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.

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.

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.

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.

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.

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.

Inhibition of NOTCH signaling pathway chemosensitizes HCC CD133+ cells to vincristine and 5-fluorouracil through upregulation of BBC3.

In hepatocellular carcinoma (HCC), the poor response to the chemotherapeutic agents is partially attributed to the chemoresistance property of cancer stem cells (CSCs). NOTCH signaling pathway plays a crucial role in the chemoresistance through the maintenance of the CSCs. We observed that the NOTCH pathway was activated in HCC CD133+ cells treated with vincristine (VIN)1 and 5-fluorouracil (5-FU)2. Therefore, we examined whether inhibition of the NOTCH can improve sensitization of HCC CD133+ cells to VIN and 5-FU. The Huh7 cell line was pre-incubated γ-secretase DAPT, as a NOTCH inhibitor, and then treated with IC50 dose of VIN or 5-FU. The CD133+ cells were then isolated and analyzed for the cell viability, apoptosis, migration and spheroid formation capacities, and gene and protein expression. It was observed that pre-incubation with DAPT significantly downregulated the expression of NOTCH-related genes and led to a significant reduction in VIN- and 5-FU-CD133+ population. In addition, DAPT pre-incubated VIN- and 5-FU-treated-CD133+ cells formed fewer spheroids in 3D culture and had a lesser migration capacity in 2D culture. Importantly, DAPT enhanced the apoptosis rate of VIN- and 5-FU-treated CD133+ cells for 3- and 2-fold, which was correlated with the enhanced expression of pro-apoptotic BBC3 (BCL-2-binding component 3) and decreased expression of HES1 that was reported to regulate BBC3 negatively. Collectively, it was observed that NOTCH inhibition sensitized the HCC CD133+ cells to VIN and 5-FU through enhancing BBC3-mediated apoptosis. The results highlighted the role of NOTCH/HES1/BBC3 axis in resistance of CD133+ cells to VIN and 5-FU. Understanding the molecular mechanisms underlying chemoresistance in HCC CD133+ cells may help in designing the novel targeted therapies to chemosensitize them.

Natural Killer Cells Contribute to Pathogenesis of Severe Alcoholic Hepatitis by Inducing Lysis of Endothelial Progenitor Cells.

BACKGROUND: Endothelial progenitor cells (EPCs) help in neovascularization and endothelial repair during injury. Patients with cirrhosis show increased number and function of EPCs in circulation. METHODS: Since natural killer (NK) cells regulate EPCs, we investigated the relationship between the 2 in alcoholic cirrhosis (AC, n = 50) and severe alcoholic hepatitis (SAH, n = 18) patients and compared with nonalcoholic cirrhosis (n = 15) and healthy controls (HC, n = 30). Levels of systemic inflammatory cytokines were measured, and coculture assays were performed between EPCs and NK cells in contact-dependent and contact-independent manner. NK cell-mediated killing of EPCs was evaluated, and expression of receptors including fractalkine (FKN) on EPCs and its cognate receptor CX3CR1 on NK cells was studied by RT-PCR assays. RESULTS: Patients with SAH had higher regulated on activation, normal T cell expressed and secreted (RANTES) (p = 0.01), vascular endothelial growth factor (VEGF) (p = 0.04), IL-1ß (p = 0.04), and IL-6 (p = 0.00) growth factors and proinflammatory cytokines as compared to AC and HC. Distinct populations of CD31+ CD34+ EPCs with low and high expression of CD45 were significantly lower in SAH than HC (CD45low , p = 0.03; CD45hi , p = 0.04) and AC (CD45low , p = 0.05; CD45hi , p = 0.02). SAH patients, however, showed increased functional capacity of EPCs including colony formation and LDL uptake. NK cells were reduced in SAH compared with AC (p = 0.002), however with higher granzyme ability (p < 0.001 and p = 0.04, respectively). In SAH, EPC-NK cell interaction assays showed that NK cells lysed the EPCs in both contact-dependent and contact-independent assays. Expression of interaction receptor CX3CR1 was significantly higher on NK cells (p = 0.0005), while its cognate receptor, FKN, was increased on EPCs in SAH patients as compared to HC (p = 0.0055). CONCLUSION: We conclude that in SAH, NK cells induce killing of EPCs via CX3CR1/FKN axis that may be one of the key events contributing to disease severity and proinflammatory responses in SAH.

Dichotomy of Notch signalling in regulating tumour immune surveillance.

Notch signalling is an evolutionarily conserved multifaceted pathway that controls diverse cellular processes. Its role in regulating development and tissue homeostasis is well established. Aberrant activation of the Notch pathway has been implicated in the initiation and progression of many types of cancers. However, although in some cancers Notch signalling acts as a tumour-promoter, in others it is reported to suppress tumour growth and progression. Accumulating evidence suggests the involvement of both the innate and adaptive immune system in the development of various tumours. Currently, extensive studies on investigating the effects of Notch signalling in tumour immune surveillance are being carried out. Interestingly, recent literature shows how the changing expression of Notch genes in different T cell subsets like CD4 and CD8 helps in controlling anti-tumour immune responses. In this review, we discuss in depth the roles of Notch signalling molecules and different immune cells in the context of the tumour microenvironment. We also outline how current knowledge can be exploited to develop novel therapies in order to control the propagation of cancer stem cells.

Increased interleukin-23 receptor (IL-23R) expression is associated with disease severity in acute-on-chronic liver failure.

BACKGROUND: Th17 cells mediated immune response is important in chronic hepatitis B (CHB) infection and inflammation associated diseases; however, little is known about their immunopathogenic role in acute-on-chronic liver failure (ACLF). Interleukin-23 receptor (IL-23R) is essential for the generation of pathogenic Th17 cells; therefore, we aimed to evaluate IL-23R expression and its correlation with disease severity in ACLF. METHODS: Forty-two patients with ACLF (HBV and alcohol-related), thirty-two with CHB and twenty healthy controls (HC) were studied. Circulating and intrahepatic profile of Th17 cells and IL-23R was investigated. Association of IL-23R with disease severity was determined. RESULTS: Circulating Th17 cells were significantly increased in both ACLF groups (P = 0.03, P = 0.006) than CHB and HC. Percentage of Th17 cells was higher in liver than peripheral blood of ACLF patients (P = 0.04). Expression of IL-23R was immensely up-regulated on Th17 cells of ACLF patients. Importantly, IL-23R not only correlated with the increased percentage of Th17 cells but also had significant association with inflammation (P = 0.03) and clinical disease severity indices including Child-Turcotte-Pugh (P = 0.001) and Model for End-Stage Liver Disease (P = 0.002) scores. The ACLF non-survivors showed higher IL-23R expression (P = 0.01). Transcription factor retinoic acid receptor-related orphan nuclear receptor gamma-t (ROR-γt) was also high in circulation and in liver of ACLF patients and it positively correlated with ALT levels (P = 0.03). Surface receptors, including CCR6, IL-17R and pro-inflammatory cytokines IL-17A, IL-22, CXCL8 and GM-CSF were highly augmented in ACLF. CONCLUSION: ACLF patients express high IL-23R on Th17 cells which induces inflammation and strongly correlates with liver disease severity.

miR-26b-5p helps in EpCAM+cancer stem cells maintenance via HSC71/HSPA8 and augments malignant features in HCC.

BACKGROUND: Targeting cancer stem cells (CSCs) in hepatocellular carcinoma (HCC) is difficult because of their similarities with normal stem cells (NSCs). EpCAM can identify CSCs from EpCAM+AFP+HCC cases, but is also expressed on NSCs. We aimed to distinguish the two using integrated protein, mRNA and miRNA profiling. METHODS: iTRAQ based protein profiling and Next Generation Sequencing (NGS) was performed on EpCAM+/EpCAM- cells isolated from HCC (Ep+CSC, Ep- HCC) and EpCAM+ cells from non-cancerous/non-cirrhotic control liver tissues (Ep+NSC). Validations were done using qRT-PCR, flowcytometry and western blotting followed by in vitro and in vivo functional studies. RESULTS: 11 proteins were overexpressed (>3 fold) in Ep+CSCs compared to Ep- HCC and Ep+NSC cells. However, RNA-sequencing confirmed the Ep+CSC specific up-regulation of only HSPA8, HNRNPC, MPST and GAPDH mRNAs among these. Database search combined with miRNA profiling revealed Ep+ CSC specific down-regulation of 29 miRNAs targeting these four genes. Of these, only miR-26b-5p was found to target both HSPA8 and EpCAM. Validation of HSPA8 overexpression and miR-26b-5p down-regulation followed by linear regression analysis established a negative correlation between the two. Functional studies demonstrated that reduced miR-26b-5p expression increased the spheroid formation, migration, invasion and tumourigenicity of Ep+ CSCs. Furthermore, anti-miR-26b-5p increased the number of Ep+ CSCs with a concomitant overexpression of stemness genes and reduction of proapoptotic protein BBC3, which is a known substrate of HSPA8. CONCLUSION: miR-26b-5p imparts metastatic properties and helps in maintenance of Ep+ CSCs via HSPA8. Thus, miR-26b-5p and HSPA8 could serve as molecular targets for selectively eliminating the Ep+ CSC population in human HCCs.

Human papillomavirus (HPV) oncoprotein E6 facilitates Calcineurin-Nuclear factor for activated T cells 2 (NFAT2) signaling to promote cellular proliferation in cervical cell carcinoma.

The calcineurin-NFAT signaling pathway regulates cell proliferation, differentiation, and development in diverse cell types and organ systems. Deregulation of calcineurin-NFAT signaling has been reported in leukaemias and few solid tumors such as breast and colon. In the present study, we found elevated calcineurin protein levels and phosphatase activity in cervical cancer cell lines and depletion of the same attenuated cell proliferation. Additionally, nuclear levels of NFAT2, a downstream target of calcineurin, viz, was found elevated in human papillomavirus (HPV) infected cells, HeLa and SiHa, compared to the HPV negative cells, HaCaT and C33A, indicative of its higher DNA binding activity. The nuclear levels of both NFAT1 and NFAT3 remain unaltered implicating they have little role in cervical carcinogenesis. Similar to the in vitro studies, the HPV infected human squamous cell carcinoma specimens showed higher NFAT2 levels compared to the normal cervical epithelium. Depletion of NFAT2 by RNAi attenuated growth of SiHa cells. Overexpression of HPV16 oncoproteins viz, E6 and E7 increased NFAT2 expression levels and DNA binding activity, while knockdown of E6 by RNAi decreased the same. Briefly, we now report an activation of calcineurin-NFAT2 axis in cervical cancer and a novel role of HPV oncoprotein in facilitating NFAT2 dependent cell proliferation.

Differentially regulated gene expression in quiescence versus senescence and identification of ARID5A as a quiescence associated marker.

In multicellular organisms majority of the cells remain in a non-dividing states of either quiescence (reversible) or senescence (irreversible). In the present study, gene expression signatures unique to quiescence and senescence were identified using microarray in osteosarcoma cell line, U2OS. It was noted that certain genes and pathways like NOD pathway was shared by both the growth arrest conditions. A major highlight of the present study was increased expression of number of chemokines and cytokines in both quiescence and senescence. While senescence-associated secretory phenotype (SASP) is well known, the quiescence-associated secretory phenotype (QASP) is relatively unknown and appeared novel in this study. ARID5A, a subunit of SWI/SNF complex was identified as a quiescence associated gene. The endogenous expression of ARID5A increased during serum starved condition of quiescence. Overexpression of ARID5A resulted in more number of cells in G0/G1 phase of cell cycle. Further ARID5A overexpressing cells when subjected to serum starvation showed a pronounced secretory phenotype. Overall, the present work has identified gene expression signatures which can distinguish quiescence from senescence.

Immune correlates of hepatitis B surface antigen spontaneous seroconversion in hepatitis B e antigen negative chronic hepatitis B patients.

BACKGROUND: Hepatitis B surface antigen (HBsAg) seroconversion in HBeAg -ve chronic hepatitis B (CHB) infection is rare, possibly due to poor antigen processing and impaired humoral response. We investigated the role of dendritic cells (DCs), T follicular helper (TFH) cells and plasma B cells in seroconversion. METHODS: HBeAg -ve (n=135) CHB patients with raised ALT at baseline were followed up. Patients undergoing HBsAg seroconversion (Gr. I, n=11) were compared with non-converters with low (Gr. II, n=17, HBV DNA<2000 IU/mL) or high HBV DNA (Gr. III, HBV DNA >2000 IU/mL, n=17). We measured cell phenotypes (TFH, B and DCs), HBV specific T-cell functionality [using pooled overlapping surface and core peptides], IL21 levels and gene expression analysis by qRT-PCR. RESULTS: Patients in Gr. I compared to Gr. II and III, had higher IL-21 levels (865 vs 276 vs 111 pg/mL, P=<.0001), TFH (CD4+ CXCR5+ ) cells (12.3 vs 4.67 vs 2.77, P=<.001), inducible T-cell co-stimulator (ICOS) expression on TFH cells (20.0 vs 13.0 vs 13.68, P=.01), HBsAg specific IL-17 (9.40 vs 2.33 vs 2.61, P=<.001) and TNF-α secreting TFH17 cells (82 vs 1.43 vs 2.33, P=<.001), plasma B (CD19+ CD38+ ) cells (15.0 vs 5.08 vs 5.57, P=<.001), myeloid (17.80 vs 5.39 vs 2.70, P=<.001) and plasmocytoid DCs (2.6 vs 0.43 vs 0.21, P=<.001). Plasma B-cell frequency (R2 =.64, P=.01) and IL-21 levels (R2 =.52, P=.003) correlated with anti-HBs titres in patients with HBsAg seroconversion. CONCLUSIONS: Dendritic cell and TFH cell mediated responses regulate humoral responses against HBV and play a major role in HBsAg seroconversion in CHB patients.

Placental expression of asialoglycoprotein receptor associated with Hepatitis B virus transmission from mother to child.

BACKGROUND: Asialoglycoprotein receptor expression on hepatocytes has been associated with endocytosis, binding and uptake of hepatitis B virus. The role of asialoglycoprotein receptor in hepatitis B virus vertical transmission and its expression on placenta has not yet been studied. PATIENTS AND METHODS: Thirty-four HBsAg+ve and 13 healthy pregnant mothers along with their newborns were enrolled. The former were categorized into transmitting and non-transmitting mothers based on their newborns being hepatitis B surface antigen and hepatitis B virus DNA positive. Expression of asialoglycoprotein receptor and hepatitis B surface antigen in placenta and isoform of asialoglycoprotein receptor on dendritic cell in peripheral and cord blood dendritic cells were analysed using flowcytometry, immune histochemistry, immune florescence and qRT-PCR. RESULTS: Twelve HBsAg+ve mothers transmitted hepatitis B virus to their newborns whereas the rest (n = 22) did not. Hepatitis B virus-transmitting mothers showed increased expression of asialoglycoprotein receptor in trophoblasts of placenta. Immunofluorescence microscopy revealed colocalization of hepatitis B surface antigen and asialoglycoprotein receptor in placenta as well as in DCs of transmitting mothers. There was no significant difference in the expression of asialoglycoprotein receptor on peripheral blood mononuclear cells or chord blood mononuclear cells between the 2 groups. However, hepatitis B virus-transmitting mothers and their HBsAg+ve newborns showed increased mRNA levels of isoform of asialoglycoprotein receptor on dendritic cell in peripheral blood mononuclear cells. Hepatitis B virus-transmitting mothers and their HBsAg+ve newborns showed an increased expression of isoform of asialoglycoprotein receptor on dendritic cell on circulating dendritic cells compared to hepatitis B virus non-transmitting mothers and their negative newborns. CONCLUSIONS: This study revealed that increased expression of asialoglycoprotein receptor in placenta and colocalization with hepatitis B surface antigen strongly indicates its role in intrauterine transmission of hepatitis B virus. Asialoglycoprotein receptor-blocking strategy can be used for therapeutic intervention of vertical transmission.

Bone marrow stem cells and their niche components are adversely affected in advanced cirrhosis of the liver.

UNLABELLED: Bone marrow (BM) is a reservoir for immune and hematopoietic cells and critical for tissue repair and regeneration. All of these functions are severely altered in cirrhosis. We investigated the cellular and functional state of BM in cirrhosis patients. We studied the histological, cellular, and molecular changes in BM of cirrhosis patients (n = 168) and controls (n = 44). Hematopoietic stem cells (HSCs) and associated niche cells, mesenchymal stem cells, Schwann cells, neural fibers, and endothelial cells were evaluated by immunohistochemistry. Cytokines and growth factors were analyzed in peripheral blood and BM plasma. Cirrhotic BM showed an inverse correlation between cluster of differentiation 34+HSCs and Model of End-Stage Liver Disease (ρ = -0.582, P < 0.001) and Child's scores (P < 0.038). BMs of cirrhosis patients with higher Model of End-Stage Liver Disease (>15) showed significantly decreased HSCs, mesenchymal stem cells, Schwann cells, and neural fibers; increased interleukin-1ß (P = 0.004), tumor necrosis factor-α (P = 0.040), and interferon-γ (P = 0.03); and decreased oncostatin M (P = 0.04), stem cell factor (P = 0.05), and stromal cell-derived factor 1 (P = 0.03) compared to those with lower Model of End-Stage Liver Disease scores (≤15). The cluster of differentiation 34+ cell population was a predictor for the development of sepsis (P < 0.001), and per unit loss increased the probability of sepsis by 16%. Cirrhosis patients with fewer HSCs had lower hemoglobin (P = 0.05) and platelet counts (P = 0.05) and showed early graft dysfunction. CONCLUSIONS: Increasing severity of cirrhosis causes derangement of the hematopoietic niche and loss of HSCs, contributing to the hematological and immunological dysfunctions and reduced potential for regeneration; restoring BM functions could provide new therapeutic options in cirrhosis. (Hepatology 2016;64:1273-1288).

Randomized controlled trial comparing lactulose plus albumin versus lactulose alone for treatment of hepatic encephalopathy.

BACKGROUND: Hepatic encephalopathy (HE) is associated with poor prognosis and treatment of HE is primarily directed at the reduction of the blood ammonia levels. The study evaluated the efficacy and safety of albumin plus lactulose versus lactulose alone for treatment of overt HE. METHODS: In prospective randomized controlled trial, 120 patients with overt HE were randomized in two groups: group A lactulose plus albumin (n = 60) and group B lactulose alone (n = 60). Primary end point was complete reversal of HE, and secondary end points were mortality and hospital stay. RESULTS: A total of 120 patients (mean age 40.4 ± 9.3 years) were included in this study. Thirty-six (30%) patients were in Child-Turcotte-Pugh (CTP) class B, and 84 (70%) were in CTP Class C. Mean CTP score was 9.8 ± 2.1, and model for end-stage liver disease score was 26.1 ± 5.3. Twenty seven (22.5%) had grade 2, 57 (47.5%) had grade 3, and 36 (30%) had grade 4 HE at the time of admission. Forty-five (75%) patients in group A compared with 32 (53.3%) patients in group B had complete reversal of HE (P = 0.03). Mortality was significantly lower in lactulose plus albumin group (11[18.3%]) versus lactulose alone (19 [31.6%], [P < 0.05]). There was significant decrease in levels of arterial ammonia, interleukin-6, interleukin-18, tumor necrosis factor-alpha, and endotoxins after treatment in both groups; however, the delta decrease was significantly higher in group A compared with group B. Hospital stay was shorter in group A. CONCLUSIONS: Combination of lactulose plus albumin is more effective than lactulose alone in treatment of overt HE.

Interleukin-28b CC genotype predicts early treatment response and CT/TT genotypes predicts non-response in patients infected with HCV genotype 3.

Response to antiviral therapy for hepatitis C virus (HCV) depends upon the genotype and host immune response. IL28b gene mutations have been shown to modulate host antiviral immune response against genotype 1. However, the predictive value of IL28b polymorphism in genotype 3 HCV patients is largely unknown. The association of IL28b polymorphism with virological response was studied in 356 patients with genotype 3 chronic HCV undergoing treatment with peg-interferon and ribavirin and was compared with matched controls. IL28b genotyping followed by DNA sequencing was performed to identify the CC, CT, or TT genotypes. Two log reduction of HCV RNA at Day 7 (Quick Viral Response, QVR) and HCV RNA negativity at Day 28 (Rapid Viral Response, RVR) were analyzed with CC and non-CC genotypes in addition to other predictors of response. The associations of alleles with the response patterns were predicted. Sustained viral response was seen in 250 (70.2%) patients and the IL28b genotype CC/CT/TT distribution was 61.1%; 30.5%; and 8.4%, respectively. The non-CC genotypes were significantly higher in non-responders when compared to responders (67.6% vs. 38.9%, P < 0.001). Interestingly, the rapid viral response in responders was observed in 72.7% with the CC genotype and in 27.2% with the non-CC genotype (P < 0.001). Multivariate analysis showed CC genotype as an independent factor predicting the sustained viral response in patients infected with HCV genotype 3. In conclusion, the IL28b CT/TT genotype strongly correlates with treatment non-response in patients infected with HCV genotype 3 and CC genotype of IL28b is associated with higher quick viral response.