A comparative study of modulatory interaction between cytokines and apoptotic proteins among Scleroderma patients with and without pulmonary involvement.

BACKGROUND: Interstitial lung disease (ILD) and pulmonary arterial hypertension (PAH) are the most eminent forms of pulmonary involvement in Scleroderma. In this study we investigate the interaction between cytokines and apoptotic proteins in treatment naive Scleroderma (SSc) patients with and without pulmonary involvement. METHODS: Newly diagnosed treatment naïve Scleroderma (SSc) patients (n = 100) and healthy controls (n = 100) were enrolled. Patients were classified as ILD-SSc, PAH-SSc and non-pulmonary SSc (np-SSc). Study variables like mRSS score, autoantibody profile, serum cytokines, serum TGF-ß (1,2,3) and apoptotic proteins were assessed for these patients. RESULTS: Scleroderma patients showed elevated levels of serum cytokines, but significantly lower IL-22 and TGF- ß1 when compared to healthy controls (p < 0.05). Apoptotic proteins were significantly elevated among Scleroderma patients, but the patient groups also showed significant lower caspase 1/3/9 levels when compared to healthy controls (p < 0.05). ILD-SSc patients reported higher mRSS score (p = 0.0436) when compared with PAH-SSc and np-SSc. In ILD-SSc patients, finger tightening (p = 0.0481) and calcinosis/lesions (p = 0.0481) were significant clinical presentations whereas, digital ulcers were significantly prominent in np-SSc patients (p = 0.0132). Elevated TGF-ß3 levels (p = 0.02) in SSC-ILD and reduced IL-4 levels (p = 0.02) in SSC-PAH were significant cytokines as compared to np-SSc. Significant correlations were obtained among serum cytokines and apoptotic proteins in Scleroderma patients with and without pulmonary involvement. (p < 0.05) CONCLUSION: Our study highlights the correlation between mRSS score, cytokines and apoptotic proteins in SSc patients with pulmonary involvement. A longitudinal follow up in these patients with assessment of these immunological parameters may be helpful in monitoring the disease.

Perturbations of immune landscape in COVID-19 associated mucormycosis.

BACKGROUND: A rise in secondary fungal infections during the COVID-19 pandemic necessitates a deeper understanding of the associated immunological perturbations. OBJECTIVES: To evaluate the clinical and immunological characteristics observed in patients with COVID-19 associated mucormycosis (CAM) infection. PATIENTS/ METHODS: Cases of mucormycosis with or post-COVID-19 infection were compared with cases of acute COVID-19 and convalescent COVID-19. Lymphocyte subsets, cytokines and other laboratory markers were compared between the groups. RESULTS: The frequency of proposed risk factors for CAM was diabetes mellitus (77%), recent history of steroid use (69%) and hypoxia during COVID-19 infection (52%). Iron metabolism was dysregulated in CAM patients with low TIBC and total iron. Further, CAM was accompanied with lymphopenia with drastic reduction in B cell counts; however, plasmablasts were not altered. Further, CAM patients had low immunoglobulin levels and antibodies specific to mucor peptide did not increase in CAM suggesting dysfunction in B-cell response. There was increase in activated effector cytotoxic CD8 T cells and NK cells in CAM compared with COVID-19 infection and healthy controls. Among T helper cells, Tregs were reduced and Th-1 frequency was increased in CAM compared with COVID-19 infection. A distinct cytokine signature was evident in CAM with increase in IL-1ß, IFN-γ, IL-6, IL-22, IL-17A, IL-10, IL-2, IL-8, IL-7, IL-21 and GM-CSF. CONCLUSION: This is the first study on immunophenotyping in CAM suggesting the need for long-term monitoring of B-cell function after SARS-CoV-2 in patients with dysregulated glycaemic control and the possible benefit of therapeutic supplementation with intravenous immunoglobulins in CAM.

Satellite Epidemic of Covid-19 Associated Mucormycosis in India: A Multi-Site Observational Study.

BACKGROUND: Sudden upsurge in cases of COVID-19 Associated Mucormycosis (CAM) following the second wave of the COVID-19 pandemic was recorded in India. This study describes the clinical characteristics, management and outcomes of CAM cases, and factors associated with mortality. METHODS: Microbiologically confirmed CAM cases were enrolled from April 2021 to September 2021 from ten diverse geographical locations in India. Data were collected using a structured questionnaire and entered into a web portal designed specifically for this investigation. Bivariate analyses and logistic regression were conducted using R version 4.0.2. RESULTS: A total of 336 CAM patients were enrolled; the majority were male (n = 232, 69.1%), literate (n = 261, 77.7%), and employed (n = 224, 66.7%). The commonest presenting symptoms in our cohort of patients were oro-facial and ophthalmological in nature. The median (Interquartile Range; IQR) interval between COVID diagnosis and admission due to mucormycosis was 31 (18, 47) days, whereas the median duration of symptoms of CAM before hospitalization was 10 (5, 20) days. All CAM cases received antifungal treatment, and debridement (either surgical or endoscopic or both) was carried out in the majority of them (326, 97.02%). Twenty-three (6.9%) of the enrolled CAM cases expired. The odds of death in CAM patients increased with an increase in HbA1c level (aOR: 1.34, 95%CI: 1.05, 1.72) following adjustment for age, gender, education and employment status. CONCLUSION: A longer vigil of around 4-6 weeks post-COVID-19 diagnosis is suggested for earlier diagnosis of CAM. Better glycemic control may avert mortality in admitted CAM cases.

A case control investigation of COVID-19 associated mucormycosis in India.

BACKGROUND: Increased occurrence of mucormycosis during the second wave of COVID-19 pandemic in early 2021 in India prompted us to undertake a multi-site case-control investigation. The objectives were to examine the monthly trend of COVID-19 Associated Mucormycosis (CAM) cases among in-patients and to identify factors associated with development of CAM. METHODS: Eleven study sites were involved across India; archived records since 1st January 2021 till 30th September 2021 were used for trend analysis. The cases and controls were enrolled during 15th June 2021 to 30th September 2021. Data were collected using a semi-structured questionnaire. Among 1211 enrolled participants, 336 were CAM cases and 875 were COVID-19 positive non-mucormycosis controls. RESULTS: CAM-case admissions reached their peak in May 2021 like a satellite epidemic after a month of in-patient admission peak recorded due to COVID-19. The odds of developing CAM increased with the history of working in a dusty environment (adjusted odds ratio; aOR 3.24, 95% CI 1.34, 7.82), diabetes mellitus (aOR: 31.83, 95% CI 13.96, 72.63), longer duration of hospital stay (aOR: 1.06, 95% CI 1.02, 1.11) and use of methylprednisolone (aOR: 2.71, 95% CI 1.37, 5.37) following adjustment for age, gender, occupation, education, type of houses used for living, requirement of ventilatory support and route of steroid administration. Higher proportion of CAM cases required supplemental oxygen compared to the controls; use of non-rebreather mask (NRBM) was associated as a protective factor against mucormycosis compared to face masks (aOR: 0.18, 95% CI 0.08, 0.41). Genomic sequencing of archived respiratory samples revealed similar occurrences of Delta and Delta derivates of SARS-CoV-2 infection in both cases and controls. CONCLUSIONS: Appropriate management of hyperglycemia, judicious use of steroids and use of NRBM during oxygen supplementation among COVID-19 patients have the potential to reduce the risk of occurrence of mucormycosis. Avoiding exposure to dusty environment would add to such prevention efforts.

Cutaneous Features, Autoantibody Profile, and Nailfold Capillaroscopy of Systemic Sclerosis: A Study of 60 Cases.

BACKGROUND: Systemic sclerosis (SSc) is an autoimmune chronic multisystem disorder with a plethora of cutaneous manifestations. These manifestations often may be the only presenting complaint. Early identification of these help in diagnosing grievous systemic manifestations and their prompt and appropriate treatment. AIMS: To study the clinical profile of SSc, modified Rodnan's skin scoring (mRSS), nailfold capillaroscopy (NFC) patterns, antibody profile in the western India population, and their association with cutaneous manifestations. METHODS: Patients of SSc fulfilling the European League Against Rheumatism (EULAR) 2013 classification of SSc criteria, who attended dermatology outpatient department (OPD) between January 2017 and September 2018 were included in the study. The demographic data, cutaneous features, autoantibody profile, mRSS, and NFC pattern were noted Results: A total of 60 patients (57 females and 3 males; mean age years) of SSc were evaluated. Clinical subtypes were 40 diffuse cutaneous SSc and 20 limited cutaneous SSc. The most common presenting symptoms were Raynaud's phenomenon (RP) (95%) and skin tightening (90%). The common cutaneous findings were sclerodactyly (86.7%), stellate scars (78.3%), parrot-beaked nose (76.7%), mask-like facies (75%), microstomia (56.7%), salt and pepper pigmentation (55%), puffy finger (46.7%), telangiectasia (46.7%), digital ulcer (38.3%), fixed flexion deformity (33.3%), and calcinosis cutis (8.33%). Limited cutaneous systemic sclerosis (lcSSc) had mRSS score of 8.3 ± 4.1 and diffuse cutaneous systemic sclerosis (dcSSc) subset had a score of 28 ± 10.4. Antinuclear antibody (ANA), Anti-topoisomerase antibody (ATA), and anti-centromere antibody (ACA) were positive in 59, 49, and 7 patients, respectively. The NFC patterns were early (23.3%), active (45%), and late (18.3%). LIMITATION: The sample size of the study was small. We were not able to determine the significance of other less common autoantibodies with scleroderma. CONCLUSION: The study highlights the importance of identifying early cutaneous findings and the role of a useful diagnostic and prognostic reproducible scoring system (mRSS) and NFC.

Mannose Binding Lectin Levels and its Association with Systemic Lupus Erythematosus Disease Severity: An Indian Report.

BACKGROUND: Dysregulated serum levels of Mannan binding lectin (MBL) has a probable role in Systemic Lupus Erythematosus (SLE) pathogenesis. OBJECTIVE: To evaluate the association between serum MBL levels in SLE patients from western India with the severity of disease Methods: SLE patients (n=70) from Western India were included. Based on MBL levels, patients were classified into four categories, viz. low (<100 ng/ml), mild (100-500 ng/ml), moderate (500-1000 ng/ml) and high (>1000 ng/ml). Correlation of serum MBL levels with disease severity was assessed using the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI). MBL levels and circulating immune complex levels were detected by ELISA. C3, C4 and CRP levels were detected by nephelometer. RESULTS: Serum MBL levels of SLE patients (1954 ± 202.4 ng/ml) was lower than that of healthy controls (2388 ± 205.0 ng/ ml). There was no significant correlation between MBL levels with severity of SLE on the basis of ACR criteria and SLEDAI scores (p> 0.05). No significant difference was observed among MBL levels and SLE patients with (1847 ± 246.7) or without (1900 ± 246.8) Lupus Nephritis. SLE patients without infections (n= 33) had low MBL levels (1700 ± 301.0 ng/ ml) as compared with SLE patients with infection (n= 37) (2189 ± 284.6 ng/ ml) (p=0.30) Conclusion: Present study indicated that low MBL levels were not associated with disease severity, haematological manifestations and infections among SLE patients from Western India.

Impact of Autoantibodies to Complement Components on the Disease Activity in SLE.

INTRODUCTION: Systemic Lupus Erythematosus (SLE) is a chronic multi-system autoimmune disease with varied clinical presentations. Complement components are the major players in disease pathogenesis. This retrospective cross-sectional study was aimed at assessing the role of autoantibodies to these complement components and their association disease activity in newly diagnosed SLE patients from India. METHOD: Clinically diagnosed SLE patients (n=57) classified as per 2015 ACR/SLICC revised criteria were enrolled between November 2016 to April 2017. Patients' sera were tested for C3 and C4 by nephelometry, while serum levels of factor H, factor P (properdin) as well as autoantibodies to C3, C4, factor H and factor P were detected by ELISA. GraphPad Prism Version 6.01 was used for statistical analysis. Mean, SD, SEM were calculated. Mann Whittney U-test, ANOVA, Chi-square test, Odd's Ratio were calculated. Pearson's correlation was used to study relativeness of the study parameters. RESULTS: Among the 57 SLE patients, low C3 were seen in 51% patients, low C4 in 49%, low factor H in 19% and low factor P in 49% patients. Positivity for autoantibodies against complement components, anti-C3 were seen in 42% patients, anti-C4 in 7%, anti-factor H in 19% and anti-factor P in 28% patients. Serum levels of C3 (p=0.0009), C4 (p=0.0031) and anti-C3 autoantibodies (p=0.0029) were significantly associated with ACR/SLICC 2015 scores. CONCLUSION: Hypocomplementemia was found to be associated with higher disease damage score in newly diagnosed SLE patients. This study adds novel arguments for the importance of the anti-C3 autoantibodies as a marker of SLE.

Cytokine genes multi-locus analysis reveals synergistic influence on genetic susceptibility in Indian SLE - A multifactor-dimensionality reduction approach.

Systemic lupus erythematosus (SLE) is a prototype autoimmune disease with unclear etiology. Several loci associated with genetic susceptibility for lupus have been described. However, it lacks reports on cytokine gene-gene interactions among SLE patients from Asian population. Epistasis interaction among single nucleotide polymorphisms (SNPs) of cytokine genes in Indian SLE patients was tested using multifactor-dimensionality reduction (MDR) analysis. A total of fourteen SNPs lacking linkage disequilibrium among different cytokines genes were genotyped in a cohort of 200 SLE patients and 201 healthy individuals as controls of Indian origin. A high degree of synergism among Lymphotoxin-α (LT-α), Interleukin-1ß (IL-1ß) and Interleukin-10 (IL-10) gene polymorphisms was detected in our SLE patients. Furthermore, by virtue of biological inter-relations among different cytokines, a high strength of interactions was observed among pro-inflammatory (IL-1ß, IL-6) and anti-inflammatory (IL-10) cytokine gene SNPs. Also, among studied pro-inflammatory cytokines and chemokines, a strong synergistic effect among Tumor Necrosis Factor-α (TNF-α), LT-α and Monocyte Chemo-attractant Protein-1 (MCP-1) SNPs was occurred. A nature of strong interaction among the candidate cytokine genes may speculate a proactive role in causing genetic susceptibility to the disease in SLE patients with Indian origin.

Fibrotic Cytokine Interplay in Evaluation of Disease Activity in Treatment Naïve Systemic Sclerosis Patients from Western India.

BACKGROUND: : Systemic sclerosis (SSc) is a demyelinating disease of skin, subcutaneous tissue, muscles and internal organs, with fibrosis as an important pathological event. AIM: : To understand cytokine interplay of IL-1ß, IL-4 and IL-6 and their association with disease activity in treatment naïve active cases of systemic sclerosis from Western India. METHODS: Twenty-five SSc patients as per ACR-EULAR 2013 criteria (classified based on pulmonary fibrosis and generalized fibrosis) and 25 age-sex matched controls were enrolled. Serum cytokine levels of IL-1ß, IL-4 and IL-6 were assessed by multiplex bead based immunoassay. RESULTS: Ten patients had Interstitial lung disease (ILD), whereas, 16 patients had generalized fibrosis. Anti-nuclear antibodies were seen in 22 patients (88%); antiScl70 in 15 patients (60%) and anti-Centromere antibodies in 5 patients (20%). Serum levels of IL-1ß in patients were significantly higher than healthy controls (p=0.0006). IL-4 levels in all SSc patients were marginally raised (p=0.0102), while IL-6 levels were significantly raised (p<0.0001). IL-4 was found to be significantly raised in SSc patients with ILD (p=0.021) as compared to patients without ILD. IL-1ß (p=0.0293) and IL-4 (p<0.0001) were significantly higher in SSc patients with fibrosis. On the contrary, IL-6 levels in patients with fibrosis were found to be lower than in patients without fibrosis. CONCLUSION: Significantly raised cytokine levels among treatment naïve systemic sclerosis patients were found to be associated with higher disease severity in our study. Higher levels of IL-1ß and IL-6 indicated an active inflammatory status, whereas significantly raised IL-4 levels indicated at higher fibrotic activity.

Association of clinical and serological parameters of systemic lupus erythematosus patients with Epstein-Barr virus antibody profile.

Epstein-Barr viral infection is one of the known environmental factors involved in development of Systemic Lupus Erythematous (SLE). Though not much is known about the exact role of Epstein-Barr virus (EBV) in SLE pathogenesis, the theory of switching of lytic and lysogenic cycles of EBV in memory B cells fits well with the periods of waning disease activity and intermittent flares in SLE patients. In this study, we investigate the association of EBV antibody profile with clinical and serological parameters in SLE. Eighty-seven clinically diagnosed SLE patients fulfilling the American College of Rheumatology (ACR) classification criteria and fifty healthy individuals were enrolled in this case control study. Anti-VCA IgM, anti-VCA IgG, and anti-EBNA IgG were detected by ELISA technique. Antibodies concentrations between two groups were compared using Mann-Whitney whereas the difference in categorical data was compared using Chi-square considering statistical significance at P < 0.05. This study demonstrated a significant increase in EBV VCA-IgG, VCA-IgM, and EBNA-IgG antibodies levels of SLE patients when compared to healthy controls (P < 0.05). High seroprevalence was seen in both the study groups for EBV VCA-IgG and EBNA-IgG antibodies when compared to VCA-IgM antibodies. A significant increase was noted in the anti-VCA-IgG levels with dsDNA autoantibody positivity (P < 0.05). Though there was no significant association between EBV antibody profile and clinical manifestations, 100% seropositivity for anti-VCA-IgG was seen in SLE patients with renal manifestations. Association of anti-VCA IgG levels with presence of anti-dsDNA antibodies suggests a possible role of EBV as an environmental trigger in pathogenesis of SLE.

Adipokine interactions promote the pathogenesis of systemic lupus erythematosus.

BACKGROUND: Adipokines are chemical mediators released from adipose tissue involved in regulation of appetite, insulin sensitivity, immune system and inflammatory responses. Adipokines contributes to low grade inflammatory response in autoimmune disease like Systemic Lupus Erythematosus (SLE) but the pathophysiology is yet not clear. The aim of this study is to understand role of adipokine interactions in SLE disease pathogenesis. METHODS: Sixty newly diagnosed treatment naïve SLE patients fulfilling the ACR criteria and forty age-sex matched healthy subjects were enrolled in thiscase-control study. Disease activity in SLE patients was evaluated using SELENA-SLEDAI. Array of adipokines, C1q circulating immune complexes (C1q-CIC), anti-C1q, anti-ribososmal P0 (anti-RibP0) and anti-mitochondrial antibodies (AMA) levels were detected by ELISA. Antinuclear antibodies (ANA) and anti-dsDNA autoantibodieswere detected by Indirect Immunofluorescence (IIF), while antigen specificities were detected by Immunoassay blot. Serum levels of C3 and C4 complement factors were assessed by nephlometer. RESULTS: Statistically significant elevation in progranulin, adipsin and resistin levels was seen among SLE patients when compared to healthy controls (p < 0.0001). Leptin and omentin levels were significantly reduced in SLE patients (p < 0.0001). There was no statistically significant difference in serum adiponectin, chemerin and visfatin levels when these two groups were compared (p > 0.05). Adiponectin, adipsin and resistin levels were elevated in SLE patients with renal manifestations (p < 0.05). Reduced leptin levels were significantly associated with presence of renal manifestations (p < 0.05). Adiponectin levels positively correlated with disease activity (r = 0.294, p = 0.027) whereas negatively correlated with C3 levels (r = -0.439, p = 0.0007). A positive correlation was observed between hypocomplementemia and leptin levels (p < 0.05). Leptin levels were negatively correlated with disease activity, anti-dsDNA, C1q-CIC and anti-C1q levels (p < 0.05). A significant positive correlation was observed between progranulin levels and anti-ribosomal P0 antibodies (r = 0.499, p < 0.0001). CONCLUSION: Adipokines levels and associated clinical manifestations suggest involvement of adipokines in disease pathogenesis of SLE. SLE disease activity and complement components may suggest regulatory effect of adipokines (adiponectin and leptin) on disease pathogenesis. Further studies on adipokines in SLE patients with renal manifestations may propose them as prognostic markers in renal damage. TRIAL REGISTRATION: NA.

A functional SNP MCP-1 (-2518A/G) predispose to renal disorder in Indian Systemic Lupus Erythematosus patients.

Systemic Lupus Erythematosus (SLE) is a clinically heterogeneous chronic, inflammatory autoimmune disorder that affects multiple organs where exact etiology of the disease is not yet clearly understood. Various evidences suggest that genetic polymorphisms in inflammatory mediators like cytokines and chemokines may influence development of the disease. Here, we investigated whether functional polymorphism at the Monocyte Chemoattractant Protein-1 (MCP-1) regulatory region associates with disease phenotype in Indian SLE patients. This case control study included 200 SLE patients and 201 ethnically matched healthy controls. Genotyping of MCP-1 (-2518 A/G) polymorphism was performed using PCR-RFLP method. Serum MCP-1 levels were detected by bead-based multiplex immunoassay. Serum MCP-1 levels were found to be higher in patients compared with healthy individuals (p<0.0001). A significant difference for MCP-1G allele frequency (OR=1.9, 95%CI=1.4-2.6, p<0.0001) was observed among SLE patients against healthy individuals. A significant difference in the distribution of MCP-1 -2518GG (OR=3.0, 95%CI=1.4-6.7, p=0.0041) and AG+GG genotypes (OR=2.0, 95%CI=1.4-3.0, p=0.0005) was also noted among SLE patients when compared with healthy individuals. A significant association was observed between A/G and G/G versus A/A genotypes with renal manifestations (p<0.0001, Pc<0.001). Serum MCP-1 levels in active LN patients were found to be significantly higher than inactive LN (p=0.0059), mild LN (p=0.0061) as well as non-LN patients (p=0.0001). These findings suggest that -2518G allele of MCP-1 -2518 A/G polymorphism is associated with renal disorders and may influence MCP-1 gene expression among Indian SLE patients.

Combined protein- and nucleic acid-level effects of rs1143679 (R77H), a lupus-predisposing variant within ITGAM.

Integrin alpha M (ITGAM; CD11b) is a component of the macrophage-1 antigen complex, which mediates leukocyte adhesion, migration and phagocytosis as part of the immune system. We previously identified a missense polymorphism, rs1143679 (R77H), strongly associated with systemic lupus erythematosus (SLE). However, the molecular mechanisms of this variant are incompletely understood. A meta-analysis of published and novel data on 28 439 individuals with European, African, Hispanic and Asian ancestries reinforces genetic association between rs1143679 and SLE [Pmeta = 3.60 × 10(-90), odds ratio (OR) = 1.76]. Since rs1143679 is in the most active region of chromatin regulation and transcription factor binding in ITGAM, we quantitated ITGAM RNA and surface protein levels in monocytes from patients with each rs1143679 genotype. We observed that transcript levels significantly decreased for the risk allele ('A') relative to the non-risk allele ('G'), in a dose-dependent fashion: ('AA' < 'AG' < 'GG'). CD11b protein levels in patients' monocytes were directly correlated with RNA levels. Strikingly, heterozygous individuals express much lower (average 10- to 15-fold reduction) amounts of the 'A' transcript than 'G' transcript. We found that the non-risk sequence surrounding rs1143679 exhibits transcriptional enhancer activity in vivo and binds to Ku70/80, NFKB1 and EBF1 in vitro, functions that are significantly reduced with the risk allele. Mutant CD11b protein shows significantly reduced binding to fibrinogen and vitronectin, relative to non-risk, both in purified protein and in cellular models. This two-pronged contribution (nucleic acid- and protein-level) of the rs1143679 risk allele to decreasing ITGAM activity provides insight into the molecular mechanisms of its potent association with SLE.

Association of Serum Ferritin Levels with Hematological Manifestations in Systemic Lupus Erythematosus Patients from Western India.

OBJECTIVE: To identify the hematological manifestations and its association with serum ferritin levels in SLE patients from Western India. METHODS: Ninety clinically diagnosed SLE patients fulfilling ACR criteria were included. Disease activity was assessed at the time of evaluation using Systemic Lupus Erythematosus Disease Activity Index (SLEDAI). Sera were tested for serum ferritin levels by ELISA (Calbiotech, USA). Autoantibodies such as ANA, anti-dsDNA by indirect immunofluorescence test (IFA- Bio-Rad, USA) and anti-cardiolipin antibodies (ACA) to IgG and IgM isotypes and Anti-ß2 GP antibodies to IgG and IgM isotypes were detected by ELISA using commercially available kits (Euroimmun, Lubeck, Germany). RESULTS: Out of 90 SLE patients studied, 41 patients (45.6%) showed hematological abnormalities, where anemia (82.9%), leucopenia (26.8%), autoimmune hemolytic anemia (AIHA) (14.6%) and idiopathic thrombocytopenic purpura (ITP) were noted in (34.1%) patients. Mean±SD serum ferritin levels among SLE patients were 270.2±266.0 ng/ml as compared to 29.0±15.8 ng/ml healthy normal controls (p<0.0001). A positive correlation between serum ferritin levels and SLEDAI scores (r= 0.2640, p=0.0124) and anti-dsDNA positivity was noted (r=0.32, p<0.0001). Serum ferritin levels were negatively correlated with hemoglobin levels (r=-0.5964, p=0.0001), WBC count (r=-0.1705, p=0.2316), platelet count ((r=-0.1701, P=0.2375), C3 levels (r=-0.4417, p=0.0034) and C4 levels (r=-0.0363, p=0.8215). CONCLUSIONS: Serum ferritin is an excellent marker of SLE which can be used for an evaluation of disease activity particularly in active stage of the disease mainly in patients having hematological and renal manifestations.

Mannose binding lectin (MBL) 2 gene polymorphism & its association with clinical manifestations in systemic lupus erythematosus (SLE) patients from western India.

BACKGROUND & OBJECTIVES: Systemic lupus erythematosus (SLE) is a prototypical autoimmune disease characterized by production of autoantibodies. Mannose binding lectin (MBL) is an important element of the innate defense system. t0 he present study was undertaken to determine whether variant alleles in MBL2 gene were associated with disease severity in SLE patients. METHODS: The MBL alleles [-550, -221, +4, Codon 52, Codon 54 and Codon 57] were studied by PCR- RFLP (restriction fragment length polymorphism) method in 100 SLE patients fulfilling ACR (American College of Rheumatology) criteria along with 100 healthy controls. SLE disease activity was evaluated using SLE Disease Activity Index (SLEDAI) score. RESULTS: Homozygosity for MBL variant allele (O/O) was observed in 24 per cent of the SLE patients compared to 16 per cent of the normal controls, while no difference was found for heterozygosity (A/O) (37 vs 35%). A significant difference was reported in incidence of double heterozygosity for mutant allele B and D (B/D) among SLE patients as against control group ( p = 0.015). MBL genotypes did not show any association with renal involvement. INTERPRETATION & CONCLUSIONS: In this study from western India, MBL gene polymorphism showed an influence as a possible risk factor for susceptibility to SLE, but had no direct effect on disease characteristics. Further studies need to be done on a larger number of SLE patients in different regions of the country.

T-regulatory cells and programmed death 1+ T cells contribute to effector T-cell dysfunction in patients with chronic obstructive pulmonary disease.

RATIONALE: Previous studies from our laboratory have shown that peripheral blood mononuclear cells (PBMCs) from patients with chronic obstructive pulmonary disease (COPD) prone to exacerbations with nontypeable Haemophilus influenzae have impaired responses to lipoprotein P6. We hypothesized that an underlying immunosuppressive network could be responsible for the defective antibacterial immunity observed in these patients. We evaluated T regulatory cells (Tregs), myeloid-derived suppressor cells (MDSC), and exhausted T effector cells (programmed death 1 [PD-1](+)) in patients with COPD, because these cells are known to play a pivotal role in suppressing immune responses. OBJECTIVES: We performed an in-depth characterization of Tregs, T effector cells, and MDSC in COPD and correlated their levels and function with disease severity. METHODS: Treg, effector T cell, and MDSC frequency from patients with COPD and healthy subjects' PBMCs were analyzed by flow cytometry. Treg immunosuppressive capacity was measured by in vitro suppression assay. The frequency of interferon-γ producing T cells and T-cell proliferation were measured after blocking CTLA-4 and PD-1. Plasma proinflammatory and immunosuppressive cytokine levels were measured. MEASUREMENTS AND MAIN RESULTS: Significantly increased levels of Tregs, MDSC, and PD-1(+) exhausted effector T cells were present in patients with COPD compared with healthy subjects. Tregs from patients with COPD suppressed P6-specific T-cell proliferation to a greater extent than Tregs from healthy subjects. Plasma levels of Treg-generated cytokines, IL-10, and transforming growth factor-ß were elevated. Blockade of CTLA-4 resulted in significant augmentation of T-cell IFN-γ production in patients with COPD. CONCLUSIONS: Functionally suppressive Tregs, MDSCs, and exhausted PD-1(+) T cells contribute to effector T-cell dysfunction in COPD.

Neuropsychiatric manifestations and associated autoantibodies in systemic lupus erythematosus patients from Western India.

Systemic lupus erythematosus with neuropsychiatric involvement (NPSLE) can be diagnosed clinically, but there is no definite serological biomarker established. The objectives of this study were to evaluate the neuropsychiatric involvement in systemic lupus erythematosus (SLE) patients and to detect the autoantibodies associated with them. Sixty NPSLE patients along with sixty SLE patients without neuropsychiatric involvement from Maharashtra, India, were included. All patients were clinically diagnosed using the American College of Rheumatology criteria. Disease activity was assessed using the systemic lupus erythematosus disease activity index. Antinuclear antibodies (ANA), anti-dsDNA, anti-neuronal antibodies were detected by indirect immunofluorescence test. Anti-ribosomal antibodies (anti-Rib-P) were tested by ELISA. NPSLE was diagnosed in age group ranging between 10 and 20 years compared with SLE patients without neuropsychiatric involvement (21-30 years). The most frequent symptoms were psychosis (75%), followed by seizures (58%), lupus headache (40%), cognitive dysfunction (36%), mood disorder (30%), cerebrovascular disease (20%), and anxiety (18%). ANA were present in all. The prevalence of anti-Rib-P was 26.6% in NPSLE and 16.6% in SLE patients without neuropsychiatric involvement. Anti-neuronal antibodies were found in 56.7% in NPSLE and 43.4% in SLE patients without neuropsychiatric involvement. Anti-neuronal antibodies were found to be highest in the patients of psychosis (66.6%) followed by central nerve system disease (63.63 %) and seizures (56.25%). There was an early onset of neuropsychiatric involvement. Anti-Rib-P antibodies as well as anti-neuronal antibodies did not show statistically significant correlation with neuropsychiatric manifestations in NPSLE patients.

Effect of proinflammatory cytokines (IL-6, TNF-α, and IL-1ß) on clinical manifestations in Indian SLE patients.

Systemic lupus erythematosus (SLE) is an inflammatory rheumatic disease characterized by production of autoantibodies and organ damage. Elevated levels of cytokines have been reported in SLE patients. In this study we have investigated the effect of proinflammatory cytokines (IL-6, TNF-α, and IL-1ß) on clinical manifestations in 145 Indian SLE patients. One hundred and forty-five healthy controls of the same ethnicity served as a control group. Clinical disease activity was scored according to SLEDAI score. Accordingly, 110 patients had active disease and 35 patients had inactive disease. Mean levels of IL-6, TNF-α, and IL-1ß were found to be significantly higher in SLE patients than healthy controls (P < 0.001). Mean level of IL-6 for patients with active disease (70.45±68.32 pg/mL) was significantly higher (P = 0.0430) than those of inactive disease patients (43.85±63.36 pg/mL). Mean level of TNF-α was 44.76±68.32 pg/mL for patients with active disease while it was 25.97±22.03 pg/mL for those with inactive disease and this difference was statistically significant (P = 0.0161). Similar results were obtained for IL-1ß (P = 0.0002). Correlation between IL-6, TNF-α, and IL-1ß serum levels and SLEDAI score was observed (r = 0.20, r = 0.27, and r = 0.38, resp.). This study supports the role of these proinflammatory cytokines as inflammatory mediators in active stage of disease.

HLA-DRß1*04 typing by simple in-house PCR-SSP technique for rheumatoid arthritis patients.

A strong association between rheumatoid arthritis (RA) and human leukocyte antigen (HLA) has been observed in many different populations and that accounts for approximately one-third of the genetic component of RA susceptibility. The greatest effect comes from DRß1 gene where the strongest association has been found with DRß1*04 (DR4) allele. As serology has some disadvantages over polymerase chain reaction (PCR)-based techniques and commercially available PCR-based kits are expensive, this study was aimed to standardize simple in-house PCR-SSP technique. Accuracy of this test was further checked with standard PCR-SSOP (RLS) results. The frequency HLA-DRß1*04 was significantly increased among RA patients when compared with normal controls. In this study, a very simple, convenient and more cost-effective in-house PCR-SSP technique was standardized for HLA-DRß1*04 typing that is helpful to RA diagnosis in developing countries like India, which can be used as a good screening test.