Estradiol changes the immunohistochemical profile of the endometrial proteins gamma PKC, AKR1B1 and alpha estradiol receptor in cattle

In cattle estradiol (E2) has an important role in the endometrial PGF2α release associated with luteolysis, however, the molecular mechanisms involved in such process are poorly understood . The PGF2α synthesis is the result of a series of intracellular events that include the participation of kinase C gamma protein (PKCγ), aldo-keto reductase family 1 member B1 (AKR1B1) and estradiol receptor α (ERα) which are present in the endometrial cells. The main hypothesis was that females treated with E2 have a modification of PKCγ, AKR1B1 and Erα endometrial protein concentration. The objective of this study was to investigate using immunohistochemistry the PKCγ, AKR1B1 and ERα proteins in endometrial tissue in Nelore cows treated or not with 3 mg of 17β - estradiol intravenously on the 15th day of the estrous cycle 0, 4 and 7 hours after injection. Nelore (n = 52), pluriparous, cyclic and non-lactating cows received 2 mg of estradiol benzoate (Sincrodiol Ourofino®, Cravinhos, Brazil) and an intravaginal progesterone device (1g; Sincrogest; Ourofino®, Cravinhos, Brazil) during 8 days. The cows received 0.5 mg of sodium cloprostenol (Sincrocio; Ourofino®, Cravinhos, Brazil) via IM, 48 hours before the withdrawal of the device and a second application the day of device removal. On day 15 of the estrous cycle (D0; estrus) the following treatments were administered: placebo (P; 5 ml of ethanol 50%; IV), estradiol (E; 5mL of 50% ethanol containing 3 mg of 17β estradiol; IV) or control (not treated). Time 0 was the moment of the treatment application. Cows were subjected to a transcervical endometrial biopsy, and according to the time of biopsy were divided into the following groups: time 0 in the control group (C; n = 10 ), time 4 hours (E4, n = 11 and P4, n = 10), and 7 hours (E7, n = 10 and P7; n = 11). The procedures performed are in accordance with the principles of the Ethics in AnimalResearch Committee of UNESP Dracena/SP (CEUA Protocol 08/2014). The tissue obtained by biopsy was placed in 4% buffered-formalin- for 24 hours and then stored in 70% alcohol until paraffin embedding. Endometrial sections were evaluated by immunohistochemistry and immunostaining was evaluated in the luminal epithelium (LE), glandular epithelium (GE) and stroma (S). The statistical differences were determined by t test and considered when P < 0.05. The results of PKCγ protein showed higher immunostaining in the LE of E4 and E7 groups compared to P4 and P7 (P < 0.05) and increased labeling in GE of E7 compared to P7 (P < 0,05). The AKR1B1 protein showed higher immunostaining in the LE of E4 and E7 groups compared to P4 and P7 (P < 0,05) and higher immunostaining in GE of E4 compared to P4 (P < 0,05). The ERα shows an higher immunostaining in the GE of P4 and P7 groups compared to E4 and E7 (P < 0.05) and higher immunostaining in LE of P4 when compared to E4 group (P < 0.05). It is concluded that E2 increases immunostaining of the PKCγ and AKR1B1 proteins and reduce of the Erα protein in endometrial tissue, thereby modifying the concentration of these endometrial receptors.