title: Eosinophil localization to the basement membrane zone is autoantibody- and complement-dependent in a human cryosection model of bullous pemphigoid.
abstract: Bullous pemphigoid (BP) is an autoimmune blistering disease characterized by antibodies (IgG and IgE) targeting cell-substrate adhesion proteins. A variety of BP models suggest that autoantibody-dependent neutrophil degranulation is essential for blister formation. However, lesional biopsies reveal a predominance of eosinophils and few neutrophils. Our goal was to evaluate the role of antibodies and complement in eosinophil localization, degranulation and split formation at the dermo-epidermal junction (DEJ) utilizing a human skin cryosection model of BP paired with a human eosinophilic cell line, 15HL-60. Expression of receptors for IgG (FcγRII), IgE (FcεRI) and complement (CR1 and CR3) was confirmed on 15HL-60 cells using flow cytometry. 15HL-60 expression of granule protein [eosinophil derived neurotoxin (EDN) and eosinophil peroxidase (EPO)] mRNA and their degranulation in vitro was confirmed using RT-PCR and ELISA, respectively. For cryosection experiments, BP or control sera or IgG and IgE antibodies purified from BP sera were utilized in combination with 15HL-60 cells ± fresh complement. Both BP serum and fresh complement were required for localization of 15-HL60 cells to the DEJ. Interestingly, eosinophil localization to the DEJ was dependent on IgG, but not IgE, and complement. However, no subepidermal split was observed. Additionally, the 15HL-60 cells did not degranulate under any experimental conditions and direct application of cell lysate to cryosections did not result in a split. Our observation that eosinophil localization to the DEJ is dependent on IgG mediated complement fixation provides additional insight into the sequence of events during the development of BP lesions.
authors.affiliation: Department of Dermatology, University of Iowa, Iowa City, IA, USA.
authors.forename: Kelly N
authors.initials: KN
authors.lastname: Messingham
authors.affiliation: Department of Dermatology, University of Iowa, Iowa City, IA, USA.
authors.forename: Jeffrey W
authors.initials: JW
authors.lastname: Wang
authors.affiliation: Department of Dermatology, University of Iowa, Iowa City, IA, USA.
authors.forename: Heather M
authors.initials: HM
authors.lastname: Holahan
authors.affiliation: Department of Dermatology, University of Iowa, Iowa City, IA, USA.
authors.forename: Rupasree
authors.initials: R
authors.lastname: Srikantha
authors.affiliation: Department of Dermatology, University of Iowa, Iowa City, IA, USA.
authors.forename: Samantha C
authors.initials: SC
authors.lastname: Aust
authors.affiliation: Department of Dermatology, University of Iowa, Iowa City, IA, USA.|Veterans Administration Medical Center, Iowa City, IA, USA.
authors.forename: Janet A
authors.initials: JA
authors.lastname: Fairley
mesh_terms: D001323:Autoantibodies / Q000097:blood; D001485:Basement Membrane / Q000378:metabolism*; D001706:Biopsy; D016022:Case-Control Studies; D002448:Cell Adhesion; D002460:Cell Line; D003165:Complement System Proteins / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D047209:Eosinophil Peroxidase / Q000378:metabolism; D004804:Eosinophils / Q000166:cytology*; D005434:Flow Cytometry; D019084:Fluorescent Antibody Technique, Indirect; D018922:HL-60 Cells; D006801:Humans; D007073:Immunoglobulin E / Q000097:blood; D007074:Immunoglobulin G / Q000097:blood; D008856:Microscopy, Fluorescence; D009498:Neurotoxins / Q000378:metabolism; D009504:Neutrophils / Q000276:immunology; D010391:Pemphigoid, Bullous / Q000276:immunology* / Q000473:pathology; D012867:Skin / Q000473:pathology
chemical_list: D001323:Autoantibodies; D007074:Immunoglobulin G; D009498:Neurotoxins; D007073:Immunoglobulin E; D003165:Complement System Proteins; D047209:Eosinophil Peroxidase
keywords: autoantibody; bullous; eosinophil; human; skin