Complement fixation by pemphigus antibody. III. Altered epidermal cell membrane integrity mediated by pemphigus antibody and complement.
The present study investigates the effects of pemphigus IgG and complement upon cell viability and/or membrane integrity using trypan blue exclusion, ethidium bromide (EB) staining, and fluorescein diacetate (FDA) conversion by living cells. Forty-eight-hour cultivated epidermal monolayers of neonatal BALB/c mice were incubated in media containing 1 mg/ml purified pemphigus IgG for 48 h in either the presence or absence of complement (absorbed AB sera). Adherent and detached cells were examined by both phase and fluorescence microscopy. Results from trypan blue exclusion showed that pemphigus IgG plus complement produced a modest decrease in exclusion of the dye compared to pemphigus IgG without complement. When FDA/EB comparisons were made, however, the differences were more substantial. When complement plus pemphigus IgG was added to cultures, the number of FDA-positive adherent cells decreased significantly and the number of EB-positive detached cells increased significantly. The effects of complement were inhibited by the use of heat-inactivated AB sera or by C1q depletion of AB sera. No significant effect on the cells was observed in the presence or absence of complement when pemphigus F(ab')2 fragments or when normal IgG was used. Plasminogen depletion of the complement source did not interfere with complement and pemphigus IgG effects as judged by the FDA/EB assay. These studies suggest that pemphigus antibody in the presence of complement alters cell membrane integrity and supports the contention that complement may play a significant role in the mechanism of acantholysis.