Jessica Fioravanti

Immunosurveillance of the Liver by Intravascular Effector CD8+ T Cells

Effector CD8(+) T cells (CD8 TE) play a key role during hepatotropic viral infections. Here, we used advanced imaging in mouse models of hepatitis B virus (HBV) pathogenesis to understand the mechanisms whereby these cells home to the liver, recognize antigens, and deploy effector functions. We show that circulating CD8 TE arrest within liver sinusoids by docking onto platelets previously adhered to sinusoidal hyaluronan via CD44. After the initial arrest, CD8 TE actively crawl along liver sinusoids and probe sub-sinusoidal hepatocytes for the presence of antigens by extending cytoplasmic protrusions through endothelial fenestrae. Hepatocellular antigen recognition triggers effector functions in a diapedesis-independent manner and is inhibited by the processes of sinusoidal defenestration and capillarization that characterize liver fibrosis. These findings reveal the dynamic behavior whereby CD8 TE control hepatotropic pathogens and suggest how liver fibrosis might reduce CD8 TE immune surveillance toward infected or transformed hepatocytes.

Eradication of large tumors expressing human papillomavirus E7 protein by therapeutic vaccination with E7 fused to the extra domain a from fibronectin.

Cervical carcinoma is one of the most common cancers in women worldwide. It is well established that chronic infection of the genital tract by various mucosatropic human papillomavirus (HPV) types causes cervical cancer. Cellular immunity to E7 protein from HPV (HPVE7) has been associated with clinical and cytologic resolution of HPV‐induced lesions. Thus, we decided to test if targeting of HPVE7 to dendritic cells using a fusion protein containing the extra domain A (EDA) from fibronectin, a natural ligand for TLR4, and HPVE7 (EDA‐HPVE7) might be an efficient vaccine for the treatment of cervical carcinoma. We found that EDA‐HPVE7 fusion protein was efficiently captured by bone marrow derived dendritic cells in vitro and induced their maturation, with the upregulation of maturation markers and the production of IL‐12. Immunization of mice with EDA‐HPVE7 fusion protein induced antitumor CD8+ T cell responses in the absence of additional adjuvants. Repeated intratumoral administration of EDA‐HPVE7 in saline was able to cure established TC‐1 tumors of 5–7 mm in diameter. More importantly, intravenous injection with EDA‐HPVE7 in combination with the TLR ligand polyinosinic‐polycytidylic acid (pIC), or with low doses of cyclophosphamide and the TLR9 ligand CpG‐B complexed in cationic lipids, were able to eradicate large established TC‐1 tumors (1.2 cm in diameter). Thus, therapeutic vaccination with EDA‐HPVE7 fusion protein may be effective in the treatment of human cervical carcinoma.

Inflammatory monocytes hinder antiviral B cell responses

LCMV evades B cell responses by recruiting inflammatory monocytes to draining lymph nodes. See related Research Articles by Fallet et al. and Moseman et al. and a Focus by Laidlaw et al. B cells hoisted by their own petard: IFN-I Certain pathogens, including HIV and hepatitis viruses, that lead to persistent infections are often associated with suboptimal antibody responses. Using lymphocytic choriomeningitis virus infection in mice, Fallet et al., Moseman et al., and Sammicheli et al. report that up-regulation of type I interferon (IFN-I) in the early phase of infection is a key contributor to premature deletion of virus-specific B cells. Blockade of IFN-I prevents B cell deletion. Although the studies agree that IFN-I does not act directly on B cells, they found that distinct immune cells mediate IFN-I–dependent deletion of B cells, depending on the system examined. Targeting of the IFN-I pathway could be used to restore B cell responses during persistent viral infections in humans. Antibodies are critical for protection against viral infections. However, several viruses, such as lymphocytic choriomeningitis virus (LCMV), avoid the induction of early protective antibody responses by poorly understood mechanisms. We analyzed the spatiotemporal dynamics of B cell activation to show that, upon subcutaneous infection, LCMV-specific B cells readily relocate to the interfollicular and T cell areas of draining lymph nodes, where they extensively interact with CD11b+Ly6Chi inflammatory monocytes. These myeloid cells were recruited to lymph nodes draining LCMV infection sites in a type I interferon– and CCR2-dependent fashion, and they suppressed antiviral B cell responses by virtue of their ability to produce nitric oxide. Depletion of inflammatory monocytes, inhibition of their lymph node recruitment, or impairment of their nitric oxide–producing ability enhanced LCMV-specific B cell survival and led to robust neutralizing antibody production. Our results identify inflammatory monocytes as critical gatekeepers that restrain antiviral B cell responses and suggest that certain viruses take advantage of these cells to prolong their persistence within the host.

Scavenger receptor class B, type I: a promising immunotherapy target

Scavenger receptor class B, type I (SR-BI) is a crucial molecule in lipid metabolism, since the interaction of high-density lipoproteins (HDLs) with SR-BI is involved in reverse cholesterol transport and cholesterol efflux. Recent findings also underscore a critical role of SR-BI in antimicrobial and immune responses. SR-BI is not only highly expressed in liver and steroidogenic glands, but also in endothelial cells, macrophages and dendritic cells. SR-BI mainly mediates anti-inflammatory responses, which may be altered by dysfunctional HDLs produced in several diseases. Moreover, SR-BI has been involved in the capture and cross-presentation of antigens from viruses, bacteria and parasites. It thus works as a pattern-recognition receptor that interacts with both damage-associated molecular patterns and pathogen-associated molecular patterns. These new findings in the microbiology and immunology fields present SR-BI as an unexplored therapeutic target that warrants further basic and applied research.

Effector CD8+ T cell-derived interleukin-10 enhances acute liver immunopathology

Background & Aims Besides secreting pro-inflammatory cytokines, chemokines and effector molecules, effector CD8+ T cells that arise upon acute infection with certain viruses have been shown to produce the regulatory cytokine interleukin (IL)-10 and, therefore, contain immunopathology. Whether the same occurs during acute hepatitis B virus (HBV) infection and role that IL-10 might play in liver disease is currently unknown. Methods Mouse models of acute HBV pathogenesis, as well as chimpanzees and patients acutely infected with HBV, were used to analyse the role of CD8+ T cell-derived IL-10 in liver immunopathology. Results Mouse HBV-specific effector CD8+ T cells produce significant amounts of IL-10 upon in vivo antigen encounter. This is corroborated by longitudinal data in a chimpanzee acutely infected with HBV, where serum IL-10 was readily detectable and correlated with intrahepatic CD8+ T cell infiltration and liver disease severity. Unexpectedly, mouse and human CD8+ T cell-derived IL-10 was found to act in an autocrine/paracrine fashion to enhance IL-2 responsiveness, thus preventing antigen-induced HBV-specific effector CD8+ T cell apoptosis. Accordingly, the use of mouse models of HBV pathogenesis revealed that the IL-10 produced by effector CD8+ T cells promoted their own intrahepatic survival and, thus supported, rather than suppressed liver immunopathology. Conclusion Effector CD8+ T cell-derived IL-10 enhances acute liver immunopathology. Altogether, these results extend our understanding of the cell- and tissue-specific role that IL-10 exerts in immune regulation. Lay summary: Interleukin-10 is mostly regarded as an immunosuppressive cytokine. We show here that HBV-specific CD8+ T cells produce IL-10 upon antigen recognition and that this cytokine enhances CD8+ T cell survival. As such, IL-10 paradoxically promotes rather than suppresses liver disease.

Interferon alpha bioactivity critically depends on Scavenger receptor class B type I function

ABSTRACT Scavenger receptor class B type I (SR-B1) binds pathogen-associated molecular patterns participating in the regulation of the inflammatory reaction but there is no information regarding potential interactions between SR-B1 and the interferon system. Herein, we report that SR-B1 ligands strongly regulate the transcriptional response to interferon α (IFNα) and enhance its antiviral and antitumor activity. This effect was mediated by the activation of TLR2 and TLR4 as it was annulled by the addition of anti-TLR2 or anti-TLR4 blocking antibodies. In vivo, we maximized the antitumor activity of IFNα co-expressing in the liver a SR-B1 ligand and IFNα by adeno-associated viruses. This gene therapy strategy eradicated liver metastases from colon cancer with reduced toxicity. On the other hand, genetic and pharmacological inhibition of SR-B1 blocks the clathrin-dependent interferon receptor recycling pathway with a concomitant reduction in IFNα signaling and bioactivity. This effect can be applied to enhance cancer immunotherapy with oncolytic viruses. Indeed, SR-B1 antagonists facilitate replication of oncolytic viruses amplifying their tumoricidal potential. In conclusion, SR-B1 agonists behave as IFNα enhancers while SR-B1 inhibitors dampen IFNα activity. These results demonstrate that SR-B1 is a suitable pharmacology target to enhance cancer immunotherapy based on IFNα and oncolytic viruses.

Pathogen‐specific B‐cell receptors drive chronic lymphocytic leukemia by light‐chain‐dependent cross‐reaction with autoantigens

Several lines of evidence indirectly suggest that antigenic stimulation through the B‐cell receptor (BCR) supports chronic lymphocytic leukemia (CLL) development. In addition to self‐antigens, a number of microbial antigens have been proposed to contribute to the selection of the immunoglobulins expressed in CLL. How pathogen‐specific BCRs drive CLL development remains, however, largely unexplored. Here, we utilized mouse models of CLL pathogenesis to equip B cells with virus‐specific BCRs and study the effect of antigen recognition on leukemia growth. Our results show that BCR engagement is absolutely required for CLL development. Unexpectedly, however, neither acute nor chronic exposure to virus‐derived antigens influenced leukemia progression. Rather, CLL clones preferentially selected light chains that, when paired with virus‐specific heavy chains, conferred B cells the ability to recognize a broad range of autoantigens. Taken together, our results suggest that pathogens may drive CLL pathogenesis by selecting and expanding pathogen‐specific B cells that cross‐react with one or more self‐antigens.

Kinetic and Dynamic Computational Model-Based Characterization of New Proteins in Mice: Application to Interferon Alpha Linked to Apolipoprotein A-I

Interferon alpha linked to apolipoprotein A-I has been recently proposed as an improved interferon-based therapy. In the present study, we aimed to develop a computational model to gain further insight into the in vivo behaviour of this new fusion protein. In order to facilitate in vivo evaluation of interferon and the fusion protein without altering their biological properties, green fluorescent protein was incorporated into their structures. Kinetic and dynamic behaviour of both compounds was successfully described after plasmid hydrodynamic administration and in situ synthesis of the studied proteins. Results from the modelling exercise showed that apolipoprotein A-I conferred a modified kinetic behaviour, varying molecule distribution and prolonging half-life without altering liver dynamic performance. However, differences in the gene expression activity were observed at brain level between both compounds. Those differences could be explained by modifications in the dynamic, but also in the biodistribution properties, which would be worth evaluating in future experiments. Therefore, the modelling approach provided a global comprehension of a complex system and allowed us to compare the in vivo behaviour of both compounds and to identify critical aspects that might be important to understand the system better and suggests a need for new model-based experiments.