A potential contribution of altered cathepsin L expression to the development of dermal fibrosis and vasculopathy in systemic sclerosis

Cathepsin L (CTSL) is a lysosomal proteolytic enzyme involved in inflammation and vascular and extracellular matrix remodelling, which are the three cardinal pathological events associated with systemic sclerosis (SSc). To elucidate the potential role of CTSL in the development of SSc, we here investigated CTSL expression in the lesional skin of patients with SSc and SSc animal models and the clinical correlation of serum CTSL levels. CTSL expression was elevated in dermal small vessels of SSc patients compared with those of healthy controls. Consistently, CTSL mRNA levels were increased in SSc lesional skin samples, but not in cultivated SSc dermal fibroblasts, compared with corresponding control samples from healthy individuals. Serum CTSL levels were significantly higher in SSc patients than in healthy controls and inversely correlated with skin score. Furthermore, the elevation of serum CTSL levels was linked to SSc vasculopathy. Supporting these results, Ctsl mRNA levels were decreased in the skin of bleomycin‐treated mice, an SSc animal model recapitulating its fibrotic aspect, and CTSL expression was enhanced in dermal small vessels of endothelial cell‐specific Fli1 knockout mice, reminiscent of SSc vasculopathy. Importantly, gene silencing of FLI1 induced CTSL mRNA expression and Fli1 occupied the CTSL promoter in human dermal microvascular endothelial cells. Collectively, these results suggest that endothelial CTSL up‐regulation partially due to Fli1 deficiency may contribute to the development of vasculopathy, while the decrease in dermal CTSL expression is likely associated with dermal fibrosis in SSc.     

Serum levels of interleukin‐18‐binding protein isoform a: Clinical association with inflammation and pulmonary hypertension in systemic sclerosis

Systemic sclerosis (SSc) is a chronic autoimmune inflammatory disease characterized by extensive tissue fibrosis and various vascular complications. A wealth of evidence suggests the substantial contribution of pro‐inflammatory cytokines to the development of SSc, but the role of interleukin (IL)‐18 signaling in this disease still remains elusive. To address this issue, we herein determined serum levels of IL‐18‐binding protein isoform a (IL‐18BPa), a soluble decoy receptor for IL‐18, by enzyme‐linked immunosorbent assay in 57 SSc patients and 20 healthy controls and evaluated their clinical correlation. Serum IL‐18BPa levels were higher in SSc patients than in healthy controls, while comparable between diffuse cutaneous SSc and limited cutaneous SSc patients. Although serum IL‐18BPa levels were not associated with dermal and pulmonary fibrotic parameters in SSc patients, there was a significant positive correlation between serum IL‐18BPa levels and right ventricular systolic pressure estimated by echocardiography. Furthermore, in 24 SSc patients who underwent right heart catheterization, serum IL‐18BPa levels positively correlated with mean pulmonary arterial pressure. As for systemic inflammatory markers, significant positive correlations of circulating IL‐18BPa levels with erythrocyte sedimentation rate and C‐reactive protein were noted. These results suggest that the inhibition of IL‐18 signaling by IL‐18BPa may be involved in the development of pulmonary vascular involvement leading to pulmonary hypertension and modulate the systemic inflammation in SSc.     

Serum interleukin‐34 levels in patients with systemic sclerosis: Clinical association with interstitial lung disease

Interleukin (IL)‐34 is a hematopoietic cytokine promoting proliferation and differentiation of macrophages. Because abnormal activation of macrophages is involved in the development of systemic sclerosis (SSc), we investigated serum IL‐34 levels in patients with SSc. Serum IL‐34 levels were significantly increased in diffuse cutaneous SSc compared with limited cutaneous SSc and healthy controls, while there were no significant differences between limited cutaneous SSc and healthy controls. In addition, SSc patients with increased serum IL‐34 levels more often had interstitial lung disease (ILD) than those with normal levels. Moreover, in SSc patients, serum IL‐34 levels negatively correlated with the percentage of predicted vital capacity, while they positively correlated with ground‐glass opacity score and fibrotic score on chest computed tomography. Collectively, increased serum IL‐34 levels were associated with greater frequency and severity of ILD in SSc patients. Serum IL‐34 levels may be a useful serological marker for SSc‐associated ILD.     

Rapid alteration of serum interleukin‐6 levels may predict the reactivity of i.v. cyclophosphamide pulse therapy in systemic sclerosis‐associated interstitial lung disease

Systemic sclerosis (SSc) is an autoimmune disorder characterized by excessive extracellular matrix deposition. Although SSc‐associated interstitial lung disease (ILD) is one of the most important complications as a cause of death in SSc, prediction factors of treatment reactivity in SSc‐ILD are still unclear. To assess relationships between interleukin (IL)‐6 and reactivity to treatment, we measured serum IL‐6 levels in 23 of active SSc‐ILD patients under i.v. cyclophosphamide (IVCY) therapy and 20 of stabilized SSc‐ILD, using the high‐sensitivity enzyme‐linked immunoassay system. Serum IL‐6 levels in active SSc‐ILD patients were significantly higher than those in stabilized SSc‐ILD patients. Among active SSc‐ILD patients, baseline serum IL‐6 levels were not significantly different between IVCY responders and non‐responders. Meanwhile, serum IL‐6 levels after three IVCY doses out of a total of six were decreased in responders but not in non‐responders. Regarding changes of parameters by the three doses of a total of six of IVCY, change in serum IL‐6 levels correlated inversely with that in values of pulmonary function test. Thus, the rapid decrease in serum IL‐6 levels during a couple of doses may predict the efficacy of IVCY therapy against SSc‐ILD.     

Clinical significance of serum soluble T‐cell immunoglobulin and mucin domain 3 levels in systemic sclerosis: Association with disease severity

T‐cell immunoglobulin and mucin domain 3 (TIM‐3) has been thought to play a crucial role in the negative regulation of immune responses. Here, we examined the levels of serum soluble TIM‐3 (sTIM‐3) in patients with systemic sclerosis (SSc) and evaluated the results with respect to the clinical features of the disease. Patients with diffuse cutaneous SSc (dcSSc) had higher levels of sTIM‐3 than those with limited cutaneous SSc and healthy individuals. Serum sTIM‐3 levels were positively correlated with the severity of skin sclerosis in early phase dcSSc. Moreover, serum sTIM‐3 levels were increased more often in patients with renal crisis and cardiac involvement than in those with normal sTIM‐3 levels. These results suggest that serum sTIM‐3 levels may be increased in patients with early phase dcSSc and associated with cardiac involvement and renal crisis. Measurement of serum sTIM‐3 may be useful for risk stratification in the early stage of the disease.     

CXCL13 produced by macrophages due to Fli1 deficiency may contribute to the development of tissue fibrosis,vasculopathy and immune activation in systemic sclerosis

CXCL13, a chemokine for B cells, follicular T cells, T helper 17 cells, and regulatory T cells, is reported to contribute to the development of systemic sclerosis (SSc), reflecting aberrant activation of immune system. To better understand the role of CXCL13 in SSc, we investigated the influence of Fli1 deficiency, a potential predisposing factor of this disease, on CXCL13 expression and assessed the clinical correlation of serum CXCL13 levels by multivariate regression analysis. Haploinsufficient loss of Fli1 remarkably induced CXCL13 expression in murine peritoneal macrophages, while gene silencing of FLI1 did not affect the expression of CXCL13 in human dermal fibroblasts and human dermal microvascular endothelial cells. Serum CXCL13 levels were elevated in SSc patients compared with healthy controls and correlated positively with skin score and negatively with pulmonary function test results. SSc patients with elevated serum CXCL13 levels had longer disease duration, diffuse cutaneous involvement, interstitial lung disease (ILD), heart involvement, pulmonary arterial hypertension, Raynaud's phenomenon, pitting scars, digital ulcers, telangiectasia, and high serum IgG levels more frequently than the other patients. In particular, serum CXCL13 levels were associated with ILD and digital ulcers by multivariate regression analysis. Taken together, these results indicate that CXCL13 expression is upregulated by Fli1 deficiency in macrophages, potentially contributing to the development of tissue fibrosis, vasculopathy and immune activation in SSc, especially ILD and digital ulcers.     

Serum levels of anti‐Fcγ receptor IIB/C antibodies are increased in patients with systemic sclerosis

Systemic sclerosis (SSc) is a systemic autoimmune disorder that results in fibrosis of the skin and multiple internal organs. Although the precise mechanism is unknown, it appears to result from the overproduction of extracellular matrix proteins and aberrant immune activations. Receptors for the Fc region of immunoglobulin (Ig)G (FcγR) are members of the Ig superfamily that modulate both activation and inhibition of immune responses. FcγRIIB is the sole inhibitory member, which has an intrinsic cytoplasmic immunoreceptor tyrosine‐based inhibitory motif. The present study was undertaken to investigate the circulating concentrations of anti‐FcγRIIB/C antibodies (Ab) in patients with SSc. Serum levels of anti‐FcγRIIB/C Ab were significantly increased in patients with SSc compared to those in controls and in patients with localized scleroderma. Serum levels of anti‐FcγRIIB/C Ab in patients with limited cutaneous SSc were similar to those in patients with diffuse cutaneous SSc. Among SSc patients, serum levels of anti‐FcγRIIB/C Ab were increased in those with nail‐fold bleeding and decreased in those with diffuse pigmentation and calcinosis. These findings support the notion that increased serum anti‐FcγRIIB/C Ab levels are involved in aberrant immune responses in SSc.     

Case of diffuse cutaneous systemic sclerosis with anti‐Ku and anti‐centromere antibodies

We report the case of a 58‐year‐old man who had an ulcer on the right middle finger that was cured by surgery 4 years before consultation with our department. A few years after the surgery, he noticed recurrence of the ulcer and sclerosis of the skin. At the initial examination, skin sclerosis was observed from the fingers to the upper arms and from the feet to the thighs. Pitting scars on the fingertips and punctured hemorrhages of the nail‐fold capillaries were also present. Gastroscopy showed slight reflex esophagitis. Laboratory findings were positive for antinuclear antibody (ANA; 1:640) with a speckled and discrete speckled pattern. Anti‐topoisomerase I (anti‐topo I) antibody and anti‐RNA polymerase III were negative, but anti‐centromere antibody was positive in an enzyme‐linked immunosorbent assay. Anti‐Ku antibody was positive in an immunoprecipitation assay using extracts of the leukemia cell line K562. Therefore, the patient was diagnosed with diffuse cutaneous systemic sclerosis with anti‐Ku and anti‐centromere antibodies. Treatment with an oral antiplatelet agent, vitamin E, a proton pump inhibitor, and i.v. lipoprostaglandin E1 were started. Subsequently, there has been repeated recurrence of finger ulcers, but no muscle involvement has been detected since his first visit. This is the first reported case of systemic sclerosis with anti‐Ku and anti‐centromere antibodies.     

Serum levels of soluble programmed death‐1 and programmed death ligand‐1 in systemic sclerosis: Association with extent of skin sclerosis

The interaction of programmed death‐1 (PD‐1) with its ligand, programmed death ligand‐1 (PD‐L1), has been considered to play a key role in the negative regulation of immune responses. Patients with diffuse cutaneous systemic sclerosis (SSc) had higher levels of soluble PD‐1 (sPD‐1) than those with limited cutaneous SSc and healthy individuals. Serum sPD‐1 levels positively correlated with the severity of skin sclerosis. In contrast, serum sPD‐L1 levels were significantly increased in patients with SSc compared with healthy individuals. Moreover, serum sPD‐L1 levels were not associated with the extent of skin sclerosis and were elevated not only in patients with diffuse cutaneous SSc, but also in those with limited cutaneous SSc. These results suggested that serum sPD‐1 levels may increase in patients with SSc and correlate with the severity of skin sclerosis. PD‐1/PD‐L1 interaction may contribute to the development of skin sclerosis in SSc.     

Demographic and clinical features of systemic sclerosis patients with anti‐RNA polymerase III antibodies

Anti‐RNA polymerase III antibody (RNAP) is primarily detected in diffuse cutaneous type systemic sclerosis (dcSSc) patients and strongly associated with renal crisis. Additionally, there has been increasing evidence that cancer in SSc patients is associated with RNAP. The aim of this study was to examine the demographic and clinical features of SSc patients with RNAP. Among 246 SSc patients, 5.7% were positive for RNAP, 20.7% were positive for anti‐topoisomerase I antibody (Topo I) alone and 39.4% were positive for anticentromere antibody (ACA) alone. The modified Rodnan total skin score (mRTSS) in SSc patients with RNAP (19.1 ± 2.6) was significantly higher than those in SSc patients with Topo I (11.5 ± 1.1) and patients with ACA (4.4 ± 0.4). Furthermore, among SSc patients with RNAP, the levels of RNAP were positively correlated with mRTSS. Renal crisis is also significantly more prevalent in SSc patients with RNAP than patients without RNAP. Male sex, dcSSc subtype, digital vasculopathy, including digital ulcers and acro‐osteolysis, interstitial lung disease and rheumatoid arthritis complications were prevalent in SSc patients with RNAP and patients with Topo‐I. Primary biliary cirrhosis and Sjögren's syndrome were more in SSc patients with RNAP and patients with ACA compared with patients with Topo 1. No significant difference in the frequency of complications, including Raynaud's phenomenon, pulmonary artery hypertension and malignancy was observed between the three groups. Thus, measurement of RNAP in SSc patients is useful for the diagnosis and risk stratification of severe manifestation, such as renal crisis and severe skin sclerosis.     

Association of anti‐RNA polymerase III antibody and malignancy in Japanese patients with systemic sclerosis

Patients with systemic sclerosis (SSc) have an increased risk of malignancy compared with the general population. Recently, SSc patients with anti‐RNA polymerase III antibody have been reported to have an increased risk of malignancy as compared with those with other disease‐specific autoantibodies in US, European and Australian populations. Therefore, we studied the relationship between disease‐specific autoantibodies and malignancy in 261 Japanese SSc patients. The prevalence of malignancy was significantly higher in patients with anti‐RNA polymerase III antibody (7/22, 31.8%) than in those with anti‐topoisomerase I antibody (2/82, 2.4%) and in those with anticentromere antibody (8/137, 5.8%). Importantly, among seven patients with anti‐RNA polymerase III antibody and malignancy, three patients (42.9%) developed malignancy from 6 months before to 12 months after SSc onset. Thus, malignancy complication in Japanese SSc patients with anti‐RNA polymerase III antibody is as high as that in other races, suggesting that SSc patients with anti‐RNA polymerase III antibody share the same pathological process among different ethnic groups.     

Clinically amyopathic dermatomyositis with rapidly progressive interstitial pneumonia: The relation between the disease activity and the serum interleukin‐6 level

A 67‐year‐old woman exhibited chilblain‐like erythema on the bilateral auricles, erythema with hyperkeratosis on the sides of fingers and the bilateral elbows, and erythema on the extensor/flexor sides of the finger joints and the iliac regions. Fine crackles were audible on the dorsal side of the bilateral lower lung regions. We diagnosed clinically amyopathic dermatomyositis accompanied by rapidly progressive interstitial pneumonia. While we initiated immunosuppressive therapy, the respiratory failure showed no responses. We performed endotoxin adsorption therapy using polymyxin B‐immobilized fiber column direct hemoperfusion (PMX‐DHP). After the first PMX‐DHP session, the respiratory condition was slightly improved but it soon deteriorated. At the second PMX‐DHP session, subcutaneous and mediastinal emphysema was noted. The patient died, showing no improvement in the respiratory condition. We measured the levels of serum interleukin (IL)‐6, tumor necrosis factor (TNF)‐α and IL‐10 before and after PMX‐DHP and compared them with previously reported levels. The levels of TNF‐α and IL‐10 showed no or slight changes, but those of IL‐6 at the beginning of the treatment were higher compared with previous reports. After the first PMX‐DHP, the IL‐6 level decreased with slight improvement in respiratory condition. However, after the second PMX‐DHP, it increased markedly with exacerbation of the respiratory failure. The serum IL‐6 level is considered to be effective for the evaluation of the disease activity and prognosis under the treatment by PMX‐DHP.     

Interferon‐γ induces upregulation and activation of the interleukin‐31 receptor in human dermal microvascular endothelial cells

Abstract: Interleukin‐31 (IL‐31), a recently discovered cytokine derived from T helper cells, is involved in chronic dermatitis and pruritus. This study demonstrates for the first time that the IL‐31 receptor complex for IL‐31 is substantially upregulated in human dermal microvascular endothelial cells after stimulation with interferon‐γ (IFN‐γ). Activation of the IL‐31 receptor complex results in the induction of the intracellular ERK1/2 signaling pathway and downregulation of IFN‐γ‐induced monokine induced by IFN‐γ expression. Inhibitor studies revealed that the IFN‐γ‐induced IL‐31RA upregulation is processed via JNK and PI3 kinase activation. In sum, our study points toward an interaction between the T_H1‐derived cytokine IFN‐γ and the T_H2‐derived cytokine IL‐31 on endothelial cells.     

No association of atherosclerosis with digital ulcers in Japanese patients with systemic sclerosis: Evaluation of carotid intima‐media thickness and plaque characteristics

Patients with systemic sclerosis (SSc) usually develop Raynaud's phenomenon, persistent digital ischemia and sometimes develop digital ulcers (DU). Several studies have reported an association of carotid artery atherosclerosis with SSc by evaluating carotid intima‐media thickness (IMT) in SSc patients. However, none of those studies analyzed the association between DU and carotid artery atherosclerosis in SSc patients. We examined the association of carotid artery atherosclerosis with digital ulcers by comparing SSc patients with (n = 48, 29.5%) and without (n = 206, 70.5%) DU. The demographic and clinical features of the SSc patients showed that young age, male sex, anti‐topoisomerase I antibody positivity, severe skin sclerosis, interstitial lung disease complication and cardiac involvements were significantly prevalent in patients with DU. In addition, diffuse cutaneous type, anti‐RNA polymerase III antibody positivity and severe skin sclerosis are more frequent in SSc patients with DU at the extensor surface of joints than SSc patients with DU at the digital tip. There were no differences in serum lipid level, carotid IMT or plaque score between SSc patients with and without DU, suggesting that atherosclerotic changes are not primarily involved in the development of DU.