bluuebunny/crossref_metadata_2025_split huggingface dataset:
An Immortalized Human Dorsal Root Ganglion Cell Line Provides a Novel Context To Study Herpes Simplex Virus 1 Latency and Reactivation
Authors: Thellman Nikki M. , Botting Carolyn , Madaj Zachary , Triezenberg Steven J. 
Published: April 2017

ABSTRACT

A defining characteristic of alphaherpesviruses is the establishment of lifelong latency in host sensory ganglia with occasional reactivation causing recurrent lytic infections. As an alternative to rodent models, we explored the use of an immortalized cell line derived from human dorsal root ganglia. HD10.6 cells proliferate by virtue of a transduced tetracycline-regulated v-
myc
oncogene. In the presence of doxycycline, HD10.6 cells mature to exhibit neuronal morphology and express sensory neuron-associated markers such as neurotrophin receptors TrkA, TrkB, TrkC, and RET and the sensory neurofilament peripherin. Infection of mature HD10.6 neurons by herpes simplex virus 1 (HSV-1) results in a delayed but productive infection. However, infection at a low multiplicity of infection (MOI) in the presence of acyclovir results in a quiescent infection resembling latency in which viral genomes are retained in a low number of neurons, viral gene expression is minimal, and infectious virus is not released. At least some of the quiescent viral genomes retain the capacity to reactivate, resulting in viral DNA replication and release of infectious virus. Reactivation can be induced by depletion of nerve growth factor; other commonly used reactivation stimuli have no significant effect.

IMPORTANCE
Infections by herpes simplex viruses (HSV) cause painful cold sores or genital lesions in many people; less often, they affect the eye or even the brain. After the initial infection, the virus remains inactive or latent in nerve cells that sense the region where that infection occurred. To learn how virus maintains and reactivates from latency, studies are done in neurons taken from rodents or in whole animals to preserve the full context of infection. However, some cellular mechanisms involved in HSV infection in rodents are different from those in humans. We describe the use of a human cell line that has the properties of a sensory neuron. HSV infection in these cultured cells shows the properties expected for a latent infection, including reactivation to produce newly infectious virus. Thus, we now have a cell culture model for latency that is derived from the normal host for this virus.
Autologous Stem Cell Transplantation Disrupts Adaptive Immune Responses during Rebound Simian/Human Immunodeficiency Virus Viremia
Authors: Reeves Daniel B. , Peterson Christopher W. , Kiem Hans-Peter , Schiffer Joshua T. 
Published: April 2017

ABSTRACT
Primary HIV-1 infection induces a virus-specific adaptive/cytolytic immune response that impacts the plasma viral load set point and the rate of progression to AIDS. Combination antiretroviral therapy (cART) suppresses plasma viremia to undetectable levels that rebound upon cART treatment interruption. Following cART withdrawal, the memory component of the virus-specific adaptive immune response may improve viral control compared to primary infection. Here, using primary infection and treatment interruption data from macaques infected with simian/human immunodeficiency virus (SHIV), we observe a lower peak viral load but an unchanged viral set point during viral rebound. The addition of an autologous stem cell transplant before cART withdrawal alters viral dynamics: we found a higher rebound set point but similar peak viral loads compared to the primary infection. Mathematical modeling of the data that accounts for fundamental immune parameters achieves excellent fit to heterogeneous viral loads. Analysis of model output suggests that the rapid memory immune response following treatment interruption does not ultimately lead to better viral containment. Transplantation decreases the durability of the adaptive immune response following cART withdrawal and viral rebound. Our model's results highlight the impact of the endogenous adaptive immune response during primary SHIV infection. Moreover, because we capture adaptive immune memory and the impact of transplantation, this model will provide insight into further studies of cure strategies inspired by the Berlin patient.

IMPORTANCE
HIV patients who interrupt combination antiretroviral therapy (cART) eventually experience viral rebound, the return of viral loads to pretreatment levels. However, the “Berlin patient” remained free of HIV rebound over a decade after stopping cART. His cure is attributed to leukemia treatment that included an HIV-resistant stem cell transplant. Inspired by this case, we studied the impact of stem cell transplantation in a macaque simian/HIV (SHIV) system. Using a mechanistic mathematical model, we found that while primary infection generates an adaptive immune memory response, stem cell transplantation disrupts this learned immunity. The results have implications for HIV cure regimens based on stem cell transplantation.
Membrane Penetration by Bacterial Viruses
Authors: Xu Jingwei , Xiang Ye 
Published: April 2017

ABSTRACT

The bacteriophage ϕ29 infects Gram-positive
Bacillus subtilis
with a short noncontractile tail. Recent studies showed that the ϕ29 tail protein gp9 forms a hexameric tube with six long loops of membrane-active peptides blocking in the tube at the distal end of the tail. The long loops exit on genome release and form a membrane pore for passage of the genome. The membrane penetration mechanism of the ϕ29 tail might be common among tailed bacteriophages.
Genome-Wide Screening Uncovers the Significance of N-Sulfation of Heparan Sulfate as a Host Cell Factor for Chikungunya Virus Infection
Authors: Tanaka Atsushi , Tumkosit Uranan , Nakamura Shota , Motooka Daisuke , Kishishita Natsuko , Priengprom Thongkoon , Sa-ngasang Areerat , Kinoshita Taroh , Takeda Naokazu , Maeda Yusuke 
Published: April 2017

ABSTRACT

The molecular mechanisms underlying chikungunya virus (CHIKV) infection are poorly characterized. In this study, we analyzed the host factors involved in CHIKV infection using genome-wide screening. Human haploid HAP1 cells, into which an exon-trapping vector was introduced, were challenged with a vesicular stomatitis virus pseudotype bearing the CHIKV E3 to E1 envelope proteins. Analysis of genes enriched in the cells resistant to the pseudotyped virus infection unveiled a critical role of N-sulfation of heparan sulfate (HS) for the infectivity of the clinically isolated CHIKV Thai#16856 strain to HAP1 cells. Knockout of NDST1 that catalyzes N-sulfation of HS greatly decreased the binding and infectivity of CHIKV Thai#16856 strain but not infectivity of Japanese encephalitis virus (JEV) and yellow fever virus (YFV). While glycosaminoglycans were commonly required for the efficient infectivity of CHIKV, JEV, and YFV, as shown by using
B3GAT3
knockout cells, the tropism for N-sulfate was specific to CHIKV. Expression of chondroitin sulfate (CS) in
NDST1
-knockout HAP1 cells did not restore the binding of CHIKV Thai#16856 strain and the infectivity of its pseudotype but restored the infectivity of authentic CHIKV Thai#16856, suggesting that CS functions at later steps after CHIKV binding. Among the genes enriched in this screening, we found that TM9SF2 is critical for N-sulfation of HS and therefore for CHIKV infection because it is involved in the proper localization and stability of NDST1. Determination of the significance of and the relevant proteins to N-sulfation of HS may contribute to understanding mechanisms of CHIKV propagation, cell tropism, and pathogenesis.

IMPORTANCE
Recent outbreaks of chikungunya fever have increased its clinical importance. Chikungunya virus (CHIKV) utilizes host glycosaminoglycans to bind efficiently to its target cells. However, the substructure in glycosaminoglycans required for CHIKV infection have not been characterized. Here, we unveil that N-sulfate in heparan sulfate is essential for the efficient infection of a clinical CHIKV strain to HAP1 cells and that chondroitin sulfate does not help the CHIKV binding but does play roles at the later steps in HAP1 cells. We show, by comparing previous reports using Chinese hamster ovary cells, along with another observation that enhanced infectivity of CHIKV bearing Arg82 in envelope E2 does not depend on glycosaminoglycans in HAP1 cells, that the infection manner of CHIKV varies among host cells. We also show that TM9SF2 is required for CHIKV infection to HAP1 cells because it is involved in the N-sulfation of heparan sulfate through ensuring NDST1 activity.
Double Plant Homeodomain Fingers 2 (DPF2) Promotes the Immune Escape of Influenza Virus by Suppressing Beta Interferon Production
Authors: Shin Dongjo , Lee Jihye , Park Ji Hoon , Min Ji-Young 
Published: April 2017

ABSTRACTThe high mutation rates of the influenza virus genome facilitate the generation of viral escape mutants, rendering vaccines and drugs against influenza virus-encoded targets potentially ineffective. Therefore, we identified host cell determinants dispensable for the host but crucial for virus replication, with the goal of preventing viral escape and finding effective antivirals. To identify these host factors, we screened 2,732 human genes using RNA interference and focused on one of the identified host factors, the double plant homeodomain fingers 2 (DPF2/REQ) gene, for this study. We found that knockdown of DPF2 in cells infected with influenza virus resulted in decreased expression of viral proteins and RNA. Furthermore, production of progeny virus was reduced by two logs in the multiple-cycle growth kinetics assay. We also found that DPF2 was involved in the replication of seasonal influenza A and B viruses. Because DPF2 plays a crucial role in the noncanonical NF-κB pathway, which negatively regulates type I interferon (IFN) induction, we examined the relationship between DPF2 and IFN responses during viral infection. The results showed that knockdown of DPF2 resulted in increased expression of IFN-β and induced phosphorylation of STAT1 in infected cells. In addition, high levels of several cytokines/chemokines (interleukin-8 [IL-8], IP-10, and IL-6) and antiviral proteins (MxA and ISG56) were produced by DPF2 knockdown cells. In conclusion, we identified a novel host factor, DPF2, that is required for influenza virus to evade the host immune response and that may serve as a potential antiviral target.IMPORTANCEInfluenza virus is responsible for seasonal epidemics and occasional pandemics and is an ongoing threat to public health worldwide. Influenza virus relies heavily on cellular factors to complete its life cycle. Here we identified a novel host factor, DPF2, which is involved in influenza virus infection. Our results showed that DPF2 plays a crucial role in the replication and propagation of influenza virus. DPF2 functions in the noncanonical NF-κB pathway, which negatively regulates type I IFN induction. Thus, we investigated the relationship between the IFN response and DPF2 in influenza virus infection. Upon influenza virus infection, DPF2 dysregulated IFN-β induction and expression of cytokines/chemokines and antiviral proteins. This study provides evidence that influenza virus utilizes DPF2 to escape host innate immunity.
Characterization of Pre-F-GCN4t, a Modified Human Respiratory Syncytial Virus Fusion Protein Stabilized in a Noncleaved Prefusion Conformation
Authors: Blais Normand , Gagné Martin , Hamuro Yoshitomo , Rheault Patrick , Boyer Martine , Steff Ann-Muriel , Baudoux Guy , Dewar Vincent , Demers Josée , Ruelle Jean-Louis , Martin Denis 
Published: April 2017

ABSTRACT
The human respiratory syncytial virus (hRSV) fusion (F) protein is considered a major target of the neutralizing antibody response to hRSV. This glycoprotein undergoes a major structural shift from the prefusion (pre-F) to the postfusion (post-F) state at the time of virus-host cell membrane fusion. Recent evidences suggest that the pre-F state is a superior target for neutralizing antibodies compared to the post-F state. Therefore, for vaccine purposes, we have designed and characterized a recombinant hRSV F protein, called Pre-F-GCN4t, stabilized in a pre-F conformation. To show that Pre-F-GCN4t does not switch to a post-F conformation, it was compared with a recombinant post-F molecule, called Post-F-XC. Pre-F-GCN4t was glycosylated and trimeric and displayed a conformational stability different from that of Post-F-XC, as shown by chemical denaturation. Electron microscopy analysis suggested that Pre-F-GCN4t adopts a lollipop-like structure. In contrast, Post-F-XC had a typical elongated conical shape. Hydrogen/deuterium exchange mass spectrometry demonstrated that the two molecules had common rigid folding core and dynamic regions and provided structural insight for their biophysical and biochemical properties and reactivity. Pre-F-GCN4t was shown to deplete hRSV-neutralizing antibodies from human serum more efficiently than Post-F-XC. Importantly, Pre-F-GCN4t was also shown to bind D25, a highly potent monoclonal antibody specific for the pre-F conformation. In conclusion, this construct presents several pre-F characteristics, does not switch to the post-F conformation, and presents antigenic features required for a protective neutralizing antibody response. Therefore, Pre-F-GCN4t can be considered a promising candidate vaccine antigen.

IMPORTANCE
Human respiratory syncytial virus (RSV) is a global leading cause of infant mortality and adult morbidity. The development of a safe and efficacious RSV vaccine remains an important goal. The RSV class I fusion (F) glycoprotein is considered one of the most promising vaccine candidates, and recent evidences suggest that the prefusion (pre-F) state is a superior target for neutralizing antibodies. Our study presents the physicochemical characterization of Pre-F-GCN4t, a molecule designed to be stabilized in the pre-F conformation. To confirm its pre-F conformation, Pre-F-GCN4t was analyzed in parallel with Post-F-XC, a molecule in the post-F conformation. Our results show that Pre-F-GCN4t presents characteristics of a stabilized pre-F conformation and support its use as an RSV vaccine antigen. Such an antigen may represent a significant advance in the development of an RSV vaccine.
Hand exercises for patients with rheumatoid arthritis: an extended follow-up of the SARAH randomised controlled trial
Authors: Williamson Esther , McConkey Christopher , Heine Peter , Dosanjh Sukhdeep , Williams Mark , Lamb Sarah E 
Published: April 2017

ObjectivesThe Stretching And strengthening for Rheumatoid Arthritis of the Hand (SARAH) randomised controlled trial evaluated the effectiveness of a hand exercise programme and demonstrated it was clinically effective and cost-effective at 12 months. The aim of this extended follow-up was to evaluate the effects of the SARAH programme beyond 12 months.MethodsUsing postal questionnaires, we collected the Michigan Hand Questionnaire hand function (primary outcome), activities of daily living and work subscales, pain troublesomeness, self-efficacy and health-related quality of life. All participants were asked how often they performed hand exercises for their rheumatoid arthritis. Mean difference in hand function scores were analysed by a linear model, adjusted for baseline score.ResultsTwo-thirds (n=328/490, 67%) of the original cohort provided data for the extended follow-up. The mean follow-up time was 26 months (range 19–40 months).There was no difference in change in hand function scores between the two groups at extended follow-up (mean difference (95% CI) 1.52 (−1.71 to 4.76)). However, exercise group participants were still significantly improved compared with baseline (p=0.0014) unlike the best practice usual care group (p=0.1122). Self-reported performance of hand exercises had reduced substantially.ConclusionsParticipants undertaking the SARAH exercise programme had improved hand function compared with baseline >2 years after randomisation. This was not the case for the control group. However, scores were no longer statistically different between the groups indicating the effect of the programme had diminished over time. This reduction in hand function compared with earlier follow-up points coincided with a reduction in self-reported performance of hand exercises. Further intervention to promote long-term adherence may be warranted.Trial registration numberISRCTN89936343; Results.
Hypoxia-inducible factor 1α predicts recurrence in high-grade soft tissue sarcoma of extremities and trunk wall
Authors: Nyström H , Jönsson M , Werner-Hartman L , Nilbert M , Carneiro A 
Published: April 2017

Background and aimSarcomas are of mesenchymal origin and typically show abundant tumour stroma and presence of necrosis. In search for novel biomarkers for personalised therapy, we determined the prognostic impact of stromal markers, hypoxia and neovascularity in high-grade soft tissue leiomyosarcoma and pleomorphic undifferentiated sarcoma.MethodWe evaluated CD163, colony-stimulating factor (CSF)-1, CD16 and hypoxia-inducible factor 1 (HIF-1)α using immunohistochemical staining and assessed microvessel density using CD31 in 73 high-grade leiomyosarcomas and undifferentiated pleomorphic sarcomas of the extremities and the trunk wall. The results were correlated to metastasis-free and overall survival.ResultsExpression of HIF-1α was associated with the presence of necrosis and independently predicted shorter metastasis-free survival (HR 3.2, CI 1.4 to 7.0, p=0.004), whereas neither expression of the stromal markers CD163, CD16 and CSF-1 nor microvessel density was prognostically relevant in this series.ConclusionsThere is increasing evidence for the prognostic role of hypoxia in high-grade soft tissue sarcoma, and these data suggest that HIF-1α expression represents a candidate prognostic biomarker for clinical application in high-grade leiomyosarcoma and undifferentiated pleomorphic sarcoma.
Human-derived gut microbiota modulates colonic secretion in mice by regulating 5-HT3 receptor expression via acetate production
Authors: Bhattarai Yogesh , Schmidt Bradley A. , Linden David R. , Larson Eric D. , Grover Madhusudan , Beyder Arthur , Farrugia Gianrico , Kashyap Purna C. 
Published: April 2017

Serotonin [5-hydroxytryptamine (5-HT)], an important neurotransmitter and a paracrine messenger in the gastrointestinal tract, regulates intestinal secretion by its action primarily on 5-HT3 and 5-HT4 receptors. Recent studies highlight the role of gut microbiota in 5-HT biosynthesis. In this study, we determine whether human-derived gut microbiota affects host secretory response to 5-HT and 5-HT receptor expression. We used proximal colonic mucosa-submucosa preparation from age-matched Swiss Webster germ-free (GF) and humanized (HM; ex-GF colonized with human gut microbiota) mice. 5-HT evoked a significantly greater increase in short-circuit current (Δ Isc) in GF compared with HM mice. Additionally, 5-HT3 receptor mRNA and protein expression was significantly higher in GF compared with HM mice. Ondansetron, a 5-HT3 receptor antagonist, inhibited 5-HT-evoked Δ Isc in GF mice but not in HM mice. Furthermore, a 5-HT3 receptor-selective agonist, 2-methyl-5-hydroxytryptamine hydrochloride, evoked a significantly higher Δ Isc in GF compared with HM mice. Immunohistochemistry in 5-HT3A-green fluorescent protein mice localized 5-HT3 receptor expression to enterochromaffin cells in addition to nerve fibers. The significant difference in 5-HT-evoked Δ Isc between GF and HM mice persisted in the presence of tetrodotoxin (TTX) but was lost after ondansetron application in the presence of TTX. Application of acetate (10 mM) significantly lowered 5-HT3 receptor mRNA in GF mouse colonoids. We conclude that host secretory response to 5-HT may be modulated by gut microbiota regulation of 5-HT3 receptor expression via acetate production. Epithelial 5-HT3 receptor may function as a mediator of gut microbiota-driven change in intestinal secretion. NEW & NOTEWORTHY We found that gut microbiota alters serotonin (5-HT)-evoked intestinal secretion in a 5-HT3 receptor-dependent mechanism and gut microbiota metabolite acetate alters 5-HT3 receptor expression in colonoids. View this article's corresponding video summary at https://www.youtube.com/watch?v=aOMYJMuLTcw&feature=youtu.be .
Glycolytic inhibition by 2-deoxy-d-glucose abolishes both neuronal and network bursts in an in vitro seizure model
Authors: Shao Li-Rong , Stafstrom Carl E. 
Published: April 2017

Neuronal activity is energy demanding and coupled to cellular metabolism. In this study, we investigated the effects of glycolytic inhibition with 2-deoxy-d-glucose (2-DG) on basal membrane properties, spontaneous neuronal firing, and epileptiform network bursts in hippocampal slices. The effect of glycolytic inhibition on basal membrane properties was examined in hippocampal CA1 neurons, which are not ordinarily active spontaneously. Intracellular application of 2-DG did not significantly alter the membrane input resistance, action-potential threshold, firing pattern, or input-output relationship of these neurons compared with simultaneously recorded neighboring neurons without intracellular 2-DG. The effect of glycolytic inhibition on neuronal firing was tested in spontaneously active hippocampal neurons (CA3) when synaptic transmission was left intact or blocked with AMPA, NMDA, and GABAA receptor antagonists (DNQX, APV, and bicuculline, respectively). Under both conditions (synaptic activity intact or blocked), bath application of 2-DG (2 mM) blocked spontaneous firing in ~2/3 (67 and 71%, respectively) of CA3 pyramidal neurons. In contrast, neuronal firing of CA3 neurons persisted when 2-DG was applied intracellularly, suggesting that glycolytic inhibition of individual neurons is not sufficient to stop neuronal firing. The effects of 2-DG on epileptiform network bursts in area CA3 were tested in Mg2+-free medium containing 50 µM 4-aminopyridine. Bath application of 2-DG abolished these epileptiform bursts in a dose-dependent and all-or-none manner. Taken together, these data suggest that altered glucose metabolism profoundly affects cellular and network hyperexcitability and that glycolytic inhibition by 2-DG can effectively abrogate epileptiform activity. NEW & NOTEWORTHY Neuronal activity is highly energy demanding and coupled to cellular metabolism. In this study, we demonstrate that glycolytic inhibition with 2-deoxy-d-glucose (2-DG) effectively suppresses spontaneous neuronal firing and epileptiform bursts in hippocampal slices. These data suggest that an altered metabolic state can profoundly affect cellular and network excitability, and that the glycolytic inhibitor 2-DG may hold promise as a novel treatment of drug-resistant epilepsy.