Innate-like Cytotoxic Function of Bystander-Activated CD8+ T Cells Is Associated with Liver Injury in Acute Hepatitis A.

Acute hepatitis A (AHA) involves severe CD8+ T cell-mediated liver injury. Here we showed during AHA, CD8+ T cells specific to unrelated viruses became activated. Hepatitis A virus (HAV)-infected cells produced IL-15 that induced T cell receptor (TCR)-independent activation of memory CD8+ T cells. TCR-independent activation of non-HAV-specific CD8+ T cells were detected in patients, as indicated by NKG2D upregulation, a marker of TCR-independent T cell activation by IL-15. CD8+ T cells derived from AHA patients exerted innate-like cytotoxicity triggered by activating receptors NKG2D and NKp30 without TCR engagement. We demonstrated that the severity of liver injury in AHA patients correlated with the activation of HAV-unrelated virus-specific CD8+ T cells and the innate-like cytolytic activity of CD8+ T cells, but not the activation of HAV-specific T cells. Thus, host injury in AHA is associated with innate-like cytotoxicity of bystander-activated CD8+ T cells, a result with implications for acute viral diseases.

Anti-inflammatory effects of intranasal cyclosporine for allergic rhinitis in a mouse model.

BACKGROUND: Although topical corticosteroids are considered a safe and effective drug for allergic rhinitis (AR), some AR patients do not show sufficient symptomatic improvement by use of topical corticosteroids. Topical cyclosporine is a safe and effective drug for patients with allergic conjunctivitis, particularly for those with steroid-resistant allergic conjunctivitis. We investigated the anti-inflammatory effects of intranasal cyclosporine for AR using a mouse model. METHODS: After establishment of allergic inflammation in 5-week-old BALB/c mice, cyclosporine was administered intranasally 3 times per week for 2 weeks. To confirm its anti-inflammatory effects, triamcinolone acetonide (TAC) was utilized as a control drug. Histopathologic changes were evaluated using Sirius red and Giemsa staining for eosinophilic and mast cell infiltration, respectively. The levels of cytokines in sinonasal tissues, including tumor necrosis factor (TNF), interleukin (IL)-4, IL-5, and IL-13, were assessed based on a cytometric bead array. RESULTS: The degree of eosinophilic infiltration was significantly decreased by instillation of cyclosporine, the potency being similar to TAC. However, the number of mast cells was not decreased by cyclosporine or TAC. The levels of TNF, IL-4, IL-5, and IL-13 were significantly decreased after treatment with cyclosporine. CONCLUSION: The anti-inflammatory effects of topical cyclosporine for AR were equivalent to those of topical corticosteroids. Topical cyclosporine may be useful for the treatment of AR, although human studies are required.

Age-related differences in human palatine tonsillar B cell subsets and immunoglobulin isotypes.

The tonsils provide defense of the upper aerodigestive tract against pathogens. Although long known to undergo functional changes with age, the precise changes occurring within tonsillar B cell populations remain undefined. In the present study, we investigated age-related changes in palatine tonsillar B cell subsets and immunoglobulin (Ig) isotypes. Palatine tonsils were obtained from forty-two tonsillectomy patients without tonsillitis who were divided into three groups: young children (4-9 years), adolescents (10-19 years), and adults (20-60 years). Tonsillar B cells were then analyzed by flow cytometry. Using expression of CD38 and IgD to define B cell subsets, we found that the frequency of germinal center (GC) B cells in the tonsils was significantly higher, and the frequency of memory B cells lower, in young children as compared to adolescents and adults. Within the GC B cell subsets, adults had a higher frequency of IgA(+) cells and a lower frequency of IgM(+) cells as compared to individuals in the younger age groups. Moreover, young children had a higher frequency of IgG(+) cells in the GC B cell subsets than did individuals in the older age groups. We also observed an abundance of IgM(+) cells among memory B cells and plasmablasts in young children and IgA(+) cells in adults. In summary, the proportion of GC B cells in palatine tonsillar B cells decreases with age, while the proportion of memory B cells increases with age. In addition, Ig isotypes in tonsils preferentially switch from IgM to IgA as individuals age.

Comparative Analysis of Liver Injury-Associated Cytokines in Acute Hepatitis A and B.

PURPOSE: Acute hepatitis A (AHA) and acute hepatitis B (AHB) are caused by an acute infection of the hepatitis A virus and the hepatitis B virus, respectively. In both AHA and AHB, liver injury is known to be mediated by immune cells and cytokines. In this study, we measured serum levels of various cytokines and T-cell cytotoxic proteins in patients with AHA or AHB to identify liver injury-associated cytokines. MATERIALS AND METHODS: Forty-six patients with AHA, 16 patients with AHB, and 14 healthy adults were enrolled in the study. Serum levels of 17 cytokines and T-cell cytotoxic proteins were measured by enzyme-linked immunosorbent assays or cytometric bead arrays and analyzed for correlation with serum alanine aminotransferase (ALT) levels. RESULTS: Interleukin (IL)-18, IL-8, CXCL9, and CXCL10 were significantly elevated in both AHA and AHB. IL-6, IL-22, granzyme B, and soluble Fas ligand (sFasL) were elevated in AHA but not in AHB. In both AHA and AHB, the serum level of CXCL10 significantly correlated with the peak ALT level. Additionally, the serum level of granzyme B in AHA and the serum level of sFasL in AHB correlated with the peak ALT level. CONCLUSION: We identified cytokines and T-cell cytotoxic proteins associated with liver injury in AHA and AHB. These findings deepen the existing understanding of immunological mechanisms responsible for liver injury in acute viral hepatitis.

Interleukin-4 enzyme-linked immunospot assay may be useful for diagnosing sensitization to house dust mite.

BACKGROUND: The skin prick test (SPT) is considered a standard test for identification of allergens, but it has some limitations in clinical practice. The multiple allergen simultaneous test (MAST), which measures allergen-specific immunoglobulin E in patients' serum, is a widely used alternative test, but is limited by its relatively low sensitivity and specificity. As a novel diagnostic test to identify allergens, we investigated the sensitivity and specificity of an interleukin-4 (IL-4) enzyme-linked immunospot (ELISpot) assay for Dermatophagoides farinae (Der f) and Dermatophagoides pteronyssinus (Der p). METHODS: Based on the symptoms and SPT results, 43 house dust mite (HDM) allergic rhinitis (AR) patients and 41 control subjects were included. Peripheral blood was drawn from each subject for IL-4 ELISpot assay and MAST. The receiver operating characteristic (ROC) curve analysis was conducted to determine the cutoff values. Sensitivity, specificity, and positive and neg predictive values were compared between the 2 tests. RESULTS: The sensitivity, specificity, and areas under the ROC curve (AUCs) of the IL-4 ELISpot assay were 88.4%, 97.6%, and 0.939 for Der f, and 95.3%, 97.5%, and 0.971 for Der p, respectively. However, the sensitivity, specificity, and AUC of MAST were 76.7%, 73.2%, and 0.777 for Der f, and 69.8%, 75.6%, and 0.788 for Der p, respectively. CONCLUSION: The IL-4 ELISpot assay showed higher sensitivity, specificity, and AUC than MAST, which indicates its clinical feasibility for diagnosing allergy for HDM. A further study is needed to determine the accuracy of the IL-4 ELISpot assay for other common allergens.

Characterization of CD8(+)CD57(+) T cells in patients with acute myocardial infarction.

Although T cells are known to be involved in the pathogenesis of coronary artery disease, it is unclear which subpopulation of T cells contributes to pathogenesis in acute myocardial infarction (MI). We studied the immunological characteristics and clinical impact of CD8(+)CD57(+) T cells in acute MI patients. The frequency of CD57(+) cells among CD8(+) T cells was examined in peripheral blood sampled the morning after acute MI events. Interestingly, the frequency of CD57(+) cells in the CD8(+) T-cell population correlated with cardiovascular mortality 6 months after acute MI. The immunological characteristics of CD8(+)CD57(+) T cells were elucidated by surface immunophenotyping, intracellular cytokine staining and flow cytometry. Immunophenotyping revealed that the CD8(+)CD57(+) T cells were activated, senescent T cells with pro-inflammatory and tissue homing properties. Because a high frequency of CD8(+)CD57(+) T cells is associated with short-term cardiovascular mortality in acute MI patients, this specific subset of CD8(+) T cells might contribute to acute coronary events via their pro-inflammatory and high cytotoxic capacities. Identification of a pathogenic CD8(+) T-cell subset expressing CD57 may offer opportunities for the evaluation and management of acute MI.

DNA immunization of Mycobacterium tuberculosis resuscitation-promoting factor B elicits polyfunctional CD8(+) T cell responses.

PURPOSE: T cell-mediated immune responses, and particularly activation of polyfunctional T cells that simultaneously produce multiple cytokines, are necessary for the control of Mycobacterium tuberculosis. In the present study, we examined if DNA immunization of Mycobacterium tuberculosis resuscitation-promoting factor B (RpfB) elicits polyfunctional T cell responses in mice. MATERIALS AND METHODS: C57BL/6 mice were immunized intramuscularly three times, at 3-week intervals, with RpfB-expressing plasmid DNA. For comparison, protein immunization was performed with recombinant RpfB in control mice. After immunization, RpfB-specific T cell responses were assessed by interferon-γ (IFN-γ) enzyme-linked immunosorbent spot assay and intracellular cytokine staining (ICS), and T cell polyfunctionality was assessed from the ICS data. RESULTS: RpfB DNA immunization induced not only humoral immune responses, but also CD8(+) and CD4(+) T cell responses. Immunodominant T-cell epitopes were identified within RpfB by assays with overlapping peptides. RpfB DNA immunization elicited a polyfunctional CD8(+) T cell response that was dominated by a functional phenotype of IFN-γ(+)/TNF-α(+)/IL-2(-)/CD107a(+). CONCLUSION: RpfB DNA immunization elicits polyfunctional CD8(+) T cell responses, suggesting that RpfB DNA immunization might induce protective immunity against tuberculosis.

Programmed death-1 (PD-1)-dependent functional impairment of CD4(+) T cells in recurrent genital papilloma.

Genital papilloma is caused by human papilloma virus (HPV) infection and recurs frequently. Although T cells are known to play a critical role in the control of HPV infection and papilloma development, the function and phenotype of these cells in the lesion remain to be elucidated. In the present study, we examined the function and phenotype of CD4(+) T cells isolated from the lesions of primary (n = 9) and recurrent (n = 11) genital papillomas. In recurrent papillomas, the frequency of proliferating (Ki-67(+)) CD4(+) T cells was significantly reduced compared with primary papillomas. Cytokine production was evaluated by intracellular cytokine staining in anti-CD3/anti-CD28-stimulated CD4(+) T cells. CD4(+) T cells from recurrent lesions showed impaired production of IL-2, IFN-γ, and TNF-α. Of interest, the frequency of cytokine-producing CD4(+) T cells significantly correlated with the frequency of Ki-67(+)CD4(+) T cells. We also studied expression of programmed death-1 (PD-1), a T-cell exhaustion marker. The frequency of PD-1(+)CD4(+) T cells was significantly increased in recurrent lesions and inversely correlated with the frequency of cytokine-producing CD4(+) T cells. The functional significance of PD-1 expression was determined in blocking assays with anti-PD-L1, which restored cytokine production of CD4(+) T cells from recurrent lesions. Taken together, in recurrent genital papilloma lesions, proliferation, and cytokine production by CD4(+) T cells are impaired and the PD-1/PD-L1 interaction is responsible for the functional impairment of CD4(+) T cells.

Immunosenescent CD8+ T cells and C-X-C chemokine receptor type 3 chemokines are increased in human hypertension.

The pathogenic role of T cells in hypertension has been documented well in recent animal studies. However, the existence of T-cell-driven inflammation in human hypertension has not been confirmed. Therefore, we undertook immunologic characterization of T cells from patients with hypertension and measured circulating levels of C-X-C chemokine receptor type 3 chemokines, which are well-known tissue-homing chemokines for T cells. We analyzed immunologic markers on T cells from patients with hypertension by multicolor flow cytometry. We then measured circulating levels of the C-X-C chemokine receptor type 3 chemokines, monokine induced by γ interferon (IFN), IFN γ-induced protein 10, and IFN-inducible T-cell α chemoattractant, in patients with hypertension and in age- and sex-matched control subjects by the cytometric bead array method. In addition, we examined histological features of IFN-inducible T-cell α chemoattractant expression from renal biopsy specimens of patients with hypertensive nephrosclerosis and control subjects. The total T-cell population from patients with hypertension showed an increased fraction of immunosenescent, proinflammatory, cytotoxic CD8(+) T cells. Circulating levels of C-X-C chemokine receptor type 3 chemokines were significantly higher in patients with hypertension than in control subjects. Furthermore, immunohistochemical staining revealed increased expression of the T-cell chemokine, IFN-inducible T-cell α chemoattractant, in the proximal and distal tubules of patients with hypertensive nephrosclerosis. Immunosenescent CD8(+) T cells and C-X-C chemokine receptor type 3 chemokines are increased in human hypertension, suggesting a role for T-cell-driven inflammation in hypertension. A more detailed characterization of CD8(+) T cells may offer new opportunities for the prevention and treatment of human hypertension.

Reduced antibody responses to the pandemic (H1N1) 2009 vaccine after recent seasonal influenza vaccination.

The vaccination program against the 2009 pandemic H1N1 influenza virus (2009 H1N1) provided a unique opportunity to determine if immune responses to the 2009 H1N1 vaccine were affected by a recent, prior vaccination against seasonal influenza virus. In the present study, we studied the immune responses to the 2009 H1N1 vaccine in subjects who either received the seasonal influenza virus vaccination within the prior 3 months or did not. Following 2009 H1N1 vaccination, subjects previously given a seasonal influenza virus vaccination exhibited significantly lower antibody responses, as determined by hemagglutination inhibition assay, than subjects who had not received the seasonal influenza virus vaccination. This result is compatible with the phenomenon of "original antigenic sin," by which previous influenza virus vaccination hampers induction of immunity against a new variant. Our finding should be taken into account for future vaccination programs against pandemic influenza virus outbreaks.

T-cell dysfunction and inhibitory receptors in hepatitis C virus infection.

Dysfunction of the virus-specific T cells is a cardinal feature in chronic persistent viral infections such as one caused by hepatitis C virus (HCV). In chronic HCV infection, virus-specific dysfunctional CD8 T cells often overexpress various inhibitory receptors. Programmed cell death 1 (PD-1) was the first among these inhibitory receptors that were identified to be overexpressed in functionally impaired T cells. The roles of other inhibitory receptors such as cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) and T cell immunoglobulin and mucin domain-containing molecule 3 (Tim-3) have also been demonstrated in T-cell dysfunctions that occur in chronic HCV patients. Blocking these inhibitory receptors in vitro restores the functions of HCV-specific CD8 T cells and allows enhanced proliferation, cytolytic activity and cytokine production. Therefore, the blockade of the inhibitory receptors is considered as a novel strategy for the treatment of chronic HCV infection.