290 CYTOKINE PROFILE IN THE AMNIOTIC FLUID OF NELORE CALVES CONCEIVED BY IN VITRO PRODUCTION AND EMBRYO TRANSFER

2010 ◽  
Vol 22 (1) ◽  
pp. 301
Author(s):  
C. F. Moya-Araujo ◽  
N. C. Preste ◽  
M. Piagentini ◽  
G. H. M. Araujo ◽  
M. G. Silva
Keyword(s):  
Amniotic Fluid ◽  
Embryo Transfer ◽  
In Vitro Production ◽  
Plastic Tube ◽  
Tnf Α ◽  
Significant Difference ◽  
Ifn Γ ◽  
The Moment ◽  
Vitro Production

Cytokines have several functions in many cells and their appropriate balance in the maternal-fetal connections is necessary to maintain a successful pregnancy. The incidence of fetal problems reported at the delivery moment in cows with conceptus conceived by in vitro production (IVP) and lack of literature in the immunology gestation area in bovine prompted this study. The aim of the present study was to determine the levels of cytokines such as tumor necrosis factor a (TNF-α), interferon-y (IFN-γ), interleukin-6 (IL-6), and IL-8 in amniotic fluid of calves from IVP and conventional embryo transfer at the moment of delivery. Forty animals were used and divided into 2 groups: 1 - Twenty Nelore cross-breed cows pregnant with Nelore calves conceived by conventional embryos transfer method (ET group); 2 - Twenty Nelore cross-breed cows pregnant with Nelore calves obtained by IVP after follicular aspiration (IVP group). Around labor, cows were transferred to a maternal paddock to permit observation of delivery. During the expulsion phase the amnion was punctured and 15 mL of fluid was collected in a plastic tube and stored in a freezer for later analysis. The cytokine levels were measured by immunoenzymatic assay (ELISA). The commercial kits used were ESS0011 (Bovine TNF Alpha), ESS0026 (Bovine IFN-gamma), and ESS0029 (Bovine IL-6) from Pierce Biotechnology™ (Rockford, IL, USA) and kit D8000C (Human IL-8) from R&D Systems™ (Minneapolis, MN, USA). The protocol was performed according to the manufacturer’s recommendations. The densities were evaluated by using an automatic optical reader ELISA. The well plates were read at 450 nm. The ANOVA on ranks was used for statistical analysis because of nonparametric distribution of data, with 5% of significance. No calves in this experiment showed signs of sepsis or death during the peripartum period. There was no significant difference in the concentration of TNF-α between the animals of the ET (250.40 pg mL-1) and IVP (398.70 pg mL-1) groups (P > 0.05). There was no difference statistically for IFN-γ; the medians of this cytokine were 35.7 pg mL-1 for the ET group and 48.22 pg mL-1 for the IVP group (P > 0.05). Only samples 15 and 18 of the IVP group had detectable concentrations of IL-6 (2227.47 and 2686.30 pg mL-1, respectively) and for IL-8 only 15 of the samples in the same group (577.6 pg mL-1). The results for TNF-α and INF-γ in this experiment can be adopted as standards for normal pregnancy in cattle, because there are no reports of such data in the literature. However, additional research in the area of gestation immunology area is necessary to describe the role of cytokines in the maintenance of pregnancy and at the moment of delivery in cattle. Supported by FAPESP, Brazil.

scholarly journals BIOCHEMICAL AND HORMONAL PROFILE OF THE AMNIOTIC FLUID FROM NELORE CALVES CONCEIVED BY DIFFERENT REPRODUCTION BIOTECHNOLOGIES

2012 ◽  
Vol 10 (2) ◽  
pp. 205
Author(s):  
Marcelo Piagentini ◽  
Nereu Carlos Prestes ◽  
Carla Fredrichsen Moya-Araujo ◽  
Gustavo Henrique Marques Araujo ◽  
Regina Kiomi Takahira ◽  
...  
Keyword(s):  
Amniotic Fluid ◽  
Chloride Concentration ◽  
In Vitro Production ◽  
Significance Level ◽  
Significant Difference ◽  
Group 3 ◽  
Lower Chloride ◽  
Commercial Kits ◽  
Vitro Production

This study aimed to quantify biochemical constituents and hormonal concentrations in the amniotic fluid of Nelore calves conceived by different reproduction biotechnologies. Sixty animals were used and divided into: 1 – Twenty Nelore cows pregnant of calves obtained by artificial insemination; 2 – Twenty cows pregnant with Nelore calves obtained by embryo transference; 3 – Twenty cows pregnant with Nelore calves obtained by in vitro production. During the expulsion phase, the amnion was punctured and 15 mL of fluid were collected. The biochemical evaluation was performed with commercial kits in accordance to the manufacturer’s recommendations. It was used the radioimmunoassay to assess the progesterone and testosterone levels. The statistical analysis used was the variance analysis and Tukey (5% significance level). Mean urea and creatinine when comparing the three groups showed that group 3 had higher values of these two metabolites, suggesting that the kidney maturity is suitable. For gamma-glutamyltransferase, group 3 had a lower value compared to group 1. This enzyme is related to metabolism, may be inferred that the metabolic activity is high in animal production in vitro. The glucose values did not differ between the groups, suggests that the maternal-fetal exchanges were suitable for the power supply to the fetus. Lower chloride concentration was showed in group 3. Sodium and potassium were found in greater quantities in group 3 when compared to other groups. There was no significant difference for the testosterone and progesterone. The results obtained in biochemical analysis can conclude that the renal and liver activity and maternal-fetal exchange are normal in the different groups.

Determination of anti-Müllerian hormone concentrations in blood as a tool to select Holstein donor cows for embryo production: from the laboratory to the farm

2012 ◽  
Vol 24 (7) ◽  
pp. 932 ◽  
Author(s):  
Charlène Rico ◽  
Laurence Drouilhet ◽  
Pascal Salvetti ◽  
Rozenn Dalbiès-Tran ◽  
Peggy Jarrier ◽  
...  
Keyword(s):  
Embryo Transfer ◽  
Plasma Samples ◽  
In Vitro Production ◽  
Experimental Station ◽  
Multiple Ovulation ◽  
Practical Information ◽  
Vitro Production ◽  
Donor Cows

High between-animal variability in the number of embryos produced by multiple ovulation and embryo transfer (MOET) and ovum pick-up and in vitro production (OPU–IVP) methods remains a major limit to the development of embryo biotechnologies in cattle. The measurement of anti-Müllerian hormone (AMH) endocrine concentrations in cows can help to predict their follicular and ovulatory responses to gonadotrophin treatment. The present study aimed to provide practical information for a simple prognostic method based on AMH measurement in Holstein cows. Accurate AMH concentrations could be measured with ELISA in blood or plasma. In cows undergoing repeated OPU protocols over 1 year, the AMH concentrations measured in plasma samples collected before each gonadotrophin treatment were found to be highly repeatable and were tightly correlated with follicular responses. From data obtained at both an experimental station and farm settings, it was possible to propose AMH cut-off values to identify low-responding cows. Gonadotrophin-stimulated cows producing fewer than 15 large follicles at oestrus and fewer than 10 embryos in MOET protocols could be discarded efficiently with plasma AMH concentrations below 87 and 74 pg mL–1, respectively. In conclusion, we propose a prognostic method based on a single AMH measurement to improve the results of embryo biotechnologies.

scholarly journals Use of Melatonin in the In Vitro Production of Bovine Embryos

2020 ◽  
Vol 21 ◽  
Author(s):  
Alan da Silva LIRA ◽  
Ricardo de Macedo CHAVES ◽  
Felipe de Jesus MORAES JUNIOR ◽  
Sergio Henrique COSTA JUNIOR ◽  
Brenda Karine Lima do AMARAL ◽  
...  
Keyword(s):  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Control Group ◽  
Bovine Embryos ◽  
In Vitro Production ◽  
Maturation Medium ◽  
Significant Difference ◽  
Maturation Rate ◽  
Vitro Production

ABSTRACT We aimed to assess the effects of melatonin in the in vitro production of bovine embryos. Our experiment was conducted at the Laboratório de Reprodução Animal of the Universidade Estadual do Maranhão. The cumulus-oocyte complexes (COCs) were distributed among treatments at concentrations of 0, 10-1, 10-3 and 10-5 µMol/L melatonin. Our experiment was further divided into two: the first was to assess the effect of different concentrations of melatonin (treatments) on the maturation rate of COCs, and the second was to assess the effects of melatonin treatments on the in vitro production of bovine embryos. The results from the first experiment demonstrated no significant difference between the in vitro maturation rate of the cultivated COCs in treatments with melatonin. In the second experiment, however, melatonin treatments yielded statistically higher cleavage, morula and blastocyst rates in the 10-5 µM group (52.9%, 52.9%, and 35.3%, respectively), and lower rates in the 10-1 µM group (19.5%, 19.5% and 7.8%, respectively), compared to the others. The control group (no melatonin) and the 10-3 µM group showed similar results. We concluded that supplementation of melatonin in the in vitro maturation medium resulted in no improvement in the oocyte maturation rate, but in the in vitro production of embryos at different concentrations, the 10-5 µM group displayed better results, but with no improvement in the variables (P < 0.05).

144 PREGNANCY RATES FOLLOWING THE TRANSFER OF IN VITRO FERTILIZED EMBRYOS TO RECIPIENTS ON DAY 6, 7, OR 8 AFTER OESTRUS

2015 ◽  
Vol 27 (1) ◽  
pp. 164
Author(s):  
R. C. Fry ◽  
K. L. Fry ◽  
H. A. McCartney ◽  
W. R. Geddes ◽  
K. Geddes
Keyword(s):  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Pregnancy Rates ◽  
Transfer Program ◽  
In Vitro Production ◽  
Chi Squared ◽  
Grade 3 ◽  
Vitro Production ◽  
Embryo Grade

The aim of this experiment was to investigate the effect of day of synchrony on the pregnancy rate of recipients following the transfer of Day 7 IVF embryos. In addition, the effect of IVF embryo grade and corpus luteum (CL) grade of recipients was determined. A total of 317 cumulus-oocyte complexes collected from 24 dry Brahman cows by TVR were matured, fertilized, and cultured under standard in vitro production procedures (Fry et al. 2003 Theriogenology 59, 446). A total of 89 (44 Grade 1, 43 Grade 2, and 2 Grade 3, IETS classification) in vitro-produced embryos were transferred to parous 4- to 9-year-old dry Brahman cross recipient cattle 7 days after IVF. Two groups of recipient cows were synchronised one day apart with an 8-day CIDR/pg protocol so that oestrous would be concentrated over 3 days with the middle day aligning with the day of IVF (Day 0). Donors that produced a large number of IVF embryos had these divided and transferred into recipients either on Day –1 or Day +1 of synchrony, and those producing less than 4 IVF embryos were transferred into recipients on Day 0. At embryo transfer the ovaries of the recipient were palpated and then scanned by rectal ultrasound and the grade of CL noted (Grade 1 = large distinct CL by palpation, Grade 2 = small distinct CL by palpation, Grade 3 = CL not distinguishable by palpation). Pregnancy was diagnosed by ultrasound scanning on Day 92. Although recipient numbers were low, differences in pregnancy rate between groups were analysed by Chi-squared. Data from the 2 Grade 3 embryos transferred were not included in the analysis (0/2 pregnant). Similar (P > 0.05) pregnancy rates were found when Day 7 IVF embryos were transferred to either Day 6 (17/32 = 53%), Day 7 (9/24 = 38%), or Day 8 (14/31 = 45%) recipients. Furthermore, neither the grade of the embryo (Grade 1: 20/44 = 45%, Grade 2: 20/43 = 47%) nor the grade of recipient CL (Grade 1: 17/45 = 38%, Grade 2: 17/29 = 59%, Grade 3: 6/13 = 46%) effected pregnancy rate (P > 0.05). This experiment demonstrates the flexibility of the IVF embryo to achieve an acceptable pregnancy rate over a range of recipient stages thereby allowing a high usage rate of good-quality recipients in an IVF embryo transfer program.

Practical applications of new research information in the practice of bovine embryo transfer

2010 ◽  
Vol 22 (1) ◽  
pp. 145 ◽  
Author(s):  
C. R. Looney ◽  
J. H. Pryor
Keyword(s):  
Embryo Transfer ◽  
New Technologies ◽  
Bovine Embryo ◽  
In Vitro Production ◽  
Research Information ◽  
Practical Applications ◽  
Cryopreservation Techniques ◽  
New Research ◽  
Vitro Production

For more than 40 years, practitioners have sought to improve all aspects of commercial bovine embryo transfer. The development of new technologies for this industry has been substantial, with recent focus on cryopreservation techniques and the in vitro production of embryos fertilised with sexed spermatozoa. When these and other new technologies are developed, the following questions remain: (1) is said technology regulated or does it require licensing; and (2) is it applicable and, if so, is it financially feasible? Computer access to published research and the advancement of data software programs conducive to the industry for data procurement have been essential for helping practitioners answer these questions by enhancing their ability to analyse and apply data. The focus of the present paper is to aid commercial embryo transfer practitioners in determining new technologies that are available and whether they can be implemented effectively, benefiting their programs.

In vitro production of IFN-γ correlates with CA repeat polymorphism in the human IFN-γ gene

1999 ◽  
Vol 26 (1) ◽  
pp. 1-3 ◽  
Author(s):  
V. Pravica ◽  
A. Asderakis ◽  
C. Perrey ◽  
A. Hajeer ◽  
P.J. Sinnott ◽  
...  
Keyword(s):  
Repeat Polymorphism ◽  
In Vitro Production ◽  
Ca Repeat ◽  
Ifn Γ ◽  
Vitro Production ◽  
Ca Repeat Polymorphism

180 DEVELOPMENTAL CAPACITY OF SINGLE CULTIVATED EMBRYOS IN SMALL AND BIG DROPLETS

2007 ◽  
Vol 19 (1) ◽  
pp. 206
Author(s):  
I. G. F. Goovaerts ◽  
J. B. P. De Clercq ◽  
M. Nichi ◽  
P. E. J. Bols
Keyword(s):  
Blastocyst Stage ◽  
Developmental Competence ◽  
In Vitro Production ◽  
Significant Difference ◽  
Group Treatments ◽  
Developmental Capacity ◽  
Droplet Sizes ◽  
The Individual ◽  
Vitro Production

An in vitro production system where a single oocyte can be followed from the ovary to the blastocyst stage would be a useful tool for studies concerning developmental competence or follicular environment. Unfortunately, until now, only low blastocyst rates could be obtained after single embryo production, and there is still discussion about the ideal droplet size. The objective of the present experiment was to compare the developmental competence of single cultivated zygotes in 20- and 500-µL droplets. Cumulus–oocyte complexes were obtained from slaughterhouse ovaries and were matured and fertilized in groups of 100 for 22 h; the presumptive zygotes were divided into 4 groups. In treatment 1, 25 zygotes were transferred into 50 µL of SOF medium supplemented with 5% serum under oil, whereas in treatment 2, 25 zygotes were transferred into 500 µL of medium. Zygotes were cultivated separately in treatments 3 and 4: in treatment 3 in 20 µL of medium under oil and in treatment 4 in 500 µL of medium. Cleavage rates and division stages were assessed after 3 days of cultivation (5% CO2, 5% O2, 90% N2); blastocyst rates were determined after 7 days. Statistical analysis was performed by logistic regression using SAS (PROC LOGISTIC). There was no difference in cleavage rates between the 2 group treatments or between the 2 single treatments. Also, the division stages were not different between the 2 single treatments (16-cell: 2.0 vs. 1.3%; 8-cell: 25.8 vs. 31.6%; 4-cell: 41.2 vs. 38.0%; and 2-cell: 31.0 vs. 29.1% for the 20 µL and the 500 µL droplet sizes, respectively). Group cultivation after 7 days in 50 µL was significantly better than in 500 µL; however, both treatments resulted in significantly higher blastocyst rates compared with the individual cultures in 20 or 500 µL, between which no significant difference could be found. Noteworthy, only 4-cell and 8-cell stages on Day 3 resulted in blastocysts on Day 7 of cultivation. In conclusion, these results indicate that cultivation in groups gives higher blastocyst rates, although the same embryo density is used as in individual cultivation (1 embryo 20 µL in treatments 2 and 3). Moreover, no significant difference could be found between single cultivation in small and big droplets. This is confirmed by the cleavage stages on Day 3, which indicate no difference in timing of cleaving between small and big droplets; time of cleaving is indicative of further developmental capacity. Table 1.Cleavage and blastocyst rates after single and group cultivation

289 IN VITRO PRODUCTION OF OVINE EMBRYOS FROM WOOL AND HAIR BREEDS

2006 ◽  
Vol 18 (2) ◽  
pp. 252
Author(s):  
E. A. Ordóñez-León ◽  
J. A. Medrano ◽  
V. O. Mejía ◽  
J. De Lucas ◽  
Y. C. Ducolomb ◽  
...  
Keyword(s):  
Physiological Saline ◽  
Relevant Information ◽  
In Vitro Production ◽  
Concentration Gradients ◽  
Independent Variables ◽  
Genetic Potential ◽  
Significant Difference ◽  
Tropical Areas ◽  
Vitro Production

In tropical areas, wool-less or haired breeds of sheep are more prolific than wool breeds. There are no reports about IVF in tropical breeds; therefore, it is not known how these respond under IVF conditions. Developing protocols for in vitro production of embryos in haired breeds could contribute to the preservation and use of their genetic potential in tropical countries where they are economically important. The aim of this study was to determine differences in IVM, IVF, and in vitro (IVD) between wool and hair breeds of sheep. A protocol for IVF (Wani 2002 Small Rum. Res. 44, 89-95) was used in wool (W) and hair (H) breeds. A total of 251 W and 251 H ewes were used. The ovaries were obtained after slaughter and transported to the laboratory in physiological saline (25�C). A total of 411 ovaries from W and 440 from H ewes were used, and 805 follicles of W and 790 of H ewes were aspirated. From these, 663 (82%) cumulus-oocyte complexes (COCs) of Wand 597 (76%) of H ewes were obtained and then used for the procedures of IVM, IVF, and IVD. The average number of COCs recovered per ovary was 1.6 forW and 1.4 for H. The average number of follicles per ovary was 1.9 for W and 1.7 for H. For IVM, COCs were incubated in TCM-199 supplemented with 20% serum from estrous ewe (SEE) for 24 h. All incubations were performed at 38.5�C in a humidified atmosphere of 5% CO2 in air. After this period, COCs were placed in fertilization medium (TALP supplemented with 200�g/mL heparin, 3�g/mL penicillamine, and 1�g/mL hypotaurine). For insemination, frozen-thawed semen from H and W rams was washed by centrifugation in two concentration gradients of a silicone solution. Oocytes and semen from the corresponding breed types were co-incubated for 18 h. For IVD, presumptive zygotes were incubated in SOF medium supplemented with 20% SEE for 7 days. Eight replicates were made. The rates of IVM, IVF, and IVD were analyzed by logistic regression, using as response variables: IVM, IVF, and IVD results, and as independent variables: breed and replicate. The percentage of recovered oocytes was 82% for W and 76% for H. For IVM and IVF, the recovered oocytes produced 535 fertilized oocytes of W and 446 of H. From these, 81% of the W oocytes and 75% of H were fertilized. Oocytes from W showed a higher percentage of IVM and IVF, with a statistically significant difference (P < 0.05). The percentage of division was 63% for W (n = 419) and 52% for H (n = 312). There were no statistically significant differences between the two groups for embryo IVD (P > 0.05). No statistical differences were found between replicates and no interactions were observed for breed � replicate (P > 0.05). It is concluded that IVM, IVF, and IVD procedures used for the development of embryos in W ewes can be used with similar results in H ewes. This is the first report of sheep IVF in Mexico that provides relevant information about the procedures of IVM, IVF, and IVD in hair and wool sheep, andsets a precedent for future investigations on in vitro embryo production in haired sheep breeds in Mexico. Funding for E.A. Ord��ez-Le�n was provided by CONACYT and UNAM.

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