• AICD
• Autoimmunity
• CD5
• Cancer
• Cytoplasmic CD5
• Membrane CD5
• Tolerance
• Tumor-infiltrating lymphocytes

• Humans
• CD5 Antigens/immunology
• CD5 Antigens/metabolism
• CD5 Antigens/genetics
• Immunotherapy
• T-Lymphocytes/immunology
• T-Lymphocytes/metabolism
• Neoplasms/immunology
• Neoplasms/therapy
• Neoplasms/genetics
• Neoplasms/metabolism
• Animals

Crohn’s disease (CD) and ulcerative colitis (UC) are inflammatory bowel diseases (IBD) resulting from the interaction of multiple environmental, genetic and immunological factors. CD5 and CD6 are paralogs encoding lymphocyte co-receptors involved in fine-tuning intracellular signals delivered upon antigen-specific recognition, microbial pattern recognition and cell adhesion. While CD5 and CD6 expression and variation is known to influence some immune-mediated inflammatory disorders, their role in IBD remains unclear. To this end, Cd5- and Cd6-deficient mice were subjected to dextran sulfate sodium (DSS)-induced colitis, the most widely used experimental animal model of IBD. The two mouse lines showed opposite results regarding body weight loss and disease activity index (DAI) changes following DSS-induced colitis, thus supporting Cd5 and Cd6 expression involvement in the pathophysiology of this experimental IBD model. Furthermore, DNA samples from IBD patients of the ENEIDA registry were used to test association of CD5 (rs2241002 and rs2229177) and CD6 (rs17824933, rs11230563, and rs12360861) single nucleotide polymorphisms with susceptibility and clinical parameters of CD (n=1352) and UC (n=1013). Generalized linear regression analyses showed association of CD5 variation with CD ileal location (rs2241002^CC) and requirement of biological therapies (rs2241002^C-rs2229177^T haplotype), and with poor UC prognosis (rs2241002^T-rs2229177^T haplotype). Regarding CD6, association was observed with CD ileal location (rs17824933^G) and poor prognosis (rs12360861^G), and with left-sided or extensive UC, and absence of ankylosing spondylitis in IBD (rs17824933^G). The present experimental and genetic evidence support a role for CD5 and CD6 expression and variation in IBD’s clinical manifestations and therapeutic requirements, providing insight into its pathophysiology and broadening the relevance of both immunomodulatory receptors in immune-mediated disorders.

• Animals
• Colitis/chemically induced
• Colitis, Ulcerative/chemically induced
• Colitis, Ulcerative/genetics
• Crohn Disease/genetics
• Dextran Sulfate/toxicity
• Inflammatory Bowel Diseases/genetics
• Mice

• 5′ untranslated region
• Hepatitis C virus
• miR-122
• miR-122-independent replication
• viral tropism

• MOP133458 CIHR
• CoMBRIDGE University of Saskatchewan
• Scholarship CanHepC
• Teaching Fellowship University of Saskatchewan

Primary Sjögren's syndrome (pSS) is an autoimmune disease triggered by a combination of environmental and host genetic factors, which results in the focal lymphocytic infiltration of exocrine glands causing eye and mouth dryness. Glandular infiltrates include T and B cell subsets positive for CD5 and/or CD6, two surface scavenger receptors involved in the fine-tuning of intracellular signals mediated by the antigen-specific receptor complex of T (TCR) and B (BCR) cells. Moreover, the epithelial cells of inflamed glands overexpress CD166/ALCAM, a CD6 ligand involved in homo and heterotypic cell adhesion interactions. All this, together with the reported association of functionally relevant single nucleotide polymorphisms (SNPs) of CD5, CD6, and CD166/ALCAM with the risk or prognosis of some immune-mediated inflammatory disorders, led us to investigate similar associations in a local cohort of patients with pSS. The logistic regression analyses of individual SNPs showed the association of CD5 rs2241002^T with anti-Ro/La positivity, CD6 rs17824933^C with neutropenia, and CD6 rs11230563^T with increased leukopenia and neutropenia but decreased peripheral nervous system EULAR Sjögren's syndrome disease activity index (ESSDAI). Further analyses showed the association of haplotypes from CD5 (rs2241002^T-rs2229177^C) with anemia and thrombocytopenia, CD6 (rs17824933^G-rs11230563^C-rs12360861^G) with cutaneous ESSDAI, and CD166/ALCAM (rs6437585^C-rs579565^A-rs1044243^C and rs6437585^C-rs579565^G-rs1044243^T) with disease susceptibility and several analytical parameters (anti-nuclear antibodies, neurological ESSDAI, and hematologic cytopenias). These results support the relevance of gene variation at loci coding for cell surface receptors involved in the modulation of T and B lymphocyte activation (CD5, CD6) and epithelial-immune cell adhesion (CD166/ALCAM) in modulating the clinical and analytical outcomes in patients with pSS.

• Ministerio de Economía y Competitividad
• Ministerio de Ciencia e Innovación
• Agència de Gestió d'Ajuts Universitaris i de Recerca
• Ministerio de Educación, Cultura y Deporte

Metabolic function plays a key role in immune cell activation, destruction of foreign pathogens, and memory cell generation. As T cells are activated, their metabolic profile is significantly changed due to signaling cascades mediated by the T cell receptor (TCR) and co-receptors found on their surface. CD5 is a T cell co-receptor that regulates thymocyte selection and peripheral T cell activation. The removal of CD5 enhances T cell activation and proliferation, but how this is accomplished is not well understood. We examined how CD5 specifically affects CD4+ T cell metabolic function and systemic metabolome by analyzing serum and T cell metabolites from CD5WT and CD5KO mice. We found that CD5 removal depletes certain serum metabolites, and CD5KO T cells have higher levels of several metabolites. Transcriptomic analysis identified several upregulated metabolic genes in CD5KO T cells. Bioinformatic analysis identified glycolysis and the TCA cycle as metabolic pathways promoted by CD5 removal. Functional metabolic analysis demonstrated that CD5KO T cells have higher oxygen consumption rates (OCR) and higher extracellular acidification rates (ECAR). Together, these findings suggest that the loss of CD5 is linked to CD4+ T cell metabolism changes in metabolic gene expression and metabolite concentration.

Since the discovery of hepatitis B virus (HBV) over five decades ago, there have been many independent studies showing presence of HBV genomes in cells of the immune system. However, the nature of HBV lymphotropism and its significance with respect to HBV biology, persistence and the pathogenesis of liver and extrahepatic disorders remains underappreciated. This is in contrast to studies of other viral pathogens in which the capability to infect immune cells is an area of active investigation. Indeed, in some viral infections, lymphotropism may be essential, and even a primary mechanism of viral persistence, and a major contributor to disease pathogenesis. Nevertheless, there are advances in understanding of HBV lymphotropism in recent years due to cumulative evidence showing that: (i) lymphoid cells are a reservoir of replicating HBV, (ii) are a site of HBV-host DNA integration and (iii) virus genomic diversification leading to pathogenic variants, and (iv) they play a role in HBV resistance to antiviral therapy and (v) likely contribute to reactivation of hepatitis B. Further support for HBV lymphotropic nature is provided by studies in a model infection with the closely related woodchuck hepatitis virus (WHV) naturally infecting susceptible marmots. This animal model faithfully reproduces many aspects of HBV biology, including its replication scheme, tissue tropism, and induction of both symptomatic and silent infections, immunological processes accompanying infection, and progressing liver disease culminating in hepatocellular carcinoma. The most robust evidence came from the ability of WHV to establish persistent infection of the immune system that may not engage the liver when small quantities of virus are experimentally administered or naturally transmitted into virus-naïve animals. Although the concept of HBV lymphotropism is not new, it remains controversial and not accepted by conventional HBV researchers. This review summarizes research advances on HBV and hepadnaviral lymphotropism including the role of immune cells infection in viral persistence and the pathogenesis of HBV-induced liver and extrahepatic diseases. Finally, we discuss the role of immune cells in HBV diagnosis and assessment of antiviral therapy efficacy.

• hepatitis B virus
• lymphotyropism
• virus genome integration
• virus variants
• woodchuck model of hepatitis B

The hepatitis C virus (HCV), an obligatory intracellular pathogen, highly depends on its host cells to propagate successfully. The HCV life cycle can be simply divided into several stages including viral entry, protein translation, RNA replication, viral assembly and release. Hundreds of cellular factors involved in the HCV life cycle have been identified over more than thirty years of research. Characterization of these cellular factors has provided extensive insight into HCV replication strategies. Some of these cellular factors are targets for anti-HCV therapies. In this review, we summarize the well-characterized and recently identified cellular factors functioning at each stage of the HCV life cycle.

• Hepatitis C virus
• Cellular factor
• Viral entry
• Translation
• Replication
• Assembly
• Release

• Animals
• Hepacivirus
• Hepatitis C/drug therapy
• Humans
• Life Cycle Stages
• Virus Assembly
• Virus Replication

Pathogens are one of the main selective pressures that ancestral humans had to adapt to. Components of the immune response system have been preferential targets of natural selection in response to such pathogen-driven pressure. In turn, there is compelling evidence showing that positively selected immune gene variants conferring increased resistance to past or present infectious agents are today associated with increased risk for autoimmune or inflammatory disorders but decreased risk of cancer, the other side of the same coin. CD5 and CD6 are lymphocytic scavenger receptors at the interphase of the innate and adaptive immune responses since they are involved in both: (i) microbial-associated pattern recognition; and (ii) modulation of intracellular signals mediated by the clonotypic antigen-specific receptor present in T and B cells (TCR and BCR, respectively). Here, we review available information on CD5 and CD6 as targets of natural selection as well as on the role of CD5 and CD6 variation in autoimmunity and cancer.

• CD5
• CD6
• autoimmunity
• cancer
• human genetics
• immune response
• infections
• natural selection
• scavenger receptors
• single nucleotide polymorphisms

• HCV extrahepatic manifestations
• Hepatitis C virus
• Interferon-free DAA regimens

• CD5
• Candida albicans
• Cryptococcus neoformans
• antifungal therapy
• fluconazole
• fungal infection
• immunotherapy
• scavenger receptors
• β-glucan

• Animals
• Antifungal Agents/pharmacology
• Candida albicans
• Cryptococcus neoformans
• Lymphocytes
• Mice
• Mycoses
• Receptors, Scavenger

• European Commission (EC)
• Ministerio de Educación, Cultura y Deporte (MECD)
• Agencia Nacional de Investigación e Innovación (ANII)
• MEC | Fundação para a Ciência e a Tecnologia (FCT)
• Ministerio de Economía y Competitividad (MEC)
• Ministerio de Economía, Industria y Competitividad, Gobierno de España (MINECO)

CD5 and CD6 are closely related signal-transducing class I scavenger receptors mainly expressed on lymphocytes. Both receptors are involved in the modulation of the activation and differentiation cell processes triggered by clonotypic antigen-specific receptors present on T and B cells (TCR and BCR, respectively). To serve such a relevant immunomodulatory function, the extracellular region of CD5 and CD6 interacts with soluble and/or cell-bound endogenous counterreceptors but also microbial-associated molecular patterns (MAMPs). Evidence from genetically-modified mouse models indicates that the absence or blockade of CD5- and CD6-mediated signals results in dysregulated immune responses, which may be deleterious or advantageous in some pathological conditions, such as infection, cancer or autoimmunity. Bench to bedside translation from transgenic data is constrained by ethical concerns which can be overcome by exogenous administration of soluble proteins acting as decoy receptors and leading to transient “functional knockdown”. This review gathers information currently available on the therapeutic efficacy of soluble CD5 and CD6 receptor infusion in different experimental models of disease. The existing proof-of-concept warrants the interest of soluble CD5 and CD6 as safe and efficient immunotherapeutic agents in diverse and relevant pathological conditions.

• autoimmunity
• cancer
• immunomodulation
• immunotherapy
• infection
• soluble CD5
• soluble CD6

• Animals
• Antigens, CD/immunology
• Antigens, Differentiation, T-Lymphocyte/immunology
• B-Lymphocytes/immunology
• CD5 Antigens/immunology
• Humans
• Immunotherapy/methods
• Receptors, Scavenger/immunology
• T-Lymphocytes/immunology

• CD5
• CD6
• Echinococcus granulosus sensu lato
• Interactome
• Ligand
• Scavenger receptor

• RNA stabilization 6
• hepatitis C virus 1
• miR-122 3
• microRNA 2
• pathogenesis 8
• replication 4
• translation 5
• tropism 7

Background: The role of regulatory T cells (Tregs) is now well established in the progression of hepatocellular carcinoma (HCC) linked to Hepatitis C virus (HCV) infection. However, nothing is known about the potential interplay between Tregs and HCV. In this pilot study, we have investigated the ability of Tregs to hang HCV on and the subsequent effect on their suppressive function and phenotype. Moreover, we have evaluated how HCV could promote the recruitment of Tregs by infected primary human hepatocytes. Methods: Tregs of healthy donors were incubated with JFH-1/HCVcc. Viral inoculation was assessed using adapted assays (RT-qPCR, Flow Citometry (FACS) and Western Blot (WB). Expression of Tregs phenotypic (CD4, CD25, CD127 and Foxp3) and functional (IL-10, GZMB, TGF-β1 and IL-2) markers was monitored by RT-qPCR, FACS and ELISA. Suppressive activity was validated by suppressive assays. Tregs recruitment by infected primary hepatic cells was evaluated using Boyden Chamber. Results: Tregs express the classical HCV receptors (CD81, CLDN1 and LDLR) and some co-receptors (CD5). HCV inoculation significantly increases the suppressive phenotype and activity of Tregs, and raises their anergy by inducing an unexpected IL-2 production. Moreover, HCV infection induces the expression of chemokines (CCL17, CXCL16, and CCL20) by primary hepatic human hepatocytes and chemokine receptors (CCR4, CXCR6 and CCR6) by Tregs. Finally, infected hepatocytes have a significantly higher potential to recruit Tregs in a seemingly CCL20-dependent manner. Conclusions: Direct interaction between HCV and Tregs represents a newly defined mechanism that could potentiate HCV immune evasion and favor intratumoral recruitment contributing to HCC progression.

• HCV
• HCV/JFH-1
• chemokines
• immune escape
• regulatory T cells

• Antigens, Differentiation/immunology
• Gene Expression Regulation/immunology
• Hepacivirus/immunology
• Hepatitis C/immunology
• Hepatitis C/pathology
• Humans
• Immune Evasion
• Liver/immunology
• Liver/pathology
• Liver/virology
• T-Lymphocytes, Regulatory/immunology
• T-Lymphocytes, Regulatory/pathology

• Clostridium perfringens enterotoxin
• Hepatitis C Virus
• claudin targeting
• claudin-1
• epidermal barrier
• reconstructed human epidermis
• viral entry

• Chromosomal aberration
• Genetic damage
• Hepatitis C virus
• Lymphomagenesis
• Mixed cryoglobulinemia
• Non-Hodgkin B-lymphoma
• Oncogenic pathways
• Pathogenesis

• B lymphocytes
• T cell responses
• T lymphocytes
• apoptosis
• signal transduction
• thymus
• tolerance/suppression/anergy

Scavenger Receptors (SRs) from the host’s innate immune system are known to bind multiple ligands to promote the removal of non-self or altered-self targets. CD5 and CD6 are two highly homologous class I SRs mainly expressed on all T cells and the B1a cell subset, and involved in the fine tuning of activation and differentiation signals delivered by the antigen-specific receptors (TCR and BCR, respectively), to which they physically associate. Additionally, CD5 and CD6 have been shown to interact with and sense the presence of conserved pathogen-associated structures from bacteria, fungi and/or viruses.

We report herein the interaction of CD5 and CD6 lymphocyte surface receptors with Echinococcus granulosus sensu lato (s.l.). Binding studies show that both soluble and membrane-bound forms of CD5 and CD6 bind to intact viable protoscoleces from E. granulosus s.l. through recognition of metaperiodate-resistant tegumental components. Proteomic analyses allowed identification of thioredoxin peroxidase for CD5, and peptidyl-prolyl cis-trans isomerase (cyclophilin) and endophilin B1 (antigen P-29) for CD6, as their potential interactors. Further in vitro assays demonstrate that membrane-bound or soluble CD5 and CD6 forms differentially modulate the pro- and anti-inflammatory cytokine release induced following peritoneal cells exposure to E. granulosus s.l. tegumental components. Importantly, prophylactic infusion of soluble CD5 or CD6 significantly ameliorated the infection outcome in the mouse model of secondary cystic echinococcosis.

Taken together, the results expand the pathogen binding properties of CD5 and CD6 and provide novel evidence for their therapeutic potential in human cystic echinococcosis.

Scavenger Receptors (SRs) are constituents of host’s innate immune system able to sense and remove altered-self and/or pathogen components. Data on their interaction with helminth parasites is scarce. In this work, we describe that CD5 and CD6 -two lymphoid SRs previously reported to interact with conserved structures from bacteria, fungi and viruses- recognize tegumental components in the cestode parasite Echinococcus granulosus sensu lato (s.l.). Moreover, both receptors differentially modulate the cytokine release by host cells exposed to E. granulosus s.l. tegumental components. Importantly, the infusion of soluble forms of CD5 or CD6 improve infection outcomes in a murine model of secondary cystic echinococcosis. In summary, our results expand the pathogen binding properties of CD5 and CD6 and suggest their therapeutic potential against helminth infections.

• Animals
• Antigens, CD/genetics
• Antigens, CD/metabolism
• Antigens, Differentiation, T-Lymphocyte/genetics
• Antigens, Differentiation, T-Lymphocyte/metabolism
• CD5 Antigens/genetics
• CD5 Antigens/metabolism
• Echinococcosis/genetics
• Echinococcosis/metabolism
• Echinococcosis/parasitology
• Echinococcus granulosus/genetics
• Echinococcus granulosus/metabolism
• Female
• Helminth Proteins/genetics
• Helminth Proteins/metabolism
• Humans
• Mice
• Mice, Inbred BALB C
• Mice, Inbred C57BL
• Protein Binding
• Proteomics
• Receptors, Scavenger/genetics
• Receptors, Scavenger/metabolism
• T-Lymphocytes/metabolism
• T-Lymphocytes/parasitology

• Innate immune response
• microbial pathogenesis
• pathogens
• scavenger receptors

• T cells
• hepatitis C virus
• lymphocytes
• lymphotropism
• virus variants

• clinicopathologic characteristics
• hepatitis C virus
• relapse
• transformed diffuse large b-cell lymphoma

• CD5
• Cancer immunotherapy
• Danger signal
• Immune-checkpoint
• Scavenger receptor
• Scavenger receptor class B type 1
• Toll-like receptors

• B-Lymphocytes/immunology
• B-Lymphocytes/virology
• B7-2 Antigen/genetics
• B7-2 Antigen/immunology
• Cell Line, Tumor
• DEAD Box Protein 58/antagonists & inhibitors
• DEAD Box Protein 58/genetics
• DEAD Box Protein 58/immunology
• Gene Expression Regulation
• Gene Library
• HEK293 Cells
• Hep G2 Cells
• Hepacivirus/immunology
• Host-Pathogen Interactions
• Humans
• Immunologic Memory
• Interferon-Induced Helicase, IFIH1/genetics
• Interferon-Induced Helicase, IFIH1/immunology
• MicroRNAs/antagonists & inhibitors
• MicroRNAs/genetics
• MicroRNAs/immunology
• Protein Binding
• RNA, Small Interfering/genetics
• RNA, Small Interfering/metabolism
• Receptors, Immunologic
• Signal Transduction
• Viral Envelope Proteins/genetics
• Viral Envelope Proteins/immunology
• Viral Tropism/immunology
• Virus Replication

• P30 CA016672 NCI NIH HHS
• R01 AI095097 NIAID NIH HHS
• R01 CA082057 NCI NIH HHS
• R24 AA012885 NIAAA NIH HHS
• R01 AA018857 NIAAA NIH HHS
• P30 DK048522 NIDDK NIH HHS
• P50 AA011999 NIAAA NIH HHS
• R21 AA025470 NIAAA NIH HHS
• P30 CA014089 NCI NIH HHS
• U19 AI083025 NIAID NIH HHS
• R01 AA025204 NIAAA NIH HHS

Current cell processing technologies for gene and cell therapies are often slow, expensive, labor intensive and are compromised by high cell losses and poor selectivity thus limiting the efficacy and availability of clinical cell therapies. We employ cell-specific on-demand mechanical intracellular impact from laser pulse-activated plasmonic nanobubbles (PNB) to process heterogeneous human cell grafts ex vivo with dual simultaneous functionality, the high cell type specificity, efficacy and processing rate for transfection of target CD3+ cells and elimination of subsets of unwanted CD25+ cells. The developed bulk flow PNB system selectively processed human cells at a rate of up to 100 million cell/minute, providing simultaneous transfection of CD3+ cells with the therapeutic gene (FKBP12(V36)-p30Caspase9) with the efficacy of 77% and viability 95% (versus 12 and 60%, respectively, for standard electroporation) and elimination of CD25+ cells with 99% efficacy. PNB flow technology can unite and replace several methodologies in an all-in-one universal ex vivo simultaneous procedure to precisely and rapidly prepare a cell graft for therapy. PNB’s can process various cell systems including cord blood, stem cells, and bone marrow.

Hepatitis C virus (HCV) frequently causes chronic hepatitis, while spontaneous recovery from infection is infrequent. Persistence of HCV after self-limited (spontaneous) resolution of hepatitis C was rarely investigated. The current study aimed to assess incidence and robustness of HCV persistence after self-resolved hepatitis C in individuals with normal liver enzymes and undetectable virus by conventional tests. Applying high sensitivity HCV RNA detection approaches, we analyzed plasma and peripheral blood mononuclear cells (PBMC) from individuals with previous hepatitis C infection. Parallel plasma and PBMC from 24 such non-viraemic individuals followed for 0.3–14.4 (mean 6.4) years were examined. Additional samples from 9 of them were obtained 4.5–7.2 (mean 5.9) years later. RNA was extracted from 250 μl plasma and, if HCV negative, from ~5 ml after ultracentrifugation, and from ex vivo stimulated PBMC. PBMC with evidence of HCV replication from 4 individuals were treated with HCV protease inhibitor, telaprevir. HCV RNA was detected in 14/24 (58.3%) plasma and 11/23 (47.8%) PBMC obtained during the first collection. HCV RNA replicative strand was evident in 7/11 (63.6%) PBMC. Overall, 17/24 (70.8%) individuals carried HCV RNA at mean follow-up of 5.9 years. Samples collected 4.5–7.2 years later revealed HCV in 4/9 (44.4%) plasma and 5/9 (55.5%) PBMC, while 4 (80%) of these 5 PBMC demonstrated virus replicative strand. Overall, 6/9 (66.7%) individuals remained viraemic for up to 20.7 (mean 12.7) years. Telaprevir entirely eliminated HCV replication in the PBMC examined. In conclusion, our results indicate that HCV can persist long after spontaneous resolution of hepatitis C at levels undetectable by current testing. An apparently effective host immune response curtailing hepatitis appears insufficient to completely eliminate the virus. The long-term morbidity of asymptomatic HCV carriage should be examined even in individuals who achieve undetectable HCV by standard testing and their need for treatment should be assessed.

• Adult
• Aged
• Base Sequence
• Female
• Follow-Up Studies
• Hepacivirus/metabolism
• Hepatitis C, Chronic/blood
• Hepatitis C, Chronic/drug therapy
• Humans
• Leukocytes, Mononuclear/metabolism
• Leukocytes, Mononuclear/virology
• Male
• Middle Aged
• Molecular Sequence Data
• Oligopeptides/administration & dosage
• RNA, Viral/blood
• Remission, Spontaneous
• Time Factors

• MOP-126056 Canadian Institutes of Health Research

B-cell receptor (BCR) signaling is essential for the development of B cells and has a critical role in B-cell neoplasia. Increasing evidence indicates an association between chronic hepatitis C virus (HCV) infection and B-cell lymphoma, however, the mechanisms by which HCV causes B-cell lymphoproliferative disorder are still unclear. Herein, we demonstrate the expression of HCV viral proteins in B cells of HCV-infected patients and show that HCV upregulates BCR signaling in human primary B cells. HCV nonstructural protein NS3/4A interacts with CHK2 and downregulates its activity, modulating HuR posttranscriptional regulation of a network of target mRNAs associated with B-cell lymphoproliferative disorders. Interestingly, the BCR signaling pathway was found to have the largest number of transcripts with increased association with HuR and was upregulated by NS3/4A. Our study reveals a previously unidentified role of NS3/4A in regulation of host BCR signaling during HCV infection, contributing to a better understanding of the molecular mechanisms underlying HCV-associated B-cell lymphoproliferative disorders.

T cell receptor (TCR) signaling is required for T-cell activation, proliferation, differentiation, and effector function. Hepatitis C virus (HCV) infection is associated with impaired T-cell function leading to persistent viremia, delayed and inconsistent antibody responses, and mild immune dysfunction. Although multiple factors appear to contribute to T-cell dysfunction, a role for HCV particles in this process has not been identified. Here, we show that incubation of primary human CD4+ and CD8+ T-cells with HCV RNA-containing serum, HCV-RNA containing extracellular vesicles (EVs), cell culture derived HCV particles (HCVcc) and HCV envelope pseudotyped retrovirus particles (HCVpp) inhibited TCR-mediated signaling. Since HCVpp’s contain only E1 and E2, we examined the effect of HCV E2 on TCR signaling pathways. HCV E2 expression recapitulated HCV particle-induced TCR inhibition. A highly conserved, 51 nucleotide (nt) RNA sequence was sufficient to inhibit TCR signaling. Cells expressing the HCV E2 coding RNA contained a short, virus-derived RNA predicted to be a Dicer substrate, which targeted a phosphatase involved in Src-kinase signaling (PTPRE). T-cells and hepatocytes containing HCV E2 RNA had reduced PTPRE protein levels. Mutation of 6 nts abolished the predicted Dicer interactions and restored PTPRE expression and proximal TCR signaling. HCV RNA did not inhibit distal TCR signaling induced by PMA and Ionomycin; however, HCV E2 protein inhibited distal TCR signaling. This inhibition required lymphocyte-specific tyrosine kinase (Lck). Lck phosphorylated HCV E2 at a conserved tyrosine (Y613), and phospho-E2 inhibited nuclear translocation of NFAT. Mutation of Y613 restored distal TCR signaling, even in the context of HCVpps. Thus, HCV particles delivered viral RNA and E2 protein to T-cells, and these inhibited proximal and distal TCR signaling respectively. These effects of HCV particles likely aid in establishing infection and contribute to viral persistence.

Globally, approximately 200 million people are persistently infected with Hepatitis C virus (HCV). Mechanisms by which HCV establishes persistent infection are complex, and several host and viral factors appear to contribute to the ability of HCV to evade immune clearance. T cell activation through the T cell receptor (TCR) is an essential first step in the generation of an adaptive immune response. Although HCV infection is associated with impaired T cell function, the mechanisms for this dysfunction are poorly understood. Here, we demonstrate that HCV particles inhibit T cell activation by interfering with proximal and distal signals that are triggered by activation through the TCR. First, HCV envelope (E2) RNA was processed into a small RNA that targeted a regulatory phosphatase, inhibiting proximal TCR signaling. Second, the lymphocyte specific Src kinase (Lck) phosphorylated HCV E2 at tyrosine 613 (Y613), and phospho-E2 inhibited nuclear translocation of activated NFAT, reducing distal TCR activation signals. The RNA and protein motifs involved are highly conserved among all HCV isolates, and mutation restored TCR signaling. Thus, HCV particles interfere with TCR signaling and impair T cell activation using two distinct mechanisms. This may contribute to HCV persistence and T cell dysfunction during HCV infection.

Hepatitis C virus (HCV) can replicate in cells of the immune system and productively propagate in primary T lymphocytes in vitro. We aimed to determine whether exposure to authentic, patient-derived HCV can modify the proliferation capacity, susceptibility to apoptosis and phenotype of T cells.

Primary total T cells from a healthy donor were used as targets and plasma-derived HCV from patients with chronic hepatitis C served as inocula. T cell phenotype was determined prior to and at different time points after exposure to HCV. T cell proliferation and apoptosis were measured by flow cytometry-based assays.

The HCV inocula that induced the highest intracellular expression of HCV also caused a greatest shift in the T cell phenotype from predominantly CD4-positive to CD8-positive. This shift was associated with inhibition of CD4+ but not CD8+ T cell proliferation and did not coincide with altered apoptotic death of either cell subset.

The data obtained imply that exposure to native HCV can have an impact on the relative frequencies of CD4+ and CD8+ T cells by selectively suppressing CD4⁺ T lymphocyte proliferation and this may occur in both the presence and the absence of measurable HCV replication in these cells. If the virus exerts a similar effect in vivo, it may contribute to the impairment of virus-specific T cell response by altering cooperation between immune cell subsets.

The online version of this article (doi:10.1186/s12985-015-0322-4) contains supplementary material, which is available to authorized users.

• Chronic hepatitis C virus infection
• Interferon alpha therapy
• Interferon alpha-induced gene expression
• Peripheral blood mononuclear cells

Hepatitis C virus (HCV) infection represents an important public health problem worldwide. Reduction of HCV morbidity and mortality is a current challenge owned to several viral and host factors. Virus molecular evolution plays an important role in HCV transmission, disease progression and therapy outcome. The high degree of genetic heterogeneity characteristic of HCV is a key element for the rapid adaptation of the intrahost viral population to different selection pressures (e.g., host immune responses and antiviral therapy). HCV molecular evolution is shaped by different mechanisms including a high mutation rate, genetic bottlenecks, genetic drift, recombination, temporal variations and compartmentalization. These evolutionary processes constantly rearrange the composition of the HCV intrahost population in a staging manner. Remarkable advances in the understanding of the molecular mechanism controlling HCV replication have facilitated the development of a plethora of direct-acting antiviral agents against HCV. As a result, superior sustained viral responses have been attained. The rapidly evolving field of anti-HCV therapy is expected to broad its landscape even further with newer, more potent antivirals, bringing us one step closer to the interferon-free era.

• Hepatitis C virus
• Evolution
• Phylogenetics
• Drug resistance
• Clinical outcome

• Animals
• Antiviral Agents/adverse effects
• Antiviral Agents/therapeutic use
• Disease Progression
• Drug Resistance, Viral
• Drug Therapy, Combination
• Evolution, Molecular
• Genotype
• Hepacivirus/drug effects
• Hepacivirus/genetics
• Hepacivirus/pathogenicity
• Hepatitis C/diagnosis
• Hepatitis C/drug therapy
• Hepatitis C/transmission
• Host-Pathogen Interactions
• Humans
• Phenotype
• Treatment Outcome

It has been reported that the direct binding of hepatitis C virus (HCV) and/or the replication of HCV in the extrahepatic organs and, especially, lymphoid cells, might affect the pathogenesis of extrahepatic diseases with HCV infection. More than one decade ago, several reports described the existence of HCV-RNA in peripheral blood mononuclear cells. Moreover, many reports describing the existence of HCV in B lymphocytes and B cell lymphoma have been published. In addition to B lymphocytes, it was reported that HCV replication could be detected in T lymphocytes and T cell lines. Among the extrahepatic diseases with HCV infection, mixed cryoglobulinemia-related diseases and autoimmune-related diseases are important for understanding the immunopathogensis of HCV persistent infection. Moreover, HCV persistent infection can cause malignant lymphoma. The biological significance of lymphotropic HCV has not yet become clear. However, several candidates have been considered for a long time. One is that lymphotropic HCV is an HCV reservoir that might contribute to the recurrence of HCV infection and difficult-to-treat disease status. The other important issue is the carcinogenesis of the lymphoid cells and disturbances of the immune responses. Therefore, the extrahepatic diseases might be induced by direct interaction between HCV and lymphoid cells. In this article, we summarize various studies showing the direct effect of HCV on lymphoid cells and discuss the biological significance of lymphotropic HCV.

• Hepatitis C virus
• Lymphotropism
• T cell
• B cell
• Immunology

• Animals
• Autoimmune Diseases/immunology
• Autoimmune Diseases/virology
• Cryoglobulinemia/immunology
• Cryoglobulinemia/virology
• Hepacivirus/immunology
• Hepacivirus/pathogenicity
• Hepatitis C/complications
• Hepatitis C/immunology
• Hepatitis C/virology
• Humans
• Immune Evasion
• Lymphocytes/immunology
• Lymphocytes/virology
• Lymphoma/immunology
• Lymphoma/virology

Resolution of chronic hepatitis C is considered when serum HCV RNA becomes repeatedly undetectable and liver enzymes normalize. However, long-term persistence of HCV following therapy with pegylated interferon-α/ribavirin (PegIFN/R) was reported when more sensitive assays and testing of serial plasma, lymphoid cells (PBMC) and/or liver biopsies was applied. Our aim was to reassess plasma and PBMCs collected during and after standard PegIFN/R therapy from individuals who became HCV RNA nonreactive by clinical testing. Of particular interest was to determine if HCV genome and its replication remain detectable during ongoing treatment with PegIFN/R when evaluated by more sensitive detection approaches. Plasma acquired before (n = 11), during (n = 25) and up to 12–88 weeks post-treatment (n = 20) from 9 patients and PBMC (n = 23) from 3 of them were reanalyzed for HCV RNA with sensitivity <2 IU/mL. Clone sequencing of the HCV 5′-untranslated region from plasma and PBMCs was done in 2 patients. HCV RNA was detected in 17/25 (68%) plasma and 8/10 (80%) PBMC samples collected from 8 of 9 patients during therapy, although only 5.4% plasma samples were positive by clinical assays. Among post-treatment HCV RNA-negative plasma samples, 9 of 20 (45.3%) were HCV reactive for up to 59 weeks post-treatment. Molecularly evident replication was found in 6/12 (50%) among PBMC reactive for virus RNA positive strand collected during or after treatment. Pre-treatment point mutations persisted in plasma and/or PBMC throughout therapy and follow-up. Therefore, HCV is not completely cleared during ongoing administration of PegIFN/R otherwise capable of ceasing progression of CHC and virus commonly persists at levels not detectable by the current clinical testing. The findings suggest the need for continued evaluation even after patients achieve undetectable HCV RNA post-treatment.

• Antiviral Agents/administration & dosage
• Antiviral Agents/therapeutic use
• Female
• Hepacivirus/genetics
• Hepacivirus/isolation & purification
• Hepacivirus/physiology
• Hepatitis C, Chronic/drug therapy
• Hepatitis C, Chronic/virology
• Humans
• Interferon-alpha/administration & dosage
• Interferon-alpha/therapeutic use
• Limit of Detection
• Male
• Polyethylene Glycols/administration & dosage
• Polyethylene Glycols/therapeutic use
• RNA, Viral/blood
• Recombinant Proteins/administration & dosage
• Recombinant Proteins/therapeutic use
• Ribavirin/administration & dosage
• Ribavirin/therapeutic use
• Virus Replication

• MOP-126056 Canadian Institutes of Health Research

Hepatitis C virus (HCV) is a major cause of liver disease worldwide. The routine diagnostics identifying HCV infection include testing for specific anti-HCV antibodies by enzyme-linked immnunosorbent assay and viral genetic material in serum or plasma. However, a small proportion of patients persistently infected with HCV, in whom anti-HCV are undetectable, constitute a serious diagnostic and possibly epidemiologic problem, as they could facilitate pathogen spread in the population. This type of infection is termed seronegative or serosilent. Seronegative HCV infection is currently of great interest to both scientists and physicians. The review presents epidemiological data concerning the prevalence of seronegative HCV infection in HIV/HCV co-infected individuals, hemodialysis patients, and blood and organ donors. The possible mechanisms behind this atypical course of infection are discussed. Furthermore, the differences between seronegative and occult infections and prolonged seroconversion are explained.

Accumulated evidence implies that hepatitis C virus (HCV) infects not only the liver but also the immune system. A lymphocyte-specific CD5 molecule was recently identified as essential for infection of T cells with native, patient-derived HCV. To assess whether the proposed hepatocyte receptors may also contribute to HCV lymphotropism, expression of scavenger receptor-class B type 1 (SR-B1), claudin-1 (CLDN-1), claudin-6 (CLDN-6), occludin (OCLN), CD5 and CD81 was examined by real-time RT-PCR and the respective proteins quantified by immunoblotting in HCV-prone and resistant T cell lines, peripheral blood mononuclear cells (PBMC), primary T cells and their subsets, and compared to hepatoma Huh7.5 and HepG2 cells. SR-B1 protein was found in T and hepatoma cell lines but not in PBMC or primary T lymphocytes, CLDN-1 in HCV-resistant PM1 T cell line and hepatoma cells only, while CLDN-6 equally in the cells investigated. OCLN protein occurred in HCV-susceptible Molt4 and Jurkat T cells and its traces in primary T cells, but not in PBMC. CD5 was displayed by HCV-prone T cell lines, primary T cells and PBMC, but not by non-susceptible T and hepatoma cell lines, while CD81 in all cell types except HepG2. Knocking-down OCLN in virus-prone T cell line inhibited HCV infection, while de novo infection downregulated OCLN and CD81, and upregulated CD5 without modifying SR-B1 expression. Overall, while no association between SR-B1, CLDN-1 or CLDN-6 and the susceptibility to HCV was found, CD5 and CD81 expression coincided with virus lymphotropism and that of OCLN with permissiveness of T cell lines but unlikely primary T cells. This study narrowed the range of factors potentially utilized by HCV to infect T lymphocytes amongst those uncovered using laboratory HCV and Huh7.5 cells. Together with the demonstrated role for CD5 in HCV lymphotropism, the findings indicate that virus utilizes different molecules to enter hepatocytes and lymphocytes.

• CD36 Antigens/genetics
• CD36 Antigens/metabolism
• CD5 Antigens/genetics
• CD5 Antigens/metabolism
• Cell Line
• Claudins/genetics
• Claudins/metabolism
• Gene Expression Regulation
• Gene Knockdown Techniques
• Hepacivirus/physiology
• Hepatitis C/genetics
• Hepatitis C/virology
• Humans
• Jurkat Cells
• Leukocytes, Mononuclear/metabolism
• Leukocytes, Mononuclear/virology
• Occludin/genetics
• Occludin/metabolism
• Receptors, Virus/genetics
• Receptors, Virus/metabolism
• T-Lymphocyte Subsets/metabolism
• T-Lymphocyte Subsets/virology
• Tetraspanin 28/genetics
• Tetraspanin 28/metabolism

• 77544-1 CIHR
• 91665-1 CIHR
• MOP-77544 CIHR

• Hepacivirus/physiology
• Hepatitis C/virology
• Hepatocytes/virology
• Host-Pathogen Interactions
• Humans
• Virus Internalization