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Study the mechanisms of action by which estradiol stimulates endometrial PGF2a synthesis in cows: assessment of its effects on the population of endometrial receptors for progesterone and estradiol

Grant number: 15/03331-3
Support type:Scholarships in Brazil - Scientific Initiation
Effective date (Start): May 01, 2015
Effective date (End): July 06, 2016
Field of knowledge:Agronomical Sciences - Veterinary Medicine - Animal Reproduction
Principal Investigator:Claudia Maria Bertan Membrive
Grantee:Barbara Piffero Mello
Home Institution: Faculdade de Medicina Veterinária e Zootecnia (FMVZ). Universidade de São Paulo (USP). São Paulo , SP, Brazil
Associated scholarship(s):15/22047-4 - Effect of level of dietary cation anion difference and duration of prepartum feeding on calcium and energy metabolism in transition cows, BE.EP.IC

Abstract

In cows the release of endometrial PGF2a can be induced in vivo by estradiol (E2), however, the role of E2 in PGF2a synthesis is still unclear. Whereas the plasma concentration of PGF2a increase 3.5 hours after the application of E2 in vivo, it is believed that E2 activates not only enzymes, but also stimulates the synthesis of essential proteins for the production of PGF2a. It is known that bovine explants from animals injected with E2 and culture of bovine endometrial cells (BEND) exposed to E2, to be treated with calcium ionophore (CI) showed a marked increase in release of PGF2 compared those not E2 received and handled only with E2 or CI. The main hypothesis of the overall study (FAPESP process 2012 / 50767-3) is that E2 stimulates the expression and / or the activity of PKC and PLA2 enzymes, both dependent on calcium for activation, involved in endometrial PGF2a synthesis. It is known that stimulation of the endometrial PGF2a synthesis involves other endocrine factors such as oxytocin (OT) and progesterone (P4). In previous analyzes recently made by this group, it was found by RT-PCR significantly increased expression of oxytocin receptors females after being treated with E2. Thus, it was hypothesized that in females treated with E2 occurs a change in the concentration of endometrial OT, P4 and E2 receptors. Thus, it is specific objective of this proposed study to investigate by immunohistochemistry the population of OT, P4 and E2 receptors in endometrial tissue in Nellore cows treated or not with 3mg of 17b-estradiol intravenously, the 15th day of the cycle estrous 0, 4 and 7 hours after injection. The tissues to be used in the present study have already been collected, so the proposal Scientific Initiation is based on the evaluation of them by immunohistochemistry technique. Thus, we used 49 Nellore cows, on the 15th day of the estrous cycle, divided into five groups: females who received no injection and were submitted to uterine biopsy at the time 0, where it was considered time 0 the time of application treatments in the other groups (n = 9; control group); females who received an injection of placebo solution (50% ethanol) at 0 hour and were subjected to uterine biopsy at hour 4 (n = 10; placebo Group 4 hours); females who received an intravenously injection of 3 mg of 17b-estradiol diluted in 50% ethanol and were subjected to uterine biopsy in 4 hours (n = 10; Estradiol Group 4 hours); females who received an injection of placebo solution (50% ethanol) at time 0 and were subjected to uterine biopsy at hour 7 (n = 10; Placebo 7 hours) and females who received an intravenously injection of 3mg diluted with 17b-estradiol 50% ethanol and were submitted to uterine biopsy in hour 7 (n = 10; Estradiol Group 7 hours). An endometrial biopsy were performed via transcervical in the ipsilateral uterine horn to the ovary containing the corpus luteum, promoted the removal of an endometrial fragment measuring about 5 x 5 mm / each. After collection of said tissue, it was placed in buffered formaldehyde solution 4% for 24 hours and then was kept in to 70% ethanol until they were embedded in paraffin. After paraffin embedding, endometrial sections will be evaluated by immunohistochemistry for OT , P4 and E2 receptors in the different treatment groups. This study will allow a better understanding of the mechanism by which E2 stimulates the synthesis of PGF2a in cows. The clarification of this mechanism may contribute to the development of anti-luteolitics strategies that could reduce embryo mortality in cows between 15 and 19 days of gestation, currently responsible for significant economic losses.