Glycine and methionine transport by bovine embryos

Zygote ◽  
1997 ◽  
Vol 5 (3) ◽  
pp. 273-276 ◽  
Author(s):  
Catherine Guyader-Joly ◽  
Chaqué Khatchadourian ◽  
Yves Ménézo
Keyword(s):  
Female Genital Tract ◽  
Bovine Embryos ◽  
Mouse Blastocyst ◽  
Cell Numbers ◽  
Significant Difference ◽  
Methionine Uptake ◽  
Methionine Transport ◽  
Female Genital

SummaryAs glycine is one of the most concentrated amino acids in the female genital tract, we investigated its uptake by bovine in vitro matured/in vitro fertilised blastocysts in the presence of increasing concentrations of radiolabelled glycine. We also determined methionine uptake by in vitro and in vivo produced embryos. In our study, the hypothesis of more than one site of enzyme activity for glycine substrate was not validated. We determined a Vmax of 23.4fmol/min per embryo and a Km value of 13.3μM. No significant difference was observed either between in vivo and in vitro derived embryos or between grade 1 and grade 2 embryos for methionine uptake. The methionine and glycine uptake of a day 7 bovine was similar to that of a day 4 mouse blastocyst. This is rather low if we consider the relative cell numbers.

In vitro capacitation of bull spermatozoa by oviductal fluid and its components

Zygote ◽  
2006 ◽  
Vol 14 (3) ◽  
pp. 259-273 ◽  
Author(s):  
Ann-Sofi Bergqvist ◽  
Joan Ballester ◽  
Anders Johannisson ◽  
Marta Hernandez ◽  
Nils Lundeheim ◽  
...  
Keyword(s):  
Female Genital Tract ◽  
Sperm Capacitation ◽  
Merocyanine 540 ◽  
Significant Difference ◽  
High Fluorescence ◽  
The Individual ◽  
Oviductal Fluid ◽  
Female Genital

SummarySperm capacitation is crucial for fertilization. However, debate continues on exactly how, where and when capacitation is elicited in the bovine female genital tract. In this study we used merocyanine-540 and the chlortetracycline (CTC) assay to test how capacitation of bull spermatozoa is affected in vitro by exposure to oviductal fluid (ODF) collected in vivo, various glycosaminoglycans (GAGs) or bicarbonate. Following different durations of exposure, spermatozoa were stained with CTC or merocyanine-540, and evaluated with epifluorescent light microscopy or flow cytometry, respectively. Incubation time did not significantly affect capacitation. Exposure (30–120 min) to ODF capacitated (p < 0.05) bull spermatozoa as measured by either merocyanine-540 or CTC. Hyaluronan was the only GAG that induced a significant increase in B-pattern spermatozoa (capacitated; p = 0.012) compared with controls. Dermatan sulphate also induced capacitation (merocyanine-540 high fluorescence; p = 0.035). Exposure to bicarbonate-enriched media also yielded an increase in merocyanine-540 high fluorescence (p < 0.0001). When bicarbonate was added to the other treatments (ODF or GAGs) an equal increase in merocyanine-540 high fluorescence was noted (p < 0.0001), compared with before addition of bicarbonate and independent of the treatment before exposure. There was no significant difference in the number of B-pattern spermatozoa when bicarbonate was added, but an significant increase in spermatozoa with an acrosome-reacted (AR)-pattern (p < 0.0001) was observed. Exposure of spermatozoa to solubilized zonae pellucidae significantly increased the AR-pattern spermatozoa (p = 0.016). In conclusion, ODF was more potent in inducing capacitation of bull spermatozoa than the individual GAGs. Our results also indicate that bicarbonate is an effector of bull sperm capacitation.

Effects of bovine oviduct epithelial cells, fetal calf serum and bovine serum albumin on gene expression in single bovine embryos produced in the synthetic oviduct fluid culture system

2005 ◽  
Vol 17 (8) ◽  
pp. 751 ◽  
Author(s):  
Mona E. Pedersen ◽  
Øzen Banu Øzdas ◽  
Wenche Farstad ◽  
Aage Tverdal ◽  
Ingrid Olsaker
Keyword(s):  
Gene Expression ◽  
Fetal Calf Serum ◽  
Calf Serum ◽  
Developmental Competence ◽  
Bovine Embryos ◽  
Glut 1 ◽  
Significant Difference ◽  
Oviduct Fluid

In this study the synthetic oviduct fluid (SOF) system with bovine oviduct epithelial cell (BOEC) co-culture is compared with an SOF system with common protein supplements. One thousand six hundred bovine embryos were cultured in SOF media supplemented with BOEC, fetal calf serum (FCS) and bovine serum albumin (BSA). Eight different culture groups were assigned according to the different supplementation factors. Developmental competence and the expression levels of five genes, namely glucose transporter-1 (Glut-1), heat shock protein 70 (HSP), connexin43 (Cx43), β-actin (ACTB) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), analysed as mRNA by using reverse transcription–polymerase chain reaction, were measured on bovine embryos cultured for 9 days. Gene expression of these in vitro-produced embryos was compared with the gene expression of in vivo-produced embryos. There was no significant difference found in embryo developmental competence between the Day 9 embryos in BOEC co-culture, FCS and BSA supplements in SOF media. However, differences in gene expression were observed. With respect to gene expression in in vivo and in vitro embryos, BOEC co-culture affected the same genes as did supplementation with FCS and BSA. HSP was the only gene that differed significantly between in vitro and in vivo embryos. When the different in vitro groups were compared, a significant difference between the BOEC co-culture and the FCS supplementation groups due to Glut-1 expression was observed.

scholarly journals Immune Pathogenesis of Asymptomatic Chlamydia trachomatis Infections in the Female Genital Tract

1995 ◽  
Vol 3 (4) ◽  
pp. 169-174 ◽  
Author(s):  
Steven S. Witkin
Keyword(s):  
Chlamydia Trachomatis ◽  
Genital Tract ◽  
Pregnancy Loss ◽  
Early Stage ◽  
Female Genital Tract ◽  
In Vivo Studies ◽  
Fallopian Tubes ◽  
Female Genital

Chlamydia trachomatis (CT) infections of the female genital tract, although frequently asymptomatic, are a major cause of fallopian-tube occlusion and infertility. Early stage pregnancy loss may also be due to an unsuspected and undetected CT infection. In vitro and in vivo studies have demonstrated that this organism can persist in the female genital tract in a form undetectable by culture. The mechanism of tubal damage as well as the rejection of an embryo may involve an initial immune sensitization to the CT 60 kD heat shock protein (HSP), followed by a reactivation of HSP-sensitized lymphocytes in response to the human HSP and the subsequent release of inflammatory cytokines. The periodic induction of human HSP expression by various microorganisms or by noninfectious mechanisms in the fallopian tubes of women sensitized to the CT HSP may eventually result in tubal scarring and occlusion. Similarly, an immune response to human HSP expression during the early stages of pregnancy may interfere with the immune regulatory mechanisms required for the maintenance of a semiallogeneic embryo.

Development of bovine embryos cultured in CR1aa and IVD101 media using different oxygen tensions and culture systems

2010 ◽  
Vol 58 (4) ◽  
pp. 465-474 ◽  
Author(s):  
Tamás Somfai ◽  
Yasushi Inaba ◽  
Yoshio Aikawa ◽  
Masaki Ohtake ◽  
Shuji Kobayashi ◽  
...  
Keyword(s):  
Inner Cell Mass ◽  
Culture Media ◽  
Cell Mass ◽  
Calf Serum ◽  
Bovine Embryos ◽  
Blastocyst Formation ◽  
Cell Numbers ◽  
Culture Systems ◽  
Significant Difference

The aim of the present study was to optimise the culture conditions for the in vitro production of bovine embryos. The development of in vitro fertilised bovine oocytes in CR1aa supplemented with 5% calf serum and IVD101 culture media were compared using traditional microdrops and Well of the Well (WOW) culture systems either under 5% or 20% oxygen tension. After 7 days of culture, a significantly higher blastocyst formation rate was obtained for embryos cultured in CR1aa medium compared to those cultured in IVD101, irrespective of O 2 tensions and culture systems. The blastocyst formation in IVD101 was suppressed under 20% O 2 compared to 5% O 2 . Despite their similar total cell numbers, higher rates of inner cell mass (ICM) cells were observed in blastocysts developed in IVD101 medium than in those developed in CR1aa, irrespective of O 2 tensions. There was no significant difference in blastocyst formation, total, ICM and trophectoderm (TE) cell numbers between embryos obtained by microdrop and WOW culture systems irrespective of the culture media and O 2 tensions used. In conclusion, CR1aa resulted in higher blastocyst formation rates irrespective of O 2 tension, whereas IVD101 supported blastocyst formation only under low O 2 levels but enhanced the proliferation of ICM cells.

scholarly journals Sperm migration in the genital tract - in silico experiments identify key factors for reproductive success

2020 ◽  
Author(s):  
Jorin Diemer ◽  
Jens Hahn ◽  
Björn Goldenbogen ◽  
Karin Müller ◽  
Edda Klipp
Keyword(s):  
Reproductive Success ◽  
In Silico ◽  
Genital Tract ◽  
Female Genital Tract ◽  
Male Gametes ◽  
Sperm Migration ◽  
Sperm Motion ◽  
Female Genital

Sperm migration in the female genital tract controls sperm selection and, therefore, reproductive success as male gametes are conditioned for fertilization while their number is dramatically reduced. Mechanisms underlying sperm migration are mostly unknown, since in vivo investigations are mostly unfeasible for ethical or practical reasons. By presenting a spatio-temporal model of the mammalian female genital tract combined with agent-based description of sperm motion and interaction as well as parameterizing it with bovine data, we offer an alternative possibility for studying sperm migration in silico. The model incorporates genital tract geometry as well as biophysical principles of sperm motion observed in vitro such as positive rheotaxis and thigmotaxis. This model for sperm migration from vagina to oviducts was successfully tested against in vivo data from literature. We found that physical sperm characteristics such as velocity and directional stability as well as sperm-fluid interactions and wall alignment are critical for success, i.e. sperms reaching the oviducts. Therefore, we propose that these identified sperm parameters should be considered in detail for conditioning sperm in artificial selection procedures since the natural processes are normally bypassed in reproductive in vitro technologies. The tremendous impact of mucus flow to support sperm accumulation in the oviduct highlights the importance of a species-specific optimum time window for artificial insemination regarding ovulation. Predictions from our extendable in silico experimental system will improve assisted reproduction in humans, endangered species, and livestock.

181 DIFFERENCES EXIST IN ZONA PELLUCIDA HARDNESS BETWEEN IN VIVO- AND IN VITRO-GENERATED BOVINE EMBRYOS

2007 ◽  
Vol 19 (1) ◽  
pp. 207 ◽  
Author(s):  
F. Itoi ◽  
T. Himaki ◽  
C. Kubota ◽  
J. Hirose ◽  
M. Miyamura ◽  
...  
Keyword(s):  
Zona Pellucida ◽  
Tactile Sensor ◽  
Bovine Embryos ◽  
Thin Glass ◽  
Standard Curve ◽  
Significant Difference ◽  
Reagent Treatment ◽  
Gelatin Sample

The zona pellucida (ZP) of mammalian ova plays an important role during maturation, fertilization, and early embryonic development. The hardness of ZP of mammalian ova has been mainly evaluated by a biochemical method, such as a difference in ova dissolution speed with an enzyme or an acid reagent treatment. However, the physical hardness of ZP in bovine embryos is largely unknown. Recently, we developed a system measuring the physical hardness of ZP of mammalian ova with a micro bio-sensor (MBS). The objective of this study was to examine the physical ZP hardness of in vivo- and in vitro-derived bovine embryos using an MBS. In the measurement system, the thin glass needle was connected at the tip of a piezo-electric ceramic tactile sensor, and the sample was pushed into a sensor at fixed speed under the computer-controlled micromanipulation system. The change in frequency at the time of displacement of the sample into the sensor was recorded using the computer program. Measurement of gelatin samples of known concentration (4 to 8%) was used to determine the basic characteristic of a sensor and to make a standard curve. In vivo-generated embryos were recovered from 6 superovulated Japanese Black cows with multiple injection of FSH. On Day 7 after insemination, morulae (M), early blastocysts (EB), and blastocysts (BL) were recovered by nonsurgical flushing of uterine horns. In vitro-generated embryos were produced as described earlier (Yoshida et al. 1998 J. Vet. Med. Sci. 60, 549–554). The M, EB, and BL at Days 5 to 7 of post-insemination in vitro were used for measurement of ZP hardness. When a sensor made contact with a harder gelatin sample, the change in frequency was large; the change in frequency has the characteristic of being small for a softer gelatin sample. By comparison with a standard curve, ZP hardness converted into gelatin concentration for each stage of bovine embryos generated in vivo was 3.95% (M: n = 9), 4.14% (EB: n = 32), and 3.92% (BL: n = 14), respectively. On the other hand, ZP hardness of bovine embryos generated in vitro was 3.42% (M: n = 56), 3.33% (EB: n = 36), and 3.25% (BL: n = 23), respectively. There was a significant difference (P &lt; 0.01) in the hardness of ZP between in vivo- and in vitro-generated bovine embryos.

154 THE EFFECT OF OXYGEN TENSION ON IN VITRO DEVELOPMENT OF BOVINE EMBRYOS IN POLYDIMETHYLSILOXANE-BASED WELL OF THE WELL DISHES PREPARED UNDER ATMOSPHERIC OR REDUCED AIR PRESSURE

2010 ◽  
Vol 22 (1) ◽  
pp. 235
Author(s):  
T. Somfai ◽  
Y. Inaba ◽  
Y. Aikawa ◽  
M. Ohtake ◽  
S. Kobayashi ◽  
...  
Keyword(s):  
Oxygen Tension ◽  
Culture Medium ◽  
Production Method ◽  
Total Cell ◽  
Air Pressure ◽  
Bovine Embryos ◽  
Cell Numbers ◽  
Significant Difference ◽  
And Control

Polydimethylsiloxane (PDMS) is a non-toxic silicon compound. Its excellent optical characteristics and easy preparation make it a good candidate material for the molding of custom-shaped dishes for embryo culture. We investigated the feasibility of PDMS-based well of the well (WOW) dishes for in vitro culture of bovine embryos under different oxygen tensions. The WOW dishes with 25 micro-wells (each of 175 μm depth and 250 μm width in diameter arranged in 5 columns and 5 rows) were molded from PDMS prepared either under atmospheric (Experiment 1) or reduced (0.1 MPa) (Experiment 2) air pressure to remove air bubbles. Presumptive zygotes obtained by the in vitro maturation and fertilization of follicular oocytes were placed and cultured for 7 days in traditional micro-drops of culture medium (Control) or in the micro-wells of PDMS-based WOW dishes (PDMS-WOW), both covered by paraffin oil. The culture medium was CR1aa supplemented with 5% calf serum. The culture drop size was 125 μL (5 μL/oocyte) in both groups. Embryo development and blastocyst cell numbers between Control and PDMS-WOW groups were compared either under 20% or 5% O2 tensions. There was no statistical difference in cleavage and blastocyst rates (ranging between 82.3-86.4% and 34.0-45.8%, respectively) between Control and PDMS-WOW embryos irrespective of oxygen tension and dish production method. In Experiment 1, the mean total cell numbers in blastocysts were lower in the PDMS-WOW group than that in Control under 20% O2 (105.0 ± 5.5 and 130.4 ± 9.9, respectively) (P < 0.05, ANOVA); however, the application of 5% O2 significantly improved the cell numbers and eliminated the difference between the PDMS-WOW and Control groups (135.4 ± 6.2 and 148.0 ± 9.0, respectively). In Experiment 2, there was no significant difference in mean total cell numbers in blastocysts between the PDMS-WOW and Control either under 20% O2 (97.2 ± 5.7 and 103.9 ± 8.9, respectively) or 5% O2 (147.5 ± 12.1 and 157.3 ± 3.9, respectively). The numbers and rates of inner cell mass and trophectoderm cells did not differ between the Control and PDMS-WOW groups, irrespective of O2 tension and production method. Our results demonstrate that bovine embryos can develop to the blastocyst stage in PDMS-based WOW dishes; however, it may express detrimental effects on embryonic cell numbers, which can be neutralized by the application of low O2 tension during culture or reduced air pressure during the PDMS preparation. This work was supported by the Research and Development Program for New Bio-Industry Initiatives.

145 CHANGES IN OXYGEN COMSUMPTION RATES OF EMBRYOS IN KOREAN CATTLE

2010 ◽  
Vol 22 (1) ◽  
pp. 231
Author(s):  
C. Y. Choe ◽  
S. R. Cho ◽  
J. K. Son ◽  
S. H. Choi ◽  
C. Y. Cho ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Embryo Quality ◽  
Blastocyst Stage ◽  
Bovine Embryo ◽  
Bovine Embryos ◽  
Cell Stage ◽  
Significant Difference ◽  
Morula Stage

Oxygen consumption has been regarded as a useful indicator for assessment of mammalian embryo quality. This study was carried out to identify whether oxygen consumption rates measured in bovine embryos using SECM can be used as a standard criteria to evaluate bovine embryo quality. Oxygen consumption of bovine embryos at various developmental stages was measured and analyzed using SECM and ANOVA analysis, respectively. We found that the oxygen consumption significantly increased in blastocyst-stage embryos compared to other stage embryos (from 2-cell stage to morula stage), indicating that oxygen consumption reflects the cell number (5.2-7.6 × 1014 mol-1 s-1 v. 1.2-2.4 × 1014 mol-1 s-1, P < 0.05). There was no significant difference between 2-cell-stage embryos and 8-cell-stage embryos. In the morula-stage embryos, the oxygen consumption of in vivo derived embryos was significantly higher than that of in vitro produced embryos (4.0 × 1014 mol-1 s-1 v. 2.4 × 1014 mol-1 s-1, P < 0.05). However, there was no significant difference in consumption of oxygen by in vivo and in vitro derived bovine blastocyst-stage embryos (P > 0.05). Good-quality embryos with grade 1 or 2 showed significantly higher oxygen consumption than grade 3 or 4 embryos. These results showed that SECM could measure oxygen consumption in bovine embryos and the oxygen consumption could reflect embryonic development stage and embryo quality.

Comparative glucose and fructose incorporation and conversion by in vitro Produced bovine embryos

Zygote ◽  
1996 ◽  
Vol 4 (2) ◽  
pp. 85-91 ◽  
Author(s):  
Catherine Guyader-Joly ◽  
Chaqué Khatchadourian ◽  
Yves Ménézo
Keyword(s):  
Free Amino ◽  
Vero Cells ◽  
Carbon Dioxide Production ◽  
Blastocyst Stage ◽  
Bovine Embryos ◽  
Cell Numbers ◽  
By Products

SummaryWe have investigated the quality of bovine IVM/IVF embryos co-cultured on Vero cells. Blastocyst cell numbers are very similar to those obtained in vivo, and higher than those obtained by co-culture with oviduct cells. The metabolism and conversion of fructose and glucose are not equivalent even though carbon dioxide production is similar and increasing from morula to blastocyst. Formation of free amino acids and incorporation into proteins are higher and faster for glucose than for fructose, but this conversion is rather stable with embryonic growth. Moreover, the by-products formed are not the same. Glucose at physiological concentrations (i.e.2 mM)seems to be a more appropriate fuel for the burst of embryonic development at the blastocyst stage in preparation for hatching.

scholarly journals Sperm migration in the genital tract—In silico experiments identify key factors for reproductive success

2021 ◽  
Vol 17 (7) ◽  
pp. e1009109
Author(s):  
Jorin Diemer ◽  
Jens Hahn ◽  
Björn Goldenbogen ◽  
Karin Müller ◽  
Edda Klipp
Keyword(s):  
Reproductive Success ◽  
In Silico ◽  
Genital Tract ◽  
Female Genital Tract ◽  
Male Gametes ◽  
Sperm Migration ◽  
Sperm Motion ◽  
Female Genital

Sperm migration in the female genital tract controls sperm selection and, therefore, reproductive success as male gametes are conditioned for fertilization while their number is dramatically reduced. Mechanisms underlying sperm migration are mostly unknown, since in vivo investigations are mostly unfeasible for ethical or practical reasons. By presenting a spatio-temporal model of the mammalian female genital tract combined with agent-based description of sperm motion and interaction as well as parameterizing it with bovine data, we offer an alternative possibility for studying sperm migration in silico. The model incorporates genital tract geometry as well as biophysical principles of sperm motion observed in vitro such as positive rheotaxis and thigmotaxis. This model for sperm migration from vagina to oviducts was successfully tested against in vivo data from literature. We found that physical sperm characteristics such as velocity and directional stability as well as sperm-fluid interactions and wall alignment are critical for success, i.e. sperms reaching the oviducts. Therefore, we propose that these identified sperm parameters should be considered in detail for conditioning sperm in artificial selection procedures since the natural processes are normally bypassed in reproductive in vitro technologies. The tremendous impact of mucus flow to support sperm accumulation in the oviduct highlights the importance of a species-specific optimum time window for artificial insemination regarding ovulation. Predictions from our extendable in silico experimental system will improve assisted reproduction in humans, endangered species, and livestock.

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