215 COMPARISON BETWEEN CONVENTIONAL DIRECT TRANSFER FREEZING AND VITRIFICATION FOR THE CRYOPRESERVATION OF IN VIVO EMBRYOS FROM BRAHMAN CATTLE

2007 ◽  
Vol 19 (1) ◽  
pp. 224 ◽  
Author(s):  
J. H. Pryor ◽  
C. R. Looney ◽  
D. Walker ◽  
G. E. Seidel, Jr ◽  
J. F. Hasler ◽  
...  
Keyword(s):  
Animal Health ◽  
Blastocyst Stage ◽  
Direct Transfer ◽  
Treatment Groups ◽  
Multiple Ovulation ◽  
Stage Of Development ◽  
Experimental Media ◽  
Commercial Company ◽  
Brahman Cattle

There is a need to develop an efficient cryopreservation technique for Brahman cattle embryos that would lead to improved success in the propagation of this breed. The objective of this study was to compare the post-thaw pregnancy rates in recipient cows after nonsurgical transfer of Brahman in vivo-derived embryos frozen in ethylene glycol (EG) and by a new vitrification (VT) method. Prior to the initial in vivo study, hatching rates were recorded 72 h post-thaw on 3 treatment groups (No treatment: 43/55, 78%; EG: 39/83, 47%; and VT: 33/103, 32%) of IVF embryos to determine if development could be achieved. This warranted Phase I: In vivo embryos were produced by multiple ovulation and nonsurgical embryo collections from Brahman cows 7 days post-estrus and AI. Embryos in morula/blastocyst stage of development were graded and maintained in holding medium (Vigro; Bioniche Animal Health, Inc., Belleville, Ontario, Canada) after collection, until randomly allocated for EG or VT protocols. Vitrification was performed using experimental media provided by a commercial company (V1: Vigro + 5 M EG; V2: Vigro + 7 M EG + 0.5 M Galactose + Ficoll; and Diluent: Vigro + 0.5 M Galactose) and by direct plunging into LN2 (Walker et al. 2005 Reprod. Fertil. Dev. 17, 153). Freezing was performed at 0.5�C min-1 from -6�C to -32�C, using a commercially available medium (Vigro Freeze Plus; Bioniche). All embryos were packaged in sterile 0.25-mL plastic straws to allow for direct transfer (DT). Embryos were stored in LN2, and later thawed and nonsurgically transferred to synchronized recipient cows on Day 7 of their cycle. EG straws were air-thawed 5 s and then thawed in 30�C H2O for 10 s. VT straws were air-thawed 10 s and then thawed in 37�C H2O for 20 s prior to shaking them down to mix columns. Pregnancy results evaluated by ultrasound on Day 35 for the presence of a fetus from 4 replicates were 27% for EG (3/11) and 8% for VT (1/13) embryos. Problems with VT straws cracking during air thawing led to a second trial (Phase II) which changed the direct plunging technique for VT to a vapor freeze of 1 to 15 min prior to plunging into LN2. Pregnancy results from 16 transfers were 38% for EG (3/8) and 50% for VT (4/8) embryos, with no straws cracking. Although the number of observations is small, the results obtained showed that VT-DT is satisfactory for producing pregnancies comparable to EG-DT. Further research is required to confirm the results obtained in this preliminary study, and to test whether this method will allow the successful production of pregnancies under different environmental and field conditions.

scholarly journals 268PREGNANCIES FROM FROZEN IVF CATTLE EMBRYOS USING SEX-SORTED AND UNSORTED SPERM

2004 ◽  
Vol 16 (2) ◽  
pp. 254 ◽  
Author(s):  
R.C. Fry ◽  
C.R. Earl ◽  
F.K. Hollinshead ◽  
D. Wild ◽  
W. Lindemans
Keyword(s):  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Blastocyst Stage ◽  
Control Group ◽  
Adult Cattle ◽  
Treatment Groups ◽  
Transfer Programs ◽  
Stage Of Development ◽  
User Friendly

Previously we demonstrated that sex-sorted sperm could produce IVF embryos from juvenile and adult cattle at rates similar to those for unsorted sperm (Fry et al., 2003 Theriogenology 52, 198). In this study we investigated the pregnancy rates of recipient cattle following the transfer of frozen/thawed IVF embryos generated from young heifers using sex-sorted and unsorted sperm. COCs collected from FSH-stimulated Senepol or Beefex heifers by TVR were matured, fertilized with either sex-sorted or unsorted Senepol sperm and cultured for 6 days under our standard laboratory conditions (Fry et al., 2003 Theriogenology 59, 446, Earl et al., 1997 Theriogenology 47, 255). Embryos reaching the blastocyst or expanded blastocyst stage of development were frozen by the CL-V method of vitrification. Briefly, embryos were equilibrated for 5–10min in HEPES-199 media containing 20% FCS (HM), placed in HM containing 10% EG, 10% DMSO for approximately 2 minutes and then in HM containing 20% EG, 20% DMSO for between 20–60sec (Vatja et al., 1997 Cryoletters 18, 191). Vitrification was achieved by collecting between 5–10 IVF embryos in a 3-μL droplet and securing this droplet to a coded CL-V holder. The droplet was vitrified using the CL-V kit (Lindemans et al., 2004 Theriogenology in press) and then sealed in a precooled ‘‘straw’’ for storage in liquid nitrogen. To thaw, the ‘‘straw’’ with specimen was removed from storage;; the specimen droplet was withdrawn from the ‘‘straw’’ and placed directly into HM containg 0.2M sucrose (SM). After approximately 5–10min each embryo was assessed, loaded into a tomcat catheter in SM and transferred surgically into a recipeint cow within 10–15min of thaw. Of 129 Brahman and Brahman cross cows receiving 2 injections of 125μg cloprostenol 11 days apart, 60 exhibited oestrus 2–4 days after the second injection and 53 were deemed suitable for embryo transfer. Pregnancy was determined by ultrasound on Day 40. No difference in pregnancy rate was found between treatment groups (P>0.05; Table 1). The low submission rate (60/129) and pregnancy rate for the in vivo control group indicate that the fertility of the recipient cows may have been compromised by the drought conditions predominating in Central Queensland. Notwithstanding, the CL-V method for the vitrification of IVF embryos produced by either sex-sorted or unsorted sperm gave similar and very promising pregnancy results of around 40%. This provides new opportunities for the rapid banking of large numbers of sexed IVF embryos generated from elite cattle by TVR for user friendly embryo transfer programs. Table 1 Pregnancies from IVF embryos derived from sex-sorted and unsorted sperm and frozen by the CL-V method of vitrification, or from in vivo embryos frozen in glycerol

Growth and viability of human blastocysts in vitro

2000 ◽  
Vol 8 (3) ◽  
pp. 241-287 ◽  
Author(s):  
GM Jones
Keyword(s):  
Blastocyst Stage ◽  
Culture Conditions ◽  
Cleavage Stage ◽  
Embryo Viability ◽  
Early Cleavage ◽  
Uterine Endometrium ◽  
Stage Of Development ◽  
Uterine Lavage

The transfer of a blastocyst established the first human clinical pregnancy following in vitro fertilization (IVF). Nine years later Cohen et al. reported pregnancies resulting from the transfer of cryopreserved human blastocysts. However, it was another six years before the first report of births resulting from the transfer of human blastocysts produced in vitro appeared in the medical literature. In the intervening period clinics have opted to transfer embryos at the early cleavage stage to the uterus, despite the fact that in vivo the embryo does not enter the uterus until two to three days later at the morula to blastocyst stage of development. The viability and potential for implantation of blastocysts is high, as indicated by the finding that more than 60% of in-vivo-derived blastocysts, recovered by uterine lavage following artificial insemination of fertile donors, implant and develop into viable fetuses when transferred to recipients. This is in stark contrast to the 10–20% of in-vitro-produced embryos transferred at the early cleavage stage of development that result in a live-birth. This reduction in viability following transfer of in-vitro-derived early cleavage stage embryos may have several possible explanations: (1) a failure of implantation due to poor synchronization between the embryo and the uterine endometrium; (2) a hostile environment in the uterus for early cleavage stage embryos; (3) sub-optimal in vitro culture conditions which result in a reduction in embryo viability; (4) the assumption that all oocytes retrieved in an IVF cycle have an equal ability to develop into viable embryos; and (5) the failure to identify the most viable embryo in a cohort. Certainly, improving culture conditions and laboratory techniques for developing high quality blastocysts routinely in vitro will not only address many of the above questions but will also improve the quality and viability of earlier stages of embryo development.

1 ARGININE SUPPLEMENTATION IN VITRO INCREASES PORCINE EMBRYO DEVELOPMENT AND AFFECTSmRNA TRANSCRIPT EXPRESSION

2011 ◽  
Vol 23 (1) ◽  
pp. 107 ◽  
Author(s):  
B. K. Bauer ◽  
L. D. Spate ◽  
C. N. Murphy ◽  
R. S. Prather
Keyword(s):  
Treatment Group ◽  
Mrna Expression ◽  
Embryo Development ◽  
Blastocyst Stage ◽  
Sequencing Analysis ◽  
Treatment Groups ◽  
Porcine Embryo ◽  
Arginine Supplementation

In vitro culture systems are suboptimal as compared to in vivo. Previous next-generation sequencing analysis of in vivo fertilized and in vitro cultured (IVC) or in vivo cultured (IVV) porcine blastocyst stage embryos identified an arginine transporter (SLC7A1) expressed 63 fold higher in IVC compared to IVV blastocysts (Bauer et al. 2010 Biol. Reprod. Epub ahead of print). Arginine catabolism may play important roles in placental and conceptus growth and development as it is a substrate for synthesis of nitric oxide synthase and polyamines. The objective of this study was to determine the effects arginine had on both embryo development and mRNA expression in in vitro fertilized embryos. Cumulus–oocyte complexes were matured for 44 h in M199 supplemented with EGF, FSH, and LH. Oocytes with a visible polar body (metaphase II) were selected and fertilized in modified Tris Buffered Medium for 5 h and then placed into one of 5 treatment groups (Porcine Zygote Medium 3 (PZM3) with 0 mM, 0.12 mM (current concentration of arginine in PZM3), 0.36 mM, 0.72 mM, or 1.69 mM arginine). Twenty-eight hours post-fertilization, cleaved embryos were selected and moved into 25 μL drops of respective culture media and cultured to day 6 in 5% CO2, 5% O2, 90% N2 at 38.5°C. To determine the effect arginine had on development the percent of embryos that made it to the blastocyst stage for each treatment group were analysed using PROC GLM in SAS (SAS Institute, Cary, NC). A least significant difference post test comparison was completed to determine if significant differences existed between treatment groups (a,b,cP < 0.05). The percentage of cleaved embryos on Day 6 that developed to blastocyst was 57.2%b,c, 50.2%c, 67.3%a,b, 67.3%a,b, 70.4%a (N = 147, 163, 150, 120, and 134) in 0 mM, 0.12 mM, 0.36 mM, 0.72 mM, and 1.69 mM arginine, respectively. Real-time PCR was then completed to assess the affect arginine supplementation had on SLC7A1 mRNA expression. Three biological replicates, each containing 10 blastocyst pools to ensure enough starting material, were collected for each treatment group. RNA was isolated from each sample and 5 μL was linearly amplified (NuGEN Ovation Pico WTA System) so multiple genes could be compared and then purified using Bio-Rad MicroSpin Columns. Expression levels were calculated relative to the reference sample and the housekeeping gene, YWHAG. The ΔΔCT values were log-transformed and analysed using PROC GLM in SAS. The expression of SLC7A1 mRNA was decreased (P = 0.0006) compared to PZM3 in the 1.69 mM arginine group. These results illustrate the positive effects that additional arginine may be having on porcine embryo development during culture from the 2-cell to the blastocyst stage. Supplementing arginine to a final concentration of 1.69 mM during culture increases development of porcine embryos to blastocyst compared to PZM3 and also decreases the expression of SLC7A1. Evaluation of the transcriptional profile appears to be a good method of letting the embryo tell us what it needs for development, and in this case arginine. Funded by F21C.

159 USE OF PHYTOHEMAGGLUTININ-L, AS AN AGGLUTINATOR AGENT, TO PRODUCE CHIMERAS OF IVF BOVINE EMBRYOS

2010 ◽  
Vol 22 (1) ◽  
pp. 238
Author(s):  
I. P. Emanuelli ◽  
B. F. Agostinho ◽  
M. P. M. Mancini ◽  
C. M. Barros ◽  
M. F. G. Nogueira
Keyword(s):  
Embryonic Stem ◽  
Blastocyst Stage ◽  
Bovine Embryos ◽  
Cell Stage ◽  
Exact Test ◽  
Fisher's Exact Test ◽  
Stage Of Development ◽  
Fisher’S Exact Test

Embryonic chimeras have been used as a tool to understand embryogenesis and organogenesis, as well as to prove, in vivo, the pluripotency of the embryonic stem cells. One of the techniques used to obtain embryonic chimeras is aggregation, which can be performed with intact or half-embryos and in different stages of the development, produced by in vivo or in vitro systems and in different wells. However, its efficiency tends to reduce when advanced stages, such as morulae and blastocysts, are used. The aim of this work was to evaluate the effect of the treatment with an agglutinating agent (phytohemagglutinin-L; PHA) in the percentage of chimeras produced with IVF bovine embryos. Bovine ovaries (from abattoir) were used to obtain 270 COC that were matured in drops (90 μL) of TCM-199 bicarbonate medium, supplemented with 10% of FCS, and incubated in vitro for 22 to 24 h. The fertilization occurred in TALP-IVF medium, and the COC were maintained in the incubator for 18 h. After fertilization, the presumptive zygotes were transferred to SOF culture medium to in vitro culture. In vitro maturation, fertilization, and culture were performed under 38.5°C, 5% CO2 in air and saturated humidity. The chimerism by aggregation was tested between 2 intact (zona-free) 8- to 16-cell stage embryos in the presence (G1, n = 16) or absence of PHA (G2, n = 14) and between one half-morula and one half-blastocyst with (G3, n = 15) or without PHA (G4, n = 12). The embryos in groups G1 and G3 were treated with PHA in a concentration of 500 μLg mL-1 for 3 min. After PHA treatment, the pairs of embryos were allocated in wells, under previously described culture conditions, until expanded blastocyst stage could be observed (Day 7 of culture). At 24 h of culture, embryonic aggregation pairs were first evaluated to detect only cohesive masses of cells. The results (chimerism rate) were 62.5%, 42.9%, 40.0%, and 25.0%, respectively, for groups G1, G2, G3, and G4. There were no significant differences neither among groups (chi-square, P = 0.252) nor between G1 and G2 (P = 0.464), G3, and G4 (P = 0.683; Fisher’s exact test). Main effects as use of PHA (G1 + G3 v. G2 + G4, P = 0.284) and stage of embryos (G1 + G2 v. G3 + G4, P = 0.183; Fisher’s exact test) were not statistically significant. However, when all groups were compared, the power of the performed test (0.354) was below the desired power of 0.800 (i.e. one must be cautious in over-interpreting the lack of difference among them). In the conditions of this study, it was concluded that the treatment with PHA did not increase the rate of aggregation in the embryonic chimera production, even for half-embryos in advanced stage of development (morulae and blastocysts). Granted by FAPESP, Brazil: 06/06491-2 and 07/07705-9 (MFGN) and 07/04291-9 (MPMM).

27 FOLLICULAR RESPONSE TO DIFFERENT eCG DOSES IN AN OVSYNCH + PROGESTERONE PROTOCOL IN BEEF HEIFERS

2010 ◽  
Vol 22 (1) ◽  
pp. 171
Author(s):  
M. F. Martínez ◽  
D. Tutt ◽  
L. Proctor ◽  
J. L. Juengel
Keyword(s):  
New Zealand ◽  
Animal Health ◽  
Follicular Development ◽  
Dominant Follicle ◽  
Treatment Groups ◽  
Follicular Wave ◽  
Multiple Ovulation ◽  
Array Transducer ◽  
Pfizer Animal Health ◽  
Follicular Response

An experiment was designed to evaluate the effect of different doses of eCG on ovarian follicular dynamics in heifers treated with a Ovsynch plus progesterone protocol. Twenty-five cyclic yearling Black Angus heifers (373.0 ± 35.7 kg), in 2 replicates, received an injection of 100 μg of GnRH (Ovurelin, Bomac Laboratories Ltd., Auckland, New Zealand) i.m. and an intravaginal progesterone device (1.38 g of progesterone; Eazi-Breed CIDR, Pfizer Animal Health, New Zealand) on Day 0 (beginning of the experiment), followed by 500 μg of cloprostenol (PGF, Estrumate, Intervet/Schering-Plough Animal Health, Auckland, New Zealand) i.m. on Day 7, and a second 100 μg of GnRH injection given i.m. on Day 9 (56 h after PGF). At the time of PGF treatment, heifers were randomly assigned to 5 treatment groups to receive 0, 300, 500, 700, or 1000 IU of eCG (Folligon, Intervet/Schering-Plough Animal Health) i.m. Heifers were monitored by ultrasonography (Aloka 900-SSD equipped with a 7.5-MHz linear-array transducer; Aloka, Tokyo, Japan) daily from Day 0 to 9 (GnRH), and then every 12 h until ovulation. Data were analyzed by one-way ANOVA or Kruskall-Wallis test, and means or ranks were compared with LSD or Wilcoxon rank sum tests, respectively. Because a replicate effect was observed (P < 0.05) in the size of the dominant follicle at the second GnRH and prior to ovulation, replicate effect was included in the analysis. The luteal area at PGF treatment was significantly greater (P < 0.01) in heifers that ovulated (750.0 ± 97 mm2) in response to the first GnRH treatment than in those that did not (301.6 ± 42.7 mm2). The diameter of the dominant follicle at the time of PGF treatment was also greater (P < 0.05) in ovulating (11.2 ± 0.4 mm) than in nonovulating (9.7 ± 0.5 mm) heifers. The interval from the first GnRH treatment to the emergence of the next follicular wave was longer (P = 0.50) and more variable in heifers that did not ovulate (2.9 ± 0.4 d; n = 27) than in those that ovulated (1.9 ± 0.2 d; n = 23). There was no effect (P < 0.37) of eCG on the interval from PGF to ovulation (86 ± 1.9 h). The number of ovulations after the second GnRH was higher (P = 0.01) in the group of heifers treated with 1000 IU of eCG (1.8 ± 0.4) than in the other groups (1.0 ± 0.0; 1.1 ± 0.1; 1.2 ± 0.1; 1.0 ± 0.1). There was an effect of day of follicular wave emergence on the number of ovulations (P < 0.01). Heifers with a wave emerging 1 to 3 days after the first GnRH (n = 37), had one ovulation (1.0 ± 0.0), whereas heifers with a wave emerging on Day 4 (3 out of 4 heifers) and Days 5 to 7 (n = 9), ovulated 2 or more follicles. In summary, the multiple ovulation effect occurred when eCG was given to heifers with a follicular wave emerging on or after Day 4, and was potentiated when heifers received 1000 IU of eCG. Although the dose of eCG given at the time of PGF treatment in an Ovsynch program has a significant effect on follicular development, the time of emergence of the dominant follicle appeared to be more important in the ovulation of preovulatory follicle/s after the eCG and the second GnRH treatment.

181 The effects of aging in the oocyte induced by trichostatin A

2020 ◽  
Vol 32 (2) ◽  
pp. 218
Author(s):  
H. A. Arena ◽  
E. Hicks ◽  
M. Mentler ◽  
B. D. Whitaker
Keyword(s):  
Embryonic Development ◽  
Trichostatin A ◽  
Germinal Vesicle ◽  
Blastocyst Stage ◽  
Blastocyst Formation ◽  
Treatment Groups ◽  
Early Maturation ◽  
Stage Of Development ◽  
Developmental Characteristics ◽  
Effects Of Aging

Successful fertilization and embryonic development decreases for an aged oocyte that is not fertilized during the optimal window of time after ovulation. If fertilization of an aged oocyte does occur, the developing embryo is prone to have a delay through meiosis and a decrease in cleavage and blastocyst formation. Trichostatin A (TSA) prohibits the breakdown of the germinal vesicle during meiosis. The objective of this study was to create an invitro model to study the effects of aging in pig oocytes. Oocytes (n=1489) were matured with or without TSA (100ngmL−1) for 24 or 48h followed by an additional 16h of maturation without TSA. A portion of the oocytes were evaluated for stage of meiosis (MI and MII) before maturation (n=95), after the first part of maturation (OMI; 24 or 48h, n=363), or after completing maturation (OMII; 16h, n=230). The remaining oocytes (n=801) were fertilized for 6 to 8h and potential embryos developed. A portion of the embryos were evaluated for fertilization characteristics 12h after IVF (n=400). Remaining embryos (n=401) were evaluated for cleavage by 48h and blastocyst formation by 144h, after IVF. Data were analysed using ANOVA and Tukey's test. Oocytes matured without TSA for 64h had a significantly higher (P&lt;0.05) percentage of oocytes at the MI stage of meiosis compared with all other treatment groups (16.00±6.80), and oocytes matured without TSA for 40h had a significantly higher (P&lt;0.05) percentage of oocytes at the MII stage of meiosis compared with all other treatment groups (14.7±11.30). The results (Table 1) indicate that supplementation of TSA to OMI significantly decreased fertilization penetration rates compared with not supplementing TSA for 24h. However, the percent of embryos cleaved by 48h after IVF was significantly higher in oocytes matured for 40h compared with those matured for 64h. The percent of embryos reaching the blastocyst stage of development by 144h after IVF was significantly lower in oocytes matured for 64h compared with those matured for 40h without TSA. In conclusion, these results suggest that supplementation of TSA during early maturation delays meiosis and decreases fertilization, and increasing maturation time causes a decrease in early embryonic development. By supplementing TSA during early maturation and increasing the length of maturation by 24h, we have created an invitro model to study the effects of aging in pig oocytes. Table 1.Fertilization and developmental characteristics of oocytes1 Amount of TSA added during OMI (ngmL−1) Length of OMI (h) Oocytes penetrated (%) Polyspermicoocytes2 (%) Oocytes with MPN2 (%) Cleavage (%) Day 7 blastocyst (%) 0 24 83.00±3.80a 15.70±4.00 75.90±4.70 77.20±4.20a 27.70±4.20a 0 48 72.00±4.50ab 20.80±4.80 76.40±5.00 58.00±5.00bc 12.00±5.00b 100 24 65.00±4.80b 30.80±5.80 73.80±5.50 68.00±4.70ab 21.00±4.70ab 100 48 56.00±5.00b 30.40±6.20 62.50±6.50 46.00±5.00c 8.00±5.00b a,bMeans with different superscripts in the same column differ (P&lt;0.05). 1TSA, trichostatin A; OMI, oocyte maturation 1. 2Percent of the number of oocytes penetrated.

Nutrient uptake and culture of Sminthopsis macroura (stripe-faced dunnart) embryos

1996 ◽  
Vol 8 (4) ◽  
pp. 685 ◽  
Author(s):  
DK Gardner ◽  
L Selwood ◽  
M Lane
Keyword(s):  
Glucose Uptake ◽  
Nutrient Uptake ◽  
Culture System ◽  
Energy Demand ◽  
Blastocyst Stage ◽  
Cell Stage ◽  
Stage Of Development ◽  
Order Of Magnitude ◽  
Sminthopsis Macroura

Glucose and pyruvate uptake by individual embryos were measured in a marsupial species (stripe-faced dunnart) and a eutherian species (mouse). At each stage of development, nutrient uptake by the dunnart embryo was around an order of magnitude greater than that of the mouse embryo. The pattern of glucose uptake by the dunnart embryo was not like that for any eutherian embryo, all of which have a low glucose uptake before the blastocyst stage. Rather, in the dunnart embryo there was a significant increase in glucose uptake after the third cleavage division, increasing from 13.6 pmol embryo h-1 at the 4-cell stage to 34.9 pmol embryo h-1 by the 8-cell stage. This increase in glucose uptake before blastocyst formation may be attributed to an increased energy demand associated with the movement of cells within the dunnart embryo. Using a new culture system, it was possible to culture 66% of dunnart embryos at the 2-4-cell stage and 80% of those at the 8-16-cell stage to the unilaminar blastocyst stage. Embryos cultured from the 2-cell to the 4-cell stage were retarded by around 12 h when they reached the blastocyst stage. Developmental retardation was also reflected in the pattern of nutrient uptake, which lagged behind that of embryos developed in vivo. The present study has shown that it is possible to culture the early marsupial embryo to the blastocyst stage in a serum-free culture system, while concomitantly quantifying embryonic nutrient requirements. Such an approach is essential for species where there is a paucity of material for study.

scholarly journals Effects of supplementation with heat shock cognate 17 KDA protein (HSC70) on in vitro bovine embryo viability

2021 ◽  
Author(s):  
Aimé Jazmín Garza Arredondo ◽  
Diana Elisa Zamora Ávila ◽  
Uziel Castillo Velásquez ◽  
Gustavo Moreno Degollado ◽  
José Fernando De La Torre Sánchez ◽  
...  
Keyword(s):  
Heat Shock ◽  
Blastocyst Stage ◽  
Vital Role ◽  
Control Group ◽  
Bovine Embryo ◽  
Bovine Embryos ◽  
Embryo Viability ◽  
Stage Of Development

Abstract Endogenous heat shock cognate 71 kDa protein (HSC70) has a vital role in early embryonic development. This study assessed the effects of exogenous HSC70 on bovine embryo development and expression of genes associated with apoptosis. Expression analyses of HSPA1A, HSPA8, Bcl-2, and Bax genes were performed in bovine embryos in vivo on day 7 of development. Subsequently, expression of HSPA1A and HSPA8 were associated with apoptotic genes (Bcl-2 and Bax) in cultured bovine embryos in vitro that were supplemented with various concentrations (0 or control group, 50, and 100 ng) of HSC70. The results indicated that the control group (0 ng) in vitro embryos had higher expression of HSPA8, Bax, and Bcl-2 genes, compared with the vivo embryos (P < 0.01). In vitro-produced embryos supplemented with 50 ng or 100 ng HSC70 had higher expression of HSPA1A, HSC70, Bcl-2, and Bax genes, compared with the control group (P < 0.01). Embryos supplemented with 100 ng had greater expression of the HSPA8 gene compared with the control group and the group supplemented with 50 ng. However, embryos supplemented with 50 ng had better characteristics (i.e., stage of development and quality) than the control and 100-ng groups. In conclusion, supplementation of in vitro culture medium with HSC70 promoted development to the blastocyst stage and improved blastocyst quality.

In vitro culture of embryos produced by in vitro maturation and fertilization of bovine follicular oocytes

1989 ◽  
Vol 1989 ◽  
pp. 14-14
Author(s):  
V. Vergos ◽  
A. Gordon ◽  
M. Gallagher ◽  
I. Gordon
Keyword(s):  
In Vitro Maturation ◽  
Present Report ◽  
Subsequent Development ◽  
Cumulus Cells ◽  
Blastocyst Stage ◽  
Fertilization In Vitro ◽  
Factors Affecting ◽  
Stage Of Development

A previous report from this laboratory dealt with the establishment of pregnancies in the early months of gestation after the non-surgical transfer of cattle embryos derived from the in vitro maturation (IVM) of primary bovine oocytes, their fertilization in vitro (IVF) and their subsequent development to the transferable stage (morula/blastocyst) using an in vivo (sheep oviduct) culture system (Lu et al.,1987). The present report deals with some factors affecting the efficiency of IVF and with the culture in vitro of zygotes to the morula/ blastocyst stage of development. Some embryos were frozen and after thawing transferred by non-surgical procedures to five recipient cattle to obtain information on their capacity to undergo further embryonic development.Primary oocytes, enclosed in cumulus cells, were recovered from vesicular follicles (2-6mm) after their dissection from the ovaries of heifers slaughtered at a local abattoir. The ovaries were brought to the laboratory within one hour of animal slaughter in medium held at 35'C.

scholarly journals Metabolism of Glucose by Preimplantation Mouse Embryos in the Presence of Glucagon, Insulin, Epinephrine, cAMP, Theophylline and Caffeine

1985 ◽  
Vol 38 (4) ◽  
pp. 421 ◽  
Author(s):  
RG Wales ◽  
NK Khurana ◽  
WR Edirisinghe ◽  
I L Pike
Keyword(s):  
Direct Effect ◽  
Glycogen Metabolism ◽  
Blastocyst Stage ◽  
Mouse Embryos ◽  
Stage Of Development ◽  
Preimplantation Mouse Embryos ◽  
Preimplantation Mouse

Neither insulin nor epinephrine influenced the incorporation of glucose into the acid-soluble or acid-insoluble glycogen pool of mouse embryos at the morula-early blastocyst stage during 5 h culture in the presence of radiolabelled glucose. During a 5 h chase culture of pulse-labelled embryos at this stage of development, acid-soluble glycogen labelled during the pulse was not utilized by the embryo but acid-insoluble glycogen was reduced. Addition of glucagon, insulin, epinephrine, cAMP, theophylline or caffeine during chase culture had no effect on the turnover of label in the glycogen pools of the embryo. These results indicate that the turnover of embryonic glycogen observed in vivo is not due to the direct effect of the hormones that regulate glycogen metabolism in the mother.

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