A three-year follow-up study of the development of joint contractures in 131 patients with systemic sclerosis.

OBJECTIVES: To analyse the correlation between the number of joint-contractures and other major clinical findings in a follow-up study of 131 patients with systemic sclerosis (SSc). METHODS: The range of motion of joints (ROM), HAQ-DI, and the major clinical characteristics were assessed. RESULTS: A high frequency of contractures (ROM<75% of the normal) were present at baseline in small joints of the hand (82%), wrists (75%), and shoulders (50%). ROM of the dominant side hand was significantly more decreased compared to the non-dominant side. The number of the upper extremity contractures correlated positively with ESR (p<0.01), CRP (p<0.01), HAQ-DI (p<0.01), and negatively with forced vital capacity (FVC) (p<0.05). The number of contractures was not significantly different in cases with early (≤ 4 years) and late disease duration in both the limited and diffuse subgroups. During the three-year follow-up period, an increase in the number of joint contractures (ROM<75%) was associated with an increase of ESR, modified Rodnan's skin score, and the European Scleroderma Study Group Activity Index by multiple linear regression analysis. Univariate analysis over a six-year period demonstrated poor outcome in patients with more than ten contractures, or more than four contractures of unilateral hand-joints. CONCLUSIONS: Contractures predominantly develop during the early years following disease onset in both SSc subgroups. Inflammation and skin-involvement are significant contributing factors for the development of contractures. The dominant hand may be more pronouncedly impaired compared to the non-dominant side. A high number of joint-contractures might be an unfavourable prognostic factor in SSc.

Follow-up of serum KL-6 lung fibrosis biomarker levels in 173 patients with systemic sclerosis.

OBJECTIVES: A single-centre retrospective longitudinal study to investigate the predictive value of KL-6 serum levels for the outcome of interstitial lung fibrosis in a large systemic sclerosis (SSc) patient cohort. METHODS: ELISA tests for the mucin like glycoprotein KL-6 were performed in sera of 173 SSc patients. The clinical and laboratory data were evaluated by a standardised protocol of chest x-ray, lung function tests, echocardiography and high-resolution computed tomography. 158 patients were 29 ± 22 months later reinvestigated, 9 patients (2 lcSSc, 7 dcSSc) died from SSc-related causes, and 6 patients were lost to follow-up. RESULTS: Serum titer of KL-6 was negatively correlated with lung function parameters, independent of the time of investigation. There was a significantly higher probability of death among patients with high level of baseline KL-6. There was no statistically significant difference in the deterioration and improvement rates between groups with normal and elevated KL-6 level at study entry, even in patients in early phase of disease (disease duration <3 years). Serum levels of KL-6 significantly decreased in patients receiving cyclophosphamide treatment in spite of the fact that the spirometry results (FVC and DLCO) did not show a significant change. CONCLUSIONS: KL-6 can be used as a lung fibrosis severity marker, but its role as a marker for disease activity is questionable. Furthermore, following cyclophosphamide treatment serum KL-6 levels may decrease independently of the lung function parameters.

Overlap of coronary disease and pulmonary arterial hypertension in systemic sclerosis.

OBJECTIVES: Pulmonary arterial hypertension (PAH) is a common complication of systemic sclerosis (SSc). Symptoms of coronary artery disease (CAD) and PAH are closely related and cardiac catheterisation is needed to confirm their diagnosis. The aim of the present work was to investigate of the extent of overlap between CAD and PAH in patients with SSc. METHODS: Based on non-invasive investigations, 20 patients out of 120 were suspected to have PAH ("suspected PAH" group). Another 10 patients showed signs of coronary disease ("suspected CAD" Group). In these 30 patients, right heart catheterisation and coronary angiography were performed, and the coronary flow reserve (CFR) was assessed by thermodilution technique. RESULTS: In the "suspected PAH" and the "suspected CAD" groups, PAH was found in 12/20 and 2/10 cases, and coronary artery stenosis in 9/20 and 6/10 cases, respectively. Severely reduced CFR was revealed in 7/20 and 3/10 cases, respectively. CONCLUSIONS: PAH, CAD and reduced CFR all show a considerable overlap in symptomatic patients with SSc. The current non-invasive investigations are neither sensitive nor specific enough to make an appropriate distinction between these different disease manifestations. A more invasive approach, such as coronary angiography at the initial catheterisation, is required to properly characterise and treat the different forms of cardiac involvement in SSc.

Cross-cultural adaptation of the disabilities of the arm, shoulder, and hand (DASH) questionnaire into Hungarian and investigation of its validity in patients with systemic sclerosis.

OBJECTIVE: To adapt and validate the Hungarian version of the DASH and the shorter QuickDASH Outcome Measures and to establish their validity in patients with systemic sclerosis (SSc). METHODS: The Hungarian adaptation of the questionnaires was performed using forward/backward translations, expert and lay reviews. 128 patients completed the DASH, the Health Assessment Questionnaire (HAQ-DI), the Modified HAQ-DI for patients with SSc (SSc-HAQ), and the Short Form Health Survey (SF-36) questionnaire. 76 patients participated in a 12-month follow-up examination. Sensitivity to change was estimated using the standardized response mean (SRM). RESULTS: Cronbach alpha in the DASH sections were between 0.94-0.97. The intraclass correlation coefficient for the test-retest reliability of DASH was 0.89. DASH scores showed a correlation with both SSc-HAQ and the physical dimensions of the SF-36 (Spearman's rho: 0.89, -0.77 and -0.42, respectively). The SRM of DASH was 0.64 among the scleroderma patients with worsening HAQ-DI status. CONCLUSIONS: The Hungarian version of the DASH and QuickDASH demonstrated equivalent reproducibility, internal consistency and validity to the originals. The strong correlations of the DASH and QuickDASH with the HAQ-DI, and with the physical dimensions of the SF-36 show that the disability of the patient with SSc is predominantly caused by the functional impairment of the upper limb. Because both questionnaires were valuable tools for measuring upper extremity function and joint damage in SSc patients, we recommend the shorter and simpler QuickDASH for everyday clinical use.

Comprehensive investigation of novel serum markers of pulmonary fibrosis associated with systemic sclerosis and dermato/polymyositis.

OBJECTIVE: To investigate the association between serum levels and clinical signs of lung fibrosis in patients with systemic sclerosis and inflammatory myopathies. METHODS: ELISA tests for a mucin-like glycoprotein KL-6, von Willebrandt factor (vWF), soluble E-selectin (sES) and surfactant protein D (SP-D) were performed in sera of 104 patients with systemic sclerosis, 31 patients with poly/dermatomyositis) and 24 patients with Raynaud's phenomenon as controls. The clinical and laboratory data were evaluated by a simple standard protocol including chest x-ray, lung function tests, echocardiography and, in selected cases, high resolution computer tomography (HRCT). Clinically significant pulmonary fibrosis (PF) defined as a simultaneous presence of radiological sign of pulmonary fibrosis and restrictive impairment. Severe PF was established if HRCT scans showed diffuse interstitial lung disease with low diffusing capacity. End stage PF was determined as severe PF with very low diffusing capacity. RESULTS: Patients with pulmonary fibrosis on chest x-ray showed significantly elevated serum levels of KL-6, SP-D and vWF. Inverse correlation was found between serum levels of KL-6/SP-D and lung function parameters, such as DLCO% and FVC. With regard to HRCT findings, patients with elevated serum level of KL-6 showed significantly more frequently ground glass opacity, diffuse and honeycombing fibrosis than patients with normal level of KL-6. The sensitivity of KL-6 for PF in SSc is increased with the severity of PF (PF on chest x-ray < severe PF < end stage of PF). Lung fibrosis occurred more frequently in patients with simultaneously elevated KL-6 and sES compared to cases with a single positivity of either KL-6 or sES. CONCLUSION: KL-6, SP-D, vWF and ES are good surrogate factors of pulmonary fibrosis but can not replace conventional diagnostic procedures. However, these markers are suitable for the assessment of progression and severity of pulmonary fibrosis in systemic autoimmune disorders once the diagnosis is established.

Survival and causes of death in 366 Hungarian patients with systemic sclerosis.

OBJECTIVE: Survival analysis of a series of 366 consecutive patients with systemic sclerosis (SSc). METHODS: Clinical and laboratory data were evaluated from 1983 until 2005 using a standard protocol. The female/male ratio was 315/51. The mean (SD) age of the patients was 56.8 (12.2) years. The duration of disease was 12 (5-19) years with a median follow-up of 6 (3-12) years. RESULTS: Kaplan-Meier univariate analysis showed that renal, cardiac involvement, pigmentation disturbances, malabsorption, a forced vital capacity <50%, diffuse scleroderma, presence of early malignancy, anaemia, and increased erythrocyte sedimentation rate (ESR) were signs of unfavourable prognosis, whereas anti-centromere antibodies were indicators of a good survival. In the multivariate Cox proportional hazards model the presence of diffuse scleroderma, renal involvement, coexistence of a malignant disease, and increased ESR were poor independent prognostic signs. Elderly age at the onset of disease also caused an unfavourable outcome. A total of 86 SSc-related deaths were recorded during the follow-up. Of them, 65% were attributed to cardiorespiratory manifestation of disease. Tumour associated early death was found in 12 cases (14%). CONCLUSIONS: In addition to the well-known factors influencing the outcome (diffuse subset, internal organ involvements, and inflammatory signs), the coexistence of scleroderma with a malignancy also causes a poor outcome.

Exposure to solvents in female patients with scleroderma.

The role of exposure to solvents was investigated in female patients with connective tissue disease and Raynaud's phenomenon using a questionnaire. Sixteen out of the 63 patients with systemic sclerosis had been exposed to solvents. A borderline significance was demonstrated compared to matched female controls (P < 0.05). Fourteen out of the 66 patients with undifferentiated connective tissue disease, 18/86 of patients with Raynaud's phenomenon, 6/45 with systemic lupus erythematosus, 1/16 with dermatopolymyositis, 1/15 with rheumatoid arthritis and 0/13 with primary Sjögren's syndrome had been exposed to solvents. None of these groups of patients showed a statistical significance compared to matched controls. Our present findings indicate that, at least in certain areas of the world, exposure to solvents may be a provoking factor in female scleroderma, but it must be emphasised that only a borderline significance was found between the scleroderma patients and controls. A large multicenter study seems to be required to clarify the importance of solvents as provoking factors of scleroderma. Furthermore, exposure to solvents does not seem to be a provoking factor among females for the other connective tissue diseases.

Elevated monocytic IL-12 and TNF-alpha production in Wegener's granulomatosis is normalized by cyclophosphamide and corticosteroid therapy.

Wegener's granulomatosis (WG) is characterized by a predominance of the type 1 T-helper cell (Th1) response. We have studied monocytic cytokine expression in untreated patients and in patients who did not respond to prior methotrexate or trimethoprim-sulphamethoxazole therapy, i.e. patients with active disease. Intracytoplasmic IL-12 and TNF-alpha expression was significantly increased in WG compared with healthy controls. IL-8 expression was not increased. Two and 12 weeks of daily standard oral cyclophosphamide and corticosteroid (CYC + GC) treatment induced a stable remission of the disease. Elevated IL-12 and TNF-alpha expression of monocytes was normalized. The active metabolite of CYC was shown to down-regulate IL-12 mRNA in vitro. Monocytic cytokines, especially IL-12, may have a role in the early determination and skewing of the immunoregulatory response towards a Th1 profile. It appears that CYC + GC exerts its effect by normalizing the Th1-driving cytokine pattern, and CYC may maintain this mode of action. Normalization of the skewed cytokine pattern may be a prerequisite and an indicator of inducing a remission in WG.

Subclinical pulmonary involvement assessed by bronchoalveolar lavage in patients with early undifferentiated connective tissue disease.

OBJECTIVE: To assess the presence of neutrophil and lymphocyte fibrosing alveolitis by bronchoalveolar lavage in patients with early undifferentiated connective tissue disease (EUCTD) and systemic sclerosis (SSc). METHOD: Bronchoalveolar lavage (BAL) was performed in 13 patients with EUCTD who exhibited signs of lung involvement by non-invasive methods including lung function tests and high resolution computed tomography. The mean age of cases was 48.1 +/- 6.6, and the mean disease duration was 1.8 +/- 0.8 years. Differential cell counts of BAL were evaluated. Eleven patients with systemic sclerosis and 5 healthy control subjects were also investigated. RESULTS: Eleven of the 13 EUCTD and 10 of the 11 SSc patients showed an elevated total cell number (above the median cell/ml of control + 2 SD) in the BAL fluid. In patients with EUCTD, the lymphocyte count was elevated in 6, and the polymorphonuclear neutrophil count in 2 patients. One of the patients with EUCTD had simultaneously elevated lymphocyte and neutrophil granulocyte counts. In the SSc group, 6 patients had an elevated lymphocyte and 6 an increased neutrophil count. Three of these cases had both increased neutrophil and elevated lymphocyte counts, simultaneously. CONCLUSION: Subclinical, predominantly lymphocyte alveolitis can be present in patients with EUCTD. Patients with SSc tend to exhibit neutrophil alveolitis.

Alveolitis may persist during treatment that sufficiently controls muscle inflammation in myositis.

Eight patients with dermato- and polymyositis (DM/PM) and two further cases with sclerodermamyositis overlap syndrome were investigated. These patients showed signs of lung manifestation by noninvasive methods. Bronchoalveolar lavage (BAL) was performed to detect alveolitis. Four of the eight DM/PM patients showed elevated neutrophil counts. All of these and one additional case had increased lymphocyte counts. Five of the DM/PM patients showed higher total cell numbers than five healthy controls. One of the cases with scleroderma-myositis overlap syndrome also developed lymphocyte alveolitis. We conclude that signs of alveolitis are often present in patients with myositis, even though the myositis was adequately controlled by corticosteroid therapy and, in four cases, with corticosteroid plus azathioprine. The need for further follow-up studies to determine the effectiveness of intensified corticosteroid/cytostatic treatment in these patients is emphasised.

Opsonization of apoptotic neutrophils by anti-neutrophil cytoplasmic antibodies (ANCA) leads to enhanced uptake by macrophages and increased release of tumour necrosis factor-alpha (TNF-alpha).

Since proteinase 3 (PR3)-ANCA interact with PR3 on the surface of apoptotic polymorphonuclear neutrophils (PMN) and ingestion of apoptotic PMN is known to modulate macrophage inflammatory reactions, we raised the question whether PR3-ANCA-opsonized apoptotic PMN influence the uptake by macrophages and their state of activation. We therefore analysed the effects of PR3-ANCA-opsonized apoptotic PMN on the uptake process by enzymatic assay. We further investigated the production of TNF-alpha, IL-10, IL-12 and the secretion of lipid inflammatory mediators (TxB2, leukotriene B4 (LTB4) and prostaglandin E2 (PGE2)) by human monocyte-derived macrophages using FACS and ELISA methods. We show that PMN-opsonization by PR3-ANCA substantially enhances phagocytosis by macrophages and thereby triggers the production of TNF-alpha and TxB2. These in vitro findings indicate that PR3-ANCA opsonization of apoptotic PMN might be an important mechanism in the pathogenesis of Wegener's granulomatosis (WG), prompting macrophages to produce proinflammatory mediators. These mediators, mainly TNF-alpha, might prime further PMN leading to perpetuation of the known priming-dependent mechanisms of ANCA action.

Investigation of the alveolar macrophages and T lymphocytes in 15 patients with systemic sclerosis.

The cell distribution and function of alveolar macrophages and T lymphocytes were investigated in the bronchoalveolar lavage (BAL) of 15 patients with systemic sclerosis (SSc). In alveolar macrophages, both spontaneous and PMA-stimulated TNF-alpha production were increased in SSc. PMA-induced IL-6 production was also elevated. Spontaneous IL-6 excretion of scleroderma alveolar macrophages was similar to the controls. Yeast and C3b-coated yeast (opsonised yeast) phagocytosis, chemotaxis and Fc receptor activity of alveolar macrophages were normal. The proportion of CD3, CD4 and CD8 T-lymphocyte subsets in the BAL was similar to the control values. The lymphocyte blast transformation index of the non-adherent cells deriving from the BAL fluid was markedly decreased.

Production and flow cytometric application of a monoclonal anti-glucocorticoid receptor antibody.

Detection and monitoring the expression and level of intracellular glucocorticoid receptor (GCR) is necessary in many clinical and experimental situations. Binding of radioactive steroids (3H dexamethasone) to the cytosolic fractions of cells has been recently used. However, it is an expensive, time-consuming technique difficult to use in routine diagnostics. In this article we describe a novel, simple method for GCR detection, using a FITC-conjugated anti-GCR monoclonal antibody (mAb) for flow cytometric measurements in permeabilized cells. The monoclonal antibody was raised against a conserved sequence (150-176 amino acids) of the regulatory part of the receptor. Synthetic peptide (called APTEK-26) fragment of the receptor conjugated to different carriers (TG, BSA) was used for immunization and screening of the hybridomas. The a-GCR 8E9, 3C8 and 5E4 clones (IgG1) were further characterized by immunoserological methods for their reactivity against overlapping synthetic peptide fragments of the receptor and by Western blot technique on cytosolic fraction of HEP G2 cells (containing the GCR). Furthermore the mAbs could be used for the FACS based detection of GCR, despite its low number of antigen structure within the cells. Solving the problem of nonspecific binding of the secondary antibodies we used our high affinity IgG1 a-GCR mAbs directly labeled with the fluorescent dye FITC. The fluorescent labeling of the GCRs in HEP G2 cell line and human peripheral blood mononuclear cells (PBMC) were demonstrated by flow cytometric analysis after fixation with 4% paraformaldehyde and permeabilization with saponin. Competition with molar excess of unlabelled antibodies and with the GCR peptide fragment confirmed the specific binding of the 8E9 and 5E4 mAbs to the GCRs. Monitoring the GCR level by flow cytometry would be useful in clinical diagnostics, e.g., in steroid-treated patients and in steroid-resistant states.