scholarly journals Effects of superstimulation with fsh on follicular population and recovery rate of oocytes in the growing phase of the first and second follicular wave

2012 ◽  
Vol 47 (No. 2 - 3) ◽  
pp. 33-37
Author(s):  
S. Čech ◽  
V. Havlíček ◽  
M. Lopatářová ◽  
M. Vyskočil ◽  
R. Doležel
Keyword(s):  
Czech Republic ◽  
Corpus Luteum ◽  
Recovery Rate ◽  
Dairy Farm ◽  
Cumulus Cells ◽  
Dominant Follicle ◽  
In Vitro Production ◽  
Follicular Wave ◽  
Vitro Production

Effectiveness of in vitro production of embryos (IVP) is limited among other factors by the recovery rate of oocytes. Gonadotropin superstimulation can improve the recovery rate of oocytes. The effect of FSH treatment on follicular population and recovery rate of oocytes at ovum pick-up (OPU) in the growing phase of the 1st as well as the 2nd follicular wave after superstimulation was the object of our experiment. Twelve unpregnant milking cows (15&ndash;20 kg milk per day) housed on a dairy farm were used in the experiment. The cows bearing corpus luteum were synchronized by PGF<sub>2 </sub>(day 0) and they were treated by FSH (Folicotropin inj. sicc. ad us vet., Spofa Prague, Czech Republic, single doses 80, 80, 80, 80, 40 and 40 UI) in 12 h intervals on days 12, 13 and 14. Transvaginal ultrasonographic puncture of oocytes in cows bearing a new corpus luteum was performed on day 7 (OPU 1, various phase of the follicular wave, removal of the dominant follicle) and it was repeated on days 10 (OPU 2, growing phase of the follicular wave &ndash; control), 16 (OPU&nbsp;3, growing phase of the first follicular wave after superstimulation) and 20 (OPU 4, growing phase of the second follicular wave after superstimulation). All follicles &gt; 2 mm were punctured. The ovarian follicles (ultrasonographically) and numbers and qualities of obtained oocytes (microscopically) were evaluated during and immediately after OPU. Follicular population was divided to small (FS, 2&ndash;5 mm), medium (FM, 5&ndash;9 mm) and large (FL, &gt; 9 mm) follicles. Oocytes were classified as 1st (intact cumulus, &gt; 3 layers of cumulus cells), 2nd (complete 1&ndash;3 layers of cumulus cells), 3rd (incomplete layers of cumulus cells, expanded cumulus mass) and 4th (absence of corona cells, degenerated oocytes) classes. Although we found the least of FS (x = 1.0) during OPU 3, significantly more FM (x = 24.7) and FL (x = 3.1) follicles were found at this procedure in comparison with others. Likewise a significantly higher number of oocytes (x = 8.1) was obtained at OPU 3 in comparison with OPU 1 and OPU 2. Significantly higher number of FM (x = 6.1) was found and non-significantly higher number of oocytes was obtained at OPU 4 in comparison with OPU 1 and 2. The results show that administration of FSH increases the number of follicles and the number of collected oocytes in the growing phase of the 1st follicular wave after superstimulation, nevertheless a higher number of follicles and a higher recovery rate of oocytes can be expected in the growing phase of the 2nd follicular wave after superstimulation as well.

221 EFFECT OF STAGE OF CORPUS LUTEUM DEVELOPMENT ON THE IN VITRO PRODUCTION OF BOVINE EMBRYOS

2011 ◽  
Vol 23 (1) ◽  
pp. 209
Author(s):  
S. Miyashita ◽  
K. Miyata ◽  
C. Tachibana ◽  
Y. Inaba ◽  
H. Koyama ◽  
...  
Keyword(s):  
Corpus Luteum ◽  
Calf Serum ◽  
Cumulus Cells ◽  
Large Mass ◽  
Bovine Embryos ◽  
In Vitro Production ◽  
Stage Of Development ◽  
The Mean ◽  
Vitro Production

The objective of this study was to investigate the effect of stage of corpus luteum (CL) development on the in vitro production of bovine embryos. Ovaries were classified according to the expected day of the oestrous cycle based on the morphology of the ovaries. Ovaries with a corpus hemorrhagicum and the remnant of the follicular lumen filled with blood were considered the early luteal stage (Days 2 to 4; Day 0 = day of ovulation, n = 46). Ovaries with a large mass of orange tissue in the CL were classified as the midluteal stage (Days 7 to 10, n = 42). Cumulus–oocyte complexes (COC) were collected by aspiration of 2- to 6-mm follicles. The COC were classified into the following grades: COC with >3 compact layers of cumulus cells and evenly granulated cytoplasm were classified into Grade 1; COC with >3 layers cumulus cells and evenly granulated cytoplasm were classified into Grade 2; COC with partially remaining cumulus cells and abnormal cytoplasm were classified into Grade 3; COC without cumulus cells or those with expanded cumulus cells were classified into Grades 4 and 5, respectively. Grades 1 and 2 COC were in vitro matured for 20 h in TCM-199 supplemented with 5% calf serum and 0.02 mg mL–1 of FSH at 38.5°C under an atmosphere of 5% CO2 in air. Matured COC were inseminated with 5 × 106 sperm for 18 h. Presumptive zygotes were cultured in CR1aa medium supplemented with 5% calf serum at 38.5°C under an atmosphere of 5% O2, 5% CO2, and 90% N2 for 9 days (fertilization = Day 0). The mean number of COC and the proportion of COC classified as Grades 1 and 2 were analysed by ANOVA. Cleavage rates on Day 3 and blastocyst rates on Days 7 to 9 were analysed by a chi-square test. The mean number of recovered oocytes in the early luteal stage (18.7 ± 9.5) was significantly higher (P < 0.05) than the number in the midluteal stage (12.2 ± 5.7). The proportion of Grades 1 and 2 oocytes in the early luteal stage [66.7% (531/789)] was significantly higher (P < 0.01) than that in the midluteal stage [51.6% (252/484)]. The cleavage and blastocyst rates in the early luteal stage [60.9% (181/297) and 32.7% (97/297), respectively] were significantly higher (P < 0.05) than those in the midluteal stage [50.7% (76/150) and 20.7% (31/150) respectively].The present study suggests that the stage of development of the CL in bovine ovaries influences the number of recovered oocytes per ovary and the development of in vitro production of bovine embryos.

scholarly journals Synchronization with estradiol benzoate in the presence of the corpus luteum in Wagyu cows increases the number of medium follicles but does not interfere with in vitro production of embryos

2021 ◽  
Vol 42 (3) ◽  
pp. 1147-1158
Author(s):  
Maria Fernanda Zamai ◽  
◽  
Fábio Luiz Bim Cavalieri ◽  
Marcia Aparecida Andreazzi ◽  
Fabio Morotti ◽  
...  
Keyword(s):  
Corpus Luteum ◽  
Estradiol Benzoate ◽  
Breeding Systems ◽  
In Vitro Production ◽  
Embryo Production ◽  
Follicular Wave ◽  
In Vitro Embryo Production ◽  
Oocyte Donors ◽  
Vitro Production

Reproductive biotechnologies are emerging as an important element for livestock; however, some strategies must be modified to adapt to different breeding systems, such as the use of follicular synchronization protocols. This study aimed to evaluate follicular synchronization using estradiol benzoate (EB), in the presence of the corpus luteum (CL) from Wagyu oocyte donors in in vitro embryo production (IVEP). Rounds of IVEP were performed in heifers and cows (n=19) that were classified into three groups: G1/CL - animals with CL, G2/WCL - animals without CL, and G3/CL + EB - animals with CL that were subjected to follicular synchronization with EB at D0. The groups G1/CL and G2/WCL were considered the control and undertook the natural process of follicular dynamics. The results showed that the synchronization of the follicular wave with the application of EB in the presence of CL, presented a smaller number of small (6.05 ± 0.55) and large follicles (0.45 ± 0.15), but increased (P < 0.05) the number of medium-sized follicles (16.20 ± 0.90). However, the results of ovum pick up showed that regardless of whether or not EB was applied, and regardless of the presence or absence of CL in the Wagyu donor, there was no difference among the groups (P > 0.05) concerning the number of viable oocytes and the viability rate. It was concluded that follicular synchronization using EB in Wagyu oocyte donors that presented a CL, increased the number of medium-sized follicles. However, there was no improvement in the efficiency of ovum pick up, in vitro embryo production, and pregnancy rate.

248 SUPERSTIMULATION STRATEGIES FOR OVUM PICKUP IN HOLSTEIN DONORS

2016 ◽  
Vol 28 (2) ◽  
pp. 256
Author(s):  
L. M. Vieira ◽  
G. A. Bó ◽  
R. J. Mapletoft
Keyword(s):  
Oocyte Quality ◽  
Control Group ◽  
Limiting Factor ◽  
Total Dosage ◽  
In Vitro Production ◽  
Total Period ◽  
Expected Time ◽  
Follicular Wave ◽  
Vitro Production

In vitro embryo production (IVP) is an important tool to enhance genetic gain in cattle. However, oocyte quality is a limiting factor for the success of IVP programs in high-producing donors. A series of studies using protocols for follicular wave synchronization and superstimulation before ovum pickup were performed to improve the efficiency of ovum pickup and in vitro production in dairy cattle. The first study evaluated superstimulation with FSH (Folltropin-V®) before ovum pickup in lactating (n = 15) and non-lactating (n = 15) Holstein donors in a crossover design. Cows underwent synchronization of follicle wave emergence (FWE) and at the expected time of FWE, the FSH group received a total dosage of 200 mg of FSH in 4 decreasing doses 12 h apart; controls received no FSH, and ovum pickup was conducted 72 h after the expected FWE in all cows. The FSH-treated cows had a higher (P < 0.01) percentage of medium-sized follicles (6 to 10 mm) at the time of ovum pickup (55.1%) than control cows (20.8%) as well as lower cumulus‐oocyte complexes (COC) recovery rates (60.0 v. 69.8%, respectively; P = 0.002). However, FSH-treated cows had a higher blastocyst production rate (34.5 v. 19.8%; P < 0.01) and more transferable embryos per ovum pickup session (3.0 ± 0.5 v. 1.8 ± 0.4; P = 0.02). Subsequent trials evaluated plasma FSH profiles in 23 heifers and in vitro production following ovum pickup in 90 non-lactating Holstein donors superstimulated with a single IM injection of FSH in 0.5% hyaluronan (HA; MAP-5®, 50 mg). Controls received no treatment, while the F200 group received 200 mg of FSH in 4 decreasing doses 12 h apart. The F200HA and F300HA groups received 200 or 300 mg of FSH in 5 or 7.5 mL, respectively, of 0.5% HA by a single IM injection. Circulating FSH area under curve (AUC) in FSH-treated animals was greater than in the control group (P = 0.02). Although the AUC for F200 group did not differ from HA groups (P = 0.56), the total period of time plasma FSH levels were elevated was greater than in the HA groups (P < 0.01). In the IVP trial, FSH-treated cows had a greater proportion of medium-sized (6–10 mm) follicles than controls (P < 0.001). Also, numbers of follicles (P = 0.01) retrieved (control: 13.1 ± 1.0; F200: 16.5 ± 1.2; F200HA: 19.5 ± 2.1; F300HA: 15.4 ± 1.4; P = 0.01) and blastocysts produced per ovum pickup session (control: 2.4 ± 0.5; F200: 3.7 ± 0.7; F200HA: 4.7 ± 0.7; F300HA: 3.1 ± 0.6; P = 0.06) were greater in cows receiving FSH, regardless of treatment. Cows in the F200HA group had a greater recovery rate (P = 0.009), number of COC cultured (P = 0.04), and blastocysts per ovum pickup session (P = 0.06) than cows in the F300HA group. In conclusion, superstimulation of Holstein donors before ovum pickup increased the efficiency of in vitro production. Additionally, a single IM dose of FSH in 0.5% HA resulted in similar plasma FSH profiles to twice-daily FSH treatment. Non-lactating donors treated with FSH produced more embryos per ovum pickup session regardless of FSH treatment. Lastly, all in vitro-produced endpoints were greater following a single dose of 200 mg of FSH in 0.5% HA than 300 mg of FSH in 0.5% HA.

scholarly journals EFFECT OF THE USE OF TWO SPERM SELECTION TECHNIQUES FOR IN VITRO PRODUCTION OF ALPACA EMBRYOS

SPERMOVA ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 67-72
Author(s):  
Mijail Contreras Huamani ◽  
◽  
Mary Naveros ◽  
Cesar Olaguivel
Keyword(s):  
In Vitro Fertilization ◽  
Cumulus Cells ◽  
Sperm Selection ◽  
Percoll Gradient ◽  
In Vitro Production ◽  
Vitro Fertilization ◽  
Maturation Medium ◽  
Blastocyst Rate ◽  
Vitro Production

The objective of this research was to evaluate the effect of the use of two sperm selection techniques for in vitro production of alpaca embryos. The ovaries and testis were collected from the local slaughterhouse and transport to 37 ° C in saline solution (0.9%) supplemented with gentamicin. Quality I, II and II oocytes were incubated in a maturation medium for 32 h at 38.5 ° C and 5% O2 and 5% CO2. For in vitro fertilization, sperm from the epididymis were selected using the Percoll gradient and Swim up technique. 18h after the oocytes were incubated with the sperm, these were denuded from the cumulus cells and cultured in SOFaa culture medium for 7 days. Morula and blastocyst rate and their morphological quality are evaluated at day 7 of culture. From a total of 370 ovaries, 1,137 oocytes were recovered, making an average of 3.6 oocytes / ovary. After the maturation and fertilization process and in vitro culture, the blastocyst rate was 8.43 ± 6.04% and 3.89 ± 1.75%, for oocytes fertilized with sperm selected with Percoll gradient and Swim up, respectively, not finding significant statistical differences (p> 0.05), between the groups. In conclusion, the in vitro fertilization of alpaca oocytes with spermatozoa selected with two selection techniques (percoll and swim up) did not significantly influence the quantity and quality of morulae and blastocysts at day 7 of embryo culture.

163IMPROVEMENT OF IN VITRO PRODUCTION OF PIG EMBRYOS BY SYNCHRONISATION OF OOCYTE MEIOTIC MATURATION WITH CYCLOHEXIMIDE

2004 ◽  
Vol 16 (2) ◽  
pp. 204 ◽  
Author(s):  
J. Ye ◽  
K.H.S. Campbell ◽  
M.R. Luck
Keyword(s):  
Cumulus Cells ◽  
Treated Group ◽  
Blastocyst Stage ◽  
Developmental Competence ◽  
Meiotic Maturation ◽  
In Vitro Production ◽  
Maturation Medium ◽  
Pre Treatment ◽  
Vitro Production

It is suggested that the relatively high rates of polyspermic fertilization and poor development of pig embryos produced in vitro are caused by asynchronous oocyte maturation. We have recently shown that pre-treatment of pig oocytes with cycloheximide (CHX) is an efficient way of synchronizing their meiotic maturation in vitro. However, it is not known whether this procedure affects fertilization or further development. The present study examined the effects of CHX-synchronised meiotic maturation on subsequent embryo development and the response to FSH. Pig ovaries were collected from a local abattoir. Cumulus-oocyte complexes (COCs) were aspirated from 3–5mm diameter follicles with a translucent appearance and extensive vascularization. COCs were first pre-incubated in defined maturation medium (DM; M199 with Earle’s salts, 25mM HEPES and sodium bicarbonate, 3mM L-glutamine, 0.1% (w/v) BSA, 0.57mM cysteine, 10ngmL−1 EGF, 0.2μgmL−1 pLH, 100μmL−1 penicillin and 0.1mgmL−1 streptomycin) or in DM supplemented with 50ngmL−1 pFSH (DMF) and 5μgmL−1 CHX for 12h. COCs were then further cultured in the same DM without CHX for 24–30h or in DMF for 36h. For controls, COCs were cultured conventionally in DM for 42h or DMF for 48h. After removal of cumulus cells, all cultured oocytes were inseminated with ejaculated sperm at a final concentration of 300000mL−1 for 6h. The IVF medium was modified Tris-buffered medium containing 0.1% BSA, 20μM adenosine and 0.2mM reduced glutathione. Putative embryos were cultured in NCSU23 without glucose but supplemented with 4.5mM Na lactate and 0.33 mM Na pyruvate for 2 days. Cleaved embryos were further cultured in normal NCSU23 for 4 days. IVM and IVF were performed in 5% CO2 in air and IVC in 5% CO2, 5% O2, 90% N2, all at 39°C and 95% RH. Three replicates with DM, with or without CHX, and one with DMF, with or without CHX, were performed with 30–50 oocytes in each replicate. Statistical comparisons were by t-test. The result with DM showed that the rate for normal cleavage at 2 days after insemination of CHX-treated oocytes (40.6±3.8%) was similar to that of controls (40.4±3.5%). However, the proportion developing to healthy blastocysts at Day 6 was significantly higher in the CHX-treated group (16.9±1.2%) than in controls (9.6±1.3%; P&lt;0.05). A significantly higher number of Day 2-cleaved embryos from CHX-treated oocytes developed to the day 6 blastocyst stage compared with controls (44.7±5.0% and 22.3±2.4%, respectively; P&lt;0.05). Supplementation of the basic maturation medium with pFSH increased the rate of cleavage in both CHX-treated oocytes (73.2%) and controls (76.9%) and increased the proportions developing to healthy blastocysts at Day 6 (CHX-treated: 39.0%; control: 11.5%). We conclude that oocytes pre-treated with CHX retain their developmental competence and that meiotic synchronization with CHX improves the efficiency of in vitro production of pig embryos. (Supported by BBSRC 42/S18810.)

Effects of lamb age, hormone stimulation and response to hormone stimulation on the yield and in vitro developmental competence of prepubertal lamb oocytes

2005 ◽  
Vol 17 (6) ◽  
pp. 593 ◽  
Author(s):  
Katherine M. Morton ◽  
Sally L. Catt ◽  
W. M. Chis Maxwell ◽  
Gareth Evans
Keyword(s):  
Recovery Rate ◽  
Blastocyst Stage ◽  
Developmental Competence ◽  
Oocyte Development ◽  
Blastocyst Formation ◽  
In Vitro Production ◽  
Hormone Stimulation ◽  
Vitro Production ◽  
High Responders

Experiments were conducted to determine the effects of lamb age, hormone stimulation (Experiment 1) and response to stimulation (Experiment 2) on the in vitro production of embryos from prepubertal lambs aged 3–4 and 6–7 weeks of age. For 3–4-week-old lambs, hormone stimulation increased the number of follicles (29.9 ± 15.3 v. 70.6 ± 8.2), oocytes per ovary (18.3 ± 6.3 v. 39.3 ± 5.8) and oocyte development to the blastocyst stage (0/192 (0.0%) v. 115/661 (17.4%); P < 0.05). Lamb age (3–4 v. 6–7 weeks old) increased oocyte development to the blastocyst stage (115/661 (17.4%) v. 120/562 (21.4%) respectively). In Experiment 2, hormone-stimulated lambs (3–4 and 6–7 weeks old) were divided into low, medium or high responders based on the number of ovarian follicles (<20, 20–50 and >100 follicles per ovary respectively). The response to hormone stimulation did not affect oocyte recovery rate, but the number of oocytes suitable for culture was increased for high-responding 3–4-week-old lambs only (P < 0.05). Oocyte development to the blastocyst stage was not affected by response to stimulation for 3–4-week-old lambs (15.2–25.6%; P > 0.05), but was reduced for high (6.7%) compared with low (19.5%) and medium (30.9%) responding 6–7-week-old lambs (P < 0.05). These results demonstrate that the production of embryos from prepubertal lambs is increased by hormone stimulation and lamb age and the response to stimulation does not affect embryo production from 3–4-week-old lambs, although by 6–7 weeks of age a high response to stimulation reduces blastocyst formation.

In vitro assessment of corpus luteum function in cattle following Ovsynch and CIDR ovulation synchronization protocols

2004 ◽  
Vol 84 (4) ◽  
pp. 721-724 ◽  
Author(s):  
M. Aali ◽  
J. A. Small ◽  
G. Giritharan ◽  
N. Ramakrishnappa ◽  
K. M. Cheng ◽  
...  
Keyword(s):  
Key Words ◽  
Corpus Luteum ◽  
Hormone Treatment ◽  
Beef Cows ◽  
In Vitro Production ◽  
Pooled Data ◽  
Progesterone Production ◽  
In Vitro Assessment ◽  
Vitro Production

Non-lactating beef cows (N = 40) were used to determine in vitro production of progesterone by CLs collected on days 6–8, 13–15 and 19–20, following Ovsynch or CIDR ovulation synchronization protocols. Progesterone released by the CL tissues into the medium was measured after 1 h of incubation (control) and after 6 h of hormone treatments (LH, PGF2α or LH + PGF2α). In vitro progesterone production did not differ (P > 0.05) between Ovsynch and CIDR ovulation synchronization protocols. Pooled data, irrespective of ovulation synchronization treatments, showed interaction (P < 0.05) between hormone treatment and stage of CL. Key words: Corpus luteum, progesterone, cows, ovulation synchronization

264 EFFECT OF FROZEN SEMEN BATCH ON THE IN VITRO PRODUCTION OF BOVINE EMBRYOS

2010 ◽  
Vol 22 (1) ◽  
pp. 289
Author(s):  
M. B. Fernandes ◽  
T. L. G. Torregrossa ◽  
R. B. Prado ◽  
R. A. Vila ◽  
F. P. Elias ◽  
...  
Keyword(s):  
Embryo Development ◽  
Bos Indicus ◽  
Cumulus Cells ◽  
Development Rate ◽  
In Vitro Production ◽  
Vitro Fertilization ◽  
Significance Level ◽  
Frozen Semen ◽  
Vitro Production

Within an in vitro production controlled system, bulls differ with respect to their semen potential in generating embryos when the variables of maternal effect are minimized (Marquant-le-Guienne and Humblot 1992 Ann. Zootech. 41, 361-370). We have tested the hypothesis that even with this variation among bulls, there is also an intra-bull variation, according to the frozen semen batch used in the in vitro fertilization, identified with the date of ejaculate and its freezing. In an embryo commercial production system, over 12 months, 10 619 viable oocytes were obtained by ultrasound-guided follicular aspiration from 642 Nelore cows (Bos indicus). The oocytes were matured in vitro for 24 h in TCM-199 supplemented with 0.5 μg mL-1 FSH, 50 μg mL-1 LH, and 10% fetal bovine serum. They were then inseminated for 18 hours in IVF-TALP medium, using the semen from 4 bulls (A to D) subdivided into 4 frozen batches (I to IV) and selected by 45/90% Percoll gradient. Putative zygotes surrounded in cumulus cells were transferred in CR2aa medium drops (Rosenkrans and First 1994 J. Anim. Sci. 72, 434-437) for 163 h at 39°C in a humidified atmosphere of 5% CO2 in air. The oocyte distribution, the total number of blastocysts, and the embryo development rate by each bull and respective batch are described in Table 1. The chi-square test was measured with a significance level of P < 0.05 and showed that there is a difference between the used batches of each bull regarding the development rate of blastocysts 163 h after IVF Therefore, there is intra-bull variation in the ability to develop in vitro embryos according to the batch of frozen semen. Table 1.Viable oocytes (VO), total blastocysts (TB), and embryo development rate (%E) by bull and batch used in IVF

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