Tear levels of IL-7, IL-1α, and IL-1ß may differentiate between IgG4-related disease and Sjögren's syndrome.

We aim to assess and compare a cytokine and chemokine profile in tears from patients with IgG4-related disease (IgG4-RD) and Sjögren's syndrome (SS), and to see if this profile could aid in differentiating these two diseases. We included 10 patients with IgG4-RD who met the Comprehensive Diagnostic Criteria for IgG4-RD and 17 patients who met the AECG criteria for primary SS. The Schirmer-I test was carried out using two standardized sterile tear strips, which were then immediately frozen at - 86 °C until assayed. The tears were extracted from the strips after they had been defrosted using a buffer containing 0.5 M NaCl and 0.5% Tween-20. The amounts (pg/ml) of the following cytokines and chemokines were then measured using luminometry: IFN-γ, TNF-α, G-CSF, IL-1-α, IL-1ß, IL-4, IL-7, IL-12p40, IL-12p70, IL-13, IL-17A, CCL2, CCL3, CCL4, CCL11, and CXCL10. In the IgG4-RD group, seven patients had lacrimal gland involvement, five had dry eye symptoms, and six had a positive Schirmer-I test. In the SS group, 16 (94.1%) had dry eyes and all had a positive Schirmer-I test. We were able to differentiate between both diseases using levels of IL-7, IL-1α, and IL-1ß; in particular, the IL-7/IL-1α and IL-7/IL-1ß ratios had the best discriminatory potential, with cut-off values of 0.32 (AUC: 0.93, sensitivity: 94%, specificity: 80%, p = 0.0003) and 12.55 (AUC: 0.96, sensitivity: 94%, specificity: 90%, p = 0.0001), respectively. Our results suggest that IL-7, IL-1α, and IL-1ß tear levels could help differentiate IgG4-RD from SS. Key Points • The lacrimal gland is frequently involved in IgG4-RD and SS. This characteristic makes both diseases mimics of one another. • Patients with IgG4-RD and SS have different profiles of tear cytokines and chemokines. • Tear IL-7, IL-1α, and IL-1ß levels may serve as helpful biomarkers in separating IgG4-RD from SS.

Dendritic cells drive profibrotic inflammation and aberrant T cell polarization in systemic sclerosis.

OBJECTIVES: SSc is a devastating autoimmune disease characterized by fibrosis and obliterative vasculopathy affecting the skin and visceral organs. While the processes mediating excessive extracellular matrix deposition and fibroblast proliferation are clear, the exact link between autoimmunity and fibrosis remains elusive. Th17 cells have been proposed as critical drivers of profibrotic inflammation during SSc, but little is known about the immune components supporting their pathogenic role. Our aim was to determine cytokine responses of stimulated monocyte-derived dendritic cells (Mo-DCs) and to determine how they influence T-cell cytokine production in SSc. MATERIAL AND METHODS: Dendritic cells (DCs) activate and shape T cell differentiation by producing polarizing cytokines. Hence, we investigated the cytokine responses of monocyte-derived DCs (Mo-DCs) from patients with limited cutaneous SSc (lcSSc), diffuse cutaneous SSc (dcSSc) and healthy controls (HCs) after stimulation with toll-like receptor (TLR) agonists. Also, using co-culture assays, we analysed T cell subpopulations after contact with autologous TLR-activated Mo-DCs. RESULTS: In general, we observed an increased production of Th17-related cytokines like IL-1ß, IL-17F, IL-21 and IL-22 by SSc compared with HC Mo-DCs, with variations between lcSSc vs dcSSc and early- vs late-stage subgroups. Noticeably, we found a significant increment in IL-33 production by Mo-DCs in all SSc cases regardless of their clinical phenotype. Strikingly, T cells displayed Th2, Th17 and dual Th2-Th17 phenotypes after exposure to autologous TLR-stimulated Mo-DCs from SSc patients but not HCs. These changes were pronounced in individuals with early-stage dcSSc and less significant in the late-stage lcSSc subgroup. CONCLUSIONS: Our findings suggest that functional alterations of DCs promote immune mechanisms favouring the aberrant T cell polarization and profibrotic inflammation behind clinical SSc heterogeneity.

Chemokine tear levels in primary Sjögren's syndrome and their relationship with symptoms.

PURPOSE: To evaluate CCL2, CXCL8, and CXCL10 in the tears of patients with Primary Sjögren's syndrome (PSS) and correlate them with ocular symptoms/discomfort and objective ocular tests. METHODS: We studied 21 patients with PSS. A single ophthalmologist, expert in dry eye, examined the patients and assessed tear film breakup time, Schirmer I test, tear meniscus height, Van Bijsterveld staining score and SICCA Ocular Staining Score. We also assessed the ESSPRI and ocular dryness VAS and the Ocular Surface Disease Index (OSDI), a 12-item scale assessing symptoms associated with dry eye disease and their impact on vision (ocular symptoms/discomfort). Tear samples collected with sterile tear flow strips were frozen at -86 °C until testing. After thawing, tears were extracted from the strips. We tested CCL2, CXCL8, and CXCL10 by luminometry. We also included 21 healthy control subjects without a dry eye. RESULTS: CXCL8 levels were similar in patients and controls. PSS patients had lower levels of CXCL10 (472.8 vs. 1652 pg/µL, p = 0.009) and CCL2 (1.08 vs. 9 pg/µL, p = 0.0001) than controls. Patients with worse ocular sicca symptoms/discomfort had the lowest CXCL10 levels (239.3 vs. 646.2 pg/µL, p = 0.02). CCL2 correlated with tear meniscus height (τ = 0.37, p = 0.02) and with OSS (τ = -0.3, p = 0.05). CONCLUSIONS: We found lower levels of CXCL10 and CCL2 in the tears of patients with PSS, associating the former with worse ocular symptoms and the latter with positive ocular target tests.

Heterogeneity of Genetic Admixture Determines SLE Susceptibility in Mexican.

Systemic Lupus Erythematosus (SLE) is an autoimmune inflammatory disorder for which Major Histocompatibility Complex (MHC) genes are well identified as risk factors. SLE patients present different clinical phenotypes, which are partly explained by admixture patterns variation among Mexicans. Population genetic has insight into the high genetic variability of Mexicans, mainly described through HLA gene studies with anthropological and biomedical importance. A prospective, case-control study was performed. In this study, we recruited 146 SLE patients, and 234 healthy individuals were included as a control group; both groups were admixed Mexicans from Mexico City. The HLA typing methods were based on Next Generation Sequencing and Sequence-Based Typing (SBT). The data analysis was performed with population genetic programs and statistical packages. The admixture estimations based on HLA-B and -DRB1 revealed that SLE patients have a higher Southwestern European ancestry proportion (48 ± 8%) than healthy individuals (30 ± 7%). In contrast, Mexican Native American components are diminished in SLE patients (44 ± 1%) and augmented in Healthy individuals (63 ± 4%). HLA alleles and haplotypes' frequency analysis found variants previously described in SLE patients from Mexico City. Moreover, a conserved extended haplotype that confers risk to develop SLE was found, the HLA-A∗29:02∼C∗16:01∼B∗44:03∼DRB1∗07:01∼DQB1∗02:02, pC = 0.02, OR = 1.41. Consistent with the admixture estimations, the origin of all risk alleles and haplotypes found in this study are European, while the protection alleles are Mexican Native American. The analysis of genetic distances supported that the SLE patient group is closer to the Southwestern European parental populace and farthest from Mexican Native Americans than healthy individuals. Heterogeneity of genetic admixture determines SLE susceptibility and protection in Mexicans. HLA sequencing is helpful to determine susceptibility alleles and haplotypes restricted to some populations.

BLK and BANK1 variants and interactions are associated with susceptibility for primary Sjögren's syndrome and with some clinical features.

BLK and BANK1 in primary Sjögren's syndrome (pSS) have scarcely been evaluated and the results are inconclusive. The aim of our study was to determine whether single nucleotide variants (SNVs) located within BLK or BANK1 are associated with susceptibility, clinical and serological features, and smoking in pSS. BLK rs13277113A/G, BANK1 rs10516487G/A and rs3733197G/A were genotyped in 203 cases and 424 controls using a TaqMan® SNP genotyping assay. The BLK rs13277113A allele showed association with pSS under the allelic (OR 1.35, p = 0.02), and recessive (OR 1.83, p = 0.003) model, while, BANK1 rs3733197G/A showed association under the dominant model (OR 2.90, p = 0.043). Interactions between BANK1 and BLK genotypes also showed association (OR 2.36, p < 0.0001). In addition, BLK rs13277113A/G was associated with protection against arthritis and BANK1 rs10516487G/A with both arthritis and keratoconjunctivitis sicca, meanwhile, BANK1 rs3733197G/A was associated with smoking in patients with pSS. This is the first study to describe an association between BLK and susceptibility to pSS in a Latin-American population. Our data also shows a first evidence of association between interactions of BLK and BANK1 in pSS, and association of BLK and BANK1with arthritis, keratoconjunctivitis sicca and smoking in patients with pSS.

Omega-3 and omega-6 fatty acids in primary Sjögren's syndrome: clinical meaning and association with inflammation.

OBJECTIVES: Lipid mediators derived from polyunsaturated fatty acids (FA), have been related to inflammation and immune response regulation. Herein we evaluated the intake and serum levels of ω-3 and ω-6 FA among patients with primary Sjögren's syndrome (pSS), and correlated with ocular/oral sicca symptoms, disease activity and a panel of chemokines/cytokines. METHODS: We included 108 patients and 100 controls. Dietary information was obtained from a food questionnaire of one-day reminder and processed using a nutritional software. Among the SS group, we measured serum ω-3 (α-linolenic acid [α-LN], eicosapentaenoic acid [EPA], docosahexaenoic acid [DHA]) and ω-6 (linoleic acid [LA], arachidonic acid [AA]) by gas chromatography flame ionization. We scored the ESSDAI, ESPRI, Schirmer-I test and NSWSF. In a subsample, we assessed the OSDI, ophthalmologic staining scores and measured CXCL8, CXCL10, CCL2, IL-22 and IL-21 in saliva, and CXCL8, CXCL10, CCL2 and CXCL9 in tears by Luminometry. RESULTS: ω-3 and ω-6 intake was lower in SS patients than controls, and did not correlate with serum levels. We found a negative correlation between α-LN and the OSDI and ESSDAI, as well as DHA and ESSDAI. In tears, AA positively correlated with CXCL9, whereas in saliva, α-LN, DHA and the ω3 sum negatively correlated with CCL2. We observed a negative correlation between the ω6 sum and IL-21. CONCLUSIONS: pSS patients had deficient omega intake. Lower ocular symptoms, ESSDAI scores and salivary CCL2 correlated with higher ω-3 levels, possible suggesting a role in chronic inflammation. Further studies are warranted to deepen in the knowledge of this association.

Immunosenescence profile and expression of the aging biomarker (p16INK4a) in testicular cancer survivors treated with chemotherapy.

BACKGROUND: Cytotoxic chemotherapy can cure advanced germ cell tumors. Nevertheless, cancer treatment may induce cellular senescence and accelerate molecular aging. The aging process implies an increase of cells expressing p16INK4a and changes in lymphocyte subpopulations. Our aim was to study the potential induction of premature immunosenescence in testicular cancer survivors (TCS) exposed to chemotherapy. METHODS: Case-control exploratory study of TCS treated with chemotherapy (≥3 BEP cycles, disease-free ≥3 months) compared with age matched healthy controls. Peripheral blood mononuclear cells were isolated, and lymphocyte subpopulations were analyzed by flow cytometry. CDKN2A/p16INK4a expression in T cells was measured using qPCR. The percentage of lymphocyte subpopulations and the CDKN2A/p16INK4a expression in TCS were compared with the control group using the Wilcoxon signed-rank test. RESULTS: We included 16 cases and 16 controls. The median age was 27 years (minimum 24, maximum 54) and the median time on surveillance was 26.5 months (minimum 3, maximum192). TCS had a lower percentage of total T cells and CD4+ T cells in total lymphocytes. Among the CD4+ T lymphocytes, TCS had less naïve CD4+ and increased memory CD4+ cells. Within the CD8+ T lymphocytes, TCS exhibited a decrease in the percentage of naïve cells and an increase in CD8 + CD45RA + CD57+ cells. TCS also exhibited decreased memory CD19+ B cells compared to the controls. The relative expression of CDKN2A/p16INK4a in T cells was increased in TCS (mean 1.54; 95% CI of the mean: 1.074-2.005; p = 0.048). CONCLUSION: In this exploratory study, TCS showed increased expression of CDKN2A/p16INK4a and a lymphocyte phenotype that has been associated with immunosenescence. Further studies are warranted to define the clinical implications of these alterations in TCS.

Aspartic acid70 in the HLA-DRB1 chain and shared epitope alleles partially explain the high prevalence of autoimmunity in Mexicans.

INTRODUCTION: Autoimmune thyroid disease (AITD) is the most common autoimmune disorder worldwide. Remarkably, it is commonly accompanied by other autoimmune diseases, such as rheumatoid arthritis (RA). The immunopathogenic mechanisms behind the coexistence of these disorders are still not completely understood. Immunogenetics influences the physiopathology of these diseases since ethnicity plays an essential role in the inheritance of susceptibility markers. METHODS: High-resolution HLA class II typing was performed using a sequence-based method. RESULTS: The allele frequency of HLA-DRB1∗04:04 and -DRB1∗03:01 were significantly increased in patients with AITD and RA compared to healthy individuals, pC â€‹= â€‹0.021, OR â€‹= â€‹2.4, 95%CI â€‹= â€‹1.19-4.75 and pC â€‹= â€‹0.009, OR â€‹= â€‹3.4, 95%CI â€‹= â€‹1.42-7.93, respectively. Remarkably, these patients have a combined risk given by susceptibility HLA-DRB1 alleles that contain the shared epitope, pC â€‹= â€‹0.03, OR â€‹= â€‹1.7, IC95% â€‹= â€‹1.07-2.76, and a lack of protective alleles carrying aspartic acid70, pC â€‹= â€‹0.009, OR â€‹= â€‹0.5, IC95% â€‹= â€‹0.32-0.84. DISCUSSION: The results suggest that patients with AITD and RA have an immunogenetic mechanism that combines the susceptibility alleles associated with both diseases. Importantly, it seems to be linked mainly to the lack of protective alleles with aspartic acid in the position 70, along with the presence of susceptibility alleles that have the sequences QRRAA, QKRAA, and RRRAA at positions 70-74. CONCLUSION: Patients with AITD and RA have a characteristic immunogenetic signature, which could be useful for determining multiple autoimmunities and assessing their relatives' risk of developing it.

The role of socioeconomic status in the susceptibility to develop systemic lupus erythematosus in Mexican patients.

INTRODUCTION: Systemic lupus erythematosus (SLE) is an autoimmune inflammatory disorder for which Major Histocompatibility Complex (MHC) genes are well-identified as risk factors. SLE patients have different phenotypes or clinical presentations, which vary among Mexicans. This variation could be explained by ethnicity and admixture. Since socioeconomic status probably limits and change the patterns of migration, this factor could favor inbreeding and homogamy in some geographic areas. Consequently, it could alter or restrict the possibilities of admixture too. Therefore, the socioeconomic status may also have implications in the susceptibility and the clinical heterogeneity of SLE in Mexican patients. METHODS: One hundred twenty-three SLE patients and 234 healthy individuals with Mexican admixed ancestry were recruited. HLA alleles were analyzed using the HLA typing method based on Sequence-based typing (SBT). RESULTS: As expected, it was found an increased frequency of the HLA-DRB1*03:01 allele in all socioeconomic groups when compared with healthy individuals. The susceptibility allele found in the low-income SLE patients was HLA-DRB1*04:05 whereas, the susceptibility alleles for the high-income SLE patients were HLA-DRB1*07:01 (pC = 0.03, OR = 2.0) and HLA-DRB1*11:04 (pC = 0.0004, OR = 5.1). Additionally, the frequencies of two protective alleles HLA-DRB1*14:06 (pC = 0.01, OR = 0.28) and HLA-DRB1*16:02 (pC = 0.04, OR = 0.22) were found diminished. These findings correlate with the admixture differences between low-income and high-income SLE patients. The clinical manifestations showed a different distribution between both groups. Arthritis and neurological disorder were prevalent in low-income SLE patients, while the hematological disorder was prevalent in high-income SLE patients. CONCLUSIONS: These findings suggest that HLA class II DRB1 genes contribute to the susceptibility and protection to develop SLE differently depending on socioeconomic status. Due to this, the clinical manifestations vary among patients and it could be related to different admixture charge.Key Point• HLA class II DRB1 genes contribute to the susceptibility and protection to develop SLE differently depending on socioeconomic status.

Value of C3d assay and IgG subclass in the prediction of the flow cytometry cross-match result for renal transplantation.

The aim of this study is to compare the association and the predictive capacity of DSA MFI, complement fixing capacity (C3d assay) and IgG subclasses determination in the prediction of FCxM result. METHODS: We used cryopreserved (70C) sera from potential renal transplant recipients, containing DSA against their respective potential donors. All patients showed negative AHG-CDC CxM and either positive or negative FCxM. Class I and Class II HLA-DSA were determined by Luminex SAB. C3d were detected by Luminex (Lifecodes®Immucor), DSA IgG 1-4 subclasses were evaluated using monoclonal antibodies specific for IgG subclasses (Luminex). RESULTS: 93 donor/recipient tests were evaluated; 32 (35.9%) patients presented at least one C3d + Ab, of which only 11 (11.8%) were donor specific. At least one IgG subclass was identified in 45 samples (48.3%). Twenty-seven FCxM tests (29%) were positive. On multivariate analysis HLA mismatches, the IgG subclass detection, DSA MFI and class II PRA remain associated to FCxM whereas C3d + Ab was not associated. For the FCxM prediction, the IgG subclass detection in combination with the DSA-MFI > 2800, had the best negative predictive value 93.9 (CI 95%, 84.2-100). CONCLUSION: Neither the C3d assay nor the IgG subclasses detection alone had an adequate predictive capacity for the FCxM. In the absence of IgG subclass detection and DSA-MFI < 2000, the probability of a negative FCxM was near 94%.

Elastographic ultrasound: an additional image tool in Sjögren's syndrome.

AIM: To evaluate the stiffness of parotid and submandibular glands using elastography ultrasound and to correlate it with B-mode ultrasonographical, clinical and serological features, salivary profibrotic and inflammatory chemokines, and salivary gland fibrosis. METHODS: We performed B-mode and elastography ultrasound of major salivary glands of 26 patients with primary Sjögren's syndrome. We registered the shear wave velocity (SWV) and correlated it with the morphologic ultrasonographic changes assessed by the Hocevar scale. We assessed the European League Against Rheumatism (EULAR) Sjögren's Syndrome Disease Activity Index (ESSDAI), EULAR Sjögren's Syndrome Patient Reported Index (ESSPRI), non-stimulated whole salivary flow rate (NSWSF), C3 and C4 levels, anti-Ro/La antibodies, salivary inflammatory (C-X-C motif ligand 13 [CXCL13], CXCL10, CXCL8, C-C motif ligand 2 [CCL2], interleukin 10 [IL-10] and IL-6) and pro-fibrotic (CXCL14, CCL28, tumor necrosis factor-related apoptosis-inducing ligand and transforming growth factor ß) chemokines and cytokines and evaluated the presence of fibrosis in the minor salivary gland. RESULTS: Ninety-two percent of patients were women; mean age was 51.1 ± 11 years; median disease duration was 6.1 years; 92.3% had oral symptoms and 26.9% fibrosis. The median B-mode score was 22.2 points and the median SWV 2.5 m/s (τ = 0.53, P = 0.001). The SWV correlated with the NSWSF (τ = -0.53, P = 0.001), ESSDAI (τ = 0.31, P = 0.03), glandular ESDDAI domain (τ = 0.36, P = 0.02), C4 levels (τ = -0.32, P = 0.04), salivary CXCL13 (τ = 0.29, P = 0.03) and CXCL10 (τ = 0.30, P = 0.003), but not with age and fibrosis. CONCLUSION: WV correlated with the B-mode ultrasound score, systemic and glandular activity and in a large degree with CXCL10, an inflammatory chemokine, but not with fibrosis. An increased SWV might represent chronic glandular inflammation rather than fibrotic changes in these patients.

C-reactive protein (CRP) polymorphisms and haplotypes are associated with SLE susceptibility and activity but not with serum CRP levels in Mexican population.

To evaluate the association of C-reactive protein (CRP) polymorphisms with risk of development SLE in a group of Mexican individuals. Five CRP polymorphisms (rs3093059, rs3093062, rs1800947, rs1130864, and rs1205) were determined by PCR-restriction fragment length polymorphism and SNP rs3093061 by refractory mutation system PCR assay in 126 SLE patients and 131 controls. Four of the polymorphisms showed differences between patients and controls. rs3093061 polymorphism was associated with a lower risk of developing lupus principally in the codominant 2 (OR = 0.219, 95% CI 0.108-0.785, P = 0.015) model. rs1130864 was associated with decreased risk mainly under codominant 1 (OR = 0.288, 95% CI 0.143-0.581, P = 0.001) model. rs1205 was associated under the over-dominant model (OR = 0.504, 95% CI 0.270-0.942, P = 0.032). The rs3091244 polymorphism was associated with decreased risk of SLE mostly under additive (OR = 0.605, 95% CI 0.393-0.931. P = 0.022) model. Our study establishes that rs3093061, rs1130864, rs1205, and rs3091244 polymorphisms are associated with decreased risk of developing SLE.

Vß T cell receptor (TCR) genes in circulating cells of patients with systemic lupus erythematosus and their healthy relatives / Genes del receptor variable beta de células T en células circulantes de pacientes con lupus eritematoso generalizado y sus familiares sanos.

Objective: We investigated the proportion of Vß T cell receptor (TCR) gene expression in peripheral CD3+ lymphocytes in familial and non-familial systemic lupus erythematosus (SLE) patients. Method: The Vß TCR repertoire was studied in 14 families in which several members had SLE. The Vß TCR usage in SLE patients (n = 27) was compared with that in healthy members of these multiplex families (n = 47), in 37 sporadic SLE patients who had no relatives with SLE, and in 15 healthy unrelated controls. Vß TCR repertoire expression was studied by multiparameter flow cytometry with the use of an array of 24 different Vß TCR gene family-specific monoclonal antibodies. Results: We found the same Vß TCR expression profile in the comparisons between sporadic SLE and familial SLE cases and healthy relatives, which included increased expression of Vß 5.2, Vß 11 and Vß 16, and lower expression of Vß 3, Vß 4, Vß 7.1 and Vß 17. Interestingly, solely Vß 17 was differentially expressed among sporadic and familial SLE. Also, increased expression of Vß 9 was the hallmark among familial SLE (casesand h ealthy relatives) in comparison to controls. Conclusion: These results highlight the notion that the final profile of the Vß TCR repertoire seen in familial and non-familial SLE seems to arise from the interaction of genetic, environmental, and immunoregulatory factors. Furthermore, it may contribute to the immunologic abnormalities affecting relatives of SLE patients.

Objetivo: Se investigó la proporción de la expresión génica del receptor variable beta de células T (Vß TCR) en linfocitos periféricos CD3+ en pacientes con lupus eritematoso generalizado (LEG) familiar y no familiar. Método: El repertorio de Vß TCR se estudió en 14 familias que presentaban más de un miembro con LEG. El uso de Vß TCR en pacientes con LEG (n = 27) se comparó con el de los miembros sanos de estas familias (n = 47), con 37 pacientes con LEG esporádico y con 15 controles sanos. La expresión del repertorio de Vß TCR se estudió por citometría de flujo multiparamétrica utilizando un arreglo de 24 diferentes anticuerpos monoclonales específicos de genes familiares para Vß TCR. Resultados: Se encontró el mismo perfil de expresión en las comparaciones entre los casos de LEG esporádico y familiar, así como en los consanguíneos sanos de las familias multicasos, que incluía una expresión incrementada de Vß 5.2, Vß 11 y Vß 16, y una menor expresión de Vß 3, Vß4, Vß 7.1 y Vß 7. De manera interesante, solo Vß 17 se expresó de modo diferente entre casos familiares y esporádicos de LEG. Igualmente, la expresión incrementada de Vß 9 fue el distintivo entre los casos de LEG familiar (casos y consanguíneos sanos) y los controles sanos. Conclusiones: Estos resultados refuerzan la noción de que el perfil final del repertorio Vß TCR observado en LEG familiar y no familiar parece surgir de la interacción de factores genéticos, ambientales e inmunorreguladores, además de que pueden explicar las alteraciones inmunitarias que se observan en los consanguíneos sanos de pacientes con LEG.

Interleukin-27 polymorphisms are associated with premature coronary artery disease and metabolic parameters in the Mexican population: the genetics of atherosclerotic disease (GEA) Mexican study.

Several studies suggest an important role of Interleukin-27 in the development of atherosclerosis. The aim of this study was to establish whether the IL-27p28 gene polymorphisms are associated with premature coronary artery disease and/or other cardiovascular risk factors. Four IL-27p28 gene polymorphisms were selected and genotyped in 1162 premature coronary artery disease cases and 1107 controls. rs26528 T and rs40837 A alleles were significantly associated with a lower risk of premature coronary artery disease under different inheritance models (Pdominant = 0.046; Pover-dominant = 0.002; Pco-dominant1 = 0.007 for rs26528T; Pover-dominant = 0.008 and Pco-dominant1 = 0.031 for rs40837). The rs40837 A allele was also associated with a lower risk of insulin resistance, in cases (Pover-dominant = 0.037) and controls (Padditive = 0.008; Pdominant = 0.047; Precessive = 0.014; Pco-dominant2 = 0.006), while the rs26528 T allele was associated with a lower risk of insulin resistance only in the control group (Precessive = 0.016; Pco-dominant2 = 0.021). Interleukin-27 plasma levels were measured in 450 controls and 450 cases, and were significantly higher in cases compared to controls (P = 0.004). However, Interleukin-27 plasma levels were not associated with IL-27p28 polymorphisms. Luciferase assays showed that co-transfection of the rs40837 A allele and miR-379-5p significantly decreased luciferase gene expression. Our study shows for the first time, that IL-27p28 gene polymorphisms are associated with premature coronary artery disease and with some metabolic parameters. The rs40837 A allele in presence of miR-379-5p significantly decreased luciferase gene expression.

HIF1A (rs11549465) and AKNA (rs10817595) Gene Polymorphisms Are Associated with Primary Sjögren's Syndrome.

Objective. To evaluate the allele and genotype frequencies of polymorphic sites of HIF1A and ANKA genes in primary Sjögren's syndrome (pSS). Methods. We included 110 patients with pSS and 141 ethnically matched healthy controls. Three HIF1A gene polymorphisms (Pro582Ser, Ala588Thr, and C191T) and two AKNA gene polymorphisms (-1372C>A and Pro624Leu) were genotyped using TaqMan probes in a Real-Time PCR instrument. Associations between pSS and genotypes, alleles, and inheritance models of the SNPs of interest were evaluated by logistic regression adjusted by age and gender. Results. The C/T genotype and the T allele of the HIF1A Pro582Ser polymorphism protected against pSS (OR = 0.22; 95% CI = 0.09-0.52; P < 0.01; OR = 0.26; 95% CI = 0.12-0.58; P < 0.01, resp.), whereas under a recessive model adjusted by age and gender, the AKNA -1372C>A polymorphism A/A genotype was associated with an increased risk of pSS (OR = 2.60; 95% CI = 1.11-6.12; P = 0.03). Conclusions. We identified HIF1A Pro582Ser T allele and C/T genotype as well as AKNA -1372C>A polymorphism A/A genotype as genetic factors associated with pSS. Further studies in other populations are needed to validate our findings and research is warranted in order to shed some light on their functional implications across biological pathways in this disease.

The rs7044343 Polymorphism of the Interleukin 33 Gene Is Associated with Decreased Risk of Developing Premature Coronary Artery Disease and Central Obesity, and Could Be Involved in Regulating the Production of IL-33.

AIM: The effect of interleukin 33 (IL-33) in the inflammatory process generates significant interest in the potential significance of IL-33 as a biomarker for coronary artery disease (CAD). Here, our objective was to analyze whether IL-33 gene polymorphisms are associated with premature CAD in a case-control association study. METHODS: Four IL-33 polymorphisms (rs7848215, rs16924144, rs16924159 and rs7044343) were genotyped by 5' exonuclease TaqMan assays in 1095 patients with premature CAD and 1118 controls. RESULTS: The rs7044343 T allele was significantly associated with a diminished risk of premature CAD (OR = 0.81, 95% CI: 0.69-0.97, Pdom = 0.020; OR = 0.85, 95% CI: 0.75-0.96, Padd = 0.019) and central obesity (OR = 0.74, 95% CI: 0.58-0.93, Pdom = 0.0007), respectively. When patients were divided into groups with and without type 2 diabetes mellitus (T2DM), the rs7044343 T allele was associated with a reduced risk of premature CAD in patients without (OR = 0.85, 95% CI: 0.73-0.99, Padd = 0.038) and with T2DM (OR = 0.61, 95% CI: 0.38-0.97, Pdom = 0.039; OR = 0.69, 95% CI: 0.49-0.97, Padd = 0.035). In order to establish the functional effect of the rs7044343 polymorphism, the production of IL-33 was determined in monocytes of selected individuals. Monocytes from individuals with rs7044343 CC genotype produced higher levels of IL-33 than monocytes from individuals with other genotypes. CONCLUSION: The results suggest that the IL-33 rs7044343 T allele could be a susceptibility marker for premature CAD and central obesity. The rs7044343 polymorphism could be involved in regulating the production of IL-33.

[Rheumatoid arthritis (RA) disease activity predicts function of ABCB1 (P-gp) and ABCG2 (BCRP1) drug-efflux transporters]. / La actividad funcional de los transportadores ABCB1 (P-gp) y ABCG2 (BCRP1) en pacientes con artritis reumatoide (AR) está determinada por la actividad de la enfermedad (AE).

BACKGROUND: P-gp and BCRP1 are transporter proteins that may confer drug resistance. OBJECTIVE: To compare P-gp and BCRP1 function in rheumatoid arthritis patients with active and inactive disease and to define their relation with disease activity. METHODS: We included 17 active patients paired (age, gender, disease duration) to 17 inactive patients. All had baseline evaluations and 27 had additional six-month follow-up. P-gp and BCRP1 functional activity was measured in peripheral mononuclear cells by flow cytometry. Percentage of lymphocytes able to extrude substrates for P-gp and BCRP1 were recorded in the presence/absence of selective inhibitors. Informed consent was obtained. Descriptive statistics and linear regression model were applied. RESULTS: Active patients had higher efflux function of both transporters than inactive patients: median (25-75 IQR) P-gp of 7.1% (1.4-29.3) vs. 1.6% (0.7-3.5), p = 0.02 and BCRP1 of 6.2% (1.3-22.4) vs. 1.3% (0.7-2), p = 0.007. At baseline, disease activity was the only predictor of both transporter functions. At follow-up, changes in disease activity correlated with shift in P-gp (r = 0.35, p = 0.07) and BCRP1 (r = 0.33, p=0.09) function. CONCLUSIONS: Patients with active rheumatoid arthritis had a higher efflux function of P-gp and BCRP1 compared to inactive patients. The behavior of P-gp and BCRP1 appeared to be conditioned by disease activity.

Rheumatoid Arthritis Disease Activity Is Determinant for ABCB1 and ABCG2 Drug-Efflux Transporters Function.

OBJECTIVE: To compare drug efflux function of ABCB1 and ABCG2 transporters in rheumatoid arthritis (RA) patients with active disease and in remission. METHODS: Twenty two active RA patients (DAS28 ≥3.2) and 22 patients in remission (DAS28<2.6) were selected from an early RA clinic. All patients were evaluated at study inclusion and six months later. ABCB1 and ABCG2 functional activity was measured in peripheral lymphocytes by flow cytometry. The percentage of cells able to extrude substrates for ABCB1 and ABCG2 was recorded. RESULTS: Active patients had higher ABCB1 and ABCG2 activity compared with patients in remission (median [interquartile range]): 3.9% (1.4-22.2) vs (1.3% (0.6-3.2), p = 0.003 and 3.9% (1.1-13.3) vs 0.9% (0.5-1.9) p = 0.006 respectively. Both transporters correlated with disease activity assessed by DAS28, rho = 0.45, p = 0.002 and rho = 0.47, p = 0.001 respectively. Correlation was observed between the time from the beginning of treatment and transporter activity: rho = 0.34, p = 0.025 for ABCB1 and rho = 0.35, p = 0.018 for ABCG2. The linear regression model showed that DAS28 and the time from the onset of treatment are predictors of ABCB1 and ABCG2 functional activity, even after adjustment for treatment. After six months we calculated the correlation between change in DAS28 and change in the functional activity in both transporters and found a moderate and significant correlation for ABCG2 (rho = 0.28, p = 0.04) and a non-significant correlation for ABCB1 (rho = 0.22, p = 0.11). CONCLUSIONS: Patients with active RA have an increased function of ABCB1 and ABCG2, and disease activity is the main determinant of this phenomena.

Absence of salivary CCL28 in primary Sjögren's syndrome.

CCL28 is a mucosa-associated epithelial-cell-produced chemokine involved in oral defense. We assessed the level of CCL28 in saliva of primary Sjögren's syndrome (pSS) patients in comparison with healthy controls and correlated it with IgA salivary levels. We included 30 non-smoker pSS patients and 30 non-smoker healthy controls paired by age (±5 years). Saliva samples were collected during the morning and kept frozen at -86 °C until the analysis. Fifty microliters of saliva was diluted 3:1 with water and analyzed for CCL28 salivary levels by ELISA method. The samples were tested in triplicate. IgA salivary levels were tested by ELISA method. We used descriptive statistics, Mann-Whitney U test and Kendall's tau correlation coefficients. pSS patients were mostly females (93.3 %), mean age 54.5 ± 13.3 years and median disease duration of 7.6 years (0.5-33). Patients with pSS had lower levels of salivary CCL28 when compared with controls [0 (0-1,272 pg/ml) vs. 94.4 (0-5,810) pg/ml, p < 0.0001]. pSS patients also had lower median levels of salivary IgA [72.55 µg/ml (0.40-297.4)] than controls [131.9 µg/ml (6.8-281.8)], although the latter results did not reach statistical significance (p = 0.51). Among the SS group, there was no correlation between CCL28 and IgA salivary levels nor between salivary IgA and disease duration, salivary flow, serum immunoglobulins or dental loss. CCL28 was absent in saliva of pSS patients; however, this finding did not correlate with salivary IgA levels.