A User Interface for Mobile Robotized Tele-echography

Ultrasound imaging allows the evaluation of the degree of emergency of a patient. However, in many situations no experienced sonographer is available to perform such echography. To cope with this issue, the OTELO project “mObile Tele-Echography using an ultra-Light rObot” (OTELO) aims to develop a fully integrated end-to-end mobile tele-echography system using an ultralight, remotely controlled six degree-of-freedom (DOF) robot. In this context, this paper deals with the user interface environment of the OTELO system, composed by the following parts: an ultrasound video transmission system providing real-time images of the scanned area at each moment, an audio/video conference to communicate with the paramedical assistant and the patient, and finally a virtual reality environment, providing visual and haptic feedback to the expert, while capturing the expert's hand movements with a one-DOF hand free input device

Characterization of antibody-mediated neutralization directed against the hypervariable region 1 of hepatitis C virus E2 glycoprotein

The hypervariable region 1 (HVR1) comprising the first 27 aa of E2 glycoprotein is a target for neutralizing antibodies against hepatitis C virus (HCV), but the mechanisms of this neutralization in the cell-culture-infectious genotype 2a strain JFH1 HCV virus (HCVcc) system are unknown. Two rabbit polyclonal sera, R1020 and R140, recognizing the HVR1 of the genotype 1a isolates H77c and Glasgow (Gla), respectively, and a Gla HVR1-specific mouse mAb AP213 have been described previously. However, attempts to generate of antibodies to the JFH1 HVR1 were unsuccessful. Therefore, this study produced chimeric JFH1 HCVcc viruses harbouring the H77c or Gla HVR1 to assess the reactivity of antibodies to this region and their effects on virus infectivity. The inter-genotypic HVR1 swap did not significantly affect virus infectivity. The genotype 1a HVR1-specific antibodies neutralized chimeric viruses in an isolate-dependent manner, underlining the role of HVR1 in HCV infection. The neutralizing antibodies reacted mainly with the C-terminal portion of HVR1, and detailed mapping identified A17, F20 and Q21 in the Gla HVR1 sequence and T21 (and possibly L20) in the corresponding H77c sequence as key epitope residues for AP213 and R140, and R1020, respectively. Importantly, none of the antibodies inhibited in vitro binding of viral envelope glycoproteins to the best-characterized HCV receptor, CD81, or to the glycosaminoglycan attachment factors. However, the HVR1 antibodies were capable of post-attachment neutralization. Overall, this study emphasizes the role of HVR1 in HCVcc entry and provides new tools to study this region further in the context of complete virions

The small compound inhibitor K22 displays broad antiviral activity against different members of the family Flaviviridae and offers potential as pan-viral inhibitor.

The virus family Flaviviridae encompasses several viruses, including (re)emerging viruses which cause widespread morbidity and mortality throughout the world. Members of this virus family are positive-strand RNA viruses and replicate their genome in close association with reorganized intracellular host cell membrane compartments. This evolutionarily conserved strategy facilitates efficient viral genome replication and contributes to evasion from host cell cytosolic defense mechanisms. We have previously described the identification of a small-compound inhibitor, K22, which exerts a potent antiviral activity against a broad range of coronaviruses by targeting membrane-bound viral RNA replication. To analyze the antiviral spectrum of this inhibitor, we assessed the inhibitory potential of K22 against several members of the Flaviviridae family, including the reemerging Zika virus (ZIKV). We show that ZIKV is strongly affected by K22. Time-of-addition experiments revealed that K22 acts during a postentry phase of the ZIKV life cycle, and combination regimens of K22 together with ribavirin (RBV) or interferon alpha (IFN-α) further increased the extent of viral inhibition. Ultrastructural electron microscopy studies revealed severe alterations of ZIKV-induced intracellular replication compartments upon infection of K22-treated cells. Importantly, the antiviral activity of K22 was demonstrated against several other members of the Flaviviridae family. It is tempting to speculate that K22 exerts its broad antiviral activity against several positive-strand RNA viruses via a similar mechanism and thereby represents an attractive candidate for development as a panviral inhibitor

Cell Culture Replication of a Genotype 1b Hepatitis C Virus Isolate Cloned from a Patient Who Underwent Liver Transplantation

The introduction of the genotype 2a isolate JFH1 was a major breakthrough in the field of hepatitis C virus (HCV), allowing researchers to study the complete life cycle of the virus in cell culture. However, fully competent culture systems encompassing the most therapeutically relevant HCV genotypes are still lacking, especially for the highly drug-resistant genotype 1b. For most isolated HCV clones, efficient replication in cultured hepatoma cells requires the introduction of replication-enhancing mutations. However, such mutations may interfere with viral assembly, as occurs in the case of the genotype 1b isolate Con1. In this study, we show that a clinical serum carrying a genotype 1b virus with an exceptionally high viral load was able to infect Huh7.5 cells. Similar to previous reports, inoculation of Huh7.5 cells by natural virus is very inefficient compared to infection by cell culture HCV. A consensus sequence of a new genotype 1b HCV isolate was cloned from the clinical serum (designated Barcelona HCV1), and then subjected to replication studies. This virus replicated poorly in a transient fashion in Huh7.5 cells after electroporation with in vitro transcribed RNA. Nonetheless, approximately 3 weeks post electroporation and thereafter, core protein-positive cells were detected by immunofluorescence. Surprisingly, small amounts of core protein were also measurable in the supernatant of electroporated cells, suggesting that HCV particles might be assembled and released. Our findings not only enhance the current method of cloning in vitro HCV replication-competent isolates, but also offer valuable insights for the realization of fully competent culture systems for HCV

NS2 Protein of Hepatitis C Virus Interacts with Structural and Non-Structural Proteins towards Virus Assembly

Growing experimental evidence indicates that, in addition to the physical virion components, the non-structural proteins of hepatitis C virus (HCV) are intimately involved in orchestrating morphogenesis. Since it is dispensable for HCV RNA replication, the non-structural viral protein NS2 is suggested to play a central role in HCV particle assembly. However, despite genetic evidences, we have almost no understanding about NS2 protein-protein interactions and their role in the production of infectious particles. Here, we used co-immunoprecipitation and/or fluorescence resonance energy transfer with fluorescence lifetime imaging microscopy analyses to study the interactions between NS2 and the viroporin p7 and the HCV glycoprotein E2. In addition, we used alanine scanning insertion mutagenesis as well as other mutations in the context of an infectious virus to investigate the functional role of NS2 in HCV assembly. Finally, the subcellular localization of NS2 and several mutants was analyzed by confocal microscopy. Our data demonstrate molecular interactions between NS2 and p7 and E2. Furthermore, we show that, in the context of an infectious virus, NS2 accumulates over time in endoplasmic reticulum-derived dotted structures and colocalizes with both the envelope glycoproteins and components of the replication complex in close proximity to the HCV core protein and lipid droplets, a location that has been shown to be essential for virus assembly. We show that NS2 transmembrane region is crucial for both E2 interaction and subcellular localization. Moreover, specific mutations in core, envelope proteins, p7 and NS5A reported to abolish viral assembly changed the subcellular localization of NS2 protein. Together, these observations indicate that NS2 protein attracts the envelope proteins at the assembly site and it crosstalks with non-structural proteins for virus assembly

HCV requires a tight junction-associated protein for cell entry

Evaluation of: Evans MJ, von Hahn T, Tscherne DM et al.: Claudin-1 is a hepatitis C virus co-receptor required for a late step in entry. Nature 446(7137), 801–805 (2007). Viruses use surface molecules on target cells as receptors to gain entry and initiate infection. Unraveling the mechanisms of the entry process is crucial for understanding the virus life cycle and pathogenesis, and for developing novel intervention strategies. HCV enters cells by endocytosis following binding to receptors via its envelope glycoproteins, E1 and E2. Several cell receptors have been proposed to be involved in this process, but these are not sufficient for productive entry, indicating that one or more entry factors are yet to be discovered. Toward this, Evans and colleagues identified claudin-1, a member of a family of transmembrane proteins that are major constituents of tight junctions between cells, as essential for HCV entry. They demonstrated that claudin-1 is a coreceptor involved in a late stage of virus entry, and identified amino acid residues located in its first external loop as critical for this process. Their findings suggest that claudin-1 does not determine species host range, but it may be a determinant of HCV tissue tropism. The discovery represents a major step forward in understanding the HCV entry mechanism and pathogenesis, and offers a novel target for future development of antiviral therapy

Lipid droplet contact sites in health and disease.

After having been disregarded for a long time as inert fat drops, lipid droplets (LDs) are now recognized as ubiquitous cellular organelles with key functions in lipid biology and beyond. The identification of abundant LD contact sites, places at which LDs are physically attached to other organelles, has uncovered an unexpected level of communication between LDs and the rest of the cell. In recent years, many disease factors mutated in hereditary disorders have been recognized as LD contact site proteins. Furthermore, LD contact sites are dramatically rearranged in response to infections with intracellular pathogens, as well as under pathological metabolic conditions such as hepatic steatosis. Collectively, it is emerging that LD–organelle contacts are important players in development and progression of disease

Rapid synchronization of hepatitis C virus infection by magnetic adsorption

Hepatitis C virus (HCV) entry into target cells is thought to be a multistep process involving several cellular factors. However, their precise role during virus entry is unclear. investigation of the mechanisms of HCV entry, such as the order of intervention by the cellular receptors, requires synchronizing infections. This study describes a new method involving magnetic adsorption of virus to nanoparticles to synchronize infection, which can be adapted to both HCV pseudoparticles and cell culture infectious HCV. By combining these particles with negatively or positively charged magnetic nanoparticles it was possible to adsorb them onto target cells under a magnetic field in only 2 min. This resulted in greater efficiency of virus adsorption to cells, and increased the infectivity of cell culture infectious virus, as compared to the standard protocol involving incubation of the virus with cells at 4°C for 1 h, or to a standard infection at 37°C. Furthermore, magnetic adsorption respected the natural entry route of the virus, making this system suitable to study the early stages of HCV infection

Robotic Systems in Current Clinical Practice

Medical robotic systems are successfully employed in various surgical specialties today. Yet, a substantial number of remarkable systems that have been developed and piloted, have failed to reach commercialization and thus adoption in clinical practice. This is partly due to the strict regulatory requirements, which typically occupy a significant amount of the development time while incurring additional costs. Pertinent to regulatory approvals is the field of Human Factors, which plays a central role in the design of safe and efficient medical devices. This study briefly introduces the FDA regulatory approval process, discusses the role of human factors in the design process and highlights specific robotic systems that have obtained approval for clinical use. The purpose is to show the status of robotic technologies in relation to the current clinical practice