Distinct CD4+CD8+ Double-Positive T Cells in the Blood and Liver of Patients during Chronic Hepatitis B and C

CD4+ and CD8+ T cells, the main effectors of adaptive cellular immune responses, differentiate from immature, non-functional CD4+CD8+ double-positive T (DPT) cells in the thymus. Increased proportions of circulating DPT lymphocytes have been observed during acute viral infections; in chronic viral diseases, the role and repartition of extra-thymic DPT cells remain largely uncharacterized. We performed a phenotypic analysis of DPT cells in blood and liver from patients chronically infected by hepatitis C (HCV) or B (HBV) viruses. The highest percentages of DPT cells, predominantly CD4highCD8low, were observed in patients infected by HCV, while HBV-infected patients mostly displayed CD4lowCD8high and CD4highCD8high DPT cells. All proportions of DPT cells were higher in liver than in blood with, for each subpopulation referred to above, a correlation between their frequencies in these two compartments. In HCV patients, intra-hepatic DPT cells displayed more heterogeneous activation, differentiation and memory phenotypes than in the blood; most of them expressed CD1a, a marker of T cell development in the thymus. Ex vivo, the inoculation of liver slices with HCV produced in cell culture was accompanied by a disappearance of CD8high cells, suggesting a direct effect of the virus on the phenotype of DPT cells in the liver. Our results suggest that, in half of the patients, chronic HCV infection promotes the production of DPT cells, perhaps by their re-induction in the thymus and selection in the liver

An Equine Model for Vaccination against a Hepacivirus: Insights into Host Responses to E2 Recombinant Protein Vaccination and Subsequent Equine Hepacivirus Inoculation

Equine hepacivirus (EqHV) is the closest known genetic homologue of hepatitis C virus. An effective prophylactic vaccine is currently not available for either of these hepaciviruses. The equine as potential surrogate model for hepacivirus vaccine studies was investigated, while equine host responses following vaccination with EqHV E2 recombinant protein and subsequent EqHV inoculation were elucidated. Four ponies received prime and booster vaccinations (recombinant protein, adjuvant) four weeks apart (day −55 and −27). Two control ponies received adjuvant only. Ponies were inoculated with EqHV RNA-positive plasma on day 0. Blood samples and liver biopsies were collected over 26 weeks (day −70 to +112). Serum analyses included detection of EqHV RNA, isotypes of E2-specific immunoglobulin G (IgG), nonstructural protein 3-specific IgG, haematology, serum biochemistry, and metabolomics. Liver tissue analyses included EqHV RNA detection, RNA sequencing, histopathology, immunohistochemistry, and fluorescent in situ hybridization. Al-though vaccination did not result in complete protective immunity against experimental EqHV inoculation, the majority of vaccinated ponies cleared the serum EqHV RNA earlier than the control ponies. The majority of vaccinated ponies appeared to recover from the EqHV-associated liver insult earlier than the control ponies. The equine model shows promise as a surrogate model for future hepacivirus vaccine research

The opposite of Dante's hell? The transfer of ideas for social housing at international congresses in the 1850s–1860s

With the advent of industrialization, the question of developing adequate housing for the emergent working classes became more pressing than before. Moreover, the problem of unhygienic houses in industrial cities did not stop at the borders of a particular nation-state; sometimes literally as pandemic diseases spread out 'transnationally'. It is not a coincidence that in the nineteenth century the number of international congresses on hygiene and social topics expanded substantially. However, the historiography about social policy in general and social housing in particular, has often focused on individual cases because of the different pace of industrial and urban development and is thus dominated by national perspectives. In this paper, I elaborate on transnational exchange processes and local adaptations and transformations. I focus on the transfer of the housing model of SOMCO in Mulhouse, (a French house building association) during social international congresses. I examine whether cross-national networking enabled and facilitated the implementation of ideas on the local scale. I will elaborate on the transmission and the local adaptation of the Mulhouse-model in Belgium. Convergences, divergences, and different factors that influenced the local transformations (personal choice, political situation, socioeconomic circumstances) will be taken into accoun

Studies on the Cobalt Deficiency in Ruminants (III) : Effects of Thiamine, Glucose and Cobalamin Injection on the Metabolism of Cobalt-deficient Sheep

International audienceN-terminal acetylation is a common protein modification in eukaryotes associated with numerous cellular processes. Inherited mutations in NAA10, encoding the catalytic subunit of the major N-terminal acetylation complex NatA have been associated with diverse, syndromic X-linked recessive disorders, whereas de novo missense mutations have been reported in one male and one female individual with severe intellectual disability but otherwise unspecific phenotypes. Thus, the full genetic and clinical spectrum of NAA10 deficiency is yet to be delineated. We identified three different novel and one known missense mutation in NAA10, de novo in 11 females, and due to maternal germ line mosaicism in another girl and her more severely affected and deceased brother. In vitro enzymatic assays for the novel, recurrent mutations p.(Arg83Cys) and p.(Phe128Leu) revealed reduced catalytic activity. X-inactivation was random in five females. The core phenotype of X-linked NAA10-related N-terminal-acetyltransferase deficiency in both males and females includes developmental delay, severe intellectual disability, postnatal growth failure with severe microcephaly, and skeletal or cardiac anomalies. Genotype–phenotype correlations within and between both genders are complex and may include various factors such as location and nature of mutations, enzymatic stability and activity, and X-inactivation in females

A New Model to Produce Infectious Hepatitis C Virus without the Replication Requirement

Numerous constraints significantly hamper the experimental study of hepatitis C virus (HCV). Robust replication in cell culture occurs with only a few strains, and is invariably accompanied by adaptive mutations that impair in vivo infectivity/replication. This problem complicates the production and study of authentic HCV, including the most prevalent and clinically important genotype 1 (subtypes 1a and 1b). Here we describe a novel cell culture approach to generate infectious HCV virions without the HCV replication requirement and the associated cell-adaptive mutations. The system is based on our finding that the intracellular environment generated by a West-Nile virus (WNV) subgenomic replicon rendered a mammalian cell line permissive for assembly and release of infectious HCV particles, wherein the HCV RNA with correct 5′ and 3′ termini was produced in the cytoplasm by a plasmid-driven dual bacteriophage RNA polymerase-based transcription/amplification system. The released particles preferentially contained the HCV-based RNA compared to the WNV subgenomic RNA. Several variations of this system are described with different HCV-based RNAs: (i) HCV bicistronic particles (HCVbp) containing RNA encoding the HCV structural genes upstream of a cell-adapted subgenomic replicon, (ii) HCV reporter particles (HCVrp) containing RNA encoding the bacteriophage SP6 RNA polymerase in place of HCV nonstructural genes, and (iii) HCV wild-type particles (HCVwt) containing unmodified RNA genomes of diverse genotypes (1a, strain H77; 1b, strain Con1; 2a, strain JFH-1). Infectivity was assessed based on the signals generated by the HCV RNA molecules introduced into the cytoplasm of target cells upon virus entry, i.e. HCV RNA replication and protein production for HCVbp in Huh-7.5 cells as well as for HCVwt in HepG2-CD81 cells and human liver slices, and SP6 RNA polymerase-driven firefly luciferase for HCVrp in target cells displaying candidate HCV surface receptors. HCV infectivity was inhibited by pre-incubation of the particles with anti-HCV antibodies and by a treatment of the target cells with leukocyte interferon plus ribavirin. The production of authentic infectious HCV particles of virtually any genotype without the adaptive mutations associated with in vitro HCV replication represents a new paradigm to decipher the requirements for HCV assembly, release, and entry, amenable to analyses of wild type and genetically modified viruses of the most clinically significant HCV genotypes

An Equine Model for Vaccination against a Hepacivirus: Insights into Host Responses to E2 Recombinant Protein Vaccination and Subsequent Equine Hepacivirus Inoculation

Equine hepacivirus (EqHV) is the closest known genetic homologue of hepatitis C virus. An effective prophylactic vaccine is currently not available for either of these hepaciviruses. The equine as potential surrogate model for hepacivirus vaccine studies was investigated, while equine host responses following vaccination with EqHV E2 recombinant protein and subsequent EqHV inoculation were elucidated. Four ponies received prime and booster vaccinations (recombinant protein, adjuvant) four weeks apart (day −55 and −27). Two control ponies received adjuvant only. Ponies were inoculated with EqHV RNA-positive plasma on day 0. Blood samples and liver biopsies were collected over 26 weeks (day −70 to +112). Serum analyses included detection of EqHV RNA, isotypes of E2-specific immunoglobulin G (IgG), nonstructural protein 3-specific IgG, haematology, serum biochemistry, and metabolomics. Liver tissue analyses included EqHV RNA detection, RNA sequencing, histopathology, immunohistochemistry, and fluorescent in situ hybridization. Al-though vaccination did not result in complete protective immunity against experimental EqHV inoculation, the majority of vaccinated ponies cleared the serum EqHV RNA earlier than the control ponies. The majority of vaccinated ponies appeared to recover from the EqHV-associated liver insult earlier than the control ponies. The equine model shows promise as a surrogate model for future hepacivirus vaccine research

Séquençage de novo de la région variable (FV) des anticorps circulants : objectif d’un logiciel de réassemblage

Humoral immunity is key for the adaptative response to infections and their prevention by vaccination. Although antibodies are at the core of the humoral response, their diversity has been much less studied than that of antibody-producing B cells. But a discrepancy has recently been reported between the repertoire of antigen receptors carried by circulating B cells and that of serum antibodies these cells produce. By enabling the de novo sequencing of antibodies, bottom-up proteomics has many assets to fill this gap. The protein cleavages that define this method markedly increase sequencing coverage. However, this requires reconstructing a posteriori the sequences of the heavy and light chains of the immunoglobulins and re-matching them. The complexity of this procedure increases considerably when analyzing complex antibody mixtures, such as those contained in serum. Therefore, we are developing software to assess the maximum antibody sequencing rate achievable with the bottom-up approach.L’immunité humorale est essentielle à la réponse adaptative aux infections et à leur prévention par la vaccination. La diversité des cellules productrices d’anticorps a été bien mieux étudiée que celle des anticorps proprement-dits, alors qu’ils constituent la pièce maitresse de la réponse humorale. Or, un décalage important a été récemment observé entre le répertoire des récepteurs antigéniques des lymphocytes B circulants et celui des anticorps du sérum. En permettant le séquençage de-novo d’anticorps, la protéomique en bottom-up présente de nombreux atouts pour combler cette lacune. Les clivages protéiques utilisés par cette méthode augmentent nettement la couverture de séquençage. Ceux-ci impliquent néanmoins de reconstruire a posteriori la séquence des chaines lourdes et légères des immunoglobulines et de les rapparier. Cette procédure devient rapidement complexe avec des mélanges d’anticorps, comme ceux issus du sérum. C’est pourquoi nous développons un logiciel pour évaluer le taux maximum de séquençage d’anticorps atteignable par l’approche bottom-up. Mots-Clés : Immunité humorale ; anticorps sériques ; spectrométrie de masse ; séquençage de novo ; reconstruction de séquences ; « One Health »

Proportions of CD4^highCD8^low, CD4^highCD8^high and CD4^lowCD8^high DPT cells in the blood and liver of patients.

<p><b>A,</b> Proportions of each DPT cell subpopulations: CD4^hiCD8^lo (1), CD4^hiCD8^hi (2) and CD4^loCD8^hi (3) in the blood of each patient (1 dot = 1 patient); percentages lower than 0.05 were plotted as 0. <b>B,</b> Same procedure as in (<b>A</b>) with LILs. Horizontal lines represent the values of the median for each DPT subpopulation in each group of patients; the three subpopulations of DPT cells correspond to the quadrants set-up in the lower right panel of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0020145#pone-0020145-g001" target="_blank">Figure 1A</a>, (numbers between brackets).</p

HCV and HBV mono-infected patients display distinct patterns of DPT cells.

<p><b>A,</b> Correlation between the proportions of CD4^hiCD8^lo DPT cell subpopulations (1) in blood and liver of HCV mono-infected patients; other subpopulations did not significantly correlate. <b>B,</b> Correlation between the proportions of CD4^loCD8^hi DPT cell subpopulations (3) in blood and liver of HBV mono-infected patients; CD4^hiCD8^hi correlated to the same extent as CD4^loCD8^hi (respectively 2 and 3) in HBV-infected patients (not shown). Each dot represents the proportions in blood and liver for a patient; significance of Pearson's correlation was tested using a t-test. The numbers between brackets correspond to the quadrants set-up in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0020145#pone-0020145-g001" target="_blank">Figure 1A</a>, lower right panel.</p