Full text
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| Author(s): |
Camila Fazzani
Total Authors: 1
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| Document type: | Master's Dissertation |
| Press: | São Paulo. |
| Institution: | Universidade de São Paulo (USP). Faculdade de Medicina (FM/SBD) |
| Defense date: | 2010-12-17 |
| Examining board members: |
José Angelo Lauletta Lindoso;
Alda Maria da Cruz;
Daniela Santoro Rosa
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| Advisor: | José Angelo Lauletta Lindoso |
| Abstract | |
Hamsters infected by Leishmania (L.) chagasi are considered one of the most remarkable models to study several characteristics related to immunosuppression, raised during active visceral leishmaniasis especially because hamsters show similar clinical manifestations as it is found in humans without surmounting the infection. In this study, hamsters infected intraperitoneally with 2x107 parasites were used to evaluate the possible factors involved in the dynamic of immunosuppression that occurs during the disease development. The evaluated parameters were the production of Th1 and Th2 cytokines profile, nitric oxide production of splenic cells and detection of apoptosis in splenic lymphocytes at early (6,20, 48 e 72 hours), intermediate (7 e 15 days) and late times (30 e 60 days) of infection. Initially, we evaluate the parasite load in the spleen and observed a progressive increase dependent on the time of infection, ratifying that infected hamsters are good models to set up the disease development. Cellular response upon mitogen or Leishmania antigen was the parameter used to evaluate the immunosuppression showing a preserved response to concanavalin A at all period of infection and a preserved response to the Leishmania antigen at all early phases however with no specific antigen response from 7 day of infection until the late phase of infection. Nitric oxide quantification was determined in the splenic cells supernatant in culture stimulated upon concanavalin A and Leishmania antigen and we observed initially low level of nitric oxide production, measured by the Griess method. Nonetheless, when the cells were stimulated upon Leishmania antigen we observed a lower level nitric oxide production at 48 and 72 hours of infection. mRNA of Th1 e Th2 cytokines profile was determined by RT-PCR at all period of infection studied in infected or non infected hamsters showing no difference at the cytokine profile in this experimental model. Detection of apoptosis in the non adherent splenic cells (probably T lymphocytes) ex-vivo and in the stimulated culture with concanavalin A and Leishmania antigen was evaluated by the following methods: annexin V-FITC, cleaved caspase 3 and TUNEL by flow cytometry analysis. There was phosphatidilserine staining by annexin method, at 6 hour of infection when cells were stimulated by mitogen or Leishmania antigen at 48 hours and 60 days of infection only when stimulated by Leishmania antigen. Splenic cells in culture stimulated by concanavalin A showed no cleaved caspase 3 staining in all period of infection studied. Otherwise, apoptosis was detected in the ex-vivo splenic cells at 20, 48 and 72 hours of infection by cleaved caspase 3 staining. We observed a low DNA break staining of non adherent splenic cells in all period analyzed, although this staining was increased in non adherent cells stimulated upon Leishmania antigen at 20 hours from 7 day of infection until 60 day of infection. Our results suggest that a visceral leishmaniasis immunosuppression in the hamsters model infected by Leishmania (L.) chagasi is antigen dependent and sets up in the intermediate phases of infection from 7 days on. This immunosuppression seems to be related to the apoptosis increase in lymphocytes already in the early period of infection probably favoring its progression especially because of its occurrence in reactive splenic antigen. Our data also suggest that the immunosuppression is not provided by dichotomization in the Th1 and Th2 cytokines profile (AU) | |
| FAPESP's process: | 07/06191-1 - Lymphocyte Apoptosis in the visceral leishmaniasis immunossupression in hamsters infected by Leishmania (Leishmania) chagasi |
| Grantee: | Camila Fazzani |
| Support Opportunities: | Scholarships in Brazil - Master |