Multifaceted activation of STING axis upon Nipah and measles virus-induced syncytia formation.

Activation of the DNA-sensing STING axis by RNA viruses plays a role in antiviral response through mechanisms that remain poorly understood. Here, we show that the STING pathway regulates Nipah virus (NiV) replication in vivo in mice. Moreover, we demonstrate that following both NiV and measles virus (MeV) infection, IFNγ-inducible protein 16 (IFI16), an alternative DNA sensor in addition to cGAS, induces the activation of STING, leading to the phosphorylation of NF-κB p65 and the production of IFNß and interleukin 6. Finally, we found that paramyxovirus-induced syncytia formation is responsible for loss of mitochondrial membrane potential and leakage of mitochondrial DNA in the cytoplasm, the latter of which is further detected by both cGAS and IFI16. These results contribute to improve our understanding about NiV and MeV immunopathogenesis and provide potential paths for alternative therapeutic strategies.

Control of Nipah Virus Infection in Mice by the Host Adaptors Mitochondrial Antiviral Signaling Protein (MAVS) and Myeloid Differentiation Primary Response 88 (MyD88).

Interferon (IFN) type I plays a critical role in the protection of mice from lethal Nipah virus (NiV) infection, but mechanisms responsible for IFN-I induction remain unknown. In the current study, we demonstrated the critical role of the mitochondrial antiviral signaling protein signaling pathway in IFN-I production and NiV replication in murine embryonic fibroblasts in vitro, and the redundant but essential roles of both mitochondrial antiviral signaling protein and myeloid differentiation primary response 88 adaptors, but not toll/interleukin-1 receptor/resistance [TIR] domain-containing adaptor-inducing IFN-ß (TRIF), in the control of NiV infection in mice. These results reveal potential novel targets for antiviral intervention and help in understanding NiV immunopathogenesis.

High Pathogenicity of Nipah Virus from Pteropus lylei Fruit Bats, Cambodia.

We conducted an in-depth characterization of the Nipah virus (NiV) isolate previously obtained from a Pteropus lylei bat in Cambodia in 2003 (CSUR381). We performed full-genome sequencing and phylogenetic analyses and confirmed CSUR381 is part of the NiV-Malaysia genotype. In vitro studies revealed similar cell permissiveness and replication of CSUR381 (compared with 2 other NiV isolates) in both bat and human cell lines. Sequence alignments indicated conservation of the ephrin-B2 and ephrin-B3 receptor binding sites, the glycosylation site on the G attachment protein, as well as the editing site in phosphoprotein, suggesting production of nonstructural proteins V and W, known to counteract the host innate immunity. In the hamster animal model, CSUR381 induced lethal infections. Altogether, these data suggest that the Cambodia bat-derived NiV isolate has high pathogenic potential and, thus, provide insight for further studies and better risk assessment for future NiV outbreaks in Southeast Asia.

Measles Virus Bearing Measles Inclusion Body Encephalitis-Derived Fusion Protein Is Pathogenic after Infection via the Respiratory Route.

A clinical isolate of measles virus (MeV) bearing a single amino acid alteration in the viral fusion protein (F; L454W) was previously identified in two patients with lethal sequelae of MeV central nervous system (CNS) infection. The mutation dysregulated the viral fusion machinery so that the mutated F protein mediated cell fusion in the absence of known MeV cellular receptors. While this virus could feasibly have arisen via intrahost evolution of the wild-type (wt) virus, it was recently shown that the same mutation emerged under the selective pressure of small-molecule antiviral treatment. Under these conditions, a potentially neuropathogenic variant emerged outside the CNS. While CNS adaptation of MeV was thought to generate viruses that are less fit for interhost spread, we show that two animal models can be readily infected with CNS-adapted MeV via the respiratory route. Despite bearing a fusion protein that is less stable at 37°C than the wt MeV F, this virus infects and replicates in cotton rat lung tissue more efficiently than the wt virus and is lethal in a suckling mouse model of MeV encephalitis even with a lower inoculum. Thus, either during lethal MeV CNS infection or during antiviral treatment in vitro, neuropathogenic MeV can emerge, can infect new hosts via the respiratory route, and is more pathogenic (at least in these animal models) than wt MeV.IMPORTANCE Measles virus (MeV) infection can be severe in immunocompromised individuals and lead to complications, including measles inclusion body encephalitis (MIBE). In some cases, MeV persistence and subacute sclerosing panencephalitis (SSPE) occur even in the face of an intact immune response. While they are relatively rare complications of MeV infection, MIBE and SSPE are lethal. This work addresses the hypothesis that despite a dysregulated viral fusion complex, central nervous system (CNS)-adapted measles virus can spread outside the CNS within an infected host.

DNA-based probes for flow cytometry analysis of endocytosis and recycling.

The internalization of proteins plays a key role in cell development, cell signaling and immunity. We have previously developed a specific hybridization internalization probe (SHIP) to quantitate the internalization of proteins and particles into cells. Herein, we extend the utility of SHIP to examine both the endocytosis and recycling of surface receptors using flow cytometry. SHIP was used to monitor endocytosis of membrane-bound transferrin receptor (TFR) and its soluble ligand transferrin (TF). SHIP enabled measurements of the proportion of surface molecules internalized, the internalization kinetics and the proportion and rate of internalized molecules that recycle to the cell surface with time. Using this method, we have demonstrated the internalization and recycling of holo-TF and an antibody against the TFR behave differently. This assay therefore highlights the implications of receptor internalization and recycling, where the internalization of the receptor-antibody complex behaves differently to the receptor-ligand complex. In addition, we observe distinct internalization patterns for these molecules expressed by different subpopulations of primary cells. SHIP provides a convenient and high throughput technique for analysis of trafficking parameters for both cell surface receptors and their ligands.

Criteria for dendritic cell receptor selection for efficient antibody-targeted vaccination.

Ab-targeted vaccination involves targeting a receptor of choice expressed by dendritic cells (DCs) with Ag-coupled Abs. Currently, there is little consensus as to which criteria determine receptor selection to ensure superior Ag presentation and immunity. In this study, we investigated parameters of DC receptor internalization and determined how they impact Ag presentation outcomes. First, using mixed bone marrow chimeras, we established that Ag-targeted, but not nontargeted, DCs are responsible for Ag presentation in settings of Ab-targeted vaccination in vivo. Next, we analyzed parameters of DEC205 (CD205), Clec9A, CD11c, CD11b, and CD40 endocytosis and obtained quantitative measurements of internalization speed, surface turnover, and delivered Ag load. Exploiting these parameters in MHC class I (MHC I) and MHC class II (MHC II) Ag presentation assays, we showed that receptor expression level, proportion of surface turnover, or speed of receptor internalization did not impact MHC I or MHC II Ag presentation efficiency. Furthermore, the Ag load delivered to DCs did not correlate with the efficiency of MHC I or MHC II Ag presentation. In contrast, targeting Ag to CD8(+) or CD8(-) DCs enhanced MHC I or MHC II Ag presentation, respectively. Therefore, receptor expression levels, speed of internalization, and/or the amount of Ag delivered can be excluded as major determinants that dictate Ag presentation efficiency in setting of Ab-targeted vaccination.

[Home safety and severe mental disorders: Developing an evaluation tool]. / Sécurité a domicile et troubles mentaux graves: elaboration d'un outil d'evaluation.

BACKGROUND: Home safety evaluation is an important issue within the context of current perspectives on accommodation for people with a serious mental illness who favour a more independent way of life. PURPOSE: This paper describes the development and content validation of the Evaluation de la sécurité a domicile et de la gestion des risques (ESGR), an occupational therapy assessment tool for people with a serious mental illness. METHOD: The ESGR was developed from scientific knowledge and clinical experience. Assessing content validity was done in two phases and involved the consultation of 11 experts. FINDINGS: In its current form, the ESGR includes 67 items organized into three categories (environment, occupation, person). IMPLICATIONS: According to the experts consulted, there is a clinical interest in using the ESGR to support occupational therapists in the assessment of home safety for people with serious mental illness. The statements are clear and representative of the concept and the target audience.

Nebulized fusion inhibitory peptide protects cynomolgus macaques from measles virus infection.

Measles is the most contagious airborne viral infection and the leading cause of child death among vaccine-preventable diseases. We show here that aerosolized lipopeptide fusion inhibitors, derived from heptad-repeat regions of the measles virus (MeV) fusion protein, block respiratory MeV infection in a non-human primate model, the cynomolgus macaque. We used a custom-designed mesh nebulizer to ensure efficient aerosol delivery of peptides to the respiratory tract and demonstrated the absence of adverse effects and lung pathology in macaques. The nebulized peptide efficiently prevented MeV infection, resulting in the complete absence of MeV RNA, MeV-infected cells, and MeV-specific humoral responses in treated animals. This strategy provides an additional shield which complements vaccination to fight against respiratory infection, presenting a proof-of-concept for the aerosol delivery of fusion inhibitory peptides to protect against measles and other airborne viruses, including SARS-CoV-2, in case of high-risk exposure, that can be readily translated to human trials.

Nebulized fusion inhibitory peptide protects cynomolgus macaques from measles virus infection.

Measles is the most contagious airborne viral infection and the leading cause of child death among vaccine-preventable diseases. We show here that aerosolized lipopeptide fusion inhibitor, derived from heptad-repeat regions of the measles virus (MeV) fusion protein, blocks respiratory MeV infection in a non-human primate model, the cynomolgus macaque. We use a custom-designed mesh nebulizer to ensure efficient aerosol delivery of peptide to the respiratory tract and demonstrate the absence of adverse effects and lung pathology in macaques. The nebulized peptide efficiently prevents MeV infection, resulting in the complete absence of MeV RNA, MeV-infected cells, and MeV-specific humoral responses in treated animals. This strategy provides an additional means to fight against respiratory infection in non-vaccinated people, that can be readily translated to human trials. It presents a proof-of-concept for the aerosol delivery of fusion inhibitory peptides to protect against measles and other airborne viruses, including SARS-CoV-2, in case of high-risk exposure.

Reprogrammed Pteropus Bat Stem Cells as A Model to Study Host-Pathogen Interaction during Henipavirus Infection.

Bats are natural hosts for numerous zoonotic viruses, including henipaviruses, which are highly pathogenic for humans, livestock, and other mammals but do not induce clinical disease in bats. Pteropus bats are identified as a reservoir of henipaviruses and the source of transmission of the infection to humans over the past 20 years. A better understanding of the molecular and cellular mechanisms allowing bats to control viral infections requires the development of relevant, stable, and permissive cellular experimental models. By applying a somatic reprogramming protocol to Pteropus bat primary cells, using a combination of ESRRB (Estrogen Related Receptor Beta), CDX2 (Caudal type Homeobox 2), and c-MYC (MYC proto-oncogene) transcription factors, we generated bat reprogrammed cells. These cells exhibit stem cell-like characteristics and neural stem cell molecular signature. In contrast to primary fibroblastic cells, these reprogrammed stem cells are highly permissive to henipaviruses and exhibit specific transcriptomic profiles with the particular expression of certain susceptibility factors such as interferon-stimulated genes (ISG), which may be related to viral infection. These Pteropus bat reprogrammed stem cells should represent an important experimental tool to decipher interactions during henipaviruses infection in Pteropus bats, facilitate isolation and production of bat-borne viruses, and to better understand the bat biology.

Biochemical and cellular characterisation of the Plasmodium falciparum M1 alanyl aminopeptidase (PfM1AAP) and M17 leucyl aminopeptidase (PfM17LAP).

The Plasmodium falciparum M1 alanyl aminopeptidase and M17 leucyl aminopeptidase, PfM1AAP and PfM17LAP, are potential targets for novel anti-malarial drug development. Inhibitors of these aminopeptidases have been shown to kill malaria parasites in culture and reduce parasite growth in murine models. The two enzymes may function in the terminal stages of haemoglobin digestion, providing free amino acids for protein synthesis by the rapidly growing intra-erythrocytic parasites. Here we have performed a comparative cellular and biochemical characterisation of the two enzymes. Cell fractionation and immunolocalisation studies reveal that both enzymes are associated with the soluble cytosolic fraction of the parasite, with no evidence that they are present within other compartments, such as the digestive vacuole (DV). Enzyme kinetic studies show that the optimal pH of both enzymes is in the neutral range (pH 7.0-8.0), although PfM1AAP also possesses some activity (< 20%) at the lower pH range of 5.0-5.5. The data supports the proposal that PfM1AAP and PfM17LAP function in the cytoplasm of the parasite, likely in the degradation of haemoglobin-derived peptides generated in the DV and transported to the cytosol.

Activation of cGAS/STING pathway upon paramyxovirus infection.

During inflammatory diseases, cancer, and infection, the cGAS/STING pathway is known to recognize foreign or self-DNA in the cytosol and activate an innate immune response. Here, we report that negative-strand RNA paramyxoviruses, Nipah virus (NiV), and measles virus (MeV), can also trigger the cGAS/STING axis. Although mice deficient for MyD88, TRIF, and MAVS still moderately control NiV infection when compared with wild-type mice, additional STING deficiency resulted in 100% lethality, suggesting synergistic roles of these pathways in host protection. Moreover, deletion of cGAS or STING resulted in decreased type I interferon production with enhanced paramyxoviral infection in both human and murine cells. Finally, the phosphorylation and ubiquitination of STING, observed during viral infections, confirmed the activation of cGAS/STING pathway by NiV and MeV. Our data suggest that cGAS/STING activation is critical in controlling paramyxovirus infection and possibly represents attractive targets to develop countermeasures against severe disease induced by these pathogens.

Nipah virus W protein harnesses nuclear 14-3-3 to inhibit NF-κB-induced proinflammatory response.

Nipah virus (NiV) is a highly pathogenic emerging bat-borne Henipavirus that has caused numerous outbreaks with public health concerns. It is able to inhibit the host innate immune response. Since the NF-κB pathway plays a crucial role in the innate antiviral response as a major transcriptional regulator of inflammation, we postulated its implication in the still poorly understood NiV immunopathogenesis. We report here that NiV inhibits the canonical NF-κB pathway via its nonstructural W protein. Translocation of the W protein into the nucleus causes nuclear accumulation of the cellular scaffold protein 14-3-3 in both African green monkey and human cells infected by NiV. Excess of 14-3-3 in the nucleus was associated with a reduction of NF-κB p65 subunit phosphorylation and of its nuclear accumulation. Importantly, W-S449A substitution impairs the binding of the W protein to 14-3-3 and the subsequent suppression of NF-κB signaling, thus restoring the production of proinflammatory cytokines. Our data suggest that the W protein increases the steady-state level of 14-3-3 in the nucleus and consequently enhances 14-3-3-mediated negative feedback on the NF-κB pathway. These findings provide a mechanistic model of W-mediated disruption of the host inflammatory response, which could contribute to the high severity of NiV infection.

Validation of the revised child version of the Assessment of Computer Task Performance.

OBJECTIVE: The purpose of this study was to examine test-retest reliability, internal consistency, and construct validity of the revised child version of the Assessment of Computer Task Performance. The subjects were 155 children of 5 to 11 years of age who did not have any visual or motor impairment. Children 5 through 9 years of age were administered the revised test, which included some additional tasks. Children 10 and 11 years of age were administered selected tasks from the adult version. A subgroup of 52 children were administered the test for the second time to examine test-retest reliability. Intra-class correlations for the mean time to complete two trials indicated moderate to good test-retest reliability (0.66-0.91). The alpha coefficient for tasks administered to children 5 through 9 years of age (0.85) supports internal consistency. Removal of one task increased the alpha coefficient to 0.79 for tasks administered to children 10 and 11 years of age. Three factors explained 84% of the total variance in scores of children 5-9 years of age and 75% of the variance in scores of children 10 and 11 years of age. The mean time to complete keyboard tasks decreased with age and the mean time to mouse tasks decreased between 5 and 9 years. CONCLUSION: The results provide evidence of test-retest reliability, internal consistency, and construct validity of the revised Assessment of Computer task Performance.

Pre- and postoperative tricuspid regurgitation in patients with severe symptomatic aortic stenosis: importance of pre-operative tricuspid annulus diameter.

Aims: Secondary tricuspid regurgitation (STR) is commonly found in patients with aortic stenosis and is associated with increased morbidity. The study sought to evaluate the prevalence of pre-operative STR and its progression after surgical aortic valve replacement (SAVR) or transcatheter aortic valve implantation (TAVI). Also, it sought to analyse the predictors of post-operative changes in STR. Methods and results: We prospectively evaluated 116 patients (aged 75.1 ± 9.8 years, predominantly male) who undergo SAVR or TAVI for severe aortic stenosis (AS) from September 2013 to April 2015. Patients with associated valve disease requiring intervention, significant coronary artery disease or left ventricular ejection fraction (LVEF) <50% were excluded. Clinical and echocardiographic data, including TR grade and right ventricular (RV) size and function, were assessed at baseline and at the 1-year follow-up. At baseline, significant TR was documented in 13 patients (11.1%) and non-significant TR was documented in 103 patients (88.9%). Atrial fibrillation (AF) was more prevalent in patients with a tricuspid annulus diameter ≥40 mm (P < 0.0051). At the 1-year follow-up, the TR grade had improved in 17 patients (14.7%), was unchanged in 68 patients (58.6%) and had worsened in 31 patients (26.7%). Moderate to severe TR was found in 30 patients (25.8%). Tricuspid annulus diameter >40 mm was the only echocardiographic predictor of significant postoperative TR (relative risk (RR) = 2.12 [1.26-3.54], P = 0.004). Right heart function and size were not independent predictors. Conclusion: Significant TR was present pre-operatively in 11.1% of patients. Post-operative progression was observed in 26.7% of patients. Only tricuspid annulus size >40 mm was an independent echocardiographic predictor of moderate to severe TR at the 1-year follow-up.

[The adaptation process of adults who sustained craniocerebral trauma]. / Le processus d'adaptation des adultes présentant des séquelles de traumatisme cranio-cérébral.

BACKGROUND: The adaptation process of adults who had sustained a traumatic brain injury is not well understood. PURPOSE: This qualitative study explores the adaptation process of adults presenting the sequelae of a traumatic brain injury. METHOD: Fifty-three community-dwelling adults with the sequelae of traumatic brain injury participated in an individual interview and answered open-ended questions. RESULTS: The qualitative analysis, inspired by the phenomenological tradition, reports an adaptation process in six stages. The elements which influenced this process as well as the effective coping strategies according to the participant's perception are described. The adaptation process of this clientele is affected primarily by the nature of the cognitive sequelae, as well as the circumstances of the traumatic event. CLINICAL IMPLICATIONS: The realization of significant occupations is one of the key elements of this process, sometimes associated to an important interior transformation from the point of view of personal standards and values.

Antibody-mediated targeting of antigen to C-type lectin-like receptors Clec9A and Clec12A elicits different vaccination outcomes.

Targeting antigen (Ag) to dendritic cell (DC) surface receptors is a potential new mode of vaccination. C-type lectin-like receptors Clec9A and Clec12A are attractive receptor targets however their targeting in vivo elicits significantly different outcomes for unknown reasons. To gain insight into the mechanisms responsible, we have examined the intrinsic capacity of Clec9A and Clec12A to elicit MHC I and MHC II Ag presentation following ex vivo targeting with primary murine DC. Both receptors exhibited high rates of internalization by CD8+ DCs, while Clec12A delivered a significantly higher Ag owing to its higher expression level. Targeting Ag to immature CD8+ DCs via both Clec9A and Clec12A failed to elicit MHC I cross-presentation above that of controls, while Clec12A was the superior receptor to target following CD8+ DC maturation. CD8- DCs were unable to elicit MHC I cross-presentation regardless of the receptor targeted. For MHC II presentation, targeting Ag to Clec12A enabled significant responses by both immature CD8+ and CD8- DCs, whereas Clec9A did not elicit significant MHC II Ag presentation by either DC subset, resting or mature. Therefore, Clec9A and Clec12A exhibit different intrinsic capacities to elicit MHC I and MHC II presentation following direct Ag targeting, though they can only elicit MHC I responses if the DC expressing the receptor is equipped with the capacity to cross-present. Our conclusions have consequences for the exploitation of these receptors for vaccination purposes, in addition to providing insight into their roles as Ag targets in vivo.

Analysis of Intracellular Trafficking of Dendritic Cell Receptors for Antigen Targeting.

Antibody-targeted vaccination aims to efficiently deliver antigen to dendritic cells by targeting specific receptors at the cell surface. The choice of receptor depends on different factors, including their capacity to induce internalization of the delivered antigen/adjuvant cargo. Assays currently used to monitor internalization in dendritic cells have several limitations. We have developed a novel DNA-based probe that allows for simple and robust high-throughput analysis of internalization. Designed for flow cytometry, the probe can also be used for fluorescence microscopy to clearly distinguish internalized from surface-bound material. Here, we describe the steps for modifying material (antibodies, proteins) with the probe, undertaking the assay, and analyzing the data obtained from flow cytometry.

Targeting dendritic cells: a promising strategy to improve vaccine effectiveness.

Dendritic cell (DC) targeting is a novel strategy to enhance vaccination efficacy. This approach is based on the in situ delivery of antigen via antibodies that are specific for endocytic receptors expressed at the surface of DCs. Here we review the complexity of the DC subsets and the antigen presentation pathways that need to be considered in the settings of DC targeting. We also summarize current knowledge about antigen delivery to DCs via DEC-205, Clec9A and Clec12A, receptor targets that strongly enhance cellular and humoral immune responses. Finally, we discuss the intracellular trafficking criteria of the targeted receptors that may impact their effectiveness as DC targets.

[Perceived self-efficacy is associated with social participation in adults with traumatic brain injury]. / La perception d'efficacité personnelle comme facteur associé à la participation sociale des adultes ayant subi un traumatisme cranio-cérébral.

BACKGROUND: Identification of the factors facilitating the social participation of adults who have sustained a traumatic brain injury can help occupational therapists with the direction for their interventions. Earlier studies centered on identifying the socio-demographic characteristics and the disabilities associated with social participation. PURPOSE: The aim of this study was to examine the association between perceived self-efficacy, a positive concept derived from social cognitive theory and social participation. METHODS: A cross-sectional and correlational research design was used with 53 adults who sustained a traumatic brain injury between 1995 and 2000 and lived in their natural environment. Two measuring tools were used: a self-administered questionnaire evaluating the perceived self-efficacy and a questionnaire evaluating social participation, administered by an examiner through a face-to-face interview. RESULTS: The results indicate that the perceived self-efficacy explains 40% of the variance of the social participation. PRACTICE IMPLICATIONS: This association suggests that social cognitive theory can constitute a reference model for occupational therapists working with this clientele.