Co-degradation of interferon signaling factor DDX3 by PB1-F2 as a basis for high virulence of 1918 pandemic influenza.

The multifunctional influenza virus protein PB1-F2 plays several roles in deregulation of host innate immune responses and is a known immunopathology enhancer of the 1918 influenza pandemic. Here, we show that the 1918 PB1-F2 protein not only interferes with the mitochondria-dependent pathway of type I interferon (IFN) signaling, but also acquired a novel IFN antagonist function by targeting the DEAD-box helicase DDX3, a key downstream mediator in antiviral interferon signaling, toward proteasome-dependent degradation. Interactome analysis revealed that 1918 PB1-F2, but not PR8 PB1-F2, binds to DDX3 and causes its co-degradation. Consistent with intrinsic protein instability as basis for this gain-of-function, internal structural disorder is associated with the unique cytotoxic sequences of the 1918 PB1-F2 protein. Infusing mice with recombinant DDX3 protein completely rescued them from lethal infection with the 1918 PB1-F2-producing virus. Alongside NS1 protein, 1918 PB1-F2 therefore constitutes a potent IFN antagonist causative for the severe pathogenicity of the 1918 influenza strain. Our identification of molecular determinants of pathogenesis should be useful for the future design of new antiviral strategies against influenza pandemics.

Entecavir-resistant hepatitis B virus decreases surface antigenicity: A full genome and functional characterization.

BACKGROUND AND AIM: Since polymerase and surface genes overlap in hepatitis B virus (HBV), an antiviral-induced mutation in the polymerase gene may alter the surface antigenicity in patients with chronic hepatitis B (CHB), but this possibility has not been clearly confirmed. This study aimed to determine the drug susceptibility and surface antigenicity of the patient-derived mutants. PATIENTS AND METHODS: Full-length HBV genomes isolated from four entecavir-resistant CHB patients were cloned and sequenced. Around 10 clones of full-length HBV obtained from each patient were analysed and registered in the NCBI GenBank. Representative clones were further characterized by in vitro drug susceptibility and surface antigenicity assays. RESULTS: The rtL180M + rtM204V mutations were common among all the clones analysed. Additionally, the ETV resistance mutations rtT184A/L, rtS202G and rtM250V were found among three patients. Most of the ETV-resistant mutants had amino acid alterations within the known epitopes recognized by T- and B-cells in the HBV surface and core antigens. The in vitro drug susceptibility assay showed that all tested clones were resistant to ETV treatment. However, they were all susceptible to ADV and TDF. More importantly, the rtI169T mutation in the RT domain, led to the sF161L mutation in the overlapping S gene, which decreased in surface antigenicity. CONCLUSIONS: The ETV resistance mutations can affect the antigenicity of the HBsAg proteins due to changes in the overlapping sequence of this surface antigen. Thus, the apparent decline or disappearance of HBsAg needs to be interpreted cautiously in patients with previous or current antiviral resistance mutations.

Identification of a quadruple mutation that confers tenofovir resistance in chronic hepatitis B patients.

BACKGROUND & AIMS: Tenofovir disoproxil fumarate (TDF) is one the most potent nucleot(s)ide analogues for treating chronic hepatitis B virus (HBV) infection. Phenotypic resistance caused by genotypic resistance to TDF has not been reported. This study aimed to characterize HBV mutations that confer tenofovir resistance. METHODS: Two patients with viral breakthrough during treatment with TDF-containing regimens were prospectively enrolled. The gene encoding HBV reverse transcriptase was sequenced. Eleven HBV clones harboring a series of mutations in the reverse transcriptase gene were constructed by site-directed mutagenesis. Drug susceptibility of each clone was determined by Southern blot analysis and real-time PCR. The relative frequency of mutants was evaluated by ultra-deep sequencing and clonal analysis. RESULTS: Five mutations (rtS106C [C], rtH126Y [Y], rtD134E [E], rtM204I/V, and rtL269I [I]) were commonly found in viral isolates from 2 patients. The novel mutations C, Y, and E were associated with drug resistance. In assays for drug susceptibility, the IC50 value for wild-type HBV was 3.8 ±â€¯0.6 µM, whereas the IC50 values for CYE and CYEI mutants were 14.1 ±â€¯1.8 and 58.1 ±â€¯0.9 µM, respectively. The IC90 value for wild-type HBV was 30 ±â€¯0.5 µM, whereas the IC90 values for CYE and CYEI mutants were 185 ±â€¯0.5 and 790 ±â€¯0.2 µM, respectively. Both tenofovir-resistant mutants and wild-type HBV had similar susceptibility to the capsid assembly modulator NVR 3-778 (IC50 <0.4 µM vs. IC50 = 0.4 µM, respectively). CONCLUSIONS: Our study reveals that the quadruple (CYEI) mutation increases the amount of tenofovir required to inhibit HBV by 15.3-fold in IC50 and 26.3-fold in IC90. These results demonstrate that tenofovir-resistant HBV mutants can emerge, although the genetic barrier is high. LAY SUMMARY: Tenofovir is the most potent nucleotide analogue for the treatment of chronic hepatitis B virus infection and there has been no hepatitis B virus mutation that confers >10-fold resistance to tenofovir up to 8 years. Herein, we identified, for the first time, a quadruple mutation that conferred 15.3-fold (IC50) and 26.3-fold (IC90) resistance to tenofovir in 2 patients who experienced viral breakthrough during tenofovir treatment.

Multiple Functions of Cellular FLIP Are Essential for Replication of Hepatitis B Virus.

Hepatitis B virus (HBV) infection is a leading cause of liver diseases; however, the host factors which facilitate the replication and persistence of HBV are largely unidentified. Cellular FLICE inhibitory protein (c-FLIP) is a typical antiapoptotic protein. In many cases of liver diseases, the expression level of c-FLIP is altered, which affects the fate of hepatocytes. We previously found that c-FLIP and its cleaved form interact with HBV X protein (HBx), which is essential for HBV replication, and regulate diverse cellular signals. In this study, we investigated the role of endogenous c-FLIP in HBV replication and its underlying mechanisms. The knockdown of endogenous c-FLIP revealed that this protein regulates HBV replication through two different mechanisms. (i) c-FLIP interacts with HBx and protects it from ubiquitin-dependent degradation. The N-terminal DED1 domain of c-FLIP is required for HBx stabilization. (ii) c-FLIP regulates the expression or stability of hepatocyte nuclear factors (HNFs), which have critical roles in HBV transcription and maintenance of hepatocytes. c-FLIP regulates the stability of HNFs through physical interactions. We verified our findings in three HBV infection systems: HepG2-NTCP cells, differentiated HepaRG cells, and primary human hepatocytes. In conclusion, our results identify c-FLIP as an essential factor in HBV replication. c-FLIP regulates viral replication through its multiple effects on viral and host proteins that have critical roles in HBV replication.IMPORTANCE Although the chronic hepatitis B virus (HBV) infection still poses a major health concern, the host factors which are required for the replication of HBV are largely uncharacterized. Our studies identify cellular FLICE inhibitory protein (c-FLIP) as an essential factor in HBV replication. We found the dual roles of c-FLIP in regulation of HBV replication: c-FLIP interacts with HBx and enhances its stability and regulates the expression or stability of hepatocyte nuclear factors which are essential for transcription of HBV genome. Our findings may provide a new target for intervention in persistent HBV infection.

Stabilization of Intrinsically Disordered DKK2 Protein by Fusion to RNA-Binding Domain.

Intrinsic disorders are a common feature of hub proteins in eukaryotic interactomes controlling the signaling pathways. The intrinsically disordered proteins (IDPs) are prone to misfolding, and maintaining their functional stability remains a major challenge in validating their therapeutic potentials. Considering that IDPs are highly enriched in RNA-binding proteins (RBPs), here we reasoned and confirmed that IDPs could be stabilized by fusion to RBPs. Dickkopf2 (DKK2), Wnt antagonist and a prototype IDP, was fused with lysyl-tRNA synthetase (LysRS), with or without the fragment crystallizable (Fc) domain of an immunoglobulin and expressed predominantly as a soluble form from a bacterial host. The functional competence was confirmed by in vitro Wnt signaling reporter and tube formation in human umbilical vein endothelial cells (HUVECs) and in vivo Matrigel plug assay. The removal of LysRS by site-specific protease cleavage prompted the insoluble aggregation, confirming that the linkage to RBP chaperones the functional competence of IDPs. While addressing to DKK2 as a key modulator for cancer and ischemic vascular diseases, our results suggest the use of RBPs as stabilizers of disordered proteinaceous materials for acquiring and maintaining the structural stability and functional competence, which would impact the druggability of a variety of IDPs from human proteome.

Suppression of interferon-mediated anti-HBV response by single CpG methylation in the 5'-UTR of TRIM22.

OBJECTIVE: Interferons (IFNs) mediate direct antiviral activity. They play a crucial role in the early host immune response against viral infections. However, IFN therapy for HBV infection is less effective than for other viral infections. DESIGN: We explored the cellular targets of HBV in response to IFNs using proteome-wide screening. RESULTS: Using LC-MS/MS, we identified proteins downregulated and upregulated by IFN treatment in HBV X protein (HBx)-stable and control cells. We found several IFN-stimulated genes downregulated by HBx, including TRIM22, which is known as an antiretroviral protein. We demonstrated that HBx suppresses the transcription of TRIM22 through a single CpG methylation in its 5'-UTR, which further reduces the IFN regulatory factor-1 binding affinity, thereby suppressing the IFN-stimulated induction of TRIM22. CONCLUSIONS: We verified our findings using a mouse model, primary human hepatocytes and human liver tissues. Our data elucidate a mechanism by which HBV evades the host innate immune system.

Intravenous Tranexamic Acid Reduces Postoperative Blood Loss After High Tibial Osteotomy.

BACKGROUND: Medial opening wedge high tibial osteotomy (HTO) entails extensive soft tissue release that may lead to substantial perioperative bleeding. Although tranexamic acid (TXA) is a well-established blood-conserving agent in total joint arthroplasty, its potential to reduce blood loss in patients undergoing HTO has not been studied extensively. QUESTIONS/PURPOSES: (1) Does TXA reduce total estimated blood loss in HTO? (2) Does TXA use in HTO affect in-hospital endpoints as measured by visual analog scale (VAS) pain scores at rest the day after surgery, wound complications in the immediate postoperative period, blood transfusions, or symptomatic deep vein thrombosis? METHODS: Between January 2015 and May 2017, a single surgeon performed 156 HTOs, all of which were done using the medial opening wedge technique. We began using intravenous TXA for all HTOs in June 2016. This left us with 89 patients who were treated during a time when no TXA was used and 67 patients who were treated when all patients received TXA. Two patients in the control group had simultaneous TKA in the contralateral leg and one patient in each group had missing data so these patients were excluded, leaving 86 (97%) patients in the control group and 66 (98.5%) in the TXA group available for analysis in this retrospective study. There were no demographic differences between the groups in terms of age, sex, body mass index, and baseline hemoglobin values. Total estimated blood loss was the primary outcome variable, which was calculated using total blood volume and decrease in hemoglobin values. Secondary outcome variables included pain VAS at rest the day after surgery, wound complications in the immediate postoperative period, allogeneic blood transfusions, and occurrence of symptomatic thromboembolic manifestations. The decision on when to transfuse was based on predetermined criteria. An orthopaedic surgeon not involved in patient care collected the patient data from electronic medical records and did chart review. RESULTS: The TXA group had less total blood loss (372 ± 36 mL versus 635 ± 53 mL, mean difference 263 mL [95% confidence interval, 248-278]; p < 0.001). Between groups, differences in VAS pain scores at rest the day after surgery favored the TXA group but were small and unlikely to be clinically important. There were two wound complications in the control group (one hematoma and one superficial wound infection) and none in the TXA group. No patients in either group received a blood transfusion, and no symptomatic thromboembolic events were detected in either group. CONCLUSIONS: This study demonstrates that the systemic administration of TXA reduces postoperative blood loss in medial opening wedge HTO; however, insofar as no transfusions were administered to patients even before the routine use of TXA in this series, and no clinically important differences in pain scores were identified, the clinical benefit of routine use of TXA in patients undergoing HTO is uncertain. Our study was too small to make safety-related claims on rare endpoints such as wound complications or thromboembolic events. Larger, and preferably randomized, trials are needed to help define whether it is important to use TXA in this setting. Our data can help inform sample size calculations for such studies. LEVEL OF EVIDENCE: Level III, therapeutic study.

Immunohistochemical mapping and transcript expression of the GPA2/GPB5 receptor in tissues of the adult mosquito, Aedes aegypti.

GPA2/GPB5 is a glycoprotein hormone found in most bilateral metazoans including the mosquito, Aedes aegypti. To elucidate physiological roles and functions of GPA2/GPB5, we aim to identify prospective target tissues by examining the tissue- and sex-specific expression profile of its receptor, the leucine-rich repeat-containing G protein-coupled receptor 1 (LGR1) in the adult mosquito. Western analyses using a heterologous system with CHO-K1 cells, transiently expressing A. aegypti LGR1, yielded a 112-kDa monomeric band and high-molecular weight multimers, which associated with membrane-protein fractions. Moreover, immunoblot analyses on protein isolated from HEK 293 T cells stably expressing a fusion construct of A. aegypti LGR1-EGFP (LGR1: 105 kDa+EGFP: 27 kDa) yielded a band with a measured molecular weight of 139 kDa that also associated with membrane-protein fractions and upon deglycosylation, migrated as a lower molecular weight band of 132 kDa. Immunocytochemical analysis of HEK 293 T cells stably expressing this fusion construct confirmed EGFP fluorescence and LGR1-like immunoreactivity colocalized primarily to the plasma membrane. Immunohistochemical mapping in adult mosquitoes revealed LGR1-like immunoreactivity is widespread in the alimentary canal. Importantly, LGR1-like immunoreactivity localizes specifically to basolateral regions of epithelia and, in some regions, appeared as punctate intracellular staining, which together indicates a potential role in feeding and/or hydromineral balance. LGR1 transcript expression was also detected in gut regions that exhibited strong LGR1-like immunoreactivity. Interestingly, LGR1 transcript expression and strong LGR1-like immunoreactivity was also identified in reproductive tissues including the testes and ovaries, which together suggests a potential role linked to spermatogenesis and oogenesis in male and female mosquitoes, respectively.

Cleaved c-FLIP mediates the antiviral effect of TNF-α against hepatitis B virus by dysregulating hepatocyte nuclear factors.

BACKGROUND & AIMS: Cytokines are key molecules implicated in the defense against virus infection. Tumor necrosis factor-alpha (TNF-α) is well known to block the replication of hepatitis B virus (HBV). However, the molecular mechanism and the downstream effector molecules remain largely unknown. METHODS: In this study, we investigated the antiviral effect and mechanism of p22-FLIP (FLICE-inhibitory protein) by ectopic expression in vitro and in vivo. In addition, to provide the biological relevance of our study, we examined that the p22-FLIP is involved in TNF-α-mediated suppression of HBV in primary human hepatocytes. RESULTS: We found that p22-FLIP, a newly discovered c-FLIP cleavage product, inhibited HBV replication at the transcriptional level in both hepatoma cells and primary human hepatocytes, and that c-FLIP conversion to p22-FLIP was stimulated by the TNF-α/NF-κB pathway. p22-FLIP inhibited HBV replication through the upregulation of HNF3ß but downregulation of HNF4α, thus inhibiting both HBV enhancer elements. Finally, p22-FLIP potently inhibited HBV DNA replication in a mouse model of HBV replication. CONCLUSIONS: Taken together, these findings suggest that the anti-apoptotic p22-FLIP serves a novel function of inhibiting HBV transcription, and mediates the antiviral effect of TNF-α against HBV replication.

Bilateral sternoclavicular joint dislocation due to sternal fracture: Is it a dislocation or a separation?

Traumatic bilateral sternoclavicular joint dislocation is very rare injury. In shoulder girdle injuries, anterior dislocation of the sternoclavicular joint accounts for 3 % and posterior sternoclavicular joint dislocation is lesser. Previous reported cases about bilateral sternoclavicular joint dislocation were result from proximal clavicle fracture with intact connection between sternum and ribs. But, the sternoclavicular joint dislocation secondary to fracture and angulation of the sternum with intact relationship between ribs and clavicle has not been reported. Authors experienced patient who has a bilateral anterior sternoclavicular joint dislocation caused by sternum fracture and anterior angulation, but intact relationship between ribs and clavicle. We report this case with satisfactory result.

Hepatocystin contributes to interferon-mediated antiviral response to hepatitis B virus by regulating hepatocyte nuclear factor 4α.

Hepatocystin/80K-H is known as a causative gene for autosomal dominant polycystic liver disease. However, the role of hepatocystin in hepatitis B virus-related liver disease remains unknown. Here, we investigated the role of hepatocystin on the cytokine-mediated antiviral response against hepatitis B virus infection. We investigated the antiviral effect and mechanism of hepatocystin by ectopic expression and RNAi knockdown in cell culture and mouse livers. Hepatocystin suppressed the replication of hepatitis B virus both in vitro and in vivo. This inhibitory effect was HBx-independent and mediated by the transcriptional regulation of viral genome via the activation of exogenous signal-regulated kinase 1/2 and the reduced expression of hepatocyte nuclear factor 4α, a transcription factor essential for hepatitis B virus replication. The amino-terminal region of hepatocystin was essential for regulation of this antiviral signaling pathway. We also found that hepatocystin acts as a critical component in interferon-mediated mitogen-activated protein kinase signaling pathway, and the interferon-induced antiviral activity against hepatitis B virus is associated with the expression levels of hepatocystin. We demonstrated that hepatocystin plays a critical role in modulating the susceptibility of hepatitis B virus to interferon, suggesting that the modulation of hepatocystin expression is important for cytokine-mediated viral clearance during hepatitis B virus infection.

The impact of the hepatitis B virus polymerase rtA181T mutation on replication and drug resistance is potentially affected by overlapping changes in surface gene.

UNLABELLED: The emergence of drug-resistant hepatitis B virus (HBV) is a major problem for antiviral treatment in chronic hepatitis B infection. In this study, we analyzed the evolution of drug-resistant mutations and characterized the effects of the rtA181T and rtI233V mutations on viral replication and drug resistance. We performed a clonal analysis of the HBV polymerase gene from serum samples during viral breakthrough treated with antiviral agents. A series of mutant clones containing rtA181T and/or rtI233V mutations were constructed and determined the effect of these mutations on the replication ability and drug resistance. An in vitro study revealed that the effect of the rtA181T mutation on viral replication and drug resistance is dependent on the mutations in the overlapping surface gene. Compared to the rtA181T surface missense mutation (rtA181T/sW172S), the introduction of rtA181T surface nonsense mutation (rtA181T/sW172*) resulted in decreased viral replication and increased drug resistance. Complementation assay revealed that the truncated PreS1 is responsible for reduced replication of rtA181T/sW172* mutant. Moreover, the rtA181T/sW172* mutant exhibited a defect in viral particle secretion. The rtI233V mutation that emerged during adefovir therapy reduced viral replication and conferred resistance to adefovir. Our data suggest that the impact of the rtA181T mutation on replication and drug resistance differs based on the mutation status of the corresponding surface gene. The rtI233V mutation also affects replication ability and drug resistance. This observation suggests the need for genotypic analysis of overlapping surface genes to manage antiviral drug resistance if clinical isolates harbor the rtA181T mutation. IMPORTANCE: The emergence of drug-resistant HBV that are no longer susceptible to nucleos(t)ide analogues is a major problem for antiviral treatment in chronic hepatitis B infection. Among drug-resistant mutations, the single rtA181T mutation is known to confer cross-resistance to antiviral drugs. This mutation causes intermediate or reduced susceptibility to tenofovir. Moreover, the clinical occurrence of the rtA181T mutation during antiviral therapy is also high. Our study revealed that the effect of the rtA181T mutation on viral replication and drug resistance is dependent on the mutations in the overlapping surface gene. This observation suggests the need for genotypic analysis of overlapping surface genes to manage antiviral drug resistance if clinical isolates harbor the rtA181T mutation. We believe that our study will not only extend the understanding of the drug resistance mechanism, but it will also ultimately provide new treatment options for patients with multidrug resistant HBV.

Hepatitis B virus inhibits liver regeneration via epigenetic regulation of urokinase-type plasminogen activator.

UNLABELLED: Liver regeneration after liver damage caused by toxins and pathogens is critical for liver homeostasis. Retardation of liver proliferation was reported in hepatitis B virus (HBV) X protein (HBx)-transgenic mice. However, the underlying mechanism of the HBx-mediated disturbance of liver regeneration is unknown. We investigated the molecular mechanism of the inhibition of liver regeneration using liver cell lines and a mouse model. The mouse model of acute HBV infection was established by hydrodynamic injection of viral DNA. Liver regeneration after partial hepatectomy was significantly inhibited in the HBV DNA-treated mice. Mechanism studies have revealed that the expression of urokinase-type plasminogen activator (uPA), which regulates the activation of hepatocyte growth factor (HGF), was significantly decreased in the liver tissues of HBV or HBx-expressing mice. The down-regulation of uPA was further confirmed using liver cell lines transiently or stably transfected with HBx and the HBV genome. HBx suppressed uPA expression through the epigenetic regulation of the uPA promoter in mouse liver tissues and human liver cell lines. Expression of HBx strongly induced hypermethylation of the uPA promoter by recruiting DNA methyltransferase (DNMT) 3A2. CONCLUSION: Taken together, these results suggest that infection of HBV impairs liver regeneration through the epigenetic dysregulation of liver regeneration signals by HBx.

Recognition of IoT-based fire-detection system fire-signal patterns applying fuzzy logic.

In Korea, the use of fire-detection systems applying IoT technology to existing analog fire-alarm systems has increased owing to the communication technology convergence, the world's best Internet network, and the proliferation of Internet of Things (IoT). Its use can be expected to increase worldwide in the future. For IoT-based fire-detection systems to exhibit the requisite reliability (based on a low false-alarm rate), research related to the analysis of detection signals should be actively promoted and conducted. However, there has been no research activity based on actual operational data, apart from the research that has been conducted in laboratory environments. The primary reason for this state of affairs has been that the installation and use of IoT-based fire-detection systems on a large scale has been rare, worldwide. Consequently, with respect to the fire-signal characteristics of IoT-based fire-detection systems, related data in this study were obtained by investigating actual fire accident cases, using fire alarm data that occurred over a period of 5 years. Based on the signal pattern analysis results using these field data, a fuzzy logic system for recognizing fire signal patterns was developed and verified. As a result, in the actual fire accidents examined, an "alarm" condition-corresponding to the high possibility of fire among the five fire alarms-was determined 30 s before the actual fire alarm. Moreover, it was also found that approximately 80% of non-fire alarms could be reduced in the actual fire alarms that occurred at Institute K during the 5-year period examined.

Comparative analysis of trap-based program/erase behaviors with different tunnel barriers.

In this study, we comparatively analyze the trap-based memory characteristics of Oxide-Nitride-Oxide (ONO) devices with different tunnel dielectrics. We fabricated two kinds of ONO devices-one is the conventional single tunnel oxide structure and the other is the bandgap engineered structure in which the modulated tunnel dielectric replaces the single tunnel oxide. The charge storage layer is 9 nm and the blocking oxide is 7 nm in both two kinds of ONO devices. Based on experimental results, we find that the memory speed is promoted to 2-4 times and 10-year data retention greatly improves in the bandgap engineered device comparing to those in the conventional device. As a result, the bandgap engineered tunnel barrier device embodies both fast P/E operation speeds and excellent long-term data retention characteristics, hence, the bandgap engineered tunnel barrier is expected to conduct performance optimization for the future scaled SONOS flash memory.

Analysis of Thermal Characteristics and Insulation Resistance Based on the Installation Year and Accelerated Test by Electrical Socket Outlets.

BACKGROUND: Electrical socket outlets are used continuously until a failure occurs because they have no indication of manufacturing date or exchange specifications. For this reason, 659 electrical fires related to electrical socket outlets broke out in the Republic of Korea at 2018 only, an increase year on year. To reduce electrical fires from electrical socket outlets, it is necessary to perform an accelerated test and analyze the thermal, insulation resistance, and material properties of electrical socket outlets by installation years. METHODS: Thermal characteristics were investigated by measured the temperature increase of electrical socket outlets classified according to year with variation of the current level. Insulation resistance characteristics was measured according to temperature for an electrical socket outlets by their years of use. Finally, to investigate the thermal and insulation resistance characteristics in relation to outlet aging, this study analyzed electrical socket outlets' conductor surface and content, insulator weight, and thermal deformation temperature. RESULTS: Analysis showed, regarding the thermal characteristics, that electrical socket outlet temperature rose when the current value increased. Moreover, the longer the time that had elapsed since an accelerated test and installation, the higher the electrical socket outlet temperature was. With respect to the insulation resistance properties, the accelerated test (30 years) showed that insulation resistance decreased from 110 °C. In relation to the installation year (30 years), insulation resistance decreased from 70 °C, which is as much as 40 °C lower than the result found by the accelerated test. Regarding the material properties, the longer the elapsed time since installation, the rougher the surface of conductor contact point was, and cracks increased. CONCLUSION: The 30-year-old electrical socket outlet exceeded the allowable temperature which is 65 °C of the electrical contacts at 10 A, and the insulation resistance began to decrease at 70 °C. It is necessary to manage electrical socket outlets that have been installed for a long time.

Factors affecting expression and transcription of uncoupling protein 2 gene.

Previous studies suggest a negative relationship between hepatic oxidative stress and productivity in beef cattle. Uncoupling protein 2 (UCP2) is involved in the disappearance of reactive oxygen species, suggesting the defensive role of UCP2 against oxidative stress. The present study examined the relationship between oxidative stress and expression levels of UCP2/Ucp2 in cultured human and mouse liver-derived cells. We also explored factors regulating bovine Ucp2 transcription. As oxidative stress inducers, hydrogen peroxide, ethanol, and cumene hydroperoxide (CmHP) were used. Expression levels of hemoxygenase 1 (HMOX1), a representative gene induced by oxidative stress, were not affected by any oxidative stress inducers in HepG2 human liver-derived cells. The levels of UCP2 mRNA were also unaffected by the oxidative stress inducers. Treatment with CmHP increased expression of Hmox1 in Hepa1-6 mouse liver-derived cells, but Ucp2 expression was not changed. Stimulus screening for regulator of transcription (SSRT) revealed that expression of p50 or p65, transcription factors conferring response to oxidative stress, did not stimulate bovine Ucp2 transcrition in HepG2 cells. SSRT also showed 11 molecules that induced Ucp2 transcription more than 4-fold; among them, endoplasmic reticulum (ER) stress-related transcription factors such as XBP1, c-JUN, JUNB, and C/EBPß were identified. However, treatment with ER stress inducers did not increase Ucp2 expression in HepG2 and Hepa1-6 cells. The present results suggest that 1) neither oxidative stress nor ER stress induces Ucp2 expression in liver-derived cells, and 2) Ucp2 transcription is stimulated by several transcription factors.

Effect of hepato-toxins in the acceleration of hepatic fibrosis in hepatitis B mice.

Chronic liver diseases such as hepatitis B viral (HBV) infection and liver fibrosis have been a major health problem worldwide. However, less research has been conducted owing to the lack of animal models. The key purpose of this study was to determine the effects of different hepatotoxins in HBV-affected liver. In this study, we successfully generated a combined liver fibrosis model by administering HBV 1.2 plasmid and thioacetamide/ethanol (TAA/EtOH). To our knowledge, this is the first study in which an increase in the liver fibrosis level is observed by the intraperitoneal administration of TAA and EtOH in drinking water after the hydrodynamic transfection of the HBV 1.2 plasmid in C3H/HeN mice. The HBV+TAA/EtOH group exhibited higher level of hepatic fibrosis than that of the control groups. The hepatic stellate cell activation in the TAA- and EtOH-administered groups was demonstrated by the elevation in the level of fibrotic markers. In addition, high levels of collagen content and histopathological results were also used to confirm the prominent fibrotic levels. We established a novel HBV mice model by hydrodynamic injection-based HBV transfection in C3H/HeN mice. C3H/HeN mice were reported to have a higher HBV persistence level than that of the C57BL/6 mouse model. All the results showed an increased fibrosis level in the HBV mice treated with TAA and EtOH; hence, this model would be useful to understand the effect of hepatotoxins on the high risk of fibrosis after HBV infection. The acceleration of liver fibrosis can occur with prolonged administration as well as the high dosage of hepatotoxins in mice.

Enantioselective synthesis of (R)-(+)- and (S)-(-)-higenamine and their analogues with effects on platelet aggregation and experimental animal model of disseminated intravascular coagulation.

Optically active tetrahydroisoquinoline alkaloids, (R)-(+)-higenamine (1R) and (S)-(-)-higenamine (1 S), and their optically active 1-naphthylmethyl analogues (2 and 3), were synthesized by enantioselective hydrogenation of the corresponding dihydroisoquinoline intermediates 7 as a key step. The evaluation of the platelet anti-aggregation effect demonstrated clearly that the (S)-(-)-enantiomers, 1S, 2S, and 3S, had higher inhibitory potency than the corresponding (R)-(+)-antipodes, 1R, 2R, and 3R, respectively, to platelet aggregation induced by epinephrine. 1S enantiomer was superior to the corresponding 1R enantiomer in attenuating all of the disseminated intravascular coagulation (DIC) and multiple organ failure (MOF) parameters tested, while the S enantiomers 2S and 3S ameliorated some of the DIC and MOF parameters more effectively than the corresponding antipodes 2R and 3R.

Generation and Characterization of a Neutralizing Human Monoclonal Antibody to Hepatitis B Virus PreS1 from a Phage-Displayed Human Synthetic Fab Library.

The hepatitis B virus (HBV) envelope contains small (S), middle (M), and large (L) proteins. PreS1 of the L protein contains a receptor-binding motif crucial for HBV infection. This motif is highly conserved among 10 HBV genotypes (A-J), making it a potential target for the prevention of HBV infection. In this study, we successfully generated a neutralizing human monoclonal antibody (mAb), 1A8 (IgG1), that recognizes the receptor-binding motif of preS1 using a phage-displayed human synthetic Fab library. Analysis of the antigen-binding activity of 1A8 for different genotypes indicated that it can specifically bind to the preS1 of major HBV genotypes (A-D). Based on Bio-Layer interferometry, the affinity (KD) of 1A8 for the preS1 of genotype C was 3.55 nM. 1A8 immunoprecipitated the hepatitis B virions of genotypes C and D. In an in vitro neutralization assay using HepG2 cells overexpressing the cellular receptor sodium taurocholate cotransporting polypeptide, 1A8 effectively neutralized HBV infection with genotype D. Taken together, the results suggest that 1A8 may neutralize the four HBV genotypes. Considering that genotypes A-D are most prevalent, 1A8 may be a neutralizing human mAb with promising potential in the prevention and treatment of HBV infection.