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| Author(s): |
Ronaldo Bragança Martins Junior
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Ribeirão Preto. |
| Institution: | Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC) |
| Defense date: | 2017-05-11 |
| Examining board members: |
Eurico de Arruda Neto;
Fernando Chahud;
Benedito Antonio Lopes da Fonseca;
Clarisse Martins Machado;
Aripuanã Sakurada Aranha Watanabe
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| Advisor: | Eurico de Arruda Neto |
| Abstract | |
The chronic adenotonsillar diseases are frequent otorhinolaryngologic conditions caused by chronic inflammation of adenoids and palatine tonsils. Rhinovirus (RV) is highly frequently detected in secretions, and tonsillar tissues from patients experience chronic tonsillar hypertrophy without symptoms of ARI, and our goal is to full understanding of viral infections in hypertrophic tonsillar tissues by RV. Of 104 enrolled patients with adenotonsillar chronic diseases, 21.1% (22/104) and 42.3% (44/104) had palatine tonsils and adenoids positive for RV by qPCR, respectively. RV Viral replication was confirmed by in situ hybridizations. Minus-strand RNA were detected in all tested samples (7 tonsils and 9 adenoids), and positive reactions were seen inside and outside of lymphoid follicles from tonsils and adenoids, in the ciliated epithelium of the adenoids and rarely in positive squamous epithelium cells from tonsils. The presence of viral structural protein VP1 and VP2 was detected within and outside of the lymphoid follicles from tonsils and adenoids, and also in epithelial cells from adenoids by immunohistochemistry (IHC). Later, by sequential immuno-peroxidase labelling and erasing protocol (SIMPLE), we saw co-localization of RV VP2 capsid protein staining with CD4 positive cells and CD20 positive cells. We confirmed that RV could infect primary culture of tonsilar mononuclear cells (TMNCs). Additionally, intracellular replication of RV in TMNCs, measured by TCID50 in HeLa cells, had an initial increase in the first 24 hours, and dropped at 48 hours post infection. Immunolocalization staining with anti-RV and TMNCs surface markers indicated that phenotypes of susceptible cells were T-cells both CD4+ and CD20+, but also, we saw co-localization of VP-2 protein with CD8+ cells, CD56+ cells and CD33+ cells. RV-16 couldn\'t infect CD123+ cell in our experiments. Finally, we were able to recover 4 rhinoviruses by inoculating WI-38 fibroblast cells and HeLa cells, confirming by the cytopathic effect and immunofluorescence positive staining with anti-VP1 antibody. Taken together, our results indicate that tonsils and adenoids of patients without ARI may be reservoirs of replicating human rhinovirus, infecting manly Tcells CD4+ and CD20+ B-cells. The high-frequency detection of RNA (-) and VP1 expression in tissues from patients with chronic adenotonsillar diseases, plus the isolation of infectious viral particles, suggests that these detected agents replicate in the adenotonsillar tissues and this specific sites may be important sources of transmission of RV in the community. (AU) | |
| FAPESP's process: | 13/06380-0 - Rhinovirus replication in explants of human tonsillar lymphoepithelial tissue infected ex vivo |
| Grantee: | Ronaldo Bragança Martins Júnior |
| Support Opportunities: | Scholarships in Brazil - Doctorate |