Fumihide Ogawa

Correlation of IgE Autoantibody to BP180 With a Severe Form of Bullous Pemphigoid

OBJECTIVE To determine the prevalence, immunoglobulin subclass distribution, and clinical correlation of antibodies (Abs), especially of IgE Abs, to BP180 and BP230 in patients with bullous pemphigoid (BP). DESIGN Retrospective case series analysis. SETTING Department of Dermatology, Nagasaki University Graduate School of Biomedical Science. PATIENTS Serum samples from 37 patients with BP, 6 with pemphigus vulgaris, 5 with pemphigus foliaceus, and 26 healthy controls (n = 26) were examined by enzyme-linked immunosorbent assay. MAIN OUTCOME MEASURES Prevalence, immunoglobulin subclass distribution, and clinical correlation of Abs, especially of IgE Abs, to BP180 and BP230. RESULTS IgG anti-BP180 and anti-BP230 Abs were detected in 35 (95%) and 26 (70%) of the 37 BP serum samples, respectively. IgG1 and IgG4 isotypes were positive in 32 (87%) and 25 (68%), respectively, of the BP serum samples for anti-BP180 Abs, while they were detected in 16 (44%) and 26 (70%), respectively, for anti-BP230 Abs. IgE anti-BP180 and anti-BP230 Abs were equally detected in 8 (22%) of the BP serum samples. Similar to IgG anti-BP180 Abs, the presence or levels of IgE anti-BP180 Abs was associated with broader skin lesions. Furthermore, patients with BP positive for IgE anti-BP180 Abs required longer duration for remission, higher dosage of prednisolone, and more intensive therapies for remission. By contrast, this was not true for those with of IgE anti-BP230 Abs. Remarkably, when analyzed in patients with BP who had a high titer of IgG anti-BP180 Abs, the presence or levels of IgE anti-BP180 Abs, but not IgG anti-BP180 Abs, were associated with a more severe form. CONCLUSIONS The present study suggests that IgE anti-BP180 Abs are related to the disease severity and activity of BP. Moreover, it may be possible to identify treatment-refractory patients with BP more specifically by assessing the presence or levels of IgE anti-BP180 Abs in those with a high IgG anti-BP180 Ab titer.

Cell Adhesion Molecules Regulate Fibrotic Process via Th1/Th2/Th17 Cell Balance in a Bleomycin-Induced Scleroderma Model

Mice s.c. injected with bleomycin, an experimental model for human systemic sclerosis, develop skin and lung fibrosis, which is mediated by inflammatory cell infiltration. This process is highly regulated by multiple adhesion molecules and does not require Ag sensitization. To assess the role of adhesion molecules in this pathogenetic process, bleomycin-induced fibrosis was examined in mice lacking adhesion molecules. L-selectin and/or ICAM-1 deficiency inhibited skin and lung fibrosis with decreased Th2 and Th17 cytokines and increased Th1 cytokines. In contrast, P-selectin deficiency, E-selectin deficiency with or without P-selectin blockade, or P-selectin glycoprotein ligand 1 (PSGL-1) deficiency augmented the fibrosis in parallel with increased Th2 and Th17 cytokines and decreased Th1 cytokines. Furthermore, loss of L-selectin and/or ICAM-1 reduced Th2 and Th17 cell numbers in bronchoalveolar lavage fluid, whereas loss of P-selectin, E-selectin, or PSGL-1 reduced Th1 cell numbers. Moreover, Th1 cells exhibited higher PSGL-1 expression and lower expression of LFA-1, a ligand for ICAM-1, whereas Th2 and Th17 cells showed higher LFA-1 and lower PSGL-1 expression. This study suggests that L-selectin and ICAM-1 regulate Th2 and Th17 cell accumulation into the skin and lung, leading to the development of fibrosis, and that P-selectin, E-selectin, and PSGL-1 regulate Th1 cell infiltration, resulting in the inhibition of fibrosis.

Treatment with rapamycin prevents fibrosis in tight‐skin and bleomycin‐induced mouse models of systemic sclerosis

OBJECTIVE Rapamycin, a novel macrolide immunosuppressive drug, is increasingly used as an agent for posttransplant immunosuppression and treatment of autoimmune disease. The molecular mechanism related to rapamycin-mediated immunosuppression is that rapamycin binds to FK-506 binding protein 12, and the formed complex inhibits the function of the mammalian target of rapamycin (mTOR), which in turn reduces protein phosphorylation, cell cycle progression, and cytokine production. The aim of this study was to examine the effect of rapamycin against the development of fibrosis and autoimmunity in 2 different types of systemic sclerosis (SSc) model mice. METHODS Tight skin (TSK/+) mice and bleomycin- induced SSc model mice were used to evaluate the effect of rapamycin on fibrosis and immunologic abnormalities. Furthermore, the antifibrotic effect of rapamycin was assessed using TSK/+ mouse fibroblasts. RESULTS Treatment with rapamycin reduced skin fibrosis of TSK/+ mice and skin and lung fibrosis of bleomycin-induced SSc model mice. The production of fibrogenic cytokines, such as interleukin-4 (IL-4), IL-6, IL-17, and transforming growth factor beta1, was attenuated by rapamycin. Hypergammaglobulinemia and anti-topoisomerase I antibody production were also reduced by rapamycin treatment in TSK/+ mice. In addition, mTOR expression levels were increased in TSK/+ mouse fibroblasts compared with those in wild-type mouse fibroblasts. Rapamycin treatment inhibited proliferation and collagen production of TSK/+ mouse fibroblasts in a dose-dependent manner. CONCLUSION This study is the first to show that rapamycin has a significant inhibitory effect on fibrosis in both TSK/+ and bleomycin-induced SSc model mice. These results suggest that rapamycin might be an attractive candidate for clinical trials in SSc patients.

The clinical relevance of serum antinuclear antibodies in Japanese patients with systemic sclerosis

Background  Systemic sclerosis (SSc) is a connective tissue disorder with excessive fibrosis of the skin and various internal organs. Although SSc is a heterogeneous disease, it has been reported that the particular antinuclear antibodies (ANA) are often indicative of clinical features, disease course and overall severity.

Clinical Significance of Serum HMGB-1 and sRAGE Levels in Systemic Sclerosis: Association with Disease Severity

IntroductionThe high mobility group box 1 protein (HMGB-1)/advanced glycation end products (RAGE) system is recently shown to play an important part in immune/inflammatory disorders. However, the association of this system in systemic sclerosis (SSc) remains unknown.Materials and MethodsTo determine clinical association of serum levels of HMGB-1 and soluble RAGE (sRAGE) in patients with SSc, sera from 70 patients with SSc and 25 healthy controls were examined by enzyme-linked immunosorbent assay. Sera from tight-skin mice and bleomycin-induced scleroderma mice, animal models for SSc, were also examined. Skin HMGB-1 and RAGE expression was assessed by immunohistochemistry.Results and DiscussionSerum HMGB-1 and sRAGE levels in SSc were higher than those in controls. Similarly, HMGB-1 and sRAGE levels in animal SSc models were higher than those in control mice. SSc patients with elevated HMGB-1 and sRAGE levels had more frequent involvement of several organs and immunological abnormalities compared to those with normal levels. Furthermore, HMGB-1 and sRAGE levels correlated positively with modified Rodnan total skin thickness score and negatively with pulmonary function test.ConclusionsHMGB-1 and sRAGE expression in the sclerotic skin was more intense than normal skin. These results suggest that elevated serum HMGB-1 and sRAGE levels are associated with the disease severity and immunological abnormalities in SSc.

The Loss of MCP-1 Attenuates Cutaneous Ischemia–Reperfusion Injury in a Mouse Model of Pressure Ulcer

The formation of pressure ulcers is dependent on multiple factors including ischemia-reperfusion (IR). This study assessed the mechanism of a previously reported murine model of cutaneous IR injury. Three cycles of IR (days 1-3) by external application of two magnetic plates were performed to induce pressure ulcer formation. Increased infiltration of neutrophils and macrophages, and augmented expression of proinflammatory cytokines and inducible nitric oxide synthase (iNOS), were observed during IR cycles. In this model, monocyte chemoattractant protein-1 (MCP-1) was remarkably increased at day 1 in the skin followed by inflammatory cell infiltration. Therefore, IR cycles were performed in MCP-1-deficient (MCP-1(-/-)) mice to evaluate the role of this chemokine in pressure ulcer development. MCP-1(-/-) mice showed reduced macrophage infiltration and expression of tumor-necrosis factor-alpha (TNF)-alpha and iNOS during IR cycles leading to attenuated apoptosis and skin injury. Importantly, MCP-1 played a role in apoptosis and injury via inducing iNOS during the reperfusion rather than the ischemic period. These findings indicate that MCP-1 may be a critical factor for macrophage recruitment and subsequent skin inflammation and injury during IR cycles. We propose that this is a useful model for investigating the mechanism of pressure ulcer formation using various transgenic mice.

CD19 regulates the development of bleomycin‐induced pulmonary fibrosis in a mouse model

OBJECTIVE The contribution of CD19 and B lymphocytes to pulmonary fibrosis is controversial. The aim of this study was to address the role of CD19 during the development of pulmonary fibrosis. METHODS Mice lacking or overexpressing the B cell surface molecule CD19, which is known as a positive regulator of B cell activation, were used in a model of bleomycin-induced pulmonary fibrosis. Ten or sixteen days after intratracheal injection of bleomycin, lung sections from mice were evaluated by histologic analysis. Seven days after instillation, the total leukocyte count and the number of B cells in bronchoalveolar lavage fluid (BALF) were determined, using a hemocytometer and flow cytometry. Bleomycin was also administered into selectin-deficient or intercellular adhesion molecule 1-deficient mouse strains. The level of CXCR3 expression on B cells was determined by flow cytometry. RESULTS CD19 deficiency significantly reduced susceptibility to intratracheal bleomycin challenge on day 16, while CD19 overexpression augmented fibrosis even on day 10. Furthermore, the survival rate and number of B cells in BALF also correlated with CD19 expression levels. The accumulation of B cells in BALF was dependent on CD19 levels, whereas there was no association with the levels of selectins or intercellular adhesion molecule 1. Additionally, CXCR3 was up-regulated in BALF B cells, while it was rarely expressed on circulating B cells. Furthermore, CD19 signaling facilitated B cell CXCR3 up-regulation in response to stimulation in vitro. CONCLUSION These results suggest that CD19 signaling is associated with the development of pulmonary fibrosis by controlling B cell infiltration into lung tissue, which may be associated with CXCR3 up-regulation.

Elevated serum BAFF levels in patients with localized scleroderma in contrast to other organ-specific autoimmune diseases

Abstract:  Serum levels of B‐cell activating factor belonging to the tumor necrosis factor family (BAFF), a potent B‐cell survival factor, are elevated in patients with systemic autoimmune diseases, such as systemic lupus erythematosus (SLE), rheumatoid arthritis and systemic sclerosis (SSc). The objective of this study was to determine serum BAFF levels and relate the results to the clinical features in patients with organ‐specific autoimmune diseases of the skin, such as localized scleroderma and autoimmune bullous diseases. Serum BAFF levels were examined by enzyme‐linked immunosorbent assay in 44 patients with localized scleroderma, 20 with pemphigus vulgaris/pemphigus foliaceus, 20 with bullous pemphigoid and 30 healthy controls. Twenty patients with SSc and 20 with SLE were also examined as disease controls. Serum BAFF levels were elevated in localized scleroderma patients compared with healthy controls. Concerning localized scleroderma subgroups, patients with generalized morphea, the severest form of localized scleroderma, had higher serum BAFF levels than linear scleroderma or morphea patients. The BAFF levels of generalized morphea were comparable with those of SSc or SLE. Furthermore, serum BAFF levels correlated positively with antihistone antibody levels and the severity of skin lesion as well as the number of skin lesions. By contrast, serum BAFF levels were not significantly elevated in patients with pemphigus or pemphigoid. These results suggest that BAFF may be contributing to autoimmunity and disease development in localized scleroderma.

Platelets control leukocyte recruitment in a murine model of cutaneous arthus reaction.

Platelets have been shown to be important in inflammation, but their role in the cutaneous Arthus reaction remains unclear. To assess the role of platelets in this pathogenetic process, the cutaneous Arthus reaction was examined in wild-type mice and mice lacking E-selectin, P-selectin, or P-selectin glycoprotein ligand-1 (PSGL-1) with or without platelet depletion by busulfan, a bone marrow precursor cell-specific toxin. Edema and hemorrhage induced by immune complex challenge significantly decreased in busulfan-treated wild-type mice compared with untreated mice. Busulfan treatment did not affect edema and hemorrhage in P-selectin- or PSGL-1-deficient mice, suggesting that the effect by busulfan is dependent on P-selectin and PSGL-1 expression. The inhibited edema and hemorrhage paralleled reduced infiltration of neutrophils and mast cells and reduced levels of circulating platelets. Increased cutaneous production of interleukin-6, tumor necrosis factor-alpha, and platelet-derived chemokines during Arthus reaction was inhibited in busulfan-treated wild-type mice relative to untreated mice, which paralleled the reduction in cutaneous inflammation. Flow cytometric analysis showed that immune complex challenge generated blood platelet-leukocyte aggregates that decreased by busulfan treatment. In thrombocytopenic mice, the cutaneous inflammation after immune complex challenge was restored by platelet infusion. These results suggest that platelets induce leukocyte recruitment into skin by forming platelet-leukocyte aggregates and secreting chemokines at inflamed sites, mainly through the interaction of P-selectin on platelets with PSGL-1 on leukocytes.

Blockade of CD40-CD40 ligand interactions attenuates skin fibrosis and autoimmunity in the tight-skin mouse

Objective : To assess the association of CD40/CD40 ligand (CD40L) interactions with the development of skin fibrosis and autoimmunity in tight-skin (TSK/+) mouse, which is a mouse model for human systemic sclerosis. Methods: Newly born TSK/+ mice were treated with murine anti-CD40L monoclonal antibody (100 μg intraperitoneally weekly). Hypodermal thickness of 8-week-old female mice (defined as the thickness of a subcutaneous loose connective tissue layer beneath the panniculus carnosus) was measured under a light microscope. All skin sections were taken from the para-midline, upper back region. Serum anti-topoisomerase I autoantibody levels, serum immunoglobulin levels and plasma soluble CD40L levels were determined by enzyme-linked immunosorbent assay. For analysis of lymphocyte surface molecules, single cell suspensions of lymphocytes were stained by monoclonal antibodies. Proliferation of TSK/+ B cells and fibroblasts to anti-CD40 antibodies was assessed by the uptake of [3H]-labelled thymidine and bromodeoxyuridine, respectively. Results: The blockade of CD40/CD40L interactions by anti-CD40L monoclonal antibody significantly reduced cutaneous fibrosis (65%) and anti-topoisomerase I autoantibody in TSK/+ mice. Anti-CD40L monoclonal antibody also normalised B lymphocyte abnormal activation in TSK/+ mice, demonstrated by hyper-γ-globulinaemia. Furthermore, augmented CD40/CD40L interactions in TSK/+ mice were suggested by upregulated expression of CD40L on CD4+ T cells, elevated plasma soluble CD40L levels. The hyperresponsiveness to CD40 stimulation was also observed in TSK/+ B cells and fibroblasts. Conclusions: Cutaneous fibrosis and autoimmunity in TSK/+ mice are closely correlated with CD40/CD40L interactions.