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| Author(s): |
Luiz Aurélio de Campos Crispin
Total Authors: 1
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| Document type: | Master's Dissertation |
| Press: | São Paulo. |
| Institution: | Universidade de São Paulo (USP). Instituto de Ciências Biomédicas (ICB/SDI) |
| Defense date: | 2021-06-18 |
| Examining board members: |
Sandra Coccuzzo Sampaio Vessoni;
Soraia Kátia Pereira Costa;
Luis Roberto de Camargo Gonçalves;
Thais Martins de Lima
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| Advisor: | Sandra Coccuzzo Sampaio Vessoni |
| Abstract | |
In the last decades, several studies have demonstrated the important anti-inflammatory action of CTX, directly assessed on the functions of neutrophils and macrophages, in addition to its potential as an immunomodulator, particularly on macrophages related to tumor progression and acute and infectious inflammatory response. This long-term immunomodulatory capacity (up to 14 days) of CTX involves phenotypic reprogramming of the macrophage, making it more pro- or anti-inflammatory, depending on the concentration of the toxin and the microenvironment stimulus. Particularly, with the local presence of macrophages, the production and release of the chemical mediators produced by them, the migration and activation of fibroblasts is intensified, leading to the deposition of large amounts of fibronectin and collagen, which can favor fibrosis. Therefore, the objective of the present project was to investigate the importance of macrophage secreting activity induced by CTX on the fibroblast functions involved in the healing process. Therefore, monocultures of macrophages differentiated from the human monocytic strain THP-1 were incubated with different concentrations of CTX, for 2 hours, washed and incubated with fresh RPMI 1640 medium, for 24 hours and after that period, human fibroblasts were incubated with the supernatants of these monocultures (cel: supernatant) or with macrophages (cel: cel) for the in vitro determination of fibroblast events, in different periods, such as: 1) proliferation assay; 2) migration test in the Wound healing model; 3) migration test in the Transwell Chamber; 4) morphology of the fibroblast actin cytoskeleton and expression of fibronectin and type I collagen secreted by these cells, visualized by confocal microscopy; 5) quantification of 15-Epi-LXA4, by immunoenzymatic assay (EIA) and 6) Proteomic and secretomic analyzes of macrophages treated with the toxin, by mass spectrometry. Together, the results presented show that macrophages treated with CTX modulate the functional activity of fibroblasts (migration and proliferation and secretion of matrix components), both in cell:cell contact, and through mediators secreted in their supernatants, in a more efficient way. pronounced to that observed for classic anti-inflammatory drugs. This study, therefore, contributed to highlight the importance of immunomodulatory capacity of CTX on events involved with healing. (AU) | |
| FAPESP's process: | 17/23974-1 - Crotoxin-induced macrophage secretory activity on fibroblast functions involved with the wound healing process. in vitro studies |
| Grantee: | Luiz Aurélio de Campos Crispin |
| Support Opportunities: | Scholarships in Brazil - Master |