A pilot study of childhood-onset Takayasu arteritis using whole exome sequencing suggests oligogenic inheritance involving classical complement, collagen, and autoinflammatory pathways.

Takayasu arteritis (TA) is a chronic granulomatous inflammatory disease affecting the aorta and its branches. Paediatric TA (pTA) may present from 6 months after birth till the adolescent age group. Genetics and pathogenesis of pTA are not fully understood. Earlier studies reported monogenic mutation in NOD2, XIAP, and STAT1 genes in patients with pTA. TA, a relatively rare disease, is more common in geographical pockets, including India. We hypothesized that South Asian patients with pTA, namely, those of Indian subcontinent origin, may have clinically relevant and unique pathogenic variants involving one or more genes, especially those linked to genetically driven vasculitic illnesses, including autoinflammatory pathologies. Children with pTA fulfilling EULAR/PRINTO/PReS classification criteria and presenting with clinical symptoms to the Paediatric Rheumatology clinic of Christian Medical College, Vellore, were included. Blood samples were collected after getting informed consent from parents or guardians and assent forms from children. DNA was extracted from whole blood using the Qiagen DNA extraction kit. Initially, the common variant in Indian population, namely, ADA2 c.139G > A; p.Gly47Arg, was screened, followed by whole exome sequencing. Fourteen children were recruited for the study. Median age of patients was 11 years (4 months-14 years) with a male-to-female ratio of 4:10. Distribution of angiographic subsets by Numano's classification of included children were as follows: type 5 (n = 7), type 4 (n = 5), and type 3 (n = 2). We identified novel variants in ten different genes. This include variants in genes of classical complement pathway, namely, C2, C3, C6, C7, and C9, and other genes, namely, CYBA, SH3BP2, GUCY2C, CTC1, COL5A1, and NLPR3. Two of 14 patients have heterozygous pathogenic variants; this implies that combination of heterozygous variants in C3 and COL5A1 might lead to disease development, suggesting digenic inheritance. One patient has a homozygous variant in CYBA. None of the patients were identified to have ADA2 variants. Whole exome sequencing reveals combination of rare variants in genes C3, COL5A1, and CYBA associated with disease development in children with Takayasu Arteritis. Key Points • We identified novel variants in genes of classical complement pathway, namely, C2, C3, C6, C7, and C9, and other genes, namely, CYBA, SH3BP2, GUCY2C, CTC1, COL5A1, and NLPR3. • Two of 14 patients have heterozygous pathogenic variants in C3 and COL5A1; this may have implications in disease development, suggesting digenic inheritance. • One patient has homozygous variant in CYBA. • None of the patients were identified to have ADA2 variants.

Gut microbiota of preterm infants in the neonatal intensive care unit: a study from a tertiary care center in northern India.

Introduction: Disruptions of the gut microbiota of preterm infants admitted to the neonatal intensive care unit (NICU) during the first 2 weeks of life are of critical importance. These infants are prone to various complications, including necrotizing enterocolitis (NEC) and sepsis. Studying the gut microbiota will improve outcomes in preterm infants. In the present study, we examined the gut microbiota of preterm infants admitted to the NICU in the first month of life. Methods: Neonates admitted to the NICU were recruited, and stool samples were collected weekly from the seventh day of the infant's life until the 30th day of life. DNA was extracted using a DNeasy Powersoil DNA isolation kit. 16S rRNA gene sequencing targeting the V3-V4 region was performed using the MiSeq platform. Sequenced reads were processed on DADA2 pipeline to obtain an amplicon sequence variant (ASV) table. All bioinformatic and statistical analyses were performed using different packages in the R statistical framework. Results: Fourteen preterm infants were recruited, and 48 samples were collected. Alpha diversity metrics, observed ASV count, and Shannon index were found to have no differences in any clinical variables. Permutational multivariate analysis of variance (PERMANOVA) showed discrimination of neonates by gestational age and administration of probiotics. Differential abundance analysis showed a decreased abundance of Bifidobacterium Breve in extremely preterm infants (gestational age <28 weeks) compared to moderate preterm infants (gestational age 29-32 weeks). Supplementation with probiotics decreased Acinetobacter and increased Bifidobacterium in the gut of preterm neonates regardless of gestational age. Conclusion: Gestational age and probiotic supplementation alter the gut microbiota of preterm infants admitted to the NICU.

Study of pathogenic T-helper cell subsets in Asian Indian patients with Takayasu arteritis.

The relapses and refractory disease are a challenge in the management of patients with Takayasu arteritis (TAK). We quantified pathogenic CD4 + memory T helper cells bearing surface markers CD161 and/or p-glycoprotein (MDR1) in patients with TAK. Peripheral blood mononuclear cells of 21 patients with TAK and 16 age-matched controls were stained with anti-CD3, anti-CD4, anti-CD45RA, anti-CD161 and anti-p-glycoprotein antibodies and subjected to flow cytometry by FACS ARIAIII. Eighteen patients underwent follow-up immunophenotyping. Intracellular staining for interleukin-17 and interferon-γ was performed for 18 patients and 11 controls. Surgical arterial biopsies of 6 TAK and 5 non-inflammatory controls were subjected to immunohistochemistry with anti-CD161 and anti-p-glycoprotein. At baseline the frequency of MDR1 + CD4 + and CD161 + MDR1 + CD4 + memory T cells was higher in TAK than controls (p = 0.002 and 0.01, respectively). After stimulation, the frequency of IFN-y + CD161 + cells was higher in TAK than controls (p = 0.028). Modal fluorescence intensity of CD161 + MDR1 + CD45RA - CD4 + cells was higher in active as compared with stable disease (p = 0.041). At 6 months, MDR1 + and CD161 + MDR1 + memory CD4 + T cells decreased significantly only in patients who had complete/partial response to treatment (p = 0.047 and 0.02, respectively). To conclude, MDR1 + and MDR1 + CD161 + CD4 + memory T-helper cells are increased in patients with TAK. These cells decreased only in patients with response to treatment during subsequent follow-up.

DNA methylation profiling of labial salivary gland tissues revealed hypomethylation of B-cell-related genes in primary Sjögren's syndrome.

The objective of this epigenetic study was to investigate the cellular proportions based on DNA methylation signatures and pathways of differentially methylated genes in labial salivary gland (LSG) tissues of individuals with Sjögren's syndrome (SS). Two methylation array datasets from the Gene Expression Omnibus repository (GSE166373 and GSE110007) were utilized, consisting of 159 LSG tissues from 77 SS cases and 82 non-SS controls. The raw data underwent analysis using the Chip Analysis Methylation Pipeline (ChAMP) in R statistical tool, which identified differential methylation probes and regions. The EpiDISH and minfi packages in R were employed to identify proportions of epithelial cells, fibroblasts, and immune cells, as well as immune cell subsets. The results showed that proportions of immune cells were increased, while proportions of epithelial cells and fibroblasts were significantly decreased in the LSG of individuals with SS compared to non-SS controls. Specifically, proportions of B-cells and CD8 T-cells were increased, while CD4 T-cells, Treg, monocytes, and neutrophils were decreased in the LSG of individuals with SS. Pathway analysis indicated that genes involved in immune responses to Epstein-Barr virus infection were significantly hypomethylated in SS, and gene set enrichment analysis highlighted the hypomethylation of genes involved in the somatic recombination of immune receptors in SS. Additionally, Disease Ontology analysis showed enriched pathways related to multiple myeloma, arthritis, and the human immunodeficiency virus. The study also revealed significant hypomethylation of the WAS gene on chromosome X in LSG tissues of individuals with SS. Overall, the findings suggest an increased proportion of B-cells and genes related to B-cell function, as well as hypomethylation of genes involved in immune responses and immune receptor recombination, in LSG tissues of individuals with SS compared to non-SS controls.

Angiogenesis related genes in Takayasu Arteritis (TAK): robust association with Tag SNPs of IL-18 and FGF-2 in a South Asian Cohort.

We performed genetic association study for genes encoding angiogenic and angiostatic proteins in patients with Takayasu arteritis (TAK). A total of 96 SNPs involving 60 genes were studied. Genotyping was performed in Fluidigm 96.96 Dynamic Array chip. All statistical analysis for SNP evaluation was performed using PLINK software. Initial analyses revealed five SNPs from three genes [IL-18 (encodes Interleukin-18), FGF2 (encodes Fibroblast Growth Factor-2), and ANGPT1 (encodes Angiopoietin-1)] as significantly different between controls and cases (uncorrected p < 0.05). After permutation-based analysis, two tag SNPs on the promoter region of IL-18 (rs187238 and rs1946518) and one 3'UTR tag SNP (rs1476217) of FGF2 were significantly associated with susceptibility to TAK, with p and OR (95% CI) of 0.0006 and 1.64 (1.25-2.17), 0.03 and 1.28 (1.02-1.64) & 0.016 and 1.33 (1.05-1.67), respectively; while, the two tag SNPs of ANGPT1 gene (rs6469101 and rs16875900) showed a trend (p = 0.055 & p = 0.051, respectively after permutation based correction). There is robust linkage disequilibrium between the two tag SNPs of IL-18 gene as validated by 1000 genome data of South Asian population; the eQTL effects of these tag SNPs of IL-18 and FGF2 genes on adjacent genes further suggest that these tag SNPs act as genetic risks for development of TAK in South Asians, with possible functional implications towards future biomarker development. Genotype phenotype study by genetic model-based analysis also revealed associations between genotype subsets and clinical features like fever, visual loss, left subclavian and coronary artery involvement in our TAK patients.

Expanding the clinical and immunological phenotype of prolidase deficiency: A case report.

Prolidase deficiency (PD) is a rare autosomal recessive disorder associated with recurrent infections, immune dysregulation, and autoimmunity. PD is characterized by persistent dermatitis, skin fragility, and non-healing ulcerations on the lower limbs as its main dermatologic characteristics. Herein, we report a boy with PD due to a novel variant in PEPD who had abnormal facies, cognitive impairment, corneal opacity, recurrent infections, and persistent non-healing leg ulcers. Th17 lymphocyte counts and phosphorylated-STAT5 expression following IL-2 stimulation were reduced in our patient as compared to healthy control.

Intravenous Immunoglobulin in Kawasaki Disease-Evolution and Pathogenic Mechanisms.

Kawasaki disease (KD) is an acute vasculitis of childhood that affects the medium vessels with a special predilection to the involvement of coronary arteries. The major morbidity of this disease is due to coronary artery aneurysm, which occurs in about 25-30% of untreated cases. For decades now, intravenous immunoglobulin (IVIg) has consistently been shown to reduce the risk of CAAs to less than 5%. However, the mechanism of immunomodulation remains unclear. Several studies on the role of IVIg in the modulation of toll-like receptor pathways, autophagy, and apoptosis of the mononuclear phagocytic system, neutrophil extracellular trap, and dendritic cell modulation suggest a modulatory effect on the innate immune system. Similarly, certain studies have shown its effect on T-cell differentiation, cytokine release, and regulatory T-cell function. In this review, we discuss the potential mechanisms underlying the immunomodulatory actions of IVIg in patients with Kawasaki disease. Furthermore, we provide a summary of the evidence regarding various infusion protocols and dosages utilized in the treatment of KD patients.

Comparison of Point-of-Care PT-INR by Hand-Held Device with Conventional PT-INR Testing in Anti-phospholipid Antibody Syndrome Patients on Oral Anticoagulation.

Evidence on agreement of point-of-care (POC) INR testing with laboratory testing in APS patients on oral anticoagulation (OAC), is scarce. This study assessed agreement of paired PT INR testing by a POC device vs. conventional platform-based laboratory test, in APS patients on OAC using a pre-determined definition of agreement. Simultaneous paired PT INR estimation in 92 APS patients was carried out, during October 2020-September 2021. POC INR was performed on capillary blood (pin prick) using the qLabs® PT-INR hand-held device, while laboratory INR estimation was performed using citrated blood (venepuncture) on STA-R Max Analyzer® using STA-NeoPTimal thromboplastin reagent®. Concordance was defined no greater than ± 30% (as per international standards ISO 17593:2007 guidelines) for each paired INR estimation. Agreement between the two was defined as ≥ 90% of paired INR measurements being concordant. 211 paired estimations were performed, within which 190 (90%) were concordant. Good correlation was seen between the 2 methods of INR estimation on Bland Altman plot analysis with an Intra-class correlation coefficient (95% CI) of 0.91(0.882, 0.932). Lab INR range > 4 (P = 0.001) was a significant predictor of higher variability between both methods of INR estimation. Lupus anti-coagulant, other anti-phospholipid antibodies (APL) or triple APL positivity did not result in any statistically significant variation in paired measurements. This study demonstrated good correlation between POC INR measurement and Lab INR estimation and agreement was ascertained between the 2 methods in APS patients on OAC.

Genome-Wide DNA Methylation Profiling in CD8 T-Cells and Gamma Delta T-Cells of Asian Indian Patients With Takayasu Arteritis.

Background: Takayasu's Arteritis (TA) is a chronic inflammatory disease that affects aorta and its main branches at their origin. Genetic, pathological and functional studies have shown that CD8 and Gamma delta (γ/δ) T-lymphocytes are involved in inflammatory processes in affected regions of arteries causing vascular damage. The molecular function of these lymphocytes remains unclear and currently no epigenetic studies are available in TA. We primarily performed genome wide methylation analysis in CD8 T cells and γδ T cells of patients with TA and compared with healthy controls. Methods: We recruited 12 subjects in each group namely TA patient and healthy controls. Blood samples were collected after obtaining informed written consent. CD8 T cells and γδ T cells were separated from whole blood. DNA extracted from these cells and were subjected to bisulfite treatment. Finally, bisulfite treated DNA was loaded in Infinium Methylation EPIC array. Bioinformatics analysis was used to identify differential methylation regions which were then mapped to genes. Results: Interleukin (IL)-32 and Lymphotoxin-A were genes significantly hypomethylated in CD8 T-cells. Anti-inflammatory cytokine genes, IL-10, IL-1RN and IL-27 were hypomethylated in γδ T cells of TA patients as compared to healthy controls. Gene enrichment analysis using Gene Ontology (GO) database and Kyoto Encyclopaedia of Genes and Genomes (KEGG) identified that genes involved in T-cell receptor signalling pathways were hypomethylated in CD8 T-cells and hypermethylated in γδ T cells of TA patients. Conclusion: CD8 T-cells might play a major role in immunopathogenesis of inflammation in TA, whereas γδ T cells may play a regulatory role.

High expression of pro-inflammatory cytokine genes IL-1ß and IL-1R2 upon TLR4 activation in Takayasu arteritis.

Toll-like receptors (TLR) 4 and its endogenous ligands are highly expressed in aorta. In the present study, we have explored the effect of TLR-4 activation by pro-inflammatory and angiogenic factors in PBMCs of patients with Takayasu Arteritis (TA). In the screening cohort, PBMCs of TA (n = 6) and healthy controls (n = 6) were stimulated with LPS and cultured. mRNA expression of 84 genes were quantitated by RT2 Profiler™ PCR Array kit in PBMCs. Validation set of additional PBMCs from TA (n = 7) and healthy controls [HC) (n = 7) were then stimulated with LPS to study expression of selected genes with delta Ct > 0.1 in the screening cohort. Significant gene expressions were correlated with Indian Takayasu arteritis activity scores (ITAS 2010). Increased expression of CCL2 was observed only in unstimulated PBMCs of patients with TA [median relative difference (RD) of 2.37] as compared to HC (RD 1.37, p < 0.03) in validation cohort, while stimulation with TLR4 ligand led to increased mRNA expression of IL-1ß (RD 7.9, p < 0.028) and IL-1R2 (RD 0.08 p < 0.013) genes as compared to that of HC [RD of 5.32 for IL-1ß and 0.01 for IL-1R2, respectively] in validation cohort. TLR4 activation also led to significantly higher expression of HPSE, TIMP1 and low expression of VEGFB, S1PR1, SERPINF1, ANGPLT4, ANGPT2, TIE1 and NOS3 genes in the screening cohort. But expression of VEGFB was not significant in validation cohort. The significant gene expressions, however, did not correlate with ITAS [ITAS2010 and ITAS-A (CRP)]. TLR4 activation leads to increased expression of IL-1ß and IL-1R2 genes in PBMCs of patients with TA.

In vitro effects of curcumin on proinflammatory cytokines and expression of their genes in minor salivary gland tissue of patients with Sjogren's syndrome.

Curcumin reduces disease severity and ameliorates lupus-like/Sjögren's Syndrome-like disease in mice model. The immunological basis of these effects is largely unknown. This study examined the effects of curcumin on pro-inflammatory cytokines secreted by minor salivary glands in patients with primary Sjögren's syndrome (pSS). Minor salivary gland (MSG) tissue samples were collected from patients undergoing biopsy for suspected pSS. The tissues were treated with phytohemagglutinin (PHA) alone as well as PHA with curcumin (30 µM) and cultured in RPMI 1640 medium for 48 h at 37 °C in CO2 incubator. After the incubation period, culture supernatant and tissues were stored in the freezer (-80 °C). IL-6 levels were measured in supernatant by ELISA kit. Gene expressions of pro-inflammatory cytokines, namely IL-6, IL-8, TNF-α, IL-1ß, IL-4, IL-10, IL-17, IL-21, and IFN-γ, were measured by qPCR. IL-6 secretion levels and gene expressions were compared statistically between groups by Student's t test. Forty-seven patients were screened. Eight patients satisfied ACR/EULAR criteria for pSS. Seven patients with absent glandular inflammation and negative serology constituted sicca controls. These 15 subjects were included in final analysis. In pSS group, but not in controls, median IL-6 levels in supernatant were less in curcumin-treated as compared to PHA-alone subset [5.5 (0.7-13.34) vs 18.3 (12-32) ng/ml; p = 0.0156]. mRNA expression levels of IL-6 were also lower in curcumin-treated samples as compared to PHA alone, when cases and controls were analyzed together as well as in cases alone (p = 0.0009 and p = 0.0078, respectively); however, mRNA expression of IL-1ß was lower in curcumin-treated samples as compared to PHA alone, only when cases and controls were analyzed together (p = 0.0215). There was no difference in other cytokine gene expression levels between the subsets under the in-vitro experimental conditions. In conclusion, curcumin reduced mRNA expression as well as secretion of IL-6 levels by salivary gland tissue of patients with pSS. Curcumin also suppressed PHA-induced mRNA expression levels of IL-6 and IL-1ß in MSG tissue of patients with pSS and controls when analyzed together as a combined group.

Salivary free light chains and salivary immunoglobulins as potential non-invasive biomarkers in primary Sjögren's syndrome.

BACKGROUND: B cells contribute significantly to the pathogenesis of primary Sjögren's syndrome (pSS). Free light chains (FLCs) are generated during the production of immunoglobulins (Igs) and are surrogates of B cell activity. We hypothesized that salivary FLCs and salivary Igs could represent salivary gland inflammation and therefore, serve as biomarkers in pSS. METHODS: Patients >18 years old fulfilling the American College of Rheumatology / European League Against Rheumatism (EULAR) 2016 criteria for pSS and age-matched healthy and disease controls (sicca non-pSS, rheumatoid arthritis, systemic lupus erythematosus) were recruited for this cross-sectional study. FLCs in saliva and serum were measured by immunoturbidimetry. Serum and salivary Igs were measured by nephelometry and enzyme-linked immunosorbent assay, respectively. Area under the receiver operator characteristic curve was determined. The factors influencing the serum and salivary FLCs in pSS were determined using backward linear regression. RESULTS: A total of 78 patients with pSS, 76 healthy controls and 62 disease controls were recruited. Median EULAR SS disease activity index (interquartile range) was 1 (3.75). Serum FLCκ and FLCλ, salivary FLCλ, serum and salivary IgG, salivary IgM was significantly higher in the pSS group compared to the controls. Areas under the curve for salivary FLCλ, serum FLCκ, serum and salivary IgG were 0.75, 0.72, 0.78 and 0.77, respectively. Regression analysis showed that salivary FLCκ, salivary FLCλ and salivary IgG were associated with positive salivary gland histopathology. Use of immunosuppressants and glucocorticoids was associated with lower values of salivary parameters. CONCLUSION: Salivary FLCλ and salivary IgG were significantly different between pSS and control groups and could be potential non-invasive biomarkers in pSS. These findings should be confirmed in larger longitudinal studies.

Update on pregnancy in Takayasu arteritis-A narrative review.

Takayasu arteritis (TA) is a chronic, idiopathic large-vessel vasculitis that affects women of reproductive age, and has significant maternal and fetal implications. Although there are contrasting data on the effect of TA on fertility, most studies have shown that fertility outcomes remain unaffected. The disease activity of TA usually either remains stable or decreases during pregnancy. The important fetomaternal complications are maternal hypertension, pre-eclampsia, prematurity, and intrauterine growth restriction. To reduce maternal and fetal morbidity, controlling the disease before conception is important. This review article discusses the various implications, challenges, and medical and endovascular management of TA during pregnancy.

Severe COVID-19, multisystem inflammatory syndrome in children, and Kawasaki disease: immunological mechanisms, clinical manifestations and management.

Multisystem inflammatory syndrome (MIS-C) is a pediatric hyperinflammation disorder caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). It has now been reported from several countries the world over. Some of the clinical manifestations of MIS-C mimic Kawasaki disease (KD) shock syndrome. MIS-C develops 4-6 weeks following SARS-CoV-2 infection, and is presumably initiated by adaptive immune response. Though it has multisystem involvement, it is the cardiovascular manifestations that are most prominent. High titres of anti-SARS-CoV-2 antibodies are seen in these patients. As this is a new disease entity, its immunopathogenesis is not fully elucidated. Whether it has some overlap with KD is still unclear. Current treatment guidelines recommend use of intravenous immunoglobulin and high-dose corticosteroids as first-line treatment. Mortality rates of MIS-C are lower compared to adult forms of severe COVID-19 disease.

Understanding immunopathological fallout of human coronavirus infections including COVID-19: Will they cross the path of rheumatologists?

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection causing coronavirus disease 2019 (COVID-19) is the biggest pandemic of our lifetime to date. No effective treatment is yet in sight for this catastrophic illness. Several antiviral agents and vaccines are in clinical trials, and drug repurposings as immediate and alternative choices are also under consideration. Immunomodulatory agents like hydroxychloroquine (HCQ) as well as biological disease-modifying anti-rheumatic drugs (bDMARDs) such as tocilizumab and anakinra received worldwide attention for treatment of critical patients with COVID-19. This is of interest to rheumatologists, who are well versed with rational use of these agents. This brief review addresses the understandings of some of the common immunopathogenetic mechanisms in the context of autoimmune rheumatic diseases like systemic lupus erythematosus (SLE) and COVID-19. Apart from demographic comparisons, the role of type I interferons (IFN), presence of antiphospholipid antibodies and finally mechanism of action of HCQ in both the scenarios are discussed here. High risks for fatal disease in COVID-19 include older age, metabolic syndrome, male gender, and individuals who develop delayed type I IFN response. HCQ acts by different mechanisms including prevention of cellular entry of SARS-CoV-2 and inhibition of type I IFN signaling. Recent controversies regarding efficacy of HCQ in management of COVID-19 warrant more studies in that direction. Autoantibodies were also reported in severe acute respiratory syndrome (SARS) as well as in COVID-19. Rheumatologists need to wait and see whether SARS-CoV-2 infection triggers development of autoimmunity in patients with COVID-19 infection in the long run.

Study of clinical utility of antibodies to phosphatidylserine/prothrombin complex in Asian-Indian patients with suspected APS.

Antiphospholipid syndrome (APS) is the most common acquired pro-thrombotic disorder, also associated with obstetric complications. Phosphatidylserine/Prothrombin complex antibody (aPSPT) though associated with various APS manifestations, is not included in the revised Sapporo Criteria. To study the prevalence of aPSPT in Asian-Indian patients with suspected APS and compare its performance with the criteria anti-phospholipid antibodies (APLs). Electronic charts of 372 individuals whose sera was tested for aPSPT in suspected APS between June 2014 and May 2016 were retrieved and analyzed. aPSPT was assayed by ELISA. aPSPT tested individuals were categorized into cases-seropositive and seronegative APS (SNAPS) and controls. aPSPT was positive in 24/58 (41.3%) cases and 17/314 (5.4%) controls (p < 0.001). aPSPT positivity was seen in 44.5%, 38.7%, and 58.4% in primary, secondary and SNAPS patients respectively. aPSPT had the best performance among all APLs, in obstetric APS with 31% sensitivity, 97.7% specificity, and an odds ratio of 18.8. It showed 41.4% sensitivity, 94.6% specificity for the classification/diagnosis of primary APS and 38.7% sensitivity, 91.5% specificity for secondary APS. Addition of aPSPT to current APS criteria to SNAPS patients led to reclassification of additional 12.1% patients as APS overall and 42.8% in obstetric APS category. In Asian-Indian patients with suspected APS, aPSPT outperformed all classical APLs in diagnosis/classification of obstetric APS and both isotypes of beta 2-glycoprotein-I antibodies in diagnosis/classification of APS. aPSPT could reclassify additional 12.1 and 42.8% patients as APS overall and obstetric APS respectively, over and above the cases satisfying revised Sapporo criteria.

High expression of S100 calgranulin genes in peripheral blood mononuclear cells from patients with Takayasu arteritis.

BACKGROUND: Toll-like receptors (TLR) 1 to 4 are highly expressed in aorta. Activation of TLR4 causes transmural arteritis in Human temporal artery-SCID chimera model. Neither TLR-4 nor its ligands have been studied in TA patients as yet. Aim of this study was to examine the expression of TLR4 and its endogenous ligands in peripheral blood mononuclear cells (PBMCs) of patients with Takayasu arteritis (TA). METHODS: mRNA expression of TLR4, RAGE and various endogenous TLR4 ligands were quantified in PBMCs of 24 TA patients and 19 sex and age matched healthy controls by real time PCR using specific primers and SYBR Green qPCR master mix. S100A8/A9 and S100A12 were measured in cell culture supernatant of PBMCs from TA patients and healthy controls, both in un-stimulated state as well as, after lipopolysaccharides (LPS) stimulated cultures for 4 h. Expression of S100A8/A9 in aortic tissues was assessed by immunohistochemistry. RESULTS: The mRNA expression of S100A8, S100A9, S100A12 and TLR4 were higher, while expression of RAGE and HSP70 were lower in TA as compared to healthy controls. Induction with LPS led to increase in secretion of both S100A8/A9 and S100A12 levels in TA as well as healthy controls. The fold of induction, measured by LPS stimulated/unstimulated control was higher in healthy controls [2.88 (1.7-3.53) fold] as compared to TA [1.345 (1-1.82) fold]; p < 0.05. Numerically, S100A8/A9 was also higher in healthy controls [2.04 (1.7-5.6) fold] as compared to TA [1.38 (1.09-3.6) fold], but it didn't reach statistical significance; p = 0.129. Mild to moderate intensity expression of S100A8/A9 protein was noted in aortic tissues from patients with TA. CONCLUSION: mRNA expression of TLR4 and its ligand S100A8, S100A9, and S100A12 in PBMCs of TA patients was higher as compared to healthy controls. LPS stimulation led to higher induction of S100A12 secretion in healthy controls as compared to TA. Expression of S100A8/A9 was detected in inflamed aortic tissues from patients with TA.

High Secretion of Interleukin-6 and Increased MINCLE Receptor Expression Upon Exposure to Mycobacterial Cord Factor Analog Trehalose-6, 6-Dibehenate (TDB) in Patients with Takayasu Arteritis.

INTRODUCTION: Suspicion on the association between Takayasu Arteritis (TA) and Tubcerculosis (TB) has been in vogue for years. Prevalence of TB in TA is reported to be higher. We aimed to study innate immune responses in patients with TA on exposure to Trehalose-6,6-dibehenate (TDB), a synthetic analogue of Trehalose-6,6-Dimycolate (TDM, also known as mycobacterial cord factor) in comparison with healthy controls. MATERIALS AND METHODS: Patients with type V TA, satisfying 1990 ACR criteria, and age and sex matched healthy controls were recruited. PBMCs were cultured with 5µg/ml, 50µg/ml or without any TDB for 48 hours in RPMI medium inside a 5% Co2 incubator. IL-6, TNF-α and IL-17 were measured in cell culture supernatant, which was separated from the cells at the end of the incubation period. Gene expressions of IL-6, IL-8, TNFα, IFN-γ, MINCLE and BCL-10 were quantified in real time PCR using specific primers and SYBR green chemistry. RESULTS: Twenty two TA patients and 21 healthy controls were recruited. Both patients and controls showed response by secreting IL-6 and TNF-α upon stimulation by TDB. Relative induction (TDB stimulated TA sample / unstimulated control) of IL-6 was significantly higher in TA [31.88(0.74-168)] patients as compared to healthy controls [1.931(0.644-8.21); p<0.002], when co-cultured with 50µg/ml TDB. The expression of MINCLE, the TDB receptor was higher in TA samples than healthy controls upon TDB stimulation. CONCLUSION: Stimulation with mycobacterial synthetic analogue led to higher secretion of IL-6 and higher expression of MINCLE in PBMCs of patients with TA as compared to healthy controls.