Prognostic significance of circulating microparticles in IgA nephropathy.

PURPOSE: Endothelial injury, involved in the pathogenesis of renal fibrosis, can generate microparticles (MPs). These are 0.1-1 µm membrane-bound vesicles shed from the damaged or activated cell surfaces. We analyzed the presence of circulating MPs and EnMPs in IgAN and correlated with markers of endothelial injury and disease activity. METHODS: The study included 30 IgAN (mean age 31.5 ± 9 years), 25 healthy controls and Lupus nephritis (n = 10) as disease controls. Circulating MPs were quantitated by Flow cytometry and EnMPs were analyzed using anti-CD31-FITC and anti-CD146-PE antibodies. Their levels were correlated with serum von Willebrand Factor, histological Oxford MEST-C score and renal outcome. A prospective validation group of 20 patients of biopsy-proven IgA nephropathy was also included. RESULTS: IgAN had significantly higher levels of MPs, EnMPs and vWF compared to controls. On multivariate analysis, plasma levels of total MPs, EnMPs and serum vWF correlated significantly with the presence of hypertension and E1 on histology. E1 and high MPs (> 130 counts/µl) were associated with shorter time to doubling of serum creatinine. MPs cutoff level of 130 counts/µl had a sensitivity of 75%, specificity of 93.3% and diagnostic accuracy of 89.5% for E1 in the validation cohort. CONCLUSION: Circulating MPs and EnMPs in IgAN correlate with E1 on histology and have a potential as non-invasive biomarkers to predict disease activity and renal outcome.

Clinical metabolomics by NMR revealed serum metabolic signatures for differentiating sarcoidosis from tuberculosis.

BACKGROUND: Pulmonary sarcoidosis (SAR) and tuberculosis (TB) are two granulomatous lung-diseases and often pose a diagnostic challenge to a treating physicians. OBJECTIVE: The present study aims to explore the diagnostic potential of NMR based serum metabolomics approach to differentiate SAR from TB. MATERIALS AND METHOD: The blood samples were obtained from three study groups: SAR (N = 35), TB (N = 28) and healthy normal subjects (NC, N = 56) and their serum metabolic profiles were measured using 1D 1H CPMG (Carr-Purcell-Meiboom-Gill) NMR spectra recorded at 800 MHz NMR spectrometer. The quantitative metabolic profiles were compared employing a combination of univariate and multivariate statistical analysis methods and evaluated for their diagnostic potential using receiver operating characteristic (ROC) curve analysis. RESULTS: Compared to SAR, the sera of TB patients were characterized by (a) elevated levels of lactate, acetate, 3-hydroxybutyrate (3HB), glutamate and succinate (b) decreased levels of glucose, citrate, pyruvate, glutamine, and several lipid and membrane metabolites (such as very-low/low density lipoproteins (VLDL/LDL), polyunsaturated fatty acids, etc.). CONCLUSION: The metabolic disturbances not only found to be well in concordance with various previous reports, these further demonstrated very high sensitivity and specificity to distinguish SAR from TB patients suggesting serum metabolomics analysis can serve as surrogate method in the diagnosis and clinical management of SAR.

Renal injury, biomarkers, and myositis, an understudied aspect of disease: prospective study in the MyoCite cohort.

Introduction: The mechanisms leading to chronic kidney disease (CKD) in patients with idiopathic inflammatory myopathies (IIMs) are poorly understood. We assessed the prevalence of subclinical renal injury in patients with IIMs, through elevation in biomarker levels of tubular injury and fibrosis (NGAL, KIM1, Activin A, CD163, and Cys-c), and assessed differences between subtypes of IIMs, and the effect of disease activity and duration. Materials and methods: Clinical data, core set measures, sera and urine were prospectively collected from all patients enrolled in the MyoCite cohort from 2017 to 2021. Twenty healthy subjects (HC) and 16 patients with acute kidney injury (AKI) were included as controls. Baseline and follow up data for IIMs were included. Enzyme-linked immunosorbent assay (ELISA) was used to measure urine NGAL (Human Lipocalin-2/NGAL Duoset ELISA, Cat no: DY1757), KIM1 (Human TIM-1/KIM 1/HAVCR Duoset ELISA, Cat.no: DY1750B), Activin A (Human Activin A Duoset ELISA, Cat no: DY338), CD163 (Human CD163 Duoset ELISA,Cat no: DY1607-05), and Cys-c (Human Cystatin C Duoset ELISA, Cat. no.: DY1196) levels, while eGFR (unit mL/min/1.73 m2) was calculated by the Cockcroft-Gault formula and CKD-EPI formula. Results: Analysis of 201 visits of 110 adult patients with IIMs indicated higher normalized biomarker levels compared to HCs, and comparable to patients with AKI, with the exception of NGAL, which was higher in the AKI group. Notably 72 (49%) patients with IIMs had eGFR<90; the levels of the 5 biomarkers were comparable between active and inactive IIMs, and different subtypes of IIMs. Similarly, a poor correlation between urine biomarker levels and core set measures of activity and damage was found. Changes in biomarker levels on follow-up did not correlate with eGFR changes. Discussion: This exploratory analysis of urinary biomarkers identified low eGFR and elevated biomarkers of CKD in nearly half of the patients with IIMs, comparable to patients with AKI and higher than HCs, indicative of potential renal damage in IIMs that may have a lead to complications in other systems.

Response Guided Slow Infusion of Albumin, Vasoconstrictors and Furosemide Improves Ascites Mobilization and Survival in Acute on Chronic Liver Failure: A Proof-of-Concept Study.

Background and Aims: Acute-on-chronic liver failure (ACLF) with increasing organ failure is associated with poor outcomes. Severely deranged systemic hemodynamics and decreased effective arterial blood volume contribute to tissue damage and organ failure. Response-guided therapy with albumin, vasoconstrictors, and furosemide may help overcome effective hypovolemia, improve diuresis and impact survival. Methods: In the observation cohort, 230 patients with ACLF (CANONIC criteria) with ascites (≥Grade II) and ACLF ≥Grade I were enrolled. A total of 136 patients (GROUP I) received response-guided (urine sodium >80mmol/day) slow albumin-furosemide infusion ±â€…terlipressin (SAFI ± T), while 94 patients (GROUP II) received standard medical therapy. Twenty-eight-day survival, ascites mobilization (nil or grade 1), and adverse events were noted. In another mechanistic cohort (n = 40), laboratory evidences for improvement in various pathophysiological alterations; gut permeability, endotoxemia, cytokine storm, neutrophil dysfunction, and hemodynamic alterations following SAFI ± T/Noradrenaline (NAdr) were evaluated. Results: Age, gender, CLIF-C-ACLF, SOFA and MELD scores, ACLF grades and urine sodium were not different between the two groups in the observation cohort. Ascites was mobilized in 102/136 in GROUP I (SAFI ± T) and 23/94 in GROUP II (p < 0.05). Twenty-eight-day survival was significantly higher in GROUP I = 103/136 (75.7%) vs GROUP II = 50/94 (53.2%), (P = <0.001). All those who were unable to reach urine sodium >80 mmol/day died. Four patients in GROUP I developed scrotal gangrene. In the mechanistic cohort, 72% of patients survived with significant improvement in gut permeability, endotoxemia, serum cytokines, neutrophil dysfunction, and hemodynamic alterations. Conclusion: Ascitic fluid mobilization by response-guided SAFI ± T/NAdr therapy improves survival by improving splanchnic and systemic hemodynamics, decreasing gut congestion, gut permeability, and endotoxemia, improving neutrophil functions, and reducing pro-inflammatory cytokines in circulation.

The comparison of cardiovascular disease risk prediction scores and evaluation of subclinical atherosclerosis in rheumatoid arthritis: a cross-sectional study.

OBJECTIVES: Primary objectives estimated prevalence of traditional cardiovascular disease (CVD) risk factors and compared different CVD risk prediction algorithms in an Indian rheumatoid arthritis (RA) population. Secondary objectives evaluated associations between carotid intima-media thickness (CIMT) and subclinical atherosclerosis (SCA) with CVD risk factors and CVD risk scores. METHODS: The presence of CVD risk factors were recorded, and 10-year CVD risk was predicted using Framingham risk scoring (FRS) using lipids (FRS-Lipids), FRS using body mass index (FRS-BMI), QRISK-2, SCORE, and the algorithm recommended by ACC/AHA (ASCVD). CIMT was measured on the far-wall of the common carotid artery. Subclinical atherosclerosis was defined as CIMT > 0.9 mm or the presence of carotid plaque. RESULTS: A total of 332 patents were enrolled, 12% had diabetes mellitus, 21.4% hypertension, and 6.9% were current/past smokers. Proportions of RA with predicted 10-year CVD risk > 10% varied from 16.2 to 41.9% between scores. Highest magnitude of risk was predicted by FRS-BMI. Agreement between scores in predicting risk was moderate in general. Mean CIMT was 0.70 ± 0.15 mm. Age, male sex, and extra-articular manifestations associated with greater CIMT. All risk scores except SCORE moderately correlated with CIMT. About one-seventh had SCA defined as CIMT > 0.9 mm or the presence of carotid plaques, associated with increasing age, male gender, or higher ratio of total cholesterol to high-density lipoprotein cholesterol. ASCVD and QRISK-2 scores had maximum area under curve for distinguishing SCA. CONCLUSION: Individual CVD risk scores predict 10-year CVD risk differently in Indian patients with RA, and require validation for predicting hard end points (CVD events, mortality). Key Points • Diabetes mellitus and hypertension are the most prevalent cardiovascular disease risk factors in Indian patients with RA. • Individual cardiovascular risk prediction scores predict risk differently in Indian patients with RA, highest risk being predicted by the FRS-BMI. • Carotid intima-media thickness in RA associated with increasing age, male sex and extra-articular manifestations. • 14% RA had subclinical atherosclerosis, associated with increasing age, male sex, and higher total cholesterol to HDL-C ratio, best distinguished by ASCVD and QRISK-2 scores.

Anti-mitochondrial antibodies in Indian patients with idiopathic inflammatory myopathies.

AIMS: Anti-mitochondrial antibodies (AMAs) are associated with distinct clinical phenotypes including cardiac and hepatic manifestations in idiopathic inflammatory myopathies (IIMs). This article studies the prevalence, clinical characteristics and outcomes of AMA in Indian patients with IIM. METHODS: Patients (97: 81 adult, 16 juvenile) clinically diagnosed with polymyositis or antibody-negative IIM were retrieved from the MyoCite bio-archive. They were tested for myositis-specific autoantibodies / myositis autoantibodies (MSAs/MAAs) using line immunoassay and antinuclear antibodies and AMAs using immunofluorescence assay (IFA). Patients were also screened for cardiac biomarkers (cardiac troponin I [c-TnI] and N terminal-pro brain natriuretic peptide [NT-pro-BNP] using immunometric immunoassay technique and enhanced chemiluminescence assay testing respectively) and hepatic manifestations using AMA testing. Results were formulated after carrying out analytical tests. RESULTS: Of the cohort, 5 adults (6.2%) (M:F 0:1) with a median age and disease duration of 37 years and 2 months respectively, tested AMA+ while the children tested negative. Dermatomyositis was the commonest phenotype, with amyopathic forms being common, often with MSA positivity. Cancer-associated myositis and polymyositis were also seen. AMA positivity is associated with Gottron's sign and calcinosis. Comparable levels of C-TnI and NT-pro-BNP and AMA testing in patients help to rule out subclinical cardiac and hepatic involvement respectively. CONCLUSION: Anti-mitochondrial antibodies are rare (6.2%) in different subtypes of IIM in the Indian population, and often coexist with MSAs. Their negative association with cardiac and hepatic involvement and probable association with Gottron's sign and calcinosis merit further investigation and long-term follow-up to understand the entire spectrum of the disease.

Novel Th17 Lymphocyte Populations, Th17.1 and PD1+Th17, are Increased in Takayasu Arteritis, and Both Th17 and Th17.1 Sub-Populations Associate with Active Disease.

Purpose: We evaluated T helper lymphocyte profile, including novel Th17 subsets Th17.1 (secrete IFN-γ, associate with corticosteroid resistance) and PD1+Th17 (secrete TGF-ß1, implicated in fibrosis), and related cytokines in peripheral blood of Takayasu arteritis (TAK). Materials and Methods: We evaluated circulating Th1, Th2, Th17, Th17.1, PD1+CD4+ T lymphocytes, PD1+Th17, and Treg lymphocytes, inflammatory (IFN-γ, IL-4, IL-6, IL-17A, IL-23, IL-1ß, TNF-α) and regulatory (IL-10, TGF-ß1) cytokines in peripheral blood of TAK (n = 57; median age 35 (interquartile range 26-45) years; 40 females) in a cross-sectional design. We studied inflammatory and regulatory cytokines in culture supernatant of peripheral blood mononuclear cells (PBMCs) from TAK following stimulation with anti-CD3/anti-CD28 and their modulation by tacrolimus (immunosuppressive) with/without tadalafil (anti-fibrotic). Furthermore, we followed up immunosuppressive-naïve active TAK (n = 16) and compared T helper lymphocyte populations and cytokines before and after immunosuppressive therapy. Healthy controls (HC, n = 21) and sarcoidosis (disease control, n = 11) were compared against TAK. Results: TAK had higher Th17, Th17.1 and PD1+Th17 lymphocytes than HC (p < 0.001), and higher PD1+CD4+ T lymphocytes than sarcoidosis (p < 0.001). Th17 lymphocytes associated with active TAK after multivariable-adjusted logistic regression (p = 0.008). TAK had greater cytokine secretion from PBMCs (IFN-γ, IL-17A, IL-10 versus HC; IL-6, TNF-α, IL-1ß versus HC or sarcoidosis) (p < 0.05). In-vitro, PBMCs from TAK showed reduced secretion of all inflammatory cytokines with tacrolimus, with synergistic reduction in IL-17A, IL-6, IL-1ß and IL-10 following addition of tadalafil to tacrolimus. Serial follow-up of immunosuppressive-naïve TAK (n = 16) showed reduction in serum IL-6 and TGF-ß1 (p < 0.05) and IL-6 in culture supernatant (p < 0.05) following immunosuppressive therapy. Conclusion: Novel Th17 sub-populations (Th17.1 and PD1+Th17) are elevated in TAK. Th17 lymphocytes associate with active TAK. In-vitro experiments on cultured PBMCs suggest promise for further evaluation of a combination of immunosuppressive tacrolimus with anti-fibrotic tadalafil (or other anti-fibrotic therapies) in clinical trials of TAK.

Tacrolimus induces remission in refractory and relapsing lupus nephritis by decreasing P-glycoprotein expression and function on peripheral blood lymphocytes.

P-glycoprotein (P-gp)-mediated efflux of corticosteroids (CS) may contribute to treatment unresponsiveness in Lupus Nephritis (LN) patients. Tacrolimus is a P-gp inhibitor and hence, may overcome this resistance. We aimed to study the response to tacrolimus, along with the expression and function of P-gp on peripheral blood lymphocytes (PBL) in patients with refractory and relapsing proliferative Lupus Nephritis. We enrolled 12 refractory/relapsing LN patients and treated them with corticosteroids and tacrolimus for 6 months. Expression and function of P-gp on PBL was measured by flow cytometry (as relative fluorescence index, RFI and Rhodamine dye efflux assay) before and 3 months after tacrolimus therapy. Renal response was assessed according to ACR response criteria after 3 and 6 months of tacrolimus therapy. 8 out of 12 refractory/relapsing LN patients achieved renal response (5 partial response, PR and 3 complete responses, CR) as early as 3 months, and 11 patients achieved renal response (7 PR and 4 CR) at 6 months from start of tacrolimus therapy. Proteinuria decreased from median urine protein creatinine ratio (UPCR) of 2.80 (2.00-3.40) at baseline to 1.20 (0.66-1.73) at 3 months (p < 0.001) and to 0.80 (0.19-1.30) at 6 months (p < 0.01). There was significant decrease in P-gp expression [RFI, 3.33 (2.87-4.97) vs 2.03 (1.25-3.86), p < 0.05) and P-gp function (RFI, 55.7 (29.7-84.1) vs 26.8 (16.1-37.0), p < 0.01) after 3 months of tacrolimus therapy. Tacrolimus achieves renal response in refractory/relapsing proliferative LN patients which may be partly related to overcoming P-glycoprotein mediated treatment unresponsiveness.

The prevalence and clinical characteristics of anti-HMGCR (anti-3-hydroxy-3-methyl-glutaryl-coenzyme A reductase) antibodies in idiopathic inflammatory myopathy: an analysis from the MyoCite registry.

This study aimed to determine the prevalence and clinical characteristics of anti-HMGCR antibodies in idiopathic inflammatory myositis (IIM) at a tertiary care centre in northern India. Data (adult and children) were retrieved from the MyoCite dataset, identifying patients with polymyositis, dermatomyositis, and antibody-negative IIM whilst fulfilling the ACR/EULAR criteria. SLE, sarcoidosis, and systemic sclerosis were included for comparison as disease controls. The baseline clinical profile, laboratory tests, and muscle biopsies were retrieved and analysed. Descriptive statistics and non-parametric statistics were used for comparison. Among 128 IIM (112 adults, 16 children, M:F 1:2.8) of age 37 (24-47) years and 6 (3-17) months disease duration, 4 (3.6%) young adults tested positive for anti-HMGCR antibodies. All children and disease control tested negative for the antibody. Anti-HMGCR + IIM exhibited higher muscle enzymes [AST (367 vs 104 IU/L, p = 0.045), ALT (502 vs 78 IU/L, p = 0.004), and CPK (12,242 vs 699 IU/L, p = 0.001] except lactate dehydrogenase with less frequent systemic features such as fatigue than antibody-negative IIM. One young girl presented with a Limb-girdle muscular dystrophy (LGMD) with chronic pattern. None of the patients exhibited rashes, statin exposure, or cancer, though one had anti-Ro52 and mild disease. Our observations depict a younger population while affirming previous literature, including NM-like presentation, and chronic LGMD-like pattern of weakness in one case. Although a small number of children were included, ours is one of the few paediatric studies that evaluated HMGCR antibodies thus far. Further investigations in a larger Indian cohort are warranted to substantiate our findings.

Prevalence of p-glycoprotein (PGP) expression, function and its effect on efficacy of rifampicin in patients with lymph node tuberculosis.

OBJECTIVE: P-glycoprotein (PGP) overexpression may be one of the operating mechanisms of suboptimal responses to antitubercular treatment (ATT) in patients with lymph node tuberculosis. This might become responsible for the development of drug resistance later due to exposure of subtherapeutic concentrations to the mycobacteria. In this study we aim to study the prevalence of PGP expression and function and its relationship with serum concentrations of Rifampicin in consecutive patients with lymph node tuberculosis. METHODS: All newly diagnosed treatment naïve subjects with a confirmed diagnosis of tubercular lymphadenopathy were included in the study and the expression and function of PGP in blood was determined by flowcytometry at baseline and after two months of treatment. Serum levels of Rifampicin was measured at 2 months by high performance liquid chromatography (HPLC). The mean net PGP expression expressed as percent and relative fluorescence indices (RFI) of PGP expression and function respectively was compared at baseline at 2 months and was also correlated with serum rifampicin levels. RESULTS: The mean net PGP expression, RFI of PGP expression and RFI of PGP function were significantly higher in patients with lymph node tuberculosis as compared to healthy controls and the mean net PGP expression and RFI of PGP expression were significantly higher at 2 months as compared to baseline (25.64 ± 5.18% vs. 27.68 ± 4.89%, 4.34 ± 1.09% vs. 4.95 ± 1.55). There was no significant difference in RFI of PGP expression and RFI of PGP function between the poor-responders and responders at baseline and 2 months however there was a trend towards significantly higher net PGP expression amongst poor responders at baseline. The mean serum rifampicin levels were 10.74 ± 2.36 µg/ml in the responder group and 7.86 ± 1.21 µg/ml in the non-responder group and the difference between the two was statistically significant (p = 0.004). CONCLUSIONS: Overexpression of PGP is common in patients with lymph node tuberculosis and leads to lower concentrations of Rifampicin in blood which subsequently may give rise to development of drug resistance. This is also responsible for poor therapeutic responses in these patients. Nonspecific inhibitors of PGP may be used in conjunction with ATT to augment therapeutic response in such cases.

Utility of neutrophil CD64 and serum TREM-1 in distinguishing bacterial infection from disease flare in SLE and ANCA-associated vasculitis.

Bacterial and opportunistic infections are a major cause of morbidity and mortality in patients with systemic lupus erythematosus (SLE) and antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis owing to treatment with immunosuppressants. Commonly used laboratory tests are unreliable in differentiating infection from active disease patients. Fc receptor (FcγR1 or CD64) expression on neutrophils and soluble TREM-1 (triggering receptor expressed on monocytes) are potential biomarkers of bacterial infections. Our aim was to measure the clinical usefulness of quantitative CD64 measurement on neutrophils and soluble TREM-1 measurements in differentiating bacterial infection from active disease in patients with SLE and ANCA vasculitis. Patients with bacterial infection (n = 25), active disease (n = 51), and healthy controls (n = 20) were included. Neutrophil CD64 expression using flow cytometry and sTREM-1 and procalcitonin levels by ELISA were studied. The percentage of neutrophils with CD64 expression and their mean fluorescence intensity in patients with infection (68.8 (56.9-86.5)%, 1037 (229-1828)) were significantly (p < 0.05) higher as compared to those without infection (7.7 (2.6-13.1)%, 456 (20-968)) and controls (7.05 (1.4-9.5)%, 99.5 (54.7-140.7)). The sensitivity and specificity of CD64 expression on neutrophils to diagnose bacterial infection (using a cutoff value of 30%) was 85% and 84%, respectively, whereas the sensitivity and specificity of procalcitonin was 75% and 85%, respectively. There was no significant difference in soluble TREM-1 levels between the two groups. Quantitative measurement of CD64 on neutrophils can distinguish between systemic infection and the flare of autoimmune diseases.

5-HT2 and 5-HT2B antagonists attenuate pro-fibrotic phenotype in human adult dermal fibroblasts by blocking TGF-ß1 induced non-canonical signaling pathways including STAT3 : implications for fibrotic diseases like scleroderma.

BACKGROUND: Release of 5-hydroxytryptamine (5-HT; serotonin) from activated platelets following microvascular injury leads to tissue fibrosis. 5-HT strongly induces extracellular matrix synthesis in dermal fibroblasts in a transforming growth factor beta 1 (TGF-ß1)-dependent manner. AIM: To evaluate anti-fibrotic properties of inhibitors of 5-HT2 and 5-HT2B (terguride, SB204741) respectively in human adult dermal fibroblasts (HADF) derived from a patient with scleroderma. METHODS: Anti-fibrotic efficacy of 5-HT2 and 5-HT2B inhibitors was evaluated as per two strategies: HADF were incubated with 5-HT (1 µM)/TGF-ß1 (10 ng/mL) for 1 hour followed by 5-HT (1 µM)/TGF-ß1 (10 ng/mL) and terguride or SB204741 (1 µM, each) for 24 hours (post-treatment strategy) and HADF were treated with terguride or SB204741 (1 µM, each) for 1 hour followed by 5-HT (1 µM)/TGF-ß1 (10 ng/mL) for 24 hours (pre-treatment strategy). Real time quantitative polymerase chain reaction for expression of pro-fibrotic (TGFΒ1, COL1A1, COL1A2, ACTA2, CTGF and FN1) and anti-fibrotic genes (MMP2/TIMP1) was performed. Expression of type I collagen, alpha smooth muscle actin (α-SMA), phosphorylation of Smad3, ERK1/2 and STAT3 was examined by immunoblotting. RESULTS: Stimulation of HADF cells with 5-HT/TGF-ß1 led to the increased expression of pro-fibrotic genes which was significantly reduced by both terguride and SB204741. Expression of anti-fibrotic genes was not affected upon incubation with the inhibitors. In 5-HT-stimulated HADF, treatment with terguride and SB204741 decreased type I collagen and α-SMA. In 5-HT/TGF-ß1 stimulated HADF, terguride and SB204741 treatment reduced ERK1/2 and STAT3 phosphorylation but did not influence Smad3 phosphorylation. CONCLUSION: Terguride and SB204741 reduce pro-fibrotic potential of HADF cells and suppress TGF-ß1-mediated non-canonical pathways, ERK1/2 and STAT3 which have been implicated in the regulation of pro-fibrotic genes and in the development of fibrosis. Taken together, our data suggest that 5-HT inhibitors might reduce fibrosis via suppression of TGF-beta1-mediated non-canonical signaling pathways. These observations have important therapeutic implications for fibrotic disorders like scleroderma.

The association of polymorphic variants, rs2267788, rs1333049 and rs2383207 with coronary artery disease, its severity and presentation in North Indian population.

BACKGROUND: The polymorphic alleles of APOA5 (rs2266788 (C), rs3135506 (G)), LPA (rs10455872 (A), rs3798220 (G)) and 9p21.3 (rs1333049 (C), rs2383207(A)) have been reported in association with susceptibility of coronary artery disease (CAD) from genome wide association studies. We aimed to assess the association of genetic variants with coronary angiogram proven CAD, severity scored with modified Gensini score and association of risk for myocardial infraction (MI) in North Indian population. METHODS: We recruited 512 angiographic proven CAD patients (mean age 58.1±10.2years) and 272 controls (mean age 50.3±11.1years) with normal coronaries from North Indian population. The genotyping technique polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was employed for rs2266788, rs3135506 and rs10455872. Amplified refractory mutation system-PCR (ARMS-PCR) was used for genotyping of rs1333049, rs2383207 and rs3798220 genetic variants. RESULTS: The polymorphic risk allele of variants rs2266788 (C), rs1333049 (C), rs2383207 (A) and heterozygous polymorphic alleles of rs2266788 (TC) were significantly associated with CAD. The homozygous alleles of rs22667788 (CC) and rs1333049 (CC) had also been significantly associated with CAD. The significance of association of rs2266788 (C, CC, TC) and rs1333049 (C, CC) increases with severity of CAD. The presence of mutant allele of rs2266788 (C) was associated with risk of MI and unstable angina (UA). Also, homozygous risk allele of rs2266788 (CC) significantly associated with risk of MI and UA in patients of chronic stable angina (CSA) patients. Whereas, the risk allele of rs1333049 (C) have shown the association with MI and UA compared to controls. The genetic variants of rs3135506 (G), rs10455872 (A) and rs3798220 (G) have low frequency in our population and reflected no association with CAD. CONCLUSION: The polymorphic variants of Apo-A5; rs2266788 (C), 9p21.3; rs1333049 (C) rs2383207 (A) are associated with CAD, its severity and exerts the risk of MI in North Indian population.

Interstitial lung disease in Systemic sclerosis: insights into pathogenesis and evolving therapies.

Interstitial lung disease (ILD) is a leading cause of mortality in systemic sclerosis (SSc). However, mortality is improving as pathogenesis is being better understood and new therapies emerge. The roles of the inflammasome and NETosis in fibrosis are being elucidated. Epigenetic targets like DNA methylation and microRNA show promise as new targets for anti-fibrotic agents. The IL17-23 pathway has been shown to be active in SSc-ILD. Newer biomarkers are being described like CCL18 and the anti-eIF2B antibody. Hypothesis-free approaches are identifying newer genes like the ALOX5AP and XRCC4 genes. Computer-aided interpretations of CT scans, screening with ultrasonography and magnetic resonance imaging (MRI) are gradually emerging into practice. Imaging can also predict prognosis. A plethora of studies has shown the benefit of immunosuppression in halting ILD progression. Extent of lung involvement and PFT parameters are used to initiate therapy. The best evidence is for cyclophosphamide and mycophenolate. Besides these, corticosteroids and rituximab are being used in cases refractory to the first line drugs. Stem cell transplant is also backed by evidence in SSc. Longer studies on maintenance therapy are awaited. The inflammation in SSc is mostly subclinical and there is great interest in developing anti-fibrotic drugs for SSc-ILD. Perfinidone and nintedanib are under trial. The last resort is lung transplantation.

Short-Term Non-Steroid Anti-Inflammatory Drug Use in Spondyloarthritis Patients Induces Subclinical Acute Kidney Injury: Biomarkers Study.

BACKGROUND: Non-steroid anti-inflammatory drug (NSAID) usage is associated with kidney injury. Rise in serum creatinine (sCr) often represents irreversible process. Thus to assess the early effects of regular NSAID use, we studied sensitive serum and urine biomarkers of kidney injury. METHODS: In a protocol-based intervention study, 103 subjects were enrolled in 3 mutually exclusive groups. Group 1 included 37 healthy controls having minimal baseline NSAID exposure as per a definition, and group 2 had 41 spondyloarthritis (SpA) patients on regular NSAID therapy for >3 months. Group 3 included 25 SpA patients having minimal NSAID exposure at baseline. Blood and urine samples were collected from all the 3 groups at baseline. Furthermore, group 3 was started on 6-week regular NSAID therapy, and blood and urine samples were re-collected at 1, 6, and 12 weeks. Baseline normal kidney function as per the definition was ensured in all the subjects. Creatinine, neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), cystatin-C, and microalbumin were measured in urine and serum samples to assess kidney injury. RESULTS: Kidney injury biomarkers were 2-3-fold higher in SpA patients using regular NSAID therapy compared to healthy controls and SpA patients having minimal NSAID exposure (uKIM-1 and uNGAL p < 0.0001, sKIM-1 and sNGAL p = 0.001). There was no difference in sCr and estimated glomerular filtration rate using Cockcroft-Gault equation between the groups. In SpA patients started on 6 weeks of regular NSAID (group 3), biomarker levels started rising at week 1 and showed a significant rise at week 6. The levels in the patients that stopped NSAID use at 6 weeks showed reversibility at 12 weeks. CONCLUSIONS: Regular NSAID use in SpA patients induces subclinical kidney injury represented by rise in biomarkers. These levels start rising as early as 7 days of regular NSAID use and are reversible on stopping the drug.

Persistent expression and function of P-glycoprotein on peripheral blood lymphocytes identifies corticosteroid resistance in patients with systemic lupus erythematosus.

Corticosteroids (CS) are the mainstay of treatment in systemic lupus erythematosus (SLE) patients. However, some patients have poor response to CS treatment. Among the multiple mechanisms of CS resistance, overexpression of P-glycoprotein (P-gp) on peripheral blood lymphocytes (PBL) may be one of them as this result in efflux of CS from lymphocytes. Thus, we evaluated the role of P-gp protein on PBLs in patients with SLE in its response to CS therapy. SLE patients (n = 42) (fulfilling ACR revised criteria) who were naïve to CS and immunosuppressive drugs were enrolled. Disease activity was assessed using SLE disease activity index (SLEDAI) and expression, and function of P-gp was evaluated by flow cytometry at baseline and after 3 months of therapy with CS. At 3 months, patients with SLEDAI >4 and SLEDAI ≤4 were grouped as nonresponders and responders, respectively. P-gp expression was significantly increased on PBLs of SLE patients as compared to healthy controls (p < 0.001). P-gp expression and function correlated with SLEDAI (r = 0.49, p = 0.005; and r = 0.49, p = 0.001, respectively). P-gp expression and function were not different in responders and nonresponders at baseline. However, at 3 months of CS therapy, P-gp expression and function decreased in responders (p < 0.001 and p < 0.005, respectively), whereas in nonresponders, it remained unchanged. Persistent overexpression and activity of P-gp are associated with poor response to CS in CS naïve patients of SLE.

Urinary Kidney injury molecule-1 can predict delayed graft function in living donor renal allograft recipients.

AIM: Delayed graft function is an early complication leading to impaired creatinine clearance, urine formation and determinant of long term graft outcome. The aim of the present study was to determine the earliest predictive cut-off value of uKIM-1 level in patients with delayed graft function and acute tubular necrosis. METHODS: We have determined the serial urinary KIM-1 normalized to urinary creatinine (uKIM-1, pg/mg) level at 0, 6, 12, 18, 24 and 48 h of post-transplant by ELISA methods. RESULT: The normalized uKIM-1 and AUC-ROC, of uKIM-1 were progressively increased up to 48 h in both delayed graft function (DGF) and immediate graft function (IGF). The u KIM-1 values were significantly high at 6, 12, 18, 24 and 48 h in patients with DGF as compared to that of IGF except at half an hour post-transplant values. Although, progressive increase in uKIM-1 values were observed in both groups of patients; there was an overlap of values between two groups up to 12 h. The earliest non-overlapping values of uKIM-1 between the groups were observed at 18 h onwards and minimum difference of 923.43 pg/mg. The earliest predictive AUC-ROC of uKIM-1 in patients with DGF without overlap with IGF was also observed at 18 h post-transplant with specificity of 100% and sensitivity of 89.9%. CONCLUSION: Serial uKIM-1 measurement can be used as non-invasive diagnostic biomarkers to predict the incident of DGF in living donor renal transplant recipients. At 18(th) post-transplant hour uKIM-1 can predict DGF with 100% specificity and 89.9% sensitivity with a cut-off value of normalized KIM-1 of 923.43 pg/mg.

Differential alteration in peripheral T-regulatory and T-effector cells with change in P-glycoprotein expression in Childhood Nephrotic Syndrome: A longitudinal study.

INTRODUCTION: Childhood Idiopathic Nephrotic Syndrome (INS) responds to glucocorticoid therapy, however, 60-80% of patients relapse and some of them become steroid non responsive. INS may occur because of T cell dysfunction, abnormal cytokines and podocytopathies which reverse on steroid treatment. The reason of relapses could be imbalances in T cells phenotypes and respective cytokines. Herein, we hypothesize that relapses in INS may occur due to imbalance in T-regulatory and T-effector cell with their respective cytokines and overexpression of P-gp on lymphocytes. METHODS: The frequency of peripheral blood CD4(+)CD25(+)FoxP3(+) Treg, CD4(+)IFN-γ(+) Th1 and CD4(+)IL-4(+) Th2 lymphocytes and their respective cytokines and P-gp expression on peripheral blood lymphocytes (PBLs) were analyzed in INS patients at baseline (n=26), during remission (n=24) and at relapse (n=15). RESULTS: Compared to baseline, the frequency of Tregs was significantly increased at remission and decreased during relapse. In contrast, the frequency of Th1 and Th2 lymphocytes was significantly decreased during remission and increased at the time of relapse. Similarly, expression of P-gp was significantly high at baseline and at the time of relapse as compared to remission. Levels of cytokines IL-10 and TGF-ß in the supernatant of stimulated PBMCs was increased during remission and decreased during relapse. In contrast, levels of IFN-γ and IL-4 were decreased during remission and increased at the time of relapse. CONCLUSIONS: Steroid therapy in INS induces decreased P-gp expression on PBLs along with increased frequency and cytokine response of T-regulatory cells, and reduced frequency and respective cytokine response of Th1 and Th2 cells during remission. However, reversal in the frequency and respective cytokines of T-regs, Th1 and Th2, and P-gp expression on PBLs occurs during relapses on follow-up.

Multidrug resistance protein-1 expression, function and polymorphisms in patients with rheumatoid arthritis not responding to methotrexate.

OBJECTIVE: To study the expression, function and polymorphism of MDR-1 protein on the peripheral blood lymphocytes in patients with RA following treatment with MTX and its relationship with response to therapy. METHODS: RA patients naïve to MTX/DMARD- and glucocorticoid were enrolled. Expression and function of MDR-1 was carried out by flow cytometry at baseline and after 4 months of therapy. MDR-1 expression was measured by relative fluorescence intensities and percentage of positive cells. MDR-1 function was assessed by Rhodamine efflux in presence or absence of verapamil. Patients with reduction in disease activity score 28 ≥1.2 were defined as responders and <1.2 as non-responders. Three single nucleotide polymorphisms in MDR-1 gene, 3435T, 1236T and 2677T/A were studied. RESULTS: Fifty-two patients of RA were grouped into responders (n = 41), and non-responders (n = 11) as per the defined criteria. There was no difference between the groups in terms of age, sex ratio or duration of illness, MTX dose and follow-up duration. The expression and function of the MDR-1 protein reduced significantly in the responder group after the treatment with MTX when compared to the baseline evaluation. The decrease was significant when compared to the non-responders at the fourth month. MDR-1 expression and function either increased or remained the same in the non-responder group after treatment with MTX. MTX unresponsiveness was not related to any of the three polymorphisms studied. CONCLUSION: Persistent expression and function of MDR-1 identifies a subset of RA patients not responding to MTX. Its early recognition may help in appropriately modulating therapy.

Magnetic-nanoparticle-doped carbogenic nanocomposite: an effective magnetic resonance/fluorescence multimodal imaging probe.

A novel and facile approach is developed to synthesize a magnetic nanoparticle (iron oxide)-doped carbogenic nanocomposite (IO-CNC) for magnetic resonance (MR)/fluorescence imaging applications. IO-CNC is synthesized by thermal decomposition of organic precursors in the presence of Fe(3) O(4) nanoparticles with an average size of 6 nm. IO-CNC shows wavelength-tunable fluorescence properties with high quantum yield. Magnetic studies confirm the superparamagnetic nature of IO-CNC at room temperature. IO-CNC shows MR contrast behavior by affecting the proton relaxation phenomena. The measured longitudinal (r(1) ) and transverse (r(2) ) relaxivity values are 4.52 and 34.75 mM(-1) s(-1) , respectively. No apparent cytotoxicity is observed and the nanocomposite shows a biocompatible nature. In vivo MR studies show both T(1) and T(2) * contrast behavior of the nanocomposite. Fluorescence imaging indicates selective uptake of IO-CNC by macrophages in spleen.