Role of serum endotoxin, FGF19, TLR2, TNF-α, IL-12 and IL-10 in NAFLD-associated T2DM pathogenesis: Insights into Th1 bias and protective mechanisms.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) in non-obese patients is pathophysiologically distinct, exhibiting common immunological link with type-2 diabetes mellitus (T2DM). This study aims to delineate the role of Toll-like receptor 2 (TLR2)-mediated immuno-modulation along with its association with fibroblast growth factor receptor 4 (FGFR4) and its ligand fibroblast growth factor 19 (FGF19) in the pathogenesis of NAFLD without or with T2DM. METHODOLOGY: Blood samples were collected from patients with NAFLD (n = 90), NAFLD with T2DM (n = 90) and healthy cohorts (n = 90) with consent and clinical records. Real-time polymerase chain reaction (PCR), enzyme-linked immunoassay (ELIZA) and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) were used to analyze messenger ribonucleic acid (mRNA), protein expression and gene polymorphism. RESULTS: The molecular genetic analysis revealed the prevalence of variant allele(A) in FGFR4 gene in both cases compared to controls. The mRNA expression of FGF19 and TLR2 exhibited significant upregulation in NAFLD without T2DM compared to NAFLD with T2DM. Tumor necrosis factor-α (TNF-α) and interleukin-12 (IL-12) showed upregulation in both disease cohorts compared to control while IL-10 showed significant downregulation in NAFLD with T2DM compared to the other two cohorts. Correlation analysis between FGF19 and TLR2 revealed significant positive association in both NAFLD with and without T2DM. The Th1:Th2 ratio showed significant upregulation in NAFLD with T2DM compared to NAFLD without T2DM. CONCLUSION: In conclusion, elevated serum endotoxin levels appear to contribute to NAFLD and T2DM development. Upregulated FGF19 seems to be protective against developing T2DM in NAFLD patients. Higher TLR2, TNF-α and IL-12 expression in NAFLD without T2DM suggests a Th1 bias in its pathogenesis, while reduced IL-10 in NAFLD with T2DM implies a more skewed Th1 state in this condition.

An investigation into the role of Notch signaling, altered angiogenesis, and inflammatory-induced preterm delivery and related complications in Northeast Indian patients.

INTRODUCTION: Notch signaling is crucial during pregnancy with ability to regulate angiogenesis and inflammatory response. Considering the enigmatic importance of Notch signaling in pregnancy including placenta development, gestational disorders and adverse pregnancy; we performed experimental analysis to identify the Notch receptor-ligands association with Preterm delivery (PTD) and linked complication. METHOD: A total of 245 cases [Term n = 135 and Preterm n = 110] were enrolled for the study from Northeast Indian Population. The differential mRNA expression of Notch receptors , ligands, its downstream target Hes1 and Immune markers (IL-10, IL-12 and TNF-α) was studied by real time polymerase chain reaction. Further the protein study of Notch1 and 4, Hes1, VEGF and TNF-α was performed by immunofluorescence. RESULTS: Placental mRNA expression of all the four notch receptors [Notch1 = 2.15 ± 1.02 fold, Notch2 = 6.85 ± 2.70 fold, and Notch3 = 1.74 ± 0.90 fold and Notch4 = 14.15 ± 6.72 fold]; ligands [JAG1 = 2.71 ± 1.22, JAG2 = 4.41 ± 2.31, DLL1 = 3.55 ± 1.38, DLL3 = 4.31 ± 2.82 and DLL4 = 3.07 ± 1.30 folds] and downstream target [Hes1 = 6.09 ± 2.89 folds] was elevated in PTD cases compared to Term delivery (TD) cases. The mRNA expression of pro-inflammatory marker (IL-12 = 3.99 ± 1.02 fold and TNF-α = 16.83 ± 2.97), was upregulated. The upregulated expression of Notch1(p < 0.001), JAG1 (p = 0.006), JAG2 (p = 0.009), DLL1 (p = 0.001), DLL4 (p < 0.001) Hes1 (p < 0.001), TNF-α (p < 0.001) and IL-12 (p = 0.006) were associated with the baby death; and Notch4 significantly inversely correlated with low birth weight (LBW). Consistently higher protein level expression of Notch1, Hes1, VEGFA and TNF-α was observed in preterm with highest expression in negative outcome cases. DISCUSSION: To conclude, the increased Notch1 expression and angiogenesis linked inflammation holds key in understanding the pathogenesis of PTD and linked complications and underlines its potential as therapeutic target for PTD interventions.

Oxidative stress-A key determinant of complications and negative outcome in hepatitis E virus infected pregnancies: A comprehensive account involving cases from northeast India.

Regulated oxidative stress (OS) is important during pregnancy. Sporadic studies suggest the significance of deregulated OS in hepatitis E virus (HEV) infected pregnancy, but with limited reactive oxygen species (ROS) or antioxidant markers. The present novel study, therefore, aimed to evaluate the significance of ROS-antioxidant imbalance and resulting altered OS in HEV infected pregnancy complications like preterm delivery (PTD) and outcome. Difference in serum levels of ROS and antioxidant panel of markers were evaluated by ELISA for HEV immunoglobulin M RNA positive genotype 1 cases (including acute [acute viral hepatitis, AVH] and fulminant [fulminant hepatic failure, FHF] cases) and healthy term delivery subjects, and analyzed statistically. Direct ROS marker H2 O2 levels and indirect OS marker for DNA damage 8-hydroxy-2'-deoxyguanosine was significantly increased in HEV-cases compared to controls, and was associated and prognostic factor for PTD and fetal death in HEV cases. A comparatively lower total serum antioxidant capacity was observed in the FHF cases compared to the control subjects and the AVH cases. Glutathione (GSH) levels and superoxide dismutase (SOD) activity were significantly associated with PTD in the FHF sub-cohorts (p = 0.017) and AVH sub-cohorts (p < 0.001), respectively, and was associated with poor prognosis in HEV cases. The serum H2 O2 levels were found to be negatively correlated with SOD activity (p = 0.016) and GSH levels (p = 0.001) in the HEV-AVH cases; and positively correlated with the viral load in HEV cases (p = 0.023). The ROS-antioxidant imbalance resulting OS plays a detrimental associative role in HEV infected pregnancy complications like PTD and adverse pregnancy outcomes; and holds therapeutic significance.

Meta-analysis of transcriptomics data identifies potential biomarkers and their associated regulatory networks in gallbladder cancer.

Aim: This study aimed to identify key genes, non-coding RNAs, and their possible regulatory interactions during gallbladder cancer (GBC). Background: The early detection of GBC, i.e. before metastasis, is restricted by our limited knowledge of molecular markers and mechanism(s) involved during carcinogenesis. Therefore, identifying important disease-associated transcriptome-level alterations can be of clinical importance. Methods: In this study, six NCBI-GEO microarray dataseries of GBC and control tissue samples were analyzed to identify differentially expressed genes (DEGs) and non-coding RNAs {microRNAs (DEmiRNAs) and long non-coding RNAs (DElncRNAs)} with a computational meta-analysis approach. A series of bioinformatic methods were applied to enrich functional pathways, create protein-protein interaction networks, identify hub genes, and screen potential targets of DEmiRNAs and DElncRNAs. Expression and interaction data were consolidated to reveal putative DElncRNAs:DEmiRNAs:DEGs interactions. Results: In total, 351 DEGs (185 downregulated, 166 upregulated), 787 DEmiRNAs (299 downregulated, 488 upregulated), and 7436 DElncRNAs (3127 downregulated, 4309 upregulated) were identified. Eight genes (FGF, CDK1, RPN2, SEC61A1, SOX2, CALR, NGFR, and NCAM) were identified as hub genes. Genes associated with ubiquitin ligase activity, N-linked glycosylation, and blood coagulation were upregulated, while those for cell-cell adhesion, cell differentiation, and surface receptor-linked signaling were downregulated. DEGs-DEmiRNAs-DElncRNAs interaction network identified 46 DElncRNAs to be associated with 28 DEmiRNAs, consecutively regulating 27 DEGs. DEmiRNAs-hsa-miR-26b-5p and hsa-miR-335-5p; and DElnRNAs-LINC00657 and CTB-89H12.4 regulated the highest number of DEGs and DEmiRNAs, respectively. Conclusion: The current study has identified meaningful transcriptome-level changes and gene-miRNA-lncRNA interactions during GBC and laid a platform for future studies on novel prognostic and diagnostic markers in GBC.

Impact of TNF-α profile in recurrent pregnancy loss pathogenesis: A patient based study from Assam.

BACKGROUND: Lacunae exist in understanding the underlying etiology in majority of recurrent pregnancy loss (RPL) cases. Given the significance of regulated immune-modulation in pregnancy, and the central role of pro-inflammatory TNF-α plays in it; this study targeted to appraise the significance of TNF-α profile in RPL pathogenesis in an ethnically distinct population from Assam, India. METHODS: Term delivery, medically terminated pregnancy (MTP) and RPL cases (based on ASRM criteria) were enrolled with no anatomical and chromosomal abnormalities or pathological infections; and blood and/or placenta/product of conceptus (POC) tissue samples were collected with informed consent. Serum level and tissue level TNF-α expression profile were screened using specific molecular tools, and was correlated with TNF-α -308 G/A genotype; for its association with RPL predisposition. RESULTS: A significant gestation specific increase in serum TNF-α levels was observed in MTP cases (19.932 ± 4.407 pg/mL) compared to term delivery subjects (p = 0.001), while a comparable levels were observed with RPL cases (22.709 ± 5.833 pg/mL) (p = 0.646). A site specific (POC) increased expression was observed in RPL compared to MTP cases at both at transcript (6.37 ± 3.714 folds) and protein levels. The TNF-α -308 variant genotype was associated with increased predisposition to RPL (OR = 1.721) compared to MTP as well as significantly increased serum TNF-α levels (p = 0.017); especially in subjects with a homozygous TNF-α -308 A/A genotype. CONCLUSION: Our data emphasizes on the importance of site specific TNF-α expression levels in RPL pathogenesis in the studied population, and underlines its importance in screening, clinical stratification, and therapeutics by molecular targeting using TNF-α inhibitors.

Increased homocysteine mediated oxidative stress as key determinant of hepatitis E virus (HEV) infected pregnancy complication and outcome: A study from Northeast India.

With the background of association of oxidative stress and Hepatitis E virus (HEV) infection in pregnancy complications the present novel study aimed to evaluate the significance of changes in maternal homocysteine levels and the related mechanism(s) in the pathophysiology of HEV related pregnancy complications and negative outcomes. Term delivery (TD, N = 194) and HEV-IgM positive pregnancy cases [N = 109] were enrolled. Serum and placental homocysteine levels were evaluated by ELISA and immunofluorescence and in turn correlated with serum Vitamin B12 levels. Distribution of variant MTHFR C➔T and TYMS1494del6bp genotyping were studied by PCR-RFLP. Differential folate receptor alpha (FR-α) expression in placenta was evaluated by real-time PCR and immunofluorescence respectively. The HEV viral load was significantly higher in both FHF and AVH cases. Higher serum homocysteine levels was associated with preterm delivery (PTD) and fetal death in HEV infected cases and was significantly inversely correlated with serum VitaminB12 levels in HEV cases. Placental homocysteine expression was upregulated in HEV cases, and in cases with negative pregnancy outcome. A Homocysteine level was associated with MTHFR C677T status. Genetic alterations in folate pathway was associated with increased risk of PTD in HEV infected pregnancy cases, disease severity, and negative pregnancy outcome in AVH and FHF groups. FR-α expression was downregulated in placental tissues of HEV infected pregnancy.Placental stress caused by HEV inflicted increased homocysteine due to alterations in maternal vitamin B12 levels and folate pathway components is detrimental mechanism in PTD and negative pregnancy outcome in HEV infected pregnancy cases and holds prognostic and therapeutic significance.

Prognostic, clinical, and therapeutic importance of RANTES-CCR5 axis in hepatitis A infection: A multiapproach study.

Fulminant hepatic failure (FHF) is a lethal manifestation of hepatitis A virus (HAV) infection, whose underlying mechanisms are poorly understood. We aimed to evaluate the importance of the modulation of the RANTES-chemokine receptor type 5 (CCR5) signaling axis and its immunomodulatory effects in directing hepatitis A disease pathogenesis using an in silico, in vitro and patient cohort-based approach. In silico interaction studies were performed using computation approaches with suitable software. Differential expression of relevant cytokines and immune cell markers were studied using real-time quantitative reverse transcription PCR (qRT-PCR), enzyme-linked immunosorbent assay, and flow-cytometry-based methods. In the HepG2 cell line, we studied inflammatory responses and susceptibility to HAV infection following RANTES stimulation and antibody blockade of CCR5. The HAV-VP3 region exhibited high interaction in CCR5: HAV complexes. RANTES levels were significantly increased in FHF cases. Reduced monocyte and T-cell activation were observed in FHF cases. RANTES expression inversely correlated with viremia but positively correlated with proinflammatory responses. Hyper Th1-biased immune responses, marked by high interleukin (IL)-12/IL-10 ratio were observed in FHF cases, which were also characterized by upregulated tumor necrosis factor-alpha (TNF-α) expression and reduced interferon-gamma expression. In vitro, RANTES was protective against HAV infection but resulted in upregulated TNF-α expression. Although viral load increased upon the regulation of inflammatory responses by CCR5 blocking, it was still significantly lower compared to control HAV-infected cells. Our study suggests the importance of RANTES-CCR5 signaling and linked-immunomodulation in HAV disease pathogenesis, as well as highlights the utility of CCR5 antagonists as a risk-reduction strategy in FHF patients. Our findings, therefore, have important implications for the management of high-risk HAV infections.

Exploring the p53 connection of cervical cancer pathogenesis involving north-east Indian patients.

BACKGROUND: As per WHO, Cervical cancer (CaCx) is a global issue, being the fourth common cancer in women with incidence rate of 13.1 per 1 lakh women globally and accounting for 311000 deaths in the year 2018 itself globally. The molecular pathogenesis in Human papillomavirus (HPV) infected cases is inconclusive. The detection of molecular factors leading to progression of CaCx can be important in the diagnosis and management of the disease. p53 a known tumor suppressor gene having a regulative role in cell cycle has been highlighted as key factor in the prevention of cancer but its significance in CaCx cases has been variably documented. The present study therefore targeted to evaluate the significance of p53 profile in CaCx cases in ethnically distinct northeast Indian population. METHODS: Blood and Tissue samples (N = 85) of cervical cancer patients were collected and screening for HPV was performed using PCR. Thereafter the differential mRNA expression(qPCR), Immunohistochemistry, Mutation (PCR direct sequencing method) of p53 was studied. Further p53 epigenetic profiling was done by Methylation specific PCR (MS-PCR) and western blotting by using p53 acetylation specific antibodies. RESULTS: Our findings revealed that the downregulation of p53 was associated with the progression of disease and the variation in downregulation based on p53 polymorphism was observed. Further hypermethylation and deacetylation of p53 was also found to be associated with the pathogenesis of CaCx. The downregulated expression and hypermethylation of p53 in lower grade of CaCx, together established its association with the progression of CaCx from lower to severe grade. CONCLUSION: Therefore, in CaCx patients of northeast Indian population, malfunctioning of p53 is found to have significant role in cervical cancer progression.

Immunoinformatics-aided identification of T cell and B cell epitopes in the surface glycoprotein of 2019-nCoV.

The 2019 novel coronavirus (2019-nCoV) outbreak has caused a large number of deaths with thousands of confirmed cases worldwide, especially in East Asia. This study took an immunoinformatics approach to identify significant cytotoxic T lymphocyte (CTL) and B cell epitopes in the 2019-nCoV surface glycoprotein. Also, interactions between identified CTL epitopes and their corresponding major histocompatibility complex (MHC) class I supertype representatives prevalent in China were studied by molecular dynamics simulations. We identified five CTL epitopes, three sequential B cell epitopes and five discontinuous B cell epitopes in the viral surface glycoprotein. Also, during simulations, the CTL epitopes were observed to be binding MHC class I peptide-binding grooves via multiple contacts, with continuous hydrogen bonds and salt bridge anchors, indicating their potential in generating immune responses. Some of these identified epitopes can be potential candidates for the development of 2019-nCoV vaccines.

Clinicopathological and immunohistochemical characteristics of breast cancer patients from Northeast India with special reference to triple negative breast cancer: A prospective study.

BACKGROUND: Molecular pathogenesis of Triple-negative breast cancer (TNBC) is inconclusively documented from resource limited countries and hence there is a lack of available targeted therapy for clinical interventions. Compared to other breast cancer subtypes, TNBC is more aggressive, higher recurrence rate, and higher prevalence in younger premenopausal women. Sporadic literature indicates predominance of TNBC in all reported breast cancer cases from Northeast India. AIM: This study was conducted to evaluate the candidature of panel of key molecular markers involved in the development and progression of TNBC for prognosis and futuristic tailored targeted therapy. MATERIALS AND METHODS: We analyzed the clinicopathological characterized and immunohistochemically screened the differential expression of key molecular markers involved in the development and progression of in TNBC cases vis-a-vis non-TNBC and autopsy-based control samples. RESULTS: TNBC tends to display at an early reproductive age and is more aggressive in nature. Further, the differential expression of 2 specific markers viz., epidermal growth factor receptor (EGFR) and FolR1 was higher in TNBC cases compared to controls and non-TNBC (both in terms of susceptibility and specificity), clinical staging in TNBC cases (severity) and mortality (outcome). Although Ki67 and vascular endothelial growth factor expression also correlated with severity and outcome of the disease but their differences in non-TNBC cases were not significantly differentiable compared to TNBC. CONCLUSIONS: The study indicates that EGFR and FolR1 could serve as useful biomarkers to determine TNBC prognosis. Further studies will be needed to evaluate EGFR and Folate pathways in order to screen out the molecular targets which may be meaningfully used for clinical stratification, intervention, and treatment.

Alterations in XRCC1 gene profile in pathogenesis of oral cavity cancer: A population-based study from Kamrup Urban District of Assam, India.

XRCC1 gene is an integral component of the base excision repair pathway regulating DNA repair, the genetic alterations in which has been documented to be associated with cancers of multiple etiologies. The present study aimed to evaluate the key polymorphisms in XRCC1 gene for its association with pathogenesis of oral cavity cancer (OCC) in Kamrup Urban District of Assam, India. Tissue biopsies (N = 152) clinicopathologically characterized OCCs were collected along with whole blood samples (N = 190) from healthy controls with all clinical and habitual details. A PCR-RFLP approach was used to study the XRCC1 polymorphisms, and statistical associations with pathogenesis were studied with SPSSv13.0 statistical software. The XRCC1 codon 194 polymorphism was significantly associated with the risk of OCC (odds ratio [OR] = 1.878, P = 0.048) and severity (OR = 2.221, P = 0.031). The presence of XRCC1 280 variant genotype increased the risk of OCC in exclusive smokers (OR = 3.818, P = 0.006), exclusive alcoholics (OR = 3.144, P = 0.027), and in exclusive areca nut chewers (OR = 3.055, P = 0.034). Human papilloma virus cases with any other habitual risk factor carrying XRCC1 280 genotype showed 3-fold significantly increased risk compared to controls (OR = 3.341, P = 0.022). The presence of XRCC1 codon 399 polymorphism was also found to be associated with significantly increased risk of oral cavity carcinoma (OR = 1.566, P = 0.049). Distribution of altered XRCC1 gene haplotype was higher in OCC cases. Polymorphisms in XRCC1 gene is associated with OCC pathogenesis in Kamrup Urban District, Assam, India, and is of prognostic significance. It is also suggestive of the importance of base excision repair pathway alterations in OCC pathogenesis.

Impact of modulation of telomerase and cancer stem-cell marker OCT4 axis in cervical cancer pathogenesis with underlying HPV16 infection.

Lacunae exist in the molecular event(s) specificity associated with cervical cancer (CaCx) pathogenesis. The present study aimed to evaluate the significance of telomerase-cervical cancer stem cells (CSCs) modulation in CaCx pathogenesis with underlying HPV16 infection. The study included HPV16 positive cases only (N = 65) of the total enrolled cases from Northeast India. The analysis of viral load and the differential messenger RNA expression of E6, E7, hTERT, hTR, and cancer stem-cell markers was studied by real-time polymerase chain reaction. Further the protein and colocalization study for E6, hTERT, and oct4 was performed by immunofluorescence. The real-time polymerase chain reaction based analysis showed an upregulation of HPV16 viral oncoprotein E6 and E7, and telomerase component hTERT and hTR expression and their correlation in CaCx susceptibility and severity. The hTERT expression correlated with viral load; while the E6 and telomerase protein expression colocalized in the nucleus. The CSCs marker octamer-binding transcription factor 4 (OCT4) was significantly upregulated in CaCx cases, was associated with CaCx susceptibility and severity, and colocalized with E6 expression in the nucleus as revealed from the immunofluorescence studies. To conclude, the telomerase-OCT4 axis modulation holds key in HPV16 CaCx pathogenesis mediated by HPV16 E6 viral oncoprotein expression, and underlines its potential for therapeutic targeting.

Significance of RANTES-CCR5 axis and linked downstream immunomodulation in Dengue pathogenesis: A study from Guwahati, India.

We aimed to evaluate the significance of the RANTES-CCR5 axis and resulting immunomodulatory status in Dengue pathogenesis involving a Guwahati, India based population where Dengue cases have increased alarmingly. An increased CC-chemokine receptor type 5 (CCR5) messenger RNA expression and CCR5 positive cell count profile was observed in Dengue cases, the highest being in severe cases. CCR5 ligand RANTES expression was significantly decreased in Dengue cases and inversely correlated with Dengue viremia fold change in severe cases. Monocytes are involved in Dengue virus homing and replication. Its levels and activation profile were higher in Dengue cases. A hyper Th1-biased immunomodulatory profile with upregulated tumor necrosis factor-α levels, and downregulated expression of antiviral cytokine interferon-γ and key regulatory Th2 anti-inflammatory cytokine interleukin 10 was observed in severe Dengue cases compared with mild Dengue cases and controls. The results, therefore, suggest the significance of RANTES-CCR5 axis deregulation and resulting altered immunomodulation in Dengue pathogenesis, and holds prognostic and therapeutic significance.

Oxidative stress and deregulations in base excision repair pathway as contributors to gallbladder anomalies and carcinoma - a study involving North-East Indian population.

Gallbladder cancer (GBC) is a fatal condition with dismal prognosis and aggressive local invasiveness; and with uncharacterised molecular pathology relating to non-specific therapeutic modalities. Given the importance of oxidative stress in chronic diseases and carcinogenesis, and the lacunae in literature regarding its role in gallbladder diseases, this study aimed to study the involvement of oxidative stress and deregulation in the base excision repair (BER) pathway in the pathogenesis of gallbladder diseases including GBC. This study involved patients from the North-East Indian population, where the numbers of reported cases are increasing rapidly and alarmingly. Oxidative stress, based on 8-OH-dG levels, was found to be significantly higher in gallbladder anomalies (cholelithiasis [CL] and cholecystitis [CS]) and GBC at the plasma and DNA level, and was associated with GBC severity. The expressions of key BER pathway genes were downregulated in gallbladder anomalies and GBC compared to controls, and in GBC compared to both non-neoplastic controls and gallbladder anomalies. Expression of XRCC1 and hOGG1 was significantly associated with both susceptibility and severity of GBC. The XRCC1 codon280 polymorphism was associated with disease susceptibility; and significantly higher oxidative stress was observed in hOGG1 genotypic variants. The genomes of GBC patients were found to be more hypermethylated compared to controls, with the promoters of XRCC1 and hOGG1 being hypermethylated and, therefore, being silenced. This study underlined the prognostic significance of the oxidative stress marker 8-OH-dG and BER pathway genes, especially hOGG1 and XRCC1, in gallbladder anomalies and GBC, as well as stated their potential for therapeutic targeting.

Genetic and expression changes in TNF-α as a risk factor for rheumatoid arthritis pathogenesis in northeast India.

Antitumour necrosis factor-alpha (TNF-α) therapy is used as a clinical intervention for rheumatoid arthritis (RA) but differences exist in response to the treatment which makes the candidature of the screening of TNF-α alteration(s) at genetic and expression levels an important agenda prior to treatment. This study aims to determine the associative role of TNF-α -308G/A polymorphism and differential expression of TNF-α in the pathogenesis of RA. A case-control study where a total of 126 RA patients were enrolled based on ACR-EULAR (2010) criteria, along with 160 community matched age and sex controls over a period of three years. The differential expression level of TNF-α mRNA and protein level was studied and TNF-α -308G/A polymorphism was screened by T-ARMS PCR assay. All statistical analysis was performed using SPSS software. mRNA expression level of TNF-α was upregulated in RA cases (avg. 15.85 ± 9.52 fold) compared to control. TNF-α protein level was found to be higher in RA cases (28.62±7.17 pg/mL) compared to control (23.14±6.91 pg/mL). TNF-α -308 variant GA genotype was higher in RA (46.03%) than in control (25%). The presence of TNF-α -308 variant A allele was associated with increased risk of RA susceptibility (odds ratio (OR) = 2.559 at 95% confidence interval (CI), P< 0.001) but not severity (OR = 1.617 at 95% CI, P = 0.571). The presence of -308 variant genotype was associated with a higher TNF-α mRNA and protein expression. The presence of TNF-α -308A allele is associated with increased risk of RA susceptibility and differential TNF-α expression, and has prognostic significance. Association of higher TNF-α pro-inflammatory cytokine levels with northeast Indian patients makes them suitable subjects for anti-TNF-α therapy.

Computational identification of hepatitis E virus-encoded microRNAs and their targets in human.

microRNAs (miRNAs) are small, noncoding RNAs which regulate eukaryotic gene expression via RNA interference pathway. Recently, miRNAs have been identified in a number of viruses with current evidence suggesting that they regulate gene expression in both virus and host. This makes viral miRNAs potential targets of clinical intervention, with the possibility of inhibiting aberrant host gene expression associated with the disease. In this study, computational approaches were taken to scan the hepatitis E virus (HEV) genome for putative pre-miRNA molecules, which were then analyzed for the presence of mature miRNAs. The 3'-untranslated region (3'-UTR) and 5'-UTR sequences targeted by these miRNAs were identified using Miranda computational tool, followed by the functional annotation of the associated messenger RNAs (mRNAs) using Gene Ontology terms and Kyoto Encyclopaedia of Genes and Genomes pathway analysis. We identified a total of nine viral encoded miRNAs in HEV. After functional annotation, the majority of the viral miRNA targets were found to be associated with cell cycle, cell differentiation, nitrogen compound metabolism, transmembrane transport, and chromosome organization. This in-silico study identified putative viral miRNAs encoded by HEV and their potential human mRNAs targets. These viral miRNAs have the potential to affect host gene expression as well as viral life cycle and pathogenesis and can, therefore, serve as potential therapeutic targets during HEV infection.

Deregulated TNF-Alpha Levels Along with HPV Genotype 16 Infection Are Associated with Pathogenesis of Cervical Neoplasia in Northeast Indian Patients.

Multiple factors are associated with human papillomavirus (HPV) infection related cervical anomalies and its progression to cervical carcinoma (CaCx), but data vary with respect to the underlying HPV genotype and with population being studied. No data are available regarding the role of immunological imbalance in HPV infected CaCx pathogenesis from Northeast India, which has an ethnically distinct population, and was aimed to be addressed through this study. The study included 76 CaCx cases, 25 cervical intraepithelial neoplasia (CIN) cases, and 50 healthy female controls. HPV screening and genotyping were performed by PCR. Differential expression of tumor necrosis factor alpha (TNF-α) was studied at serum level by enzyme-linked immunosorbent assay and tissue level by immunohistochemistry and messenger RNA (mRNA) level by real-time PCR. The data were correlated with interferon gamma (IFN-γ) and NF-κßp65 levels at protein level, as well as HPV16 E6 and E7 expression at transcript level statistically. HPV infection and HPV16 genotype were predominant in the studied cohort. TNF-α was found to be downregulated at both mRNA and protein levels in CaCx cases compared to controls; and the gradient downregulation correlated with progression of the disease from normal→CIN→CaCx. TNF-α expression correlated with insufficient modulation of both IFN-γ and NF-κßp65. The HPV16 E6 and E7 transcripts were found to be sharply upregulated in CaCx cases strongly inversely correlated with the TNF-α expression. Significant role of TNF-α downregulation associated with insufficient IFN-γ and total NF-κßp65 modulation and the resulting significant upregulation of viral transcripts E6 and E7 are key to the HPV16 infection mediated CaCx pathogenesis in northeast Indian patients.

Associative role of TYMS6bpdel polymorphism and resulting hyperhomocysteinemia in the pathogenesis of preterm delivery and associated complications: A study from Northeast India.

Aberrations including genetic alterations in folate pathway are detrimental in multiple disease pathogenesis, including pregnancy. The present study is based on the screening of the associative role of TYMS 14946bp deletion(del) polymorphism and associated hyperhomocysteinemia in susceptibility to preterm delivery (PTD), which is strongly associated with neonatal mortality and morbidity. METHODS: A total of 209 PTD cases {extremely preterm (n=22), very preterm (n=43) and moderately preterm (n=144)} and 194 term delivery cases were evaluated for TYMS 14946bp deletion and its association with preterm delivery, pregnancy outcome, baby birth weight and homocysteine estimation. RESULTS: The results showed that the distribution of TYMS 14946bp del/del genotype significantly increased the risk of PTD [OR=2.801, p=0.002] and is associated with fetal death. The TYMS 6bp ins/del and 6bp del/del genotype was associated with low birth weight (LBW) compared to 6bp ins/ins genotype in both term and PTD groups, and in case of very (p=0.024) and moderately (p=0.045) sub-cohorts of PTD significantly. Elevated serum homocysteine levels were significantly associated with PTD (p<0.001) and fetal death (p=0.013); and was also found to significantly correlate with TYMS 14946bp del/del genotype in all the pregnancy cases (p=0.008). TYMS 6bp del/del genotype was associated with higher homocysteine levels compared to ins/ins (p=0.005) and ins/del (p=0.062) genotypes within the PTD group. CONCLUSION: The study provides crucial information regarding the importance of TYMS6bpdel/del genotype and associated hyperhomocysteinemia in susceptibility to PTD, fetal death and LBW; and thus indicating their prognostic significance of TYMS 6bp del/del genotype in PTD which is of clinical importance.

Imbalance in leptin-adiponectin levels and leptin receptor expression as chief contributors to triple negative breast cancer progression in Northeast India.

BACKGROUND: Triple-Negative breast cancer (TNBC), accounts for a large percentage of breast cancer cases in India including Northeast India. TNBC has an unclear molecular aetiology and hence limited targeted therapies. Human breast is comprised of glandular, ductal, connective, and adipose tissues. Adipose tissue is composed of adipocytes. The adipocytes apart from being energy storage depots, are also active sources of adipocytokines and/or adipokines. The role of adipokines in breast cancer including TNBC has been sporadically documented. Two adipokines in particular, leptin and adiponectin, have come to be recognized for their influence on breast cancer risk and tumour biology. Therefore, the aim of this study was to understand the association of differential expression of critical adipokines and associated cellular mechanism in the susceptibility and severity of TNBC in northeast Indian population. MATERIALS AND METHODS: We collected 68 TNBC and 63 controls cases and examined for serum leptin and adiponectin levels using enzyme linked immunosorbent assay (ELISA). Leptin Receptor (Ob-R) mRNA expression was determined by real-time polymerase chain reaction (RT-PCR) assay. Differential Ob-R mRNA expression and correlation with cancer stem cell (CSC) markers was evaluated, and correlated with severity. RESULTS: The serum leptin levels were significantly associated with TNBC severity, while the adiponectin levels were comparative. The serum leptin levels correlated inversely with the adiponetin levels. Serum leptin levels were unaffected with difference in parity. The difference in leptin levels in pre and post menopausal cases were found to be statistically non-significant. Higher leptin levels were also found to be associated obesity, mortality and recurrence. Obesity was found to be a factor for TNBC pathogenesis and severity. Increased Ob-R mRNA expression was associated with TNBC, significantly with TNBC severity, and was significantly higher in obese patients with higher grade TNBC cases. The Ob-R gene mRNA expression was significantly higher in the obese TNBC cases showing recurrence or mortality. The higher Ob-R gene mRNA expression correlated significantly with higher serum leptin levels and lower serum adiponectin levels in TNBC cases. The Ob-R mRNA expression with associated with modulation of CSC oct4 and nanog. CONCLUSIONS: In conclusion, the present study is first of its kind on TNBC from northeast India, indicates that adipocytokines does play a role in TNBC pathogenesis. Thus, the understanding of molecular mechanisms of both leptin and adiponectin and their interplay in TNBC offer the prospects for new therapeutic approaches targeting similar signalling pathways.

Preterm delivery and associated negative pregnancy outcome - A tale of faulty progesterone receptor signalling pathway and linked derailed immunomodulation: A study from Northeast India.

Preterm delivery (PTD) is one of the potent contributor of neonatal mortality and morbidity, and the underlying cause in some situation is elusive. This study attempts to delineate the association of deregulation in progesterone receptor (PR) pathway and deleterious immune responses in predisposing patients to PTD in Northeast India, a region with high rate of PTD cases. A total of 109 cases of PTD and 100 term delivery cases were enrolled with all clinical details. The PTD cases were stratified based on gestation age at delivery. The differential expression of PR and key downstream effectors and cytokines were evaluated for correlation with PTD susceptibility, gestational period, and pregnancy outcome. The results indicated a sharp downregulation in PR expression is associated with PTD susceptibility, lower gestational period and negative pregnancy outcome. The PR downstream effector PIBF was also found to be downregulated in PTD, and is associated with gestational period and negative pregnancy outcome. The downregulation of PR and PIBF expression was found to correlate with a predominant Th1 state with higher CD56+NK cell counts and pro-inflammatory burst lead by hyper TNF-α, NF-kB and IFNγ expression, and complicated by lower IL10 expression, contributing to PTD as well as negative pregnancy outcome in the PTD cases. TNF-α expression in placenta inversely correlated with placental PR expression. To conclude, deregulation in PR pathway is a hallmark of preterm delivery and negative pregnancy outcome. Differential expression of several markers such as PR, PIBF and TNF-α has prognostic significance, and hence is of clinical significance.