Resectable lung lesions malignancy assessment and cancer detection by ultra-deep sequencing of targeted gene mutations in plasma cell-free DNA.

BACKGROUND: Early detection of lung cancer to allow curative treatment remains challenging. Cell-free circulating tumour (ct) DNA (ctDNA) analysis may aid in malignancy assessment and early cancer diagnosis of lung nodules found in screening imagery. METHODS: The multicentre clinical study enrolled 192 patients with operable occupying lung diseases. Plasma ctDNA, white cell count genomic DNA (gDNA) and tumour tissue gDNA of each patient were analysed by ultra-deep sequencing to an average of 35 000× of the coding regions of 65 lung cancer-related genes. RESULTS: The cohort consists of a quarter of benign lung diseases and three quarters of cancer patients with all histopathology subtypes. 64% of the cancer patients are at stage I. Gene mutations detection in tissue gDNA and plasma ctDNA results in a sensitivity of 91% and specificity of 88%. When ctDNA assay was used as the test, the sensitivity was 69% and specificity 96%. As for the lung cancer patients, the assay detected 63%, 83%, 94% and 100%, for stages I, II, III and IV, respectively. In a linear discriminant analysis, combination of ctDNA, patient age and a panel of serum biomarkers boosted the overall sensitivity to 80% at a specificity of 99%. 29 out of the 65 genes harboured mutations in the patients with lung cancer with the largest number found in TP53 (30% plasma and 62% tumour tissue samples) and EGFR (20% and 40%, respectively). CONCLUSION: Plasma ctDNA was analysed in lung nodule assessment and early cancer detection, while an algorithm combining clinical information enhanced the test performance. TRIAL REGISTRATION NUMBER: NCT03081741.

Spatio-temporal release of NGF and GDNF from multi-layered nanofibrous bicomponent electrospun scaffolds.

Scaffolds capable of providing dual neurotrophic factor (NTF) delivery with different release kinetics, spatial delivery of NTFs at different loci and topographical guidance are promising for enhanced peripheral nerve regeneration. In this study, we have designed and fabricated multi-layered aligned-fiber scaffolds through combining emulsion electrospinning, sequential electrospinning and high-speed electrospinning (HS-ES) to modulate the release behavior of glial cell line-derived growth factor(GDNF) and nerve growth factor (NGF). GDNF and NGF were incorporated into poly(lactic-co-glycolic acid) (PLGA) fibers and poly(D,L-lactic acid) (PDLLA) fibers, respectively. Aligned fibers were obtained in each layer of multi-layered scaffolds and relatively thick tri-layered and tetra-layered scaffolds with controlled layer thickness were obtained. Their morphology, structure, properties, and the in vitro release of growth factors were examined. Dual and spatio-temporal release of GDNF and NGF with different release kinetics from multi-layered scaffolds was successfully demonstrated. High separation efficiency by PDLLA fibrous barrier layer for spatial neurotrophic factor delivery from both tri-layered scaffolds and tetra-layered scaffolds was achieved.

Development and validation of a subcritical 1,1,1,2-tetrafluoroethane extraction technique: Gas chromatography-mass spectrometry method for the simultaneous determination of polycyclic aromatic hydrocarbons, polychlorinated biphenyls, and polybrominated biphenyl ethers in aquatic products.

A simple, rapid, and green method was developed for the simultaneous analysis of polycyclic aromatic hydrocarbons, polychlorinated biphenyls, and polybrominated biphenyl ethers in aquatic products using subcritical 1,1,1,2-tetrafluoroethane extraction coupled with gas chromatography and mass spectrometry. Effects of the extraction temperature, pressure, and cosolvent volume on the extraction efficiency were investigated by extracting spiked oyster samples. The results show that the maximum extraction efficiency was obtained at 40°C, 12 MPa, and a cosolvent (dichloromethane) volume of 5.0 mL. Under these conditions, the calibration curves had good linearity with square of the correlation larger than 0.998 in the concentration range of 5-800 ng/mL; limit of detection and limit of quantitation were 0.16-2.83 and 0.55-9.43 ng/g, respectively. At spiked levels of 10, 30, and 50 ng/g, the average recoveries were 70.4-80.4% for polycyclic aromatic hydrocarbons, 74.0-83.6% for polychlorinated biphenyls, and 66.9-78.0% for polybrominated biphenyl ethers, with average relative standard deviations of less than 16.3%. The established method has no significant differences in recovery compared to traditional methods and is suitable for the analysis of real samples.

Prospect of research on anti-atherosclerosis effect of main components of traditional Chinese medicine Yiqi Huoxue Huatan recipe through gut microbiota: A review.

The incidence and mortality rates of cardiovascular diseases are on the rise globally, posing a severe threat to human health. Atherosclerosis (AS) is considered a multi-factorial inflammatory disease and the main pathological basis of cardiovascular and cerebrovascular diseases, as well as the leading cause of death. Dysbiosis of the gut microbiota can induce and exacerbate inflammatory reactions, accelerate metabolic disorders and immune function decline, and affect the progression and prognosis of AS-related diseases. The Chinese herbal medicine clinicians frequently utilize Yiqi Huoxue Huatan recipe, an effective therapeutic approach for the management of AS. This article reviews the correlation between the main components of Yiqi Huoxue Huatan recipe and the gut microbiota and AS to provide new directions and a theoretical basis for the prevention and treatment of AS.

Improving resistant starch content of cassava starch by pulsed electric field-assisted esterification.

This study aims to investigate the effect of pulsed electric field (PEF) assisted OSA esterification treatment on the multi-scale structure and digestive properties of cassava starch and structure-digestion relationships. The degree of substitution (DS) of starch dually modified at 1.5-4.5 kV/cm was 37.6-55.3 % higher than that of starch modified by the conventional method. Compared with native starch, the resistant starch (RS) content of esterified starch treated with 3 kV/cm significantly increased by 17.13 %, whereas that of starch produced by the conventional method increased by only 5.91 %. Furthermore, assisted esterification at low electric fields (1.5-3 kV/cm) promotes ester carbonyl grafting on the surface of starch granules, increases steric hindrance and promotes the rearrangement of the amorphous regions of starch, which increases the density of the double-helical structure. These structural changes slow down starch digestion and increase the RS content. Therefore, this study presents a potential method for increasing the RS content of starch products using PEF to achieve the desired digestibility.

Mechanism of Yiqi Huoxue Huatan recipe in the treatment of coronary atherosclerotic disease through network pharmacology and experiments.

In recent years, with population aging and economic development, morbidity and mortality of atherosclerotic cardiovascular disease associated with atherosclerosis (AS) have gradually increased. In this study, a combination of network pharmacology and experimental verification was used to systematically explore the action mechanism of Yiqi Huoxue Huatan Recipe (YHHR) in the treatment of coronary atherosclerotic heart disease (CAD). We searched and screened the active ingredients of Coptis chinensis, Astragalus membranaceus, Salvia miltiorrhiza, and Hirudo. We also searched multiple databases for related target genes corresponding to the compounds and CAD. STRING was used to construct the protein-protein interaction (PPI) network of genes. Metascape was used to perform gene ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis for common targets to analyze the main pathways, and finally, the molecular docking and main possible pathways were verified by experimental studies. Firstly, a total of 1480 predicted target points were obtained through the Swiss Target Prediction database. After screening, merging, and deleting duplicate values, a total of 768 targets were obtained. Secondly, "Coronary atherosclerotic heart disease" was searched in databases such as the OMIM, GeneCards, and TTD. 1844 disease-related targets were obtained. Among PPI network diagram of YHHR-CAD, SRC had the highest degree value, followed by AKT1, TP53, hsp90aa1 and mapk3. The KEGG pathway bubble diagram was drawn using Chiplot, the Signal pathways such as NF kappa B signaling pathway, Lipid and AS, and Apelin signaling pathway are closely related to the occurrence of CAD. The PCR and Western blot methods were used to detect the expression of NF-κB p65. When compared with that in the model group, the expression of NF-κB p65mRNA decreased in the low-concentration YHHR group, with P < .05, while the expression of NF-κB p65mRNA decreased significantly in the high-concentration YHHR group, with P < .01. On the other hand, when compared with that in the model group, the expression of NF-κB p65 decreased in the low-concentration YHHR group, but was not statistically significant, while the expression of NF-κB p65 was significant in the high-concentration YHHR group, and has statistical significance with P < .05. YHHR has been shown to resist inflammation and AS through the SRC/NF-κB signaling pathway.

Pulsed electric field-assisted esterification improves the freeze-thaw stability of corn starch gel by changing its molecular structure.

The influence of pulsed electric field (PEF) combined with octenyl succinic anhydride (OSA) on the freeze-thaw stability of corn starch gel was investigated. After five freeze-thaw cycles, the syneresis value of OSA starch treated with PEF-assisted esterification for 15 min was lower by 29.5 %, while that of OSA starch without PEF treatment was lower by 10.17 %, compared to that of native starch. Low-field nuclear magnetic resonance data showed that the introduction of OSA groups greatly increased the water-holding capacity of starch. Results from differential scanning calorimetry (DSC) and X-ray diffraction (XRD) showed that the PEF-assisted esterification markedly hindered the re-formation of the helical structure of starch during freeze-thaw cycles. Moreover, PEF-assisted esterification improved the viscoelastic properties of the starch gel. It is found that the freeze-thaw stability of the PEF-modified starch depends not only on the degree of substitution but also on the starch molecular fine structure. PEF-assisted OSA starch with a high degree of substitution, a low content of amylose, and a high content of short amylopectin chains were found to have high freeze-thaw stability. This study shows that PEF-assisted esterification is a promising technique that should be used for preserving the quality of frozen foods.

Enhanced encapsulation of lutein using soy protein isolate nanoparticles prepared by pulsed electric field and pH shifting treatment.

In this study, soy protein isolate (SPI) was modified by a pulsed electric field (PEF) combined with pH shifting treatment (10 kV/cm, pH 11) to prepare SPI nanoparticles (PSPI11) for efficient loading of lutein. The results showed that when the mass ratio of SPI to lutein was 25:1, the encapsulation efficiency of lutein in PSPI11 increased from 54% to 77%, and the loading capacity increased by 41% compared to the original SPI. The formed SPI-lutein composite nanoparticles (PSPI11-LUTNPs) had smaller, more homogeneous sizes and larger negative charges than SPI7-LUTNPs. The combined treatment favored the unfolding of the SPI structure and could expose its interior hydrophobic groups to bind with lutein. Nanocomplexation with SPIs significantly improved the solubility and stability of lutein, with PSPI11 showing the greatest improvement. As a result, PEF combined with pH shifting pretreatment is an effective method for developing SPI nanoparticles loaded and protected with lutein.

Binding affinity of curcumin to bovine serum albumin enhanced by pulsed electric field pretreatment.

The present study investigated the effect of pulsed electric field (PEF) pretreatment on the interaction between bovine serum albumin (BSA) and curcumin. Fluorescence quenching results showed that proper PEF pretreatment significantly increased the binding affinity of curcumin and BSA, the binding constant increased by 6.77 times under the conditions of 15 kV/cm for 0.51 ms. However, at higher PEF strength (≥25 kV/cm) and longer processing time (≥0.68 ms), the binding affinity was weakened. PEF pretreatment made the protein structure more disordered and induced partial unfolding of BSA, exposing more hydrophobic regions, thereby increasing the binding affinity to curcumin. PEF-treated BSA (PBSA) possessed better encapsulation efficiency (95.19%) and loading capacity (5.25 mg/g) for curcumin, and the storage stability of curcumin were enhanced by the formation of a complex with PBSA. This study provides new insights into the design of BSA-based delivery systems for curcumin and other hydrophobic nutrients.

Succinylated whey protein isolate-chitosan core-shell composite particles as a novel carrier: Self-assembly mechanism and stability studies.

Single protein [whey protein isolate (WPI) or succinylated whey protein isolate (SWPI)] and composite particles of proteins with chitosan (CS) were tested for their ability to encapsulate and protect curcumin (CUR). Combining protein and CS resulted in changes in zeta-potential and surface hydrophobicity, particularly in the SWPI-H (high degree of succinylation, 90 %) and CS composite particle (H-CS). Furthermore, the secondary and tertiary structures were dramatically altered using Fourier transform infrared (FTIR), circular dichroism (CD), and X-ray diffraction (XRD). Scanning electron microscopy (SEM) and atomic force microscope (AFM) analyses revealed that H-CS exhibited a soft core-rigid shell morphology due to electrostatic interactions, hydrophobic interactions, and H-bond interactions. Fluorescence quenching results demonstrated that H-CS had a higher binding constant (K, 1.69 ×104 M-1) and encapsulation effectiveness (EE, 88.3 %) of CUR. Because of increased binding sites and steric hindrance, CUR was stabilized more effectively in H-CS in photostability and thermostability tests,. These results show that SWPI-CS composite particles can be utilized to build a protection system for water-insoluble nutritional supplements.

Protective effect of baicalein on DNA oxidative damage and its binding mechanism with DNA: An in vitro and molecular docking study.

In this work, the protective effect of baicalein on DNA oxidative damage and its possible protection mechanisms were investigated. 2-thiobarbituric acid (TBA) colorimetry and agarose gel electrophoresis study found that baicalein protected the deoxyribose residue and double-stranded backbone of DNA from the damage of hydroxyl radicals. Antioxidant analysis results showed that baicalein has excellent radicals scavenging effects and Fe2+ chelating ability, which might be the mechanism of baicalein protecting DNA. DNA binding studies indicated that baicalein bound to the minor groove of DNA with moderate binding affinity (K = (7.35 ±â€¯0.91) × 103 M-1). Hydrogen bonding and van der Waals forces played a major role in driving the binding process. Molecular docking further confirmed the experimental results. This binding could stabilize DNA double helix structure, thereby protecting DNA from oxidative damage. This study may provide theoretical basis for designing new functional foods of baicalein for DNA damage protection.

Pulsed electric field assisted modification of octenyl succinylated potato starch and its influence on pasting properties.

The physicochemical properties and structural changes of potato starch esterified with octenyl succinic anhydride (OSA) assisted with pulsed electric field (PEF) were investigated. Results showed that PEF treatment during esterification resulted in a significant modification of pasting properties. The pasting temperature at 2-6 kV/cm reduced by 7.6-15.1 °C for PEF-assisted OSA starches but only by 3 °C for OSA modified starch without PEF treatment as compared to that of native starch. PEF-assisted esterification could reduce the reaction time and improve the reaction efficiency over the control by 6.1-39.1 %. A novel schematic model on structure-functionality relationship for PEF-assisted OSA modified starch was proposed. Structural disorganizations of starch induced lower pasting temperature and paste viscosity. The results suggest that PEF could be a potential eco-friendly and cost-effective physical technique to prepare starch products with desired paste behaviors and to broaden its application area especially in papermaking and textile industries.

Enhanced synthesis of succinylated whey protein isolate by pulsed electric field pretreatment.

The objective of this study was to investigate the feasibility of pulse electric field (PEF) as a pretreatment for whey protein isolate (WPI) before its succinylation. The degree of succinylation (DS) of WPI increased from 88.31% for native WPI to 93.45% for PEF-pretreated WPI (PWPI, initial pH 10.0) for the same succinic anhydride (SA) to WPI ratio (1:1). Fourier transform infrared spectroscopy and scanning electron microscopy analysis proved the successful succinylation of WPI. For PWPIs, the surface hydrophobicity, exposed sulphydryl, and total sulphydryl decreased, which indicates the occurrence of changes in protein structures with more hydrophilic groups and better protein dispersion. Moreover, PEF may expose more amino acid residues binding sites that are present inside the protein, which is more suitable for succinylation. Therefore, the PEF pretreatment of proteins can improve their efficient use that is expected to play a critical role in succinylation industry.

Review of the application of pulsed electric fields (PEF) technology for food processing in China.

With the improvement of living standards, growing consumer demand for high-quality and natural foods has led to the development of new mild processes to enhance or replace conventional thermal and chemical methods for food processing. Pulsed electric fields (PEF) is an emerging and promising non-thermal food processing technology, which is ongoing from laboratory and pilot plant level to the industrial level. Chinese researchers have made tremendous advances in the potential applications of PEF for processing a wide range of food commodities over the last few years, which contributes to the current understanding and development of PEF technology. The objective of this paper is to conduct a systematic review on the achievements of PEF technology used for food processing in China and the corresponding processing principles. Research on the applicability of PEF in food processing suggests that PEF can be used alone or in combination with other methods, not only to inactivate microorganisms and extract active constituents, but also to modify biomacromolecules, enhance chemical reactions and accelerate the aging of fermented foods, which are mainly related to permeabilization of biomembranes, occurrence of electrochemical and electrolytic reactions, polarization and realignment of molecules, and reduction of activation energy of chemical reactions induced by PEF treatments. In addition, some of the most important challenges for the successful implementation of large-scale industrial applications of PEF technology in the food industry are discussed. The results bring out the benefits of both researchers and the industry.

Dynamic formation of cellular aggregates of chondrocytes and mesenchymal stem cells in spinner flask.

OBJECTIVES: Cellular aggregates are readily applicable in cell-based therapy. The effects of agitation and inoculation density on the aggregation of cells in spinner flask and the molecular mechanism of aggregation were investigated. MATERIALS AND METHODS: The aggregation kinetics of cells in spinner flask was evaluated with bovine articular chondrocytes (bACs), rabbit bone marrow-derived mesenchymal stem cells (rMSCs) and their mixture. The morphology of cellular aggregates was studied with scanning electron microscopy and gene expression of cell adhesion-related molecules was analysed. RESULTS: It was shown that suspension culture in spinner flask induced the aggregation of bACs and rMSCs. Both cells exhibited increased aggregation rate and aggregate size with decreasing agitation rate and increasing cell inoculation density. Additionally, aggregate size increased with extended culture time. By analysing gene expression of integrin ß1 and cadherin, it was indicated that these molecules were potentially involved in the aggregation process of bACs and rMSCs, respectively. Aggregates composed of both bACs and rMSCs were also prepared, showing rMSCs in the core and bACs in the periphery. CONCLUSIONS: Cellular aggregates were prepared in dynamic suspension culture using spinner flask, the key parameters to the aggregation process were identified, and the molecular mechanism of aggregation was revealed. This would lay a solid foundation for the large-scale production of cellular aggregates for cell-based therapy, such as cartilage regeneration.

Genome-Wide Screening and Functional Analysis Identifies Tumor Suppressor Long Noncoding RNAs Epigenetically Silenced in Hepatocellular Carcinoma.

Long noncoding RNAs (lncRNA) play critical roles in the development of cancer, including hepatocellular carcinoma (HCC). However, the mechanisms underlying their deregulation remain largely unexplored. In this study, we report that two lncRNAs frequently downregulated in HCC function as tumor suppressors and are epigenetically silenced by histone methyltransferase EZH2. lncRNAs TCAM1P-004 and RP11-598D14.1 were inhibited by EZH-mediated trimethylation of H3K27me3 at their promoters. Downregulation of TCAM1P-004 and RP11-598D14.1 was frequently observed in HCC tumors compared with adjacent normal tissues. Both lncRNAs inhibited cell growth, cell survival, and transformation in HCC cells in vitro as well as tumor formation in vivo. Using RNA pull-down and mass spectrometry, we demonstrated that TCAM1P-004 bound IGF2BP1 and HIST1H1C, whereas RP11-598D14.1 bound IGF2BP1 and STAU1. These lncRNA-protein interactions were critical in regulating p53, MAPK, and HIF1α pathways that promoted cell proliferation in HCC. Overexpression of EZH2 was critical in repressing TCAM1P-004 and RP11-598D14.1, and EZH2-TCAM1P-004/RP11-598D14.1-regulated pathways were prevalent in human HCC. Aberrant suppression of TCAM1P-004 and RP11-598D14.1 led to loss of their tumor-suppressive effects by disrupting the interaction with IGF2BP1, HIST1H1C, and STAU1, which in turn promoted HCC development and progression. Collectively, these findings demonstrate the role of TCAMP1P-004 and RP11-598D14.1 in suppressing tumor growth and suggest that EZH2 may serve as a therapeutic target in HCC. SIGNIFICANCE: EZH2-mediated loss of lncRNAs TCAM1P-004 and RP11-598D14.1 hinders the formation of tumor suppressor lncRNA-protein complexes and subsequently promotes HCC growth.

Incorporation and release of dual growth factors for nerve tissue engineering using nanofibrous bicomponent scaffolds.

Electrospun fibrous scaffolds have been extensively used as cell-supporting matrices or delivery vehicles for various biomolecules in tissue engineering. Biodegradable scaffolds with tunable degradation behaviors are favorable for various resorbable tissue replacements. In nerve tissue engineering, delivery of growth factors (GFs) such as nerve growth factor (NGF) and glial cell line-derived neurotrophic factor (GDNF) from scaffolds can be used to promote peripheral nerve repair. In this study, using the established dual-source dual-power electrospinning technique, bicomponent scaffolds incorporated with NGF and GDNF were designed and demonstrated as a strategy to develop scaffolds providing dual GF delivery. NGF and GDNF were encapsulated in poly(D, L-lactic acid) (PDLLA) and poly(lactic-co-glycolic acid) (PLGA) nanofibers, respectively, via emulsion electrospinning. Bicomponent scaffolds with various mass ratios of GDNF/PLGA fibers to NGF/PDLLA fibers were fabricated. Their morphology, structure, properties, and the in vitro degradation were examined. Both types of core-shell structured fibers were evenly distributed in bicomponent scaffolds. Robust scaffolds with varying component ratios were fabricated with average fiber diameter ranging from 307 ± 100 nm to 688 ± 129 nm. The ultimate tensile stress and elastic modulus could be tuned ranging from 0.23 ± 0.07 MPa to 1.41 ± 0.23 MPa, 11.1 ± 3.0 MPa to 75.9 ± 3.3 MPa, respectively. Adjustable degradation was achieved and the weight loss of scaffolds ranged from 9.2% to 44.0% after 42 day degradation test. GDNF and NGF were incorporated with satisfactory encapsulation efficiency and their bioactivity were well preserved. Sustained release of both types of GFs was also achieved.

Inactivation mechanism of Vibrio parahaemolyticus via supercritical carbon dioxide treatment.

The effects of supercritical carbon dioxide (SC-CO2) treatments on Vibrio parahaemolyticus cells were determined using viable plate count method at different treatment times (10 and 40min), pressures (10-25MPa), and temperature (40°C). Using the changes in the physical (absorbance, transmission electron microscope and contents of fatty acids) and chemical indexes (pH value, activity of Na+K+-ATPase, SDS-PAGE) were for further understand the mechanisms of bacterial inactivation under SC-CO2. The result showed that 25MPa treatment for 40min in 40°C could significantly (P<0.05) enhance inactivation of V. parahaemolyticus. The pH value and activity of Na+K+-ATPase of SC-CO2 treated groups significantly (P<0.05) decreased compared with blank group. Damage to the cell membrane and cytoplasmic components can be observed on transmission electron microscope images. Results of SDS-PAGE and UV-absorbing substances also showed that the leakage of proteins and cytoplasmic materials increased with the SC-CO2 treatment time and pressure. Therefore, our results indicate that SC-CO2 can be applied to inactivate V. parahaemolyticus by causing a low pH, as well as severe damage to key substances and structures.

Statistical analysis of mutant allele frequency level of circulating cell-free DNA and blood cells in healthy individuals.

Cell-free DNA (cfDNA) in plasma has emerged as a potential important biomarker in clinical diagnostics, particularly in cancer. However, somatic mutations are also commonly found in healthy individuals, which interfere with the effectiveness for cancer diagnostics. This study examined the background somatic mutations in white blood cells (WBC) and cfDNA in healthy controls based on sequencing data from 821 non-cancer individuals and several cancer samples with the aim of understanding the patterns of mutations detected in cfDNA. We determined the mutation allele frequencies in both WBC and cfDNA using a panel of 50 cancer-associated genes that covers 20 K-nucleotide region and ultra-deep sequencing with average depth >40000-fold. Our results showed that most of the mutations in cfDNA were highly correlated to WBC. We also observed that the NPM1 gene was the most frequently mutated gene in both WBC and cfDNA. Our study highlighted the importance of sequencing both cfDNA and WBC to improve the sensitivity and accuracy for calling cancer-related mutations from circulating tumour DNA, and shedded light on developing a strategy for early cancer diagnosis by cfDNA sequencing.

Hepatitis B virus X protein promotes hepatocellular carcinoma transformation through interleukin-6 activation of microRNA-21 expression.

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide, and chronic hepatitis B virus (HBV) infection is the major risk factor of HCC. The virus encodes HBV X (HBx) protein that plays a critical role in the development of HCC. Studies have revealed numerous HBx-altered genes and signalling pathways that heavily contribute to tumourigenesis of non-tumour hepatocytes. However, the role of HBx in regulating other critical gene regulators such as microRNAs is poorly understood, which impedes the exploration of a complete HBx-associated carcinogenic network. Besides, critical microRNAs that drive the transformation of non-tumour hepatocytes are yet to be identified. Here, we overexpressed C-terminal truncated HBx protein in a non-tumour hepatocyte cell line MIHA, and measured a panel of cancer-associated miRNAs. We observed that oncogenic miR-21 was upregulated upon ectopic expression of this viral protein variant. HBx-miR-21 pathway was prevalent in HCC cells as inhibition of HBx in Hep3B and PLC/PRF/5 cells significantly suppressed miR-21 expression. Subsequently, we showed that the upregulation of miR-21 was mediated by HBx-induced interleukin-6 pathway followed by activation of STAT3 transcriptional factor. The high dependency of miR-21 expression to HBx protein suggested a unique viral oncogenic pathway that could aberrantly affect a network of gene expression. Importantly, miR-21 was essential in the HBx-induced transformation of non-tumour hepatocytes. Inhibition of miR-21 effectively attenuated anchorage-independent colony formation and subcutaneous tumour growth of MIHA cells. Our study suggested that overexpression of miR-21 was critical to promote early carcinogenesis of hepatocytes upon HBV infection.