5-FU promotes HBV replication through oxidative stress-induced autophagy dysfunction.

BACKGROUND & AIMS: Hepatitis B virus (HBV) reactivation is a major problem that must be overcome during chemotherapy for HBV-related hepatocellular carcinoma (HCC). However, the mechanism underlying chemotherapy-associated HBV reactivation is still not fully understood, hindering the development of improved HBV-related HCC treatments. METHODS: A meta-analysis was performed to assess the HBV reactivation risk during transcatheter arterial chemoembolization (TACE). To investigate the regulatory effects and mechanisms of 5-FU on HBV replication, an HBV mouse model was established by pAAV-HBV1.2 hydrodynamic injection followed by intraperitoneal 5-FU injection, and different in vitro models (HepG2.2.15 or Huh7 cells) were established. Realtime RT‒qPCR, western blotting, luciferase assays, and immunofluorescence were used to determine viral parameters. We also explored the underlying mechanisms by RNA-seq, oxidative stress evaluation and autophagy assessment. RESULTS: The pooled estimated rate of HBV reactivation in patients receiving TACE was 30.3 % (95 % CI, 23.1%-37.4 %). 5-FU, which is a chemotherapeutic agent commonly used in TACE, promoted HBV replication in vitro and in vivo. Mechanistically, 5-FU treatment obviously increased autophagosome formation, as shown by increased LC3-II levels. Additionally, 5-FU impaired autophagic degradation, as shown by marked p62 and mCherry-GFP-LC3 upregulation, ultimately promoting HBV replication and secretion. Autophagy inhibition by 3-methyladenine or chloroquine significantly altered 5-FU-induced HBV replication. Furthermore, 5-FU-induced autophagy and HBV replication were markedly attenuated with a reactive oxygen species (ROS) scavenger. CONCLUSIONS: Together, our results indicate that ROS-induced autophagosome formation and autophagic degradation play a critical role in 5-FU-induced HBV reactivation.

TRAF2 inhibits senescence in hepatocellular carcinoma cells via regulating the ROMO1/ NAD+/SIRT3/SOD2 axis.

The suppression of tumor proliferation via cellular senescence has emerged as a promising approach for anti-tumor therapy. Tumor necrosis factor receptor-associated factor 2 (TRAF2), an adaptor protein involved in the NF-κB signaling pathway and reactive oxygen species (ROS) production, has been implicated in hepatocellular carcinoma (HCC) proliferation. However, little is currently known about whether TRAF2 promotes HCC development by inhibiting cellular senescence. Replicative senescence model and IR-induced mouse model demonstrated that TRAF2 expression was decrease in senescence cells or liver tissues. Depletion of TRAF2 could inhibit proliferation and arrest the cell cycle via activating p53/p21WAF1 and p16INK4a/pRb signaling pathways in HCC cells and eventually lead to cellular senescence. Mechanistically, TRAF2 deficiency increased the expression of mitochondrial protein reactive oxygen species modulator 1 (ROMO1) and subsequently activated the NAD+/SIRT3/SOD2 pathway to promote the production of ROS and cause mitochondrial dysfunction, which eventually contributed to DNA damage response (DDR). Our findings demonstrate that TRAF2 deficiency inhibits the proliferation of HCC by promoting senescence. Therefore, targeting TRAF2 through various approaches holds therapeutic potential for treating HCC.

Association between whole-grain consumption and carotid atherosclerosis: the Tianjin chronic low-grade systemic inflammation and health (TCLSIH) cohort study.

Background: Whole-grain contains a range of beneficial nutrients, which are thought to play a role in the prevention of chronic diseases. However, the association between whole-grain consumption and the risk of developing carotid atherosclerosis (CA) has not been sufficiently elucidated. We, therefore, conducted this study to investigate the relationship between whole-grain consumption and CA in the general adult population. Methods: This prospective cohort study included a total of 2166 participants (19.2-84.6 years, 55.0% men) without a history of cardiovascular disease, cancer, and CA at baseline. A validated food frequency questionnaire was used to assess whole-grain consumption. Measurements of CA include carotid intima-media thickness (IMT) and carotid plaque. IMT thickening is defined as: IMT ≥ 1.0 mm or a carotid bifurcation IMT ≥ 1.2 mm. Carotid plaque is defined as: distinct area protruding ≥1.5 mm into the vascular lumen of the carotid artery. Cox proportional hazards regression models were used to examine the association of whole-grain consumption with incident CA. Results: A total of 538 (341 men) first incident cases of CA occurred during 5585 person-years of follow-up (median follow-up: 4.2 years). After adjusting for demographic characteristics, lifestyle factors, dietary intake, individual and family history of disease, the multivariable HRs (95% CIs) for incident CA were 1.00 (reference) for <1 time per week, 1.10 (0.85, 1.43) for 1 time per week, 0.95 (0.75, 1.20) for 2-6 times per week, and 1.12 (0.80, 1.56) for ≥1 times per day, respectively (P for trend = 0.99). Similar results were observed in stratified analyses by the main covariates and sensitivity analyses. Conclusion: Our data indicate that whole-grain consumption had no significant association with the risk of CA in an adult Chinese population. In our study population, there is a low consumption of whole-grain, which may limit our ability to see an association. Further cohort studies or randomized controlled trials are needed to confirm our results.

The TRAF2-p62 axis promotes proliferation and survival of liver cancer by activating mTORC1 pathway.

TRAF2 (Tumor necrosis factor receptor-associated factor 2) is a dual function protein, acting as an adaptor protein and a ubiquitin E3 ligase, which plays an essential role in mediating the TNFα-NFκB signal pathway. Dysregulated expression of TRAF2 has been reported in a variety of human cancers. Whether and how TRAF2 regulates the growth of liver cancer cells remains elusive. The goal of this study is to investigate potential dysregulation of TRAF2 and its biological function in liver cancer, and to elucidate the underlying mechanism, leading to validation of TRAF2 as an attractive liver cancer target. Here, we reported TRAF2 is up-regulated in human liver cancer cell lines and tissues, and high TRAF2 expression is associated with a poor prognosis of HCC patients. Proteomics profiling along with Co-immunoprecipitation analysis revealed that p62 is a new substrate of TRAF2, which is subjected to TRAF2-induced polyubiquitination via the K63 linkage at the K420 residue. A strong negative correlation was found between the protein levels of p62 and TRAF2 in human HCC samples. TRAF2 depletion inhibited growth and survival of liver cancer cells both in vitro and in vivo by causing p62 accumulation, which is partially rescued by simultaneous p62 knockdown. Mechanistically, TRAF2-mediated p62 polyubiquitylation activates the mTORC1 by forming the p62-mTORC1-Rag complex, which facilitates the lysosome localization of mTORC1. TRAF2 depletion inhibited mTORC1 activity through the disruption of interaction between p62 and the mTORC1 complex. In conclusion, our study provides the proof-of-concept evidence that TRAF2 is a valid target for liver cancer.

Inhibition of BTK improved APAP-induced liver injury via suppressing proinflammatory macrophages activation by restoring mitochondrion function.

BACKGROUND: Acetaminophen (APAP) overdose can cause severe liver injury and APAP-induced liver injury (AILI) is one of the leading causes of acute liver failure (ALF). Bruton's tyrosine kinase (BTK) is a key tyrosine kinase in immune responses, which plays an important role in many inflammatory diseases. However, its effect on AILI is still not clear. Here, we aimed to assess the effect of BTK on AILI and explore its underlying mechanism. METHODS: In our study, western blot and immunohistochemistry were used to detect the expression of BTK in AILI. The C57BL/6 mice were used to check the protective effect of BTK inhibition on AILI and the activation of BTK was confirmed in mice macrophages treated with APAP. Immunofluorescence, immunohistochemistry, oxygen consumption rate (OCR) detection, polymerase chain reaction (PCR), flow cytometry and western blot were used to determine the role of BTK in mitochondrial dynamics and function of macrophages and the underlying mechanisms in AILI. RESULTS: Our results showed that BTK upregulated in AILI. BTK inhibition protected mice from AILI and BTK was activated in mice macrophages in response to APAP. Mechanically, BTK inhibition promoted mitochondrial fusion and restored mitochondrial function through phospholipase C gamma 2 (PLCγ2)-reactive oxygen species (ROS)-Optic Atrophy 1(OPA1) pathway in macrophages and finally suppressed the release of proinflammatory cytokines. CONCLUSIONS: In conclusion, we found that BTK inhibition protected mice from AILI by restoring the mitochondrial function of macrophages through the improvement of the mitochondrial dynamic imbalance via PLCγ2-ROS-OPA1 signaling pathway, which indicated that BTK might be a potential therapeutic target of AILI.

Trends in Incidence and Prognostic Factors of Two Subtypes of Primary Liver Cancers: A Surveillance, Epidemiology, and End Results-Based Population Study.

OBJECTIVES: The objective of our study was to investigate and compare the epidemiologic characteristics, prognostic factors, and survival between hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC) patients. METHODS: Age-adjusted incidence rates were evaluated from 1975 to 2016 using the Surveillance, Epidemiology, and End Results (SEER) database. Overall survival (OS) was investigated using the Kaplan-Meier method and log-rank test. Univariate and multivariate Cox regression analyses were performed to identify the independent prognostic factors for OS. RESULTS: In the last 10 years, the incidence rate of ICC increased rapidly by 109% (annual percentage change (APC) = 8.24, 95% CI = 6.64 to 9.86; P < .001), compared with a much more modest 12% increase in the incidence of HCC (APC = 1.59, 95% CI = .56 to 2.62; P < .001). This trend persisted throughout the study across different age groups, sexes, and races. Males older than 70 years and of other races (non-African American and non-Caucasian) showed the highest incidence rates of HCC and ICC. Multivariate Cox regression analysis demonstrated that other race, married status, later year of diagnosis, more examined lymph nodes, and surgery were significant protective factors of OS in HCC patients. In contrast, the race and year of diagnosis were not independent prognostic factors, but radiation and chemotherapy were protective factors of OS in ICC patients. The median OS was 18 months and 12 months in HCC and ICC patients, respectively. CONCLUSION: In the last 10 years, the incidence of HCC had a slow growth in the United States, whereas ICC showed a remarkable increase. The 5-year OS of the former has improved in recent years while that of the latter showed no significant improvement. Therefore, surgery could contribute to superior survival outcomes as compared to other treatments.

Neuroendocrine Carcinoma as an Independent Prognostic Factor for Patients With Prostate Cancer: A Population-Based Study.

Background: Neuroendocrine carcinoma (NEC) is a rare and highly malignant variation of prostate adenocarcinoma. We aimed to investigate the prognostic value of NEC in prostate cancer. Methods: A total of 530440 patients of prostate cancer, including neuroendocrine prostate cancer (NEPC) and adenocarcinoma from 2004 to 2018 were obtained from the national Surveillance, Epidemiology, and End Results (SEER) database. Propensity score matching (PSM), multivariable Cox proportional hazard model, Kaplan-Meier method and subgroup analysis were performed in our study. Results: NEPC patients were inclined to be older at diagnosis (Median age, 69(61-77) vs. 65(59-72), P< 0.001) and had higher rates of muscle invasive disease (30.9% vs. 9.2%, P < 0.001), lymph node metastasis (32.2% vs. 2.2%, P < 0.001), and distal metastasis (45.7% vs. 3.6%, P < 0.001) compared with prostate adenocarcinoma patients. However, the proportion of NEPC patients with PSA levels higher than 4.0 ng/mL was significantly less than adenocarcinoma patients (47.3% vs. 72.9%, P<0.001). NEPC patients had a lower rate of receiving surgery treatment (28.8% vs. 43.9%, P<0.001), but they had an obviously higher rate of receiving chemotherapy (57.9% vs. 1.0%, P<0.001). A Cox regression analysis demonstrated that the NEPC patients faced a remarkably worse OS (HR = 2.78, 95% CI = 2.34-3.31, P < 0.001) and CSS (HR = 3.07, 95% CI = 2.55-3.71, P < 0.001) compared with adenocarcinoma patients after PSM. Subgroup analyses further suggested that NEPC patients obtained significantly poorer prognosis across nearly all subgroups. Conclusion: The prognosis of NEPC was worse than that of adenocarcinoma among patients with prostate cancer. The histological subtype of NEC is an independent prognostic factor for patients with prostate cancer.

Neddylation inhibitor MLN4924 has anti-HBV activity via modulating the ERK-HNF1α-C/EBPα-HNF4α axis.

Hepatitis B virus (HBV) infection is a major public health problem. The high levels of HBV DNA and HBsAg are positively associated with the development of secondary liver diseases, including hepatocellular carcinoma (HCC). Current treatment with nucleos(t)ide analogues mainly reduces viral DNA, but has minimal, if any, inhibitory effect on the viral antigen. Although IFN reduces both HBV DNA and HBsAg, the serious associated side effects limit its use in clinic. Thus, there is an urgent demanding for novel anti-HBV therapy. In our study, viral parameters were determined in the supernatant of HepG2.2.15 cells, HBV-expressing Huh7 and HepG2 cells which transfected with HBV plasmids and in the serum of HBV mouse models with hydrodynamic injection of pAAV-HBV1.2 plasmid. RT-qPCR and Southern blot were performed to detect 35kb mRNA and cccDNA. RT-qPCR, Luciferase assay and Western blot were used to determine anti-HBV effects of MLN4924 and the underlying mechanisms. We found that treatment with MLN4924, the first-in-class neddylation inhibitor currently in several phase II clinical trials for anti-cancer application, effectively suppressed production of HBV DNA, HBsAg, 3.5kb HBV RNA as well as cccDNA. Mechanistically, MLN4924 blocks cullin neddylation and activates ERK to suppress the expression of several transcription factors required for HBV replication, including HNF1α, C/EBPα and HNF4α, leading to an effective blockage in the production of cccDNA and HBV antigen. Our study revealed that neddylation inhibitor MLN4924 has impressive anti-HBV activity by inhibiting HBV replication, thus providing sound rationale for future MLN4924 clinical trial as a novel anti-HBV therapy.

Neddylation: A Versatile Pathway Takes on Chronic Liver Diseases.

Neddylation is a ubiquitin-like posttranslational modification that conjugates neural precursor cell expressed developmentally downregulated-8 (Nedd8) to specific substrates for regulation of protein activity. In light of current researches, the neddylation pathway is aberrant in the pathogenesis of many diseases. In our review, we summarize the versatile roles of neddylation in chronic liver diseases (CLDs). CLDs are one of the leading causes of chronic disease-associated deaths worldwide. There are diverse etiologic agents causing CLDs, mainly including hepatitis B virus (HBV) infection, nonalcoholic fatty liver disease (NAFLD), chronic exposure to alcohol or drugs, and autoimmune causes. So far, however, there remains a paucity of effective therapeutic approach to CLDs. In this review, we summarized the role of the neddylation pathway which runs through the chronic hepatitis B/NAFLD-liver fibrosis-cirrhosis-hepatocellular carcinoma (HCC) axis, a canonical pattern in the process of CLD development and progression. The dysregulation of neddylation may provide a better understanding of CLD pathology and even a novel therapeutic strategy. Correspondingly, inhibiting neddylation via MLN4924, a small molecule compound targeting NEDD8-activating enzyme (NAE), can potently alleviate CLD progression and improve the outcome. On this basis, profiling and characterization of the neddylation pathway can provide new insights into the CLD pathology as well as novel therapeutic strategies, independently of the etiology of CLD.

DHX15 Inhibits Autophagy and the Proliferation of Hepatoma Cells.

Autophagy is a highly conserved process by which superfluous or harmful components in eukaryotic cells are degraded by autophagosomes. This cytoprotective mechanism is strongly related to various human diseases, such as cancer, autoimmune diseases, and diabetes. DEAH-box helicase 15 (DHX15), a member of the DEAH box family, is mainly involved in RNA splicing and ribosome maturation. Recently, DHX15 was identified as a tumor-related factor. Although both autophagy and DHX15 are involved in cellular metabolism and cancer progression, their exact relationship and mechanism remain elusive. In this study, we discovered a non-classic function of DHX15 and identified DHX15 as a suppressive protein in autophagy for the first time. We further found that mTORC1 is involved in DHX15-mediated regulation of autophagy and that DHX15 inhibits proliferation of hepatocellular carcinoma (HCC) cells by suppressing autophagy. In conclusion, our study demonstrates a non-classical function of DHX15 as a negative regulator of autophagy related to the mTORC1 pathway and reveals that DHX15-related autophagy dysfunction promotes HCC cell proliferation, indicating that DHX15 may be a target for liver cancer treatment.

Exosomal miRNAs in hepatocellular carcinoma development and clinical responses.

Hepatocellular carcinoma remains the sixth most lethal malignancy in the world. While HCC is often diagnosed via current biomarkers at a late stage, early detection of HCC has proven to be very difficult. Recent studies have focused on using exosomal miRNAs in clinical diagnostics and therapeutics, because they have improved stability in exosomes than as free miRNAs themselves. Exosomal miRNAs act through novel mechanisms for inducing cellular responses in a variety of biological circumstances. Dysregulated expression of miRNAs in exosomes can also accelerate HCC progression, including cell proliferation and metastasis, via alteration of a network of genes. Growing evidence demonstrates that exosomal miRNAs can affect many aspects of physiological and pathological conditions in HCC and indicates that miRNAs in exosomes can not only serve as sensitive biomarkers for cancer diagnostics and recurrence but can also potentially be used as therapeutics to target HCC progression. In this review, we summarize the latest findings between exosomal miRNAs and HCC, in order to better comprehend the functions and applications in HCC. Moreover, we discuss critical issues to consider when developing anti-tumor exosomal miRNAs as a novel therapeutic strategy for treating HCC in the clinic.

[Study of Cine-MRI for the soft palate in patients with obstructive sleep apnea hypopnea syndrome].

OBJECTIVE: To study dynamic change and pathophysiology of airway obstruction of the soft palate in patients with obstructive sleep apnea hypopnea syndrome (OSAHS) during wakefulness and natural sleep. METHOD: Sixteen patients who were diagnosed as OSAHS by sleep questionnaires, medical examination and polysomnography were enrolled in this study in Shanghai Tenth People' Hospital from May to December during 2007. All patients were requested to keep awake prior to examination. Sequential midline sagittal images of the upper airway were obtained during awake and asleep state with Cine-MRI and been transmitted to portable computer. Morphologic change of the soft palate, the anterior-posterior pendulum angle of the soft palate, the anteroposterior diameter and the length of soft palate were measured. Statistical analysis was performed with paired t-test. RESULT: During wakefulness: soft palate caused obstruction by floating backwards and widening anteroposterior diameter(distance between hard palate and uvula P > 0.05, included angle of hard palate and segmental vente of uvula P < 0.05, included angle of hard palate and segmental dorsum of uvula P < 0.01, difference of included angle P < 0.01). Main obstruction site was on retropalatal region. During natural sleep: soft palate caused obstruction by lengthening down and widening anteroposterior diameter (distance between hard palate and uvula P < 0.01), included angle of hard palate and segmental vente of uvula P > 0.05, included angle of hard palate and segmental dorsum of uvula P > 0.05, difference of included angle P < 0.01). Main obstruction site was on retroglottal region. CONCLUSION: Morphologic change of soft palate in patients with OSAHS is multiple, and level of obstruction is deeper during natural sleep than during wakefulness. Main reason of airway obstruction is distinct during different state. The obstruction of upper airway of patients with OSAHS during wakefulness can't replace that during natural sleep.

Optimization of b value in diffusion-weighted MRI for the differential diagnosis of benign and malignant vertebral fractures.

OBJECTIVE: The objective was to explore the optimal b value in diffusion-weighted imaging (DWI) of MRI for differential diagnosis of benign and malignant vertebral fractures. MATERIALS AND METHODS: Thirty-four consecutive patients with vertebral compression fractures underwent sagittal diffusion-weighted imaging (DWI) with different b values. The group included 14 patients with 18 benign vertebral fractures due to osteoporosis and/or trauma and 20 patients with 27 malignant vertebral fractures due to malignancy. The quality of the images was analyzed qualitatively on a three-point scale and quantitatively by measurement of the signal-to-noise ratio (SNR). Apparent diffusion coefficient (ADC) values were also calculated. RESULTS: Smaller b values correlated with better DW image quality. We found significant differences in the qualitative points values among the DW images with different b values (F=302.18, p<0.001). The mean SNR of the images ranged from 21.75+/-3.64 at a b value of 0 s/mm2 to 5.31+/-3.17 at a b value of 800 s/mm2. The SNR of DWI with a b value of 300 s/mm2 (18.62+/-2.47) was significantly different from that with other b values (p<0.01). The mean combined ADC values of malignant fractures were significantly lower than those of benign ones on DWI with a b value of 300 s/mm2 (t=9.097, p<0.01). Four cases of benign vertebral fractures were misdiagnosed as being malignant when b values of 0 s/mm2 and 100 s/mm2 were used. CONCLUSIONS: When DWI with multiple b values is used to differentiate benign from malignant vertebral compression fractures, b values within the range of around 300 s/mm2 are recommended, taking into account both SNR and diffusion weighting of water molecules.