title: Exploring innate immune induction and interference mechanisms of Hepatitis A and C Viruses in human hepatocytes: a comparative analysis
creator: Colasanti, Ombretta
subject: ddc-570
subject: 570 Life sciences
subject: ddc-610
subject: 610 Medical sciences Medicine
description: This dissertation focuses on the host innate immune response in the context of hepatitis A virus (HAV) and hepatitis C virus (HCV) infections in human hepatocytes.   Innate immunity is the body's first line of defense against invading pathogens, functioning via a network of cells and molecules that quickly respond to infections. Central to this response are pattern recognition receptors (PRRs), which recognize pathogen-associated molecular patterns (PAMPs) and trigger the production of Interferons (IFNs) and the expression of Interferon-Stimulated Genes (ISGs), critical to antiviral defenses. In the case of viral infections like HAV and HCV, the host's immune response plays a pivotal role in disease outcome, and these viruses, in turn, have developed strategies to evade and manipulate the immune system. Thus, the interplay between viruses and the host's immune response is a key aspect of viral pathogenesis.  HAV and HCV are both positive-strand RNA (+ssRNA) hepatotropic viruses with a strict human tropism. Despite sharing a number of similarities in terms of genome structure and replication, they have unique characteristics and induce different immune responses, with HAV being cleared and HCV establishing chronic infections in >70% of the cases, making them intriguing subjects for comparative studies.   Notably, the only comparative in vivo study conducted on HAV and HCV-infected chimpanzees revealed that HAV, albeit replicating to a higher degree than HCV, barely elicited an innate immune response, as opposed to that solidly mounted upon HCV infection. This was attributed to a potent counteractive activity by the HAV protease 3C, and their precursors 3CD and 3ABC, towards the PRRs pathways. Therefore, I aimed at elucidating how HAV and HCV triggered, and counteracted, the intrinsic immune response in the liver, potentially associating these mechanisms to their strikingly different infection outcomes.   To this aim, I employed relevant cell culture models to examine the efficiency with which HAV and HCV were sensed by endoplasmic Toll-Like-Receptor 3 (TLR3) and the cytoplasmic Rig-I-Like-Receptors (RLRs). I furthermore quantified the proteolytical cleavage of the TLR3 adaptor TIR-domain-containing adapter-inducing interferon-β (TRIF) and the RLR adaptor Mitochondrial Antiviral-Signaling protein (MAVS) via transient and stable expression of the respective viral proteases. My results indicated that both HAV and HCV infections induced similar levels of ISGs and IFNs in human hepatocytes, signaling the activation of innate immunity. HAV sensing primarily involved Melanoma Differentiation-Associated protein 5 (MDA5) and Laboratory of Genetics and Physiology 2 (LGP2), with limited interference to MAVS and to TRIF by the HAV proteases 3ABC and 3CD. In contrast, HCV sensing by Retinoic acid-Inducible Gene I (RIG-I) and MDA5 was entirely blocked by the efficient cleavage of the RLRs adaptor protein MAVS, leading to the activation of TLR3 as the primary source of ISG response for HCV. TRIF, the adaptor protein for TLR3, was not cleaved by HCV NS3-4A, contrary to previous reports. In addition, my research found that Huh7 cells were found to be defective on the MDA5-LGP2 axis.   My approach then shifted to an in vivo examination of SCID Alb-uPA humanized mice infected with patient-derived as well as cell culture adapted HAV and HCV, where I observed that both HAV and HCV triggered ISGs to similar extents, confirming the results obtained in vitro.   Overall, my results suggest that while HCV primarily triggers a TLR3-mediated response, HAV relies on an MDA5-mediated sensing mechanism, which is not sufficiently blocked. This challenges the established understanding that HAV does not induce IFN, or fully inhibits them, in infected hepatocytes both in vitro and in vivo. However, caution must be applied when interpreting both the results in vitro, due to models with limited physiological relevance, and in vivo, due to a limited number of animals. These findings underscore the need for fully immunocompetent animal models to carry out comprehensive investigations of the determinants of viral persistence and clearance, which are currently lacking.
date: 2024
type: Dissertation
type: info:eu-repo/semantics/doctoralThesis
type: NonPeerReviewed
format: application/pdf
identifier: https://archiv.ub.uni-heidelberg.de/volltextserverhttps://archiv.ub.uni-heidelberg.de/volltextserver/34625/1/Colasanti_PhDThesis_HeidelbergUniversity.pdf
identifier: DOI:10.11588/heidok.00034625
identifier: urn:nbn:de:bsz:16-heidok-346252
identifier:   Colasanti, Ombretta  (2024) Exploring innate immune induction and interference mechanisms of Hepatitis A and C Viruses in human hepatocytes: a comparative analysis.  [Dissertation]     
relation: https://archiv.ub.uni-heidelberg.de/volltextserver/34625/
rights: info:eu-repo/semantics/openAccess
rights: Please see front page of the work (Sorry, Dublin Core plugin does not recognise license id)
language: eng