scholarly journals 63INTERFERON-TAU EXPRESSION FROM PRIMARY TROPHECTODERM OUTGROWTHS OF BOVINE BLASTOCYSTS: COMPARISON BETWEEN IVP, NT, AND PARTHENOGENIC EMBRYOS

2004 ◽  
Vol 16 (2) ◽  
pp. 153
Author(s):  
O.M. Ocon ◽  
N.C. Talbot ◽  
A.M. Powell ◽  
A.D. Ealy
Keyword(s):  
Nuclear Transfer ◽  
Culture Medium ◽  
Mixed Linear Model ◽  
Bovine Embryos ◽  
Interferon Tau ◽  
Vitro Fertilization ◽  
Fetal Bovine ◽  
Conditioned Cell Culture Medium ◽  
Bovine Blastocyst

Interferon-tau (IFN-tau) is expressed soon after bovine blastocyst formation and might be useful as a marker of appropriate biological function in embryos produced by nuclear transfer. To assess this possibility we have compared IFN-tau levels in the conditioned medium of primary trophectoderm cultures derived from IVP, nuclear transfer (NT), or parthenogenic bovine embryos. Embryos were produced from in vitro-matured cumulus-oocyte complexes processed from local slaughterhouse ovaries or obtained from Bomed, Inc. (Madison, WI, USA). In vitro fertilization, NT, and parthenogensis were as previously described (Talbot et al., 2000 Tissue and Cell, 32, 9–27) except that embryo culture was in G1/G2 medium in 5% oxygen (Lane et al., 2003 Theriogenology, 60, 407–419). Each 8–11-day embryo was cultured individually in a 4-well plate well (Nunc) with STO feeder cells using DMEM medium containing 10% fetal bovine serum as previously described (Talbot et al., 2000 Biol. Reprod. 62, 235–247). Any contaminating epiblast or endoderm was physically dissected and discarded so as to produce pure trophectoderm outgrowths. The success/failure ratio for colony formation was similar for IVP and NT embryos (IVP=155/29; NT=104/25), but was significantly different (P<0.05) for parthenogenic embryos (54/43). Trophectoderm colonies reached diameters of 1 to 1.5cm in 3–4wk, and, at this time, 72-h-conditioned cell culture medium was harvested, frozen, and measured for IFN-tau anti-viral activity as previously described (Talbot et al., 2000 Biol. Reprod. 62, 235–247). From 313 observations, IFN-tau production was analyzed as a two-factor mixed linear model. Differences in IFN-tau production by type of embryo were statistically significant (F=42.61; P<0.0001; df=2). Mean comparisons were done with Sidak adjusted P-values so that the experiment-wise error was 0.05. IFN-tau production means for IVP-, NT-, and parthenogenic-derived trophectoderm were 4311IUmL−1 (n=155), 626IUmL−1 (n=104), and 1595IUmL−1 (n=54), respectively. The results show that mean IFN-tau production from trophectoderm cultures derived from NT embryos is significantly reduced in comparison to IVP- and parthenogenote-derived cultures. Parthenogenote-derived cultures also produced significantly less IFN-tau than IVP embryos on average. IFN-tau production from trophectoderm outgrowths may be a useful measure of NT reprogramming success.

Secretion of interferon-tau by bovine embryos in long-term culture: comparison of in vivo derived, in vitro produced, nuclear transfer and demi-embryos

1999 ◽  
Vol 55 (3-4) ◽  
pp. 151-162 ◽  
Author(s):  
M Stojkovic ◽  
M Büttner ◽  
V Zakhartchenko ◽  
J Riedl ◽  
H.-D Reichenbach ◽  
...  
Keyword(s):  
Nuclear Transfer ◽  
Bovine Embryos ◽  
Interferon Tau ◽  
Long Term Culture

Effect of protein supplementation on development to the hatching and hatched blastocyst stages of cat IVF embryos

2002 ◽  
Vol 14 (5) ◽  
pp. 291 ◽  
Author(s):  
N. W. Kurniani Karja ◽  
Takeshige Otoi ◽  
Masako Murakami ◽  
Minori Yuge ◽  
Mokhamad Fahrudin ◽  
...  
Keyword(s):  
Bovine Serum ◽  
Culture Medium ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Early Embryo ◽  
Protein Supplementation ◽  
Vitro Fertilization ◽  
Fetal Bovine ◽  
The Mean

The effects of protein supplementation in culture medium on development to the hatching and hatched blastocyst stages of cat in vitro-fertilized embryos were investigated. In the first experiment, presumptive zygotes derived from in vitro maturation and in vitro fertilization (IVF) were cultured in modified Earle's balanced salt solution (MK-1) supplemented with 0.4% bovine serum albumin (BSA) or 5% fetal bovine serum (FBS) for 9 days. There were no significant differences between the BSA and FBS groups with respect to the proportion of cleavage and development to the morula and blastocyst stages of zygotes. However, the presence of FBS in the medium enhanced development to the hatching blastocyst stage of zygotes compared with the BSA group (31.4% v. 7.8%). Moreover, 2.9% of zygotes cultured with FBS developed to the hatched blastocyst stage. The mean cell number of blastocysts derived from zygotes cultured with FBS was significantly higher (P<0.01) than that from zygotes cultured with BSA (136.6 v.101.5). In the second experiment, embryos at the morula or blastocyst stage, which were produced by culturing in MK-1 supplemented with 0.4% BSA after IVF, were subsequently cultured in MK-1 with 0.4% BSA or 5% FBS. Significantly more morulae developed to the blastocyst (P<0.05) and hatching blastocyst stages (P<0.01) in the FBS group than in the BSA group (71.5% and 53.6% v. 44.9% and 6.0%, respectively). Although none of the morulae cultured with BSA developed to the hatched blastocyst stage, 11.5% of morulae cultured with FBS developed to the hatched blastocyst stage. Moreover, the proportion of development to the hatching blastocyst stage of blastocysts was significantly higher (P<0.01) in the FBS group than in the BSA group (68.7% v. 9.8%). None of the blastocysts cultured with BSA developed to the hatched blastocyst stage, whereas 7.3% of blastocysts cultured with FBS developed to the hatched blastocyst stage. The results of the present study indicate that supplementation with FBS at different stages of early embryo development promotes development to the hatching and hatched blastocyst stages of cat IVF embryos.

scholarly journals Expression Profile of Genes as Indicators of Developmental Competence and Quality of In Vitro Fertilization and Somatic Cell Nuclear Transfer Bovine Embryos

PLoS ONE ◽  
2014 ◽  
Vol 9 (9) ◽  
pp. e108139 ◽  
Author(s):  
Maria Jesús Cánepa ◽  
Nicolás Matías Ortega ◽  
Melisa Carolina Monteleone ◽  
Nicolas Mucci ◽  
German Gustavo Kaiser ◽  
...  
Keyword(s):  
In Vitro Fertilization ◽  
Somatic Cell ◽  
Expression Profile ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Developmental Competence ◽  
Bovine Embryos ◽  
Vitro Fertilization

134 EFFECT OF REPLACING FETAL BOVINE SERUM BY DIFFERENT GROWTH FACTORS IN THE PRE-IMPLANTATION DEVELOPMENT OF BOVINE EMBRYOS GENERATED BY IN VITRO FERTILIZATION

2014 ◽  
Vol 26 (1) ◽  
pp. 180
Author(s):  
R. Felmer ◽  
T. Vargas ◽  
R. Sanchez ◽  
M. E. Arias
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Culture Medium ◽  
Fetal Bovine Serum ◽  
Fibroblast Growth Factor 2 ◽  
Bovine Embryos ◽  
Epidermal Growth ◽  
Fetal Bovine ◽  
Defined Culture

Different culture systems have been studied that support pre-implantation development of bovine embryos up to the blastocyst stage. However, the use of chemically defined culture systems has been less studied. The objective of the present study was to evaluate the effect, in the developmental potential of in vitro-produced bovine embryos, of replacing fetal bovine serum (FBS) by different growth factors in the maturation and embryo culture media. In experiment 1, oocytes collected by aspiration of ovaries from a local slaughterhouse were matured in standard TCM-199 culture medium at 38.5°C, 5% CO2, and saturation humidity. The effect of insulin-like growth factor 1 (100 ng mL–1), epidermal growth factor (10 ng mL–1), and fibroblast growth factor 2 (500 ng mL–1) was evaluated at 24 h by the presence of a polar body after removal of cumulus-oocyte complexes. In experiment 2, oocytes matured in vitro in the presence of FBS were fertilized by co-incubation with commercial sperm (mL) for 18 h in standard fertilization medium (Fert-TALP). The presumptive zygotes were denuded and randomly allocated in a chemically defined culture medium based on KSOM supplemented with polyvinyl alcohol (PVA), fructose, and each of the growth factors listed previously. Undefined cultured medium was based on KSOM supplemented with 5% FBS. Embryos were cultured at 38.5°C in a mixture of gases and saturation humidity. Cleavage and blastocyst rates were recorded on Days 3 and 7, respectively. Analysis of variance was used to test for statistically significant differences between groups (P < 0.05) using Stat Graphics Plus 2 Software. In cases where statistically significant differences were observed, a multiple comparison test was run using Tukey's test. In experiment 1, a similar maturation rate was observed in all treatments relative to the undefined maturation medium (range = 88–91%). In experiment 2, no differences were observed in the cleavage (79, 87, 85, and 85%) and the blastocyst rates (24, 25, 26, and 30%) for the epidermal growth factor, insulin-like growth factor 1, fibroblast growth factor 2, and FBS treatments, respectively. In conclusion, we demonstrated that maturation of bovine oocytes can be achieved in chemically defined conditions by replacing FBS by each of the growth factors evaluated herein. Furthermore, chemically defined KSOM medium supplemented by any of these growth factors can generate a similar rate of blastocyst than the undefined medium containing FBS. Analyses are under way to evaluate the effect of completely defined culture conditions (maturation and embryo culture) on the pre-implantation development of embryos produced in the presence of these growth factors.

scholarly journals Fetal bovine serum is associated with polar body degeneration after in vitro maturation of bovine oocytes

2018 ◽  
Vol 68 (3) ◽  
pp. 279
Author(s):  
B. MACÍAS-GARCÍA ◽  
S. MACEDO ◽  
A. ROCHA ◽  
L. GONZÁLEZ-FERNÁNDEZ
Keyword(s):  
Bovine Serum ◽  
Culture Medium ◽  
Fetal Bovine Serum ◽  
Polar Body ◽  
Chromatin Conformation ◽  
Prolonged Incubation ◽  
Vitro Fertilization ◽  
Fetal Bovine ◽  
Bovine Oocytes

In vitro fertilization (IVF) in cattle is commonly used worldwide. Although extensive research has been conducted using different additives in the different IVF steps, little is known regarding how protein type may affect bovine oocytes during the fertilization period. In addition, unlike Tissue Culture Medium 199 (TCM), fertilization medium may induce oocytes’ chromatin degeneration during prolonged incubation in the horse (Modified Whitten’s medium). Thus, in the present work TCM-199 supplemented with either 7 mg/ml of Bovine Serum Albumin (TCM+BSA) or 10% Fetal Bovine Serum (v/v; TCM+FBS) was used. Bovine oocytes were matured in vitro and placed in the previously mentioned media for further 18 hours, in the absence of added sperm (sham fertilization) and their chromatin conformation was evaluated. After IVM, 78.9% of the initial oocytes had reached the MII stage. After sham fertilization, 58.6% of the oocytes in TCM+BSA while just 28.3% in TCM+FBS maintained the MII chromatin conformation (p < 0.05). Subsequent experiments run using PB extruded oocytes and incubated in TCM+BSA and TCM+FBS during sham fertilization, demonstrated that FBS was consistently associated with polar body dissolution or degeneration.

Biochemical markers for pregnancy in the spent culture medium of in vitro produced bovine embryos

2021 ◽  
Author(s):  
Gabriela de Oliveira Fernandes ◽  
Marcella Pecora Milazzotto ◽  
Andrei Antonioni Guedes Fidelis ◽  
Taynan Stonoga Kawamoto ◽  
Ligiane de Oliveira Leme ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Biochemical Markers ◽  
Culture Medium ◽  
Culture Media ◽  
Ionization Mass ◽  
Metabolic Profiles ◽  
Bovine Embryos ◽  
The Media

Abstract The present study aimed to identify biomarkers to assess the quality of in vitro produced (IVP) bovine embryos in the culture media. IVP embryos on Day (D) 5 of development were transferred to individual drops, where they were maintained for the last 48 h of culture. Thereafter, the medium was collected and the embryos were transferred to the recipients. After pregnancy diagnosis, the media were grouped into the pregnant and nonpregnant groups. The metabolic profiles of the media were analyzed via electrospray ionization mass spectrometry, and the concentrations of pyruvate, lactate, and glutamate were assessed using fluorimetry. The spectrometric profile revealed that the media from embryos from the pregnant group presented a higher signal intensity compared to that of the nonpregnant group; the ions 156.13 Da [M + H]+, 444.33 Da [M + H]+, and 305.97 Da [M + H]+ were identified as biomarkers. Spent culture medium from expanded blastocysts (Bx) that established pregnancy had a greater concentration of pyruvate (p = 0.0174) and lesser concentration of lactate (p = 0.042) than spent culture medium from Bx that did not establish pregnancy. Moreover, pyruvate in the culture media of Bx can predict pregnancy with 90.9% sensitivity and 75% specificity. In conclusion, we identified markers in the culture media that helped in assessing the most viable IVP embryos with a greater potential to establish pregnancy.

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