Naoki Kuwabara

The role of T lymphocytes in patients with food-sensitive atopic dermatitis

The role of T lymphocytes was assessed in patients with food-sensitive atopic dermatitis (AD). T lymphocytes plus monocytes responded well to ovalbumin or bovine serum albumin (BSA) in children with AD who were sensitive to hens egg or cows milk compared with healthy children and children with immediate allergic symptoms who are sensitive to hens egg or cows milk. The responding cells were shown to be predominantly CD4+ T lymphocytes. Interleukin-2 activity and interferon-gamma concentrations in culture supernatants of ovalbumin-stimulated peripheral blood mononuclear cells (PBMCs) from patients with AD who were sensitive to hens egg were significantly higher than those of healthy children and patients sensitive to hens egg with immediate symptoms. Expression of Fc epsilon R II on B lymphocytes in cultures of ovalbumin-stimulated PBMCs from patients with AD was significantly higher than that of healthy children, but it tended to be lower than that of patients with immediate symptoms. These results suggest that, in patients with AD who are food sensitive, CD4+ T lymphocytes stimulated by food antigens secrete lymphokines such as interleukin-2 and interferon-gamma that are secreted from TH1 clones in mice, and express Fc epsilon R II on B lymphocyte that is induced by interleukin-4 secreted from TH2 clones in mice. Taken together, cell-mediated immunity may also occur in addition to IgE-mediated hypersensitivity in patients with food-sensitive AD.

Long‐term study of the immunodeficiency of Bloom's syndrome

The immune state was evaluated over a 10‐year period in two individuals with Blooms syndrome. In both patients, serum concentrations of IgM were markedly low. Mildly decreased serum concentrations of IgG and IgA increased significantly with age, whereas the IgM levels remained low. From assessments of B‐cell and T‐cell functions in pokeweed mitogen‐induced immunoglobulin production, the IgM deficiencies were thought to result from B‐cell dysfunction. T‐cell function appeared intact. Moreover, although the percentages of surface IgM‐bearing cells were not reduced, the numbers of IgM‐secreting cells were reduced. These findings suggest that the IgM deficiency is due to an abnormality in the maturation of surface IgM‐bearing B cells into IgM‐secreting cells.

METHYLATION PATTERNS OF I ε REGION IN B CELLS STIMULATED WITH INTERLEUKIN 4 AND EPSTEIN-BARR VIRUS IN PATIENTS WITH A HIGH LEVEL OF SERUM IgE

Human IgE synthesis requires the presence of both interleukin 4 (IL‐4) and T‐cells. However, it is not clear what role IL‐4 and T‐cells play in the induction of IgE synthesis at the level of gene regulation. B cells that were obtained from patients with a high level of serum IgE and from healthy donors were immortalized by Epstein‐Barr virus. We examined IgE production of these B cells stimulated with IL‐4. Supernatant IgE levels of patients B cells cultured with or without IL‐4 were higher than those of healthy donors B cells. Our results indicated that B cells stimulated with IL‐4 from patients produced IgE, germline C ε transcript, and S μ S ε recombination. The germline C e transcript was dose‐dependently induced in the presence of IL‐4 and related to the supernatant IgE level. In B cells stimulated with IL‐4 that were obtained from patients, (some of the) DNA near or within the I e region was (already partly) unmethylated, unlike those from healthy donors, and there was a loss of methyl groups of the DNA upon the addition of IL‐4 in B cells from both patients and normal donors. IgE synthesis of B cells stimulated with IL‐4 in patients with a high level of serum IgE is due to an accessibility in the immunoglobulin heavy‐chain isotype switch, and this may reflect the accessibility in synthesis of germline C ε transcript, which may be caused by the increase of opening chromatin structures because of their unmethylation in the I ε region.

Improvement of food‐sensitive atopic dermatitis accompanied by reduced lymphocyte responses to food antigen following natural measles virus infection

Five patients with atopic dermatitis (AD) who were sensitive to hens egg were observed before and after natural measles virus infection. Within 4 weeks of natural measles virus infection, the eczematous lesions clearly improved in four of the five patients in whom neither offending foods were eliminated, nor anti‐allergic drugs, systemic steroids and steroid ointment administered. This was accompanied by reduced proliferative responses of peripheral blood mononuclear cells (PBMCs) to ovalbumin (OA). Another patient showed a transient improvement of AD symptoms, from severe to mild, and thereafter returned to severe accompanied by increased proliferative responses of PBMCs to OA. Radioallergosorbent test (RAST) scores for hens egg in all five patients did not change in each level in each patient, except the transiently decreased RAST scores for hens egg in one patient, after the infection. Thus, in patients with AD who are sensitive to food, the improvement of AD symptoms that appeared within 4 weeks of natural measles virus infection was related to reduced proliferative responses of PBMCs to the food antigen following the infection.

Timing of onset of allergic symptoms as a response to a double-blind, placebo-controlled food challenge in patients with food allergy combined with a radioallergosorbent test and the evaluation of proliferative lymphocyte responses.

The timing of onset of allergic symptoms in a double-blind, placebo-controlled food challenge (DBPCFC) was combined with a radioallergosorbent test (RAST) and evaluation of proliferative responses of peripheral blood mononuclear cells (PBMCs) in 27 atopic dermatitis patients with hens egg allergy. Of the 27 patients, 11 showed reactions within 2 h (immediate group), and 16 showed reactions after more than 2 h (nonimmediate group) after ingestion of hens egg. The RAST scores for hens egg in the immediate group were significantly (p < 0.01) higher than those in the nonimmediate group, while the proliferative responses of PBMCs to ovalbumin in the nonimmediate group were significantly (p < 0.01) higher than those in the immediate group. These results suggest that the RAST values are related to immediate allergic symptoms in DBPCFC and the proliferative responses of PBMCs are related to nonimmediate allergic symptoms in DBPCFC. The timing of onset of allergic symptoms in DBPCFC will render DBPCFC more useful in the diagnosis of food allergy, including the late-onset reactions to foods.

Interferon-γ and interleukin-4 production of ovalbumin-stimulated lymphocytes in egg-sensitive children

BACKGROUND The causal relation between egg allergy and cytokines derived from lymphocytes is unknown. OBJECTIVE Interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) production of ovalbumin-stimulated and interleukin-2 (IL-2)-stimulated peripheral blood mononuclear cells from egg-sensitive patients was investigated and compared with that of stimulated peripheral blood mononuclear cells from nonatopic healthy children. METHODS Peripheral blood mononuclear cells from egg-sensitive patients and nonatopic healthy children were cultured with ovalbumin and IL-2 for seven days. The IFN-gamma and IL-4 concentrations in culture supernatants of the peripheral blood mononuclear cells were investigated. RESULTS The levels of IFN-gamma production of only IL-2-stimulated or both ovalbumin-stimulated and IL-2-stimulated peripheral blood mononuclear cells from egg-sensitive patients with atopic dermatitis was significantly higher than that of healthy children and that of egg-sensitive patients with immediate allergic symptoms. CONCLUSIONS Increased IFN-gamma production by lymphocytes after IL-2 and antigen stimulation has important implications for the mechanism of food-sensitive atopic dermatitis.

Genetic Analysis of IgE and the IGHE, IGHEP1 and IGHEP2 Genes in Atopic Families

The familial occurrence of allergic diseases was studied in 478 individuals and their family members. The results showed that there was pronounced familial clustering. Total serum IgE concentrations of atopic patients and their parents were well correlated. Moreover, the concentration of specific IgE of the patients and their parents was also well correlated, suggesting that IgE production was genetically determined. To determine if major structural abnormalities of IGHE, IGHEP1 and IGHEP2 genes might lead to aberrant control and subsequent increase in IgE concentration, genomic DNAs from 55 individuals, i.e., 31 atopic patients and their family members, were examined. We detected the IGHE, IGHEP1 and IGHEP2 genes in all 55 leukocyte DNAs. We could not find any large deletions or duplications in the IGHE, IGHEP1 and IGHEP2 genes of atopic patients with high serum IgE concentrations.

Common variable immunodeficiency with increased surface IgM-positive double-bearing B cells.

We report a case of common variable immunodeficiency (CVI) that shows low levels of IgG and IgA, but a normal quantitative or qualitalive level of IgM. T‐cell functions were not disturbed. Increased numbers of surface IgM (sIgM) and sIgD, sIgM and sIgA, sIgM and sIgA double bearing B cells were observed as compared with a control. No IgG and IgA induction upon stimulation with Staphylococcus aureus Cowan I (SAC)and recombinant interleukin‐2(rIL‐2), or pokeweed mitogen (PWM) and rIL‐4 or rIL‐6 was observed, although there was proliferation. Although; μ mRNA was expressed as much as in a healthy control, transcription of γ mRNA and α mRNA was very low. Furthermore, no enhanced effects of γ mRNA and α mRNA were recognized upon stimulation with rIL‐4 and rIL‐6. These results suggest that the patients B cells might be detective at the switching process from μ, μ and δ, μ and γ to γ or μ and α to α.

Identification of monocyte chemotactic factors in supernatants of ovalbumin‐stimulated lymphocytes from patients with atopic dermatitis who are sensitive to hen's egg

Monocyte chemotactic activities in supernatants of ovalbumin (OA)‐stimulated peripheral blood mononuclear cell (PBMC) cultures were studied in patients with atopic dermatitis (AD) who were sensitive to hens egg. The monocyte chemotactic activities in hens egg‐sensitive AD patients were significantly higher than those of non‐atopic healthy controls and patients with immediate allergic symptoms. However, the monocyte chemotactic activities were not detected in bovine serum albumin‐Stimulated PBMC culture supernatants in patients with AD who were sensitive to hens egg. but not to cows milk. Furthermore, there was significant correlation between the monocyte chemotactic activities and proliferative responses of PBMCs to OA in hens egg‐sensitive AD patients, whereas there was no significant correlation between the monocyte chemotactic activities and radioallergosorbent test values. These results suggest that PBMCs stimulated with food antigens produce monocyte chemotactic factors which relate to the pathogenesis of AD in food‐sensitive AD patients and that the pathogdenesis of AD may be related to cell‐mediated immune responses.

Immunoglobulin secreting cells in lymphocytes of patients with IgA deficiency or hyper-IgA

To assess the ability of immunoglobulin production in vivo, we enumerated the immunoglobulin secreting cells in the peripheral blood of patients with an IgA deficiency and of those with hyper-IgAemia. All seven patients with primary IgA deficiency and two of the three patients with secondary IgA deficiency had low numbers of IgA secreting cells. In all five patients with hyper-IgA the number of IgA secreting cells was increased. Our results suggest that measurement of immunoglobulin secreting cells in PBMCs is useful in the assessment of ability of immunoglobulin production in vivo.