Bullous Pemphigoid Autoantibodies Directly Induce Blister Formation without Complement Activation

Abstract

Complement activation and subsequent recruitment of inflammatory cells at the dermal/epidermal junction are thought to be essential for blister formation in bullous pemphigoid (BP), an autoimmune blistering disease induced by autoantibodies against type XVII collagen (COL17); however, this theory does not fully explain the pathological features of BP. Recently, the involvement of complement-independent pathways has been proposed. To directly address the question of the necessity of the complement activation in blister formation, we generated C3-deficient COL17-humanized mice. First, we show that passive transfer of autoantibodies from BP patients induced blister formation in neonatal C3-deficient COL17-humanized mice without complement activation. By using newly generated human and murine mAbs against the pathogenic noncollagenous 16A domain of COL17 with high (human IgG1, murine IgG2), low (murine IgG1), or no (human IgG4) complement activation abilities, we demonstrate that the deposition of Abs, and not complements, is relevant to the induction of blister formation in neonatal and adult mice. Notably, passive transfer of BP autoantibodies reduced the amount of COL17 in lesional mice skin, as observed in cultured normal human keratinocytes treated with the same Abs. Moreover, the COL17 depletion was associated with a ubiquitin/proteasome pathway. In conclusion, the COL17 depletion induced by BP autoantibodies, and not complement activation, is essential for the blister formation under our experimental system.

Dysregulation of mCD46 and sCD46 contribute to the pathogenesis of bullous pemphigoid

“…BP is an autoimmune subepidermal blistering disease caused by IgG autoantibodies targeting the noncollagenous 16A (NC16A) domain of human collagen 17 21, 22 . Previous studies using a passive transfer mouse model demonstrated that these autoantibodies mediated the pathogenic mechanisms of BP, including autoantibody binding, complement fixation and activation, immune cell infiltration, proteinase secretion, and subepidermal blister formation.…”

Type XVII collagen coordinates proliferation in the interfollicular epidermis

“…The following antibodies were used: anti-E-cadherin (Cell Signaling Technology, Danvers, Massachusetts, USA; 24E10, RRID: AB_10694492), anti-phospho histone H3 (Ser10) (Merck Millipore, Billerica, Massachusetts, USA, RRID: AB_11210699), anti-PCNA (Dako, Santa Clara, California, USA; PC10, RRID: AB_2160651), anti-BrdU (Dako; M0744, RRID:AB_10013660), anti-loricrin (Covance, Princeton, New Jersey, USA, RRID:AB_10064155), anti-cleaved caspase-3 (Cell Signaling Technology, RRID:AB_2341188), FITC-conjugated anti-CD3e (BioLegend, San Diego, California; 145–2 C11, RRID:AB_394595), Alexa Fluor 488-conjugated anti-F4/80 (Affymetrix, Santa Clara, California, USA; BM8, RRID: AB_893479), FITC (fluorescein isothiocyanate)-conjugated anti-Ly-6G (Beckman coulter, Brea, California, USA; RB6-8C5,RRID: AB_394643), anti-TGFβ1 (Santa Cruz Biotechnology, Dallas, Texas, USA, RRID: AB_632486), anti-p-Smad2 (Cell Signaling Technology; 138D4, RRID:AB_490941), anti-LEF1 (Cell Signaling Technology; C12A5, RRID: AB_823558), anti-β-catenin (BD; 14/Beta-catenin, RRID: AB_397554), anti-hCOL17 NC16A domain (TS39-3, homemade) (Ujiie et al, 2014), anti-extracellular portion of hCOL17 (mAb233, homemade) (Nishizawa et al, 1993), anti-murine COL17 NC14A domain (MoNC14A, homemade) (Nishie et al, 2015), anti-cytoplasmic portion of COL17 (Abcam, Cambridge, UK; ab186415, 1A8c, homemade (Nishizawa et al, 1993)), anti-plectin (Abcam; ab32528, RRID: AB_777339), anti-BP230 (Cosmo bio, Tokyo, Japan; 239, RRID:AB_1961833), anti-desmogleins 1 and 2 (PROGEN, Wieblingen, Heidelberg, Germany; DG3.10, RRID: AB_1284107), anti-integrin β1 (Chemicon International, Billerica, Massachusetts, USA; MB1.2, RRID: AB_2128202), anti-integrin α6 (BD Pharmingen; GoH3, RRID: AB_2296273), anti-ELANE (Abcam), anti-phospho aPKC (Santa Cruz; sc-271962, RRID:AB_10708397), and anti-survivin (Cell Signaling Technology; 71G4B7, RRID:AB_10691694), anti-laminin β1 (Abcam; LT3, RRID: AB_775971). …”

Membranous glomerulonephropathy in a patient with bullous pemphigoid

“…IgG1 is more prevalent in patients at the early stage of BP than IgG4, but IgG4 increases in patients with chronic BP. Ujiie et al reported that autoantibodies to BP induced blister formation in the absence of complement activation and that IgG4 was predominantly elevated and associated with the induction of blister formation in BP [17]. Our patient developed MN secondarily to blister formation during the chronic stage of BP.…”