Keiji Okubo

Effect of antianaphylactic agents on substance-P induced histamine release from rat peritoneal mast cells

SummarySubstance P is known to be a potent histamine liberator for mast cells. The influence of antianaphylactic agents, disodium cromoglycate (DSCG), ketotifen, and tranilast was studied on substance-P and compound 48/80-induced histamine release from rat peritoneal mast cells. Substance-P induced histamine release was inhibited by these agents, while compound 48/80-induced histamine release was not inhibited by tranilast. Our findings suggest that these antianaphylactic agents are assumed to be effective for cutaneous diseases which might be concerned with substance P and histamine.

Purification of rat cutaneous mast cells with Percoll density centrifugation

SummaryThe skin is the major site on anaphylaxis, and cutaneous mast cells have an important role in its reactions. The isolation and purification of rat cutaneous mast cells are described here. Rat abdominal skin was digested with collagenase and hyaluronidase, and centrifuged with Percoll. The buoyant density of cutaneous mast cells was high, and relatively pure mast cells were obtained. The purity of cutaneous mast cells was 7.4%±2.4% before and 50.0%±6.4% after Percoll density centrifugation; peritoneal mast cells revealed 5.8%±1.3% purity before and 61.0%±10.6% purity after the same procedure. The isolated cutaneous cells released 21.3%±3.8% histamine and the peritoneal mast cells released 55.5%±3.8% histamine upon stimulation with 10 μg/ml compound 48/80. These findings suggest that there are functional subsets of connective tissue mast cells.

Alterations in membrane fluidity during keratinocyte differentiation measured by fluorescence polarization

SummaryThe epidermis shows a distinctive pattern of differentiation wherein keratinocytes proliferate in the basal cell layer and mature into spinous and granular cells. Using a discontinuous density-gradient centrifugation method, guinea-pig keratinocytes were separated into high (HDF), intermediate (IDF), and low (LDF) density fractions. Morphological and flow cytometrical observations demonstrated that HDF, IDF, and LDF were basal, spinous, and granular cell-rich fractions, respectively. Membrane fluidity of the fractionated keratinocytes was measured by diphenylhexatriene fluorescence polarization. Polarization (p)-value of keratinocytes was negatively correlated with temperature. At each temperature, HDF cells showed a lower p-value than IDF or HDF cells except at 40° C. Since a low p-value indicates a high degree of Brownian motion, membrane fluidity is higher in basal cells and lower in spinous and granular cells. Our results indicate that membrane fluidity of guinea-pig keratinocytes decreases during their maturation.

Epidermal Keratinocytes of Bullous Pemphigoid Express Intercellular Adhesion Molecule‐1 (ICAM‐1)

Intercellular adhesion molecule‐1 (ICAM‐1) is the ligand for lymphocyte function associated antigen‐1 (LFA‐1), mediating the adhesion of lymphocytes to vascular endothelium. Keratinocytes are known to express ICAM‐1 in some inflammatory dermatoses. Using an indirect immunofluorescence method, we examined the patterns of ICAM‐1 and LFA‐1 staining in bullous pemphigoid (BP) lesions and compared them to pemphigus vulgaris (PV) cases. ICAM‐1 was expressed on the cell surface and in the cytoplasm of epidermal keratinocytes at the sites of erythematous and bullous lesions of BP. LFA‐1 molecules were expressed on T cells at the basement membrane zone. In addition, HLA‐DR‐positive keratinocytes were observed in the basal layer. ICAM‐1 was not, however, expressed on epidermal keratinocytes in uninvolved skin from BP patients, PV or normal control skin. It is known that ICAM‐1 is expressed on keratinocytes at the site of lymphoid infiltration in cutaneous dermatoses and that LFA‐1‐positive T cells can bind to interferon gamma‐induced ICAM‐1‐positive keratinocytes. Our results suggest that cellular immunity involving ICAM‐1 and LFA‐1 may play a part in the pathogenesis of bullous pemphigoid.