Coculture of porcine epithelial oviductal cells and in vitro produced embryos: effect on embryo development and quality

LORENZO, MS.; TEPLITZ, G.; CRUZANS, PR.; LUCHETTI, CG.; LOMBARDO, DM.

Bologna

Congreso; 19th International Congress on Animal Reproduction; 2020

ICAR Committee

The oviduct is involved in many reproductive functions including early embryo development. The epithelial cells that cover the oviduct produce the oviductal fluid and could be used to recreate the in vivo environment into which embryo development takes place. This study aimed to evaluate the coculture of porcine embryos with a monolayer of porcine oviductal epithelial cells (POEC) and its effect on embryo development and quality. The POEC were obtained by pressing the itsmus usign slides and performing 3 cycles of vortexing and decanting in DMEM-F12 medium. Passage 1 was used for these experiments (POEC-1). Oocytes were obtained from follicular aspiration of slaughterhouse´s ovaries. Oocytes were in vitro matured for 44 h in TCM 199 supplemented with human menopausal gonadotropin and dAMPc during the first 22 h. In vitro fertilization was performed with 17°C-refrigerated boar semen for 4 h in 100 µL-drop of TCM 199 with caffeine, BSA, sodium lactate, and sodium pyruvate (20 denuded oocytes per drop, 1x106 spermatozoa/mL). Presumptive zygotes were washed and randomly assigned to one of the groups for in vitro culture: control (50 µL-drop of NCSU 23 with sodium pyruvate and lactate), POEC-1 (same than the control + POEC-1 50000 cells/mL), POEC-1 + SFB (same than the control + POEC-1 50000 cells/mL and 2.5% of bovine fetal serum). Culture conditions were 7 % O2, 5% CO2, 39 °C and humidity. On day 2, cleavage rate was registred and embryos were transferred to drops of NCSU 23 with glucose and without cells. The blastocyst rate was registred on day 7. Embryo quality was assessed by counting the number of cells per blastocyst (Hoescht) and the apoptosis index (TUNEL positive cells/total cells). Coculture with POEC-1 significantly increased blastocyst rate (control: 14%; POEC-1 + SFB: 10%; POEC-1: 28%; p < 0.05 Chi-square test) and allowed embryo hatching. However, there was no difference in the number of cells per blastocyst (control: 58.6 ± 6; POEC-1 + SFB: 50.3 ± 3.7; POEC-1: 50.6 ± 4.8) neither in the apoptosis index (control: 8.1; POEC + SFB 8.3; POEC: 7.4; nonparametric ANOVA). The use of POEC-1 during the first 2 days of embryo culture enhance embryo development and improved the conditions to allow embryo hatching. Other parameters of embryo quality should be evaluated in the future.