Pure ocular cicatricial pemphigoid: A distinct immunopathologic subset of cicatricial pemphigoid

ObjectiveTo determine whether ocular cicatricial pemphigoid (OCP) may represent a distinct immunopathologic disease when it is pure ocular cicatricial pemphigoid (POCP) (e.g., only confined to the conjunctiva) or when it is associated with skin or extraocular mucous membrane lesions or both (OCP⁺).DesignProspective, immunologic, and immunopathologic study with special emphasis on direct immunoelectron microscopy.ParticipantsSix patients with POCP and seven patients with OCP⁺.InterventionAfter informed consent was obtained, a conjunctival biopsy was performed in all patients. Skin and extraocular mucosa biopsy specimens were harvested in selected cases only.Main outcome measuresResults of direct immunofluorescence and direct immunoelectron microscopy without freezing on conjunctival and skin biopsy specimens, indirect immunofluorescence, and Western immunoblotting analysis were analyzed.ResultsResults of direct immunoelectron microscopic examination of the conjunctiva showed the presence of immune deposits in the upper lamina lucida of the basement membrane zone in the six patients with POCP, whereas the immune reactants were located in the lower part of the lamina lucida and in the lamina densa of the basement membrane zone (conjunctiva, buccal mucosa, and skin) in the seven patients with OCP⁺. Direct immunofluorescence was positive in the biopsy specimens of three patients with POCP (50%) and the seven patients with OCP⁺ (100%). Results of indirect immunofluorescence study showed circulating autoantibody levels only in two patients with OCP⁺, and results of Western immunoblot analysis were negative.ConclusionsResults of direct immunoelectron microscopic examination of the conjunctiva support the hypothesis that POCP may be a disease entity distinct from mucocutaneous cicatricial pemphigoid.     

Immunoelectron microscopic study of the conjunctiva in cicatricial pemphigoid

· Background: Immune deposits can be found on the conjunctival basement membrane zone of patients affected by cicatricial pemphigoid using immunofluorescence technique. The purpose of this study was to perform direct immunoelectron microscopy on the conjunctiva of patients with scarring conjunctivitis associated with cicatricial pemphigoid · Methods: Conjunctival and skin biopsies were performed in six patients who presented with presumed autoimmune cicatrizing conjunctivitis associated with cicatricial pemphigoid. Specimens were processed for direct immunofluorescence and direct immunoelectron microscopy. · Results: Direct immunofluorescence was positive in all skin samples and in three of six conjunctival samples. Direct immunoelectron microscopy showed immune deposits in the lamina lucida and the lamina densa of all skin and conjunctival samples. · Conclusions: Direct immunoelectron microscopy can be performed on the conjunctiva. It shows the precise localization of cicatricial pemphigoid target antigens within the conjunctival basement membrane zone. Received: 29 April 1997 Revised version received: 16 February 1998 Accepted: 9 March 1998     

Two cases of cicatricial pemphigoid showing atypical immunofluorescent findings.

We describe two patients with the clinical symptoms of cicatricial pemphigoid (CP). Biopsy specimens of the conjunctiva were taken. Histologic examination revealed subepidermal bullae and infiltration of inflammatory mononuclear cells. Direct immunofluorescent study showed immunoglobulins bound to the basement membrane zone (BMZ) in these patients. The patients also had intercellular immunoglobulin deposition in the conjunctival epithelium. No circulating anti-BMZ antibodies were detected, but one patient had a circulating antiintercellular antibody. Rare cases of CP with atypical immunofluorescent findings are reported.     

Linear IgA bullous disease limited to the eye: a diagnostic dilemma: response to intravenous immunoglobulin therapy

PURPOSE: To report on a diagnostic dilemma and treatment challenge in a patient with chronic cicatrizing conjunctivitis without involvement of skin and other mucous membranes persisting for 6 years and not responding to topical and systemic steroids. DESIGN: Interventional case report. METHODS: We performed direct immunofluorescence of the conjunctiva with fluorescein-conjugated rabbit antihuman antibodies against immunoglobulin A, G, and M, complement 3 component, and fibrinogen. To investigate the presence of circulating antibodies in patient's serum, indirect immunofluorescence using normal human conjunctiva, normal human skin, and monkey esophagus as substrate was done. In addition, we did immunoblot analysis using normal human epidermis as substrate to determine the molecular weight of an antigen. The patient was treated with intravenous immunoglobulin (IVIg). The correlation between the titer of circulating antibodies and the activity of conjunctival inflammation at various intervals during the course of IVIg therapy was demonstrated by immunoblot assay with serial dilutions of the patient's serum. The highest dilution at which the binding was visible was considered the titer. RESULTS: Direct immunofluorescence of the conjunctiva and indirect immunofluorescence with both salt split skin and conjunctiva as substrate disclosed linear deposition of immunoglobulin A (IgA) at the epithelial basement membrane. Immunoblot analysis demonstrated the presence of IgA circulating antibodies in patient's serum directed against a 97kDa protein in human epidermis. A continuous decrease in the titer of these antibodies correlating to improvement of clinical symptoms was observed during IVIg therapy. CONCLUSIONS: Use of a nonconventional diagnostic tool (immunoblot analysis), in addition to conventional immunohistologic studies, might be helpful in establishing the diagnosis of patients with chronic cicatrizing conjunctivitis. On the basis of results of these laboratory tests and clinical presentation, we believe that this patient has linear IgA bullous disease limited to the eye. IVIg therapy decreased the titer of circulating antibodies and induced a remission in this patient.     

Conjunctival involvement in cicatricial and bullous pemphigoid: a clinical and immunopathological study.

Patients with bullous pemphigoid were found to have significant ocular abnormalities. In a group of 18 patients one had conjunctival shrinkage, and 11 of 15 (73%) had positive linear direct immunofluorescence on conjunctival biopsy from a clinically uninvolved site. Our ocular findings in a group of 14 with cicatricial pemphigoid are also reported and compared with those from a control group of 20. Our findings suggest there is overlap between the pemphigoid groups and raise further questions about the pathogenicity of immunoreactants within the basement membrane zone. Bulbar conjunctival biopsy was simple and well tolerated, and the rate of immunofluorescence positivity of conjunctiva was twice that of skin in both pemphigoid groups.     

Antibasement membrane antibody-mediated experimental conjunctivitis

In ocular cicatricial pemphigoid, the binding of circulating antibodies to conjunctiva is believed to initiate an antibody-mediated cytotoxic response that results in inflammation and tissue damage. To develop a model of antibody-mediated conjunctival inflammation, we examined the effect on conjunctiva of local or systemic administration of a murine monoclonal antibody against basement membrane of stratified squamous epithelium. Neonatal rabbits were given either a single subconjunctival or intraperitoneal injection of the antibody. Eyes were graded clinically for inflammation and conjunctival biopsies were performed. After subconjunctival injection, clinical and histologic inflammation as well as murine antibody and rabbit complement binding to conjunctival basement membrane were detected. With systemic administration there was post-injection clinical inflammation, and conjunctival basement membrane-bound murine antibody was detected. There was no difference observed in conjunctival mitotic rate or goblet cell frequency between treatment groups and controls, following either route of administration. We have created, therefore, a model for antibody-mediated conjunctivitis in rabbits by local or systemic administration of a monoclonal antibody against a component of stratified squamous epithelial basement membrane.     

Ocular involvement in anti-epiligrin cicatricial pemphigoid

PURPOSE: To report an anti-epiligrin cicatricial pemphigoid (AECP) patient with severe ocular involvement and to provide a practical approach to distinguishing AECP patients from those with other subepidermal blistering diseases. METHODS: Techniques included direct and indirect immunofluorescence microscopy, Western blot and immunoprecipitation studies, as well as interdisciplinary examinations of mucous membranes and skin. RESULTS: This study describes a patient with clinical features of cicatricial pemphigoid, circulating anti-basement membrane zone IgG antibodies, and subepidermal blisters. Histopathology and immunofluorescence analysis suggested the diagnosis of a cicatricial pemphigoid-like type of epidermolysis bullosa acquisita. However, Western blot and immunoprecipitation studies demonstrated that the patient's serum contained autoantibodies against laminin 5 alpha3 subunit, leading to the diagnosis of an AECP. CONCLUSION: Since patients with AECP have an increased relative risk for malignant tumors, it is important to distinguish this entity within the spectrum of cicatricial pemphigoid patients by additional studies such as Western blot or immunoprecipitation.     

The role of antibody to human beta4 integrin in conjunctival basement membrane separation: possible in vitro model for ocular cicatricial pemphigoid.

PURPOSE: To demonstrate the specific binding of autoantibodies present in the sera of patients with ocular cicatricial pemphigoid (OCP) to human beta4 integrin present in the normal human conjunctiva (NHC) and to study the role of OCP autoantibodies and antibody to human beta4 integrin in the pathogenesis of subepithelial lesion formation in OCP. METHODS: Indirect immunofluorescence assay and in vitro organ culture method using NHC were used. Sera and IgG fractions from 10 patients with OCP; immunoaffinity-purified OCP autoantibody; antibodies to human beta4, beta1, alpha6, and alpha5 integrins; and sera from patients with pemphigus vulgaris, bullous pemphigoid (BP), and chronic atopic and chronic ocular rosacea cicatrizing conjunctivitis; and normal human serum (NHS) were used. RESULTS: Nine of 10 OCP sera or IgG fractions, immunoaffinity-purified OCP autoantibody, antibodies to human beta4 and alpha6 integrins, and sera from patients with BP showed homogenous, smooth linear binding along the basement membrane zone (BMZ) of the NHC. NHS, antibodies to other integrins, and sera from patients with chronic cicatrizing conjunctivitis from other causes showed no such binding. When NHC was first absorbed with OCP sera and then reacted with anti-beta4 antibodies or vice versa, the intensity of the BMZ binding was dramatically reduced or completely eliminated, indicating that there were autoantibodies in OCP sera specific for the beta4 integrin. BMZ separation developed 48 to 72 hours after addition of total OCP sera, IgG fractions from OCP sera, immunoaffinity-purified autoantibodies from sera of patients with OCP, or anti-beta4 antibodies to the NHC cultures, but not after addition of normal control sera, sera from patients with chronic cicatrizing conjunctivitis from causes other than OCP, or sera from patients with OCP in clinical remission. CONCLUSION: Circulating anti-beta4 integrin antibody may have an important role in the pathogenesis of OCP.     

Ocular Cicatricial Pemphigoid

The characteristic feature of ocular cicatricial pemphigoid (OCP) is progressive shrinkage of the conjunctiva. In our series of 78 patients with OCP, 21% had cutaneous involvement and 50% had involvement of the oral mucosa. Immunoglobulins and the third component of complement are found bound to the conjunctival epithelium and basement membrane of patients with OCP. Circulating antibodies which bind to the conjunctival and corneal epithelium but not to the conjunctival basement membrane have also been demonstrated. OCP is associated with an increased prevalence of HLA-B12. The lids and conjunctiva of patients with OCP demonstrate an increased incidence of potential pathogens when compared with age- and sex-matched controls. When followed for a period averaging 22 months, the majority of patients not treated with systemic immunosuppressives or topical corticosteroids progress. However, OCP has a variable course because there were patients in all stages who did not progress. The acute manifestations of OCP may cause rapid shrinkage of the conjunctiva and may be suppressed with systemic corticosteroids.     

The value of biopsies in the evaluation of chronic progressive conjunctival cicatrisation

Background: Chronic progressive conjunctival cicatrisation is poorly understood, and therapy of this condition remains difficult. This study assessed the value of immunohistochemical investigations in the evaluation of patients who present with chronic cicatrising conjunctivitis similar to cicatricial pemphigoid (CP) Methods: Bulbar conjunctival biopsies from 36 patients with acute (n = 5), subacute (n =13) and chronic (n=18) ocular disease were studied. The biopsy was retaken in 7 patients to evaluate the present immunological findings in comparison with a biopsy more then 5 years ago. All the specimens were investigated for the presence of immunoglobulins and complement at the epithelial basement membrane, and the phenotype of the inflammatory cellular infiltrate was analysed. Twenty-nine patients were evaluated for the presence of circulating IgG-anti-basement membrane zone antibodies Results: CP was confirmed by immunoglobulins and/or complement deposition at the epithelial basement membrane in 11 patients (31%). IgA was found to be the most frequent deposit. Eleven CP patients, mainly those with active or burnt-out disease, showed absence of immunoglobulins and/or complement at the conjunctival basement membrane. In 14 of 36 patients, conjunctival cicatrisation was subsequently felt to be caused by conditions other than CP. The cellular phenotype in the subepithelial conjunctiva was unspecific, but in CP the disease activity was reflected by the number of neutrophils and macrophages. Circulating IgG antibodies were found in none of the patients' serum Conclusion: Immunoglobulin and/or complement deposition at the epithelial basement membrane confirms the diagnosis of mucous membrane pemphigoid. Their absence, however, does not rule it out and is a frequent feature in very active conjunctival disease or after immunosuppressive treatment. The analysis of the cellular phenotype in mucous membrane pemphigoid may be useful in the assessment of disease activity but does not help in determining the underlying disease process causing the cicatrising conjunctivitis     

Linear IgA disease. The ocular manifestations

The ocular history and examination of a 54-year-old Filipino woman with linear IgA disease is described. Results of the eye examination were consistent with chronic cicatricial conjunctivitis, showing subconjunctival fibrosis and symblepharon formation. Direct immunofluorescence of the conjunctiva was positive for IgA and C3 in a linear pattern along the epithelial basement membrane. The ophthalmologic and dermatologic findings in linear IgA disease are compared with those of dermatitis herpetiformis, bullous pemphigoid, and cicatricial pemphigoid. This is the first documented case report of the ocular manifestations of linear IgA disease in the American literature.     

Immunophenotypic analysis of the inflammatory infiltrate in ocular cicatricial pemphigoid. Further evidence for a T cell-mediated disease

Ocular cicatricial pemphigoid (OCP) is characterized by the deposition of immunoglobulin and complement along the conjunctival epithelial basement membrane zone (BMZ). In order to further elucidate the cellular populations of the local inflammatory infiltrates, the authors used a panel of monoclonal antibodies in cryostat tissue sections to delineate T cell subsets, B lymphocytes, dendritic cells, and macrophages in six patients with OCP. In comparison with matched controls of the epibulbar conjunctiva, the authors discovered a threefold increase in T lymphocytes within the epithelium and a 20-fold increase within the substantia propria. In contrast with the normal-standing population of conjunctival T lymphocytes, there were activated interleukin 2 receptor (IL-2R)-positive lymphocytes in both the epithelium and the substantia propria. Macrophages were the second most common cells in the substantia propria, accounting for 12.7% of the mononuclear population--a threefold increase over the normal percentage. B cells and plasma cells, normally absent from epibulbar conjunctiva, were the next most prominent populations, constituting 6.9 and 4.6%, respectively, of all mononuclear cells. Dendritic cells which process antigen locally constituted only 1.2% of the mononuclear cell population, but were increased 25-fold over normal controls. By elaborating cytokines that promote fibroplasia, the T cells in OCP may be effector cells along with macrophages and other inflammatory cells in bringing about scarification of the substantia propria, and may furthermore be responsible for an immunoregulatory defect that allows local B lymphocytes to produce autoantibodies to the BMZ.     

Diagnostics and pharmacological treatment of ocular cicatrical pemphigoid

Ocular cicatricial pemphigoid (OCP) is an autoimmune disease characterize by mucous membrane fibrosis and skin changes resulting with scarring. The pathogenic mechanisms of ocular cicatricial pemphigoid are incompletely understood. Antibasement membrane antibodies which lead to subepithelial blistering, granulation tissue and inflammatory infiltrate formation in the substantia propria are thought to be the main pathophysiological mechanisms in cicatricial pemphigoid. It has been found eosinophils and increased collagen type I and III. Human leukocyte antigen HLA-DR2, HLA-DR4 and DQw7 genotypes have been identified as conferring increased susceptibility to the development of this disease. Ocular cicatrical pemphigoid (OCP) is one of the forms of bullous pemphigoid. Initial symptoms of ocular pemfigoid are not characteristic. Conjunctival fibrosis may cause severe entropion, trichiasis, symblepharon, dry eye syndrome, corneal epithelial erosions or ulceration. Secondary glaucoma is one of the most frequent complications. Ocular cicatricial pemphigoid may be chronic, acute, or subacute disease with periodic exacerbation of conjunctival inflammation. The treatment in this disease are topical drops or ointment (lubricants, corticosteroids, antibiotics, antiglaucomatous). Oral dapsone and corticosteroids may control the activity of the disease. In other progressive cases immunosuppressive drugs must be used (azathioprine, cyclophosphamide, methotrexate, mycophenolan mofetil, daclizumab, intravenous immunoglobulin therapy). To make an early diagnosis of ocular cicatricial pemphigoid, biopsy and immunohistochemical analysis of conjunctiva should be performed in every case of persistent conjunctival inflammation.     

Immunopathology of cicatricial pemphigoid affecting the conjunctiva

Conjunctival biopsy specimens from 13 patients with cicatricial pemphigoid and from 13 age-matched healthy individuals undergoing cataract surgery were analyzed by light microscopy and immunohistochemical techniques, including a panel of monoclonal antibodies used to characterize inflammatory mononuclear cell phenotypes. Results of histologic examination of cicatricial pemphigoid specimens showed typical squamous metaplasia, vasculopathy, increased numbers of mast cells, and abundant plasma cells. All cicatricial pemphigoid specimens demonstrated immunoreactants at the epithelial basement membrane zone (BMZ). Epithelium of cicatricial pemphigoid conjunctiva showed significantly more T-helper cells (CD4+), dendritic cells (CD1+), and macrophages (CD14+), and a significantly higher helper/suppressor ratio than did controls. In the substantia propria, pemphigoid specimens showed dramatically increased inflammatory infiltrate with significantly more cells staining, in order of frequency, for T cells (CD3+, CD5+), T-helper cells (CD4+), T-suppressor cells (CD8+), macrophages (CD14+, Mac-1+), and dendritic cells (CD1+, HLA-DR+). Ten percent of these cells expressed interleukin-2 receptor protein (CD25+), indicating T-cell activation.     

Mucous membrane pemphigoid: an update

PURPOSE OF REVIEW: To review articles on mucous membrane pemphigoid, published between June 2004-May 2005. RECENT FINDINGS: Decreased glycosylation of mucin was found in patients with ocular cicatricial pemphigoid. A unique antigen in oral mucous membrane pemphigoid has not yet been identified. Increased vascular cell adhesion molecule and intercellular adhesion molecule 1 expression was found in skin of patients affected by mucous membrane pemphigoid. Autoreactive T cells to an epitope of bullous pemphigoid antigen 180 kilodaltons were identified in the blood of some patients with mucous membrane pemphigoid. Circulating IgA against an antigen in mucous membrane pemphigoid was found in about 20% of patients, without prognostic significance. Enhanced sensitivity for direct immunofluorescence was reported if skin biopsy specimens were stored for 24 hours in saline. An enzyme-linked immunosorbent assay for detection of circulating autoantibodies against laminin-5 was developed. Sensitivity was higher than indirect immunofluorescence on salt-split skin and immunoblotting. Patients with younger onset (<60 years) of ocular cicatricial pemphigoid were found to have disease evolution similar to that of an older group (>70 years) but were visually impaired earlier in life. Intravenous immunoglobulin as treatment of ocular cicatricial pemphigoid was found to be superior to conventional immunosuppressants, with fewer side effects and better long-term outcome for halting disease activity. Intraoperative adjunction of mitomycin C during fornix reconstruction with amniotic membrane resulted in achieving a deeper fornix in 83% of patients with various cicatrizing conjunctivitis. Transplantation of cultured epithelial cells of oral mucosa in corneal limbal stem cell deficiency was successful in improving visual acuity and reestablishing corneal transparency in mid- to advanced ocular cicatricial pemphigoid. SUMMARY: Further advances have been achieved in the field of mucous membrane pemphigoid.     

Autoimmune phenomena of the external eye.

The immunologic status of patients with ocular pemphigoid, Mooren's ulcer, chronic herpetic keratitis, and staphylococcal peripheral corneal ulcers was studied. Although tissue-fixed and circulating antibodies to the conjunctival epithelium were found in all groups, patients with Mooren's ulcer demonstrated these findings most consistently. Immunoglobulins bound to the conjunctival basement membrane were found not only in ocular pemphigoid but also in patients with Mooren's and staphylococcal ulcers. Approximately one half of the patients with ocular pemphigoid and Mooren's ulcer demonstrated elevations in serum IgA levels. An increased prevalence of HLA-B12 was found in ocular pemphigoid.     

Mucous membrane pemphigoid with fatal bronchial involvement in a seventeen-year-old girl

PURPOSE: This study was designed to report a case of biopsy-proven mucous membrane pemphigoid with severe bronchial involvement in a young woman. METHODS: Case report of a 17-year-old girl who presented with worsening dyspnea, skin rash, and bilateral ocular injection, symblepharon, and fornix foreshortening. Conjunctival, skin, and bronchial biopsies were performed along with imaging and serological tests in an effort to establish a diagnosis for this unusual constellation of findings. The surprising occurrence of a cerebrovascular accident during her hospitalization also prompted a search for a concurrent coagulation disorder. RESULTS: Immunofluorescence studies of conjunctival, skin, and bronchial tissue specimens revealed deposition of multiple antibody classes at the basement membrane zone. The patient also possessed circulating basement membrane zone antibodies in her serum and a significant titer of antiphospholipid antibodies. She underwent dilation and stent placement for subglottic tracheal and left bronchial stenosis and was treated with immunosuppressive agents. After a favorable initial response, the patient experienced progressive bronchial stenosis and respiratory compromise, culminating in her death from bronchospasm and cardiopulmonary arrest. CONCLUSION: To our knowledge, this is the first report of mucous membrane pemphigoid involving the lower airways that was confirmed by immunofluorescence analysis. It highlights the potentially lethal, systemic nature of mucous membrane pemphigoid and underscores the need to question patients about symptoms of respiratory dysfunction.     

Identification of ocular cicatricial pemphigoid antibody binding site(s) in human beta4 integrin

PURPOSE: To identify specific site(s) on human ss4 molecule to which sera from ocular cicatricial pemphigoid (OCP) patients bind and to determine its role in the process of blister formation. METHODS: Clone the fragments representing the extracellular and intracellular domain of ss4 molecule from normal human conjunctival mRNA into an expression vector; map the region to which sera from OCP patients bind by Western blot analysis. Determine the role of the immunodominant region in pathogenesis by demonstrating the ability of the rabbit antibody to the immunodominant region to produce separation of basement membrane zone (BMZ) from the basal epithelial layer when incubated with normal human conjunctiva in an in vitro organ culture model. RESULTS: Majority of the OCP sera tested bound to the C-terminal end of the intracellular domain (IC3.0) of the human ss4 integrin. Further subcloning of IC3.0 demonstrated that a smaller fragment extending from 1489 aa to 1572 aa (IC3.4) was responsible for this binding. This region may have multiple antibody binding sites. Antibody to human IC3.0 and IC3.4 produced in rabbit, resulted in BMZ separation, histologically identical with that observed when normal human conjunctiva was cultured with OCP sera in an human conjunctival organ culture model. CONCLUSIONS: These observations identify IC3.4 as the antibody binding site for sera of OCP patients and suggest a possible role for it in blister formation. Indirectly it highlights certain important aspects of the structural and functional dynamics of the biology of the hemidesmosomes and basement membranes.     

Lichen planus and cicatrizing conjunctivitis: characterization of five cases

PURPOSE: To report the clinical and immunopathologic features and the response to therapy in a series of six patients with cicatrizing conjunctivitis due to lichen planus. DESIGN: Retrospective case series. METHODS: All six patients were seen in an ocular pemphigoid clinic. Clinical, immunopathologic, and serologic features were evaluated and therapeutic response in each patient was monitored. RESULTS: All six patients had evidence of conjunctival scarring. Five patients had lichen planus of the oral mucosa and gingiva; one patient had involvement of the skin. Histologic findings consisted of thickened epithelium and an interface lymphocytic infiltrate along the lamina propria. In three patients, electron microscopy of the conjunctiva revealed thickening, fragmentation, and duplication of the basement membrane zone. Direct immunofluorescence examination of the conjunctiva and oral mucosa demonstrated linear and shaggy fibrinogen deposition along the basement membrane zone, confirming the diagnosis of lichen planus. All six patients were placed on immunosuppressive therapy with control of the disease. However, only one patient was able to discontinue the anti-inflammatory medication and have the lichen planus remain in remission. CONCLUSIONS: Lichen planus should be included in the differential diagnosis of cicatrizing conjunctivitis. Performing appropriate investigations to distinguish conjunctival lichen planus from other autoimmune diseases such as mucous membrane pemphigoid is critical to managing the patient with cicatrizing conjunctivitis appropriately. Oral cyclosporine effectively controlled the conjunctival lichen planus in four of the six cases.     

Detection of ocular mucus in normal human conjunctiva and conjunctiva from patients with cicatricial pemphigoid using lectin probes and histochemical techniques

Conjunctival biopsies from patients with cicatricial pemphigoid affecting the conjunctiva and patients undergoing cataract surgery (normal conjunctiva) were snap-frozen, cryostat sectioned and incubated with fluorescein-conjugated lectins; peanut agglutinin (PNA), Helix pomatia agglutinin (HPA), soybean agglutinin (SBA), wheat germ agglutinin (WGA) and succinylated wheat germ agglutinin (S-WGA). Controls consisted of preincubating the lectins with the appropriate blocking sugars before applying the lectins to the sections. PNA and HPA stained the mucus granules contained in the conjunctival goblet cells but did not stain mucus or glycocalyx at the ocular surface distal to the goblet cells. Native WGA and S-WGA had high affinities for conjunctival goblet cells and the apical epithelial cell layers. Native WGA stained mucus and glycocalyx at the ocular surface. This staining of the ocular surface by WGA was confirmed at the transmission electron microscopic level using WGA conjugated to ferritin. Cicatricial pemphigoid patients in this study had reduced numbers of goblet cells; however, those goblet cells which were observed in cicatricial pemphigoid conjunctiva stained positively with HPA, PNA, WGA, and SWGA as did goblet cells in normal conjunctiva.