Bo Qin

Suppression of USP18 Potentiates the Anti-HBV Activity of Interferon Alpha in HepG2.2.15 Cells via JAK/STAT Signaling

Ubiquitin-specific protease 18 (USP18, also known as UBP43) has both interferon stimulated gene 15 (ISG15) dependent and ISG15-independent functions. By silencing the expression of USP18 in HepG2.2.15 cells, we studied the effect of USP18 on the anti-HBV activity of IFN-α and demonstrated that knockdown of USP18 significantly Inhibited the HBV expression and increased the expression of ISGs. Levels of hepatitis B virus surface antigen (HBsAg), hepatitis B virus e antigen (HBeAg), HBV DNA and intracellular hepatitis B virus core antigen (HBcAg) were dramatically decreased with or without treatment of indicated dose of IFN-α. Suppression of USP18 activated the JAK/STAT signaling pathway as shown by the increased and prolonged expression of phosphorylated signal transducer and activator of transcription 1 (p-STAT1) in combination with enhanced expression of several interferon stimulated genes (ISGs). Our results indicated that USP18 modulates the anti-HBV activity of IFN-α via activation of the JAK/STAT signaling pathway in Hepg2.2.15 cells.

ORF3 of Hepatitis E Virus Inhibits the Expression of Proinflammatory Cytokines and Chemotactic Factors in LPS-Stimulated Human PMA-THP1 Cells by Inhibiting NF-κB Pathway

Hepatitis E virus (HEV) is one of the primary causative agents of acute hepatitis. It is noteworthy that HEV can develop chronic infection and even lead to liver cirrhosis; however, the mechanism has not been revealed. In this study, the ELISA assay was used to detect protein levels, and we found that HEV open reading frame 3 (ORF3) protein inhibited the expression of proinflammatory cytokines (tumor necrosis factor-alpha [TNF-α], interleukin [IL]-1β, IL-6, IL-8, IL-12p40, and IL-18) and chemotactic factors (nitric oxide [NO], interferon-inducible protein-10 (IP-10), macrophage inflammatory protein (MIP)-1α, monocyte chemoattractant protein-1 (MCP-1), granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF)] in lipopolysaccharide (LPS)-stimulated human PMA-THP1 cells. Further study showed that mRNA and protein levels of pattern recognition receptors (PRRs), such as Toll-like receptor 4 (TLR4), TNF receptor-associated factor 6 (TRAF6), and nucleotide-binding oligomerization domain containing 2 (NOD2), decreased after infection of pLL3.7-ORF3 (pORF3); moreover, the inhibition produced corresponding upregulation of IκBα and downregulation of phosphorylated IκB kinase IKKɛ (p-IKKɛ) and phosphorylated nuclear factor (NF)-κB (p-NF-κB), but little variation was found in the concentration of IKKɛ and NF-κB. Taken together, our results demonstrated that HEV ORF3 attenuated LPS-induced cytokine production and chemotactic factors, predominantly by inhibiting various PRRs-mediated NF-κB signaling pathways. The anti-inflammatory properties might be of great importance to clarify the role and mechanism of macrophages in chronic HEV infection and cirrhosis.

Comparison of Three Intervention Models for Promoting Circumcision among Migrant Workers in Western China to Reduce Local Sexual Transmission of HIV

Objective Three models for promoting male circumcision (MC) as a preventative intervention against HIV infection were compared among migrant worker populations in western China. Methods A cohort study was performed after an initial cross-sectional survey among migrant workers in three provincial level districts with high HIV prevalence in western China. A total of 1,670 HIV seronegative male migrants were cluster-randomized into three intervention models, in which the dissemination of promotional materials and expert- and volunteer-led discussions are conducted in one, two, and three stage interventions. Changes in knowledge of MC, acceptability of MC, MC surgery uptake, and the costs of implementation were analyzed at 6-month and 9-month follow-up visits. Results All three models significantly increased the participants’ knowledge about MC. The three-stage model significantly increased the acceptability of MC among participants and led to greatest increase in MC uptake. At the end of follow-up, 9.2% (153/1,670) of participants underwent MC surgery; uptake among the one-, two-, and three-stage models were 4.9%, 9.3%, and 14.6%, respectively. Multivariable Cox regression analysis showed that three-stage model was the most effective method to scale up MC, with RR = 2.0 (95% CI, 1.3-3.1, P=0.002) compared to the on-site session model. The two-stage intervention model showed no significant difference with either the on-site session model (RR=1.5, 95% CI, 0.92-2.4, P=0.12) or three-stage model (P=0.10). Conclusions A three-stage intervention with gradual introduction of knowledge led to the significantly increase in MC uptake among migrant workers in western China, and was also the most cost-effective method among the three models.

Development of Self-Management Indicators for Chronic Hepatitis B Patients on Antiviral Therapy: Results of a Chinese Delphi Panel Survey

Objective This study aimed to develop a set of indicators that could be used to measure and monitor the self-management performance for chronic hepatitis B (CHB) patients on antiviral therapy in China. Methods A two-round Delphi study via e-mail correspondence was conducted, with a group of 30 Chinese experts. The Delphi questionnaire consisted of 53 indicators identified from a literature review. Experts rated and scored the importance of indicators on a five-point Likert scale. Consensus was considered to be reached if a median score in the top tertile (4-5) and ≥80% of panel ratings in the top tertile (4-5) after Round 2. The included indicators were validated with a group of 106 CHB patients. Results The response rates for the first and second rounds were 90.9% (n=30) and 86.7% (n=26), respectively. Three new indicators were suggested in the first round. 55 indicators were included in the second round after modified. 45 (81.8%) indicators achieved on the level of consensus, all of which had an inter-quartile range of 1 or below. The final set included 4 domains and 45 indicators which were well accepted and understandable by CHB patients. Conclusion This Delphi study produced a set of 45 self-management indicators for CHB patients on antiviral therapy in China. These indicators could be used to measure and monitor the patients’ self-management performance, with the goal of improving the quality of life in this population.

Factors associated with acceptability of circumcision among male drug users in western China: a cross-sectional study

This study is aimed to investigate whether male circumcision (MC) is feasible among IDUs in China. 1304 drug users who attended methadone maintenance therapy clinics in Guangxi, Chongqing and Xinjiang were selected for participation by using convenience sampling, and completed a self-administered questionnaire. The factors associated with the acceptability of MC were examined via multiple logistic regression models. 45.2% (589/1304) of the participants reported an acceptance of MC. Many of the participants who were initially not willing to accept MC (715/1304) had changed their mind when they were informed that MC would reduce the risk of HIV and STDs (43.4%; 310/715), that MC is associated with few surgery-related complications (23.1%; 165/715), that the surgical procedure could be arranged free of charge (20.1%, 144/715). In the multivariate analysis, higher acceptability of MC was associated with knowledge of the hazards of phimosis (OR=2.22), the presence of phimosis (OR=14.87), and knowledge that MC can prevent AIDS and STDs (OR=1.49); while lower acceptability was associated with residing in Chongqing province (OR=0.41) and an educational level of junior (OR=0.64) and senior high (OR=0.63) school. The MC policy targeting IDUs in China should take into account these factors associated with MC acceptability.

Screening and identification of key biomarkers in hepatocellular carcinoma: Evidence from bioinformatic analysis

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Intense efforts have been made to elucidate the pathogeny, but the molecular mechanisms of HCC are still not well understood. To identify the candidate genes in the carcinogenesis and progression of HCC, microarray datasets GSE19665, GSE33006 and GSE41804 were downloaded from Gene Expression Omnibus (GEO) database. The differentially expressed genes (DEGs) were identified, and function enrichment analyses were performed. The protein-protein interaction network (PPI) was constructed and the module analysis was performed using STRING and Cytoscape. A total of 273 DEGs were identified, consisting of 189 downregulated genes and 84 upregulated genes. The enriched functions and pathways of the DEGs include protein activation cascade, complement activation, carbohydrate binding, complement and coagulation cascades, mitotic cell cycle and oocyte meiosis. Sixteen hub genes were identified and biological process analysis revealed that these genes were mainly enriched in cell division, cell cycle and nuclear division. Survival analysis showed that BUB1, CDC20, KIF20A, RACGAP1 and CEP55 may be involved in the carcinogenesis, invasion or recurrence of HCC. In conclusion, DEGs and hub genes identified in the present study help us understand the molecular mechanisms underlying the carcinogenesis and progression of HCC, and provide candidate targets for diagnosis and treatment of HCC.

Impact of Educational Interventions on Acceptance and Uptake of Male Circumcision in the General Population of Western China: A Multicenter Cohort Study

To compare different intervention models for promoting male circumcision (MC) to prevent HIV transmission in Western China. A total of 1690 male participants from multiple study sites were cluster randomly allocated to three-stage (Model A), two-stage (Model B), and one-stage (Model C) educational interventions. In all three interventions models, knowledge about MC significantly increased and the reported willingness to accept MC increased to 52.6% (255/485), 67.0% (353/527), and 45.5% (219/481) after intervention, respectively (P < 0.05). Rate of MC surgery uptake was highest (23.7%; 115/485) among those who received Model A intervention, compared to those who received Model B (17.1%; 90/527) or Model C (9.4%; 45/481) interventions (P < 0.05). Multivariable Cox regression analysis identified that Model A or Model B had twice the effect of Model C on MC uptake, with relative risks of 2.4 (95%CI, 1.5–3.8) and 2.2 (95%CI, 1.3–3.6), respectively. Model B was the most effective model for improving participants’ willingness to accept MC, while Model A was most successful at increasing uptake of MC surgery. Self-reported attitude towards MC uptake was not strongly correlated with actual behavior in this study focusing on the general male population in Western China.

Acceptability of Male Circumcision among College Students in Medical Universities in Western China: A Cross-Sectional Study

Background Male circumcision (MC) has been shown to reduce the risk of female to male transmission of HIV. The goal of this survey was to explore MC’s acceptability and the factors associated with MC among college students in medical universities in western China. Methods A cross-sectional study was carried out in three provinces in western China (Guangxi, Chongqing and Xinjiang) to assess the acceptability of MC as well as to discover factors associated with the acceptability among college students in medical universities. A total of 1,790 uncircumcised male students from three medical universities were enrolled in this study. In addition, 150 students who had undergone MC were also enrolled in the survey, and they participated in in-depth interviews. Results Of all the uncircumcised participants (n = 1,790), 55.2% (n = 988) were willing to accept MC. Among those who accepted MC, 67.3% thought that MC could improve their sexual partners’ hygiene, 46.3% believed that HIV and sexually transmitted diseases (STDs) could be partially prevented by MC. The multivariable logistic regression indicates that MC’s acceptability was associated with three factors: the redundant foreskin (OR = 10.171, 95% CI = 7.629–13.559), knowing the hazard of having a redundant foreskin (OR = 1.597, 95% CI = 1.097–2.323), and enhancing sexual pleasure (OR = 1.628, 95% CI = 1.312–2.021). The in-depth interviews for subjects who had undergone MC showed that the major reason for having MC was the redundant foreskin (87.3%), followed by the benefits and the fewer complications of having MC done. In addition, most of these participants (65.3%) said that the MC could enhance sexual satisfaction. Conclusions MC’s acceptance among college students in medical universities is higher than it is among other populations in western China. An implementation of an MC programme among this population is feasible in the future.

HEV ORF3 downregulates TLR7 to inhibit the generation of type I interferon via impairment of multiple signaling pathways

Hepatitis E is the most common type of acute hepatitis prevalent worldwide. The open reading frame 3 protein of HEV (HEV ORF3) is proposed to create a favorable environment for viral replication and pathogenesis. However, the mechanisms by which HEV overcomes the effects of host immunity, particularly the role of ORF3, remain to be established. Expression of IFNα and IFNβ in supernatant and cell samples was examined via ELISA and quantitative RT-PCR. The protein levels of specific signaling factors in cells overexpressing HEV ORF3 were examined via western blot. Analyses of cells transfected with vectors expressing ORF3 demonstrated that HEV ORF3 significantly impairs the generation of endogenous type I interferon through downregulating TLR3 and TLR7 as well as their corresponding downstream signaling pathways. Moreover, inhibition of NFκB, JAK/STAT and JNK/MAPK signaling pathways contributed significantly to suppression of increased levels of TLR7. Levels of p-P65, p-STAT1 and p-JNK were markedly impaired in ORF3-expressing cells, even upon treatment with the respective agonists. HEV ORF3 inhibits the production of endogenous type I interferon through downregulation of TLR3 and TLR7. Furthermore, suppression of TLR7 is achieved through impairment of multiple signaling pathways, including NFκB, JAK/STAT and JNK/MAPK.

Gene expression profile after knockdown of USP18 in Hepg2.2.15 cells

In our previous work, we found that the expression of ubiquitin‐specific protease 18 (USP18), also known as UBP43, is associated with the efficiency of interferon alpha (IFN‐α) treatment in patients with chronic hepatitis B (CHB). To elucidate the influence of USP18 on hepatitis B virus (HBV) replication and the mechanism of this activity, we silenced USP18 by introducing short hairpin RNA (shRNA) into Hepg2.2.15 cells. To identify the changed genes and pathways in Hepg2.2.15‐shRNA‐USP18 cells, we performed a microarray gene expression analysis to compare the Hepg2.2.15 stably expressing USP18‐shRNA cells versus control cells using the Affymetrix Human Transcriptome Array (HTA) 2.0 microarrays. Microarray analysis indicated that genes involved in regulation of thyroid hormone signaling pathway, complement, and coagulation cascades, PERK‐mediated unfolded protein response, and insulin‐like growth factor‐activated receptor activity were significantly altered after USP18 knockdown for 72 h. Furthermore, genes involved in hepatocyte proliferation, liver fibrosis, such as cell cycle regulatory gene CCND1, were also altered after USP18 knockdown in Hepg2.2.15 cells. In conclusion, USP18 is critical for regulating the replication of HBV in Hepg2.2.15 cells, which suggest that USP18 may be a candidate target for HBV treatment.