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.

Chelidonine reduces IL-1β-induced inflammation and matrix catabolism in chondrocytes and attenuates cartilage degeneration and synovial inflammation in rats

Chondrocyte inflammation and catabolism are two major features in the progression of osteoarthritis (OA). Chelidonine, a principal alkaloid extracted from Chelidonium majus, is suggested to show anti-inflammation, anti-apoptosis, and anti-oxidation activities in various diseases. However, its potential effects on OA cartilage degeneration remains unclear. To evaluate the effect of chelidonine on OA and its underlying mechanism, we incubated chondrocytes with interleukin (IL)-1β and chelidonine at varying concentrations. Then, we performed the CCK-8 assay, fluorescence immunostaining, reverse transcription PCR, ELISA, and western blotting to evaluate cell viability, catabolic/inflammatory factors, levels of extracellular matrix (ECM)-related proteins, and the involved pathways. H&E and Safranin-O staining and ELISA were performed to measure cartilage degradation and synovial inflammation. Chelidonine suppressed the IL-1β-mediated catabolism and inflammation of chondrocytes. Chelidonine suppressed the NF-κB pathway activation. Similarly, our in vivo experiment showed that chelidonine partially attenuated cartilage degradation while inhibiting synovial inflammation. Chelidonine inhibited inflammation and catabolism through modulation of NF-κB pathways in vitro, thereby avoiding rat cartilage degeneration and synovial inflammation within OA.

[Molecular mechanism of Ganoderma against gastric cancer based on network pharmacology and experimental test].

This study aims to explore the molecular mechanism of Ganoderma against gastric cancer based on network pharmacology, molecular docking, and cell experiment. The active components and targets of Ganoderma were retrieved from Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP), and gastric cancer-related targets from GeneCards and Online Mendelian Inheritance in Man(OMIM). The protein-protein interaction(PPI) network of the common targets was constructed with STRING, followed by Gene Ontology(GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analysis of the common genes based on Bioconductor and R language. The medicinal-disease-component-target network and medicinal-disease-component-target-pathway network were established by Cytoscape. Molecular docking was performed between ß-sitosterol(the key component in Ganoderma) and the top 15 targets in the PPI network. Cell experiment was performed to verify the findings. A total of 14 active components and 28 targets of Ganoderma were retrieved, and the medicinal and the disease shared 25 targets, including caspase-3(CASP3), caspase-8(CASP8), caspase-9(CASP9), and B-cell lymphoma-2(BCL2). The common targets involved 72 signaling pathways and apoptosis and p53 signaling pathway may play a crucial role in the effect of Ganoderma against gastric cancer. ß-sitosterol had strong binding activity to the top 15 targets in the PPI network. The in vitro cell experiment demonstrated that ß-sitosterol inhibited gastric cancer AGS cell proliferation by inducing cell apoptosis and cell cycle arrest in the S phase, which might be related to the regulation of the p53 pathway. This study shows the multi-component, multi-target, and multi-pathway characteristics of Ganoderma against gastric cancer, which lays a scientific basis for further research on the molecular mechanism.

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.

The Crosstalk Between Regulatory Non-Coding RNAs and Nuclear Factor Kappa B in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a highly lethal type of malignancies that possesses great loss of life safety to human beings worldwide. However, few effective means of curing HCC exist and its specific molecular basis is still far from being fully elucidated. Activation of nuclear factor kappa B (NF-κB), which is often observed in HCC, is considered to play a significant part in hepatocarcinogenesis and development. The emergence of regulatory non-coding RNAs (ncRNAs), particularly microRNAs (miRNAs) and long non-coding RNAs (lncRNAs), is a defining advance in cancer biology, and related research in this branch has yielded many diagnostic and therapeutic opportunities. Recent studies have suggested that regulatory ncRNAs act as inhibitors or activators in the initiation and progression of HCC by targeting components of NF-κB signaling or regulating NF-κB activity. In this review, we attach importance to the role and function of regulatory ncRNAs in NF-κB signaling of HCC and NF-κB-associated chemoresistance in HCC, then propose future research directions and challenges of regulatory ncRNAs mediated-regulation of NF-κB pathway in HCC.

MiR-144-3p-mediated dysregulation of EIF4G2 contributes to the development of hepatocellular carcinoma through the ERK pathway.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common cancers with high incidence and mortality. However, the underlying mechanisms of HCC still remain unclear. Eukaryotic translation initiation factors (eIFs) have a substantial effect on tumor development. In this study, we were aimed to investigate the role of eukaryotic translation initiation factor 4 gamma 2 (EIF4G2) in HCC. METHODS: Western blot (WB) of 30 paired HCC tissues and tissue microarrays (TMAs) conducted by immunohistochemistry (IHC) in 89 paired HCC samples were performed to assess EIF4G2 expression. Clone formation, real-time cell analysis (RTCA), wound healing and transwell assays were adopted to evaluate the role of EIF4G2 on HCC cell proliferation, migration and invasion abilities. The function of EIF4G2 in HCC tumor growth was assessed in a xenograft nude mouse model in vivo. The regulation of EIF4G2 by miR-144-3p was performed by luciferase reporter assay and WB. RESULTS: The EIF4G2 protein was clearly upregulated in HCC tissues, and high EIF4G2 expression was closely related to HCC prognosis. EIF4G2 silencing could inhibit HCC cell growth and metastasis in vitro, and suppress tumorigenesis in vivo by repressing the ERK signaling pathway. The results of luciferase reporter assays, WB and IHC staining verified that EIF4G2 was negatively regulated by miR-144. And re-expression of EIF4G2 could partially reverse the inhibiting effect of miR-144 in HCC. CONCLUSION: In summary, our study revealed the role of EIF4G2 in HCC development via the activation of the ERK pathway. We also found that EIF4G2 could be negatively regulated by the tumor suppressor miR-144. Our investigations indicated that EIF4G2 might be a promising therapeutic target in HCC.

Separation of epigallocatechin gallate and epicatechin gallate from tea polyphenols by macroporous resin and crystallization.

Epigallocatechin gallate (EGCG) and epicatechin gallate (ECG) are the most abundant ester catechins of green tea polyphenols (GTPs) with numerous potential bioactivities, which have wide application prospects in the fields of medicine and functional foods. In this study, a new method using macroporous resin and crystallization was established to separate and purify EGCG and ECG. Two resins with high adsorption and desorption capacities for EGCG and ECG were screened through static adsorption/desorption tests, and the LX-20B resin was selected through column chromatography due to its best separation effect. Moreover, the column separation parameters of LX-20B resin (sample amount, ethanol elution concentration, elution volume, and elution flow rate) were optimized. After resin purification, the EGCG and ECG purity were 70.08 ± 2.55% and 74.97 ± 2.66%, respectively, and the recovery rates were 68.07 ± 2.43% and 74.28 ± 2.24%, respectively. After crystallization, the EGCG purity reached 95.87 ± 0.89%, with a total recovery rate of 58.66%, and the ECG purity reached 95.55 ± 1.30%, with a total recovery rate of 62.45%. The separation efficiency of the resin showed no significant change after 6 cycles. These results show the proposed method to be a simple, eco-friendly, and cost-effective separation method for the industrial separation and purification of EGCG and ECG from GTPs.

Screening and identifying of α-amylase inhibitors from medicine food homology plants: Insights from computational analysis and experimental studies.

There is a growing interest in screening α-amylase inhibitors from natural products for application in the development of new antidiabetic drugs or functional foods. In this study, a structure-based virtual screening was applied to rapidly identify the α-amylase inhibitors from medicine food homology (MFH) plants. Similarity search, docking & scoring were used for further filter small molecules. As a result, 21 corresponding potential α-amylase inhibitors from MFH plants were obtained. And, six polyphenol compounds (curcumin, procyanidins, epicatechin gallate (ECG), epigallocatechin gallate (EGCG), hesperidin, and puerarin) were highlighted for further verification after a thorough assessment of the classification of hit molecules as well as docking scores. The results of the enzyme inhibition test showed that ECG, EGCG, and procyanidins had the better binding ability of α-amylase among these six polyphenols. The Ki values of ECG, EGCG, and procyanidins on α-amylase were 0.70, 1.68, and 0.24, respectively. The CD spectra results indicated that the three polyphenols can cause conformational changes in α-amylase. PRACTICAL APPLICATIONS: A structure-based virtual screening method for rapid identifying α-amylase inhibitors from MFH plants was developed successfully in this study. These findings suggested that natural polyphenols such as ECG, EGCG, and procyanidins may be a potential inhibitor of α-amylase which could be used as a nutrient supplement for the prevention of diabetes mellitus or can be further used in the development of hypoglycemic drugs. At the same time, it can provide theoretical guidance for the better utilization and development of medicine food homology plants containing these potential α-amylase inhibitors. Moreover, this work may provide ideas and references for the screening of other target protein inhibitors.

Extracellular vesicles participate in macrophage-involved immune responses under liver diseases.

The liver serves as a central participant in immune system owing to its particular blood supply and large amounts of immune cells, in which macrophages play a significant role in liver homeostasis and disorders. Extracellular vesicles (EVs), membrane-defined nanometer-sized vesicles released by cells in a tightly controlled manner, have attracted intensive research attention as a critical vehicle for cell-cell communication in the pathophysiology of liver. Accumulating evidence has proved that extracellular vesicles are frequently involved in macrophage-mediated biological behaviors. Not only can macrophages produce and secrete EVs containing multifarious cargo themselves to exert immunomodulatory functions, but also macrophages may serve as target cells of EVs from other cells eliciting the alteration of their phenotype and function. Since both macrophage as well as EVs show pleiotropic and central effects in the progression of liver diseases, their roles in adjusting innate immunity of liver often present a crossover. In this review we are dedicated to deciphering the complex immunological network constituted by macrophages and EVs in several common liver diseases, including acute liver injury or failure and a set of chronic liver diseases such as viral hepatitis B and C, metabolic and alcoholic liver diseases, as well as hepatocellular carcinoma (HCC). From the aspect of immunology, we integrate the mechanism of EVs and hepatic macrophages in the setting of liver diseases and show a promising significance of utilizing this association into clinical immunotherapy.

Impact of Nrf2 on tumour growth and drug sensitivity in oncogenic K-ras-transformed cells in vitro and in vivo.

K-ras is one of the most common oncogenes in human cancers, and its aberrant activation may lead to malignant transformation associated with oxidative stress and activation of the transcription factor Nrf2 that regulates multiple detoxification enzymes. The purpose of this research was to use gene editing technology to evaluate the role of Nrf2 in affecting tumour growth and drug sensitivity of K-rasG12V-transformed cells. We showed that induction of K-rasG12V caused a significant activation of Nrf2 associated with increased expression of its target genes NAD(P)H:quinone oxidoreductase 1 (NQO1) and haem oxygenase-1 (HO-1). Interestingly, knock-out of Nrf2 by CRISPR/Cas9 in K-rasG12V-expressing cells only impacted the expression of NQO1 but not HO-1. We also found that Nrf2 knock-out caused high reactive oxygen species (ROS) stress, suppression of cell proliferation, increased apoptosis in vitro, and a decrease of tumour growth in vivo. Furthermore, abrogation of Nrf2 significantly increased the sensitivity of K-rasG12V cells to multiple anticancer agents including phenethyl isothiocyanate (PEITC), doxorubicin, etoposide, and cisplatin. These results show that genetic abrogation of Nrf2 impairs the malignant phenotype of K-RasG12V-transformed cells in vitro and in vivo, and demonstrates the critical role of Nrf2 in promoting cell survival and drug resistance in cells harbouring oncogenic K-ras. As such, inhibition of Nrf2 would be an attractive strategy to increase the therapeutic effect and overcome drug resistance in cancer with oncogenic K-ras activation.

Synthesis and biological evaluation of 20-epi-amino-20-deoxysalinomycin derivatives.

To improve the druggability of salinomycin, a 20-epi-amino-20-deoxysalinomycin derivatives library was synthesized with high efficacy from which a few salinomycin derivatives with high potency and selectivity were identified through comprehensive cytotoxicity assay, including a fluorine-19 magnetic resonance sensitive tool molecule. Using a K-ras cellular model, salinomycin and its derivatives showed different molecular mode of action from literature reports. These results would be valuable for developing salinomycin-based cancer therapy.

Regulation of stem-like cancer cells by glutamine through ß-catenin pathway mediated by redox signaling.

BACKGROUND: Cancer stem cells (CSCs) are thought to play an important role in tumor recurrence and drug resistance, and present a major challenge in cancer therapy. The tumor microenvironment such as growth factors, nutrients and oxygen affect CSC generation and proliferation by providing the necessary energy sources and growth signals. The side population (SP) analysis has been used to detect the stem-like cancer cell populations based on their high expression of ABCG2 that exports Hoechst-33342 and certain cytotoxic drugs from the cells. The purpose of this research is to investigate the effect of a main nutrient molecule, glutamine, on SP cells and the possible underlying mechanism(s). METHODS: Biochemical assays and flow cytometric analysis were used to evaluate the effect of glutamine on stem-like side population cells in vitro. Molecular analyses including RNAi interfering, qRT-PCR, and immunoblotting were employed to investigate the molecular signaling in response to glutamine deprivation and its influence on tumor formation capacity in vivo. RESULTS: We show that glutamine supports the maintenance of the stem cell phenotype by promoting glutathione synthesis and thus maintaining redox balance for SP cells. A deprivation of glutamine in the culture medium significantly reduced the proportion of SP cells. L-asparaginase, an enzyme that catalyzes the hydrolysis of asparagine and glutamine to aspartic acid and glutamate, respectively, mimics the effect of glutamine withdrawal and also diminished the proportion of SP cells. Mechanistically, glutamine deprivation increases intracellular ROS levels, leading to down-regulation of the ß-catenin pathway. CONCLUSION: Glutamine plays a significant role in maintaining the stemness of cancer cells by a redox-mediated mechanism mediated by ß-catenin. Inhibition of glutamine metabolism or deprivation of glutamine by L-asparaginase may be a new strategy to eliminate CSCs and overcome drug resistance.