Structural basis for the interaction of human herpesvirus 6B tetrameric glycoprotein complex with the cellular receptor, human CD134.

A unique glycoprotein is expressed on the virus envelope of human herpesvirus 6B (HHV-6B): the complex gH/gL/gQ1/gQ2 (hereafter referred to as the HHV-6B tetramer). This tetramer recognizes a host receptor expressed on activated T cells: human CD134 (hCD134). This interaction is essential for HHV-6B entry into the susceptible cells and is a determinant for HHV-6B cell tropism. The structural mechanisms underlying this unique interaction were unknown. Herein we solved the interactions between the HHV-6B tetramer and the receptor by using their neutralizing antibodies in molecular and structural analyses. A surface plasmon resonance analysis revealed fast dissociation/association between the tetramer and hCD134, although the affinity was high (KD = 18 nM) and comparable to those for the neutralizing antibodies (anti-gQ1: 17 nM, anti-gH: 2.7 nM). A competition assay demonstrated that the anti-gQ1 antibody competed with hCD134 in the HHV-6B tetramer binding whereas the anti-gH antibody did not, indicating the direct interaction of gQ1 and hCD134. A single-particle analysis by negative-staining electron microscopy revealed the tetramer's elongated shape with a gH/gL part and extra density corresponding to gQ1/gQ2. The anti-gQ1 antibody bound to the tip of the extra density, and anti-gH antibody bound to the putative gH/gL part. These results highlight the interaction of gQ1/gQ2 in the HHV-6B tetramer with hCD134, and they demonstrate common features among viral ligands of the betaherpesvirus subfamily from a macroscopic viewpoint.

Tetrameric glycoprotein complex gH/gL/gQ1/gQ2 is a promising vaccine candidate for human herpesvirus 6B.

Primary infection of human herpesvirus 6B (HHV-6B) occurs in infants after the decline of maternal immunity and causes exanthema subitum accompanied by a high fever, and it occasionally develops into encephalitis resulting in neurological sequelae. There is no effective prophylaxis for HHV-6B, and its development is urgently needed. The glycoprotein complex gH/gL/gQ1/gQ2 (called 'tetramer of HHV-6B') on the virion surface is a viral ligand for its cellular receptor human CD134, and their interaction is thus essential for virus entry into the cells. Herein we examined the potency of the tetramer as a vaccine candidate against HHV-6B. We designed a soluble form of the tetramer by replacing the transmembrane domain of gH with a cleavable tag, and the tetramer was expressed by a mammalian cell expression system. The expressed recombinant tetramer is capable of binding to hCD134. The tetramer was purified to homogeneity and then administered to mice with aluminum hydrogel adjuvant and/or CpG oligodeoxynucleotide adjuvant. After several immunizations, humoral and cellular immunity for HHV-6B was induced in the mice. These results suggest that the tetramer together with an adjuvant could be a promising candidate HHV-6B vaccine.

An Animal Model That Mimics Human Herpesvirus 6B Pathogenesis.

Human herpesvirus 6B (HHV-6B), a T-lymphotropic virus, infects almost exclusively humans. An animal model of HHV-6B has not been available. Here, we report the first animal model to mimic HHV-6B pathogenesis; the model is based on humanized mice in which human immune cells were engrafted and maintained. For HHV-6B replication, adequate human T-cell activation (which becomes susceptible to HHV-6B) is necessary in this murine model. Here, we found that an additional transfer of human mononuclear cells to humanized mice resulted in an explosive proliferation of human activated T cells, which could be representative of graft-versus-host disease (GVHD) because the primary transfer of human cells was not sufficient to increase the number and ratio of human T cells. Mice infected with HHV-6B became weak and/or died approximately 7 to 14 days later. Quantitative PCR analysis revealed that the spleen and lungs were the major sites of HHV-6B replication in this model, and this was corroborated by the detection of viral proteins in these organs. Histological analysis also revealed the presence of megakaryocytes, indicating HHV-6B infection. Multiplex analysis of cytokines/chemokines in sera from the infected mice showed secretions of human cytokines/chemokines as reported for both in vitro infection and clinical samples, indicating that the secreted cytokines could affect pathogenesis. This is the first animal model showing HHV-6B pathogenesis, and it will be useful for elucidating the pathogenicity of HHV-6B, which is related to GVHD and idiopathic pneumonia syndrome.IMPORTANCE Human herpesvirus 6B (HHV-6B) is a ubiquitous virus that establishes lifelong latent infection only in humans, and the infection can reactivate, with severe complications that cause major problems. A small-animal model of HHV-6B infection has thus been desired for research regarding the pathogenicity of HHV-6B and the development of antiviral agents. We generated humanized mice by transplantation with human hematopoietic stem cells, and here, we modified the model by providing an additional transfer of human mononuclear cells, providing the proper conditions for efficient HHV-6B infection. This is the first humanized mouse model to mimic HHV-6B pathogenesis, and it has great potential for research into the in vivo pathogenesis of HHV-6B.

Humanization of Murine Neutralizing Antibodies against Human Herpesvirus 6B.

Exanthem subitum is a common childhood illness caused by primary infection with human herpesvirus 6B (HHV-6B). It is occasionally complicated by febrile seizures and even encephalitis. HHV-6B reactivation also causes encephalitis, especially after allogeneic hematopoietic stem cell transplantation. However, no adequate antiviral treatment for HHV-6B has yet been established. Mouse-derived monoclonal antibodies (MAbs) against the HHV-6B envelope glycoprotein complex gH/gL/gQ1/gQ2 have been shown to neutralize the viral infection. These antibodies have the potential to become antiviral agents against HHV-6B despite their inherent immunogenicity to the human immune system. Humanization of MAbs derived from other species is one of the proven solutions to such a dilemma. In this study, we constructed chimeric forms of two neutralizing MAbs against HHV-6B to make humanized antibodies. Both showed neutralizing activities equivalent to those of their original forms. This is the first report of humanized antibodies against HHV-6B and provides a basis for the further development of HHV-6B-specific antivirals.IMPORTANCE Human herpesvirus 6B (HHV-6B) establishes lifelong latent infection in most individuals after the primary infection. Encephalitis is the most severe complication caused by both the primary infection and the reactivation of HHV-6B and is the cause of considerable mortality in patients, without any established treatments to date. The humanization of the murine neutralizing antibodies described in this research provided a feasible way to reduce the inherent immunogenicity of the antibodies without changing their neutralizing activities. These newly designed chimeric antibodies against HHV-6B have the potential to be candidates for antivirals for future use.

Crystal Structure of Human Herpesvirus 6B Tegument Protein U14.

The tegument protein U14 of human herpesvirus 6B (HHV-6B) constitutes the viral virion structure and is essential for viral growth. To define the characteristics and functions of U14, we determined the crystal structure of the N-terminal domain of HHV-6B U14 (U14-NTD) at 1.85 Å resolution. U14-NTD forms an elongated helix-rich fold with a protruding ß hairpin. U14-NTD exists as a dimer exhibiting broad electrostatic interactions and a network of hydrogen bonds. This is first report of the crystal structure and dimerization of HHV-6B U14. The surface of the U14-NTD dimer reveals multiple clusters of negatively- and positively-charged residues that coincide with potential functional sites of U14. Three successive residues, L424, E425 and V426, which relate to viral growth, reside on the ß hairpin close to the dimer's two-fold axis. The hydrophobic side-chains of L424 and V426 that constitute a part of a hydrophobic patch are solvent-exposed, indicating the possibility that the ß hairpin region is a key functional site of HHV-6 U14. Structure-based sequence comparison suggests that U14-NTD corresponds to the core fold conserved among U14 homologs, human herpesvirus 7 U14, and human cytomegalovirus UL25 and UL35, although dimerization appears to be a specific feature of the U14 group.

GenDet: Meta Learning to Generate Detectors From Few Shots.

Object detection has made enormous progress and has been widely used in many applications. However, it performs poorly when only limited training data is available for novel classes that the model has never seen before. Most existing approaches solve few-shot detection tasks implicitly without directly modeling the detectors for novel classes. In this article, we propose GenDet, a new meta-learning-based framework that can effectively generate object detectors for novel classes from few shots and, thus, conducts few-shot detection tasks explicitly. The detector generator is trained by numerous few-shot detection tasks sampled from base classes each with sufficient samples, and thus, it is expected to generalize well on novel classes. An adaptive pooling module is further introduced to suppress distracting samples and aggregate the detectors generated from multiple shots. Moreover, we propose to train a reference detector for each base class in the conventional way, with which to guide the training of the detector generator. The reference detectors and the detector generator can be trained simultaneously. Finally, the generated detectors of different classes are encouraged to be orthogonal to each other for better generalization. The proposed approach is extensively evaluated on the ImageNet, VOC, and COCO data sets under various few-shot detection settings, and it achieves new state-of-the-art results.

Constitutive Model of Isotropic Magneto-Sensitive Rubber with Amplitude, Frequency, Magnetic and Temperature Dependence under a Continuum Mechanics Basis.

A three-dimensional nonlinear constitutive model of the amplitude, frequency, magnetic and temperature dependent mechanical property of isotropic magneto-sensitive (MS) rubber is developed. The main components of MS rubber are an elastomer matrix and magnetizable particles. When a magnetic field is applied, the modulus of MS rubber increases, which is known as the magnetic dependence of MS rubber. In addition to the magnetic dependence, there are frequency, amplitude and temperature dependencies of the dynamic modulus of MS rubber. A continuum mechanical framework-based constitutive model consisting of a fractional standard linear solid (SLS) element, an elastoplastic element and a magnetic stress term of MS rubber is developed to depict the mechanical behavior of MS rubber. The novelty is that the amplitude, frequency, magnetic and temperature dependent mechancial properties of MS rubber are integrated into a whole constitutive model under the continuum mechanics frame. Comparison between the simulation and measurement results shows that the fitting effect of the developed model is very good. Therefore, the constitutive model proposed enables the prediction of the mechanical properties of MS rubber under various operating conditions with a high accuracy, which will drive MS rubber's application in engineering problems, especially in the area of MS rubber-based anti-vibration devices.

[Prevention of traditional Chinese medicine Gubiao Pixie prescription for nosocomial infection in elderly population].

OBJECTIVE: To observe the curative effect of traditional Chinese medicine (TCM) Gubiao Pixie prescription on the prevention of nosocomial infection in elderly patients with susceptible factors. METHODS: A prospective observational study was conducted. The elderly patients aged over 70 years admitted to the Department of Integrated TCM and Western Medicine of Tianjin First Center Hospital from March 2015 to March 2016 were enrolled. The patients were divided into experiment group and control group by random number table. The patients in control group were given conventional treatment without special intervention; and those in experimental group were given Gubiao Pixie prescription particles (prescription composition: Radix Astragali 30 g, Bran fried Rhizoma Atracylodis 20 g, Raidix Saposhnikoviae 12 g, Radix Scutellariae 10 g, Fructus Tsaoko 6 g) on the basis of conventional treatment. The Gubiao Pixie prescription particles were taken in warm water 300 mL, twice a day in morning and evening respectively, taking half an hour after meals, and were increased or decreased according to the disease condition. The changes in immune function parameters before and after treatment, as well as the incidence of nosocomial infection, the abnormal increase rate of body temperature, white blood cell (WBC), and C-reactive protein (CRP) after 10 days of treatment in the two groups were observed. RESULTS: A total of 110 elderly patients with susceptible factors during hospitalization were included. After the exclusion of vomiting, abdominal distension and failure to conform the trial requirements, hospitalization time less than 10 days of patients, a total of 100 patients were enrolled in the analysis finally, with 50 patients in control group and in experimental group respectively. There were no significant differences in immune function parameters including IgA, IgG, IgM before treatment between the two groups. After 10 days of treatment, the immune function parameters showed no significant improvement in control group, and those in experiment group were improved significantly, and IgA (g/L: 1.59±0.32 vs. 1.29±0.29), IgG (g/L: 12.07±2.37 vs. 10.23±1.91), and IgM (g/L: 1.01±0.29 vs. 0.88±0.24) were significantly increased as compared with those of control group (all P < 0.01). Compared with the control group, the incidence of nosocomial infection (20% vs. 38%) and the abnormal increase rate of body temperature (24% vs. 44%), WBC (28% vs. 52%), and CRP (28% vs. 50%) 10 days after treatment in experimental group were significantly decreased (all P < 0.05). CONCLUSIONS: TCM Gubiao Pixie prescription has a role in enhancing immune function and antibacterial and bactericidal effect. It has certain preventive effect on nosocomial infection in susceptible people.

Human herpesvirus 6 U11 protein is critical for virus infection.

All herpesviruses contain a tegument layer comprising a protein matrix; these proteins play key roles during viral assembly and egress. Here, liquid chromatography and tandem mass spectrometry analysis (LC-MS/MS) of proteins from human herpesvirus 6 (HHV-6)-infected cells revealed a possible association between two major tegument proteins, U14 and U11. This association was verified by immunoprecipitation experiments. Moreover, U11 protein was expressed during the late phase of infection and incorporated into virions. Finally, in contrast to its revertant, a U11 deletion mutant could not be reconstituted. Taken together, these results suggest that HHV-6 U11 is an essential gene for virus growth and propagation.

Cytoplasmic tail domain of glycoprotein B is essential for HHV-6 infection.

Human herpesvirus 6 (HHV-6) glycoprotein B (gB) is an abundantly expressed viral glycoprotein required for viral entry and cell fusion, and is highly conserved among herpesviruses. The present study examined the function of HHV-6 gB cytoplasmic tail domain (CTD). A gB CTD deletion mutant was constructed which, in contrast to its revertant, could not be reconstituted. Moreover, deletion of gB cytoplasmic tail impaired the intracellular transport of gB protein to the trans-Golgi network (TGN). Taken together, these results suggest that gB CTD is critical for HHV-6 propagation and important for intracellular transportation.