Lourdes Mozo

Epidemiology of systemic lupus erythematosus in a northern Spanish population: gender and age influence on immunological features.

The present work was planned to research epidemiological and immunological features of systemic lupus erythematosus (SLE) in a Caucasian population from the north of Spain (Asturias). There is only one specialized immunology laboratory in this region where samples from all patients with a plausible or a firm diagnosis of SLE are referred for immunological analysis. Since 1992 we have reviewed registered data from samples submitted to the immunology laboratory with a firm, definitive diagnosis of SLE, based on the fulfillment of the American College of Rheumatology (ACR) criteria. We have constructed a database, which included 367 SLE patients. The point prevalencewas 34.12/100 000 (95% CI: 30.63-37.61/100 000), whereas the incidence rate calculated during the last five years was 2.15/100 000/year (95% CI: 1.76-2.54/100 000/year). The female/male ratio varied according to the age at diagnosis, being maximum (50: 1) between 22 and 28 years. The median age at diagnostis was significantly lower in females than in males. Immunological features also yielded sex and age peculiarities. The percentage of patients with anti-SSa antibodies yielded significant differences between males (18.6%) and females (34.6%). Anti-RNP and anti-Sm antibodies were more frequently present in childhood-onset patients, the difference with the oldest-onset group being statistically significant. Other analyses did not show significant differences, although, as a whole, we observed a trend towards a higher presence of autoantibodies related to an early disease onset.

Glucocorticoids increase IL-10 expression in multiple sclerosis patients with acute relapse

High doses of glucocorticoids (GCs) are widely employed to treat acute attacks in relapsing-remitting multiple sclerosis (MS) patients. Their beneficial effects are partially due to their capacity to regulate the cytokine network. In the present work, we have examined the effect of GCs on the production of the immunosuppressor cytokine IL-10. Blood samples from MS patients suffering an acute relapse were obtained immediately before initiating therapy and after receiving a daily dose of 1 g intravenous methylprednisolone (MP) for four days. Levels of IL-10 mRNA in PBMC were semiquantified by RT-PCR, whereas protein concentration in serum and in cell culture supernatant was measured by ELISA. Our results show that 7 out of the 9 patients studied displayed increased IL-10 mRNA expression as well as higher serum IL-10 concentration following steroid treatment. In contrast, mRNA expression of two inflammatory cytokines, TNFalpha and IFNgamma, decreased following steroid therapy. In vitro experiments employing normal PBMC showed that methylprednisolone (MP) upregulated IL-10 expression as determined by measuring mRNA levels, flow cytometry of intracytoplasmic protein concentration, and the amount of secreted protein. Peak responses of secreted IL-10 by PBMC cultured cells treated with MP were obtained at 48 h. The effect was steroid-specific as IL-10 expression reversed to baseline levels in the presence of the glucocorticoid receptor antagonist RU486. Contrary to the effect of MP on the spontaneous expression of IL-10, this drug downregulated LPS-induced IL-10 synthesis. In fact, the concentration of IL-10 in LPS-induced IL-10 secretion from normal PBMC decreased upon addition of MP to cell cultures. Thus, it seems that MP exerts an opposite effect on the spontaneous and LPS-induced IL-10 production. Our studies indicate that GCs may control inflammatory responses by upregulating production of the immunosuppressor cytokine IL-10.

TNFalpha and IL-10 gene polymorphisms in inflammatory bowel disease. Association of -1082 AA low producer IL-10 genotype with steroid dependency.

OBJECTIVES:An altered production of cytokines underlies inflammatory bowel disease (IBD) susceptibility. Various polymorphisms at the IL-10 and TNFα gene promoters control cytokine production levels. The influence of these polymorphisms on susceptibility to ulcerative colitis (UC) and Crohns disease (CD) and their association with clinical features were analyzed.SUBJECTS AND METHODS:Genetic polymorphisms of TNFα (−308 G/A) and IL-10 (−1082 G/A, −812 C/T, and −592 C/A) were determined using the LightCycler system with hybridization probes matched with one sequence variant. The study population included 99 UC patients, 146 CD patients, and 343 matched controls.RESULTS:We did not find association between TNFα or IL-10 gene polymorphisms and UC or CD susceptibility, though a slight influence of −1082*G allele in UC appearance was observed. In a stratified analysis, a highly significant association between the −1082 AA IL-10 genotype and the steroid dependency was observed in IBD (p < 0.0001), contributing both UC (p = 0.004) and CD (p = 0.003) to this association. In contrast, TNFα genotypes did not influence steroid dependency in IBD. Further, the contribution of cytokine genotypes and of clinical features to the appearance of steroid-dependent status (dependent variable) was studied by multivariate analysis. The steroid-dependent phenotype correlated in UC with extensive disease (p = 0.010) and with the low producer −1082 AA IL-10 genotype (p = 0.002) and in CD with penetrating disease (p = 0.010), arthritis (p = 0.011), and the −1082 AA IL-10 genotype (p = 0.006).CONCLUSIONS:The main conclusion is that carriage of the −1082 AA IL-10 genotype (low producer) is a relevant risk factor for developing steroid-dependent IBD.

Glucocorticoids up-regulate constitutive interleukin-10 production by human monocytes

Background IL‐10 plays an immunosuppressive role in inflammatory responses. Increased plasma levels of IL‐10 have been detected in patients under glucocorticoid (GC) therapy, indicating that steroids may exert their suppressive effect, in part, by increasing IL‐10 production.

Glucocorticoids inhibit IL-4 and mitogen-induced IL-4Rα chain expression by different posttranscriptional mechanisms

BACKGROUND A high level of expression of IL-4Ralpha chain on the surface of lymphocytes has been described in certain allergic and inflammatory autoimmune diseases. Progression of these diseases are usually controlled by steroid treatment. One mechanism by which these drugs exert their antiinflammatory and immunosuppressive effects is by widely repressing or enhancing the production of cytokines and their receptors. OBJECTIVES The effect of glucocorticoids on IL-4Ralpha chain expression has not been previously studied, and this is the aim of the present report. For this purpose, human lymphocytes were induced to express IL-4Ralpha chain by means of protein kinase C (PKC) activation with phorbol myristate acetate (PMA) or by triggering the Janus kinase-Stat pathway with IL-4 in the presence or absence of pharmacologic doses of dexamethasone. METHODS IL-4Ralpha cell surface expression was studied by flow cytofluorometry. The levels and stability of mRNA were assessed by Northern blot analysis. The effect of dexamethasone on the IL-4Ralpha rate of transcription was determined by nuclear run-on experiments. RESULTS Dexamethasone significantly downregulated PMA-induced IL-4Ralpha mRNA and protein levels in total peripheral blood mononuclear cells and in isolated T cells. The mechanism involved a posttranscriptional regulation of IL-4Ralpha expression because dexamethasone decreased the PMA-induced IL-4Ralpha mRNA half-life. However, we found that PMA did not influence the transcription rate of IL-4Ralpha gene, irrespective of the presence or absence of dexamethasone. This immunosuppressor also diminished the IL-4-induced IL-4Ralpha expression on the surface of isolated T and B lymphocytes but, interestingly, without modifying mRNA levels that indicates that dexamethasone downregulated IL-4-dependent IL-4Ralpha expression by acting at a translational or posttranslational level. In fact, we observed that the drug did not affect IL-4-induced IL-4Ralpha gene transcription rate nor did it shorten mRNA half-life. The effect of dexamethasone on the IL-4Ralpha was steroid specific because it was totally reversed by the glucocorticoid receptor antagonist RU486. CONCLUSION Our results support that dexamethasone may influence the course of allergic and inflammatory diseases by downregulating the expression of IL-4Ralpha.

Systemic lupus erythematosus in Asturias, Spain: clinical and serologic features.

Abstract: Asturias is an autonomous region in the north of Spain with historical and anthropologic peculiarities. In the current report, we examine the main clinical and immunologic features of 363 patients with systemic lupus erythematosus (SLE), virtually the entire population of SLE patients in Asturias. We constructed a database with the clinical and immunologic features of all patients fulfilling the American College of Rheumatology criteria, based on the review of hospital records corresponding to blood samples received for antinuclear antibodies testing since 1992. Arthritis was the most frequently observed main clinical feature and neuropathy was the rarest. Male patients had a disease more frequently characterized by serositis (p < 0.05) and neurologic disorder (p < 0.01) than females, while children presented malar rash (p < 0.05), fever (p < 0.05), and kidney involvement (p < 0.01) more often than adults. Late-onset patients were characterized by lower frequencies of malar rash (p < 0.01), neurologic disorder (p < 0.05), alopecia (p < 0.01), and lymphadenopathy (p < 0.05) than young adults. Numerous direct and inverse significant associations were found among clinical and immunologic features. The most relevant significant associations were neurologic disorder with lupus anticoagulant (p < 0.01); kidney involvement with serositis (p < 0.01) and DNA antibodies (p < 0.05); and thrombosis with DNA antibodies (p < 0.05), cardiolipin antibodies (p < 0.01), and lupus anticoagulant (p < 0.01). A low mortality was found in our series, although kidney involvement (p < 0.05) and cardiolipin antibodies (p < 0.05) are factors associated with poor survival. Abbreviations: ACR = American College of Rheumatology, CI = confidence intervals, dsDNA = double-stranded DNA, OR = odds ratio, RNP = ribonucleoprotein, SLE = systemic lupus erythematosus, SM = Smith, SSA = Sjögren syndrome A, SSB = Sjögren syndrome B.

Cytokine polymorphisms influence treatment outcomes in SLE patients treated with antimalarial drugs

Antimalarial agents have been widely used as disease-modifying antirheumatic drugs in the treatment of systemic lupus erythematosus (SLE) and other rheumatological diseases, although their mechanism of action has not yet been fully defined. It is known, however, that effective response to treatment is variable among patients. Thus, the identification of genetic predictors of treatment response would provide valuable information for therapeutic intervention. The aim of the present study was to analyze the effect of antimalarial treatment on tumor necrosis factor (TNF)α serum levels and evaluate the possible influence of TNFα and IL-10 functional genetic polymorphisms on the response to antimalarial drugs. To this end, TNFα serum levels were quantified in 171 SLE patients and 215 healthy controls by ELISA techniques and polymorphisms at positions -1,082 and -308 of the IL-10 and TNFα gene promoterswere determined by PCR amplification followed by hybridization with fluorescent-labeled allele-specific probes in 192 SLE patients and 343 matched controls. Data were related to clinical features and treatment at the time of sampling and during the course of the disease. Results showed a significantly higher amount of serum TNFα in the entire SLE population compared with controls. However, TNFα serum levels correlated negatively with the use of antimalarial treatment during at least three months before sampling. Patients under single or combined treatment with these drugs had TNFα serum levels similar to healthy controls, whereas untreated patients and those under corticosteroid or immunosuppressive therapies had increased amounts of this cytokine. This suggests, however, that antimalarial-mediated inhibition of TNFα was only significant in patients who were genetically high TNFα or low IL-10 producers. In addition, evaluation of SLE patients administered antimalarial drugs for three or more years who did not require any other specific SLE treatment indicates that patients with the combined genotype low IL-10/high TNFα are the best responders to antimalarial therapy, developing mild disease with a good course under this treatment. In conclusion, we proposed that an antimalarial-mediated downregulation of TNFα levels in SLE patients is influenced by polymorphisms at IL-10 and TNFα promoters. Our results may thus find important clinical application through the identification of patients who are the most likely to benefit from antimalarial therapy.

Generation of CD4 + CD45RA + Effector T Cells by Stimulation in the Presence of Cyclic Adenosine 5′-Monophosphate- Elevating Agents

After TCR cross-linking, naive CD4+CD45RA+ T cells switch to the expression of the CD45RO isoform and acquire effector functions. In this study we have shown that cAMP-elevating agents added to anti-CD3- and anti-CD28-stimulated cultures of T lymphocytes prevent acquisition of the CD45RO+ phenotype and lead to the generation of a new subpopulation of primed CD4+CD45RA+ effector cells (cAMP-primed CD45RA). These cells displayed a low apoptotic index, as the presence of dibutyryl cAMP (dbcAMP)-rescued cells from CD3/CD28 induced apoptosis. Inhibition of CD45 splicing by dbcAMP was not reverted by addition of exogenous IL-2. cAMP-primed CD45RA cells had a phenotype characteristic of memory/effector T lymphocytes, as they showed an up-regulated expression of CD2, CD44, and CD11a molecules, while the levels of CD62L Ag were down-regulated. These cells also expressed the activation markers CD30, CD71, and HLA class II Ags at an even higher level than CD3/CD28-stimulated cells in the absence of dbcAMP. In agreement with this finding, cAMP-primed CD45RA cells were very efficient in triggering allogenic responses in a MLR. In addition, cAMP-primed CD45RA cells produce considerable amounts of the Th2 cytokines, IL-4, IL-10, and IL-13, whereas the production of IFN-γ and TNF-α was nearly undetectable. The elevated production of IL-13 by neonatal and adult cAMP-primed CD45RA cells was specially noticeable. The cAMP-dependent inhibition of CD45 splicing was not caused by the production of immunosuppressor cytokines. These results suggest that within the pool of CD4+CD45RA+ cells there is a subpopulation of effector lymphocytes generated by activation in the presence of cAMP-elevating agents.

Clinical associations of anti-SSA/Ro60 and anti-Ro52/TRIM21 antibodies: Diagnostic utility of their separate detection

Clinical associations of anti-SSA/Ro60 and anti-Ro52/TRIM21 antibodies are not yet fully established. In order to analyse the diagnostic utility of their separate detection, we retrospectively revised the clinical data of 200 anti-SSA/Ro60 and/or anti-Ro52/TRIM21 positive patients identified by line immunoassay during ANA routine detection. Anti-SSA/Ro60 positive patients showed a significantly higher prevalence of autoimmune diseases (AIDs) independently on the presence of anti-Ro52/TRIM21 (OR 3.13, 95% CI 1.10–8.88, p = 0.032). Anti-SSA/Ro60 was independently associated with systemic lupus erythematosus (SLE) when comparing with Sjögrens syndrome (SS) and other systemic AIDs (OR 3.46, 95% CI 1.08–11.06, p = 0.036). The more frequent specificity found in cutaneous lupus erythematosus (CLE) was also anti-SSA/Ro60. In contrast, detection of isolated anti-Ro52/TRIM21 was characteristic of SS (7/35, 20.0%), diffuse cutaneous systemic sclerosis (dcSSc) (3/4, 75.0%), primary biliary cirrhosis (PBC) (4/5, 80.0%) and, specially, of polymyositis/dermatomyositis (PM/DM) (6/6, 100%). In fact, anti-Ro52/TRIM21 was the only antibody detected in 4 out of the 6 PM/DM patients. Malignancies mainly account for the observed high prevalence of mono-specific anti-Ro52/TRIM21 in patients with non-AIDs (10/15, 62.5%). In conclusion, this retrospective study supports the routine distinction of anti-SSA/Ro60 and anti-Ro52/TRIM21 due to their different clinical associations.

Diagnostic value of anti-deamidated gliadin peptide IgG antibodies for celiac disease in children and IgA-deficient patients.

Objectives: The aim of the study was to analyze the diagnostic performance of anti-deamidated gliadin peptide (dGp) immunoglobulin (Ig) G and IgA regarding the age at celiac disease (CD) diagnosis and the anti-dGp IgG usefulness for diagnosing CD IgA-deficient patients. Methods: Anti-dGp IgG and IgA and anti-native gliadin (nGlia) IgA were determined by enzyme fluoroimmunoassay in 100 newly diagnosed anti-tissue transglutaminase (tTG) IgA-positive pediatric and adult patients with CD and in 100 age-matched patients with other digestive pathologies. Anti-dGp IgG was evaluated in 6 CD IgA-deficient patients. Results: When analyzing all of the patients, the anti-dGp IgG assay showed higher diagnostic accuracy (area under receiver operating characteristic curve), specificity, and positive predictive value than anti-dGp IgA and anti-nGlia IgA. All of the diagnostic parameters corresponding to anti-dGp IgG reached the same values as anti-tTG IgA in children 7 years or younger. In older patients, both anti-dGp isotypes showed an inverse behavior, IgG having a higher specificity and positive predictive value but a lower sensitivity and negative predictive value than IgA. Anti-dGp levels were associated with the severity of intestinal lesions, and an inverse association was found regarding age at diagnosis. Both anti-dGp IgG and IgA were found to be positive in the 9 patients with minimal intestinal changes included in the study. All of the patients with CD with IgA deficiency were positive for anti-dGp IgG. Conclusions: The diagnostic performance of anti-dGp depends on the antibody isotype and on the age at CD diagnosis, anti-dGp IgG being a serological marker at least as useful as anti-tTG IgA for detecting CD in children ages 7 years or younger. Our data also indicate that anti-dGp IgG can improve the diagnosis of IgA-deficient patients.