123 Efficiency of embryo production using ovum pickup oocytes recovered from stimulated and nonstimulated cows

2021 ◽  
Vol 33 (2) ◽  
pp. 169
Author(s):  
R. Y. Chinarov ◽  
G. N. Singina ◽  
V. Havlicek ◽  
N. P. Taradajnic ◽  
T. E. Taradajnic ◽  
...  
Keyword(s):  
Animal Health ◽  
Calf Serum ◽  
Prostaglandin F2α ◽  
Essential Element ◽  
Developmental Competence ◽  
Ultrasound Guided ◽  
Russian Science ◽  
Hormonal Stimulation ◽  
Follicular Aspiration ◽  
Significant Difference

Recovery of oocytes from live animals through ultrasound-guided transvaginal follicular aspiration (ovum pickup, OPU) is an essential element of modern livestock development. To increase the number of OPU oocytes retrieved, hormonal stimulation is broadly applied; however, the results are ambiguous. The aim of the present study was to compare recovery rate, quality of oocytes, and efficiency of invitro production of cattle embryos using OPU oocytes derived from hormonal treated and untreated cows. The study was performed in Simmental heifers at the age of 17 to 23 months. The heifers in the first group (n=7) were previously synchronized using prostaglandin F2α (Estrumate, MSD Animal Health) and gonadotrophin-releasing hormone (GnRH; Receptal, MSD Animal Health) and then underwent superstimulation with Stimofol (Reprobiol SPRL, Belgium) in a total amount of 0.25mg of FSH and 0.05mg of LH in a treatment of 6 injections on 3 consecutive days starting on Day −4 before OPU. The heifers of the second group (n=2) underwent OPU once a week for 5 weeks (10 OPU sessions). Groups of recovered cumulus–oocyte complexes (COCs) were matured in TCM-199 containing 10% fetal calf serum, 1mM Na-pyruvate, 50μg mL−1 gentamycin, 10μg mL−1 FSH, and 10μg mL−1 LH for 24h, fertilized in Fert-TALP (Tyrode’s-albumin-lactate-pyruvate) for 18 to 20h, and cultured in CR1aa medium for 7 days. All steps of IVP were performed at 38.5°C with 5% CO2 and 90% humidity. The cleavage and blastocysts rates were assessed on Day 2 and 7, respectively. In addition, Day-7 blastocysts were fixed, and the total cell number was determined using 4′,6-diamidino-2-phenylindole (DAPI) staining. The data were analysed by ANOVA using SigmaStat software package. As expected, the number of aspirated follicles per session in the first group of cows was higher than that in the second group: 14.6±1.2 vs. 6.8±0.5 (P<0.05). In total, 54 and 42 COCs were collected, which corresponds to recovery rates of 54.5±7.7 and 61.3±4.9% for stimulated and nonstimulated cows, respectively. After the quality evaluation, 36 COCs of SS heifers (67.2±1.9%) and 31 COCs of non-SS heifers (75.3±5.0%) were selected for IVP. We did not observe a significant difference in cleavage rate between two groups: 77.8% in the SS group vs. 71.0% in the non-SS group, whereas the rate of blastocyst production was higher (P<0.05) for SS heifers (25.0%) compared with non-SS animals (16.1%). The total number of cells in Day 7 blastocysts was significantly (P<0.01) higher in the first group (89.8±1.3) compared with the second group (70.6±3.2). Thus, superstimulation of heifers before OPU increases the efficiency of oocyte retrieval by ultrasound-guided transvaginal follicular aspiration as well as the developmental competence of OPU-derived bovine oocytes invitro. These studies were performed under financial support of the Russian Science Foundation (project No. 19-16-00115) and the Ministry of Science and Higher Education of Russia.

Effects of bovine oviduct epithelial cells, fetal calf serum and bovine serum albumin on gene expression in single bovine embryos produced in the synthetic oviduct fluid culture system

2005 ◽  
Vol 17 (8) ◽  
pp. 751 ◽  
Author(s):  
Mona E. Pedersen ◽  
Øzen Banu Øzdas ◽  
Wenche Farstad ◽  
Aage Tverdal ◽  
Ingrid Olsaker
Keyword(s):  
Gene Expression ◽  
Fetal Calf Serum ◽  
Calf Serum ◽  
Developmental Competence ◽  
Bovine Embryos ◽  
Glut 1 ◽  
Significant Difference ◽  
Oviduct Fluid

In this study the synthetic oviduct fluid (SOF) system with bovine oviduct epithelial cell (BOEC) co-culture is compared with an SOF system with common protein supplements. One thousand six hundred bovine embryos were cultured in SOF media supplemented with BOEC, fetal calf serum (FCS) and bovine serum albumin (BSA). Eight different culture groups were assigned according to the different supplementation factors. Developmental competence and the expression levels of five genes, namely glucose transporter-1 (Glut-1), heat shock protein 70 (HSP), connexin43 (Cx43), β-actin (ACTB) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), analysed as mRNA by using reverse transcription–polymerase chain reaction, were measured on bovine embryos cultured for 9 days. Gene expression of these in vitro-produced embryos was compared with the gene expression of in vivo-produced embryos. There was no significant difference found in embryo developmental competence between the Day 9 embryos in BOEC co-culture, FCS and BSA supplements in SOF media. However, differences in gene expression were observed. With respect to gene expression in in vivo and in vitro embryos, BOEC co-culture affected the same genes as did supplementation with FCS and BSA. HSP was the only gene that differed significantly between in vitro and in vivo embryos. When the different in vitro groups were compared, a significant difference between the BOEC co-culture and the FCS supplementation groups due to Glut-1 expression was observed.

132 The Developmental Competence of Bovine Oocytes Exposed to Progesterone and Luteotrophic Hormones During the Second Step of Two-Step Maturation

2018 ◽  
Vol 30 (1) ◽  
pp. 206
Author(s):  
G. Singina ◽  
I. Lebedeva ◽  
T. Taradajnic ◽  
E. Shedova ◽  
A. Lopukhov ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Calf Serum ◽  
Developmental Competence ◽  
Second Step ◽  
Russian Science ◽  
Cleavage Rate ◽  
System 1 ◽  
And Control ◽  
Bovine Oocytes

Data on effects of progesterone (P4) during in vitro maturation of bovine oocytes on their capacity for embryonic development are contradictory. Our study was aimed at characterising effects of P4 and 2 luteotropic hormones, prolactin (PRL) and LH, on bovine oocyte developmental competence during the second step of two-step maturation (from metaphase (M)I to MII). Slaughterhouse-derived cumulus-enclosed oocytes (CEO) were matured for 12 or 24 h [one-step (OS) Control] in TCM-199 containing 10% fetal calf serum (FCS), 10 μg mL−1 porcine FSH, and 10 μg mL−1 ovine LH at 38.5°C and 5% CO2. The CEO cultured for 12 h were transferred to the following culture systems: (1) TCM-199 containing 10% FCS (Control 1) or (2) a monolayer of granulosa cells (GC) precultured for 12 h in TCM-199 containing 10% FCS (Control 2); then, the oocytes were matured for next 12 h. In both systems, the medium of experimental groups was supplemented with either P4 (50 ng mL−1) or bovine PRL (25 and 50 ng mL−1) or ovine LH (5 μg mL−1). All treatments were repeated 5 to 6 times using 138 to 196 oocytes per group. Following IVM, all oocytes underwent IVF as described previously (Singina et al. 2014 Reprod. Fertil. Dev. 26, 154). Embryos were cultured in CR1aa medium until Day 5 post-insemination and then transferred to the same medium supplemented with 5% FCS and cultured to Day 7. Embryo development was evaluated at Days 2 and 7 for cleavage and blastocyst formation. Apoptosis was detected by the TUNEL method using 26 to 47 blastocysts per group (from 4 to 5 separate experiments). For each system, arcsine-transformed data were analysed by one-way ANOVA. In OS Control, the cleavage and blastocyst rates were 68.9 ± 4.4% and 22.0 ± 2.4%, respectively. Regardless of the system or medium of two-step culture, the cleavage rate did not differ from that for OS Control, varying between 57.6 and 68.4%. In the absence of GC (System 1), the blastocyst yield in the P4 group (30.4 ± 0.8%) was greater (P < 0.05) than in OS Control and Control 1 (20.2 ± 2.7%) as well as in the groups treated with LH (19.1 ± 3.0%) and 25 ng mL−1 PRL (20.1 ± 2.7%). In the presence of GC, P4 raised the yield from 16.7 ± 2.3% (Control 2) to 27.7 ± 2.4% (P < 0.05). Furthermore, in System 2, the blastocyst rate in groups treated with P4 and 50 ng mL−1 PRL (25.0 ± 2.8%) was higher (P < 0.05) than in the LH group (13.9 ± 2.6%). Meanwhile, the proportion of apoptotic nuclei (2.3-6.9%) was not associated with the system of oocyte maturation or effects of hormones studied. Our data indicate that P4 (50 ng mL−1) can enhance the developmental competence of bovine oocytes during the second step of two-step maturation regardless of the presence of granulosa cells, whereas the similar effect of PRL (50 ng mL−1) is less pronounced and depends on the granulosa-conditioned environment. This research was supported by the Russian Science Foundation (project 16-16-10069).

scholarly journals In vivo ultrasound guided transvaginal oocyte collection in the cow

2018 ◽  
Vol 49 (3) ◽  
pp. 195
Author(s):  
G. S. AMIRIDIS (Γ.Σ. ΑΜΟΙΡΙΔΗΣ) ◽  
M. SALAHEDDINE ◽  
I. A. JEFFCOATE ◽  
E. VAINAS (Ε. ΒΑΪΝΑΣ) ◽  
L. ROBERTSON
Keyword(s):  
Recovery Rate ◽  
Transvaginal Ultrasound ◽  
Oestrous Cycle ◽  
Ultrasound Guided ◽  
Follicular Aspiration ◽  
Oocyte Recovery ◽  
Oocyte Collection ◽  
Significant Difference

This paper describes the results of the in vivo ultrasound guided follicular aspiration for ovum pick υρ (OPU) in the cow. Twelve non pregnant dry cows aged 4-6 years were used in this experiment. Eight cows underwent OPU during three successive oestrous cycles and another four cows were used as controls having only transvaginal ultrasound scanning of their ovaries. Oocyte collection took place three times during the luteal phase of each natural oestrous cycle (days 3-4,9-11 and 14-17). The content of 326 follicles with a diameter of 4-15mm was aspirated and 104 oocytes were collected (recovery rate 31.9% or 1.55 oocytes per cow and session). The oocyte recovery rate increased after the first three sessions (from 13.04% to 35.0%) and reached levels of υρ to 52.6%. More follicles were aspirated on days 9-11 (133 follicles 40.8%) compared to 111 (34%) follicles on days 14-17 and 82 (25%) on days 3-4) (P<0.05). The evaluation of the collected oocytes revealed that 60 oocytes (57.7%) were suitable for further in vitro manipulation. Neither the origin of the oocyte (left or right ovary) nor the stage of the oestrous cycle affected the recovery rate or the quality of the collected oocytes. There was no significant difference either in the length of the oestrous cycle between the experimental animals and the controls (21.6± 1.4 vs. 22.37±1.0 respectively), or in plasma progesterone concentration in daily collected blood samples from the animals of the two groups. The results of this study are compared to those from the international literature and to the results from endoscopical methods for oocyte recovery. The feasibility of application of this technique to projects designed to improve the genetic merit of cows is discussed.

366 THE BOVINE MODEL FOR REPRODUCTIVE AGING: SUPEROVULATORY RESPONSE

2006 ◽  
Vol 18 (2) ◽  
pp. 290
Author(s):  
P. S. Malhi ◽  
G. P. Adams ◽  
R. J. Mapletoft ◽  
J. Singh
Keyword(s):  
Body Weight ◽  
Animal Health ◽  
Ovarian Response ◽  
Reproductive Aging ◽  
Ultrasound Guided ◽  
Expected Time ◽  
Follicular Wave ◽  
Follicular Aspiration ◽  
The Difference ◽  
Follicular Response

Two experiments were done to test the hypotheses that aging in cattle is associated with a reduction in the number of follicles recruited into an ovarian follicular wave, and a reduction in the ovarian response following gonadotropin treatment for superovulation. Old cows (13 to 16 years old; n = 6 to 9) and their young daughters (3 to 6 years old; n = 7 to 9) were treated to induce an ovarian superstimulatory response twice in each of two consecutive years (i.e. 31 and 33 superstimulatory cycles, respectively). In Experiment 1, follicular wave emergence was synchronized using a single treatment of estradiol (5 mg) combined with progesterone (100 mg) at the time of CIDR-B (Bioniche Animal Health, Belleville, Ontario, Canada) insertion. Superstimulatory treatment consisted of a total dose of 50 mg NIH-FSH-P1 (Folltropin-V; Bioniche Animal Health) per 100 kg body weight divided into twice daily intramuscular injections over 4 days. CIDR devices were removed at the time of the last FSH treatment and 500 �g cloprostenol (Estrumate; Schering-Plough Animal Health, Montreal, Quebec, Canada) was administered twice along with last two FSH treatments to induce luteolysis. Ovulation was induced using 25 mg porcine LH (Lutropin-V; Bioniche Animal Health) administered 24 h after the last FSH treatment. Ovulations were detected by ultrasonography at 24, 48, and 72 h after LH treatment. In Experiment 2, follicular wave emergence was synchronized with ultrasound-guided follicle ablation in the presence of a functional corpus luteum. Superstimulatory treatment was given as in Experiment 1. Ultrasound-guided follicle aspirations were performed 48 to 72 h after the last FSH for the purpose of another experiment. In both experiments, the ovarian follicular response was monitored daily by ultrasonography. There were no differences (P > 0.3) in mean body weight and dose of FSH treatment between old cows and their daughters. Fewer 2-5-mm follicles (P < 0.01) were detected at the expected time of wave emergence in old cows than in their daughters. In Experiment 1, the difference in mean number of ovulations in old and young cows (32 � 4 vs. 40 � 3) did not reach significance (P = 0.11), but fewer follicles e6 mm (P < 0.01) were detected on the day before ovulation in old cows than in their daughters. The number of follicles e6 mm detected just before ovulation (Experiment 1) or follicular aspiration (Experiment 2) in successive superstimulatory cycles was correlated within animals (r > 0.8; P < 0.0001). Similarly, the number of ovulations in successive superstimulatory cycles (Experiment 1) was correlated within animals (r = 0.6; P = 0.04). In conclusion, aging was associated with fewer 2-1 mm follicles at the time of wave emergence, a lower follicular response, and a tendency for a lower ovulatory response subsequent to gonadotropin treatment. The follicular and ovulatory response to superstimulatory treatment was repeatable within individuals.

224 EFFECT OF LENGTHENING THE SUPERSTIMULATORY TREATMENT PROTOCOL ON THE OVARIAN RESPONSE AND NUMBERS OF TRANSFERABLE EMBRYOS IN BEEF COWS

2012 ◽  
Vol 24 (1) ◽  
pp. 224
Author(s):  
A. Garcia Guerra ◽  
A. Tribulo ◽  
J. Yapura ◽  
J. Singh ◽  
R. J. Mapletoft
Keyword(s):  
Total Dose ◽  
Collection Efficiency ◽  
Animal Health ◽  
Treatment Protocol ◽  
Prostaglandin F2α ◽  
Ovarian Response ◽  
Beef Cows ◽  
Control Group ◽  
Corpora Lutea ◽  
Significant Difference

The present study was designed to test the hypothesis that an increase in the length of exposure of ovulatory follicles to progesterone and FSH during ovarian superstimulation will increase the number of ovulations and viable embryos in cattle. Twenty-four beef cows were initially subjected to follicle ablation at random stages of the oestrous cycle to determine the number of follicles at wave emergence; cows with comparable numbers of follicles were randomly allocated to groups by replicate. A single dose of prostaglandin F2α (PGF; Estroplan, Vétoquinol, QC, Canada) was given IM 7 to 9 days after follicle ablation and daily ultrasound examinations were performed to detect ovulation. Follicular ablation of all follicles ≥5 mm was done 5 to 8 days after ovulation and a progesterone-releasing intravaginal device (PRID, Vétoquinol) was inserted. The Control group (n = 12) was treated with 4 days of FSH and cows in the Long group (n = 12) were given 7 days of FSH treatment. The FSH treatments were initiated 1.5 days after ablation (Day 0) with a total dose of 400 mg of NIH-FSH-P1 (Folltropin-V; Bioniche Animal Health, Belleville ON, Canada). The total dose of FSH was distributed equally over 8 (Control) or 14 (Long) IM injections at 12-h intervals. Prostaglandin F2α was administered twice (at 12-h intervals) on Day 2 (Control) or Day 5 (Long) and PRID were removed 12 h after the last PGF. Both groups received 25 mg of porcine LH (pLH) IM (Lutropin-V; Bioniche Animal Health) 24 h after PRID removal and AI with frozen–thawed semen of proven fertility was done 12 and 24 h later. A third AI was done 12 h later in cows with 2 or more follicles ≥9 mm (12/12 and 9/12 in Control and Long groups, respectively; P = 0.22). All animals were subjected to transrectal ultrasonography every other day and at the time of ova or embryo collection. Ova or embryos were collected nonsurgically 7 days after the pLH injection and evaluated following IETS guidelines. Embryos were defined as transferable (Grades 1, 2 and 3) and freezable (Grades 1 and 2). Procedure Genmod was used to compare variables between treatments and results are presented as means ± SEM (Table 1). There was no significant difference in the total numbers of ova/embryos recovered, but there were more ovulations, corpora lutea (CL), fertilized ova and transferable and freezable embryos in the Long group (P < 0.05). Collection efficiency (number of ova/embryos over the number of CL) was lower in the Long group (P < 0.05). In summary, lengthening of the superstimulatory treatment resulted in an increased number of ovulations without a decrease in oocyte or embryo competence. Data suggest that the traditional 4-day superstimulatory treatment protocol provides inadequate time to maximize ovulatory response. Table 1.Superovulatory response (mean ± SEM) of cows subjected to a conventional 4-day (Control) or a 7-day (Long) treatment protocol

236 KINETICS AND PATTERN OF THE FIRST CLEAVAGE OF IN VITRO-FERTILIZED EMBRYOS BY IN VIVO-MATURED OOCYTES AND X-SORTED SPERMATOZOA IN DAIRY CATTLE

2013 ◽  
Vol 25 (1) ◽  
pp. 266
Author(s):  
S. Matoba ◽  
S. Sugimura ◽  
H. Matsuda ◽  
Y. Aikawa ◽  
M. Ohtake ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Calf Serum ◽  
Blastocyst Stage ◽  
Developmental Competence ◽  
Holstein Cows ◽  
Cleavage Pattern ◽  
Cell Cleavage ◽  
Significant Difference

Recently, we reported that high rates of good-quality blastocysts can be produced by IVF of in vivo-matured oocytes, obtained by ovum pick-up (OPU) after superstimulation in Holstein cows, with X-sorted sperm [Matoba et al. 2012 Reprod. Domest. Anim. 47(Suppl. 4), 515]. However, we have limited knowledge concerning the normality of embryonic cleavages in such embryos. The present study examined their kinetics and pattern of the first cell cycle. In vivo-matured oocytes were collected by OPU from non-lactating Holstein cows just before ovulation after superstimulation and ovulation induction by gonadotropin-releasing hormone. The oocytes were inseminated with 5 × 106 sperm mL–1 of X-sorted sperm and cultured in CR1aa supplemented with 5% newborn calf serum and 0.25 mg mL–1 of linoleic acid albumin at 38.5°C in 5% CO2, 5% O2, and 90% N2 for 216 h. Embryo kinetics were observed individually using a microwell culture dish and time-lapse cinematography (CCM-1.4MZS, Astec, Fukuoka, Japan) (Sugimura et al. 2010 Biol. Reprod. 83, 970–978). Photographs of each embryo were taken every 15 min during the in vitro culture period, and images were analysed by CCM-1.4 software (Astec). The cleavage pattern was categorised into normal cleavage (2 even blastomeres without fragment or protrusion) or abnormal cleavage (those with 2 uneven blastomeres, with fragments or protrusions and those dividing into 3 to 5 blastomeres at the first cleavage). Data were analysed by ANOVA, chi-square, and discriminant function. A total of 117 embryos were examined; of this number, 63.2% developed to the blastocyst stage and the rest were degenerated. A high rate of normal cleavage and a low rate of abnormal cleavage, including those with 2 uneven blastomeres and those with fragments or protrusions in the first cleavage pattern, were recorded in embryos that could develop to blastocysts compared with degenerated ones (P < 0.01 or P < 0.05, respectively; Table 1). No significant difference was found in those dividing into 3 to 5 blastomeres between the blastocysts and degenerated embryos (Table 1). Embryos developing to the blastocyst stage had a shorter duration of the first cell cycle [27.2 ± 2.3 h post-insemination (hpi)] compared with those undergoing degeneration (30.6 ± 5.7 hpi; P < 0.001). The threshold of duration of the first cell cycle was calculated by (X – 27.2)/2.3 = (30.6 – X)/5.7, resulting in X = 28.2. Blastocysts with a short duration of the first cell cleavage (≤28.2 hpi) showed a higher frequency of the normal cleavage pattern than those with a duration of the first cell cleavage longer than 28.2 hpi (71.7 and 53.6%, respectively; P < 0.05). Our results revealed that those IVF embryos that finished their first cleavage before 28.2 h of IVF and showed a normal cleavage pattern had superior developmental competence. Table 1.The first cleavege pattern reflects the developmental competence: blastocysts versus degenerated embryos This work was supported by the Research and Development Projects for Application in Promoting New Policy of Agriculture, Forestry and Fisheries (22016).

263 IN VITRO FERTILIZATION AND EMBRYO DEVELOPMENT OF CUMULUS - OOCYTE COMPLEXES COLLECTED BY ULTRASOUND-GUIDED FOLLICULAR ASPIRATION IN LLAMAS TREATED WITH GONADOTROPIN

2010 ◽  
Vol 22 (1) ◽  
pp. 288 ◽  
Author(s):  
M. A. Berland ◽  
A. von Baer ◽  
V. Parraguez ◽  
P. Morales ◽  
G. P. Adams ◽  
...  
Keyword(s):  
In Vitro Fertilization ◽  
Embryo Development ◽  
Animal Health ◽  
Ovarian Response ◽  
Ultrasound Guided ◽  
Vitro Fertilization ◽  
Follicular Aspiration ◽  
Oocyte Collection ◽  
Health Canada

We have previously documented that both FSH and eCG are equally effective in inducing ovarian superstimulation in llamas, resulting in the recovery of a high number of expanded COC suitable for in vitro fertilization (Ratto et al. 2005 Theriogenology 63, 2445-2457). The objective of the study was to evaluate the ovarian response, morphology, and competence of COC collected by ultrasound-guided follicular aspiration in llamas treated with FSH or eCG. Llamas were assigned randomly into 2 groups (n = 16 per group) and treated for 48 h after follicle ablation with (1)25 mg of FSH (Folltropin, Bioniche Animal Health Canada Inc., Belleville, Canada) i.m. twice daily for 4 d; or (2) 1000IU of eCG (Novormon, Bioniche Animal Health Canada) as a single i.m. dose. The starting of gonadotropin treatment was considered Day 0. Both groups were given an i.m. dose of 5 mg of Armour Standard LH (Lutropin, Bioniche Animal Health Canada) on Day 6, and COC were collected by transvaginal ultrasound follicle aspiration of all follicles ≥7 mm on Day 7. The ovarian response was assessed by transrectal ultrasonography using a 7.5-MHz linear-array transducer (Aloka SSD-500, Clinics, Santiago, Chile) immediately before oocyte collection at 24 to 26 h after LH treatment in both groups. The COC were classified as expanded, compact, denuded, or degenerated. Expanded COC collected from FSH- (n = 147) and eCG-treated llamas (n = 141) were fertilized in vitro using epididymal sperm as previously described (Ratto et al. 2006 Anim. Reprod. Sci. 97, 246-257). Gametes were co-incubated at 38.5°C in air with 5% CO2 and high humidity for 18 h. After in vitro fertilization, presumptive zygotes were co-culture in SOF medium supplemented with 0.6% of BSA with llama granulosa cells at 39°C, 5% CO2, 5% O2, and 90% N2 for 7 days. Embryo development was evaluated on Days 2, 5, and 7 of in vitro culture (Day 0 = IVF). Data were analyzed by Student’s t-test or Fisher’s exact test and presented as mean ± SEM. The FSH and eCG treatment groups did not differ with respect to the number of follicles ≥7 mm at the time of COC collection (16.0 ± 2.7 v. 14.0 ± 1.9; P = 0.5), the number of COC collected (11.5 ± 1.9 v. 9.7 ± 1.2; P = 0.4), or the collection rate per follicle aspirated (77.0 v. 71.5%; P = 0.2). No difference was detected between FSH and eCG-treated llamas in the number of expanded COCs (9.8 ± 1.4 v. 9.4 ± 1.2; P = 0.8). The percentage of presumptive zygotes to develop into 2 to 8 cells on Day 2 (65.3 v. 63.1), morulas on Day 5 (46.2 v. 42.5), and blastocyst stage on Day 7 (23.1 v. 20.5) did not differ (P > 0.05) between FSH and eCG-treated llamas, respectively. In conclusion, FSH and eCG treatments were equally effective for ovarian superstimulation and oocyte collection. The recovery of a high number of expanded COC can be used directly for in vitro fertilization and their competence is not affected by gonadotropin treatment. The study was supported by Convenio Desempeño en Investigacion (2007-DGI-CDA-04), Universidad Catolica de Temuco.

159 RELATIONSHIP BETWEEN CLEAVAGE PATTERNS OF FIRST CELL CYCLE AND POST-TRANSFER VIABILITY IN BOVINE EMBRYOS OBTAINED BY OVUM PICKUP AND IN VITRO FERTILIZATION

2012 ◽  
Vol 24 (1) ◽  
pp. 191
Author(s):  
K. Imai ◽  
S. Sugimura ◽  
T. Somfai ◽  
Y. Inaba ◽  
Y. Aikawa ◽  
...  
Keyword(s):  
Cell Division ◽  
Calf Serum ◽  
Development Program ◽  
Time Lapse ◽  
The Other ◽  
Developmental Competence ◽  
Vitro Fertilization ◽  
Significant Difference ◽  
Conception Rates

More than 300 000 embryos have been transferred all over the world (Stroud 2010 IETS Newsl. 27(4), 11–21). We have reported that embryos that showed the abnormal cleavage pattern at the first cell division can develop to the blastocyst stage (Somfai et al. 2010 J. Reprod. Dev. 56, 200–207). However, we have limited knowledge about the consequences of the pattern of first embryonic cleavage on their post-transfer developmental competence. The present study was conducted to determine the developmental competence of bovine blastocysts showing different cleavage patterns at their first cell division. Cumulus–oocyte complexes were collected by ovum pickup from Japanese Black cows and were subjected to in vitro maturation and IVF as reported previously (Imai et al. 2006 J. Reprod. Dev. 52, S19–S29 suppl). Inseminated oocytes were cultured in CR1aa medium supplemented with 5% calf serum covered by mineral oil at 38.5°C in 5% CO2 in air with micro-droplets or 5% CO2, 5% O2 and 90% N2. The kinetics of embryo development were analysed by time-lapse cinematography for 168 h after IVF by using a Cultured Cell Monitoring System (CCM-M1.4ZS, Astec, Fukuoka, Japan). A total of 673 photographs of each embryo were taken (1 photograph in every 15 min) during in vitro culture. Image stacks were analysed by the CCM-M1.4 software. Embryos were classified in 5 groups according to the pattern of first cleavage as normal cleavage (NC), direct cleavage from 1 cell to 3 to 4 blastomeres (3–4BL), unequal blastomeres (UB), multiple fragments (MF) and protrusion formation (PT). Blastocysts developing from each group were transferred into the ipsilateral uterine horn of each synchronized recipient on Day 7 or 8 after oestrus. Data on conception at Day 60, abortion and delivery were then recorded. Data were analysed by chi-square test and Student's t-test. In total, 43 embryos were transferred, 17 conceptions (39.5%) were established and 16 recipients (94.1%) were delivered. Only 1 abortion was detected at Day 223 in the NC group. The highest conception rate was observed in the NC group (55%, n = 20) and the 3–4BL (n = 12), UB (n = 6) and PT (n = 3) groups showed similar conception rates of 33.3% (1 implanted embryo belonged to 2 classes in UB and PT) and none of the embryos derived from the MF group (n = 3) could cause conception. There was a significant difference (P < 0.05) in conception rates between the NC group and totals of each of the other cleavage groups. No significant difference was found in gestation lengths and birth weights between the NC group (282.2 ± 4.4 days, 30.6 ± 3.8 kg, respectively) and totals of each of the other cleavage groups (282.8 ± 5.3 days, 30.3 ± 1.9 kg, respectively). These results indicate that embryos showing abnormal cleavage patterns at first cell division can develop to normal calves with normal gestation lengths and birth weights; however, their post-transfer viability is lower than for NC embryos. This work was supported by the Research and Development Program for New Bio-industry Initiatives.

scholarly journals 164 INFLUENCE OF LINEAGE OF OOCYTE DONOR ON THE YIELD AND MORPHOLOGY OF OOCYTES RECOVERED BY ULTRASOUND-GUIDED FOLLICULAR ASPIRATION IN NELLORE COWS

2012 ◽  
Vol 24 (1) ◽  
pp. 194
Author(s):  
A. Martins ◽  
R. S. Calegari ◽  
D. M. Paschoal ◽  
D. G. Souza ◽  
M. J. Sudano
Keyword(s):  
Body Condition Score ◽  
Genetic Lineage ◽  
Ultrasound Guided ◽  
Genetic Traits ◽  
Oocyte Donor ◽  
Follicular Aspiration ◽  
Significant Difference ◽  
The Mean ◽  
The 1960S

Several factors exert considerable influence on oocyte production and quality, such as donor age, nutrition, season and others. However, very few studies have been done to verify the effects of breed on the quantity or quality of oocytes, or both. In Brazil, the Nellore breed has been developed as a result of many years of selective breeding from 6 major ancestor bulls imported from India in the 1960s. These animals have characterised their respective lineages, as evidenced by the well-defined genetic traits. This study was carried out to investigate the influence of the genetic lineage of oocyte donors on the number and quality of oocytes obtained through ultrasound-guided follicular aspiration in Nellore cows derived from 2 lineages of bulls [Karvadi (K) and Taj Mahal (T)]. Both maternal (Km and Tm) and paternal (Kp and Tp) lineages were investigated. Oocyte aspirations were repeatedly performed with an aspiration interval of 15 days without the superovulation procedure in 56 donor females over a period of 2 years. At the beginning of the experiment, the animals were on average 78 ± 12 months old with a body condition score of 3.6 ± 0.08 (on a 5-point scale). Recovered cumulus–oocyte complexes were counted, morphologically examined and classified into 7 categories (grades I to VII) according to the number of layers of the cumulus–oocyte and the appearance of the cytoplasm. Data were analysed using ANOVA and Tukey's test after transformation into log(x + 1). Values are reported as means ± s.e.m. and differences (P > 0.05) were taken as significant. The mean number of oocytes retrieved from donors of lineage Tp-Tm was higher (n = 9; 28.23 ± 1.92; P < 0.05) than that obtained for lineages Kp-Tm (n = 11; 21.34 ± 1.32), Kp-Km (n = 15; 21.28 ± 1.73) and Tp-Km (n = 21; 16.72 ± 1.31). There was no significant difference in the mean number of recovered oocytes between donors of lineages Kp-Tm and Kp-Km; however, lineage Tp-Km yielded a lower number of oocytes. Donors of lineage Kp yielded more (P < 0.05) oocytes of grade III than lineage Tp (10.11 ± 0.66 vs 8.79 ± 0.58, respectively). These lineages produced a higher mean number of oocytes graded as I, II and III (3.14 ± 0.21, 4.93 ± 0.33 and 10.11 ± 0.66 vs 3.19 ± 0.21, 5.59 ± 0.44 and 8.79 ± 0.58, respectively) than any other categories. However, more oocytes (P < 0.05) of grades I, II and III were obtained for lineage Tm (3.64 ± 0.25, 5.9 ± 0.42 and 11.67 ± 0.67) than for lineage Km (2.85 ± 0.19, 4.92 ± 0.40 and 7.73 ± 0.56, respectively), with similar results for oocytes of grades IV, V, VI and VII. In conclusion, the genetic lineage of the oocyte donor influenced the production and quality of oocytes obtained through ultrasound-guided follicular aspiration in Nellore cows. Selection of donor cows based on their oocyte morphology and availability can further improve IFV programs.

248 IN VITRO EMBRYO PRODUCTION USING IN VIVO-MATURED OOCYTES COLLECTED TRANSVAGINALLY FROM WOOD BISON (BISON BISON ATHABASCAE)

2015 ◽  
Vol 27 (1) ◽  
pp. 213
Author(s):  
M. P. Cervantes ◽  
J. M. Palomino ◽  
M. Anzar ◽  
R. J. Mapletoft ◽  
G. Mastromonaco ◽  
...  
Keyword(s):  
In Vitro Maturation ◽  
Animal Health ◽  
Calf Serum ◽  
Developmental Competence ◽  
High Humidity ◽  
Embryo Production ◽  
Wood Bison ◽  
In Vitro Embryo Production

Reproductive technologies are being developed to help conserve the genetic diversity of wood bison, a threatened species. To date, the efficiency of in vitro embryo production in bison is very low and appears to be related to inadequate in vitro conditions for oocyte maturation. Recently, we have attempted to circumvent the problem by inducing oocyte maturation in vivo and found that more than one-third of superstimulated oocytes collected 30 h after administration of hCG were at metaphase II (Cervantes et al. 2013 Reprod. Fertil. Dev. 25, 283; Cervantes et al. 2014 Reprod. Fertil. Dev. 26, 199). We hypothesise that additional maturation time in vitro, after in vivo maturation, will allow the remaining oocytes to reach the MII stage, and thus improve in vitro embryo production in wood bison. The objective of this study was to determine the effect of an additional 4 h of in vitro maturation on the developmental competence of oocytes collected 30 h after hCG treatment. Wood bison cows (n = 24) were superstimulated by the administration of 300 mg of FSH (Folltropin-V) diluted in 0.05% hyaluronan on the day of follicular wave emergence and 100 mg of FSH in hyaluronan 2 days later. Bison were administered 2500 IU of hCG (Chorulon) IM 2 days after the last dose of FSH. Transvaginal ultrasound-guided follicle aspiration was performed 30 h after hCG treatment to collect cumulus-oocyte complexes (COC). Expanded COC (with no evidence of degeneration) were selected and assigned randomly to 2 groups (n = 38 COC/group) in which IVF was done immediately, or after 4 h of in vitro maturation in TCM 199 with 5% calf serum, 5 μg mL–1 pLH, 0.5 μg mL–1 pFSH, and 0.05 μg mL–1 gentamicin, at 38.5°C, 5% CO2 and high humidity. In vitro fertilization (Day 0) was done with frozen-thawed wood bison semen (dose 5 × 106 sperm mL–1) in Brackett-Oliphant medium at 38.5°C, 5% CO2, and high humidity. Presumptive zygotes were cultured in CR1aa plus 5% calf serum, at 38.5°C and in 5% CO2, 5% O2, and 90% N2 and high humidity. Cleavage was recorded on Day 3, and blastocyst formation was recorded on Days 7 and 8. Cleavage and blastocyst rates (calculated from the total number of oocytes submitted to IVF) were compared between groups by chi-square analysis. No difference was detected between groups (immediate fertilization v. after an additional 4 h in vitro) in cleavage rate on Day 3 (55.3 v. 60.5%, respectively, P = 0.82), or blastocyst rate on Day 7 (13.2 v. 23.7%, respectively, P = 0.37). However, the blastocyst rate on Day 8 was higher in the COC group exposed to an additional 4 h of in vitro maturation (18.4 v. 44.7%, respectively, P = 0.03). Results support the hypothesis that an additional short period of in vitro maturation improves the developmental competence of oocytes collected after 30 h of in vivo maturation.We thank Bioniche Animal Health for providing FSH (Folltropin-V) and hyaluronan (MAP-5), and Merck Animal Health for hCG (Chorulon).

Sign in / Sign up

Export Citation Format

Share Document