Autoantibodies from Patients with Cicatricial Pemphigoid Target Different Sites in Epidermal Basement Membrane

Abstract

Indirect immunogold electron microscopy studies of cryofixed, freeze-substituted, and post-embedded normal human skin were performed to localize precisely the ultrastructural binding site of circulating autoantibodies from two groups of patients with cicatricial pemphigoid. One group of patients had circulating IgG autoantibodies that bound the dermal side of 1 M NaCl-split skin and immunoprecipitated epiligrin. The other group of patients had circulating IgG autoantibodies directed against the epidermal side of 1 M NaCl-split skin and showed no specific reactivity to any keratinocyte polypeptide by immunoprecipitation. IgG autoantibodies from all patients with anti-epiligrin cicatricial pemphigoid bound the lowermost aspect of the lamina lucida at its interface with the lamina densa; the greatest staining was seen beneath and beside hemidesmosomes. In contrast, IgG from cicatricial pemphigoid patients whose autoantibodies bound the epidermal side of 1 M NaCl-split skin localized to hemidesmosomes and the junction between hemidesmosomes and the plasma membranes of basal keratinocytes. Although the latter staining pattern is similar to that observed with anti-BPAG2 autoantibodies, sera from our patients with cicatricial pemphigoid did not bind BPAG2 in immunoprecipitation studies of radiolabeled human keratinocyte extracts or show immunoblot reactivity to a fusion protein corresponding to the immunodominant epitope of this polypeptide. These studies demonstrate the following: 1) Autoantibodies from patients with anti-epiligrin cicatricial pemphigoid consistently bind the lower lamina lucida at its interface with the lamina densa; and 2) other patients with the same phenotype may have IgG autoantibodies against yet-unknown epitopes in basal keratinocytes.

Context-Dependent Regulation of Collagen XVII Ectodomain Shedding in Skin

“…We and another group have reported that the C-terminus of COL17 binds with laminin 332 in vitro 23,29 . Because laminin 332 is always present on the dermal side of 1-M NaCl-split skin 3,30 , the present data indicate that another binding molecule, or molecules, on the epidermal side is present in the ECM, which binds with the shed COL17 ectodomain more strongly than laminin 332 does. A candidate molecule is integrin α6, which is known to be able to bind with the ectodomain of COL17 31,32 .…”

The Autoantibodies to α6β4 Integrin of Patients Affected by Ocular Cicatricial Pemphigoid Recognize Predominantly Epitopes Within the Large Cytoplasmic Domain of Human β4

“…The sera used by two investigators bound to the dermal side of salt split skin on indirect immunofluorescence assay and lamina densa on IEM contained Abs to laminin-5 (34). In the second group, investigators demonstrate that immunodeposits occur on hemidesmosomes and basal keratinocyte cytoplasm or the junction between hemidesmosomes and the inner plasma membrane of keratinocytes (33)(34)(35). These investigators observed that sera that produced such immune deposition did not contain Abs to laminin-5 or BP (33).…”

“…In the first group of studies, authors report deposition of immunoreactants on the lower lamina lucida and lamina densa (31)(32)(33). The sera used by two investigators bound to the dermal side of salt split skin on indirect immunofluorescence assay and lamina densa on IEM contained Abs to laminin-5 (34).…”

“…In the second group, investigators demonstrate that immunodeposits occur on hemidesmosomes and basal keratinocyte cytoplasm or the junction between hemidesmosomes and the inner plasma membrane of keratinocytes (33)(34)(35). These investigators observed that sera that produced such immune deposition did not contain Abs to laminin-5 or BP (33). In this study, using transfected cell lines, we demonstrated that patients with CP whose autoantibody binds to the epidermal side of salt split skin have Abs that bind to the cytoplasmic domain of ␤ 4 integrin.…”

New insights into the immunoultrastructural organization of cutaneous basement membrane zone molecules

“…Recently, mutations of LAMA3, LAMB3, or LAMC2 genes were detected in patients with Herlitz junctional EB in which the expression of laminin 5 is completely absent or markedly reduced (3,45). Furthermore, development of autoantibodies to laminin 5 leads to an acquired autoimmune blistering disease, such as one subtype of cicatricial pemphigoid (46,47). These anti-laminin 5 autoantibodies, directed to the a 3 chain of this molecule (48), were shown to directly produce dermoepidermal separation in animal models (49).…”