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| Author(s): |
Flavia Caroline Destro
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Botucatu. 2016-02-03. |
| Institution: | Universidade Estadual Paulista (Unesp). Faculdade de Medicina Veterinária e Zootecnia. Botucatu |
| Defense date: | 2016-01-22 |
| Advisor: | João Carlos Pinheiro Ferreira |
| Abstract | |
The general objective of this study was to characterize the production of progesterone (P4) by the cells of the bovine corpus luteum (CL) subjected to different culture and coculture media in vitro. To this end, three experiments described in two articles were conducted. Article I: CLs belonging to the middle phase (10-12 days, n=5) were collected and processed in the laboratory. The luteal cells (LCs) were cultured for 07 days in a humid atmosphere of 5% CO2, with or without the addition of 10% fetal bovine serum (FBS), with their respective treatments: CONTROL; CONA (concanavalin-A) (10 μg/mL); LH (100 μg/mL); CONALH; LHPGFβα (10 ng/mL); CONALHPGFβα. Samples of culture medium were collected on D1 and D7 for further P4 measurement. P values <0.05 were considered statistically different. Culture of LCs in the presence of CONA decreases the ability of LC to secrete P4, and this effect was reversed by addition of LH in the culture medium in D1, but not in D7. The suppressive action was more evident in CONA cultures without FBS. In Article II, Experiment 1, CLs of middle phase of the estrous cycle (10-12 days, n = 4) were used. In the laboratory, the CLs were processed and the LCs were cultured for 07 days in a humidified atmosphere of 5% CO2. The LCs received the respective treatments with or without LH (100 μg/mL): Control; PGFβα (10 ng/mL); PGFβα (100 ng/mL); PGFβα (354,5 ng/mL). The samples that received LH produced more P4, and the administration of different doses of PGFβα showed no change in P4 secretion. In Experiment β, CLs of middle phase of the estrous cycle (10-12 days, n = 4) were used. In the laboratory, the CLs were processed and the LCs, luteal endothelial cells (ECs) and immune cells (ICs) were isolated and co-cultured for 07 days in a humidified atmosphere of 5% CO2. The groups were divided with or without PGFβα: Control: LC on bottom of the plate; Control: EC on bottom of the plate; Co-culture: LC + EC + IC on bottom of the plate; LC (bottom of the plate) + EC + IC (inside the insert); EC (bottom of the plate) + LC + IC (inside the insert). It was not observed variation in the production of P4, when added PGFβα in culture and co-culture of cells from CL. In both experiments, samples of culture medium were collected on D1 and D7 for further P4 measurement. P values <0.05 were considered statistically different. We conclude that the in vitro culture and co-culture of bovine CL cells are alternative tools for the study of luteal function, but there is a need for further methodological adjustments in order to mimic physiological processes related to steroidogenesis and luteolysis. (AU) | |
| FAPESP's process: | 13/00992-3 - In vitro culture of bovine steroidogenic luteal cells and non-steroidogenic. Effect of PGF2a on the expression of genes and proteins associated with steroidogenesis and apoptosis |
| Grantee: | Flavia Caroline Destro |
| Support Opportunities: | Scholarships in Brazil - Doctorate |