CircPIAS1 promotes hepatocellular carcinoma progression by inhibiting ferroptosis via the miR-455-3p/NUPR1/FTH1 axis.

BACKGROUND: The role of circRNAs in hepatocellular carcinoma (HCC) progression remains unclear. CircPIAS1 (circBase ID: hsa_circ_0007088) was identified as overexpressed in HCC cases through bioinformatics analysis. This study aimed to investigate the oncogenic properties and mechanisms of circPIAS1 in HCC development. METHODS: Functional analyses were conducted to assess circPIAS1's impact on HCC cell proliferation, migration, and ferroptosis. Xenograft mouse models were employed to evaluate circPIAS1's effects on tumor growth and pulmonary metastasis in vivo. Bioinformatics analysis, RNA immunoprecipitation, and luciferase reporter assays were utilized to elucidate the molecular pathways influenced by circPIAS1. Additional techniques, including RNA pulldown, fluorescence in situ hybridization (FISH), chromatin immunoprecipitation (ChIP), qPCR, and western blotting, were used to further explore the underlying mechanisms. RESULTS: CircPIAS1 expression was elevated in HCC tissues and cells. Silencing circPIAS1 suppressed HCC cell proliferation and migration both in vitro and in vivo. Mechanically, circPIAS1 overexpression inhibited ferroptosis by competitively binding to miR-455-3p, leading to upregulation of Nuclear Protein 1 (NUPR1). Furthermore, NUPR1 promoted FTH1 transcription, enhancing iron storage in HCC cells and conferring resistance to ferroptosis. Treatment with ZZW-115, an NUPR1 inhibitor, reversed the tumor-promoting effects of circPIAS1 and sensitized HCC cells to lenvatinib. CONCLUSION: This study highlights the critical role of circPIAS1 in HCC progression through modulation of ferroptosis. Targeting the circPIAS1/miR-455-3p/NUPR1/FTH1 regulatory axis may represent a promising therapeutic strategy for HCC.

Mesenchymal Stem Cells Promote Polarization of M2 Macrophages in Mice with Acute-On-Chronic Liver Failure via Mertk/JAK1/STAT6 Signaling.

Acute-on-chronic liver failure (ACLF) is a severe disease with a high mortality. Macrophage-related inflammation plays a crucial role in ACLF development. Mesenchymal stem cells (MSCs) treatment was demonstrated to be beneficial in ACLF in our previous study; however, the underlying mechanisms remain unknown. Therefore, mouse bone marrow-derived MSCs were used to treat an ACLF mouse model or cocultured with RAW264.7/J774A.1 macrophages that were stimulated with LPS. Histological and serological parameters and survival were analyzed to evaluate efficacy. We detected changes of Mer tyrosine kinase (Mertk), JAK1/STAT6, inflammatory cytokines, and markers of macrophage polarization in vitro and in vivo. In ACLF mice, MSCs improved liver function and 48-h survival of ACLF mice and alleviated inflammatory injury by promoting M2 macrophage polarization and elevated Mertk expression levels in macrophages. This is significant, as Mertk regulates M2 macrophage polarization via the JAK1/STAT6 signaling pathway.

Mesenchymal stem cell-regulated miRNA-mRNA landscape in acute-on-chronic liver failure.

BACKGROUND: Acute-on-chronic liver failure (ACLF) is a major challenge in the field of hepatology. While mesenchymal stem cell (MSC) therapy can improve the prognosis of patients with ACLF, the molecular mechanisms through which MSCs attenuate ACLF remain poorly understood. We performed global miRNA and mRNA expression profiling via next-generation sequencing of liver tissues from MSC-treated ACLF mice to identify important signaling pathways and major factors implicated in ACLF alleviation by MSCs. METHODS: Carbon tetrachloride-induced ACLF mice were treated with saline or mouse bone marrow-derived MSCs. Mouse livers were subjected to miRNA and mRNA sequencing. Related signal transduction pathways were obtained through Gene Set Enrichment Analysis. Functional enrichment, protein-protein interaction, and immune infiltration analyses were performed for the differentially expressed miRNA target genes (DETs). Hub miRNA and mRNA associated with liver injury were analyzed using LASSO regression. The expression levels of hub genes were subjected to Pearson's correlation analysis and verified using RT-qPCR. The biological functions of hub genes were verified in vitro. RESULTS: The tricarboxylic acid cycle and peroxisome proliferator-activated receptor pathways were activated in the MSC-treated groups. The proportions of liver-infiltrating NK resting cells, M2 macrophages, follicular helper T cells, and other immune cells were altered after MSC treatment. The expression levels of six miRNAs and 10 transcripts correlated with the degree of liver injury. miR-27a-5p was downregulated in the mouse liver after MSC treatment, while its target gene E2f2 was upregulated. miR-27a-5p inhibited E2F2 expression, suppressed G1/S phase transition and proliferation of hepatocytes, in addition to promoting their apoptosis. CONCLUSIONS: This is the first comprehensive analysis of miRNA and mRNA expression in the liver tissue of ACLF mice after MSC treatment. The results revealed global changes in hepatic pathways and immune subpopulations. The miR-27a-5p/E2F2 axis emerged as a central regulator of the MSC-induced attenuation of ACLF. The current findings improve our understanding of the molecular mechanisms through which MSCs alleviate ACLF.

The impact of serum thyroid-stimulation hormone levels on the outcome of hepatitis B virus related acute-on-chronic liver failure: an observational study.

BACKGROUND: Thyroid dysfunction has been reported in severe liver diseases. The aim of this study was to analyze the impact of serum thyroid-stimulation hormone (TSH) levels on the prognosis of patients with hepatitis B virus (HBV)-related acute-on-chronic liver failure (ACLF). METHODS: This retrospective cohort study included 1,862 patients with HBV-related ACLF. Risk factors associated with 30-day and 90-day survival, hazard ratios (HRs), and 95% confidence intervals (CIs) for TSH were estimated using Cox proportional hazards regression. The Area Under the ROC curve (AUROC) analysis was carried out, and the cut-off values were calculated. After grouping by the cut-off value, survival was compared between the groups using the log-rank test. This study data is from the "Survival Cohort Study (SCS)", which has been registered at ClinicalTrials.gov (NCT03992898). RESULTS: Multivariate analysis indicated that an elevated TSH level was a highly significant predictor for 30-day survival (HR = 0.743, 95% CI: 0.629-0.878, P < 0.001) and 90-day survival (HR = 0.807, 95% CI: 0.717-0.909, P < 0.001). The AUROC of TSH level for 30-day and 90-day mortality were 0.655 and 0.620, respectively, with the same best cut-off values of 0.261 µIU/mL. Log-rank test showed that the group with higher TSH level had higher 30-day (78.5%, 95% CI: 76.1%-80.9% vs. 56.9%, 95% CI: 53.4%-60.4%; P < 0.001) and 90-day survival rate (61.5%, 95% CI: 58.6%-64.4% vs. 42.8%, 95% CI: 39.3%-46.3%; P < 0.001). Similar findings were observed in subgroups analysis. After adjusting for age and other risk factors, the higher level of TSH remained associated with 30-day survival (HR = 0.602, 95% CI: 0.502-0.721, P < 0.001) and 90-day survival (HR = 0.704, 95% CI, 0.609-0.814, P < 0.001). CONCLUSIONS: Serum TSH level significantly correlate with HBV-related ACLF patients' survival and may be of value for predicting 30-day and 90-day survival of patients with HBV-related ACLF.

Risk factors for underlying comorbidities and complications in patients with hepatitis B virus-related acute-on-chronic liver failure.

Hepatitis B virus-related acute-on-chronic liver failure (HBV-ACLF) is a severe and life-threatening complication, characterised by multi-organ failure and high short-term mortality. However, there is limited information on the impact of various comorbidities on HBV-ACLF in a large population. This study aimed to investigate the relationship between comorbidities, complications and mortality. In this retrospective observational study, we identified 2166 cases of HBV-ACLF hospitalised from January 2010 to March 2018. Demographic data from the patients, medical history, treatment, laboratory indices, comorbidities and complications were collected. The mortality rate in our study group was 47.37%. Type 2 diabetes mellitus was the most common comorbidity, followed by alcoholic liver disease. Spontaneous bacterial peritonitis, pneumonia and hepatic encephalopathy (HE) were common in these patients. Diabetes mellitus and hyperthyroidism are risk factors for death within 90 days, together with gastrointestinal bleeding and HE at admission, HE and hepatorenal syndrome during hospitalisation. Knowledge of risk factors can help identify HBV-ACLF patients with a poor prognosis for HBV-ACLF with comorbidities and complications.

Associations between nephrolithiasis and diabetes mellitus, hypertension and gallstones: A meta-analysis of cohort studies.

AIM: To review and clarify the strengths and directions of associations between nephrolithiasis and hypertension (HTN), diabetes mellitus (DM) and gallstones (GS) given the inconsistent results reported in cohort studies. METHODS: Relevant literature was searched in PubMed and EMBASE from inception to July 2019, for cohort studies that examined the relationships between kidney stones and these three diseases among adults. Pooled relative risks (RRs) were calculated by maximally adjusted risk estimates using a random effect model. Subgroup analysis, meta-regression and sensitivity analysis were conducted whenever appropriate. RESULTS: Of 3537 papers, 21 articles with each including 1 to 3 cohorts were identified. In this meta-analysis, nephrolithiasis was reciprocally linked to HTN, DM and GS. Kidney stones were significantly associated with 31%, 33% and 46% higher risks of incident HTN, DM and GS whereas GS was associated with a significantly higher risk of nephrolithiasis (RR: 1.49; 95% CI, 1.28-1.73), followed by HTN (RR: 1.30; 95% CI, 1.11-1.52) and DM (RR: 1.18; 95% CI, 1.07-1.29). Also, females with DM (RR: 1.29; 95% CI, 1.08-1.55) were more likely to develop kidney stones than diabetic male patients (RR: 0.91; 95% CI, 0.75-1.10). CONCLUSION: Although additional studies are needed to confirm these findings and elucidate the mechanisms, this study revealed possible bidirectional associations between nephrolithiasis and HTN, diabetes and GS, which reinforced the notion of nephrolithiasis as a systemic disease that requires comprehensive investigations.

Dietary and lifestyle factors for primary prevention of nephrolithiasis: a systematic review and meta-analysis.

BACKGROUND: Dietary and lifestyle factors may play an important role in the increasing prevalence of nephrolithiasis. We aimed to review and quantify the associations between lifestyle factors and incident nephrolithiasis and suggest lifestyle changes for the primary prevention of nephrolithiasis. METHODS: PubMed, EMBASE, and Cochrane Library were searched up to May 2019, for observational studies and randomized controlled trials (RCTs) that assessed modifiable lifestyle factors and risk of nephrolithiasis in adults. Pooled relative risks (RRs) and 95% confidence intervals (CIs) were computed using a random effects model. The I2 statistic was employed to evaluate heterogeneity. Subgroup analysis, sensitivity analysis and meta-regression were also conducted whenever possible. RESULTS: Fifty relevant articles with 1,322,133 participants and 21,030 cases in total were identified. Prominent risk factors for incident stones were body mass index (1.39,1.27-1.52), dietary sodium (1.38, 1.21-1.56), fructose, meat, animal protein, and soda. In contrast, protective factors included fluid intake (0.55, 0.51-0.60), a Dietary Approaches to Stop Hypertension (DASH) style diet (0.69, 0.64-0.75), alcohol (0.69, 0.56-0.85), water, coffee, tea, vegetables, fruits, dietary fiber, dietary calcium (0.83, 0.76-0.90), and potassium. Vitamin D (1.22, 1.01-1.49) and calcium (1.16, 1.00-1.35) supplementation alone increased the risk of stones in meta-analyses of observational studies, but not in RCTs, where the cosupplementation conferred significant risk. CONCLUSIONS: Several modifiable factors, notably fluid intake, dietary patterns, and obesity, were significantly associated with nephrolithiasis. Long-term RCTs are required to investigate the cost-effectiveness of dietary patterns for stone prevention. The independent and combined effects of vitamin D and calcium supplementation on nephrolithiasis need further elucidation.

Allogeneic bone marrow-derived mesenchymal stromal cells for hepatitis B virus-related acute-on-chronic liver failure: A randomized controlled trial.

Mortality from hepatitis B virus (HBV)-related acute-on-chronic liver failure (ACLF) is high due to limited treatment options. Preclinical and clinical investigations have proved that treatment with mesenchymal stromal cells (MSCs) is beneficial for recovery from liver injury. We hypothesized that the outcome of HBV-related ACLF would be improved by MSC treatment. From 2010 to 2013, 110 patients with HBV-related ACLF were enrolled in this open-label, nonblinded randomized controlled study. The control group (n = 54) was treated with standard medical therapy (SMT) only. The experimental group (n = 56) was infused weekly for 4 weeks with 1.0 to 10 × 105 cells/kg allogeneic bone marrow-derived MSCs and then followed for 24 weeks. The cumulated survival rate of the MSC group was 73.2% (95% confidence interval 61.6%-84.8%) versus 55.6% (95% confidence interval 42.3%-68.9%) for the SMT group (P = 0.03). There were no infusion-related side effects, but fever was more frequent in MSC compared to SMT patients during weeks 5-24 of follow-up. No carcinoma occurred in any trial patient in either group. Compared with the control group, allogeneic bone marrow-derived MSC treatment markedly improved clinical laboratory measurements, including serum total bilirubin and Model for End-Stage Liver Disease scores. The incidence of severe infection in the MSC group was much lower than that in the SMT group (16.1% versus 33.3%, P = 0.04). Mortality from multiple organ failure and severe infection was higher in the SMT group than in the MSC group (37.0% versus 17.9%, P = 0.02). CONCLUSION: Peripheral infusion of allogeneic bone marrow-derived MSCs is safe and convenient for patients with HBV-related ACLF and significantly increases the 24-week survival rate by improving liver function and decreasing the incidence of severe infections. (Hepatology 2017;66:209-219).

Dexamethasone in outcome of patients with hepatitis B virus-related acute-on-chronic liver failure.

BACKGROUND AND AIM: Acute-on-chronic liver failure (ACLF) caused by hepatitis B virus (HBV) is a severe disease with high mortality. Immune injury plays an important role during the early stage of the disease. Our research aimed to investigate the safety and efficacy of dexamethasone therapy for patients with HBV-related ACLF. METHODS: A total of 134 inpatients with HBV-induced ACLF were enrolled from January 2009 to December 2012. All the patients received the standard medicine treatment (SMT), among whom 31 cases underwent additional dexamethasone injection for three times (dexamethasone treatment [DMT] Group). A total of 35 patients (SMT Group) matched for baseline characters served as controls. Both the groups were followed up for 12 weeks. The survival rates, liver functions, and complications were recorded. RESULTS: The 12-week cumulative survival rates were 45.7% (16/35)and 48.4% (15/31) for SMT Group and DMT Group, respectively, and no significant differences were found (P = 0.959). There were no dramatic differences in liver function and model for end-stage liver disease (MELD) score at 1, 2, 4, 8, and 12 weeks between two groups. There were no significant differences in the incidence of complications (i.e. infection, gastrointestinal bleeding, encephalopathy, hepatorenal syndrome, and ascites) from 1 to 12 weeks between Group SMT and Group DMT. More than 40 ages, MELD score more than 28 and encephalopathy were independent risk factors for the mortality of patients. CONCLUSIONS: Dexamethasone cannot improve liver functions and 12-week survival rates of patients with HBV-related ACLF. Age, MELD score, and encephalopathy are independent risk factors.

Med1 inhibits ferroptosis and alleviates liver injury in acute liver failure via Nrf2 activation.

BACKGROUND: Extensive hepatocyte mortality and the absence of specific medical therapy significantly contribute to the unfavorable prognosis of acute liver failure (ALF). Ferroptosis is a crucial form of cell death involved in ALF. In this study, we aimed to determine the impact of Mediator complex subunit 1 (Med1) on ferroptosis and its potential hepatoprotective effects in ALF. RESULTS: Med1 expression is diminished in the liver of lipopolysaccharide (LPS)/D-galactosamine (D-GalN)-induced ALF mice, as well as in hepatocytes damaged by H2O2 or TNF-α/D-GalN in vitro. Med1 overexpression mitigates liver injury and decreases the mortality rate of ALF mice by ferroptosis inhibition. The mechanism by which Med1 inhibits erastin-induced ferroptosis in hepatocytes involves the upregulation of nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream antioxidant genes heme oxygenase-1 (HO-1), glutamate cysteine ligase catalytic (GCLC), and NAD(P)H quinone oxidoreductase 1 (NQO1). Furthermore, Med1 overexpression suppresses the transcription of proinflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in the liver of mice with LPS/D-GalN-induced ALF. CONCLUSION: Overall, our research findings indicate that Med1 suppresses ferroptosis and alleviates liver injury in LPS/D-GalN-induced ALF through the activation of Nrf2. These findings substantiate the therapeutic viability of targeting the Med1-Nrf2 axis as a means of treating individuals afflicted with ALF.

Imbalance of interleukin-17-producing CD4 T cells/regulatory T cells axis occurs in remission stage of patients with hepatitis B virus-related acute-on-chronic liver failure.

BACKGROUND AND AIM: Although regulatory T cells (Treg) and interleukin-17-producing CD4 T cells (Th17) have been demonstrated to play opposing roles in inflammation-associated diseases, their frequency and balance in different stages of hepatitis B virus (HBV)-related acute-on-chronic liver failure (ACLF) remain unknown. METHODS: Fourteen patients with HBV-associated ACLF were studied and defined into different stages according to disease activity. Circulating Th17 cells and Treg cells were analyzed by flow cytometry, and the cytokines were quantitated by enzyme-linked immunosorbent assay. Results were correlated with temporal changes in viral load, disease progression and compared with 30 chronic hepatitis B (CHB) subjects and 18 healthy subjects. RESULTS: We showed a significantly higher frequency of circulating Th17 cells in the remission stage of ACLF when compared with the progression stage, the CHB group, or normal controls. However, the frequency of circulating Treg cells was significantly lower in the remission stage of ACLF when compared with the progression stage or the CHB group. The increase in Th17 cells and concomitant decrease in Treg cells created an imbalance in the remission stage of ACLF patients, which negatively correlated with disease progression. In addition, we showed that ACLF patients in the remission stage had an altered profile of cytokines that regulated the induction of Th17 cells and Treg cells. CONCLUSIONS: ACLF patients in the remission stage had an imbalance of Th17 to Treg cells, which could be used as a prognostic marker to predict disease progression. This imbalance could play a role in the immunopathogenesis of HBV-related ACLF.

[Up-regulator of cell proliferation predicts poor prognosis in hepatocellular carcinoma].

OBJECTIVE: To explore the potential value of up-regulator of cell proliferation (URGCP) as a biomarker for predicting the prognosis of hepatocellular carcinoma (HCC). METHODS: The expression of URGCP was analyzed in 15 HCC cell lines and in 10 pairs of HCC and adjacent tissues with reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. The expression of URGCP in 278 paraffin-embedded, archived clinical HCC samples was analyzed by immunohistochemistry (IHC) and statistic analysis conducted to examine the relationship of prognosis and URGCP expression. RESULTS: IHC analysis revealed a high expression of URGCP in all HCC cell lines and in 122/278 (43.8%) paraffin-embedded archived HCC specimens. The expression level of URGCP was significantly correlated with clinical staging and poor patient survival of HCC in the study cohort and in various clinical subgroups, but not correlated with HCC patient age, tumor size, tumor number or alpha-fetoprotein level. CONCLUSION: URGCP plays an important role in promoting the proliferation and tumorigenesis of HCC and may represent a novel prognostic biomarker and therapeutic target for the disease.

Autologous bone marrow mesenchymal stem cell transplantation in liver failure patients caused by hepatitis B: short-term and long-term outcomes.

UNLABELLED: Our study aimed to investigate the short-term efficacy and long-term prognosis of liver failure patients caused by hepatitis B after a single transplantation with autologous marrow mesenchymal stem cells (MMSCs). A total of 527 inpatients with liver failure caused by hepatitis B were recruited and received the same medical treatments, among whom 53 patients underwent a single transplantation with autologous MMSCs. A total of 105 patients matched for age, sex, and biochemical indexes, including alanine aminotransferase (ALT), albumin, total bilirubin (TBIL), prothrombin time (PT), and Model for End-Stage Liver Disease (MELD), comprised the control group. A total of 120 mL of bone marrow was obtained from each patient and then diluted and separated. Then, the MMSC suspension was slowly transfused into the liver through the proper hepatic artery. The success rate of transplantation was 100%, without serious side effects or complications. Levels of ALB, TBIL, and PT and MELD score of patients in the transplantation group were markedly improved from 2-3 weeks after transplantation, compared with those in the control group. At 192 weeks of follow-up, there were no dramatic differences in incidence of hepatocellular carcinoma (HCC) or mortality between the two groups. Additionally, there were no significant differences in the incidence of HCC or mortality between patients with and without cirrhosis in the transplantation group. CONCLUSION: Autologous MMSC transplantation is safe for liver failure patients caused by chronic hepatitis B. Short-term efficacy was favorable, but long-term outcomes were not markedly improved. In respect to several parameters, this method is preferable for patients with liver cirrhosis and may have potential for reducing their incidence of HCC and mortality.

Safety and efficacy of Thymosin α1 in the treatment of hepatitis B virus-related acute-on-chronic liver failure: a randomized controlled trial.

BACKGROUND/PURPOSE OF THE STUDY: Mortality from hepatitis B virus (HBV)-related acute-on-chronic liver failure (ACLF) is high. Severe infection is the most important complication that affects the outcomes of ACLF patients. Thymosin α1 (Tα1) can improve immune imbalance and this study aimed to investigate the safety and efficacy of Tα1 treatment for HBV-related ACLF. METHODS: From 2017 to 2019, 120 patients with HBV-related ACLF were enrolled in this open-label, randomized, and controlled clinical trial (ClinicalTrial ID: NCT03082885). The control group (N = 58) was treated with standard medical therapy (SMT) only. The experimental group (N = 56) was subcutaneously injected with 1.6 mg of Tα1 once a day for the first week and then twice a week from week 2 to week 12. RESULTS: The 90-day cumulated liver transplantation free survival rate of the Tα1 group was 75.0% (95% confidence interval 63.2-86.8%) versus 53.4% (95% confidence interval 39.7-67.1%) for the SMT group (p = 0.030). No significant difference was found in the survival using competitive risk analysis. The incidences of new infection and hepatic encephalopathy in the Tα1 group were much lower than those in the SMT group (32.1% vs 58.6%, p = 0.005; 8.9% vs 24.1%, p = 0.029, respectively). Mortality from severe infection in the SMT group was higher than in the Tα1 group (24.1% vs 8.9%, p = 0.029). CONCLUSION: Tα1 is safe for patients with HBV-related ACLF and significantly improves the 90-day liver transplantation-free survival rate. There may be a subgroup which may benefit from Tα1 therapy by the mechanism of preventing infection.

[Safety evaluation and quality control of MSCs from hepatitis B patient in vitro].

OBJECTIVE: To get mesenchymal stem cells (MSCs) from hepatitis B patient and to valuate the safety and quality after long-term culture in vitro. METHODS: The cells obtained directly from bone marrow and cultured in Mesen Pro medium supplemented with FGF, and the morphology of MSCs was observed. Surface antigens of the MSCs were analyzed by flow-cytometry. The bacteria, virus, endotoxin and residual serum of cell suspension were detected. The MSCs and perpheral blood T lymphocytes were co-cultured in 48 well plates for 72 h and the T lymphocyte proliferation was measured by using MTT reduction method and the effect of MSCs on T lymphocyte transformation stimulated by PHA was also observed. The oncogenicity of MSCs was verified by the tumorigenesis test in sofo agar. The genetic stability of MSCs was examined by karyotype analysis. RESULT: The MSCs from hepatitis B patient could be passaged to many generations and had strong abilities of proliferation. They expressed stem cell-surface antigens and maintained normal karyotype, prevented the pollution of bacteria and viruses, inhibited the immune response of allogenic T lymphocytes and no oncogenicity found. CONCLUSION: The MSCs have proliferative potentials, can be passaged in long-term cultures in Mesen Pro medium without oncogenicity, can maintain normal karyotype, can inhibit the immune response of T lymphocytes and can alleviate the grafe-versus diseases. The MSCs can be served as a new type of cells in cell and gene therapy.

[Study on HBV antigens and IL-12 affecting T cell-mediated immunity in HBsAg carriers].

OBJECTIVE: To investigate the effect of HBV antigens and pathological mechanism of chronic HBV infection by analyzing the cellular immune function of peripheral blood mononuclear cells (PBMCs) from HBsAg carriers. METHODS: PBMCs were prepared from individuals with chronic asymptomatic HBV infection and cultured in the presence of different antigens and/ or cytokines. The levels of cytokines in culture supernatants were detected by ELISA method. The phenotype of the cells was detected by FACS. RESULTS: The levels of IFN y secreted by PBMCs from HBsAg carriers were (48.3+/-19.8) pg/ml, significantly lower than that from healthy controls (t = 3.023, P less than 0.05); The IFN y produced by PBMCs from HBeAg positive patients due to HBsAg and HBcAg stimulation were (50.4+/-51.6) pg/ml and (63.2+/-36.9) pg/ml, significantly lower than that of HBeAg negative patients (t = 2.468 and 3.184, P less than 0.05, respectively). The IL-12p70 secreted by PBMCs from HBeAg positive patients was also significantly lower than that of HBeAg negative patients (P less than 0.05); Exogenous IL-12 promoted significantly PBMCs to secrete IFN y (P less than 0.01) and IL-12 combined with HBV antigens activated CD8+CD45RA+CCR7+ and CD8+CD45RA-CD62L+ cells. IL-12 secreted by PBMCs decreased in HBeAg positive patients, which may be the crucial reason of viral persistence in chronic HBV carriers. Exogenous IL-12 combined with specific HBV antigen could promote the central memory CD8+ T cells to produce IFN y.

Rapid diagnosis of disseminated Mycobacterium mucogenicum infection in formalin-fixed, paraffin-embedded specimen using next-generation sequencing: A case report.

BACKGROUND: Mycobacterium mucogenicum (M. mucogenicum) belongs to the group of rapidly growing Nontuberculous mycobacteria. This microorganism is associated with a wide spectrum of infectious diseases. Due to a low detection rate or the time required for conventional culture methodology, a rapid and broad-spectrum method is necessary to identify rare pathogens. CASE SUMMARY: A 12-year-old immunocompetent girl presented with painful masses for five months. The first mass was found in the right upper quadrant of the abdomen, and was about 1 cm × 1.5 cm in size, tough but pliable in texture, with an irregular margin and tenderness. An abscess gradually formed and ulcerated with suppuration of the mass. Three new masses appeared on the back one by one. Chest computed tomography showed patchy and streaky cloudy opacities in both lungs. Needle aspiration of the abscess was performed, but the smear and conventional culture were negative, and the pathological examination showed no pathogens. We then performed next-generation sequencing using a formalin-fixed, paraffin-embedded specimen to identify the pathogen. A significantly high abundance of M. mucogenicum was detected. The patient's abscesses gradually decreased in size, while inflammation in both lungs improved following 12-wk of treatment. No recurrence was observed four months after the end of the one-year treatment period. CONCLUSION: Next-generation sequencing is a promising tool for the rapid and accurate diagnosis of rare pathogens, even when using a formalin-fixed, paraffin-embedded specimen.

Gut microbiota as prognosis markers for patients with HBV-related acute-on-chronic liver failure.

The gut microbiota in the hepatitis B virus related acute-on-chronic liver failure (HBV-ACLF) is poorly defined. We aim to uncover the characteristics of the gut microbiota in HBV-ACLF and in other HBV associated pathologies. We analyzed the gut microbiome in patients with HBV-ACLF or other HBV associated pathologies and healthy individuals by 16S rRNA sequencing and metagenomic sequencing of fecal samples. 212 patients with HBV-ACLF, 252 with chronic hepatitis B (CHB), 162 with HBV-associated cirrhosis (HBV-LC) and 877 healthy individuals were recruited for the study. CHB and HBV-LC patients are grouped as HBV-Other. We discovered striking differences in the microbiome diversity between the HBV-ACLF, HBV-Other and healthy groups using 16S rRNA sequencing. The ratio of cocci to bacilli was significantly elevated in the HBV-ACLF group compared with healthy group. Further analysis within the HBV-ACLF group identified 52 genera showing distinct richness within the group where Enterococcus was enriched in the progression group whilst Faecalibacterium was enriched in the regression group. Metagenomic sequencing validated these findings and further uncovered an enrichment of Lactobacillus casei paracasei in progression group, while Alistipes senegalensis, Faecalibacterium prausnitzii and Parabacteroides merdae dominated the regression group. Importantly, our analysis revealed that there was a rapid increase of Enterococcus faecium during the progression of HBV-ACLF. The gut microbiota displayed distinct composition at different phases of HBV-ACLF. High abundance of Enterococcus is associated with progression while that of Faecalibacterium is associated with regression of HBV-ACLF. Therefore, the microbiota features hold promising potential as prognostic markers for HBV-ACLF.