Identification of a new putative functional IL18 gene variant through an association study in systemic lupus erythematosus.

Interleukin-18 (IL-18) is a proinflammatory cytokine that plays an important role in chronic inflammation and autoimmune disorders. In this study, we aimed to determine the potential role of the IL18 gene in SLE. To define the genetic association of the IL18 and SLE, we have genotyped nine SNPs in an independent set of Spanish cases and controls. The IL18 polymorphisms were genotyped by PCR, using a predeveloped TaqMan allele discrimination assay. Two SNPs were still significant after fine mapping of the IL18 gene. The SNP (rs360719) surviving correction for multiple tests was genotyped in two replication cohorts from Italy and Argentina. After the analysis, a significance with rs360719 C-allele remained across the sets and after the meta-analysis (Pooled OR = 1.37, 95% CI 1.21-1.54, combined P = 3.8E-07, Pc = 1.16E-06). Quantitative real-time PCR was performed to assess IL18 mRNA expression in PBMC from subjects with different IL18 rs360719 genotypes. We tested the effect of the IL18 rs360719 polymorphism on the transcription of IL18 by electrophoretic mobility shift assay and western blot. We found a significant increase in the relative expression of IL18 mRNA in individuals carrying the rs360719 C-risk allele; in addition we show that the polymorphism creates a binding site for the transcriptional factor OCT-1. These findings suggest that the novel IL18 rs360719 variant may play an important role in determining the susceptibility to SLE and it could be a key factor in the expression of the IL18 gene.

Identification of HAVCR1 gene haplotypes associated with mRNA expression levels and susceptibility to autoimmune diseases.

Human HAVCR1 gene maps on 5q33.2, a region linked with susceptibility to allergic and autoimmune diseases. The aims of the present study were to define the haplotypes of HAVCR1 gene taking into account both HapMap Project SNP haplotypes and exon 4 variants, to investigate a possible relationship between these haplotypes and mRNA expression levels, and to assess whether HAVCR1 gene is involved in susceptibility to rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). Genotyping of three ins/del variants in the exon 4 was performed by fragment length analysis. Five tag SNPs genotypes and mRNA levels were determined using TaqMan assays. We defined four major haplotypes in our population: the two major haplotypes (named haplotypes A and B) bear both the 5383_5397del variant and the two most common SNP sets found in the CEU population. Quantification analysis revealed that genotype B/B had the highest median of mRNA expression levels (vs. BX + XX, p < 0.0001). Additionally, frequency of the genotype BB was significantly higher in RA patients than in controls (12.3 vs. 5.9% in controls, p = 0.0046, p (c) = 0.014, OR = 2.23, 95% CI 1.23-4.10). Our results support a relationship between HAVCR1 haplotypes and mRNA expression levels, and suggest an association of this gene with autoimmune diseases.

[Clinical and epidemiological features and HLA association in patients with pars planitis]. / Características clínicas, epidemiológicas y asociación con el sistema HLA en pacientes con pars planitis.

BACKGROUND AND OBJECTIVES: Epidemiological studies on North American patients reported an association between HLA DR15 and pars planitis. This association has not been studied in the Spanish population. The objectives of the present study were to describe the clinical and epidemiological features of patients with pars planitis diagnosed in our hospital as well as the prevalence of multiple sclerosis and HLA class I and II. PATIENTS AND METHODS: Twenty four patients with pars planitis were identified among 226 patients with uveitis diagnosed in the Ophtahlmology Department of our center from January 1992 to October 2006. Twenty four patients and 194 healthy controls underwent HLA A, B and DR genotyping. RESULTS: The most frequent complication was cystic macular edema. Most patients needed many medical treatments. No statistical association was found between pars planitis and HLA. CONCLUSIONS: Epidemiological data were consistent with previously reported studies. There appears to be no association between the occurrence of pars planitis and HLA DR 15 or other known HLA genotypes in Spanish patients. However, the small sample size could have limited the power of this study.

[Severe infections in a cohort of patients with systemic lupus erythematosus]. / Infecciones graves en una cohorte de pacientes con lupus eritematoso sistémico.

BACKGROUND AND OBJECTIVE: To study severe infectious complications in a cohort of patients with systemic lupus erythematosus (SLE). PATIENTS AND METHOD: Retrospective study of 705 patients followed from January 1980 to January 2008. Data are expressed in percentages. RESULTS: The frequency of severe infectious was 38.6%. The etiology was bacterial 54.4%, viric 30.4% and opportunist in 15.2% patients. Involved organs were: Lung 38.2%, kidney 48.9%, central nervous system 43%. 43.75% patients received pulsed ciclofosfamide therapy and 88.6% received glucocorticoids (39.7% pulsed). The mortality was 27.7%. CONCLUSIONS: At present, infection is an important cause of mortality in patients with SLE. Early diagnosis of infectious complications is very important in SLE.

[Evaluation of the effectiveness of treatment with rituximab associated to cyclophosphamide in patients with resistant idiopathic inflammatory myopathy]. / Evaluación de la eficacia del tratamiento con rituximab asociado a ciclofosfamida en pacientes con miopatía inflamatoria idiopática refractaria.

BACKGROUND AND OBJECTIVE: We report our experience with rituximab plus cyclophosphamide in the treatment of patients with resistant idiopathic inflammatory myopathies. PATIENTS AND METHOD: Open-label uncontrolled prospective study on 17 patients. RESULTS: Evaluation was completed after 1, 6 and 12 months in 95'2, 85'7 y 52'4% of cycles, respectively. Total or partial remission was achieved after 1, 6 and 12 months in 65, 100 y 63'6% of evaluated cycles, respectively. Absolute depletion of B lymphocites from peripheral blood was found in the 18 cases with available data. There were 5 relapses; median of time to relapse: 11 months; treatment was repeated in 4. Four patients (6 cycles) had impaired pulmonary function; one (with a multifactorial etiology) did not improve but the other 3, with interstitial pneumonia associated or not with respiratory muscle weakness, did. Five patients with positive anti-Jo-1 antibodies (6 cycles) displayed similar results. The only adverse event observed was a case of meningitis caused by Corynebacterium, with good results. CONCLUSION: Rituximab seems a valid alternative for the treatment of patients with resistant polymyositis or dermatomyosytis.

[Thyroid dysfunction in patients with pulmonary arterial hypertension. A cohort study of 58 patients]. / Disfunción tiroidea en pacientes con hipertensión arterial pulmonar. Estudio de una cohorte de 58 pacientes.

BACKGROUND AND OBJECTIVE: Thyroid disease (TD) is more prevalent in patients with pulmonary arterial hypertension (PAH) than in the general population. The frequency and the cause of this association are not well established. We aimed to quantify and analyze the incidence and characteristics of TD in a cohort of PAH patients (idiopathic or preferentially associated with systemic diseases) and review the literature. PATIENTS AND METHOD: Fifty eight PAH patients were prospectively studied, according to a previously established protocol (that included right heart catheterization); TSH, T(4), and antithyroglobulin and antiperoxidase antibodies were determined. TD was defined as an abnormal TSH level and/or elevated antithyroid antibodies (TAbs). Clinical, biological and hemodynamic variables were compared between patients with and without TD. RESULTS: TD was detected in 30 patients (51%): high TSH levels were observed in 21 (36,21%); hyperthyroidism in 2 (3,45%); and TAbs in 16 of 54 (27,59%), 7 of whom were euthyroid. In the TD group, PAH evolution time was longer (4,62 vs 2,61 years; P=.005, CI 95%, 0,63-3,38), more patients were in functional class IV (13;43,3% vs 5;15,8%, P=.046, CI 95% ,0,05-11,75), cardiac output was lower (P=.032, CI 95%, 3,16-4,89) and epoprostenol treatment was more frequently used (14;46,6% vs 4;14,3%, P=.008, CI 95%, 1,46-18,85; OR=5,25). CONCLUSIONS: The frequency of TD detected in this PAH cohort reaches 51% and it was associated with a longer evolution time of PAH and worse hemodynamic situation. Although epoprostenol was used more frequently in TD patients, a causal relationship with TD could not be established.

[Experience with imatinib to treat pulmonary arterial hypertension]. / Experiencia en el tratamiento de la hipertensión arterial pulmonar con imatinib.

Despite advances in the treatment of patients with pulmonary arterial hypertension (PAH), survival has not improved greatly (is still very affected). Imatinib, an antagonist of platelet-derived growth factor with antiproliferative activity, has been effective in experimental models and clinically in several published reports. We report the results of imatinib therapy in 4 patients with PAH (functional class IV) who were refractory to treatment with drug combinations for this condition. The final outcome was favorable in only 1 of the 4 cases. In this case, the patient was in functional class III and his hemodynamic parameters had improved significantly within 5 months after starting therapy. However, the patient died as a result of severe toxic hepatitis in which imatinib may have played a role. The present report adds to the few already in the literature (4 cases) and suggests that care should continue to be shown when using imatinib to treat PAH.

Comparison of two ELISA assays for anti-Sp100 determination.

Antibodies to Sp100 have been described not only in primary biliary cirrhosis (PBC), but also in other diseases. Two assays for detection of Sp100 levels by enzyme-linked immunosorbent assay (ELISA) have been compared in a cohort of patients from our area: (a) Sp100 kit produced by IMTEC, Immunodiagnostica GmbH, and (b) Quanta Lite Sp100 kit produced by INOVA Diagnostics. We analyze here the correlation between the two assays and compare their efficiency in diagnosing PBC. We also comment on the exceptions derived from reactivity with other diseases. We studied 78 sera by IIF with the typical multiple nuclear dots (MND) pattern from patients who suffered from PBC, hepatopathies different from PBC, systemic lupus erythematosus (SLE), other connective tissue diseases (CTD), skeletal diseases, lung diseases, hematological disorders, a miscellaneous group, and a healthy IIF negative control group. The tests work equally well despite their different quantification system: (a) it is based on a standard curve; and (b) it is based on a single-point antigen-specific calibration. Some discrepancies could be explained by differences in the immunodominant epitope used in the ELISA. The main finding of this study is that the presence of MND/Sp100-positive antibodies were detected not only in hepatic diseases, mainly PBC, but also in other clinical conditions, confirmed by both tests. Diagnosis of PBC must be established in the right clinical context, because other diseases recognizing the same epitope, mainly SLE, may also show high Sp100 levels. Sera from PBC patients with antimitochondrial antibodies (AMA) showed higher anti-Sp100 than the AMA-negative group.

[Rituximab for treatment of patients with systemic autoimmune diseases]. / Utilidad del rituximab en el tratamiento de pacientes con enfermedades sistémicas autoinmunitarias.

BACKGROUND AND OBJECTIVE: To assess the value of rituximab in systemic autoimmune diseases which are refractory to others treatments. PATIENTS AND METHOD: Prospective study on 12 patients -7 with systemic lupus erythematosus (SLE), 4 with Wegener's granulomatosis (WG), and 1 with overlapping connective disease and autoimmune thrombocytopenia-, controlled in a specialized unit of a tertiary hospital. Four weekly doses of rituximab, 2 biweekly doses of cyclophosphamide, and glucocorticoids were administered to all patients, and other immunosuppressants were also administered as considered necessary in each case. RESULTS: Mean follow up after treatment with rituximab was 12.8 moths for SLE patients and 12.3 for WG patients. In SLE patients, proteinuria was reduced below 1 g daily in 5 cases (83%), with a clear parallel improvement in the urinary sediment. Serositis was resolved in both cases. One patient required 3 treatment cycles to obtain an adequate response and another required a second cycle for relapse. Only one patient with WG had a favorable response. The patient treated for autoimmune thrombocytopenia had a favorable response, with no relapses, and creatine-kinase levels also tended to return to normal. There were 2 serious adverse events (terminal renal failure and serious colitis in a patient with SLE, and death of one patient with WG), that were not adjudicated directly to rituximab. Immunoglobulin levels did not change substantially. There were no infusion reactions or associated infections. CONCLUSIONS: Rituximab was useful in patients with SLE refractory to other immunosuppressants. On the contrary, its efficacy in WG was limited. The response of thrombocytopenia was complete and maintained.

[Screening of pulmonary hypertension in a Spanish cohort of patients with systemic sclerosis]. / Cribado de hipertensión pulmonar en una cohorte española de pacientes con esclerosis sistémica.

BACKGROUND AND OBJECTIVE: Pulmonary arterial hypertension (PAH) is an important cause of morbimortality in systemic sclerosis (SSc). Evolution is worse than that of subjects with idiopathic PAH, but prognosis improves when PAH is diagnosed early. The aim of this research is to describe results of a screening program for diagnosis of pulmonary hypertension (PH) carried out in a cohort of Spanish patients with SSc. PATIENTS AND METHOD: PH screening was performed by transthoracic doppler echocardiography (TTDE) in 184 patients with SSc. Patients with systolic pulmonary arterial pressure estimated by TTDE>35 mmHg were evaluated per protocol to confirm diagnosis and type of PH. RESULTS: PAH was diagnosed in 25 patients (13.6%). Patients with diffuse and limited SSc developed PAH in a similar degree, 9/60 (15%) vs. 16/100 (16%), with no cases among patients with SSc "sine scleroderma" or "pre-scleroderma" (P<.001). The only clinical or epidemiological data characterizing patients with PAH were older age (mean age 67 years for patients with PAH vs. 56 years for those without PAH, P=.007), limited SSc, a trend toward shorter evolution of the underlying disease (median 8 years for patients with PAH vs. 10 years for those without PAH, P=.73), and a higher frequency of positive anticentromere antibodies (16 patients [64%] with PAH vs. 70 (48,3%) without PAH, P=.19). CONCLUSIONS: Prevalence of PAH in SSc was high and supports the implementation of a regular screening program.

CD38 polymorphisms in Spanish patients with systemic lupus erythematosus.

The ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase (CD38) gene is a positional and functional candidate gene for the susceptibility to systemic lupus erythematosus (SLE) because CD38 gene maps in the described SLE risk region 4p15 and CD38 molecule is a leukocyte activation antigen and ectoenzyme involved in numerous immune functions. The aim of this study was to investigate the possible association of the polymorphisms located at positions 182 of intron 1 (C/G) and 418 (C/T, located in exon 3) of the CD38 gene with the susceptibility and clinical features of SLE. Genotyping of 276 Spanish patients with SLE and 194 controls was performed by polymerase chain reaction amplification-refractory mutation system techniques. No association between the polymorphisms studied and the susceptibility to SLE was found. However, when patients were stratified according to their clinical manifestations, a significant increase of CC individuals and a significant decrease of CG individuals among patients with discoid rash (67.9% vs. 53.1% in controls p = 0.02, pc > 0.05, odds ratio [OR] = 1.87, 95% confidence interval [95% CI] 1.05-3.35; and 23.5% vs. 40.2% in controls, p = 0.008, pc = 0.024, OR = 0.46 95% CI 0.24-0.85) were found. Logistic regression analysis identified CC genotype as an independent risk factor for discoid rash among patients with SLE (p = 0.01, OR = 2.23, 95% CI 1.19-4.18). In conclusion, a slight contribution of the polymorphism located in intron 1 of the CD38 gene in the clinical features of SLE could be postulated.

CTLA4 polymorphism in Spanish patients with systemic lupus erythematosus.

The cytotoxic T-lymphocyte antigen 4 (CTLA4, CD152) gene is a positional and functional candidate gene to susceptibility to systemic lupus erythematosus (SLE) because CTLA4 gene maps in the described SLE risk region 2q33 and CTLA4 molecule has an inhibitory effect on T-cell activation. Several polymorphisms have been described in CTLA4 gene, among them, a T/C change at position -1722, a C/T transition at position -319, and another A/G transition at position +49. The aim of this study was to investigate the possible association of these polymorphisms with the susceptibility to SLE in 276 Spanish autochthonous patients using a healthy control group composed of 194 ethnically matched volunteer bone marrow donors. Genotyping of these CTLA4 positions was performed in SLE patients and controls using a polymerase chain reaction amplification refractory mutation system. The genotypic frequencies were in Hardy-Weinberg equilibrium in all patients. No differences in the distribution of the genotype frequencies between patients and controls were found in any case. Our results from the Spanish autochthonous population differ from those found in the Korean population regarding the involvement of the polymorphism located at -1722 in the susceptibility to SLE.

Salivary and serum beta2-microglobulin and gamma-glutamyl-transferase in patients with primary Sjögren syndrome and Sjögren syndrome secondary to systemic lupus erythematosus.

BACKGROUND: Sialochemistry has been proposed as a simple and useful tool for the diagnosis of Sjögren syndrome (SS). Although many changes have been detected in several constituents of saliva from patients with SS, none are individually sensitive or specific enough for diagnosing SS. The aim of this study was to assess the value of the combined determination of beta2-microglobulin (beta2m) and gamma-glutamyl-transferase (GGT) activity in serum and saliva as a diagnostic instrument for differentiating primary and secondary [to systemic lupus erythematosus (SLE)] SS patients from normal subjects. METHODS: Nineteen primary SS (pSS) patients, 15 patients with SS secondary to SLE, and 25 SLE patients without SS were studied. Thirty healthy subjects were included in the study as control group. RESULTS: By means of a mathematical model, (a) 84.1%, (b) 85.7%, and (c) 87.0% of patients were correctly classified as SS or normal when (a) salivary beta2m and GGT values, (b) serum beta2m and salivary GGT values, and (c) salivary beta2m and GGT along with serum beta2m values, respectively, were considered. To differentiate between pSS and sSS by means of the mathematical model, the combination of serum beta2m and salivary GGT values achieved that 81.8% of the patients were correctly classified. CONCLUSION: Since sialochemistry is an easy, safe and reliable test, the combined determination of beta2m and GGT in saliva and serum was useful for differentiating SS patients from normal subjects, but not excessively good for differentiating pSS from sSS patients.

[Iloprost for severe Raynaud's phenomenon and ischaemic ulcers related with systemic diseases]. / Iloprost en el fenómeno de Raynaud grave y las úlceras isquémicas de las enfermedades sistémicas.

BACKGROUND AND OBJECTIVE: To evaluate the long term efficacy of treatment with intravenous iloprost for severe Raynaud's phenomenon (RP) and ischemic leg ulcers in patients with autoimmune systemic diseases. PATIENTS AND METHOD: Prospective observational study over 2 years with iloprost (intravenous infusions, 0.5 to 2 ng/kg/min, initial cycle of 5 consecutive days and maintenance infusions during 24 h monthly, lengthened when it was needed) in patients with severe RP and ischemic leg ulcers whithout response to conventional therapy. Treatment was halted in patients with a good response after one year of treatment, with regular clinical controls. RESULTS: We treated 23 patients. Iloprost reduced significantly the mean number (SD) of monthly episodes of RP (150.38 [102.04] initially and 40.05 [78.06] at the end; p < 0.0005), the mean highest duration of episodes of RP (21.86 [26.96] min initially and 7.14 [9.87] min at the end; p = 0.013), the associated pain (p = 0.005), and the mean number of ischemic digital (4.25 [2.86] initially and 0.63 [2.25] at the end; p = 0.003) and leg ulcers (1.67 [0.52] initially and 0.33 [0.52] at the end; p = 0.01). Articular symptoms and inflammatory markers did not improve. Treatment was stopped in 8 patients (in 5 for a very good evolution and in 3 for other causes), and only 1 of them needed to be treated again. Side effects were seen in all cases but always disappeared after slowing infusion. CONCLUSIONS: Iloprost was effective in the long term treatment of severe RP and ischemic leg ulcers in patients with autoimmune systemic diseases.

[Combined treatment with intravenous prostacyclin and sildenafil in patients with pulmonary hypertension: report of 4 cases]. / Tratamiento combinado con prostaciclina intravenosa y sildenafilo en pacientes con hipertensión arterial pulmonar: descripción de 4 casos.

BACKGROUND AND OBJECTIVE: Here we report the experience obtained from a combined treatment with intravenous (i.v) prostacyclin and oral sildenafil in patients with severe pulmonary hypertension (PHT) who had a poor response to prior treatment with prostacyclin alone. PATIENTS AND METHOD: Sildenafil was added to the treatment in four patients with PHT (primary in two patients and secondary to collagenosis in the other two) with no adequate response to i.v. prostacyclin treatment. The clinical course, 6minutes walking test and echocardiogram were evaluated. RESULTS: Initial sildenafil dose was 12.5 mg three times daily, which was increased up to 50 mg three times daily in one patient and up to 50 mg four times daily in the other three. The symptoms of right heart failure were controlled in all cases. Before the start of sildenafil administration, two patients had class III dyspnea and two patients had class IV dyspnea. Two patients converted to class I (previously class III and IV), and the other two converted to class II. The distance walked within 6 minutes increased (average increase 55%) and systolic pulmonary artery pressure decreased in all patients (average reduction 27%). Effects of sildenafil were substained. The only side effect seen was mild headache. CONCLUSIONS: Our experience supports the value of sildenafil in the treatment of PHT and suggests that combined treatment is useful for rescuing patients who fail to respond to initial treatment with i.v. prostacyclin.

[Usefulness of brain SPECT with HMPAO-99mTc and psychological tests for diagnosis of neurological involvement in Behçet's disease]. / Utilidad de la SPECT cerebral con HMPAO-99mTc y de los tests psicométricos en el diagnóstico de la afección neurológica de la enfermedad de Behçet.

BACKGROUND: The purpose of this study was to establish the usefulness of single photon emission computed tomography (SPECT) and psychological tests for diagnostic of neuro-Behçet (NB) and to evaluate the clinical significance of neurological symptoms that are difficult to interpret and asymptomatic abnormalities in diagnostic tests. PATIENTS AND METHOD: Forty patients with Behçet's disease (BD) were enrrolled for being studied with magnetic resonance imaging (MRI), SPECT and psychological tests. RESULTS: MRI findings were abnormal in 52,9% of patients with neurological involvement and 23.1% without it (p < 0.1), whereas SPECT findings were abnormal in 82.3% and 61.5%, respectively (no significant difference). The difference between MRI and SPECT findings was significant (p < 0.02 for the complete group; p < 0.05 for patients without neurological symptoms; p < 0.08 for patients with them). The mean follow-up period was 42.6 months, and no patient without neurological involvement or those only with neurological symptoms that are difficult to interprete developed definite neurological involvement. The results of cognitive tests were not significantly different among patients with or without neurological involvement, neither among patients and controls. The scale 2 (depression) of the personality test was more frequent in patients with definite neurological involvement (p < 0.05). CONCLUSIONS: SPECT seems more sensible and less specific than MRI for diagnostic of NB. Although SPECT findings were frequently abnormal in patients with BD without neurological involvement or with neurological symptoms hard to interpret, no patient from this group developed a NB flare after a long follow-up period. A characteristic personality was found for patients with BD.