Hepatitis C virus (HCV) is the main cause of chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. HCV is a single-stranded, positive-sense RNA virus that is classified into the Flaviviridae family, Hepacivirus genus (21). More than 170 million people worldwide are chronically infected with HCV. The 9.6-kb HCV genome encodes a polyprotein of approximately 3,010 amino acids (aa). The polyprotein is cleaved co-and posttranslationally into at least 10 proteins by viral proteases and cellular signalases: the structural proteins core, E1, E2, and p7 and the nonstructural proteins NS2, NS3, NS4A, NS4B, NS5A, and NS5B (21).Persistent HCV infection causes not only intrahepatic diseases but also extrahepatic manifestations, such as type 2 diabetes. Clinical and experimental data suggest that HCV infection is an additional risk factor for the development of diabetes (26,29,30). HCV-related glucose metabolic changes and insulin resistance have significant clinical consequences, such as accelerated fibrogenesis, reduced virological response to alpha interferon (IFN-␣)-based therapy, and increased incidence of hepatocellular carcinoma (29). Therefore, the molecular mechanism of HCV-related diabetes needs to be clarified.We have sought to identify a novel mechanism of HCV-induced diabetes. We previously demonstrated that HCV suppresses hepatocytic glucose uptake through downregulation of cell surface expression of glucose transporter 2 (GLUT2) in a human hepatoma cell line (19). The uptake of glucose into cells is conducted by facilitative glucose carriers, i.e., glucose transporters (GLUTs). GLUTs are integral membrane proteins that contain 12 membrane-spanning helices. To date, a total of 14 isoforms have been identified in the GLUT family (24). GLUT2 is expressed in the liver, pancreatic ␤-cells, hypothalamic glial cells, retina, and enterocytes. Glucose is transported into hepatocytes by GLUT2 (34). We previously reported that GLUT2 expression was reduced in hepatocytes obtained from HCV-infected patients (19). We also demonstrated that GLUT2 mRNA levels were lower in HCV replicon cells and in HCV J6/JFH1-infected cells than in the control cells. GLUT2 promoter activity was suppressed in HCV-replicating cells. However, the molecular mechanism of HCV-induced suppression of GLUT2 gene expression remains to be elucidated.In the present study, we aimed to clarify molecular mechanisms of HCV-induced suppression of GLUT2 gene expression. We analyzed transcriptional regulation of the GLUT2 promoter in HCV replicon cells. We demonstrate that HCV infection downregulates hepatocyte nuclear factor 1␣ (HNF-1␣) expression at both transcriptional and posttranslational levels, resulting in suppression of GLUT2 promoter. We propose that HCV-induced downregulation of HNF-1␣ may play a crucial role in glucose metabolic disorders caused by HCV. MATERIALS AND METHODSCell culture. The human hepatoma cell line Huh-7.5 (4) was kindly provided by Charles M. Rice (The Rockefeller University, New York, NY).

Background Recently, high-intensity intermittent training (HIIT) appears to have the same beneficial effects or even superior to those of continuous moderate-intensity training (CMIT) on body fat mass reduction. Exercise may induce myokine secretion such as irisin, which plays a role as a mediator of beiging process, and thus might contribute as treatment of obesity. However, the effects of those exercise formulas on irisin level changes as beiging agent are not known. In addition, metabolic states may affect the irisin responses to those exercise formulas. Therefore, this study was aimed to determine the different effects of exercises using HIIT and CMIT on circulating and tissue irisin levels in normal and abnormal metabolic conditions (obese). Methods Sixteen male Sprague-Dawley rats (8 weeks of age) were randomized to 4 groups according to training regimens (HIIT and CMIT) and metabolic conditions (normal and abnormal/obese). The groups are (1) HIIT on normal metabolic (n=4), (2) CMIT on normal metabolic (n=4), (3) HIIT on abnormal metabolic (n=4), and (4) CMIT on abnormal metabolic (n=4). Abnormal metabolic condition was induced with high fat diet (19% fat) for 8 weeks in obese rats. Irisin levels in serum, skeletal muscle, and white adipose tissue were evaluated by ELISA. Results Serum irisin levels were shown significantly higher in normal metabolic compared to abnormal metabolic condition (P<0.001). The effect of interaction between metabolic condition and exercise formula was found (P<0.01) on adipose irisin levels. The effect of HIIT was shown significantly more effective on adipose irisin levels, compared with CMIT in abnormal metabolic conditions. However, no significant differences of skeletal muscle irisin levels were found in both normal and abnormal metabolic subjects (P>0.05). Regarding exercise formula, no different effects were found between HIIT and CMIT on skeletal muscle irisin levels in both metabolic conditions (P>0.05). The similar findings were observed in serum irisin levels (P>0.05). Conclusions The exercise effects in abnormal metabolic condition might be more adaptable in maintaining the irisin levels in skeletal muscle and induce the irisin uptake from circulation into adipose tissue. In addition, HIIT might be more involved to induce irisin uptake into adipose tissue; thus it might have the significant role in beiging process. However, further research about how the HIIT formula affects the regulation mechanisms of irisin uptake into adipose tissue is still warranted.

Hepatitis C virus (HCV) infection often causes extrahepatic manifestations, such as type 2 diabetes. We previously reported that HCV infection induces the lysosomal degradation of the transcription factor HNF-1a via an interaction with viral NS5A, thereby suppressing GLUT2 gene expression. However, the molecular mechanism of NS5A-induced degradation of HNF-1a is largely unknown. We aimed to identify the determinants necessary for the degradation of HNF-1a induced by NS5A. Coimmunoprecipitation analysis revealed that the POU specific (POU s ) domain spanning from aa 91 to 181 of HNF-1a is responsible for the interaction of NS5A. We also found that the region from aa 121 to 126 of NS5A, which is known as the binding motif of the HCV replication factor FKBP8, is important for the degradation of HNF-1a. A NS5A V121A mutation disrupted the NS5A-HNF-1a interaction as well as the degradation of HNF-1a. Our findings suggest that NS5A Val 121 is crucial for viral pathogenesis.

Hepatitis C virus (HCV) NS5A protein plays crucial roles in viral RNA replication, virus assembly, and viral pathogenesis. Although NS5A has no known enzymatic activity, it modulates various cellular pathways through interaction with cellular proteins. HCV NS5A (and other HCV proteins) are reportedly degraded through the ubiquitin-proteasome pathway; however, the physiological roles of ubiquitylation and deubiquitylation in HCV infection are largely unknown. To elucidate the role of deubiquitylation in HCV infection, an attempt was made to identify a deubiquitinase (DUB) that can interact with NS5A protein. An ovarian tumor protein (OTU), deubiquitinase 7B (OTUD7B), was identified as a novel NS5A-binding protein. Co-immunoprecipitation analyses showed that NS5A interacts with OTUD7B in both Huh-7 and HCV RNA replicon cells. Immunofluorescence staining revealed that HCV NS5A protein colocalizes with OTUD7B in the cytoplasm. Moreover, HCV infection was found to enhance the nuclear localization of OTUD7B. The OTUD7B-binding domain on NS5A was mapped using a series of NS5A deletion mutants. The present findings suggest that the domain I of NS5A is important and the region from amino acid 121 to 126 of NS5A essential for the interaction. Either V121A or V124A mutation in NS5A disrupts the NS5A-OTUD7B interaction. The results of this in vivo ubiquitylation assay suggest that HCV NS5A enhances OTUD7B DUB activity. Taken together, these results suggest that HCV NS5A protein interacts with OTUD7B, thereby modulating its DUB activity.

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BackgroundTransferring critically ill patients with COVID-19 is a challenging task; therefore, well-trained medical team is needed. This study aimed to determine the role of in situ simulation training during pandemic by using high-fidelity manikin to improve interprofessional communication, skills and teamwork in transferring critically ill patients with COVID-19.MethodsThis single-blinded randomised control trial included 40 subjects allocated into standard low-fidelity simulator (LFS) and high-fidelity simulator (HFS) groups. Subjects, who were not members of multiprofessional team taking care of patients with COVID-19, in each group were assigned into small groups and joined an online interactive lecture session, two sessions of in-situ simulation and a debriefing session with strict health protocols. The first simulation aimed to teach participants the skills and steps needed. The second simulation aimed to assess transfer skills, communication and teamwork performance, that participants had learnt using a validated, comprehensive assessment tool. Data were analysed using unpaired t test or Mann-Whitney test.ResultsThe HFS group showed significantly better overall transfer and communication skills than LFS group (89.70±4.65 vs 77.19±3.6, <0.05 and 100 vs 88.34 (63.33–100), p=0.022, respectively). The HFS group also demonstrated significantly better teamwork performance than the standard LFS group (90 (80–900) vs 80 (70–90), p=0.028).ConclusionIn situ simulation training using HFS significantly showed better performance than the standard training using LFS in regards to overall transfer and communication skills as well as teamwork performance. The training using HFS may provide a valuable adjunct to improve interprofessional skills, communication and teamwork performance in transferring critically ill patients with COVID-19.Trial registration numberNCT05113823.

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Background Obesity is a cause of Fgf21 resistance, which affects the browning and thermogenesis process of the adipose tissue. Decreased receptor expression is influenced by microRNA 34a (miR-34a), whose expression is increased in obesity. While Fgf21-based therapies have been widely investigated, the potential activity of Hibiscus sabdariffa Linn extract (HSE) against Fgf21 resistance is unknown. This study aims to determine the effects of HSE on the expression of miR-34a and Fgf21 receptors in white adipose tissue. Methods This experimental study used 24 male Sprague-Dawley rats and divided into four groups: Control (N); diet-induced-obesity rats (DIO); DIO rats with HSE 200 mg/kgBW/day and DIO rats with HSE 400 mg/kgBW/day. Rats were fed a high-fat diet for 17 weeks. HSE was administered daily for five weeks. The administration of HSE 400 mg/kg BW/day resulted in the equivalent expression of miR-34a to that of the control (p > 0.05). Results Fgfr1 receptor expression was also similar to controls (p > 0.05). Beta-klotho expression was significantly lower than that of Control (p < 0.05) but equivalent to that of DIO rats (p < 0.05). Conclusions H. sabdariffa has the potential to reduce Fgf21 resistance in DIO rats through the suppression of miR-34a expression and an increase in the number of Fgfr1 and beta-klotho receptors in adipose tissue.