Active Ocular Cicatricial Pemphigoid Research Articles

Elevated serum BAFF in patients with ocular cicatricial pemphigoid

ObjectifL'efficacité de la déplétion lymphocytaire B dans le traitement de la pemphigoïde oculaire cicatricielle (POC) grave met en évidence le rôle clé que jouent les lymphocytes B dans l'immunopathogénèse de la POC. Le facteur d'activation des lymphocytes B (BAFF) est un puissant facteur de croissance des lymphocytes B et agit comme molécule de costimulation de la production d'immunoglobines. La hausse des taux sériques de BAFF est associée à des maladies auto-immunes généralisées, notamment le lupus érythémateux disséminé, la polyarthrite rhumatoïde et la pemphigoïde bulleuse. Nous postulons que les taux sériques de BAFF sont également élevés en présence de POC. MéthodesOn a prélevé le sérum de 30 patients qui présentaient une POC évolutive d'apparition récente, de 9 patients en rémission, de 10 patients en période de récidive et de 15 témoins en bonne santé. Le test ELISA (enzyme-linked immunosorbent assay) a servi à mesurer les taux sériques de BAFF. RésultatsLes taux de BAFF étaient significativement plus élevés chez les patients qui présentaient une POC évolutive d'apparition récente (700,8 ± 181,8 pg/mL) que chez les témoins en bonne santé (564,1 ± 133,2 pg/mL; p = 0,014). On n'a pas observé de différence significative entre les patients dont la POC était en rémission (585,4 ± 216,2 pg/mL) et les témoins en bonne santé. Les patients en période de récidive qui ont reçu le rituximab présentaient des taux extrêmement élevés de BAFF (1721,9 ± 790,8 pg/mL). L'analyse longitudinale des taux sériques de BAFF provenant de 6 patients a révélé que les taux de BAFF diminuaient à mesure que la maladie évoluait, passant de 895,0 ± 240,8 pg/mL à 625,4 ± 199,8 pg/mL (p = 0,003) entre le moment d'apparition et la rémission de la POC. ConclusionsLe BAFF contribue à l'inflammation active observée dans la POC évolutive. L'administration d'un antagoniste du BAFF peut avoir des effets bénéfiques dans la POC réfractaire, surtout en présence d'une résistance au rituximab.

Membrane Array Analysis of Tear Proteins in Ocular Cicatricial Pemphigoid

To explore non-invasive, protein-based, membrane array technology as a means to evaluate the global immune and angiogenic profile of tear proteins in patients with active ocular cicatricial pemphigoid (OCP). Forty-three proteins consisting of cytokines, angiogenic/growth factors, and immunoinflammatory modulators were measured by membrane array in tear samples of four control patients and four OCP patients during active disease and after treatment. Signals for several distinct and consistent molecular entities were upregulated in all four active OCP tear samples relative to controls. In particular, interleukin-8 and matrix metalloproteinase-9 were elevated during active disease and decreased after systemic immunomodulatory therapy. Protein array analysis may provide a well-tolerated assay to monitor levels of inflammatory markers in the tears of OCP patients in response to therapy.

Biology of interleukin-5 in ocular cicatricial pemphigoid.

Ocular cicatricial pemphigoid (OCP) is an autoimmune disease characterized by the presence of autoantibodies, T-cell dysregulation, and abnormal serum levels of cytokines such as interleukin-6, interleukin-1, and tumor necrosis factor-alpha. The purpose of the present study was to investigate levels of interleukin-5 (IL-5) in the sera, eosinophil counts in the peripheral blood, and eosinophil and mast cell counts in the inflamed conjunctivae of patients with active OCP. Seven patients diagnosed in the active phase of OCP presenting with chronic cicatrizing conjunctivitis were studied. The serum levels of IL-5 were compared to a group of seven age-, race-, and sex-matched normal human subjects. Eosinophil and mast cell counts in the patients' conjunctivae were compared to those in normal conjunctivae harvested during cataract surgery from seven normal individuals. In addition, eosinophil counts in peripheral blood of patients with active OCP were compared to those in normal individuals. The mean serum level of IL-5 in patients with active OCP was higher (67.23 pg/ml, range 46.33-98.26 pg/ml) than that in normal individuals (12.18 pg/ml, range 7.66-18.86). The difference was statistically significant ( P<0.001). On light microscopy the biopsied conjunctivae stained with hematoxylin and eosin revealed statistically significant differences ( P<0.001) in the mean numbers of eosinophils in the substantia propria between the patients with active OCP (6.8 cells/cm(2), range 4.8-8.2 cells/cm(2)) and normal controls (0.91 cells/cm(2), range 0.4-1.8 cells/cm(2)). The average number of mast cells found in the substantia propria of the biopsied conjunctivae was statistically significantly higher in patients with OCP (13.79 cells/cm(2), range 6.6-19.4) than in normal individuals (4.34 cells/cm(2), range 3.2-7.8; P<0.01). The average number of eosinophils in the peripheral blood of patients with active OCP (6.6x10(7)/l, range 2.9 - 9.3x10(7)/l) was statistically significantly higher ( P<0.01) than in normal controls (2.09x10(7)/l, range 0 - 4.5x10(7)/l). The results suggest that IL-5 may play an important role in the pathogenesis of OCP.

Ocular cicatricial pemphigoid antigen: partial sequence and biochemical characterization.

Ocular cicatricial pemphigoid (OCP) is an autoimmune disease that affects mainly conjunctiva and other squamous epithelia. OCP is histologically characterized by a separation of the epithelium from underlying tissues within the basement membrane zone. Immunopathological studies demonstrate the deposition of anti-basement membrane zone autoantibodies in vivo. Purified IgG from sera of patients with active OCP identified a cDNA clone from a human keratinocyte cDNA library that had complete homology with the cytoplasmic domain of beta 4-integrin. The sera recognized a 205-kDa protein in human epidermal, human conjunctiva, and tumor cell lysates that was identified as beta 4-integrin by its reaction with polyclonal and monoclonal antibodies to human beta 4-integrin. Sera from patients with bullous pemphigoid, pemphigus vulgaris, and cicatricial pemphigoid-like diseases did not recognize the 205-kDa protein, indicating the specificity of the binding. These data strongly implicate a role for human beta 4-integrin in the pathogenesis of OCP. It should be emphasized that multiple antigens in the basement membrane zone of squamous epithelia may serve as targets for a wide spectrum of autoantibodies observed in vesiculobullous diseases. Molecular definition of these autoantigens will facilitate the classification and characterization of subsets of cicatricial pemphigoid and help distinguishing them from bullous pemphigoid. This study highlights the function and importance of beta 4-integrin in maintaining the attachment of epithelial cells to the basement membrane.

Collagen Abnormalities in Conjunctiva of Patients with Cicatricial Pemphigoid

The purpose of this study was to analyze the distribution and types of collagen in the substantia propria of the conjunctiva of patients with ocular cicatricial pemphigoid (OCP). Biopsy specimens were collected from 10 patients with active OCP, five patients with active Behçet's disease, nine patients with atopic keratoconjunctivitis, five patients with chronic rosacea blepharoconjunctivitis, and six normal patients undergoing cataract surgery. Cryostat tissue sections were cut and stained using an indirect immunofluorescence technique, employing a panel of primary antibodies directed against seven collagen types. Differences between OCP, Behçet's, and normal conjunctiva were seen in the staining for collagen types III, IV, and VII. The intensity of staining for type III collagen was increased in the substantia propria of OCP conjunctiva as compared to the other groups. The basement membrane zone (BMZ) of OCP patients was typically disrupted and fragmented in appearance when stained for type IV collagen, a finding not seen in the non-OCP specimens. The BMZ staining pattern for type VII collagen in OCP conjunctiva was even more disrupted than that seen for type IV collagen, particularly on the posterior side, which was thickened and reduplicated with short fibers extending into the superficial stroma. The production of type III collagen by fibroblasts of the substantia propria is a common feature of diseases associated with subepithelial fibrosis. The damage to the epithelial BMZ and the subsequent attempt at repair with aberrant type IV and VII collagen production are unique to conjunctiva affected by OCP.

Detection of Cellular Adhesion Molecules in Inflamed Human Corneas

The expression of cellular adhesion molecules in 31 penetrating keratoplasty specimens from a broad range of corneal inflammatory diseases was studied using monoclonal antibodies and an immunoperoxidase technique. Corneas were divided into noninflamed, mild to moderately inflamed, and severely inflamed groups based on histologic findings. The panel of adhesion molecules studied included HLA-ABC, HLA-DR, CD3, LFA-1, MAC-1, ICAM-1, PECAM-1, VCAM-1, and E-selectin-1. The adhesion molecules ICAM-1, HLA-DR, PECAM-1, CD3, VCAM-1, LFA-1, and MAC-1 were selectively expressed in areas of corneal inflammation. In general, HLA-DR and intercellular adhesion molecule ICAM-1 were co-expressed in similar regions. PECAM-1 was restricted to zones of marked inflammation and vascularization. E-selectin-1 was detected only in the stroma of a graft melt in a patient with active ocular cicatricial pemphigoid, and may reflect a primary regulatory dysfunction in this disorder. The ICAM-1 ligand was, in general, more diffusely distributed than its receptor LFA-1, a beta-2 integrin found on leukocyte cell membranes. The localization of the integrin MAC-1, present on macrophages, neutrophils, and some lymphocytes, did not always parallel the staining pattern of ICAM-1, suggesting promiscuity in its binding to other ligands besides ICAM-1. Adhesion molecules are detected readily at sites of corneal inflammation and may play a critical role in facilitating the recruitment of immune regulatory cells to these areas. Future efforts to block or modulate the expression of intercellular adhesion molecules may provide new therapeutic options in the treatment of corneal inflammatory diseases.

Detection and partial characterization of ocular cicatricial pemphigoid antigens on COLO and SCaBER tumor cell lines.

Ocular cicatricial pemphigoid (OCP) is a chronic autoimmune inflammatory disease which affects the conjunctiva and other squamous epithelial mucous membranes resulting in a scarring process. It is characterized by the deposition of an anti-basement membrane zone (BMZ) antibody in vivo. Sera from 11 patients with active OCP were studied. Using monkey esophagus and normal skin as substrate, weak staining of the BMZ was observed in conventional indirect immunofluorescence (IIF) assay. Using salt split human skin as substrate, the OCP sera demonstrated binding to the epidermal side of the split, in low titers with weak staining. Ten of the 11 sera were positive on an immunoblot assay using COLO and SCaBER tumor cell lysates demonstrating 230, 205, 160, and 85 kD proteins. Sera from six bullous pemphigoid (BP) patients, with only cutaneous involvement and high titer of anti-BMZ antibody, as detected by IIF, also bound to 230, 160, and 85 kD proteins on both lysates in comigration experiments. Serum from five normal individuals and two patients each with severe atopic conjunctival disease, erythema multiforme with chronic conjunctivitis and systemic lupus erythematosus (SLE), did not demonstrate those bands. When the two lysates were first absorbed with BP sera and then the same lysates were immunoblotted with OCP sera, in all ten OCP sera the 230, 160, and 85 kD bands were eliminated and only a single 205 kD band was uniformly present. These results indicate that OCP sera recognize peptide(s) present in 230, 205 and 160 kD proteins in lysates from COLO and SCaBER tumor cells. These proteins contain the immunodominant region of the BMZ molecule(s) in which the OCP antigen(s) reside. The OCP antigen(s) appears to be distinct from the BP antigen(s).

Systemic Chemotherapy for Ocular Cicatricial Pemphigoid

The records of 105 patients treated with three different chemotherapeutic agents for ocular cicatricial pemphigoid (OCP) were reviewed to compare long-term efficacies, side effects, and tolerance of different regimens. For the entire group, OCP progressed in 6% of eyes in 10% of patients (follow-up 35 months). More than half of the treatment failures occurred in patients intolerant of chemotherapy. Diaminodiphenylsulfone (DAP), as initial agent, failed to control disease in 2% of patients, compared with 8% after cyclophosphamide (CYC) and 9% after azathioprine (AZA) (p less than 0.05). Stratification of results revealed that DAP was the most effective initial agent for modestly active OCP, whereas CYC was the most effective initial choice for highly active cases. In patients treated with a single agent exclusively for 10 months or more, failure to control disease occurred in 4% of DAP, 4% of CYC, and 15% of AZA patients (p less than 0.01). Recommendations for a sequential approach to chemotherapy for OCP are presented.

Scanning Electron Microscopy of Conjunctival Surfaces in Patients With Ocular Cicatricial Pemphigoid

The conjunctival surfaces of ten patients with active, ocular cicatricial pemphigoid, three patients with drug-controlled ocular cicatricial pemphigoid, and six patients with normal conjunctivas were studied using scanning electron microscopy. A homogeneous granular sheet of amorphous mucin-like material was observed covering extensive areas of the conjunctiva in eight of ten patients with active ocular cicatricial pemphigoid. This sheet of amorphous material was absent on drug-controlled ocular cicatricial pemphigoid and normal conjunctival specimens. Our study demonstrates that patients with active ocular cicatricial pemphigoid possess ocular surface mucus that appears thicker and more continuous than normal ocular mucus when observed with scanning electron microscopy. This observation is in agreement with clinical observations of thick mucus strands in the inferior fornix of patients with active ocular cicatricial pemphigoid.

Ultrastructural characteristics of conjunctiva in cicatricial pemphigoid.

Conjunctival biopsy specimens were obtained from three patients with clinically active ocular cicatricial pemphigoid and from two of these patients after successful immunosuppressive therapy. Compared with specimens from normal subjects, the pretreatment specimens showed profound alteration in interepithelial adhesion as evidenced by a dramatic increase in desmosomes. Unusually prominent tonofilaments and tonofibrils were seen throughout the cytoplasm. The basal lamina showed areas of discontinuity, duplication, and focal thickening. The lamina propria was thickened and demonstrated an infiltration of inflammatory cells and disorganized collagen fibrils. Post-treatment specimens had a relative absence of desmosomes between the widely separated epithelial cells, less prominent tonofilaments and tonofibrils among epithelial cells, and, in one specimen, goblet cells. The enhanced network of desmosome-tonofilament complexes serves to tightly bind epithelial cells, perhaps providing a clue to the pathophysiology of conjunctival surface changes.