Diagnostic Accuracy of Anticarbamylated Protein Antibodies in Established Rheumatoid Arthritis: A Monocentric Cross-Sectional Study.

OBJECTIVE: To evaluate the diagnostic accuracy of anticarbamylated protein antibodies (CarP), alone and in combination with traditional biomarkers (rheumatoid factor [RF] and anticitrullinated peptide antibodies [ACPA]), in established rheumatoid arthritis (RA). METHODS: A commercially available enzyme-linked immunosorbent assay (ELISA) kit was used to assess CarP concentrations in serum samples of 200 established RA and 206 controls (115 healthy donors and 55 patients with other rheumatic diseases). Main outcome measures were sensitivity, specificity, and area under the curve (AUC; 95% confidence interval [CI]). Difference in accuracy was evaluated by comparison of the respective AUCs. RESULTS: A serum CarP cut-off of 1.47 ng/ml or more differentiated patients with RA from controls with 30% sensitivity, 97.1% specificity, and good accuracy (AUC[95%CI] = 0.83[0.79-0.86], P < 0.0001). However, it showed moderate diagnostic accuracy in seronegative RA patients: sensitivity 17.9%, specificity 96.9%, and AUC (95% CI) = 0.69 (0.63-0.75). The diagnostic accuracy of CarP_ACPA and CarP_RF combinations was significantly superior to that of ACPA and RF alone (P < 0.0001 and P = 0.015, respectively), but not to that of ACPA_RF combination (P = 0.089) In addition, the CarP_ACPA_RF combination did not improve the diagnostic accuracy of the ACPA_RF combination (AUC mean difference [95% CI] = 0.006 [-0.001 to 0.015], P = 0.10). The number of positive autoantibodies (0, 1, 2, or 3) was not significantly associated with moderate-severe disease (Disease Activity Score-28 [DAS-28] > 3.2) in adjusted multiple regression analysis. CONCLUSION: CarP has good diagnostic accuracy in established RA but not in seronegative RA. The addition of CarP to ACPA and RF alone or in combination does not significantly enhance the diagnostic accuracy of ACPA_RF combination.

Peripheral blood CD8+ T-cell profiles in patients with psoriatic arthritis: a cross-sectional case-control study.

OBJECTIVE: While CD4+ T-cells are traditionally regarded as the main pathogenic T-cell subpopulation in psoriatic arthritis (PsA), the role of circulating CD8+ T-cells remains poorly characterized. We evaluated the differential representation of CD8+ T-cell subpopulations in peripheral blood (PB) of PsA patients. PATIENTS AND METHODS: CD8+IL-17+, CD8+IFNγ+ and CD8+IL-17-IL-22+ T-cells were evaluated by flow-cytometry in 25 consecutive PsA patients, 7 rheumatoid arthritis (RA) patients, 16 patients with psoriasis, and 26 healthy controls (HC). RESULTS: We observed a significant expansion of circulating IFN-γ producing CD8+ T-cells in PsA when compared to psoriasis [21.2 (6.9-55.8)% vs. 3.8 (0.7-11.8)%, p < 0.0001] and HC samples [21.2 (6.9-55.8)% vs. 4.05 (0.44-19.8)%, p < 0.0001]. A frequency of circulating IFN-γ producing CD8+T-cells ≥ 9% distinguished PsA from psoriasis patients with a specificity of 84% and a sensitivity of 87.5% [AUC = 0.9 (0.80-0.99), p < 0.0001]. In addition, we found a significant expansion of circulating IL-17 producing CD8+ T cells in RA patients when compared to PsA, psoriasis and HC samples. By contrast, there were no significant between-group differences in the prevalence of circulating IL-22 producing CD8+ T-cells. In PsA patients there was a significant correlation between number of swollen joints and frequency of circulating IFN-γ producing CD8+ T-cells, and between extent and severity of psoriasis and frequency of circulating IL-17 producing CD8+ T-cells. CONCLUSIONS: Circulating IFNγ-producing CD8+ T-cells are raised in PsA when compared to psoriasis, suggesting a potential pathogenetic involvement of CD8+ T-cells and IFNγ production in chronic joint inflammation and damage. The significant enrichment of circulating IL-17 producing CD8+ T-cells in RA when compared to PsA warrants functional characterization and confirmation in larger studies. We found no significant enrichment of circulating IL-22 producing CD8+ T-cells in PsA, RA and psoriasis.

Identification of a HERV-K env surface peptide highly recognized in Rheumatoid Arthritis (RA) patients: a cross-sectional case-control study.

Endogenous retroviruses (HERV) are believed to be pathogenic in several autoimmune diseases. Among them, HERV-K viruses have been reported recently to be involved in the pathogenesis of rheumatoid arthritis (RA). In this study we have explored the role of humoral immune response against HERV-K as a potential pathogenetic mechanism in RA. Four different peptides from the extracellular portion of the env protein of HERV-K (env-su19-37 , env-su109-126 , env-su164-186 , env-su209-226 ) were selected by bioinformatic analysis on the basis of their putative immunogenicity. Indirect enzyme-linked immunosorbent assay (ELISA) was then carried out to quantify antibodies against those peptides on blood samples of 70 consecutive RA patients and 71 healthy controls (HC). Differences between the two groups were analysed using the Mann-Whitney test. Potential correlations between RA laboratory, clinical descriptors and immunoglobulin (Ig)G levels were explored by bivariate regression analysis. Serum autoantibodies against one of four tested peptides of HERV-K (env-su19-37 ) were significantly higher in RA than in HC (19 versus 3%, P = 0·0025). Subgroup analysis showed no association between anti-HERV-K peptide humoral response and clinical, serological and clinimetric RA disease descriptors. Serum from RA patients in our series reacted significantly against HERV-K env-su19-37 peptide in comparison to the general population suggesting a role for the HERV-K- related, secondary antigenic-driven immune response in the pathogenesis of RA. Further studies are needed to confirm these results and to explore the role of this HERV-K surface peptide as a potential therapeutic target.

Mycobacterium tuberculosis lipoarabinomannan antibodies are associated to rheumatoid arthritis in Sardinian patients.

Little is known regarding the environmental factors at play in igniting rheumatoid arthritis (RA) autoimmunity, although an association between Mycobacteria and RA has been documented. This pilot study focused on examining a possible involvement of Mycobacterium tuberculosis (MTB) and Mycobacterium avium ss. paratuberculosis (MAP) in RA. We measured out the serum levels of IgG antibody against different mycobacterial antigens in Sardinian patients and controls, by an enzyme-linked immunosorbent assay. The population study was composed of 61 RA patients under different therapies and 52 healthy controls, whereas the antigens tested were MTB lipoarabinomannan (ManLAM), MAP heath shock protein 70, and MAP protein tyrosine phosphatase. The frequencies of anti-ManLAM antibodies were higher in the RA group (23 %) compared to the healthy controls (5.7 %) (AUC = 0.7; p < 0.0001), whereas serum reactivity to MAP antigens was not observed. ManLAM antigen was also detected in the plasma of three RA patients (which were anti-ManLAM antibody positive) by Western blot analysis using anti-Man-LAM monoclonal antibodies. The data produced corroborate the hypothesis of a potential association between MTB ManLAM and RA disease, but so far, further studies are necessary to understand its role in RA pathogenesis.

Genetics of Behçet's disease in Sardinia: two distinct extended HLA haplotypes harbour the B*51 allele in the normal population and in patients.

OBJECTIVES: To define the contribution of HLA genes and extended HLA haplotypes to the susceptibility to Behçet's disease (BD) in Sardinia. METHODS: Forty-five unrelated Sardinian patients with BD, diagnosed according to the ISG criteria, 45 HLA-B*51 positive and 185 unselected healthy controls were enrolled in the study. DNA samples were typed for HLA class I and class II alleles and genotyped for microsatellites (MICA-TM) and single-nucleotide polymorphisms (rs1264457 HLA-E; rs2281820 motilin; rs1799724 at -857, rs361525 at -238 TNF-alpha) spanning the HLA region. RESULTS: HLA-B*5101 was confirmed as conferring susceptibility to BD (pc=0.0042; OR=4.4; 95% CI=2.0 to 9.6). It is noteworthy that in Sardinia this allele was found more frequently within a haplotype (HLA-A2; -Cw2; -B*5101; -DRB1*11; -DQA1*05; - DQB1*03) that reached its highest frequency in patients with BD. Linkage disequilibrium analysis showed the existence of an additional B*51 haplotype (HLA-A2; -Cw2; -B*5101; -DRB1*04; -DQA1*03; -DQB1*03) not associated with susceptibility to the disease. CONCLUSIONS: In Sardinia, the BD-associated HLA-B*5101 allele is inherited as part of two distinctive haplotypes differently distributed in patients and controls. These findings can be interpreted as suggestive of the presence of additional genes within the MHC region conferring susceptibility to BD. The hypothesis that an environmental pressure could have contributed to the preservation of the BD-associated HLA haplotype in Sardinia is also discussed.

Antioxidant effect of Iloprost: current knowledge and therapeutic implications for systemic sclerosis.

Many lines of independent research have pointed out the role of oxidative stress as a fascinating pathogenic link between the three main hallmarks of systemic sclerosis (SSc), viz. humoral and cellular immunity activation, microvascular damage and widespread tissue fibrosis. Therefore counteracting oxidative stress may have a favourable impact on clinical features and progression of disease. It is becoming clearer that Iloprost, a synthetic stable analogue of prostacycline, currently employed in the treatment of SSc vascular features, also possess anti-oxidative properties beside its prostaglandin-like vasodilatory and antiaggregant effects. This brief review is aimed to discuss available clinical evidences supporting Iloprost antioxidant action and focus on putative molecular pathways underlying it.

Hemarthrosis as acute presentation of acquired hemophilia in a patient with systemic lupus erythematosus: successful treatment and long-lasting remission.

Hemorrhagic events due to production of antibodies directed against coagulation factors are rarely observed in systemic lupus erythematosus (SLE). We report the case of a patient with clinically quiescent SLE who developed factor VIII inhibitor in acquired hemophilia presenting as hemarthrosis. Initial treatment with FVII, FVIII and FIX plasma concentrate, metilprednisolone and immunoglobulins i.v. were started but new hemorrhagic manifestation occurred. Plasma exchange was also administered, but it was discontinued early due to partial efficacy. In addition, pulse cyclophosphamide 0.5 g/m(2) was started. Eight weeks later, FVIII and FIX activity returned within normal ranges, FVIII and FIX inhibitors decreased significantly and hemorrhagic manifestations disappeared. The rare occurrence of acquired hemophilia due to the presence of anti-factor VIII antibodies associated to SLE, which was reviewed, might explain the lack of therapeutic guide-lines; indeed therapeutic options are available but the outcome in each single patient is not predictable.

Association of MICA alleles with psoriatic arthritis and its clinical forms. A multicenter Italian study.

OBJECTIVE: Analysis of the association between psoriatic arthritis (PsA) clinical forms and MICA gene transmembrane polymorphisms. METHODS: Patients were classified as having peripheral asymmetric oligoarthritis (AO), peripheral symmetric poly-arthritis (PA) and spondylitis (SP), or disease combinations (PA/SP, OA/SP). Two hundred and twenty-six patients with PsA were typed for MICA exon 5 microsatellite (TM) by heteroduplex analysis and compared with 225 normal controls. RESULTS: MICA-TM microsatellite typing revealed that, among the different clinical forms of PsA, only the combined PA/SP subset shows a significant positive association with MICA-A9 and a lower frequency of MICA-A4, A5 genotype in PsA patients with a decrease, only in the PA/SP cohort, of all MICA-A5 combinations except MICA-A5, -A9. CONCLUSION: These results suggest a role for genes within the HLA region in the pathogenesis of PsA, and reinforce the idea that the different forms of PsA may have heterogeneous genetic basis.

Effect of rituximab on clinical and laboratory features of antiphospholipid syndrome: a case report and a review of literature.

Antiphospholipid syndrome (APS) is an autoimmune disorder characterized by a hypercoagulable state related to persistently elevated levels of antiphospholipid antibodies (aPL). Current treatment for APS is only partially effective and new therapies are strongly needed. We report on a case of a 50 years old man with APS who suffered from recurrent thromboembolic episodes despite conventional anticoagulant treatment. Eight years after the first thrombotic manifestation he was diagnosed with a large B cell non-Hodgkin lymphoma. Treatment with CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone) plus rituximab was started with partial clinical remission of lymphoma and normalization of aPL levels with a three years follow-up period free of thrombotic episodes.A review of the literature revealed that only 12 case reports on the use of rituximab in patients with primary, secondary and catastrophic APS have been published. Current knowledge clearly suggests the need for clinical trials to evaluate the effect of rituximab in the treatment of resistant APS.

Iloprost therapy acutely decreases oxidative stress in patients affected by systemic sclerosis.

BACKGROUND: Oxidative stress has been considered a leading factor in the pathogenesis of systemic sclerosis (SSc). Consistently with this hypothesis the determination of urinary isoprostanes, a reliable method for evaluation of oxidative stress, has recently showed increased levels of isoprostanes in SSc patients. Data about the effect on oxidative stress of accepted therapies for SSc such as iloprost therapy are lacking. OBJECTIVE: The aim of this prospective study was to verify whether iloprost therapy in patients with SSc acutely reduces oxidative stress assessed by determination of 8-Iso PGF2alpha urinary levels. METHODS: urine samples were obtained before and after a five-day cycle of iloprost infusion and urinary 8-Iso PGF2alpha levels were determined using a commercially available enzyme immunoassay. RESULTS: Consistent with previous reports, we found an increased level of oxidative stress in SSc patients with respect to healthy controls. Basal urinary 8-iso PGF2alpha levels in SSc patients were significantly higher than those in healthy controls [2002(1122-3575) pg/mg creatinine vs. 334(225.7-441) pg/mg creatinine, p<0.001]. Moreover, as expected, urinary 8-iso PGF2alpha levels after iloprost therapy were significantly lower than basal levels [1277.5 pg/mg creatinine (742.7-2017.3) vs. 2002 pg/mg creatinine (1122-3575), p=0.001] but persisted significantly elevated respect to the levels of healthy controls (p<0.001). The effect of iloprost on oxidative stress appeared significant in patients with early and limited form of disease. CONCLUSIONS: This prospective open-label explorative study suggests that standard course of iloprost therapy may acutely reduce oxidative stress in SSc patients. This effect appears to be more consistent in the early phases and in the limited subset of disease. Further larger trials are needed to confirm our results and to explain the pathway of such reduction, its clinical significance and potential therapeutic implications.

Genetics of psoriasis and psoriatic arthritis.

Psoriasis and psoriatic arthritis are linked diseases characterised by (distinct ?) immune-mediated pathogenetic mechanisms and by a genetic background interacting with environmental factors. Some candidate susceptibility genes have been studied extensively; they include HLA genes, genes within the HLA region and genes outside the HLA region; among them corneodesmosin and other genes of PSORS1 region, MICA and TNF-a polymorphisms. The main findings in the literature are discussed.

HLA-DPB1 alleles association of anticardiolipin and anti-beta2GPI antibodies in a large series of European patients with systemic lupus erythematosus.

Our objective was to determine the HLA-DPB1 allele associations of anticardiolipin (aCL) and anti-beta2GPI (a(beta)2GPI) antibodies, and of clinical manifestations of the antiphospholipid syndrome (APS), in systemic lupus erythematosus (SLE). We studied 577 European patients with SLE. aCL and a(beta)2GPI antibodies were measured by ELISA. Molecular typing of HLA-DPB1 locus was performed by polymerase chain reaction-sequence specific oligonucleotide probe (PCR-SSOP) method. aCL showed positive association with -DPB1*1501 (P = 0.005, OR = 7.4), and -DPB1*2301 (P = 0.009, OR = 3.3). a(beta)2GPI showed positive association with -DPB1*0301 (P = 0.01, OR = 1.9), and -DPB1*1901 (P = 0.004, OR = 8.1). In addition, livedo reticularis was associated with -DPB1*1401, and Raynaud's phenomenon with -DPB1*2001. In conclusion, HLA-DPB1 locus may contribute to the genetic predisposition to develop antiphospholipid antibodies and clinical manifestations of the APS in patients with SLE.

Detection of early impairment of coronary flow reserve in patients with systemic sclerosis.

OBJECTIVE: To investigate whether coronary flow reserve (CFR), measured by a new non-invasive method, is impaired early in patients with systemic sclerosis (SSc) and whether CFR impairment correlates with clinical or functional measures, or both. METHODS: 27 patients with SSc without clinical evidence of ischaemic heart disease and 23 control group subjects matched for age and sex were studied. CFR was evaluated in the left anterior descending coronary artery (LAD) with a new non-invasive method: contrast (Levovist) enhanced transthoracic Doppler during adenosine infusion. The pulsed wave Doppler examination of blood flow velocity was recorded in the LAD at rest and after maximum vasodilatation by adenosine infusion. RESULTS: In patients with SSc, without clinical evidence of ischaemic heart disease, CFR was impaired (p=0.0001). 14/27 patients with SSc had severe reduction of the CFR (< or =2.5) compared with controls (p=0.002). A non-significant trend between mean CFR and the severity and duration of the disease was also seen. CONCLUSIONS: CFR is often reduced in patients with SSc, suggesting early preclinical cardiac involvement in SSc. This impairment in coronary microvasculature is detectable by a non-invasive echocardiographic method and in this study was more common in the diffuse form of SSc.

Increased level of HLA-B27 expression in ankylosing spondylitis patients compared with healthy HLA-B27-positive subjects: a possible further susceptibility factor for the development of disease.

OBJECTIVE: In B27 transgenic rats, susceptibility to the development of a spondyloarthropathy-like disease has been shown to correlate with the level of B27 transgene expression on lymphoid cells. The aim of this work was to study HLA-B27 molecule expression in peripheral blood mononuclear cells (PBMCs) from patients with ankylosing spondylitis (AS) and from normal controls (NC). METHODS: Twenty B27(+) AS patients and 16 B27(+) NC were studied. HLA-B27 whole molecules and free heavy chains (HCs) and total HLA class I molecules were evaluated at the surface of PBMCs by immunofluorescence and flow cytometry. B27 subtypes were defined with the PCR-SSP (polymerase chain reaction-sequence-specific primer) technique. Cellular activation was evaluated by the expression of CD69, CD25 molecules and interferon gamma (IFN-gamma) production. RESULTS: B27 expression was 55,536.3+/-18,961.0 MESF (molecules of equivalent soluble fluorochromes) units in AS and 25,936.0+/-12,117.5 MESF in NC (P=0.00009), total HLA class I expression was 448,840.2+/-136,293.8 MESF in AS and 533,494.4+/-232,931.1 MESF in NC (not significant), HC expression was 10,593.4+/-6,396.1 MESF in AS and 14,843.0+/-7,544.2 MESF in NC (not significant). The higher B27 expression in the SA group was not due to higher cell activation as it was not correlated with CD69 and CD25 expression in PBMCs or with the level of IFN-gamma. HLA-B27 expression did not correlate with indexes of disease status [Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), Bath Ankylosing Spondylitis Functional Index (BASFI) and Bath Ankylosing Spondylitis Metrology Index (BASMI)]. CONCLUSIONS: We found greater expression of HLA-B27 molecules in patients with AS than in healthy subjects. This phenomenon was not accompanied by general up-regulation of HLA class I molecules or by greater expression of classical T-cell activation markers. On this basis we propose that the higher expression of the HLA-B27 molecules is a further predisposing factor for the development of AS.

[The role of HLA-B27 molecules in the pathogenesis of ankylosing spondylitis]. / Il ruolo delle molecole HLA-B27 nella patogenesi della spondilite anchilosante.

Ankylosing Spondylitis (AS) is characterised by the strongest association with an HLA antigen ever described for any disease. It represents therefore the ideal model for the understanding of the link between immune-mediated diseases and the HLA system. The role of HLA-B27 in the pathogenesis of AS will be discussed focusing on the recently described higher expression of these molecules in patients with AS compared with healthy controls.

Sustained normalization of cerebral blood-flow after iloprost therapy in a patient with neuropsychiatric systemic lupus erythematosus.

We report the case of a 30-year-old caucasian woman affected by SLE who developed neurological symptoms (prosopagnosia and visual-spatial agnosia) after nine years of disease. Brain MRI showed no abnormalities while a brain SPECT scan showed diffuse uptake defects and hypoperfusion areas in the right and left frontal-parietal regions. At that time the patient was on hydroxychloroquine (400 mg/day) and oral prednisolone (0.5 mg/kg/day) as maintenance therapy. One year later the patient showed worsening of Raynaud's phenomenon with digital dystrophic lesions and was therefore treated with an intravenous infusion of Iloprost (1.5 ng/kg/min per 6h/day for 10 days consecutively), while baseline treatment remained unchanged. One month later the patient showed a dramatic improvement in her cognitive function and subsequent SPECT scans showed the gradual disappearance of perfusion abnormalities. This first report of Iloprost treatment in CNS lupus suggests the potential therapeutic usefulness of this drug in patients with SLE and functional CNS involvement.