Miyuki Azuma

Studies on Murine IgE with Monoclonal Antibodies

Rat monoclonal antibodies were constructed by fusion of immunized rat spleen cells with a nonsecreting mouse myeloma cell. Two monoclonal antibodies (6HD5 and HMK-12) were selected for further study. Both reacted with various IgE molecules of different specificities and different allotypes, but did not react with immunoglobulins of other isotypes and with light chains. These antibodies were therefore anti-isotypic (IgE) and not anti-allotypic or anti-idiotypic. It was shown by competition studies that these antibodies recognize different epitopes on the FcR epsilon fragment. A sensitive ELISA for the quantitation of murine IgE was developed with these monoclonal antibodies; the sensitivity was between 2 and 250 ng/ml for detection of serum IgE levels. Good correlation was obtained with protein amounts as determined by enzyme-linked immunosorbent assay (ELISA) and passive cutaneous anaphylaxis (PCA) activities. Both monoclonal antibodies were used to study anaphylactic reactions elicited by IgE antibodies. Both could inhibit PCA reactions and both could elicit reverse PCA reactions.

Functional CD86 (B7-2/B70) is predominantly expressed on Langerhans cells in atopic dermatitis.

Recently, we reported the functional expression of CD86 on cultured human Langerhans cells derived from normal epidermis. In the present study, we investigated the expression and function of co‐stimulatory molecules in the pathogenesis of atopic dermatitis. In immunohistochemical analysis, CD80 and/or CD86 were detected on dendritic‐shaped cells not only in the epidermis but also in the dermis in the inflammatory lesions of atopic dermatitis (n = 12). CD80 was expressed in only five cases (42%), while CD86 was expressed in all cases (100%). These molecules were not detected in normal control subjects (n = 8). In non‐lesional skin of atopic dermatitis (n = 4). CD86 but not CD80 was detected in one case. CD86 was preferentially induced on dendritic‐shaped cells in positive patch test sites to Dermatophagoides pteronyssinus or house dust allergen in atopic dermatitis (n = 4). The CD80‐ or CD86‐positive cells were confirmed as Langerhans cells by double immunostaining using anti‐CD1a monoclonal antibody. Neither CD86 over that CD80 was detected n keratinocytes. Similar results of the stronger expression of CD86 over that of CD80 were obtained from psoriasis vulgaris (n = 11) and from contact dermatitis (n=7), although CD86 was expressed only in 57% of the contact dermatitis cases. The percentage of Langerhans cells positive for CD86 was higher than for CD80, i.e. 48% compared with 9%, respectively, in the epidermis of lesional skin of atopic dermatitis (n=8). The expression rate of these molecules on Langerhans cells increased in the dermis. To investigate the function of co‐stimulatory molecules on Langerhans cells in atopic dermatitis, we conducted an inhibition test with antibodies. Anti‐CD86 monoclonal antibody almost completely nhibited T‐cell proliferation stimulated with crude extract of D. pteronyssinus in the presence of epidermal cells as antigen‐presenting cells, whereas anti‐CD80 monoclonal antibody produced less of an inhibitory effect. These data indicate that CD86 expressed on Langerhans cells may play an important part in the pathogenesis of atopic dermatitis.for Investigative Dermatology. Washington, DC (1–5 May 1996).

Soluble Fas molecule in the serum of patients with systemic lupus erythematosus

The serum level of soluble Fas (sFas) molecules in 35 patients with SLE was determined by enzyme-linked immunosorbent assay (ELISA) and its relation to other lymphocyte activation markers and clinical parameters was examined. The level of sFas increased significantly compared to that in normal subjects, consistent with previous reports. There was a significant correlation between the level of sFas and that of sCD4, suggesting some relation between sFas and activation of CD4+ T cells. Patients with lymphopenia tended to have low levels of sFas, making it possible to hypothesize that sFas protects against apoptosis. Although the change in the level of sFas with steroid therapy was variable, some relation to the differential activation of T cell subsets was suggested.

Interferon-gamma differentially regulates CD80 (B7-1) and CD86 (B7-2/B70) expression on human Langerhans cells

CD80 (B7‐1) and CD86 (B7‐2/B70) have recently been identified in cultured human Langerhans cells (LCs), although their role and regulatory properties remain unclear. We present our comparison of the expression of the molecules, mRNAs and the function between CD80 and CD86 in human LCs treated by interferon γ (IFN‐γ). We examined the regulatory properties of CD80 and CD86 expression in human LCs pretreated with IFN‐γ. Flow cytometric analysis indicated that the mean fluorescence intensity of CD86 but not CD80 was enhanced. However, the percentage modulation of both CD80 and CD86 positive cells were significantly up‐regulated in a dose‐dependent manner, after 48‐h culturing with IFN‐γ. The regulatory properties of CD80 and CD86 mRNA expressions in human LC were studied using polymerase chain reaction methods. We found that both CD80 and CD86 mRNA of enriched LCs following IFN‐γ pretreatment for 12 h were higher than those without pretreatment.