Xiang Ding, Luis A. Diaz, Janet A. Fairley, George J. Giudice, Zhi Liu

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The Anti-Desmoglein 1 Autoantibodies in Pemphigus Vulgaris Sera are Pathogenic Xiang Ding, Luis A. Diaz, Janet A. Fairley, George J. Giudice, Zhi Liu Journal of Investigative Dermatology Volume 112, Issue 5, Pages 739-743 (May 1999) DOI: 10.1046/j.1523-1747.1999.00585.x Copyright © 1999 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Schematic diagram of the recombinant extracellular domain of Dsg1.His (rDsg1.His) and Dsg3.His (rDsg3.His). rDsg1.His and rDsg3.His were constructed and expressed in a baculovirus expression system. Both proteins contain a signal sequence (S), a propeptide (P), and the entire extracellular domain (EC1–5) linked to a six-histidine tail. The black strips represent the Ca2+-binding sites. Journal of Investigative Dermatology 1999 112, 739-743DOI: (10.1046/j.1523-1747.1999.00585.x) Copyright © 1999 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Immunoblotting analysis of recombinant extracellular domains of Dsg1 and Dsg3. rDsg1 and rDsg3 were electrophoresed by 10% SDS–PAGE, transferred to a nitrocellular filter and blotted with a polyclonal rabbit anti-Dsg1 (lanes 1 and 2) or rabbit anti-Dsg3 antibody (lanes 3 and 4). Lanes 1 and 3, Hi-5 cell culture medium; lane 2, medium from Hi-5 cells infected with rDsg1.His baculovirus; lane 4, medium from Hi-5 cells infected with rDsg3.His baculovirus. Journal of Investigative Dermatology 1999 112, 739-743DOI: (10.1046/j.1523-1747.1999.00585.x) Copyright © 1999 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 The immune specificity of affinity purified anti-Dsg1 and anti-Dsg3 from PV patients. Anti-Dsg1 and anti-Dsg3 antibodies were purified from two PV patients by using a rDsg1 or rDsg3 affinity column, respectively. The purified anti-Dsg1 or anti-Dsg3 antibodies were used to immunoprecipitate 125I-labeled rDsg1.His (a) and rDsg3.His (b). The precipitates were fractionated on a 10% SDS–PAGE and visualized by autoradiography. Lanes 1 and 6, positive control sera (P) (a, PF serum; b, PV serum); lanes 2 and 7, normal control sera (N); lanes 3 and 8, total IgG from PV patients; lanes 4 and 9, affinity purified anti-Dsg3 antibodies; lanes 5 and 10, affinity purified anti-Dsg1 antibodies. Journal of Investigative Dermatology 1999 112, 739-743DOI: (10.1046/j.1523-1747.1999.00585.x) Copyright © 1999 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 Pathogenicity of affinity purified anti-Dsg1 and anti-Dsg3. Neonatal BALB/C mice were injected i.d. with anti-Dsg1 (A–C) or anti-Dsg3 (D–F) (0.56–0.8 mg per g body weight), respectively. The clinical, immunologic, and histologic examinations were performed 20 h post IgG injection. Nikolskey sign was seen in mice injected with both anti-Dsg1 (A) and anti-Dsg3 antibodies (D). Direct IF showed the binding of anti-Dsg1 antibodies to the intercellular spaces of subcorneal keratinocytes (B), whereas the binding of anti-Dsg3 were shown to locate to the suprabasal layer of the murine epidermis (E). Hematoxylin and eosin examination demonstrated that anti-Dsg1 antibodies induce subcorneal acantholysis (C), whereas anti-Dsg3 antibodies produce suprabasilar acantholysis (F) in neonatal mice. Site of basal keratinocytes (arrow in B–F). Original magnification, ×200. Journal of Investigative Dermatology 1999 112, 739-743DOI: (10.1046/j.1523-1747.1999.00585.x) Copyright © 1999 The Society for Investigative Dermatology, Inc Terms and Conditions