Human herpesvirus 6 major immediate early promoter has strong activity in T cells and is useful for heterologous gene expression

<p>Abstract</p> <p>Background</p> <p>Human herpesvirus-6 (HHV-6) is a beta-herpesvirus. HHV-6 infects and replicates in T cells. The HHV-6-encoded major immediate early gene (MIE) is expressed at the immediate-early infection phase. Human cytomegalovirus major immediate early promoter (CMV MIEp) is commercially available for the expression of various heterologous genes. Here we identified the HHV-6 MIE promoter (MIEp) and compared its activity with that of CMV MIEp in various cell lines.</p> <p>Methods</p> <p>The HHV-6 MIEp and some HHV-6 MIEp variants were amplified by PCR from HHV-6B strain HST. These fragments and CMV MIEp were subcloned into the pGL-3 luciferase reporter plasmid and subjected to luciferase reporter assay. In addition, to investigate whether the HHV-6 MIEp could be used as the promoter for expression of foreign genes in a recombinant varicella-zoster virus, we inserted HHV-6 MIEp-DsRed expression casette into the varicella-zoster virus genome.</p> <p>Results</p> <p>HHV-6 MIEp showed strong activity in T cells compared with CMV MIEp, and the presence of intron 1 of the MIE gene increased its activity. The NF-κB-binding site, which lies within the R3 repeat, was critical for this activity. Moreover, the HHV-6 MIEp drove heterologous gene expression in recombinant varicella-zoster virus-infected cells.</p> <p>Conclusions</p> <p>These data suggest that HHV-6 MIEp functions more strongly than CMV MIEp in various T-cell lines.</p

An Angiotensin II Type 1 Receptor Blocker Prevents Renal Injury via Inhibition of the Notch Pathway in Ins2 Akita Diabetic Mice

Recently, it has been reported that the Notch pathway is involved in the pathogenesis of diabetic nephropathy. In this study, we investigated the activation of the Notch pathway in Ins2 Akita diabetic mouse (Akita mouse) and the effects of telmisartan, an angiotensin II type1 receptor blocker, on the Notch pathway. The intracellular domain of Notch1 (ICN1) is proteolytically cleaved from the cell plasma membrane in the course of Notch activation. The expression of ICN1 and its ligand, Jagged1, were increased in the glomeruli of Akita mice, especially in the podocytes. Administration of telmisartan significantly ameliorated the expression of ICN1 and Jagged1. Telmisartan inhibited the angiotensin II-induced increased expression of transforming growth factor β and vascular endothelial growth factor A which could directly activate the Notch signaling pathway in cultured podocytes. Our results indicate that the telmisartan prevents diabetic nephropathy through the inhibition of the Notch pathway

Plasma Corticosterone Activates SGK1 and Induces Morphological Changes in Oligodendrocytes in Corpus Callosum

Repeated stressful events are known to be associated with onset of depression. Further, stress activates the hypothalamic–pituitary–adrenocortical (HPA) system by elevating plasma cortisol levels. However, little is known about the related downstream molecular pathway. In this study, by using repeated water-immersion and restraint stress (WIRS) as a stressor for mice, we attempted to elucidate the molecular pathway induced by elevated plasma corticosterone levels. We observed the following effects both, in vivo and in vitro: (1) repeated exposure to WIRS activates the 3-phosphoinositide-dependent protein kinase (PDK1)–serum glucocorticoid regulated kinase (SGK1)–N-myc downstream-regulated gene 1 (NDRG1)–adhesion molecule (i.e., N-cadherin, α-catenin, and β-catenin) stabilization pathway via an increase in plasma corticosterone levels; (2) the activation of this signaling pathway induces morphological changes in oligodendrocytes; and (3) after recovery from chronic stress, the abnormal arborization of oligodendrocytes and depression-like symptoms return to the control levels. Our data strongly suggest that these abnornalities of oligodendrocytes are possibly related to depression-like symptoms

Varicella-Zoster Virus Keratitis with Asymptomatic Conjunctival Viral Shedding in the Contralateral Eye

Purpose: To report a case of varicella-zoster virus (VZV) keratitis with detection of VZV DNA in the tear fluid of not only the symptomatic eye but also the contralateral asymptomatic eye by polymerase chain reaction (PCR). Methods: This is a case report. A 63-year-old otherwise healthy woman presented with circular corneal ulcer and stromal opacity with infiltration accompanied by mild conjunctival and ciliary injections in the left eye. Bacterial cultures of the corneal scrapings and virus PCR analyses of tear fluid from both eyes were performed. Results: No pathogen was found by bacterial cultures. PCR was negative for Acanthamoeba, herpes simplex virus and cytomegalovirus, but positive for VZV. VZV DNA was also detected in the unaffected eye. Based on the diagnosis of VZV keratitis, oral valacyclovir and acyclovir eye ointment were administered to the corneal infected eye. The infected eye was healed and VZV DNA turned negative in the tear fluid of the treated eye after 6 months of treatment; however, VZV DNA was still positive in the tear fluid of the contralateral eye. Conclusions: To our knowledge, this is the first case report of the detection of VZV DNA in the tear fluid of both affected and unaffected eyes in a patient with VZV keratitis. Asymptomatic conjunctival shedding of VZV may continue in the healthy unaffected eye in VZV keratitis patients

Iron overload may be critical for liver dysfunction in anorexia nervosa, and the role of haematocrit-adjusted albumin in assessing nutritional status: a case report

Abstract Background Anorexia nervosa (AN) is frequently associated with liver dysfunction, but the precise mechanism remains undefined. Since the nutritional marker albumin has a low correlation with changes in body weight in AN, and patients with AN often have dehydration as a complication, we also examined whether haematocrit (HCT)-adjusted serum albumin could be a better nutritional marker in AN. Case presentation We describe a 15-year-old girl with severe weight loss and liver damage whose liver enzymes normalized after 1.5 months of hospitalization and weight gain. We found a significant correlation between body weight (BW) and HCT-adjusted serum albumin (Spearman’s rank correlation coefficient (rs) = 0.66, P = 5.28 × 10−3) and between BW and alanine aminotransferase (ALT) (rs = -0.825, P = 8.45 × 10−5). After division by HCT, correlations between serum albumin and ALT (rs = -0.835, P = 5.24 × 10−5) and between the iron-storage protein ferritin and the liver enzyme gamma-glutamyl transferase (rs = 1.0, P = 0.017) were also statistically significant. Conclusion These results suggest that improvement of the nutritional status in AN could relieve liver dysfunction and facilitate iron transport. Since a decrease in the iron-transport protein transferrin presumably increases labile non-transferrin-bound iron, resulting in excess reactive oxygen species production, a defect in iron transport due to malnutrition could be one of the causes of liver injury in AN. In addition, HCT-adjusted albumin could be a better marker than its raw data to assess changes in nutritional status in AN

Characterization of susceptibility variants of poliovirus grown in the presence of favipiravir

Background: T-705 (favipiravir) is a potent inhibitor of RNA-dependent RNA polymerases of influenza viruses and no favipiravir-resistant virus has been isolated. Poliovirus RNA polymerase has been well characterized and isolation of resistant virus was examined in poliovirus. Methods: Susceptibility variants of poliovirus I (Sabin strain) were isolated during passages in the presence of favipiravir and characterized for their susceptibility and the sequence of RNA polymerase. Results: Five variants with 0.47–1.88 times the 50% inhibitory concentration for plaque formation of the parent poliovirus had amino acid variations in the 3D gene of the RNA polymerase. The distribution of amino acid variations was not related to ribavirin resistance, and two amino acid variation sites were found near the finger domain. Conclusion: Favipiravir as a chain terminator would not be incorporated and replicate to cause lethal mutagenesis as a mutagen like ribavirin, and resistant mutants were not isolated. A high replication level would generate mutations leading to favipiravir resistance as ribavirin resistance was generated, but generated mutations would be lethal to the RNA polymerase function. Keywords: Poliovirus, T-705, Favipiravir, Mutants, Resistance, RNA dependent RNA polymeras

Human Herpesvirus 6 Open Reading Frame U14 Protein and Cellular p53 Interact with Each Other and Are Contained in the Virion

A mass spectroscopic analysis of proteins from human herpesvirus 6 (HHV-6)-infected cells showed that the HHV-6 U14 protein coimmunoprecipitated with the tumor suppressor p53. The binding of U14 to p53 was verified by coimmunoprecipitation experiments in both Molt-3 cells infected with HHV-6 and 293 cells cotransfected with U14 and p53 expression vectors. Indirect immunofluorescence assays (IFAs) showed that by 18 h postinfection (hpi) U14 localized to the dot-like structures observed in both the nucleus and cytoplasm where p53 was partly accumulated. Despite Northern blotting evidence that U14 follows late kinetics, the U14 protein was detected immediately after infection (at 3 hpi) by IFA. In addition, by Western blotting, U14 was detected at 0 hpi or in the presence of cycloheximide which completely abolished the expression of IE1 protein. In addition to U14, p53 was detected at 0 hpi although it was not detected in mock-infected cells. Furthermore, both U14 and p53 were clearly detected in the viral particles by Western blotting and immunoelectron microscopy, supporting the idea that U14 and p53 are incorporated into virions. Our study provides the first evidence of the incorporation of cellular p53 into viral particles and suggests that p53 may play an important role in viral infection