Appearance of “Pemphigus Acantholysis Factor” in Human Skin Cultured with Pemphigus Antibody

Abstract

These studies deal with the mechanism of pemphigus IgG-induced epidermal acantholysis. When normal human skin was culted with defined medium containing IgG from pemphigus serum, extensive epidermal acantholysis developed and heat-labile proteolytic enzyme(s) were recovered in the culture medium. The enzyme(s) displayed maximal activity at pH 6.5 when a 3H-amino acid-labeled, insoluble epidermal cell material was used as substrate. The enzyme activity increased during the first 3 days of culture and the appearance of maximal activity coincided with the time of onset of acantholysis. Acantholysis did not occur in control cultures incubated with normal IgG and the enzyme did not appear in the medium or in aqueous extracts of cultured tissues. The enzyme(s) is probably not of lysosomal origin because low pH-active proteases characteristic of these organelles remained within the cells. The effects of puromycin on appearance of enzyme activity, acantholysis and cell viability was studied. At cytotoxic concentrations, the appearance of the enzyme(s) and acantholysis were prevented, whereas at less toxic concentrations enzyme activity and acantholysis were not prevented. Because inhibition of protein synthetic rates by puromycin could not be dissociated from the cytotoxic effects, it is uncertain whether enzyme appearance and acantholysis were dependent upon living tissue or on specific protein synthesis. After pemphigus IgG was removed from the conditioned medium by DEAE cellulose and affinity column chromatography, the remaining material contained enzyme activity and caused acantholysis in fresh skin explants. Similar activities were not present in normal IgG-containing conditioned medium or unfractionated epidermal extracts from normal skin. These data indicate that when the pemphigus IgG autoantibody interacts with epidermal cell surface antigens, the cell responds by synthesis or activation of a non-IgG "pemphigus acantholysis factor" (PAF) which may be a nonlysosomal proteolytic enzyme. It is suggested that PAF causes loss of adhesion between keratinocytes and ultimately produces the characteristic acantholytic cells of pemphigus.

Distinction between epidermal antigens binding pemphigus vulgaris and pemphigus foliaceus autoantibodies.

“…In vitro models using organ culture of skin have demonstrated that PV or PF IgG, in the absence of complement or inflammatory cells, can cause acantholysis (5)(6)(7). It is thought that the pemphigus IgG bound to the surface of the epidermal cell causes the release of one or more proteolytic enzymes, which results in acantholysis (7,(10)(11)(12)(13)(14).…”

Pemphigus Antibodies Identify a Cell Surface Glycoprotein Synthesized by Human and Mouse Keratinocytes

“…Although light microscopic tissue immunofluorescence patterns appear to be intercellular, immunofluorescence performed on cell suspensions and immunoelectron microscopy have demonstrated that pemphigus antibodies bind to the keratinocyte cell surface (4)(5)(6)(7)(8) (9). The binding of pemphigus IgG to the keratinocyte cell surface probably results in the release of proteolytic enzymes that can cause acantholysis (6,(10)(11)(12). Thus, the interaction of pemphigus antibody with a cell surface antigen or antigens is probably a mediator of disease.…”

Characterization of keratocalmin, a calmodulin-binding protein from human epidermis.

“…The loss of staining is not likely to be due to autoantibody blocking because the staining of ROC 129.1 is preserved in perilesional skin, an area in which PF antibodies are bound (33). Destruction ofdesmosomal components by the ensuing inflammatory events after PF antibodies are bound may explain the augmentation ofblister formation by complement and proteases found in some experimental systems (34)(35)(36)(37)(38) and the loss of ROC 129.1 reactivity in the base of the lesions.…”