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K B Yancey

Publications and source records attributed to K B Yancey.

At least 19 recordsLinked to original sources
PubMed1999

Anti-epiligrin cicatricial pemphigoid with antibodies against the gamma2 subunit of laminin 5.

BACKGROUND: Cicatricial pemphigoid (CP) is a scarring subepithelial mucocutaneous blistering disease characterized by anti-basement membrane zone autoantibodies. Anti-epiligrin CP is an uncommon variant that has been recently characterized. Severe laryngeal involvement is infrequently observed in all forms of CP and has been documented in only 2 patients with anti-epiligrin CP. OBSERVATIONS: We report a case of CP exhibiting extensive laryngeal and ocular involvement. Histological, immunofluorescence, and immunoprecipitation studies confirmed the diagnosis of anti-epiligrin CP. Immunoblotting studies demonstrated the presence of antibodies against the alpha3 and the gamma2 subunit of laminin 5. CONCLUSION: This article expands the diversity of the clinical and immunopathologic features of this newly characterized variant of CP.

Aged↗
PubMed1999

Anti-epiligrin cicatricial pemphigoid with IgG autoantibodies to the beta and gamma subunits of laminin 5.

Anti-epiligrin cicatricial pemphigoid is an autoimmune subepithelial blistering disorder of mucous membranes and skin. By immunoblot analyses, sera of most patients with antiepiligrin cicatricial pemphigoid have been shown to react specifically with the alpha3 chain of laminin 5. We describe the first patient with anti-epiligrin cicatricial pemphigoid in whom circulating IgG autoantibodies directed against the beta3 and gamma2-chains of laminin 5 were detected. Treatment with oral prednisolone was beneficial in controlling the disease.

Aged↗
PubMed1998

Analysis of antigens targeted by circulating IgG and IgA autoantibodies in 50 patients with cicatricial pemphigoid.

In this study we investigated sera from 50 typical cicatricial pemphigoid (CP) patients. By indirect immunofluorescence on 1 M NaCl-split human skin sections, IgG of 17 sera and IgA of 22 sera reacted with the epidermal side of the split, while IgG of two sera reacted with the dermal side. These latter two sera were later confirmed to be anti-epiligrin CP. By immunoblotting of epidermal extracts, IgG of 14 sera reacted with the 230 kD bullous pemphigoid (BP) antigen (BP230). IgG of 15 sera and IgA of 11 sera reacted with the 180 kD BP antigen (BP180). Interestingly, a bacterial fusion protein containing the BP180 NC16a domain was recognized by IgG of 18 sera but not by IgA of any sera. Fusion proteins containing the C-terminal region of BP180 were recognized by IgG of 20 sera, but it was detected by IgA of only two sera. Our results suggest that, although CP sera show very low titers of autoantibodies, a considerable number of sera contain IgG antibodies to BP180 (either NC16a or C-terminal domain), confirming previous studies. In addition, we showed that greater numbers of IgA antibodies react with BP180, seemingly with different types of epitopes from those for IgG antibodies. Because the specificity of IgG antibodies is not very different from those in BP, IgA antibodies may play a specific role for the development of characteristic clinical features in CP. Future studies should elucidate the pathogenic role of the IgA antibodies in CP.

Animals↗
PubMed1998

Antiepiligrin cicatricial pemphigoid represents an autoimmune response to subunits present in laminin 5 (alpha3beta3gamma2).

Sera from 20 patients with antiepiligrin cicatricial pemphigoid were studied to define the specific reactivity of their IgG autoantibodies. IgG from all patients bound exclusively to the dermal side of 1 mol/L NaCl split skin and immunoprecipitated laminin 5 (alpha3beta3gamma2) from extracts of human keratinocytes (HKs). Immunoblot studies on purified laminin 5 subunits demonstrated that patient IgG bound alpha3 alone in 16 patients. In two patients, IgG autoantibodies were directed predominantly to the gamma2 subunit, yet showed trace reactivity to alpha3 as well. Sera from two patients did not immunoblot any laminin 5 subunits, their IgG presumably immunoprecipitating laminin 5 via a conformational epitope. Sera from patients with alpha3 subunit-specific IgG immunoprecipitated all subunits of laminin 5 as well as polypeptides of 190 and 200 kDa from the conditioned media of HKs. Preclearance studies and experiments utilizing affinity-purified patient IgG demonstrated that the latter signified laminin 6 (alpha3beta1gamma1) that was bound by cross-reactive alpha3 subunit-specific patient IgG. Sera from patients with gamma2 subunit-specific IgG showed no reactivity to laminin 6, except for faint reactivity provided by low levels of their alpha3 subunit-specific IgG. Taken together, these findings indicate that antiepiligrin cicatricial pemphigoid signifies an autoimmune response to subunits present in laminin 5.

Autoantibodies↗
PubMed1998

A deletion mutation in COL17A1 in five Austrian families with generalized atrophic benign epidermolysis bullosa represents propagation of an ancestral allele.

Patients with generalized atrophic benign epidermolysis bullosa, a usually nonlethal form of junctional epidermolysis bullosa, have generalized blistering, nail dystrophy, patchy alopecia, and dental abnormalities. Skin fragility in most cases is due to mutations in the gene encoding type XVII collagen (COL17A1). Recently, we reported five Austrian families with generalized atrophic benign epidermolysis bullosa who share the same COL17A1 mutation. Affected individuals in three families are homozygous for 4003delTC, whereas those in two others are compound heterozygotes. To determine if the occurrence of 4003delTC in these unrelated families signifies propagation of an ancestral allele or a mutational hot spot, haplotypes were determined for polymorphisms both within and flanking COL17A1. Five intragenic polymorphisms were chosen based on their informativeness. One of these, not previously reported, was 2988 A or C that introduces a new restriction site for Eco0109 I. All the 4003delTC alleles showed the same haplotype for these five polymorphic markers. Fourteen microsatellite polymorphisms were selected based on their high heterozygosity and their location within 10q23-q25 near COL17A1. Three families shared microsatellite polymorphisms covering at most 19 cM, whereas the others shared smaller regions consistent with cross-over events during passage of this mutation through several generations. These results indicate that 4003delTC occurs on a single ancestral allele.

Alleles↗
PubMed1998

Cycloheximide facilitates the identification of aberrant transcripts resulting from a novel splice-site mutation in COL17A1 in a patient with generalized atrophic benign epidermolysis bullosa.

Patients with generalized atrophic benign epidermolysis bullosa often show decreased expression of type XVII collagen, a transmembrane hemidesmosomal protein encoded by COL17A1. This report documents a novel splice-site mutation in COL17A1 in a patient with generalized atrophic benign epidermolysis bullosa, and applies a new methodology to define and characterize the resulting mRNA splice variants. Mutational analysis of COL17A1 identified a maternally inherited G-to-T transversion at the -1 position of exon 32. This acceptor splice-site mutation led to the formation of aberrant transcripts present at extremely low levels. Based on our recent finding that cycloheximide stabilized mutant COL17A1 transcripts in keratinocytes homozygous for a frameshift mutation, the effects of the splice-site mutation on splicing of COL17A1 transcripts were determined using reverse transcriptase polymerase chain reaction of total RNA from keratinocytes incubated for 2.5 h in the presence or absence of 10 microg cycloheximide per ml. Using this approach, an abnormally spliced transcript was identified that contains an extra 264 bases upstream from exon 32, resulting in a premature termination codon 27 bp downstream from the cryptic splice site. Three other splice variants, including one derived from the skipping of exon 32, were also identified. These results indicate the usefulness of cycloheximide treatment in evaluating the abnormal processing of mRNA due to splice-site mutations, because: (i) aberrant splicing often generates a premature termination codon, (ii) transcripts with premature termination codons can occur at low or undetectable levels due to nonsense-mediated mRNA decay, and (iii) the levels of these transcripts can be increased by cycloheximide.

Adult↗
PubMed1998

97 kDa linear IgA bullous dermatosis antigen localizes in the lamina lucida between the NC16A and carboxyl terminal domains of the 180 kDa bullous pemphigoid antigen.

Linear IgA bullous dermatosis is an autoimmune blistering disease characterized by circulating IgA anti-basement membrane autoantibodies. A 97 kDa protein (97-LAD), which localizes at the basement membrane zone of normal human skin, is one of the major autoantigens associated with this disease and possesses multiple regions of amino acid identity with the extracellular domain of the 180 kDa bullous pemphigoid antigen, BPAG2. To investigate further the relationship between 97-LAD and BPAG2, immunogold electron microscopy was performed on cryo-ultrathin sections of normal human skin using a series of polyclonal and monoclonal antibodies. Gold particles immunolabeling two newly developed monoclonal antibodies against 97-LAD were localized to the lamina lucida. This immunolabeling pattern was associated with hemidesmosomes and localized at a mean distance of 28 nm beneath the plasma membrane of basal keratinocytes. In contrast, polyclonal antibodies against a fusion protein containing the NC16A domain of BPAG2 immunolabeled the plasma membrane of the hemidesmosomal complex, whereas polyclonal antibodies against the carboxyl terminus mainly immunolabeled the lower lamina lucida with a mean distance of 42 nm beneath the plasma membrane. By double immunolabeling, 97-LAD was localized as if being sandwiched between the NC16A and the carboxyl terminal domains of BPAG2. These results clearly demonstrated the co-localization of 97-LAD and the extracellular portion of BPAG2 in the lamina lucida, and suggested that 97-LAD is closely related to, and/or forms a complex with, the extracellular domain of BPAG2.

Autoantibodies↗
PubMed1998

Anti-epiligrin cicatricial pemphigoid: a case associated with gastric carcinoma and features resembling epidermolysis bullosa acquisita.

A 48-year-old woman with anti-epiligrin cicatricial pemphigoid (CP) who showed clinical features resembling epidermolysis bullosa acquisita was found to have adenocarcinoma of the stomach. Histological examination of lesional skin demonstrated a subepidermal blister. Direct immunofluorescence microscopy of perilesional skin revealed linear deposits of IgG and C3 at the basement membrane zone. The patient's serum contained IgG autoantibodies that bound to the dermal side of 1 mol/L NaCl-split normal human skin as determined by indirect immunofluorescence microscopy, and the lamina lucida as determined by indirect immunoelectron microscopy. The patient's serum immunoprecipitated laminin-5 from extracts and media of biosynthetically radiolabelled human keratinocytes. Immunoblot studies showed that the patient's autoantibodies specifically bound the alpha3 subunit of this laminin isoform. Fragility of the skin and bullous lesions disappeared after total gastrectomy, but soon reappeared possibly in association with metastatic disease in a lymph node. The possibility that anti-epiligrin CP may develop paraneoplastically in some patients is discussed.

Anti-Inflammatory Agents↗
PubMed1997

Premature termination codons are present on both alleles of the bullous pemphigoid antigen 2/type XVII collagen gene in five Austrian families with generalized atrophic benign epidermolysis bullosa.

Patients with generalized atrophic benign epidermolysis bullosa (GABEB), an inherited subepidermal blistering disease, often have no immunologically detectable bullous pemphigoid antigen 2 (BPAG2) in their epidermal basement membrane. Recently, we analyzed the BPAG2 gene (GenBank no. M91669) in an Austrian family with GABEB and identified a homozygous deletion mutation, 4003delTC, that results in a downstream premature termination codon (PTC). This mutation has now been identified in additional descendants, suggesting transmission of this mutant allele through at least six generations. Screening of four other Austrian GABEB families revealed that affected members were homozygous for 4003delTC in two cases and heterozygous in two others. In the latter, mutational analysis identified two novel nonsense mutations, Q1403X and G803X, that were confirmed by restriction endonuclease digestions. Thus, PTCs on both alleles of BPAG2 are present in all of these GABEB families. Immunoprecipitation and northern blot studies of cultured keratinocytes from homozygous GABEB patients show that 4003delTC results in undetectable levels of BPAG2 protein and mRNA-findings consistent with the process of nonsense-mediated mRNA decay. Incubating keratinocytes with cycloheximide increased BPAG2 mRNA to a level detectable by northern analysis. When the latter was used in reverse transcription-PCR studies, the mutation was demonstrated, suggesting that cycloheximide may allow mutational analysis in cases where low transcript levels have previously thwarted RT-PCR studies. These findings account for the absence of BPAG2 in GABEB patients and attest to the importance of this protein in adhesion of epidermis to epidermal basement membrane.

Alleles↗
PubMed1997

The extracellular domain of BPAG2 localizes to anchoring filaments and its carboxyl terminus extends to the lamina densa of normal human epidermal basement membrane.

Bullous pemphigoid antigen 2 (BPAG2) is a 180 kDa type II transmembrane protein associated with hemidesmosomes (HDs) in basal keratinocytes. To better understand how BPAG2 promotes keratinocyte adhesion to epidermal basement membrane (BM), purified IgG against a baculovirus-encoded recombinant was used to localize its carboxyl terminus in human skin by immunogold electron microscopy (IEM). A 2.1-kb BPAG2 cDNA encoding the distal extracellular domain and carboxyl terminus of BPAG2 was used in a baculovirus expression system to create virus that produced a 70-kDa recombinant form of BPAG2 (BV4). BV4 was purified, characterized, and used to raise high-titer specific rabbit IgG. Purified anti-BV4 IgG bound the epidermal side of 1 M NaCl split skin and bound only BPAG2 on immunoblots containing extracts of human keratinocytes. In IEM studies of pre- and post-embedded skin, the distal ectodomain of BPAG2 localized beneath HDs in basal keratinocytes; there was no evidence of BPAG2 beneath melanocytes. Anti-BV4 IgG extensively bound anchoring filaments on the epidermal side of 1 M NaCl split skin; this staining extended along anchoring filaments to their ends. In post-embedded skin, the carboxyl terminus of BPAG2 was localized within the lamina densa, 41 nm (mean of 400 determinations) beneath plasma membranes of basal keratinocytes. BPAG2 thus extends from the intracellular HD plaque of basal keratinocytes to the lamina densa of human epidermal BM.

Animals↗
PubMed1997

Noncomplement fixing, IgG4 autoantibodies predominate in patients with anti-epiligrin cicatricial pemphigoid.

This study characterized the specific reactivity, IgG subclass, and complement fixing ability of anti-laminin-5 IgG from 12 patients with anti-epiligrin cicatricial pemphigoid. Circulating IgG from all patients bound the dermal side of 1 M NaCl split skin, immunoprecipitated laminin-5 produced by biosynthetically radiolabeled human keratinocytes, and (in 10 of 12 cases) immunoblotted the laminin-alpha3 subunit. Analysis of the distribution of IgG subclasses in these patients' circulating anti-laminin-5 autoantibodies by semiquantitative indirect immunofluorescence microscopy using the HP series of subclass-specific monoclonal antibodies revealed: (i) IgG4 predominant autoantibodies in seven of 11 sera; (ii) IgG1 and IgG2 at substantially lower levels in a smaller number of sera; and (iii) no specific IgG3 anti-laminin-5 autoantibodies in any patients. The same IgG4-dominant profile of anti-laminin-5 autoantibodies was found in enzyme-linked immunosorbent assay studies of purified human laminin 5. Direct immunofluorescence microscopy of six skin biopsies from three patients found that IgG4 was also the predominant subclass of IgG in epidermal basement membranes in situ. Consistent with these findings, sera from 11 of 11 patients with anti-laminin-5 IgG autoantibodies did not fix C3 to epidermal basement membranes in vitro. These immunochemical studies suggest that complement activation does not play a major role in the pathophysiology of this disease and that subepidermal blisters in these patients may develop via a direct effect of anti-laminin-5 IgG itself.

Autoantibodies↗
PubMed1997

Generalized atrophic benign epidermolysis bullosa.

Junctional EB was once considered a uniformly fetal autosomal recessive skin disease. One of the first and best characterized forms of generalized JEB that has a more favorable prognosis was GABEB. Since its initial description in 1976, many additional cases have been compiled, all sharing the features of chronic blistering from birth, nail dystrophy, hair loss, and abnormal teeth. In addition, some patients have large melanocytic nevi that form at sites of prior blisters. The first clues as to the protein defective in these patients was provided by immunofluorescence microscopy studies, which showed absent or decreased expression of type XVII collagen in these patient's epidermal BM. This protein was a promising candidate because it is located in the ultrastructural plane where blisters from in patients with GABEB. It is also the same hemidesmosomal protein against which antibodies are directed in patients with autoimmune blistering diseases: bullous, gestational, and cicatricial pemphigoid. The demonstration of decreased expression of type XVII collagen in patients with GABEB pointed the way to a crucial distinction that could not be based on clinical and histologic findings--that of discriminating infants with GABEB from those with the lethal Herlitz variant of JEB. These two diseases may be differentiated because decreased expression of type XVII collagen is found exclusively in GABEB, whereas decreased expression of laminin 5 is most likely associated with Herlitz JEB. The molecular basis of GABEB has been determined. The immunofluorescence microscopy studies mentioned above directed mutation studies to the COL17A1 gene. Several mutations in COL17A1 have been described in patients with GABEB, almost all of which result in a PTC. As a consequence of the PTC, mutant COL17A1 transcripts are rapidly degraded by nonsense-mediated mRNA decay, blocking production of type XVII collagen from the mutant allele. Heterozygous carriers of the mutation are thus rendered untouched by the mutation. In the homozygous state, however, this results in the absence of this vital adhesion protein, leading to chronic blistering and skin fragility observed in these patients. These studies also significantly increase our knowledge about the normal functions of type XVII collagen. The fact that patients deficient in type XVII collagen have fragile skin attests to the role of this protein in the adhesion of basal keratinocytes to epidermal BM. The severity of GABEB, however, is much less than that observed in patients with a complete deficiency of laminin 5, suggesting that type XVII collagen functions cooperatively with other hemidesmosomal proteins in promoting adhesion. Studies using GABEB keratinocytes will help characterize these kinds of interactions. The ultimate prospect of mutation analysis in these patients is the hope of correcting their genetic defect and curing their disease. Until then, the physician plays an important role in making the diagnosis, providing general supportive skin care, advising avoidance of trauma, treating infections, and supporting the patient's psychosocial development. Further studies of patients with GABEB will lead to new therapeutic approaches, as well as reveal additional complex functions of type XVII collagen not only in skin, but also in hair, teeth, and nails.

Collagen↗
PubMed1996

Passive transfer of anti-laminin 5 antibodies induces subepidermal blisters in neonatal mice.

Patients with a recently identified subepithelial blistering disease have IgG anti-laminin 5 autoantibodies. To determine if such antibodies can be pathogenic in vivo, we developed and characterized rabbit anti-laminin 5 IgG, and passively transferred these antibodies to neonatal mice. Immune rabbit IgG specifically bound human and murine epidermal basement membranes, immunoblotted and immunoprecipitated all laminin 5 subunits from extracts of human and murine keratinocytes, and showed no reactivity to other keratinocyte proteins or epithelial basement membranes that do not contain laminin 5. Mice (n = 29) receiving purified anti-laminin 5 IgG developed, in a dose-related fashion, circulating anti-laminin 5 antibodies, deposits of rabbit IgG and murine C3 in epidermal basement membranes, and subepidermal blisters of skin and mucous membranes. No alterations developed in controls (n = 14) receiving identical amounts of normal rabbit IgG. Passive transfer of anti-laminin 5 (but not control) IgG to neonatal C5- (n = 3) or mast cell-deficient (n = 3) mice produced subepidermal blisters with the same clinical, histologic, and immunopathologic features as those documented in BALB/c mice. These studies establish an animal model of a human blistering disease that can be used to define disease mechanisms and treatment modalities.

Animals↗
PubMed1996

Antiepiligrin cicatricial pemphigoid: the first case report from Japan.

We describe a Japanese man with antiepiligrin cicatricial pemphigoid and typical clinical features, including ocular involvement. Direct immunofluorescence showed IgG deposition at the basement membrane zone. Indirect immunofluorescence of 1M sodium chloride-split skin showed circulating antibasement membrane zone antibodies of IgG class reactive with the dermal side of the split. Immunoblotting of human epidermal and dermal extracts, as well as a bacterial fusion protein of BP180 NC16a domain, showed no specific reactivity. In contrast, with immunoprecipitation of either culture medium or cell lysate from normal keratinocytes, the patient's serum clearly reacted with the protein epiligrin, a laminin isoform present in the lamina lucida of the human epidermal basement membrane zone. This is the first confirmed case of a Japanese patient with this disease entity.

Basement Membrane↗
PubMed1996

Reactivity of autoantibodies from patients with defined subepidermal bullous diseases against 1 mol/L salt-split skin. Specificity, sensitivity, and practical considerations.

BACKGROUND: Circulating IgG anti-basement membrane autoantibodies from patients with subepidermal bullous diseases can be categorized on the basis of their pattern of reactivity against 1 mol/L sodium chloride (NaCl)-split skin. OBJECTIVE: The purpose of this study was to define by immunochemical techniques the specific antigen(s) targeted by IgG autoantibodies from a group of patients with subepidermal blistering diseases and then (1) prospectively determine which side(s) of 1 mol/L NaCl-split skin is (are) bound by these patients' autoantibodies, (2) compare the sensitivity of intact and 1 mol/L NaCl-split skin for the detection of these autoantibodies; and (3) devise a practical method to distinguish patients with antiepiligrin cicatricial pemphigoid from those with other subepidermal blistering diseases. METHODS: Investigative techniques included direct and indirect immunofluorescence microscopy, immunoprecipitation studies, and immunoblotting. RESULTS: These studies identified 14 patients whose sera immunoprecipitate bullous pemphigoid antigens 1, 2, or both. These patients' circulating IgG anti-basement membrane autoantibodies bind the epidermal (n = 11), epidermal and dermal (n = 2), or dermal (n = 1) sides of 1 mol/L NaCl-split skin by indirect immunofluorescence microscopy. In contrast, IgG from all patients with autoantibodies directed against type VII collagen (n = 5) or epiligrin (n = 6) bind only the dermal side of 1 mol/L NaCl-split skin. In all but one patient in this series, 1 mol/L NaCl-split skin proved to be a more sensitive test substrate than intact human skin for detection of circulating IgG anti-basement membrane autoantibodies. Patients with antiepiligrin cicatricial pemphigoid were distinguished from other patients in that their circulating autoantibodies bound epidermal basement membrane in the skin of primates but not small mammals. CONCLUSION: NaCl-split skin (1 mol/L) of various species is a sensitive and practical indirect immunofluorescence microscopy test substrate for the evaluation of patients with IgG anti-basement membrane autoantibodies and evaluation of subepidermal bullous diseases.

Antigens↗