scholarly journals Oocyte developmental competence in a bovine model of reproductive aging

Reproduction ◽  
2007 ◽  
Vol 134 (2) ◽  
pp. 233-239 ◽  
Author(s):  
Pritpal S Malhi ◽  
Gregg P Adams ◽  
Reuben J Mapletoft ◽  
Jaswant Singh

The study was designed to test the hypothesis that aging in cattle is associated with reduced developmental competence of oocytes. The hypothesis was tested by comparing embryo production and pregnancy rates between 13- to 16-year-old cows (n = 6 in Year 1 and n = 9 in Year 2) and their 3- to 6-year-old young daughters (n = 8 in Year 1 and n = 9 in Year 2) after superovulation and transfer of embryos into an unrelated group of young recipients. Embryos were transferred into 2- to 5-year-old recipient cows (n = 99) as singletons (n = 45) or in pairs (n = 54 pairs). Embryo survival in recipients was determined by ultrasonography and by the number of calves born. Between old versus young cows, the number of ovulations (31 ± 4 vs 38 ± 3; P = 0.2) and the number of corpora lutea (25 ± 3 vs 29 ± 2; P = 0.3) did not differ, but fewer (P = 0.04) embryos were recovered from old cows (6 ± 2) than their daughters (12 ± 2). A higher proportion (P < 0.0001) of unfertilized oocytes/uncleaved zygotes were recovered from old cows (222/312, 71%) than their daughters (119/316, 38%). Among the embryos recovered, the proportion of International Embryo Transfer Society Grades 1–2 embryos was similar (P = 0.9) between old (59/90, 66%) and young cows (130/194, 67%). The survival of embryos after transfer into recipients, and the proportion of calves born were also similar between old and young cows. In conclusion, recovery of fewer embryos and a greater proportion of unfertilized oocytes/uncleaved zygotes suggest reduced developmental competence of oocytes from old cows, but there was no difference between age groups in embryo survival after the morula/blastocyst stage.

Author(s):  
Batara Sirait ◽  
Budi Wiweko ◽  
Ahmad Aulia Jusuf ◽  
Dein Iftitah ◽  
R. Muharam

Oocyte developmental competence is one of the determining factors that influence the outcomes of an IVF cycle regarding the ability of a female gamete to reach maturation, be fertilized, and uphold an embryonic development up until the blastocyst stage. The current approach of assessing the competency of an oocyte is confined to an ambiguous and subjective oocyte morphological evaluation. Over the years, a myriad of biomarkers in the cumulus-oocyte-complex has been identified that could potentially function as molecular predictors for IVF program prognosis. This review aims to describe the predictive significance of several cumulus-oocyte complex (COC) biomarkers in evaluating oocyte developmental competence. A total of eight acclaimed cumulus biomarkers are examined in the study. RT-PCR and microarray analysis were extensively used to assess the significance of these biomarkers in foreseeing oocyte developmental competence. Notably, these biomarkers regulate vital processes associated with oocyte maturation and were found to be differentially expressed in COC encapsulating oocytes of different maturity. The biomarkers were reviewed according to the respective oocyte maturation events namely: nuclear maturation, apoptosis, and extracellular matrix remodeling, and steroid metabolism. Although substantial in vitro evidence was presented to justify the potential use of cumulus biomarkers in predicting oocyte competency and IVF outcomes, the feasibility of assessing these biomarkers as an add-on prognostic procedure in IVF is still restricted due to study challenges.


2006 ◽  
Vol 18 (2) ◽  
pp. 271 ◽  
Author(s):  
T. S. Hussein ◽  
R. B. Gilchrist ◽  
J. G. Thompson

Paracrine factors secreted by the oocyte (oocyte-secreted factors, OSFs) regulate a broad range of cumulus cell functions including proliferation, differentiation, and apoptosis. The capacity of oocytes to regulate their own microenvironment by OSFs may in turn contribute to oocyte developmental competence. The aim of this study was to determine if OSFs have a direct influence on bovine oocyte developmental competence during in vitro maturation (IVM). Cumulus-oocyte complexes (COCs) were obtained by aspiration of >3-mm follicles from abattoir-derived ovaries. IVM was conducted in Bovine VitroMat (Cook Australia, Eight Mile Plains, Brisbane, Australia) supplemented with 0.1 IU/mL rhFSH for 24 h under 6% CO2 in air at 38.5�C. In the first experiment, COCs were co-cultured with denuded oocytes (DOs, 5/COC in 10 �L) beginning at either 0 or 9-h of IVM. To generate the 9-h DO group, COCs were first cultured intact for 9-h and then denuded. In the second experiment, specific OSFs, recombinant bone morphogenetic protein-15 (BMP-15) and growth differentiation factor 9 (GDF-9), were prepared as partially purified supernatants of transfected 293H cells, and used as 10% v/v supplements in Bovine VitroMat. Treatments were: (1) control (no supplement), (2) BMP-15, (3) GDF-9, (4) BMP-15 and GDF-9, and (5) untransfected 293H control. Following maturation, in vitro production of embryos was performed using the Bovine Vitro system (Cook Australia) and blastocysts were examined on Day 8 for development. Developmental data were arcsine-transformed and analyzed by ANOVA, followed by Tukey's test. Cell numbers were analyzed by ANOVA. Co-culturing intact COCs with DOs from 0 or 9 h did not affect cleavage rate, but increased (P < 0.001) the proportion of cleaved embryos that reached the blastocyst stage post-insemination (50.6 � 1.9 and 61.3 � 1.9%, respectively), compared to COCs cultured alone (40.7 � 1.4%). Therefore, paracrine factors secreted by DOs increased the developmental competence of oocytes matured as COCs. OSFs also improved embryo quality, as co-culture of COCs with DOs (0 or 9 h) significantly increased total cell (156.1 � 1.3 and 159.1 � 1.3, respectively) and trophectoderm (105.7 � 1.3 and 109.8 � 0.4, respectively) numbers, compared to control COCs (total = 148 � 1.2, trophectoderm = 98.2 � 0.8, P < 0.001). BMP-15 alone or with GDF-9 also significantly (P < 0.001) increased the proportion of oocytes that reached the blastocyst stage post insemination (57.5 � 2.4% and 55.1 � 4.5%, respectively), compared to control (41.0 � 0.9%) and 293H-treated (27.1 � 3.1%) COCs. GDF-9 also increased blastocyst yield (49.5 � 3.9%) but not significantly. These results are the first to demonstrate that OSFs, and particularly BMP-15 and GDF-9, directly affect bovine oocyte developmental competence. These results have far-reaching implications for improving the efficiency of IVM in domestic species and human infertility treatment, and support the role of OSF production by oocytes as a diagnostic marker for developmental competence.


2008 ◽  
Vol 20 (1) ◽  
pp. 215
Author(s):  
J. L. Altermatt ◽  
T. K. Suh ◽  
J. E. Stokes ◽  
L. F. Campos-Chillon ◽  
E. M. Carnevale

Reduced fertility in aged mares is associated with delayed early embryo development and lower pregnancy rates, potentially related to oocyte developmental competence. Human oocyte morphology has been associated with developmental potential, although comparative evidence is lacking in the mare. Exogenous FSH may be beneficial in obtaining more oocytes; however, effects on oocyte morphology and competence are unknown. Objectives were to determine if zona pellucida thickness (ZPT), ooplasm volume (OV), and perivitelline space volume (PVSV) were related to mare age or FSH treatment and to cleavage, blastocyst, and pregnancy rates after intracytoplasmic sperm injection (ICSI). Cycles with and without eFSH treatment were alternated; eFSH treatments began in diestrus with a cohort of follicles ≥20 mm. Oocytes were collected by transvaginal aspiration from follicles >30 mm from young (4 to 9 years) and old (>20 years) mares at 20 to 24 h after administration of recombinant eLH. Oocytes were cultured for 18 h in TCM-199 at 38.5�C in 6% CO2 in air. Sperm were injected 40 � 1 h after eLH, using frozen sperm from a single ejaculate. Presumptive zygotes were incubated in Dulbecco's modified Eagle's medium/F12 + 10% fetal calf serum at 38.5�C in 5% CO2, 5%O2, and 90% N2. Cleavage (≥2 cells) was recorded 48 h after ICSI. Blastocysts considered viable (formation before 9 d and good quality) were transferred nonsurgically into recipients 3 to 7 days after ovulation. Only pregnancies of fetuses with heart beats were included. Morphological parameters of oocytes (old, n = 40; young, n = 37) were obtained from photographic images taken at ICSI and analyzed by computer-assisted measurement using digital calipers (Spot Software, Diagnostic Instruments, Inc., Sterling Heights, MI, USA). Zona pellucida thickness was averaged from 2 measurements 90� to 180� apart. Ooplasm volume was calculated (4/3πr3) from the average of 2 diameters of the ooplasm 90� apart; and PVSV was calculated as the difference of the vitelline membrane volume and that of the volume at the inner volume of the ZP calculated as an oblate spheroid (4/3πa2b) from the average of 2 diameters. Zona pellucida thickness, OV, and PVSV were analyzed using 2-way ANOVA for main effects of age and treatment and 3-way ANOVA by adding cleavage as a factor. Zona pellucida thickness was less (P = 0.007) for old compared with young (least squares mean SEM of 11.4 � 0.2 and 12.3 � 0.2 µm, respectively) with no effect on cleavage, blastocyst, or pregnancy rates. Ooplasm volume was not different (P = 0.14) between old and young (309 036 � 5373 and 320 544 � 5639 µm3, respectively) and did not affect cleavage, blastocyst, or pregnancy rates. The PVSV was greater (P = 0.001) in old compared with young (157 505 � 10 853 and 102 161 � 11 388 µm3, respectively) and may be related to the lower cleavage (P = 0.03), blastocyst (P = 0.02), and pregnancy (P = 0.05) rates. Treatment with FSH had no effect (P > 0.1) on morphology or embryo development. In this study, ZPT and PVSV differed with mare age and could be of predictive value for oocyte developmental competence.


2018 ◽  
Vol 30 (1) ◽  
pp. 163
Author(s):  
S. Ledda ◽  
J. M. Kelly ◽  
S. K. Walker ◽  
Y. Natan ◽  
A. Arav

To advance the use of embryo vitrification technology in veterinary practice, we developed a system in which embryo vitrification, warming, and dilution can be performed within a straw. An in-straw embryo cryopreservation method reduces the need for equipment and technical skills and can facilitate direct embryo transfer to the uterus. This study proposes the use of a new device named “Sarah” that is designed to permit all in-straw embryo cryopreservation procedures. Ovine in vitro-produced (IVP) embryos were vitrified at either early blastocyst stage (EB, n = 65, 6 days post-IVF) or fully expanded blastocyst stage (FB, n = 168, 7 days post-IVF). The vitrification procedure using Sarah constituted a 0.25-mL straw with a capsule having 50-µm pores inserted at one end. Embryos at each stage (EB and FB) were divided into 2 subgroups and vitrified by 1 of 2 methods: (1) multi-step (MS) group-a straw containing 2 embryos was sequentially loaded vertically into 1.5-mL tubes containing 6 different vitrification solutions: 10, 20, 40, 60, 80, or 100% ES (with 100% ES being 7.5% DMSO +7.5% EG + 20% FCS in TCM-199; 90 s each step) followed by 30 s each in 75 and 100% VS (100% VS being 18% DMSO +18% EG + 0.5 M trehalose + BSA in TCM-199); and (2) two-step (TS) group-the straw (2 embryos/straw) was loaded with 100% of ES (5 min), followed by 100% VS solution for 30 s. For both methods, at the end of the preparation steps, the straws were plunged directly into liquid N2. Non-vitrified embryos were maintained in in vitro culture as a control group (n = 102). The warming procedure consisted of placing the straws directly into 5-mL tubes containing 100, 50, 25% WS (WS = 1 M sucrose in TCM-199+ 20% FCS) at 38.6°C (for first solution) and at room temperature for all the rest (5 min each), before being placed into the holding medium. Embryos were recovered from the straws, incubated at 38.6 C in 5% CO2 in air in TCM 199 + 5% FCS, and evaluated for blastocoel re-expansion, embryo survival, and hatching rate at 2, 14, 48 h post-warming. Blastocyst re-expansion (2 h) after warming increased as the developmental stage progressed and was not affected by the vitrification method. In fact, it was significantly (P < 0.05) higher for FB vitrified in the MS and TS methods (77.90% and 71.25%, respectively) compared with the EB method (62.5% and 48.50%, respectively). At 24 h, survival rate of vitrified FB was significantly higher (P < 0.05) in the MS system (95.35%) compared with those in TS (86.25%). Survival rates of FB embryos for both methods (MS and TS) were significantly higher (P < 0.001) than EB embryos vitrified in MS (56.25%) and TS (56.55) methods. After 48 h of culture, the hatching rate for FB vitrified in the MS system (87.21%) was comparable with TS (77.5%) and control (85.3%) groups but significantly higher (P < 0.001) than vitrified EB in MS (43.75%) and TS (36.36%). In conclusion, we showed that a high survival rate of IVP embryos can be achieved by this new in-straw vitrification and warming device (“Sarah”), with hatching rates in vitro comparable with that of control fresh embryos. This method has the potential for use in direct embryo transfer in field conditions.


2020 ◽  
Vol 32 (2) ◽  
pp. 180
Author(s):  
L. Ferré ◽  
M. Kjelland ◽  
T. Stroud ◽  
P. Ross

Invitro embryo production (IVP) has become a reliable alternative for genetic improvement in beef and dairy herds. Maximizing embryo yield and pregnancy per donor are key factors. The aim of this study was to compare ovum pickup (OPU) yields, developmental competence of cumulus-oocyte complexes (COCs), and pregnancy rates from Angus donors in a commercial IVP setting. Donors (&gt;4-year-old pluriparous open dry cows) were handled under the same feeding and environmental conditions. Treatment groups were organised as follows: Group 1: no synchronization (SYNCH; n=5); Group 2: SYNCH with no superstimulation (SOV; n=5); Group 3: SYNCH + SOV (n=5) and OPU 36h after last FSH injection; Group 4: SYNCH + SOV (n=5) and OPU 48h after last FSH injection; and Group 5: SYNCH + SOV (n=5) and OPU 72h after last FSH injection. Follicular waves in groups 2, 3, 4, and 5 were synched by gonadotrophin-releasing hormone (GnRH), prostaglandin F2α (PGF), and controlled internal drug release (CIDR). No pre-synch was used. Injections of FSH (pFSH=180mg, Folltropin) were performed IM twice a day, for three days. A minimum of three replicates were performed for each donor. A Mindray DP30V equipped with a micro-convex transducer 5.0-8.5MHz probe, disposable 20-gauge needle, and a flow rate of 15mL min−1 were used for OPU. All visible follicles (Foll) were punctured and retrieved into a 50-mL 36°C warmed tube with media (phosphate-buffered saline, bovine serum albumin (BSA), and heparin). Viable oocytes were classified according to IETS guidelines. The COCs were matured in 100µL of M199 medium supplemented with ALA-glutamine (0.1mM), Na pyruvate (0.2mM), gentamicin (5µgmL−1), epidermal growth factor (50ngmL−1), oFSH (50ngmL−1), bLH (3μgmL−1), cysteamine (0.1mM), and 10% fetal bovine serum (FBS) for 22 to 24h. Fertilization (Day 0) was carried out using highly fertile sires selected by discontinuous 40%/80% layers (PureSperm) and diluted to a final concentration of 1×106 spermmL−1. Matured oocytes were fertilized in 50µL of modified synthetic oviductal fluid (SOF) media supplemented with fructose (90µgmL−1), penicillamine (3µgmL−1), hypotaurine (11µgmL−1), and heparin (10µgmL−1). After 18h, presumptive zygotes were denuded and cultured under low oxygen tension in 50-µL drops of SOF-BSA for 7 days. On Day 3.5, 2% of FBS was added. On Day 7, fresh transferable (grade 1 and 2, IETS standards) blastocysts were implanted into synchronized recipient cows. Around Day 30, ultrasound diagnosis was performed to determined pregnancy rate (PR). We used ANOVA for comparisons of mean values and X2 test for proportions, α=0.05 (Table 1). In conclusion, synchronization, FSH stimulation, and 48-h coasting before OPU in Angus cows increased the number of collected viable oocytes and embryo development rates. More transferrable embryos and higher rates of PR per OPU were obtained using 36- and 48-h coasting, respectively. Table 1.Follicles (Foll), viable oocytes, cleavage, blastocysts, and pregnancy rates (PR) in Angus cows after ovum pickup (OPU) or invitro embryo production (IVP) SYNCH1 FSH Coasting Foll/OPU Oocytes/OPU Viable oocytes Cleavage,% Embryos,% Embryos/OPU PR,% PR/OPU NO NO NO 10.8a 9.0a 8.9a 63a 25a 2.3a 45a 1.0a YES NO NO 10.1a 8.4a 8.1a 73b 27b 2.2a 51b 1.1a YES YES 36H 16.5b 13.8b 13.6b 70c 29b 4.0b 46a 1.8b YES YES 48H 16.1b 13.4b 13.3b 69c 28b 3.8c 52b 2.0b YES YES 72H 15.6b 13.0b 12.6b 64a 22c 2.8d 55b 1.6c a-dValues with different superscripts in the same column differ (P&lt;0.05). 1SYNCH=synchronized.


2008 ◽  
Vol 20 (9) ◽  
pp. 71
Author(s):  
M. Bertoldo ◽  
P. K. Holyoake ◽  
G. Evans ◽  
C. G. Grupen

Reduced farrowing rate caused by embryonic mortality is a manifestation of seasonal infertility in pigs. The ability of the oocyte to mature, be fertilised and sustain embryonic development is acquired gradually by the oocyte throughout folliculogenesis. This study was undertaken to determine if seasonal differences in ovarian morphology are associated with reduced reproductive performance displayed during seasonal infertility. Sows culled after weaning were sourced from two genetically distinct herds (Farms A and B). Pairs of ovaries were collected from sows 4 days post-weaning during winter (n = 131) and summer (n = 275). Ovarian weight (Farm A only) and the numbers of small (3–4 mm) and large (5–8 mm) follicles were assessed (Farms A and B). Data did not follow normality and was analysed using the Mann–Whitney test. Mean ovarian weight per sow during winter (20.0 ± 1.3 g) was significantly heavier than that during summer (15.3 ± 0.8 g; P < 0.05). Farm A ovaries had a greater total number of antral follicles in winter compared with summer, and a greater number of antral follicles in winter compared with Farm B ovaries (P < 0.05). In Farm A ovaries, the proportion of follicles that were large was greater in summer compared with winter (78% v. 66%; P < 0.05), but the follicular distribution did not change with season in Farm B ovaries. While the findings demonstrate that ovarian weight was greater in winter compared with summer, they suggest that this difference was not the result of changes in the number or distribution of surface antral follicles. The difference in ovarian weight is possibly due to differences in ovarian tissue resulting from regressed corpora lutea. Further studies are being undertaken to assess the effect of season on oocyte developmental competence and the steroid content of follicular fluid isolated from small and large follicles.


Medicina ◽  
2019 ◽  
Vol 55 (6) ◽  
pp. 293 ◽  
Author(s):  
Kontopoulos ◽  
Simopoulou ◽  
Zervomanolakis ◽  
Prokopakis ◽  
Dimitropoulos ◽  
...  

Background and Objective: During the last few years, a trend has been noted towards embryos being transferred at the blastocyst stage, which has been associated with improved rates regarding implantation and clinical pregnancy in comparison to cleavage stage embryo transfers. There is a limited number of studies investigating this notion in oocyte donation cycles employing cryopreserved embryos. The aim of this study is to evaluate the implantation potential and clinical pregnancy rates between the day 3 cleavage stage and blastocyst stage embryo transfers in oocyte donation cycles employing vitrified embryos. Methods: This is a retrospective evaluation of oocyte donation frozen–thawed transfers completed in our clinic from January 2017 to December 2017. Intracytoplasmic sperm injection was conducted for all oocytes. Following fertilization, all embryos were cryopreserved either at the cleavage or blastocyst stage. Embryo transfer of two embryos was performed under direct sonographic guidance in all cases. Results: Our results confirmed a 55.6% clinical pregnancy (CP) resulting from day 3 embryo transfers, a 68.8% CP from day 5, and 71.4% CP from day 6. Significantly improved pregnancy rates were related to embryo transfers at the blastocyst stage when compared to cleavage stage transfers (68.9% and 55.6% respectively, p = 0.016). The risk with regards to multiple pregnancies was similar. Conclusion: Our findings indicate that in oocyte donation cycles employing vitrified embryos, embryo transfer at the blastocyst stage is accompanied with a significant improvement in pregnancy rates and merits further investigation.


2004 ◽  
Vol 16 (2) ◽  
pp. 212 ◽  
Author(s):  
T. Nishisouzu ◽  
M. Sugawara ◽  
S. Aoki ◽  
K. Kishida ◽  
M. Moriyoshi ◽  
...  

Treatments with GnRH and PGF2α for synchronization of ovulation has resulted in acceptable pregnancy rates after fixed-time artificial insemination in dairy cows without estrus detection. The objective of the present study was to evaluate the practicability of ovulation synchronization (Ovsynch, Pursley JR et al. 1995 Theriogenology 44, 915–923) in dairy cattle using GnRH and PGF2α for the embryo transfer recipients. Dairy cattle (cows; n=100, heifers; n=88) were randomly allocated to one of two groups. The control group (cows; n=45, heifers; n=37) was composed of cows in natural estrus. The ovulation synchronization group (cows; n=55, heifers; n=51) was treated with an intramuscular injection of 100μg of GnRH at a random stage of the estrous cycle. Seven days later, the cattle received PGF2α (Cows; 25–30mg) or PGF2α analog (Heifers; 0.5mg) in order to regress the corpora lutea (CL). Forty-eight hours later, cows and heifers received a second injection of 100μg GnRH. Embryo transfer was carried out 7 days after the second injection of GnRH in the ovsynch group and 7 days after estrus in the control group. The cattle judged to have CL 17mm were classified as acceptable recipients. The size of the follicles and the CL were determined to be of estrus stage and embryo transfer by means of ultrasonography. The mean numbers of follicles and CL were analyzed by ANOVA, while pregnancy rates were analyzed by chi-square test. The results are presented in the Table. The proportion of cows and heifers determined to be acceptable embryo transfers was not different between the control group and the ovsynch group. There were no differences in the proportion of acceptable embryo transfers between the control group and the ovsynch group. Follicle diameter at the time of estrus in the control group (cows; 20.7±0.7mm, heifers; 16.8±0.5mm) were significantly larger than that of the ovsynch group (cows; 18.0±1.0mm, heifers; 14.7±0.2mm) (P&lt;0.05). Although CL diameter at the time of embryo transfer in heifers showed no differences between the control group and the ovsynch group (25.0±1.0mm v. 22.8±1.5mm), The CL diameter of the control cow group was larger than that of the ovsynch group (29.8±0.7mm v. 26.1±1.0mm, P&lt;0.05). However, no differences in pregnancy rate were seen between the control group and the ovsynch group. These results suggest that ovsynch can be effectively applied in an embryo transfer program for cattle. Table 1 Proportion of acceptable embryo transfer recipients and pregnancy rate in dairy cattle in the control ovsynch groups


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