Effects of hepatitis C virus core protein and nonstructural protein 4B on the Wnt/ß-catenin pathway.

BACKGROUND: Hepatitis C virus (HCV) core protein and nonstructural protein 4B (NS4B) are potentially oncogenic. Aberrant activation of the Wnt/ß-catenin signaling pathway is closely associated with hepatocarcinogenesis. We investigated the effects of HCV type 1b core protein and NS4B on Wnt/ß-catenin signaling in various liver cells, and explored the molecular mechanism underlying HCV-related hepatocarcinogenesis. RESULTS: Compared with the empty vector control, HCV core protein and NS4B demonstrated the following characteristics in the Huh7 cells: significantly enhanced ß-catenin/Tcf-dependent transcriptional activity (F = 40.87, P < 0.01); increased nuclear translocation of ß-catenin (F = 165.26, P < 0.01); upregulated nuclear ß-catenin, cytoplasmic ß-catenin, Wnt1, c-myc, and cyclin D1 protein expression (P < 0.01); and promoted proliferation of Huh7 cells (P < 0.01 or P < 0.05). Neither protein enhanced ß-catenin/Tcf-dependent transcriptional activity in the LO2 cells (F = 0.65, P > 0.05), but they did significantly enhance Wnt3a-induced ß-catenin/Tcf-dependent transcriptional activity (F = 64.25, P < 0.01), and promoted the nuclear translocation of ß-catenin (F = 66.54, P < 0.01) and the Wnt3a-induced proliferation of LO2 cells (P < 0.01 or P < 0.05). Moreover, activation of the Wnt/ß-catenin signaling pathway was greater with the core protein than with NS4B (P < 0.01 or P < 0.05). CONCLUSIONS: HCV core protein and NS4B directly activate the Wnt/ß-catenin signaling pathway in Huh7 cells and LO2 cells induced by Wnt3a. These data suggest that HCV core protein and NS4B contribute to HCV-associated hepatocellular carcinogenesis.

[Effect of NF-κB on the pathogenic course of non-alcoholic fatty liver disease].

With the economic growth and better standards of living, the prevalence of non-alcoholic fatty liver disease (NAFLD) is expected to increase dramatically worldwide. NAFLD is a common chronic inflammation disease. NF-κB is a transcription factor that plays crucial roles in inflammation, immunity, cell proliferation and apoptosis. It can facilitate the occurrence and development of NAFLD, and the underlying mechanisms are related to insulin resistance, oxidative stress, alteration of intestinal flora, activation of renin angiotensin system, etc.

[Screening of differentially expressed genes and gene pathways in hepatitis C virus 1b type nonstructural protein 4B stably expressed L02 cell line].

OBJECTIVE: To screen differentially expressed genes and gene pathways in L02 cell line stably expressing hepatitis C virus (HCV) Ib type nonstructural protein 4B (NS4B) mediated by lentiviral system, and to provide a basis for further research of molecular biological mechanism of NS4B gene in chronic hepatitis C and hepatocarcinogenesis. METHODS: NS4B stably overexpressed L02 cell line and negative control stable L02 cell line, designated as L02-NS4B and L02-mkate2 respectively, were resurrected and amplified in vitro. The differentially expressed genes between L02-NS4B and L02-mkate2 were determined by gene expression microarray from Human Gene 1.0ST. The significant pathways of the differential genes were selected by the Fisher's exact test and χ2 test according to kyoto encyclopedia of genes and genomes (KEGG) database. The differential expression levels of 5 selected genes including protein kinase C delta binding protein (PRKCDBP), tumor protein p53 (TP53), v-akt murine thymoma viral oncogene homolog 1 (AKT1), baculoviral IAP repeat containing 3 (BIRC3) and B-cell lymphoma 2-like1 (BCL2L1) from cDNA microarray data were further verified by real-time quantitative polymerase chain reaction (real-time QPCR). RESULTS: Between L02-NS4B and L02-mkate2, the genes with flurescence intensity ratio >1.2 or <0.8 were considered as differentially expressed genes. A total of 2 682 differentially expressed genes in the known 28 869 human genes were detected in L02-NS4B, 1 446 genes were upregulated and 1 236 genes were downregulated. A total of 41 involved pathways of up-regulated differential genes were identified by KEGG database, mainly including apoptosis, extracellular matrix receptor interaction, cell cycle, pathways in cancer and Toll-like receptor signaling pathway; and 20 involved pathways of down-regulated differential genes were identified, mainly including pathways in cancer, Wnt signaling pathway and cell cycle pathway. Of the 5 upregulated genes selected from cDNA microarray data, 3 genes showed the same differential expression pattern by real-time QPCR as that shown in cDNA microarray data, namely AKT1, BIRC3 and BCL2L1. The confirmation rate of real-time QPCR was 60%. CONCLUSION: HCVNS4B can up-regulate or down-regulate the expression of many genes in L02 cells, thus affecting multiple signaling pathways relevant to cell apoptosis, cell cycle and carcinogenesis.

A comprehensive review of the anticancer effects of decursin.

Cancer is a globally complex disease with a plethora of genetic, physiological, metabolic, and environmental variations. With the increasing resistance to current anticancer drugs, efforts have been made to develop effective cancer treatments. Currently, natural products are considered promising cancer therapeutic agents due to their potent anticancer activity and low intrinsic toxicity. Decursin, a coumarin analog mainly derived from the roots of the medicinal plant Angelica sinensis, has a wide range of biological activities, including anti-inflammatory, antioxidant, neuroprotective, and especially anticancer activities. Existing studies indicate that decursin affects cell proliferation, apoptosis, autophagy, angiogenesis, and metastasis. It also indirectly affects the immune microenvironment and can act as a potential anticancer agent. Decursin can exert synergistic antitumor effects when used in combination with a number of common clinical anticancer drugs, enhancing chemotherapy sensitivity and reversing drug resistance in cancer cells, suggesting that decursin is a good drug combination. Second, decursin is also a promising lead compound, and compounds modifying its structure and formulation form also have good anticancer effects. In addition, decursin is not only a key ingredient in several natural herbs and dietary supplements but is also available through a biosynthetic pathway, with anticancer properties and a high degree of safety in cells, animals, and humans. Thus, it is evident that decursin is a promising natural compound, and its great potential for cancer prevention and treatment needs to be studied and explored in greater depth to support its move from the laboratory to the clinic.

A new perspective on structural characterisation and immunomodulatory activity of arabinogalactan in Larix kaempferi from Qinling Mountains.

In this study, crude polysaccharide (LAG-C) and homogeneous arabinogalactan (LAG-W) were isolated from Qinling Larix kaempferi of Shaanxi Province. Bioactivity assays showed that LAG-W and LAG-C enhanced the phagocytic ability, NO secretion, acid phosphatase activity, and cytokine production (IL-6, IL-1ß, and TNF-α) of RAW264.7 macrophages. Notably, LAG-W exhibited a significantly stronger immunomodulatory effect than LAG-C. The primary structure of LAG-W was characterised by chemical methods (monosaccharide composition, methylation analysis, and alkali treatment) and spectroscopic techniques (gas chromatography-mass spectrometry, high-performance liquid chromatography-mass spectrometry, and 1D/2D nuclear magnetic resonance). LAG-W was identified as a 22.08 kilodaltons (kDa) neutral polysaccharide composed of arabinose and galactose at a 1:7.5 molar ratio. Its backbone consisted of repeated →3)-ß-Galp-(1→ residues. Side chains, connected at the O-6 position, were mainly composed of T-ß-Galp-(1→ and T-ß-Galp-(1→6)-ß-Galp-(1→ residues. And it also contained small amounts of T-ß-Arap-(1→, T-α-Araf-(1→6)-ß-Galp-(1→6)-ß-Galp-(1→, and T-α-Araf-(1→3)-α-Araf-(1→6)-ß-Galp-(1→ residues. By structurally and functionally characterising L. kaempferi polysaccharides, this study opens the way for the valorisation of this species.

Quasi-Hyperbolic Framework Graphite Foam-Based Composites with High Thermal Conductivity and Electromagnetic Shielding Properties Fabricated by an Electrochemical Expansion Method.

Nowadays, the rapid development of electronic devices requires composites with high thermal conductivity and good electromagnetic shielding properties. The key challenge lies in the construction of high-performance conductive networks. Herein, an electrochemical expansion graphite foam (EEG) with a quasi-hyperbolic framework was prepared by an electrochemical expansion method, and then the epoxy resin (EP) was filled to fabricate the composites. The graphite plate was first electrochemically intercalated and then foamed, in which plasticization was caused by weak oxidation in intercalation and the quasi-hyperbolic framework was induced by foaming during expansion. These processes were characterized by Fourier transform infrared (FTIR), micro-Raman, X-ray photoelectron spectroscopy (XPS), and so on. Based on the highly efficient quasi-hyperbolic framework and high-quality graphite structure, the thermal conductivity of the composite reached 43.523 W/(m·K), and total electromagnetic interference (EMI) shielding (SET) reached 105 dB. The heat transfer behavior was simulated by finite element analysis (FEA) in detail. This method of preparing high thermal conductivity and electromagnetic shielding materials has a good application prospect.

A nanoparticle-based sonodynamic therapy reduces Helicobacter pylori infection in mouse without disrupting gut microbiota.

Infection by Helicobacter pylori, a prevalent global pathogen, currently requires antibiotic-based treatments, which often lead to antimicrobial resistance and gut microbiota dysbiosis. Here, we develop a non-antibiotic approach using sonodynamic therapy mediated by a lecithin bilayer-coated poly(lactic-co-glycolic) nanoparticle preloaded with verteporfin, Ver-PLGA@Lecithin, in conjunction with localized ultrasound exposure of a dosage permissible for ultrasound medical devices. This study reveals dual functionality of Ver-PLGA@Lecithin. It effectively neutralizes vacuolating cytotoxin A, a key virulence factor secreted by H. pylori, even in the absence of ultrasound. When coupled with ultrasound exposure, it inactivates H. pylori by generating reactive oxygen species, offering a potential solution to overcome antimicrobial resistance. In female mouse models bearing H. pylori infection, this sonodynamic therapy performs comparably to the standard triple therapy in reducing gastric infection. Significantly, unlike the antibiotic treatments, the sonodynamic therapy does not negatively disrupt gut microbiota, with the only major impact being upregulation of Lactobacillus, which is a bacterium widely used in yogurt products and probiotics. This study presents a promising alternative to the current antibiotic-based therapies for H. pylori infection, offering a reduced risk of antimicrobial resistance and minimal disturbance to the gut microbiota.

Online LC-ESI-MS/MS comparative analysis of N/O-glycopatterns in human colostrum from different ethnic groups in Northwest China.

Human milk oligosaccharides, including free oligosaccharides and glycoconjugates, exert a key role in neonatal health and development. Changes in free oligosaccharides of milk from different ethnic groups have been documented. In this study, human milk was collected from Han, Hui, and Tibetan populations in northwest China, and differences in N/O-glycome among these three ethnic groups were systematically compared using online high-performance liquid chromatography-tandem mass spectrometry. Among the 63 detected N-glycans, 35 showed significant differences between the three ethnic groups (p < 0.05). Among the 70 detected O-glycans, four neutral O-glycans and six acidic O-glycans exhibited significant differences among the three ethnic groups (p < 0.05), with six acidic O-glycans reported for the first time. Overall, the extent of milk N/O-glycosylation was higher in the Han population than in the Hui or Tibetan groups. This trend was particularly pronounced for the main sialylated N/O-glycans. Except for sulfated O-glycans, which were higher in the milk from Tibetan mothers, the other types of N/O-glycans were present in similar proportions across all ethnic groups. Understanding the composition of N/O-glycans in human milk can help research on the structure-function relationship of glycans.

Impact of structurally diverse polysaccharides on colonic mucin O-glycosylation and gut microbiota.

Understanding how dietary polysaccharides affect mucin O-glycosylation and gut microbiota could provide various nutrition-based treatments. Here, the O-glycan profile of the colonic mucosa and gut microbiome were investigated in C57BL/6J mice fed six structurally diverse dietary polysaccharides and a mixture of six fibers. Dietary polysaccharides increased total O-glycans, mainly by stimulating neutral glycans. Highly branched arabinogalactan promoted terminally fucosylated core 1 O-glycans; whereas linear polysaccharides, including pectin, konjac glucomannan, inulin, and the fiber mixture, favored terminally di-fucosylated O-glycans. The last three polysaccharides also lowered the level of sulfated O-glycans and sialylated mono-fucosylated O-glycans. Varied monosaccharide composition in mixed polysaccharides had a synergistic beneficial effect, boosting fucosylated neutral glycans, decreasing acidic glycans, and stimulating microbial richness and diversity. Dietary polysaccharides containing arabinose and sulfate groups enhanced the relative abundances of Akkermansia and Muribaculaceae, respectively. The present comparison reveals the relationship between dietary polysaccharide structure, mucin O-glycan composition, and intestinal microorganisms.

A gas-solid reaction growth of dense TiO2 nanowire arrays on Ti foils at ambient atmosphere.

In this paper, a facile method is demonstrated to directly fabricate dense titania nanowire arrays on titanium foils under the atmosphere without extra moist conditions. The influences of temperature, time, different catalysts, and concentrations of the respective catalysts on the growth of titania nanowires are discussed in detail. The morphology, composition and crystal structure of the titania nanostructures are revealed by scanning electron microscopy (SEM), powder-X-ray diffraction (XRD) and transmission electron microscopy (TEM) techniques, by which a gas-solid reaction mechanism is suggested to explain the growth process of TiO2 nanowires on Ti substrate.

Randomized controlled study of plasma exchange combined with molecular adsorbent re-circulating system for the treatment of liver failure complicated with hepatic encephalopathy.

BACKGROUND/AIMS: To evaluate the use of plasma exchange (PE) combined with the molecular adsorbent re-circulating system (MARS) for the treatment of liver failure complicated with hepatic encephalopathy. METHODOLOGY: A prospective randomized controlled study was conducted to compare the therapeutic effect of MARS treatment (MARS group, n=60) with that of PE combined with MARS treatment (PE+MARS group, n=60) in patients with liver failure complicated with hepatic encephalopathy. RESULTS: The serum total bilirubin and blood ammonia levels were significantly decreased compared with pretreatment levels after 3 days of both the MARS treatment (p=0.0001, p<0.001) and PE+MARS treatment (both p<0.0001) and the Glasgow coma scale score was significantly increased (both p<0.0001). The 30-day mortality rate was 10.0% (6/60) in the MARS group and 11.7% (7/60) in the PE + MARS group. The per capita cost of treatment was significantly lower in the PE + MARS group than in the MARS group (p=0.0003). CONCLUSIONS: Both MARS and PE + MARS therapy can safely and effectively be used to treat liver failure complicated with hepatic encephalopathy, but PE + MARS therapy reduces serum total bilirubin level more effectively and is more cost-effective.

A facile chemical approach for preparing a SERS active silver substrate.

This communication describes a new surface-enhanced Raman scattering (SERS) active silver substrate prepared by iodination of the evaporated silver foil. After iodination, the morphology of the silver substrate undergoes a self-evolution process in which it displays accordingly the UV-vis absorption shift as well as the AFM topological test. Rhodamine 6G (R6G) is used as the probe molecule to evaluate the enhancement efficiency of the silver substrate at different self-evolution time intervals. The SERS intensity of R6G increases up to ∼29-fold and reaches a maximum after the substrate evolved for 24 h. This method is feasible for the production of an efficient SERS silver substrate.

Clinical characteristics and therapeutic analysis of invasive fungal infection in chronic severe hepatitis patients.

OBJECTIVE: To investigate clinical features and antifungal therapeutic effect of chronic severe hepatitis (CSH) patients with invasive fungal infection (IFI), and to improve the diagnosis and treatment. METHODS: Clinical manifestation, blood routine, imageology and mycetology characteristic, antifungal treatment perscription and therapeutic effect of 79 CSH patients with IFI were retrospectively analyzed. Antifungal therapeutic effect was compared between fluconazole and voriconazole. RESULTS: Thirteen (16.5%) patients received glucocorticoid or other immunodepressants for a relatively long time, 40 (50.6%) patients had invasive operation, and 61 (77.2 %) patients were administered 1-6 kinds of broad-spectrum antibiotics. Seventy-three patients had fever. Leucocytes and neutrophilic granulocyte increased in 96.2% of the patients. Lung (31.6%), intestinal tract (26.2%) and oral cavity (14%) infections were common. Fungus was found in 70.9% of the patients. Candida albicans (40.9%) and aspergillus (21.1%) were often seen. Halo signs and crescent signs on lung CT were relatively specific in 40% of the patients with fungal pneumonia. Voriconazole was more effective than fluconazole(71.4% vs. 39.0%, P<0.05). Twelve patients with lung aspergillus infection were administered voriconazole, 8 (66.7%) patients of whom was effective, and the other 4 (33.3%) patients died. CONCLUSION: There are high risk factors in major CSH patients with IFI. The most common clinical manifestations of CSH patients with IFI are fever, leukocytosis, lung and intestinal tract infection. Candida albicans and aspergillus infection are common. Voriconazole is more effective than fluconazole, and can increase the survival rate of CSH patients with IFI.

Molecular Mechanisms of the Action of Myricetin in Cancer.

Natural compounds, such as paclitaxel and camptothecin, have great effects on the treatment of tumors. Such natural chemicals often achieve anti-tumor effects through a variety of mechanisms. Therefore, it is of great significance to conduct further studies on the anticancer mechanism of natural anticancer agents to lay a solid foundation for the development of new drugs. Myricetin, originally isolated from Myrica nagi, is a natural pigment of flavonoids that can inhibit the growth of cancer cells (such as liver cancer, rectal cancer, skin cancer and lung cancer, etc.). It can regulate many intracellular activities (such as anti-inflammatory and blood lipids regulation) and can even be bacteriostatic. The purpose of this paper is to outline the molecular pathways of the anticancer effects of myricetin, including the effect on cancer cell death, proliferation, angiogenesis, metastasis and cell signaling pathway.

A bulky bis-pocket manganese(V)-oxo corrole complex: observation of oxygen atom transfer between triply bonded Mn(V)[triple bond]O and alkene.

The highly bulky bis-pocket corrole 5,10,15-tris(2,4,6-triphenylphenyl)corrole (H(3)TTPPC) has been synthesized. Resonance Raman spectroscopy revealed a triply bonded Mn[triple bond]O moiety in its manganese(V)-oxo complex. Direct oxygen atom transfer from (TTPPC)Mn[triple bond]O to styrene was confirmed by an (18)O-labeling experiment. The (TTPPC)Mn(III) complex also exhibits significant shape selectivity in the catalytic epoxidation of nonconjugated dienes.

Impact of main vessel calcification on procedural and clinical outcomes of bifurcation lesion undergoing provisional single-stenting intervention: a multicenter, prospective, observational study.

BACKGROUND: Few data on the combined effects of bifurcation and calcification on coronary artery disease (CAD) patients undergoing percutaneous coronary intervention (PCI) are available. This study evaluated the impact of main vessel (MV) calcification on the procedural and long-term outcomes in patients with CAD who underwent provisional single stent PCI. METHODS: This is a multicenter, prospective, observational study. Patients with bifurcation lesions were enrolled at 10 PCI centers in China from January 2015 to December 2017. Intravascular ultrasound or optical coherence tomography was performed in all patients to evaluate the MV calcification. Patients were treated with provisional single stent strategy using drug eluting stents and followed-up at 1 month, 6 months and 12 months after discharge by telephone contact or outpatient visit. Repeated coronary imaging was performed within one year. We compared the procedural success rates in MV and in side branch (SB), and target lesion failure (TLF), defined as a composite of cardiac death, non-fatal myocardial infarction, definite or possible stent thrombosis and target lesion revascularization between patients with and without MV calcification. RESULTS: A total of 185 subjects were enrolled according to the inclusion and exclusion criteria of this study. MV calcification was detected in 119 (64.3%, calcification group) and not found in 66 (35.7%, non-calcification group) patients. The angiographic success rate of MV was 95.8% in the calcification group and 97.0% in the non-calcification group (P = 0.91); the angiographic success rate of SB was 32.8% in the calcification group and 53.0% in the non-calcification group (P < 0.05). During the one-year follow-up period, TLF occurred in 14 (11.8%) patients in the calcification group and in 13 (19.7%) in the non-calcification group (P = 0.31). Multivariate regression analysis showed the same result (HR = 1.23, 95% CI: 0.76-1.52, P = 0.47). Calcification on group had higher recurrent angina than non-calcification group (13.51% vs. 17.65%, P < 0.05). CONCLUSIONS: In patients with coronary bifurcation lesion treated with provisional one stent approach, calcification of MV is associated with lower SB procedural success rate, it could increase recurrence of angina; however, it was not associated with an increased risk of TLF.

Mycophenolate mofetil prevents lethal acute liver failure in mice induced by bacille Calmette-Guérin and lipopolysaccharide.

BACKGROUND/AIMS: To investigate the effect of mycophenolate mofetil (MMF) on acute liver injury induced by bacille Calmette-Guérin (BCG) and lipopolysaccharide (LPS). METHODS: Acute liver failure was induced in male Kunming strain mice by injecting the animals with BCG 2.5 mg per mouse, and LPS 10 microg per mouse 10 days later. The mice in the treatment groups were given MMF 2 h before, simultaneous with, or 2 h after administration of LPS, and the mice in the control group were given the same dose of saline. The 24-h survival rate, serum alanine aminotransferase (ALT), and aspartate aminotransferase (AST) levels were compared. Serum levels of tumor necrosis factor alpha (TNF-alpha), interferon gamma (IFN-gamma), and interleukin 6 (IL-6) were measured and the expressions of TNF-alpha, IFN-gamma, and IL-6 mRNA in the liver tissue were determined by reverse transcription-polymerase chain reaction (RT-PCR). Concanavalin A (Con A)-induced splenocyte proliferation were determined by methods of methyl thiazolyl tetrazolium. RESULTS: Injecting a small dose of LPS into BCG-primed mice caused a lethal hepatic injury mimicking acute hepatitis, from which 16 of the 20 mice died within 24 h (20% survival rate). Massive necrosis of parenchymal hepatocytes with marked inflammatory cell infiltration was observed by histological examination. In parallel, serum ALT and TNF-alpha, IFN-gamma, and IL-6 levels were increased. Expression of TNF-alpha, IFN-gamma, and IL-6 mRNA in the liver were significantly increased also. Treatment with MMF markedly reduced the death rate in a dose-dependent manner. It reached its maximal effect at the dosage of 150 mg per kg of body weight when pretreated 2 h before LPS injection, with improvement of histological feather and survival rate (84.2%, 16/19). MMF significantly inhibited serum levels of TNF-alpha, IFN-gamma, and IL-6, and significantly reduced TNF-alpha, IFN-gamma, and IL-6 expression in the liver, which increased after BCG and LPS injection. Moreover, splenocyte proliferation response induced by Con A was also inhibited by MMF treatment. CONCLUSIONS: Treatment with MMF has a protective effect on endotoxin-induced fatal liver failure by regulating the production of inflammatory cytokines and T-cell proliferation.

Low-temperature synthesis of single crystalline Ag2S nanowires on silver substrates.

We report on the successful synthesis of silver sulfide (Ag(2)S) nanowires by a simple and mild gas-solid reaction approach. For the nanowire synthesis, a preoxidized silver substrate is exposed to an atmosphere of an O(2)/H(2)S mixture at room temperature or slightly above. The resulting Ag(2)S nanowires are phase pure with a monoclinic crystal structure and have diameters of a few tens of nanometers and lengths up to 100 mum. The influence of reaction conditions on the diameter, length, and morphology of the Ag(2)S nanowires has been studied by a number of structural and spectroscopic techniques. The nanowire growth mechanism on the Ag substrate has been discussed, which is likely characterized by continuous deposition at the tip. Additionally, we demonstrate thinning and cutting of individual Ag(2)S nanowires with electron beams and laser beams, which are potentially useful for nanowire manipulation and engineering.

[Role of platelet activating factor in hepatorenal syndrome].

OBJECTIVE: To investigate the relationship between platelet activating factor (PAF) and occurrence of hepatorenal syndrome (HRS). METHODS: A total of 57 cases of patients with liver cirrhosis or hepatitis gravis were enrolled. The serum concentrations of PAF, lipopolysaccharide (LPS) and prothrombin activity (PTA), total bilirubin, and serum creatinine were detected in patients combined with HRS and in patients without HRS in hospital at the term. The correlations among the PAF, LPS, and the function of liver and kidney were analyzed. RESULTS: The concentrations of PAF and LPS in the patients combined with HRS were significantly higher than those patients without HRS. There were linear correlations between the PAF and LPS level, the total bilirubin level and PAF level, the PAF level and serum creatinine level. CONCLUSION: The PAF associates with the occurrence of HRS. The PAF may mediate the occurrence of HRS induced by LPS. The PAF level may forecast whether patients with liver failure will take place HRS.

Inhibition of expression of hepatitis C virus 1b genotype core and NS4B genes in HepG2 cells using artificial microRNAs.

The present study aimed to evaluate the silencing effect of artificial microRNAs (amiRNAs) against the hepatitis C virus (HCV) 1b (HCV1b) genotype core (C) and non-structural protein 4B (NS4B) genes. pDsRed-monomer-Core and pDsRed-monomer-NS4B plasmids, containing the target genes were constructed. A total of eight artificial micro RNA (amiRNA)-expressing plasmids, namely, pmiRE-C-mi1 to -mi4 and pmiRE-NS4B-mi1 to -mi4, were designed and constructed to interfere with various sites of the core and NS4B genes, and the amiRNA interfering plasmid and the corresponding target gene-expressing plasmid were co-transfected into HepG2 cells. At 48 h after transfection, HCV core and NS4B gene expression levels were detected using fluorescence microscopy, flow cytometry, reverse transcription quantitative polymerase chain reaction and western blot analysis. Fluorescence microscopy revealed that the target gene-transfected cells expressed red fluorescent protein, whereas the interfering plasmid-transfected cells exhibited expression of green fluorescent protein. The percentage of red fluorescent proteins and mean fluorescence intensity, as well as protein expression levels of the core and NS4B genes within the cells, which were co-transfected by the amiRNA interfering plasmid and the target gene, were significantly decreased. The results of the present study confirmed that amiRNAs may effectively and specifically inhibit the expression of HCV1b core and NS4B genes in HepG2 cells, potentially providing a novel therapeutic strategy for the treatment of HCV.