Reproduction in Domestic Animals - Facultad de Ciencias Veterinarias

16 t h International Congress on Animal Reproduction
Poster Abstracts 97
that in wild. Parturition timing and endocrine patterns show
similarities to those in domestic camelids.
P208
In vitro maturation of dromedary camel (Camelus
dromedarius) oocytes: effect of different protein
supplementations and epidermal growth factor
Wani, NA*, Skidmore, JA
Camel Reproduction Center, United Arab Emirates
The present experiment was aimed to compare the effect of different
protein supplementation sources, fetal calf serum (FCS), estrous
dromedary serum (EDS) and BSA, and the effect of different
concentrations of epidermal growth factor (EGF) on in vitro nuclear
maturation of dromedary camel oocytes. Ovaries collected from a
local slaughterhouse were brought to the laboratory in a thermos flask
containing warm normal saline solution (NSS) and cumulus oocyte
complexes (COCs) were harvested by aspirating the visible follicles
using an 18G hypodermic needle attached to a 20 mL syringe
containing PBS supplemented with 5% FCS. Pooled COCs were
randomly distributed to 4-well culture plates containing 400 μL of the
maturation medium and cultured at 38.5 0 C in an atmosphere of 5%
CO 2 in air for 36 h. The basic maturation medium consisted of TCM-
199 supplemented with 0.1 mg/mL L-glutamine, 0.8 mg/mL sodium
bicarbonate, 0.25mg/mL pyruvate, 50 μg/mL gentamicine, 10 μg/mL
bFSH, 10 μg/mL bLH and 1 μg/mL estradiol. In experiment 1, this
medium was supplemented with either 10% FCS, 10% EDS or 0.4%
BSA whereas, in experiment 2, the maturation medium was
supplemented with 0, 10, 20 or 50 ng/mL of EGF. At the end of the
culture period all intact COCs were denuded of cumulus cells. The
oocytes with a visible polar body, considered to be in metaphase-II
stage, were used for other experiments, while as all other oocytes
were fixed in ethanol: acetic acid (3:1) for 24 h and stained with 1%
(w/v) aceto-orcein stain. The slides were examined under phase
contrast microscope at magnification of 400X to evaluate the status of
nuclear maturation. Oocytes were classified as germinal vesicle (GV),
diakinesis (DK), metaphase-I (M-I), metaphase-II (M-II) or others
(those with degenerated, fragmented, scattered, activated or without
visible chromatin). In experiment 1, no difference (P < 0.05) was
observed in the proportion of oocytes reaching M-II stage between the
media supplemented with FCS (71.5 ± 4.8), EDS (72.8 ± 2.9) and
BSA (72.7 ± 6.2). In experiment 2, a high proportion (P < 0.05) of
oocytes reached M-II stage when the maturation medium was
supplemented with 20 ng/mL of EGF (81.4 ± 3.2) compared with the
media supplemented with 10 ng/mL (66.9 ± 4.1) and control (67.2 ±
7.1) groups. It may be concluded that all the three protein
supplementation sources used in this study are capable of supporting
oocyte nuclear maturation equally and a supplementation of 20 ng/mL
of EGF increases the maturation rate of oocytes in this species.
Poster 05 - Equine Reproduction
P209
Effect of an immunomodulator on estrogen alpha and
progesterone receptor expression in endometrial tissue
of healthy, endometritis resistant mares during the
estrous cycle
Acuña, S 1 *, Tasende, C 2 , Rivulgo, M 3 , Alzola, R 3 , Felipe, A 3 , Rogan, D 4 ,
Fumuso, E 5
1Cellular and Molecular Biology, Veterinary Faculty, Uruguay; 2 University of
the Republic, Uruguay, Uruguay; 3 Argentina; 4 Bioniche Life Sciences Inc.,
Canada; 5 Universidad Nacional del Centro de la Provincia de Buenos Aires,
Argentina
The estrogen alpha and progesterone receptor (ERα and PR)
expression was investigated in endometrial biopsies of healthy,
endometritis resistant mares treated by intrauterine administration at
estrous (ovarian follicles >29mm, folds and endometrial edema) with
1500 μg Mycobacterial Cell Wall-DNA Complex (MCC). The
follicular dynamic was followed by ultrasonography. Endometrial
biopsies were taken repeatedly from the same mares at diestrous (in
the previous estrous cycle on day 6 post ovulation, n=3, group D),
estrous (immediately before treatment, n=7, group E), 24 h post
treatment (n=7, group 24hPT), ovulation (n=7, group OvPT) and
diestrous (6 days post treatment, n=7, group DPT). An
immunoperoxidase staining technique was used to visualize ERα and
PR immunoreactivity. The immunoreactivity was analyzed in
Luminal Epithelium (LE), Glandular Epithelium (GE) and Stromal
(St) cells. Ten fields were analyzed for each cell types at a
magnification of 1000x. The average staining for each cell types was
calculated according to the following procedure = 1 x n (SI1) + 2n
(SI2) + 3n (SI3), where n = amount of cells per field exhibits SI (1),
moderate (2) and intense (3). The total positive cells (LE + GE + St)
and average staining was analyzed by ANOVA test. The model
included the effect of group, cell types and the interactions between
them. The level of significance was considered to be P