Original Article

Journal of Investigative Dermatology (1986) 86, 29–33; doi:10.1111/1523-1747.ep12283762

Complement Fixation by Pemphigus Antibody. III. Altered Epidermal Cell Membrane Integrity Mediated by Pemphigus Antibody and Complement

Seiji Kawana1, William D Geoghegan2 and Robert E Jordon

  1. 1Department of Dermatology, Kitasato University School of Medicine, 1-15-1 Kitasato, Sagamihara, Kanagawa 228, Japan
  2. 2Cutaneous Immunopathology Unit and Department of Dermatology, University of Texas Health Science Center, Houston, Houston, Texas, U.S.A.

Received 13 May 1985; Accepted 12 August 1985.

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Abstract

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 arid 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 depiction 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.

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References

  1. Beutner, EH, Jordon, RE: Demonstration of skin antibodies in sera of pemphigus vulgaris patients by indirect immunofluorescence staining. Proc Soc Exp Biol Med 1964 177: 505–510,
  2. Diaz, LA, Marcelo, CL: Pemphigoid and pemphigus antigens in cultured epidermal cells. Br J Dermatol 1978 98: 631–637,  | PubMed | ChemPort |
  3. Schiltz, JR, Michel, B, Papay, R: Pemphigus antibody interaction with human epidermal cells in culture: A proposed mechanism for pemphigus acantholysis. J Clin Invest 1978 62: 778–788,  | PubMed | ChemPort |
  4. Singer, KH, Sawka, NJ, Samowitz, HR, Lazarus, GS: Proteinase activation: A mechanism for cellular dyshesion in pemphigus. J Invest Dermatol 1980 74: 363–367,  | Article | PubMed | ChemPort |
  5. Hashimoto, K, Shafran, KM, Webber, PS, Lazarus, GS, Singer, KH: Anti-cell surface pemphigus autoantibody stimulates plasminogen activator activity of human epidermal cells. A mechanism for the loss of epidermal cohesion and blister formation. J Exp Med 1983 157: 259–272,  | Article | PubMed | ISI | ChemPort |
  6. Nishikawa, T, Kurihara, S, Harada, T, Sugawara, M, Hatano, H: Capability of complement fixation of pemphigus antibodies in vitro. Arch Dermatol Res 1977 260: 1–6,  | PubMed | ChemPort |
  7. Hashimoto, T, Sugiura, M, Kurihara, S, Nishikawa, T: In vitro complement activation by intercellular antibodies. J Invest Dermatol 1982 78: 316–318,  | Article | PubMed | ChemPort |
  8. Kawana, S, Janson, M, Jordon, RE: Complement fixation by pemphigus antibody. I. In vitro fixation to organ and tissue culture skin. J Invest Dermatol 1984 82: 506–510,  | Article | PubMed | ChemPort |
  9. Kawana, S, Geoghegan, WD, Jordon, RE: Complement fixation by pemphigus antibody. II. Complement enhanced detachment of epidermal cells. Clin Exp Immunol 1985 61: 517–525,  | PubMed | ChemPort |
  10. Cram, DL, Fukuyama, K: Immunochemistry of ultraviolet-induced pemphigus and pemphigoid lesions. Arch Dermatol 1972 106: 819–824,  | Article | PubMed | ISI | ChemPort |
  11. Jordon, RE, Schroeter, AL, Rogers, RS III, Perry, HO: Classical and alternate pathway activation of complement in pemphigus vulgaris lesions. J Invest Dermatol 1974 63: 256–259,  | Article | PubMed | ISI | ChemPort |
  12. Joost, T, Cormane, RH, Pondman, KW: Direct immunofluorescent study of the skin on occurrence of complement in pemphigus. Br J Dermatol 1972 87: 466–474,  | PubMed |
  13. Janson, M, Geoghegan, WD, Jordon, RE: Separation of human fore-skin epidermal cells on a discontinuous Percoll gradient (abstr). J Invest Dermatol 1982 78: 342,
  14. Beutner, EH, Chorzelski, TP, Jordon, RE: Autosensitivity in Pemphigus and Bullous Pemphigoid. 1970 Springfield, IL, Charles C Thomas, 194 pp
  15. Nisonoff, A, Wissler, FC, Lipman, LN, Woernley, DL: Separation of univalent fragments from the bivalent rabbit antibody molecule by reduction of disulfide bonds. Arch Biochem Biophys 1960 89: 230–244,  | Article | PubMed | ISI | ChemPort |
  16. Kolb, W, Kolb, LM, Podack, HR: C1q: Isolation from human serum in high yield by affinity chromatography and development of a highly sensitive hemolytic assay. J Immunol 1979 122: 2103–2111,  | PubMed | ChemPort |
  17. Deutsch, DH, Mertz, ET: Plasminogen purification from human plasma by affinity chromatography. Science 1970 170: 1095–1096,  | PubMed | ISI | ChemPort |
  18. Dankberg, F, Persidsky, MD: A test of granulocyte membrane integrity and phagocytic function. Cryobiology 1976 13: 430–432,  | Article | PubMed | ChemPort |
  19. Martel, J, Jaramilo, S, Allen, FM Jr, Rubinstein, P: Serology for automated cytotoxicity assays. Vox Sang 1974 27: 13–20,  | PubMed | ChemPort |
  20. Black, L, Berenbaum, MC: Factors affecting the dye exclusion test for cell viability. Exp Cell Res 1964 35: 9–13,  | Article | PubMed | ISI | ChemPort |
  21. Jones, KH, Senft, JA: An improved method to determine cell viability by simultaneous staining with fluorescein diacetate-propidium iodide. J Histochem Cytochem 1985 33: 77–79,  | PubMed | ChemPort |
  22. Tennant, JR: Evaluation of the trypan blue technique for determination of cell viability. Transplantation 1964 2: 685–694,

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