Alterations in CD4+ T-cell Subsets in Living Donor Liver Transplantation Associated With Graft Rejection.

Background and objectives: Regulatory T-cells (Tregs) play a key role in immune homeostasis after organ transplantation. However, the role of CD4+ T cell subsets in early acute rejection is still not well understood. Therefore, our aim was to determine changes in CD4+ T-cell subsets in living donor liver transplantation (LDLT). Methods: LDLT patients were assessed for T-cell subsets, Tregs frequencies and their functionality by flow-cytometry at peri- and post-transplant in the span of 1 year. Results: 33 patients were followed up and 11 (33%) patients have developed early acute cellular rejection (ACR). At peri-transplant time point, MFI of Foxp3+ Tregs was significantly increased compared to HC (P = 0.04). However, CD4+CD25+Foxp3+/CD127- Tregs numbers and IL-10, IL-17 and TGF-ß secreting functional Tregs were significantly decreased at 3 months compared to peri-transplant (P = 0.003). But in patients with rejection, CD4+CD25+FOXP3+ and CD4+CD25+CD127- Tregs were significantly decreased at day 3 compared to no rejection group (P = 0.048). Patients with rejection also showed significantly decreased numbers of IL-17 and TGF-ß secreting CD4+CD25+FOXP3+ Tregs at peri-transplant time (P = 0.04, P = 0.03) compared to no rejection. Further, rejection group showed decreased terminally differentiated effector memory (TEMRA) at peri-transplant and day 7 (P = 0.048 and P = 0.01). Additionally, CD4+ central memory (CM) was decreased at peri-transplant (P = 0.05), 1 month (P = 0.04), and 3 to 6 month (P = 0.02). Interpretation and conclusion: Tregs frequencies were significantly decreased in peri-TX in rejection patients. Further, decreased frequencies of CD4+ TEMRA and CD4+ CM at day 7 and 1 month were associated with rejection.

Long Term HBsAg Responses to Peg-Interferon Alpha-2b in HBeAg Negative Chronic Hepatitis B Patients Developing Clinical Relapse after Stopping Long-Term Nucleos(t)ide Analogue Therapy.

Background and aims: A high proportion of hepatitis B e antigen (HBeAg)-negative chronic hepatitis B (CHB) patients develop clinical relapse after stopping long-term nucleotide analogues (NAs). The aim of this study was to assess the efficacy of pegylated interferon (PEG-IFN) alpha 2b in inducing hepatitis B surface antigen (HBsAg) loss in such patients. Methods: NAs were stopped in 118 HBeAg-negative CHB patients fulfilling the Asian Pacific Association for the Study of Liver (APASL) 2015 criteria for stopping NAs; they had received NAs for a median interquartile range (IQR) of 60 (48-84) months. Results: Overall, 82 of 118 (69.5%) patients developed clinical relapse after stopping NAs; 44 within 12 months (and treated with PEG-IFN alpha 2b 1.5 mcg/kg weekly subcutaneous injections for 48 weeks); and 38 after 12 months [and treated with tenofovir alafenamide fumarate (TAF) 25 mg daily] of follow-up. The decision to treat with either PEG-IFN or TAF was not a time-bound decision but was due to logistical problems.During the median IQR follow-up of 48 (43.5-52.5) months after the start of PEG-IFN, 14 of 44 (31.8%) patients developed clinical relapse after stopping PEG-IFN and were started on TAF. At the last follow-up visit, HBsAg was found to be negative in 7/44 (15.9%) of patients receiving PEG-IFN.Among 38 patients treated with TAF for clinical relapse, during the median IQR follow-up of 18 (12-30) months after start of TAF, no patient became HBsAg negative.36 patients did not develop clinical relapse during the follow-up, and after a median IQR follow-up of 60 (60-60) months after stopping NAs, HBsAg negative was found in 1/36 (2.8%) of patient at the last follow-up. Conclusions: Among patients with HBeAg-negative chronic hepatitis B who developed clinical relapse after stopping long-term NAs therapy and were subsequently treated with PEG-IFN alpha 2b, 15.9% achieved HBsAg loss on long-term follow-up.

Metabolomic analysis shows dysregulation in amino acid and NAD+ metabolism in palmitate treated hepatocytes and plasma of non-alcoholic fatty liver disease spectrum.

There is an alarming increase in incidence of fatty liver disease worldwide. The fatty liver disease spectrum disease ranges from simple steatosis (NAFL) to steatohepatitis (NASH) which culminates in cirrhosis and cancer. Altered metabolism is a hallmark feature associated with fatty liver disease and palmitic acid is the most abundant saturated fatty acid, therefore, the aim of this study was to compare metabolic profiles altered in hepatocytes treated with palmitic acid and also the differentially expressed plasma metabolites in spectrum of nonalcoholic fatty liver. The metabolites were analyzed by liquid chromatography-mass spectrometry (LC-MS) platform. Hepatocyte cell lines PH5CH8 and HepG2 cells when treated with 400 µM dose of palmitic acid showed typical features of steatosis. Metabolomic analysis of lipid treated hepatocyte cell lines showed differential changes in phenylalanine and tyrosine pathways, fatty acid metabolism and bile acids. The key metabolites tryptophan, kynurenine and carnitine differed significantly between subjects with NAFL, NASH and those with cirrhosis. As the tryptophan-kynurenine axis is also involved in denovo synthesis of NAD+, we found significant alterations in the NAD+ related metabolites in both palmitic acid treated and also fatty liver disease with cirrhosis. The study underscores the importance of amino acid and NAD+supplementation as promising strategies in fatty liver disorder.

Immune predictors of hepatitis B surface antigen seroconversion in patients with hepatitis B reactivation.

BACKGROUND: Hepatitis B surface antigen (HBsAg) seroconversion is sometimes observed in hepatitis B reactivation (rHBV), probably due to immune resetting and differentiation. AIMS: To investigate sequential immune differentiation and abrogation of tolerance in patients with rHBV who achieved HBsAg seroconversion. METHODS: We included 19 patients with chronic hepatitis B (CHBV; HBV DNA log103-8 ), 67 with rHBV (raised ALT [>5XULN], HBV DNAlog104-8 ) and 10 healthy controls. Immune differentiation, tolerance and functional status of CD4, CD8, T regulatory cells (Tregs), B cells and follicular T helper (Tfh) cells were assessed at baseline and 24 weeks. RESULTS: At 24 weeks, 81% rHBV (n = 67) lost HBV DNA and HBeAg (41%), and 12 (19%) lost HBsAg and made anti-HBs titers >10 IU/ml. rHBV patients had higher Th1/17, TEM , Tfh, Tfh1/17, plasma and ATM B cells, and lower Tregs, Th2, Th17 and TEMRA expression. rHBV showed lower PD1, TIM3, LAG3, SLAM and TOX compared to CHBV. There was a significant increase in CD8, CD8EM, Tfh, Tfh1/17 and plasma B cells in seroconverters than non-seroconverters. At 24 weeks, we also observed increased plasma B cell frequency in seroconverters. While non-seroconverters showed higher expression of PD1, TIM3, LAG3, SLAM and TOX on CD4/CD8 T cells, blockade of PD1, TIM3, LAG3 and CTLA4 significantly enhanced IFN-γ, TNF-α, IL-4 and IL-21 expression on CD4/CD8 and Tfh cells in non-seroconverters. CONCLUSIONS: Non-seroconverters have increased inhibitory markers on CD4/CD8 T cells. There is a critical play of CD8, Tfh and B cells and subsets in seroclearance, along with checkpoint molecules as a potential therapy for non-seroconverters in HBV infection.

Presence of entry receptors and viral markers suggest a low level of placental replication of hepatitis B virus in a proportion of pregnant women infected with chronic hepatitis B.

The transplacental route of vertical transmission of Hepatitis B Virus (HBV) has been known for over a decade. Here we present evidence which suggest HBV can replicate in placenta. Forty-one HBsAg positive and 10 control pregnant women were enrolled in the study after obtaining informed consent. HBV positives were further divided in the High Viral Load (HVL) Group and Low Viral Load (LVL) Group according to INASL guidelines 2018. The Presence of the HBV DNA and expression of NTCP in the placenta was analyzed by qPCR/RT-qPCR and/or immunohistochemistry (IHC). The presence of cccDNA was assessed using Digital Droplet PCR while the presence of pre-genomic (pg) RNA was assessed through qRT-PCR and sequencing. The presence of HBeAg and HBcAg in the placenta was assessed by IHC. Immunostaining of NTCP, HBeAg and HBcAg on trophoblasts along with the presence of total HBV DNA, cccDNA and pgRNA indicated, that these cells are not only susceptible to HBV infection but may also support viral replication. This is further supported by the finding that trophoblasts of the several HBeAg seronegative samples harbored the HBeAg. Although, we did not find any correlation in NTCP expression and viral markers with viral load indicates placental replication may not aping hepatocytes. The presence of the HBV receptor, NTCP along with the presence of cccDNA, pgRNA, and HBeAg in placenta of HBV infected females without circulating HBeAg suggest that placenta act as a replication host.

Secretome of senescent hepatoma cells modulate immune cell fate by macrophage polarization and neutrophil extracellular traps formation.

Presence of dysfunctional senescent hepatocytes is a hallmark feature of liver cirrhosis which finally culminates in liver cancer. We now report the presence of senescent hepatocytes (p21 and p53 positive) in the vicinity of infiltrated immune cells in hepatocellular carcinoma tissue specimens by immunohistochemistry. Hence, we evaluated in vitro, the relevance of senescent hepatoma cells in altering the fate of monocytes and neutrophils by assaying for macrophage polarization and extracellular trap (NETs) formation, respectively. Premature senescence was induced in hepatoma cells (HepG2 and Huh7 cells) by treating cells with doxorubicin. Senescent hepatoma cells showed strong inflammatory phenotype with induced expression of cytokines (IL1ß, IL6, IL8 and IL13) as evaluated by flow cytometry. The senescent secretome from hepatoma cells when incubated with healthy monocytes caused it to differentiate predominantly towards M2 fate (CD80low CD86low CD163high CD206high) when analysed by flow cytometry. This was corroborated by the finding in clinical samples where human hepatocellular carcinoma harbouring senescent hepatocytes showed presence of M2 macrophages, while M1 macrophages were predominant in non-tumorous region. Additionally, the senescent secretome from Huh7 cells enhanced the NETs formation, while HepG2 secretome had an inhibitory effect. In conclusion, the "pro-inflammatory" senescent secretome drives non-inflammatory type M2 macrophage polarization and modulated neutrophil traps which in turn can influence the tumor microenvironment.

Plasma Proteomic Analysis Identified Proteins Associated with Faulty Neutrophils Functionality in Decompensated Cirrhosis Patients with Sepsis.

Decompensated cirrhosis (DC) is susceptible to infections and sepsis. Neutrophils and monocytes provide the first line of defense to encounter infection. We aimed to evaluate proteins related to neutrophils functionality in sepsis. 70 (DC), 40 with sepsis, 30 without (w/o) sepsis and 15 healthy controls (HC) plasma was analyzed for proteomic analysis, cytokine bead array, endotoxin, cell free DNA and whole blood cells were analyzed for nCD64-mHLADR index, neutrophils-monocytes, functionality and QRT-PCR. nCD64-mHLADR index was significantly increased (p < 0.0001) with decreased HLA-DR expression on total monocytes in sepsis (p = 0.045). Phagocytic activity of both neutrophils and monocytes were significantly decreased in sepsis (p = 0.002 and p = 0.0003). Sepsis plasma stimulated healthy neutrophils, showed significant increase in NETs (neutrophil extracellular traps) and cell free DNA (p = 0.049 and p = 0.04) compared to w/o sepsis and HC. Proteomic analysis revealed upregulated- DNAJC13, TMSB4X, GPI, GSTP1, PNP, ANPEP, COTL1, GCA, APOA1 and PGAM1 while downregulated- AHSG, DEFA1,SERPINA3, MPO, MMRN1and PROS1 proteins (FC > 1.5; p < 0.05) associated to neutrophil activation and autophagy in sepsis. Proteins such as DNAJC13, GPI, GSTP1, PNP, ANPEP, COTL1, PGAM1, PROS1, MPO, SERPINA3 and MMRN1 showed positive correlation with neutrophils activity and number, oxidative burst activity and clinical parameters such as MELD, MELD Na and Bilirubin. Proteomic analysis revealed that faulty functionality of neutrophils may be due to the autophagy proteins i.e., DNAJC13, AHSG, TMSB4X, PROS1 and SERPINA3, which can be used as therapeutic targets in decompensated cirrhosis patients with sepsis.

Granulocyte-Macrophage Colony-Stimulating Factor Modulates Myeloid-Derived Suppressor Cells and Treg Activity in Decompensated Cirrhotic Patients With Sepsis.

Background: Decompensated cirrhosis patients are more prone to bacterial infections. Myeloid-derived suppressor cells (MDSCs) expand in sepsis patients and disrupt immune cell functions. Granulocyte-macrophage colony-stimulating factor (GM-CSF) therapy helps in restoring immune cell functions and resolving infections. Its role in MDSC modulation in cirrhosis with sepsis is not well understood. Methods: A total of 164 decompensated cirrhotic-62 without (w/o), 72 with sepsis, and 30 with sepsis treated with GM-CSF-and 15 healthy were studied. High-dimensional flow cytometry was performed to analyze MDSCs, monocytes, neutrophils, CD4 T cells, and Tregs at admission and on days 3 and day 7. Ex vivo co-cultured MDSCs with T cells were assessed for proliferation and apoptosis of T cells and differentiation to Tregs. Plasma factors and mRNA levels were analyzed by cytokine-bead assay and qRT-PCR. Results: Frequencies of MDSCs and Tregs were significantly increased (p = 0.011 and p = 0.02) with decreased CD4 T cells (p = 0.01) in sepsis than w/o sepsis and healthy controls (HCs) (p = 0.000, p = 0.07, and p = 0.01) at day 0 and day 7. In sepsis patients, MDSCs had increased IL-10, Arg1, and iNOS mRNA levels (p = 0.016, p = 0.043, and p = 0.045). Ex vivo co-cultured MDSCs with T cells drove T-cell apoptosis (p = 0.03, p = 0.03) with decreased T-cell proliferation and enhanced FOXP3+ expression (p = 0.044 and p = 0.043) in sepsis compared to w/o sepsis at day 0. Moreover, blocking the MDSCs with inhibitors suppressed FOXP3 expression. GM-CSF treatment in sepsis patients significantly decreased MDSCs and FOXP3+ Tregs but increased CD4 T-cell functionality and improved survival. Conclusion: MDSCs have an immunosuppressive function by expanding FOXP3+ Tregs and inhibiting CD4+ T-cell proliferation in sepsis. GM-CSF treatment suppressed MDSCs, improved T-cell functionality, and reduced Tregs in circulation.

The Clock-NAD+ -Sirtuin connection in nonalcoholic fatty liver disease.

Nonalcoholic or metabolic associated fatty liver disease (NAFLD/MAFLD) is a hepatic reflection of metabolic derangements characterized by excess fat deposition in the hepatocytes. Identifying metabolic regulatory nodes in fatty liver pathology is essential for effective drug targeting. Fatty liver is often associated with circadian rhythm disturbances accompanied with alterations in physical and feeding activities. In this regard, both sirtuins and clock machinery genes have emerged as critical metabolic regulators in maintaining liver homeostasis. Knockouts of either sirtuins or clock genes result in obesity associated with the fatty liver phenotype. Sirtuins (SIRT1-SIRT7) are a highly conserved family of nicotinamide adenine dinucleotide (NAD+)-dependent deacetylases, protecting cells from metabolic stress by deacetylating vital proteins associated with lipid metabolism. Circadian rhythm is orchestrated by oscillations in expression of master regulators (BMAL1 and CLOCK), which in turn regulate rhythmic expression of clock-controlled genes involved in lipid metabolism. The circadian metabolite, NAD+ , serves as a crucial link connecting clock genes to sirtuin activity. This is because, NAMPT which is a rate limiting enzyme in NAD+ biosynthesis is transcriptionally regulated by the clock genes and NAD+ in turn is a cofactor regulating the deacetylation activity of sirtuins. Intriguingly, on one hand the core circadian clock regulates the sirtuin activity and on the other hand the activated sirtuins regulate the acetylation status of clock proteins thereby affecting their transcriptional functions. Thus, the Clock-NAD+-Sirtuin connection represents a novel "feedback loop" circuit that regulates the metabolic machinery. The current review underpins the importance of NAD+ on the sirtuin and clock connection in preventing fatty liver disorder.

Asian Pacific association for the study of liver (APASL) guidelines: hepatitis B virus in pregnancy.

Hepatitis B virus (HBV) infection still remains a major public health issue in the Asia-Pacific region. Most of the burden of HBV-related disease results from infections acquired in infancy through perinatal or early childhood exposure to HBV in Asia-Pacific. Hepatitis B during pregnancy presents unique management issues for both the mother and fetus. These APASL guidelines provide a comprehensive review and recommendations based on available evidence in the literature, for the management of females with HBV infection through every stage of pregnancy and postpartum. These also address the concerns, management challenges, and required follow-up of children born to hepatitis B-positive mothers.

Immune Surveillance by Myeloid-Derived Suppressor Cells in Liver Diseases.

BACKGROUND: Myeloid-derived suppressor cells (MDSCs) are immunosuppressive in nature, originate in the bone marrow, and are mainly found in the blood, spleen, and liver. In fact, liver acts as an important organ for induction and accumulation of MDSCs, especially during infection, inflammation, and cancer. In humans and rodents, models of liver diseases revealed that MDSCs promote regeneration and drive the inflammatory processes, leading to hepatitis, fibrogenesis, and cirrhosis, ultimately resulting in hepatocellular carcinoma. SUMMARY: This brief review is focused on the in-depth understanding of the key molecules involved in the expansion and regulation of MDSCs and their underlying immunosuppressive mechanisms in liver diseases. KEY MESSAGE: Modulated MDSCs can be used for therapeutic purposes in inflammation, cancer, and sepsis.

Application of CRISPR-Cas9 based gene editing to study the pathogenesis of colon and liver cancer using organoids.

Two breakthrough techniques that have totally revolutionized biology in last 1 decade are the discovery of genome editing tools and growing the stem cells/primary tissue explants in defined 3D culture. In this regard the discovery of CRISPR-Cas9 as a specific gene editing tool and organoid culture from adult stem cell has provided easy handy tools to uncover the process of organ development and also modeling cancer. Genetically modified organoids have been developed by sequential knockout and knockin of driver mutations by genome editing followed by niche-based selection. The modified organoids when xenotransplanted in animal models faithfully recapitulate the neoplastic events of human tumors. The present review focuses on the merging of these two powerful technologies in understanding the complexities of colon and liver cancer.

Higher circulating natural killer cells and lower lactate levels at admission predict spontaneous survival in non-acetaminophen induced acute liver failure.

Massive cellular necrosis in acute liver failure (ALF) is dominantly immune mediated and innate immune cells are major pathophysiological determinants in liver damage. In fifty ALF and fifteen healthy, immune cells phenotyping by flow-cytometry, DAMPs using ELISA were analysed and correlated with clinical and biochemical parameters. ALF patients (aged 27 ± 9 yr, 56% males, 78% viral aetiology) showed no difference in neutrophils and classical monocytes, but significantly increased intermediate monocytes (CD14+CD16+) (p < 0.01), decreased non-classical monocytes (CD14-CD16+) and CD3-veCD16+CD56+ NK cells compared to HC. ALF patients who survived, showed higher NK cells (9.28 vs. 5.1%, p < 0.001) among lymphocytes and lower serum lactate levels (6.1 vs. 28, Odds ratio 2.23, CI 1.27-3.94) than non- survivors had higher. Logistic regression model predicted the combination of lactate levels with NK cell percentage at admission for survival. In conclusion, Combination of NK cell frequency among lymphocytes and lactate levels at admission can reliably predict survival of ALF patients.

Mechanistic and physiological approaches of fecal microbiota transplantation in the management of NAFLD.

Non-alcoholic fatty liver disease (NAFLD) is a multifaceted disease allied with various metabolic disorders, obesity and dysbiosis. Gut microbiota plays an influential role in the pathogenesis of NAFLD and other metabolic disorders. However, recent scientific upsurge emphasizes on the utility of beneficial gut microbiota and bacteriotherapy in the management of NAFLD. Fecal microbiota transplantation (FMT) is the contemporary therapeutic approach with state-of-the-art methods for the treatment of NAFLD. Other potential therapies include probiotics and prebiotics supplements which are based on alteration of gut microbes to treat NAFLD. In this review, our major focus is on the pathological association of gut microbiota with progression of NAFLD, historical aspects and recent advances in FMT with possible intervention to combat NAFLD and its associated metabolic dysfunctions.

Impact of human leukocyte antigen compatibility on outcomes of living donor liver transplantation: Experience from a tertiary care center.

INTRODUCTION: The role of HLA compatibility in kidney, heart, and stem cell transplantation is well known, but with regard to living donor liver transplantation (LDLT), there is a different scenario. In the present study, we aim to examine the effects of donor-recipient HLA mismatches at A, B, and DR loci on various outcomes of LDLT-like graft survival, early allograft dysfunction (EAD), acute rejection, length of hospital (LOH) stay, sepsis, and cytomegalovirus (CMV) reactivation. METHODS: This is a retrospective single center study of a cohort of adult patients who underwent first time ABO-compatible (ABOc) LDLT between January 2010 and December 2018. Transplants with incomplete records or without HLA typing data were excluded. Donor-recipient HLA-A, B, and DR mismatches were assessed in the host versus graft (HVG) direction and were correlated with various post-transplant outcomes. RESULTS: Among 140 transplants being evaluated, approximately two third had total HLA mismatches between 2 and 3. HLA mismatches at each locus as well as cumulative HLA mismatches did not show any association with overall graft survival, EAD, acute rejection episodes, and LOH stay. However, the presence of minimum one mismatch at HLA-A and DR loci was associated with the development of CMV reactivation (P = .03) and sepsis (P = .02) post-LDLT respectively. CONCLUSION: HLA mismatch is not associated with acute rejection, early graft dysfunction, and overall survival in LDLT. Its impact on CMV reactivation and sepsis needs further evaluation.

The role and delicate balance of Host Immunity in Coronavirus Disease-19.

Severe Acute Respiratory Syndrome (SARS) associated with SARS-CoV-2, causes a severe form of the respiratory illness known as Coronavirus Disease-19 (COVID-19). COVID-19 has emerged as a worldwide pandemic with a high number of fatalities. Approximately 112,654,202 people have been infected so far with this disease which has led to the death of more than one point seven million (2,496,749) till 24th Feb, 2021. Measures to counter this disease have led to a global economic slowdown. Multiple drug trials are ongoing and several putative candidates for vaccination against the virus have been approved and are in the pipeline. Many studies have also characterized the immunological profile of patients infected with COVID-19. Some studies suggest that the severity of the COVID-19 infection is directly associated with the cytokine storm. In this review, we aim to compile the available knowledge and describe the nature of immune responses in patients infected with COVID-19 in different age groups, comorbidity, and immune-compromised state and their association with disease severity.

Humoral Immune Responses and Hepatitis B Infection.

BACKGROUND: Chronicity or seroclearance of hepatitis B virus (HBV) antigens is determined by the host immune responses. Current approaches to treat HBV patients are based on inhibition of replication using different antivirals (nucleoside or nucleotide analogs) as monotherapy, or along with immune modulators as combination therapy is being used worldwide for reducing the viral load. Understanding the role of immune cellular therapies with currently available treatments for persistent viral-mediated responses in HBV patients is unexplored. However, the generation of antibodies against a surface (HBs) and envelop (HBe) antigen of hepatitis B remains an issue for future studies and needs to be explored. SUMMARY: Humoral immunity, specifically T follicular helper (TFh) cells, may serve as a target for therapy for HBsAg seroconversion. In this review, we have been engrossed in the importance and role of the humoral immune responses in CHBV infection and vertical transmission. Key Message: TFh cells have been suggested as the potential target of immunotherapy which lead to seroconversion of HBe and HBs antigens of HBV. HBsAg seroconversion and eradication of covalently closed circular DNA are the main challenges for existing and forthcoming therapies in HBV infection.

PNPLA3 polymorphisms are associated with raised alanine aminotransferase levels in hepatitis C virus genotype 3.

BACKGROUND AND STUDY AIMS: Hepatitis C virus (HCV) infection is one of the leading causes of end-stage liver diseases. This study aimed to determine the association between polymorphisms in interleukin 28B (IL28B), PNPLA3, toll-like receptor 7 (TLR7), nucleotide-binding oligomerization domain-containing protein 2 (NOD2) and retinoic inducible gene-I (RIG-I) and HCV genotype and clinical presentation in an Indian population. PATIENTS AND METHODS: A total of 500 patients with chronic HCV were enrolled in 19 centres across India. Genomic DNA was extracted from whole blood samples, and single nucleotide polymorphisms (SNPs) for IL28B, PNPLA3, TLR7, NOD2 and RIG-I genes were genotyped by real-time PCR using a TaqManSNP genotyping assay. RESULTS: The mean age of the patients was 45 + 13 years, and the most common genotype observed was HCV genotype 3 (54%), followed by genotype 1 (24%). Although the allelic frequencies of TLR7, NOD2 and RIG-I were in significant disequilibrium in HCV patients compared with those in controls, the PNPLA3 polymorphism correlated significantly with higher viral load and alanine aminotransferase (ALT) levels in genotype 3 patients. Patients with PNPLA3 CG/GG genotypes, along with IL28B genotype CC, had higher levels of ALT than those with other genotypes. CONCLUSION: These results indicate that PNPLA3 polymorphisms are associated with higher ALT levels in HCV genotype 3 patients in India and can help in identifying people who are at greater risk of developing HCV-associated liver diseases.

Role of Microbiota in Pathogenesis and Management of Viral Hepatitis.

Hepatitis is a condition that can be self-limiting or can progress to fibrosis (scarring), cirrhosis, or liver cancer. These days, gut microbiota becomes an important part of our immune system, which is important for disease progression or recovery. Translocation of gut microbial and metabolic products causes intestinal inflammation by modulating immune cells leading to impairment of the primary barrier. But there are limited studies discussing pathogenesis and management of hepatitis with gut microbiota. In this review, we have discussed the role of gut microbiota in pathogenesis and management of various hepatitis, especially hepatitis B and C. We have discussed the role of bacterial products, LPS-TLR4 pathway, and unmethylated CpG DNA, which ultimately affects downstream NF-kB signaling in hepatitis. Finally, we have discussed the role of fecal microbiota transplantation in the management of hepatitis.

In vitro targeted screening and molecular docking of stilbene, quinones, and flavonoid on 3T3-L1 pre-adipocytes for anti-adipogenic actions.

In metabolic disorders like obesity, NAFLD and T2DM, adipocytes are dysfunctional. Hence, pharmacological interventions have importance in preventing differentiation of adipocytes and stimulating lipid uptake. We, therefore, investigated the effects of arbutin (ARB), purpurin (PUR), quercetin (QR), and pterostilbene (PTS) on adipocyte differentiation and lipid uptake using 3T3-L1 adipocytes. Further, in silico docking studies were achieved to investigate interactions of ARB, PUR, QR, and PTS with beta-ketoacyl reductase (KR) and thioesterase (TE) domains of fatty acid synthase (FAS) enzyme. Mature 3T3-L1 adipocytes were used to investigate the anti-adipogenic effect of selected pharmacological agents by Oil Red O staining and in vitro fatty acid uptake analysis. Molecular docking studies were performed to predict the binding interactions of selected compounds with KR and TE domains of FAS enzyme. All these agents significantly decrease the adipocyte differentiation and showed the stimulatory effect on fatty acid uptake in 3T3-L1 adipocytes. However, PTS and PUR proved to be anti-adipogenic, whereas ARB and QR showed significant effect on fatty acid uptake, compared to others. Similarly, all the compounds displayed significant binding interactions with KR and TE domains of FAS enzyme, supporting the results of in vitro studies. Graphical abstract.