Macrophages exploit the mannose receptor and JAK-STAT1-MHC-II pathway to drive antigen presentation and the antimycobacterial immune response after BCG vaccination.

Tuberculosis (TB), caused by Mycobacterium tuberculosis ( M. tb), remains one of the leading causes of fatal infectious diseases worldwide. The only licensed vaccine, Mycobacterium bovis Bacillus Calmette-Guérin (BCG), has variable efficacy against TB in adults. Insufficiency of immune cell function diminishes the protective effects of the BCG vaccine. It is critical to clarify the mechanism underlying the antimycobacterial immune response during BCG vaccination. Macrophage mannose receptor (MR) is important for enhancing the uptake and processing of glycoconjugated antigens from pathogens for presentation to T cells, but the roles of macrophage MR in the BCG-induced immune response against M. tb are not yet clear. Here, we discover that macrophage MR deficiency impairs the antimycobacterial immune response in BCG-vaccinated mice. Mechanistically, macrophage MR triggers JAK-STAT1 signaling, which promotes antigen presentation via upregulated MHC-II and induces IL-12 production by macrophages, contributing to CD4 + T cell activation and IFN-γ production. MR deficiency in macrophages reduces the vaccine efficacy of BCG and increases susceptibility to M. tb H37Ra challenge in mice. Our results suggest that MR is critical for macrophage antigen presentation and the antimycobacterial immune response to BCG vaccination and offer valuable guidance for the preventive strategy of BCG immunization.

PE_PGRS: Vital proteins in promoting mycobacterial survival and modulating host immunity and metabolism.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (M. tb), is the leading infectious cause of mortality worldwide. One of the key reasons for M. tb pathogenesis is the capability of M. tb to evade immune elimination and survive in macrophage, eventually causing chronic infection. However the pathogenicity mechanism of M. tb is not unclear yet, and thus diagnosis and therapy for TB remains a challenge. The genome of M. tb, encodes a unique protein family known as the PGRS family, with largely unexplored functions. Recently, an increasing number of reports have shown that the PE_PGRS proteins play critical roles in bacterial pathogenesis and immune evasion. The PE_PGRS protein family, characterized by a special N-terminal PE (Pro (P)-Glu (E) motif) domain and a C-terminal PGRS (Polymorphic GC-rich Repetitive Sequences) domain, is restricted mainly to pathogenic mycobacteria. Here we summarize current literature on the PE_PGRS as vital proteins in promoting bacterial survival and modulating host immunity, cell death and metabolism. We also highlight the potential of PE_PGRS as novel targets of anti-mycobacterial interventions for TB control.

A Novel Imidazolinone Derivative with 5-Arylidene and 2-Substituted Mercapto Group as Potent Anti-Hepatitis C Virus Compound.

Here six novel imidazolinone derivatives have been synthesized and the compound 4b containing 5-para-methoxy-phenylidene and 2-thioalkylation terminal substitution with 3'-cyano-2',6'-dimethylphenyl showed the best anti-HCV activity and the lowest cytotoxicity. Selectivity index (SI=CC50 /IC50 ) for the 4b was determined as 36, indicating that compound 4b was highly selective towards HCV.

Development of Near-Infrared Nucleic Acid Mimics of Fluorescent Proteins for In Vivo Imaging of Viral RNA with Turn-On Fluorescence.

GFP-like fluorescent proteins and their molecular mimics have revolutionized bioimaging research, but their emissions are largely limited in the visible to far-red region, hampering the in vivo applications in intact animals. Herein, we structurally modulate GFP-like chromophores using a donor-acceptor-acceptor (D-A-A') molecular configuration to discover a set of novel fluorogenic derivatives with infrared-shifted spectra. These chromophores can be fluorescently elicited by their specific interaction with G-quadruplex (G4), a unique noncanonical nucleic acid secondary structure, via inhibition of the chromophores' twisted-intramolecular charge transfer. This feature allows us to create, for the first time, FP mimics with tunable emission in the near-infrared (NIR) region (Emmax = 664-705 nm), namely, infrared G-quadruplex mimics of FPs (igMFP). Compared with their FP counterparts, igMFPs exhibit remarkably higher quantum yields, larger Stokes shift, and better photostability. In a proof-of-concept application using pathogen-related G4s as the target, we exploited igMFPs to directly visualize native hepatitis C virus (HCV) RNA genome in living cells via their in situ formation by the chromophore-bound viral G4 structure in the HCV core gene. Furthermore, igMFPs are capable of high contrast HCV RNA imaging in living mice bearing a HCV RNA-presenting mini-organ, providing the first application of FP mimics in whole-animal imaging.

Neutralization of IL-10 produced by B cells promotes protective immunity during persistent HCV infection in humanized mice.

Chronic HCV infection can lead to cirrhosis and is associated with increased mortality. Interleukin (IL)-10-producing B cells (B10 cells) are regulatory cells that suppress cellular immune responses. Here, we aimed to determine whether HCV induces B10 cells and assess the roles of the B10 cells during HCV infection. HCV-induced B10 cells were enriched in CD19hi and CD1dhi CD5+ cell populations. HCV predominantly triggered the TLR2-MyD88-NF-κB and AP-1 signaling pathways to drive IL-10 production by B cells. In a humanized murine model of persistent HCV infection, to neutralize IL-10 produced by B10 cells, mice were treated with pcCD19scFv-IL-10R, which contains the genes coding the anti-CD19 single-chain variable fragment (CD19scFv) and the extracellular domain of IL-10 receptor alpha chain (sIL-10Ra). This treatment resulted in significant reduction of B10 cells in spleen and liver, increase of cytotoxic CD8+ T-cell responses against HCV, and low viral loads in infected humanized mice. Our results indicate that targeting B10 cells via neutralization of IL-10 may offer a novel strategy to enhance anti-HCV immunotherapy.

Binding of cellular nucleolin with the viral core RNA G-quadruplex structure suppresses HCV replication.

Hepatitis C virus (HCV) infection is a major cause of human chronic liver disease and hepatocellular carcinoma. G-quadruplex (G4) is an important four-stranded secondary structure of nucleic acids. Recently, we discovered that the core gene of HCV contains a G4 RNA structure; however, the interaction between the HCV core RNA G4 and host cellular proteins, and the roles of the HCV core RNA G4 in HCV infection and pathogenesis remain elusive. Here, we identified a cellular protein, nucleolin (NCL), which bound and stabilized the HCV core RNA G4 structure. We demonstrated the direct interaction and colocalization between NCL and wild-type core RNA G4 at both in vitro and in cell physiological conditions of the alive virus; however no significant interaction was found between NCL and G4-modified core RNA. NCL is also associated with HCV particles. HCV infection induced NCL mRNA and protein expression, while NCL suppressed wild-type viral replication and expression, but not G4-modified virus. Silencing of NCL greatly enhanced viral RNA replication. Our findings provide new insights that NCL may act as a host factor for anti-viral innate immunity, and binding of cellular NCL with the viral core RNA G4 structure is involved in suppressing HCV replication.

The Role of Glycosylation in Infectious Diseases.

Glycosylation plays an important role in infectious diseases. Many important interactions between pathogens and hosts involve their carbohydrate structures (glycans). Glycan interactions can mediate adhesion, recognition, invasion, and immune evasion of pathogens. To date, changes in many protein N/O-linked glycosylation have been identified as biomarkers for the development of infectious diseases and cancers. In this review, we will discuss the principal findings and the roles of glycosylation of both pathogens and host cells in the context of human important infectious diseases. Understanding the role and mechanism of glycan-lectin interaction between pathogens and hosts may create a new paradigm for discovering novel glycan-based therapies that can lead to eradication or functional cure of pathogens infection.

Quantitative analysis of serum-based IgG agalactosylation for tuberculosis auxiliary diagnosis.

Tuberculosis (TB) is the leading infectious cause of mortality worldwide, especially in developing countries. However, effective means for TB diagnosis, especially for bacillus-negative (Bn) TB laboratory diagnosis, are urgently needed. In the present study, serum IgG from each tuberculosis patients and healthy controls was purified using affinity chromatography. The samples were then analyzed using mass spectrometry (MS) and ultraperformance liquid chromatography (UPLC) methods. We quantitatively assessed the changes of serum IgG galactosylation in 567 human serum samples including 377 pulmonary TB patients and 190 healthy donors (HDs). We found significantly more agalactosylated (G0) vs monogalactosylated (G1) and digalactosylated (G2) N-glycans of IgG in TB patients, including smear-negative TB patients, than in HDs. The detection rate of TB diagnostic performance by MS for IgG-Gal ratio G0/(G1 + G2 × 2) is 90.48% for bacillus-positive (Bp) and 73.16% for Bn TB patients. Further, combination of MS method with other routine laboratory TB diagnostic methods significantly increased the detection rate to 91.01%-98.39%. Similar results were observed in Mycobacterium tuberculosis (M. tb) infection mouse models. The decrease in galactosylation of IgG in TB patients was also qualitatively confirmed using specific lectin blot assay. Using the above techniques, we can discriminate the content of IgG G0 with terminal N-acetylglucosamine and IgG-Gal ratio G0/(G1 + G2 × 2) between TB patients and HDs. Our data suggest that quantitative analysis of serum-based IgG-Gal ratio G0/(G1 + G2 × 2) could be used for TB auxiliary diagnosis with high effectiveness and feasibility and its combination with other routine laboratory TB diagnostic methods could remarkably improve the detection rate.

Mannose-capped lipoarabinomannan-induced B10 cells decrease severity of dextran sodium sulphate-induced inflammatory bowel disease in mice.

Inflammatory bowel disease (IBD) is a chronic, non-specific, inflammatory gastrointestinal disease that mainly consists of Crohn's disease and ulcerative colitis. However, the aetiology and pathogenesis of IBD are still unclear. B10 (IL-10 producing regulatory B) cells, a subset of regulatory B cells, are known to contribute to intestinal homeostasis and the aberrant frequency of B10 cells is associated with IBD. We have recently reported that B10 cells can be induced by ManLAM (mannose-capped lipoarabinomannan), a major cell-wall lipoglycan of M tb (Mycobacterium tuberculosis). In the current study, the ManLAM-induced B10 cells were adoptively transferred into IL(interleukin)-10-/- mice and the roles of ManLAM-induced B10 cells were investigated in DSS (dextran sodium sulphate)-induced IBD model. ManLAM-induced B10 cells decrease colitis severity in the mice. The B10 cells downregulate Th1 polarization in spleen and MLNs (mesenteric lymph nodes) of DSS-treated mice. These results suggest that IL-10 production by ManLAM-treated B cells contributes to keeping the balance between CD4+ T cell subsets and protect mice from DSS-induced IBD.

Lighting up the Native Viral RNA Genome with a Fluorogenic Probe for the Live-Cell Visualization of Virus Infection.

RNA viruses represent a major global health threat, and the visualization of their RNA genome in infected cells is essential for virological research and clinical diagnosis. Due to the lack of chemical toolkits for the live-cell imaging of viral RNA genomes, especially native viral genomes without labeling and genetic modification, studies on native virus infection at the single-live-cell level are challenging. Herein, taking hepatitis C virus (HCV) as a representative RNA virus, we propose that the innate noncanonical G-quadruplex (G4) structure of viral RNA can serve as a specific imaging target and report a new benzothiazole-based G4-targeted fluorescence light-up probe, ThT-NE, for the direct visualization of the native RNA genome of HCV in living host cells. We demonstrate the use of the ThT-NE probe for several previously intractable applications, including the sensitive detection of individual virus-infected cells by small-molecule staining, real-time monitoring of the subcellular distribution of the viral RNA genome in live cells, and continuous live-cell tracking of the infection and propagation of clinically isolated native HCV. The fluorogenic-probe-based viral RNA light-up system opens up a promising chemical strategy for cutting-edge live-cell viral analysis, providing a potentially powerful tool for viral biology, medical diagnosis, and drug development.

Tumor-Derived Exosomal Long Noncoding RNAs as Promising Diagnostic Biomarkers for Prostate Cancer.

BACKGROUND/AIMS: Exosomal circulating long non-coding RNAs (lncRNAs) in blood are emerging as clinically useful and non-invasive biomarkers for tumor diagnosis. However, normal cells can also secrete exosomes, so it is a prerequisite to obtain tumor-derived exosomes for better understanding of their diagnostic impacts in cancer. In this study, a dual-antibody-functionalized immunoaffinity system was established to isolate exosomes and investigate their lncRNAs expression pattern and clinical significance in prostate cancer (PCa). METHODS: A commercially available kit was used to isolate total exosomes, which were then purified by a dual-antibody-functionalized immunoaffinity system. RT-qPCR was performed to detect the expression of exosomal lncRNAs. Receiver operating characteristic (ROC) curves were plotted to assess the diagnostic value. RESULTS: Expression levels of two lncRNAs in tumor-derived exosomes were significantly higher than those in total exosomes. The levels of SAP30L-AS1 were upregulated in benign prostatic hyperplasia (BPH), and SChLAP1 levels were significantly higher in PCa than in BPH and healthy individuals. The area under the ROC curve indicated that SAP30L-AS1 and SChLAP1 had adequate diagnostic value to distinguish PCa from controls. Two lncRNAs separately combined with prostate specific antigen (PSA) possessed a moderate ability for discrimination. SAP30L-AS1 expression level was related to PSA values and tumor invasion. SChLAP1 expression was significantly higher in patients with higher Gleason scores, and was also effective in differentiating between BPH and PCa when the concentration of PSA was in the gray zone. CONCLUSION: The isolation of tumor-derived exosomes by dual-antibody-functionalized immunoaffinity systems and detection of their lncRNAs in plasma may lead to the identification of suitable biomarkers, with potential diagnostic utility.

A strategy of targeting B10 cell by CD19scFv-IL10R for tumor therapy.

IL-10 producing B (B10) cells, a subset of regulatory B (Breg) cells, produce IL-10 and play immunosuppressive roles in antitumor immunity. B10 cells are associated with enhanced tumor-aggressiveness and a poorer prognosis. To specifically inhibit the IL-10 secreted by B cells, we constructed the recombinant plasmid pcCD19scFv-IL10R, which contained the gene of anti-CD19 single-chain variable fragment (CD19scFv) and the extracellular domain of IL-10R1. Soluble CD19scFv-IL10R protein was identified in vitro and in vivo after the cells were transfected with pcCD19scFv-IL10R plasmid or the mice were injected with the plasmid. The fusion protein had the bispecific ability to target both IL-10 and CD19 molecules in vitro. Intramuscularly (i.m.) injecting mice with pcCD19scFv-IL-10R plasmid inhibited hepatocellular carcinoma growth in vivo. Mice treated with pcCD19scFv-IL-10R showed a significant reduction in B10 cells and regulatory T (Treg) cells, but an increase in the anti-tumor Th1 immune response and the cytotoxic CD8+ T cell response. Thus, targeting B10 cells by CD19scFv-IL10R molecule may offer a new avenue for tumor therapy.

The ubiquitin ligase RNF5 regulates antiviral responses by mediating degradation of the adaptor protein MITA.

Viral infection activates transcription factors NF-kappaB and IRF3, which collaborate to induce type I interferons (IFNs) and elicit innate antiviral response. MITA (also known as STING) has recently been identified as an adaptor that links virus-sensing receptors to IRF3 activation. Here, we showed that the E3 ubiquitin ligase RNF5 interacted with MITA in a viral-infection-dependent manner. Overexpression of RNF5 inhibited virus-triggered IRF3 activation, IFNB1 expression, and cellular antiviral response, whereas knockdown of RNF5 had opposite effects. RNF5 targeted MITA at Lys150 for ubiquitination and degradation after viral infection. Both MITA and RNF5 were located at the mitochondria and endoplasmic reticulum (ER) and viral infection caused their redistribution to the ER and mitochondria, respectively. We further found that virus-induced ubiquitination and degradation of MITA by RNF5 occurred at the mitochondria. These findings suggest that RNF5 negatively regulates virus-triggered signaling by targeting MITA for ubiquitination and degradation at the mitochondria.

N-glycosylation-mutated HCV envelope glycoprotein complex enhances antigen-presenting activity and cellular and neutralizing antibody responses.

BACKGROUND: The development of an efficient vaccine and broadly cross-neutralizing antibodies of hepatitis C virus (HCV) remains a priority. The heavily glycosylated viral envelope glycoprotein E1E2 complex is a candidate vaccine antigen. Bacteria-derived unmethylated CpG DNA, a potent stimulator of immune cells, is important for vaccine research. METHODS: Here, the immunogenicities of wild type (WT) E1E2, five N-glycosylation site mutated E1E2 glycoproteins, and five CpG-coupled E1E2 N-glycosylation mutated glycoproteins were analyzed in BALB/c mice by DNA vaccination using in vivo electroporation. RESULTS: The E1E2 protein expression levels were examined and shown to be unaffected by these N-glycosylation mutations. We found that a CpG-coupled E1-N209D-E2-N430D DNA vaccine (named CpG-E1E2-M4) induced the highest cellular immune response compared to the WT E1E2, CpG-E1E2, and other mutants. Furthermore, the CpG-E1E2-M4 anti-serum effectively neutralized the infection of cell-cultured HCV (HCVcc, genotype 2a)- and HCV pseudo particles (HCVpp, genotypes 1 to 7) to Huh-7.5.1 hepatocytes. Additionally, CpG-E1E2-M4 enhanced the Interleukin-12 (IL-12) production and antigen-presenting activity of CD11c(+) dendritic cells (DCs) by inducing CD4(+) Th1 polarization and the production of perforin and granzyme B (GrB) in CD8(+) T cells. CONCLUSIONS: As our knowledge this is the first study revealing that the naturally poor immunogenicity of E1E2 can be enhanced by the deletion of N-glycans combined with the addition of immune activator CpG by DNA vaccination. GENERAL SIGNIFICANCE: Deletion of N-glycans can enhance viral immunogenicity. The selected CpG-E1E2-M4 mutant is a novel potential HCV DNA vaccine that elicits enhanced CD4(+) Th1 and CD8(+) T cell responses and neutralizing antibody production against HCV infection. This article is part of a Special Issue entitled "Glycans in personalised medicine" Guest Editor: Professor Gordan Lauc.

Ficolin-A/2, acting as a new regulator of macrophage polarization, mediates the inflammatory response in experimental mouse colitis.

Human ficolin-2 (FCN-2) and mouse ficolin-A (FCN-A, a ficolin-2-like molecule in mouse) are activators of the lectin complement pathway, present in normal plasma and usually associated with infectious diseases, but little is known about the role of FCN-A/2 in inflammatory bowel disease (IBD). In our present study, we found that patients with IBD exhibited much higher serum FCN-2 levels than healthy controls. In the dextran sulphate sodium-induced acute colitis mouse model, FCN-A knockout mice showed much milder disease symptoms with less histological damage, lower expression levels of pro-inflammatory cytokines [interleukin-6 (IL-6), IL-1ß and tumour necrosis factor-α (TNF-α)], chemokines (CXCL1/2/10 and CCL4) and higher levels of the anti-inflammatory cytokine IL-10 compared with wild-type mice. We demonstrated that FCN-A/2 exacerbated the inflammatory pathogenesis of IBD by stimulating M1 polarization through the TLR4/MyD88/MAPK/NF-κB signalling pathway in macrophages. Hence, our data suggest that FCN-A/2 may be used as a novel therapeutic target for IBD.

Ficolin-2 triggers antitumor effect by activating macrophages and CD8+ T cells.

Ficolin-2 is an important serum complement lectin. Here, we describe novel findings indicating that serum ficolin-2 concentrations in multiple tumor patients are significantly lower than those in healthy donors. Administration of exogenous ficolin-2 or ficolin-A (a ficolin-2-like molecule in mouse), with only once, could remarkably inhibit the tumor cells growth in murine tumor models via early macrophages, dendritic cells (DCs) and CD8+ T cells, but not CD4+ T cells. Ficolin-A (FCN-A) knockout (KO) mice exhibits significantly increased tumor cell growth. Ficolin-2 induces macrophage activation, promotes M1 polarization and facilitates proliferation and antigen-specific cytotoxicity of CD8+ T cells. Ficolin-2 binds to Toll-like receptor 4 (TLR4) on macrophages and DCs and promotes their antigen-presenting abilities to CD8+ T cells. Our findings provide a new therapeutic strategy for tumors based on the triggering of immune-mediated antitumor effect by ficolin-2.

Alteration of N-glycan expression profile and glycan pattern of glycoproteins in human hepatoma cells after HCV infection.

BACKGROUND: Hepatitis C virus (HCV) infection causes chronic liver diseases, liver fibrosis and even hepatocellular carcinoma (HCC). However little is known about any information of N-glycan pattern in human liver cell after HCV infection. METHODS: The altered profiles of N-glycans in HCV-infected Huh7.5.1 cell were analyzed by using mass spectrometry. Then, lectin microarray, lectin pull-down assay, reverse transcription-quantitative real time PCR (RT-qPCR) and western-blotting were used to identify the altered N-glycosylated proteins and glycosyltransferases. RESULTS: Compared to uninfected cells, significantly elevated levels of fucosylated, sialylated and complex N-glycans were found in HCV infected cells. Furthermore, Lens culinaris agglutinin (LCA)-binding glycoconjugates were increased most. Then, the LCA-agarose was used to precipitate the specific glycosylated proteins and identify that fucosylated modified annexin A2 (ANXA2) and heat shock protein 90 beta family member 1 (HSP90B1) was greatly increased in HCV-infected cells. However, the total ANXA2 and HSP90B1 protein levels remained unchanged. Additionally, we screened the mRNA expressions of 47 types of different glycosyltransferases and found that α1,6-fucosyltransferase 8 (FUT8) was the most up-regulated and contributed to strengthen the LCA binding capability to fucosylated modified ANXA2 and HSP90B1 after HCV infection. CONCLUSIONS: HCV infection caused the altered N-glycans profiles, increased expressions of FUT8, fucosylated ANXA2 and HSP90B1 as well as enhanced LCA binding to Huh7.5.1. GENERAL SIGNIFICANCE: Our results may lay the foundation for clarifying the role of N-glycans and facilitate the development of novel diagnostic biomarkers and therapeutic targets based on the increased FUT8, fucosylated ANXA2 and HSP90B1 after HCV infection.

A single-stranded DNA aptamer against mannose-capped lipoarabinomannan enhances anti-tuberculosis activity of macrophages through downregulation of lipid-sensing nuclear receptor peroxisome proliferator-activated receptor γ expression.

Mannose-capped lipoarabinomannan (ManLAM) is an immunomodulatory epitope of Mycobacterium tuberculosis (Mtb). An aptamer (ZXL1) that specifically binds to ManLAM from the virulent Mtb H37Rv strain was previously generated and it was found that ZXL1 functions as an antagonist, inhibiting the ManLAM-induced immunosuppression of DCs. In the present study, it was found that ZXL1 inhibits Mtb entry into murine macrophages and that ZXL1 enhances IL-1ß and IL-12 mRNA expression and cytokine production in ManLAM-treated macrophages but decreases IL-10 production. Inducible nitric oxide synthase expression in macrophages was upregulated in the presence of ZXL1 after stimulation with ManLAM. ZXL1 was also found to inhibit expression of lipid-sensing nuclear receptor peroxisome proliferator-activated receptor γ (PPAR-γ). These results suggest that ZXL1 promotes anti-tuberculosis activity through downregulation of PPAR-γ expression, which may contribute to M1 macrophage polarization and Mtb killing by macrophages.

A Single ssDNA Aptamer Binding to Mannose-Capped Lipoarabinomannan of Bacillus Calmette-Guérin Enhances Immunoprotective Effect against Tuberculosis.

Because Mycobacterium bovis, termed bacillus Calmette-Guérin (BCG), the only available used tuberculosis (TB) vaccine, retains immunomodulatory properties that limit its protective immunogenicity, there are continuous efforts to identify the immunosuppression mechanism as well as new strategies for improving the immunogenicity of BCG. Here, an ssDNA aptamer "antibody" BM2 specifically bound to the mannose-capped lipoarabinomannan (ManLAM) of BCG was selected. BM2 significantly blocked ManLAM-mannose receptor (MR) binding, triggered ManLAM-CD44 signaling, and enhanced M1 macrophage and Th1 activation via cellular surface CD44 in vitro and in vivo. BM2 enhanced immunoprotective effects of BCG against virulent Mycobacterium tuberculosis H37Rv infection in mice and monkeys models. Thus, we report a new mechanism of the interaction between ManLAM and CD44 on macrophages and CD4(+) T cells and reveal that ManLAM-binding membrane molecule CD44 is a novel target for the enhancement of BCG immunogenicity, and BM2 has strong potential as an immune enhancer for BCG.

Ficolin-2 inhibits hepatitis C virus infection, whereas apolipoprotein E3 mediates viral immune escape.

Human ficolin-2 (L-ficolin/p35) is a lectin-complement pathway activator that is present in normal human plasma and is associated with infectious diseases; however, little is known regarding the roles and mechanisms of ficolin-2 during chronic hepatitis C virus (HCV) infection. In this study, we found that ficolin-2 inhibits the entry of HCV at an early stage of viral infection, regardless of the viral genotype. Ficolin-2 neutralized and inhibited the initial attachment and infection of HCV by binding to the HCV envelope surface glycoproteins E1 and E2, blocking HCV attachment to low-density lipoprotein receptor (LDLR) and scavenger receptor B1, and weakly interfering with CD81 receptor attachment. However, no interference with claudin-1 and occludin receptor attachment was observed. The C-terminal fibrinogen domain (201-313 aa) of ficolin-2 was identified as the critical binding region for the HCV-E1-E2 N-glycans, playing a critical role in the anti-HCV activity. More importantly, we found that apolipoprotein E (ApoE)3, which is enriched in the low-density fractions of HCV RNA-containing particles, promotes HCV infection and inhibits ficolin-2-mediated antiviral activity. ApoE3, but not ApoE2 and ApoE4, blocked the interaction between ficolin-2 and HCV-E2. Our data suggest that the HCV entry inhibitor ficolin-2 is a novel and promising antiviral innate immune molecule, whereas ApoE3 blocks the effect of ficolin-2 and mediates an immune escape mechanism during chronic HCV infection. HCV may be neutralized using compounds directed against the lipoprotein moiety of the viral particle, and ApoE3 may be a new target to combat HCV infection.