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| Author(s): |
Pierina Lorencini Parise
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Campinas, SP. |
| Institution: | Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia |
| Defense date: | 2024-06-27 |
| Examining board members: |
José Luiz Proença Módena;
Iranaia Assunção Miranda;
Luciana Barros de Arruda;
Luciana Pereira Ruas;
Marielton dos Passos Cunha
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| Advisor: | José Luiz Proença Módena; Carla Cristina Judice |
| Abstract | |
Oropouche virus (OROV) is an emerging arbovirus responsible for thousands of cases of exanthematous febrile illness in the Amazon region. As other arboviruses, OROV can cause neurological complications in a portion of infected patients. Although it has been demonstrated that OROV can cross the blood-brain barrier (BBB) ??and cause infection in the central nervous system (CNS) of neonate mice and hamsters, little is known about the pathogenetic mechanisms associated with the breakdown of this barrier. It has already been demonstrated that IRF-5 (Interferon regulatory factor 5), a transcription factor activated after recognition of PAMPs (pathogen-associated molecular patterns), is essential in controlling the neuroinvasion of OROV and other arboviruses such as WNV and LACV. Therefore, the objective of this project was to characterize the role of IRF-5 expression in endothelial cells in controlling neuroinvasion by OROV and explore the factors associated with viral infection in the CNS. For this, we analyzed mortality, viral tropism and leukocyte infiltration in the CNS of animals that do not express IRF-5 in the endothelium, using the Cre/LoxP recombination system. We also characterized OROV infection and the expression of innate immune response genes and endothelial adhesion factors in established endothelial cell lines. Furthermore, we examined the effect of IRF-5 on the integrity and passage of viruses across the endothelial barrier and through complex models that mimic the BBB. Finally, we used bio-plex and RNA-seq techniques to explore the response of human CNS cells to OROV infection. The results showed that 35% of the conditional knockout animals are vulnerable to infection, showing signs of neurological impairment as loss of balance and limb paralysis, high viral load in the CNS (between 104 and 108 eqFFU/g) and increased number and adhesion of immune cells to the brain microvasculature, even when infection occurs through the transference of circulating infected immune cells. Therefore, we conclude that the expression and activation of IRF-5 in endothelial cells is essential to restrict access to the CNS and the consequent breakdown of the BBB during OROV infection in vivo. Furthermore, OROV productively infects established human (HBEC-5i) and murine (BEND3) endothelial cell lines. While HBEC-5i cells are highly susceptible and permissive to OROV infection, reaching titers as high as 105 and 106 FFU/mL, BEND3 cells are partially resistant to infection reaching a maximun of 104 FFU/mL, with lower extracellular amounts of virus and no apparent cytopathic effect. It was also possible to observe a strong antiviral response in infected endothelial cells, with increased expression of signaling pathways, interferon-stimulated genes (ISGs) and endothelial adhesion factors (ICAM, VCAM, e-selectin and p-selectin). Still, there was no modulation in the expression of tight junctions or disruption of the endothelial barrier in vitro, despite it being possible to recover virus in the lower chamber in transwell models, indicating that the virus is capable of crossing the endothelial barrier without causing cellular damage. Finally, human glial and neuronal cells present an intense response to viral infection, with the release of cytokines, chemokines and modulation of cellular pathways (AU) | |
| FAPESP's process: | 17/26908-0 - The role of IRF-5 in Oropouche virus pathogenesis and neurovirulence |
| Grantee: | Pierina Lorencini Parise |
| Support Opportunities: | Scholarships in Brazil - Doctorate (Direct) |