On the other hand, studies on the infiltrating lymphocytes in the skin lesion of AD revealed oligoclonal T-cell accumulation. However, it is not clear exactly what antigen(s) the accumulating T cells are exactly recognize in situ. Therefore, this study attempted to determine whether or not the clonally expanded T-cell clones in the diseased skin recognize HDM. Specifically, peripheral blood mononuclear cells (PBMC) obtained from six patients with AD, who revealed high titers of anti-HDM IgE, were stimulated with HDM antigens purified from Dermatophagoides pteronyssinus (Dp). T-cell clonotypes expanded by the stimulation were then identified by the analysis of their T-cell receptor (TCR) B-gene sequences using a combination of the reverse transcription-polymerase chain reaction and subsequent single strand conformation polymorphism separation. The Dp-responding T-cell clonotypes were compared with those that accumulated in the AD skin lesion in vivo. Nucleotide sequences of the TCR were also determined. As a result, the Dp stimulation induced oligoclonal T-cell expansion from the originally heterogeneous peripheral T-cell population of AD patients. However, only a small part of the Dp-reacting T-cell clonotypes detected in PBMC was identical to those accumulated in the AD skin lesion in vivo, and vice versa. This indicates that the frequency of the clonal expansion of Dp-specific T-cell clonotypes in the skin lesion of AD would be rather limited.

In view of recent studies on the mechanisms of the survival of peripheral memory T cells, we tested the biologic role of pectate lyase, a pectin-degrading enzyme, as the cross-reactive antigen required for the recurring survival signals for human T cells specific for Cha o 1, a pollen allergen molecule of the Japanese cypress. We determined a 16-mer epitope peptide for the T-cell clone, and prepared synthetic oligopeptides of homologous regions in putative pectate lyase of other plants. Of these homologous peptides, ZePel (Zinnia elegans), ban 17 (banana), and Amb a 1.1 (short ragweed) induced strong proliferative responses of the Cha o 1-specific T-cell clone in vitro. In addition, suboptimal doses of peptide homologs derived from banana and short ragweed enhanced the survival potency of this T-cell clone without detectable proliferative responses to the peptides. When there was no antigen stimulation, the T-cell clone decreased in viable cell number and lost antigen-specific proliferation activity on day 6 during in vitro incubation. On the other hand, T-cell clones incubated with these survival-inducing peptides maintained proliferative activity to Cha o 1 even on day 9. Serum derived from the donor patient did not contain detectable levels of IgE specific to banana or short ragweed by CAP-RAST. These results show that human T cells specific for pollen allergen seem to use cross-reactive pectate lyase peptides to deliver survival signals even in the absence of pollen allergen, and memory T cells maintained in such a manner might be functioning at the onset of allergic pollinosis, although pollen allergens are seasonal.

Background: CD80 (B7-1) and CD86 (B7-2) play an important role in antigen presentation to effector cells. Recent studies have demonstrated that these costimulatory molecules are also expressed on activated T cells. However, the functional role of CD80 and CD86 expressed on allergen-specific T cells in atopic diseases has not yet been clarified., Objective: We sought to determine the functional role of CD80 and CD86 expressed on allergen-specific T cells in atopic diseases., Methods: We assayed the expression of CD80 and CD86 on allergen-specific T-cell lines from patients with perennial allergic rhinitis stimulated by Dermatophagoides farinae-crude (Der f-c) antigen, 1 of the major allergens causing house dust mite allergy. T-cell proliferation induced by Der f-c-specific T-T cell interactions was measured, and the role of CD80 and CD86 in this proliferation was examined. In addition, we compared the proportion of CD45RO+CD86(+) T cells in primary culture of PBMCs stimulated by Der f-c antigen between patients with perennial allergic rhinitis and control subjects., Results: On T-cell activation, CD86 antigen was upregulated earlier than CD80. Both CD80 and CD86 expressed on Der f-c-specific T cells could provide costimulatory signals to induce allergen-specific T-cell proliferation that was partially inhibitable by both anti-CD80 and anti-CD86 mAbs. The proportion of CD45RO+CD86(+) T cells in primary culture from atopic patients was significantly higher than that from control subjects., Conclusion: These results suggest that costimulatory molecules, such as CD80 and CD86, expressed on allergen-specific T cells may be involved in the amplification of allergen-specific immune responses through T-T cell interactions in atopic diseases.

Background: MSP-1 of Plasmodium falciparum induces strong proliferative T cell responses even in malaria-nonexposed individuals. Epitopes recognized by malaria-nonimmune T cells have not been identified, and immunological mechanisms inducing such T cell responses remain to be uncovered. MSP-1 is a vaccine candidate, and it should be understood whether those epitopes have any roles in MSP-1-mediated protective immunity. The T epitopes-inducing malaria-naive T cell response was analyzed in the hope of understanding the underlying mechanisms., Methods: Human T cell lines and clones reactive to MSP-1 of P. falciparum were established from malaria-nonexposed Japanese donors in vitro, and epitope peptides were identified. Sequences of those epitope peptides were compared to unrelated peptides in the data base. One of those peptides was tested for both binding to HLA-DR molecules and inducing proliferative responses of MSP-1-reactive T cells., Results: There are at least 6 epitopes recognized by malaria-naive T cells under the restriction by HLA-DRB1*1502 or 0802. Important amino acids for the T cell recognition were identified for an MSP-1 peptide. A yeast peptide which shared those residues induced proliferative responses of MSP-1-reactive T cells., Conclusion: We identified T epitopes in the N-terminal region of MSP-1, some of which showed molecular similarities with unrelated environmental antigens, suggesting the presence of cross-reactive T epitopes in MSP-1. Cytokine production in response to those epitopes suggests regulatory functions of those T cells during primary infection with P. falciparum.

Responses of peripheral blood mononuclear cells to synthetic oligopeptides of Der p II, one of the major allergen molecules of Dermatophagoides pteronyssinus, were compared between allergic and nonallergic subjects. Healthy subjects showed positive responses to crude extracts of D. pteronyssinus, but only allergic subjects showed elevated cellular responses to Der p II. We synthesized three oligopeptides of Der p II in which motifs of a possible T-cell epitope were included. Of 14 subjects with positive response to Der p II, three responded to all three peptides, while five did not respond to any peptide tested. In 11 allergic patients who showed positive response to Der p II, responsiveness to the peptide K33-T47 was significantly higher than that to other peptides (P < 0.05). All the responding patients were also positive for scratch test to Der p II, suggesting that those epitopes induced IgE-promoting helper T-cell response in allergic persons. On the other hand, the in vitro cellular responses were not necessarily correlated to IgE production against Der p II in healthy subjects.

T-cell responses to pathogenic free-living amoebae, Acanthamoeba sp., were analyzed in healthy Japanese individuals. Of 20 healthy subjects, 10 (50%) showed significant proliferative responses of peripheral blood mononuclear cells to the soluble amoebic antigens in vitro. The antigens used were not mitogenic, and no evidence of amoebic superantigens was available. We established human T-cell clones reactive to Acanthamoeba, all of which were CD3- and CD4-positive, CD8-negative, and TCR-alpha beta-positive. We isolated two strains of Acanthamoeba from two patients, one from a patient with meningoencephalitis (CSF strain) and the other from a patient with keratitis (K strain). Of 13 clones, 11 were reactive to the K-strain as well as to the CSF-strain antigen under human leukocyte antigen (HLA)-DR restriction, whereas the other two were specific for the K-strain antigen. All but one clone tested showed TH1-equivalent functions because these cells produced interferon (IFN)-gamma in response to the amoebic antigen but produced no detectable level of interleukin 4 (IL-4). These results suggest that immunocompetent hosts might have acquired protective immunity mediated by Acanthamoeba-specific T-cells during natural sensitization.

In the present study, we investigated the cellular basis of house dust mite-driven immune responsiveness in an atopic individual with perennial rhinitis. We established 40 human T-cell clones (CD3+, 4+, 8-) reactive to Dermatophagoides pteronyssinus (Dp) antigen under the restriction of HLA-DR. By using the crude Dp antigen and its 14 molecular weight (MW) fractions, we analyzed the distribution of T-cell-recognizing sites in the whole Dp extract. We tested T-cell-mediated immunity through two parameters; the identification of Dp fractions inducing T-cell proliferation, and the ability of T-cell clones to secrete IL-2, IL-4, and IFN-gamma. According to a prominent peak among fraction-driven T-cell proliferation, we observed that T-cell clones that recognized 45,000- to 95,000-MW fractions were common, while clones reactive to 15,000- to 25,000-MW fractions were less frequent. Several clones were also reactive to antigens of Dermatophagoides farinae or other insects. Based on the responses of cloned T cells, we observed at least 9 distinct T epitopes in crude Dp antigen. These T-cell clones had a heterogenous secretory pattern of cytokines. T-cell clones showed no association between their ability to produce regulatory cytokine and their recognition of particular Dp fractions.

To analyse the cross-reactivity of T-cell-mediated immunity between Dermatophagoides pteronyssinus (Dp) and Chironomus yoshimatsui (Cy), the most common allergens in Japan, we established antigen-specific human T-cell lines and clones. Some but not all of the Cy-induced T-cell lines showed a significant proliferative response not only to Cy, but also to Dp. No T-cell line responded to other unrelated antigens. When we stimulated the Dp-induced T-cell clones with Cy, 3 of the 40 clones (7.5%) showed a significant proliferation, and 2 of the 3 clones produced interleukin-4 and interferon-gamma, indicating their helper function. Cross-reactivity was diminished significantly after the absorption of Dp antigen in an anti-Cy affinity column. The cross-reactive epitopes were thought to be expressed on the Dp molecule of 45-53 kD. The presence of helper T cells reactive to both allergens suggests a possibility that this cross-reactivity might be involved in part in the high incidence of allergy to the 2 major allergens.

Serum-mediated regulation of T-cell responses specific for soluble egg antigen (SEA) of Schistosoma japonicum was tested in human hosts. When we added autologous serum to SEA-specific human T-cell lines (CD3+, 4+, 8-), we observed suppression of T-cell proliferation, and this suppressive activity was detected in the immunoglobulin-G2 (IgG2) subclass. Suppression was dose-dependent and antigen-specific. T-cell proliferation induced by only one SEA fraction of greater than 18 kDa was modulated in the presence of 100 micrograms/ml autologous as well as allogeneic infected IgG2. This SEA fraction-driven proliferation was also regulated by suppressor T-cells through distinct suppressive mechanisms. Our results suggest that T-cell responses to a particular component(s) of SEA are strictly regulated through both cellular and humoral mechanisms in human chronic S. japonicum infection.

This study investigated the cellular response of human lymphocytes to Dermatophagoides pteronyssinus crude antigen and 14 molecular weight (MW) fractions. The cells were derived from atopic patients and healthy individuals who were skin test-positive or skin test-negative to mite antigen. When stimulated with crude antigen, the group of patients showed elevated proliferation and production of lymphokine in comparison with the healthy skin test-negative individuals (P less than 0.01). By stimulation with fractions, there was a remarkable variety in the responding patterns to each fraction. However, when expressed as a mean value, the patient group exhibited a sharp and high peak at 95,000 MW, which is different from IgE responses. In the other two groups, no apparent peak response was observed. Lymphokine production by fraction stimulation was studied in six distinct individuals. The most important fractions for inducing lymphokine production differed in each individual tested. Moreover, fractions which induced active lymphokine production were not necessarily the main targets of proliferative response in atopic patients.

We analysed regulatory mechanisms of the human T cell response to soluble egg antigen (SEA) of Schistosoma japonicum in vitro. SEA is a crude antigen mixture containing numerous epitopes. We obtained SEA-induced T cell lines from five patients with chronic schistosomiasis japonica, and tested their proliferative response to molecular weight fractions of SEA. Although all T cell lines showed strong responses to crude SEA, there was a heterogeneity in fraction-driven responsiveness. All but one T cell line tested failed to respond to SEA fraction I (mol. wt greater than 18 kD). One patient who was typed as HLA-DQw1/w4, did not show proliferation of CD4+ T cells to fraction I; however, a fraction I-driven helper T cell response was observed when we added HU-11 monoclonal antibody specific for HLA-DQw1/w4. This indicated that the patient had helper T cells to the fraction even though their response was suppressed. Because HLA-DQ had an effect on functional expression of suppressor T cells, it was suggested that there was epitope-specific regulation of the T cell response to SEA, and HLA-DQ-controlled immune suppression might be involved in the regulatory system in human chronic schistosome infection.

To analyze the possible major T cell recognition site(s) of chironomid antigens, we established human T cell lines and clones (CD3+ 4+ 8-) reactive to soluble extracts of the adult midge of Tokunagayusurika akamusi (TAA) and/or Chironomus yoshimatsui (CYA). All T cell lines and clones proliferated heavily in response to relatively large molecular weight fractions of TAA (MW greater than or equal to 15,000). Nine clones reactive to TAA were classified into 3 groups according to reactivity, indicating the existence of at least 3 distinct T cell recognition sites in TAA. Five T cell clones responded to both TAA and CYA, although the two chironomid antigens were serologically distinct. We conclude that T cell recognition sites of chironomid antigens are different from B cell recognition sites in humans.

T cell lines specific for Schistosoma japonicum egg Ag were established in vitro from patients with chronic schistosomiasis japonica, and investigated their possible immunopathologic roles by testing lymphokines production and in vitro granuloma formation assay. All lines tested had surface phenotypes of CD3+ CD4+ CD8-, and showed S. japonicum soluble egg Ag (SEA)-specific proliferation requiring HLA-DR-restricted Ag presentation. Of these fractions of SEA separated by gel filtration, Fraction II (m.w. 7,000 to 18,000) and III (m.w. 7,000) induced strong proliferation of T cell lines, whereas fraction I (m.w. 18,000+) failed to induce detectable proliferation to any T cell lines tested. One of the T cell lines was cloned by micromanipulation: two of eight clones responded only to fraction II, and six to both fractions II and III. We observed that four of eight clones tested produced IL-2 in response to SEA, and three of them were able to transfer S. japonicum egg-specific granulomatous hypersensitivity in vitro to an HLA haplo-identical individual without previous schistosome infection. These immunopathologic functions of T cell clones seemed to be activated by at least two distinct epitopes of SEA. Our present observations suggest that at least two distinct CD4+ human T cells, both of which recognize epitopes expressed on SEA molecules of less than 18 kDa, might have critical roles in granulomatous hypersensitivity to eggs of S. japonicum in humans.

To characterize the mechanisms of induction and regulation of the cell population involved in granuloma formation around eggs of Schistosoma japonicum, we utilized a simple method of in vitro experiments. Lyt1+2-T cells were essential for in vitro responses to the intact S. japonicum eggs, which were assumed to be comparable to in vivo granulomatous responses. T-cell responses seemed to be macrophage-dependent, and responding T cells produced IL-2-like activities in the culture supernatant. Sera taken from chronically infected mice acted as regulatory factors to these T-cell responses as was the case in in vivo granuloma formation. The method used here was simple and highly informative for the studies on pathogenesis of schistosomiasis japonica.

Antigens that produce an antibody response in some members of a species may fail to do so in others. The response to an antigen is controlled by a gene termed the immune response (Ir) gene, which is transmitted as a single dominant trait. We have provided evidence for similar immune suppression (Is) genes which control non-responsiveness through the antigen specific suppressor T cell. The non-responsiveness is also dominantly inherited and the Is genes are linked to the histocompatibility (HLA) antigen system. Here we report that the HLA-DR2 molecule from a non-responder haplotype (HLA-Dw12-DR2-DQwl) is required for the proliferative T cell response to schistosoma japonicum (Sj) antigen, as a restriction element, indicating that the HLA-DR2 is the product of the Ir gene, and that the HLA-DQwl molecule of the non-responder haplotype is important in the antigen-specific suppression of the response to this antigen, suggesting that it is the product of the Is gene. We therefore conclude that the HLA-DR and DQ molecules, which are controlled by the distinct genes in the MHC multigene family, regulate immune response and immune suppression and that the gene for HLA-DQ is epistatic to that for HLA-DR in controlling the immune response to schistosomal antigen in humans.

Immune responsiveness to schistosomal worm antigen was investigated in the individuals infected with Schistosoma japonicum by measuring the antigen-specific proliferative response of the peripheral T lymphocytes in vitro. Out of 57 infected individuals, 10 (17.5%) were low responders, whereas 47 (82.5%) were high responders to this antigen. The strong association between the HLA-Aw24-Bw52-Dw12 haplotype and the low responder group was demonstrated. Because the association of the low responder group was strongest with the HLA-D specificity, Dw12, and because the HLA-D region was assumed to be comparable to the I region of the murine H-2 complex, it was suggested that the low responsiveness to schistosomal worm antigen was controlled by a single dominant immune suppression gene that was in strong linkage disequilibrium with HLA-Dw12.