scholarly journals Characterization of secreted proteins of 2-cell mouse embryos cultured in vitro to the blastocyst stage with and without protein supplementation

2014 ◽  
Vol 31 (6) ◽  
pp. 757-765 ◽  
Author(s):  
Tanya Burch ◽  
Liang Yu ◽  
Julius Nyalwidhe ◽  
Jose A. Horcajadas ◽  
Silvina Bocca ◽  
...  
Keyword(s):  
Blastocyst Stage ◽  
Protein Supplementation ◽  
Mouse Embryos ◽  
Secreted Proteins

Viability and growth of mouse embryos after in vitro culture and fusion

Development ◽  
1970 ◽  
Vol 23 (3) ◽  
pp. 693-704
Author(s):  
Patricia Bowman ◽  
Anne McLaren
Keyword(s):  
In Vitro Culture ◽  
Success Rate ◽  
Zona Pellucida ◽  
Growth Regulation ◽  
Excess Mortality ◽  
Blastocyst Stage ◽  
Mouse Embryos ◽  
Foster Mothers ◽  
Mouse Eggs

About 80 % of 8-cell mouse eggs developed to the blastocyst stage in culture, whether the zona pellucida was left intact, or removed with pronase (pre-incubated and dialysed) and the eggs then cultured singly or as fused pairs. When pronase was used without prior incubation and dialysis, the success rate was reduced to 50 %. After transfer to uterine foster-mothers, 20–30 % of apparently normal blastocysts cultured with or without the zona, singly or fused, developed into live foetuses, compared with over 50 % of control blastocysts taken directly from the uterus. Some of the excess mortality of cultured embryos took place before implantation and some soon after. The foetuses derived from cultured blastocysts averaged 0·1 g lighter than those derived from control uterine blastocysts similarly transferred. No differences in the weights of the placentae were observed. Foetal and placental weights were unaffected by whether the eggs had been cultured singly or fused, implying that growth regulation of fused embryos is complete by the 17th day of gestation. The longer the eggs were maintained in culture, the lower was their viability after transfer, and the lighter were the foetuses derived from them.

Normal adenylate ribonucleotide content in mouse embryos homozygous for the t12 mutation

Development ◽  
1983 ◽  
Vol 78 (1) ◽  
pp. 43-51
Author(s):  
Horst Spielmann ◽  
Robert P. Erickson
Keyword(s):  
In Vitro Culture ◽  
Normal Values ◽  
Firefly Luciferase ◽  
Blastocyst Stage ◽  
Mouse Embryos ◽  
Luciferase Assay ◽  
Preimplantation Mouse Embryos ◽  
Preimplantation Mouse

The recently improved firefly luciferase assay was used to determine ATP, ADP or AMP in single preimplantation mouse embryos from crosses yielding lethal t12/t12 embryos. Normal values of the three adenylate ribonucleotides were found in freshly collected 2-cell and 4-cell embryos and during in vitro culture to the blastocyst stage. A decrease in adenylate ribonucleotide content was seen in putative t12/t12 embryos only when they were degenerating.

Lethality of a tritiated amino acid in early mouse embryos

Development ◽  
1985 ◽  
Vol 88 (1) ◽  
pp. 209-217
Author(s):  
Janet L. Wiebold ◽  
Gary B. Anderson
Keyword(s):  
Specific Activity ◽  
Subsequent Development ◽  
Blastocyst Stage ◽  
Mouse Embryos ◽  
Radiation Induced ◽  
Specific Activities ◽  
Tritiated Amino Acids ◽  
Early Mouse

2- to 4-cell and morula- to blastocyst-stage mouse embryos were cultured for 1 h in tritiated leucine at two specific activities and their subsequent development followed in vitro and in vivo (after transfer to recipients), respectively. 2- to 4-cell embryos that incorporated an average of 42 d.p.m. per embryo were impaired in their ability to develop to the morula and blastocyst stage. Recipients receiving morulae and blastocysts that had incorporated an average of 384 d.p.m. per embryo failed to produce young. Reduction of the specific activity improved the viability of embryos both in vitro and in vivo but development was still less than that of unlabelled embryos. Protein degradation curves were different for both 2- to 4-cell and morulato blastocyst-stage embryos labelled at the two different specific activities. Most studies using tritiated amino acids have employed higher specific activities than those used here and they may have to be reevaluated due to the possibility of radiation-induced artifacts.

Localization of cytoskeletal proteins in preimplantation mouse embryos

Development ◽  
1980 ◽  
Vol 55 (1) ◽  
pp. 211-225
Author(s):  
E. Lehtonen ◽  
R. A. Badley
Keyword(s):  
Blastocyst Stage ◽  
Cytoskeletal Proteins ◽  
Mouse Embryos ◽  
Trophectoderm Cells ◽  
Apparent Concentration ◽  
Preimplantation Mouse ◽  
Early Mouse ◽  
Filament Protein

The immunofluorescence technique was used to detect the presence and distribution of actin, alpha-actinin, tubulin and 10 nm filament protein in early mouse embryos. Actin and alpha-actinin stainings showed a distinct concentration to a peripheral layer in the cleavage-stage blastomeres and in trophectoderm cells. Dots of fluorescence appeared in this cortical staining pattern. The distribution of tubulin staining in the blastomere cytoplasm was relatively even with apparent concentration at the perinuclear region and frequently at wide intercellular contact areas. 10 nm filament protein was distributed evenly in the blastomere cytoplasm without cortical concentration of the label. At the blastocyst stage, the trophectoderm cells in blastocyst outgrowths in vitro developed well organized cytoskeletons including both microfilament, microtubule and 10 nm filament elements. Comparable structures were not observed in blastocysts in vivo, or in late hatched blastocysts cultured in suspension. The morphogenetic significance of the observations is discussed.

Potential of two-cell mouse embryos to develop to term despite partial damage after cryopreservation

1995 ◽  
Vol 29 (3) ◽  
pp. 320-326 ◽  
Author(s):  
Th. Rülicke ◽  
P. Autenried
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Blastocyst Stage ◽  
Mouse Embryos ◽  
Cell Stage ◽  
Freeze Thaw ◽  
Foster Mothers ◽  
Vital Stains ◽  
Partial Damage

Approximately 18% of cryopreserved 2-cell mouse embryos of 26 different batches showed various degrees of morphological damage after the freeze-thaw process. Normal and damaged morphology were assessed by light microscopy and the ability of an embryo to develop in vitro to a blastocyst, or to develop to term, after transfer to foster mothers. Using vital stains such as Fluorescein-diacetate (FDA) and 4',6-Diamidino-2-Phenylindole (DAPI) it was found that in approximately 82% of the cases, both of the 2 blastomeres of the cryopreserved embryos survived the freeze-thaw process; in 10% only one cell survived the process; and in 8% none survived. Normally, only intact 2-cell embryos are considered for transfer. Here it was shown that over 60% of the partially damaged embryos developed in vitro to the blastocyst stage and, of those, 26% developed to term after transfer to suitable foster mothers. Although the inner cell mass (ICM) appeared to remain smaller during culture after the transfer of partially damaged 2-cell stage embryos, no difference during gestation period was found compared with intact embryos.

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.

The effect of microtubule- and microfilament-disrupting drugs on preimplantation mouse embryos

Development ◽  
1980 ◽  
Vol 60 (1) ◽  
pp. 71-82
Author(s):  
G. Siracusa ◽  
D. G. Whittingham ◽  
M. De Felici
Keyword(s):  
Blastocyst Stage ◽  
Mouse Embryos ◽  
Microtubule Assembly ◽  
Continuous Exposure ◽  
Blastocyst Formation ◽  
Multipolar Spindles ◽  
Preimplantation Mouse Embryos ◽  
Preimplantation Mouse ◽  
Continuous Presence

The sensitivity of early preimplantation mouse embryos to drugs which disrupt microfilament function (cytochalasin B-CB and cytochalasin D-CD) and microtubule assembly (colchicine, colcemid, vinblastine and griseofulvin) was examined. CD inhibited cleavage at a concentration 35-fold lower than CB (3 × 10−7 M ν. 1 × 10−5 M). Treatment of 2-cell embryos for 6 h with 1 × 10−5 M CB or 1 × 10−6 M CD or continuous exposure to lower concentrations of CB or CD did not affect development to the blastocyst stage in vitro. Vinblastine inhibited cleavage at a concentration tenfold lower than colcemid or colchicine (1 × 10−8 M ν. 1 × 10−7 M). The continuous presence of colcemid at 10−8 M did not affect the development of 2-cell embryos to the blastocyst stage, but development was reduced with vinblastine at 1 × 10−8 M and completely inhibited with colchicine at 1 × 10−8 M. The drugs produced similar responses when 2-cell embryos were treated for 6 h with concentrations that inhibited cleavage. Complete inhibition of cleavage was obtained after only a 2 h exposure to 2 × 10−7 M colchicine. A similar concentration of lumicolchicine did not affect cleavage or blastocyst formation. Embryos were less sensitive to griseofulvin; the first cleavage division was unaffected by concentrations as high as 3 × 10−4 M and only 50% of 2-cell embryos failed to cleave in 1 × 10 1 and 3 × 10−4 M griseofulvin. At these concentrations a small proportion of 1-cell embryos and the majority of the 2-cell embryos showed unequal cytoplasmic division probably caused by the formation of multipolar spindles. The continuous exposure of 2-cell embryos to 3 × 10−5 M griseofulvin did not affect blastocyst formation.

A simplified biopsy method for precompacted mouse embryos: A technical report

2002 ◽  
Vol 50 (4) ◽  
pp. 469-479 ◽  
Author(s):  
S. Bodó ◽  
L. Laczkó ◽  
Gabriella Horváth ◽  
Keyword(s):  
Genetic Diagnosis ◽  
In Utero ◽  
Blastocyst Stage ◽  
Mouse Embryos ◽  
Embryo Biopsy ◽  
Control Groups ◽  
Technical Report ◽  
Significant Difference ◽  
Biopsy Method

This article presents a new, simple and rapid embryo biopsy method. The blastomere for genetic analysis can be separated from a precompacted mouse embryo after a partial zona digestion with the use of a holding pipette. For the micromanipulation only two microcapillaries and micromanipulators are needed. The development of the biopsied embryos was studied during in vitro culture and in utero following embryo transfer. There was no significant difference between the treated and the control groups in the ratio of embryos that developed to the blastocyst stage, although the biopsied embryos were delayed in their development because they contained significantly fewer cells compared to the control ones at the same stage. Although there was no difference in the ratio of implantation, the development of the biopsied embryos in utero was also delayed 12-24 hours on the 9th day of pregnancy. No difference in development was visible from the 13th day of pregnancy. Statistically, no differences were found in the developmental ratio (number of developed fetuses/transferred embryos) of the control and treated embryos during gastrulation (9th day of pregnancy), at the beginning of organogenesis (13th day of pregnancy) and before birth (19th day of pregnancy). The embryo biopsy method presented here can be a new and useful tool for preimplantation genetic diagnosis.

Open-pulled straw (OPS) vitrification of in vitro fertilised mouse embryos at various stages

2008 ◽  
Vol 56 (2) ◽  
pp. 245-253 ◽  
Author(s):  
Chang-Liang Yan ◽  
Qi-En Yang ◽  
Guang-Bin Zhou ◽  
Yun-Peng Hou ◽  
Xue-Ming Zhao ◽  
...  
Keyword(s):  
Survival Rate ◽  
Developmental Stages ◽  
Developmental Rate ◽  
Blastocyst Stage ◽  
Significant Loss ◽  
Mouse Embryos ◽  
Cell Stage ◽  
Fresh Embryos

The present study was designed to investigate the cryotolerance of in vitro fertilised (IVF) mouse embryos at various preimplantation developmental stages. IVF mouse embryos were vitrified by the open-pulled straw (OPS) method. After warming, embryos were morphologically evaluated and assessed by their development to blastocysts, hatched blastocysts or term. The results showed that a high proportion (93.3–100.0%) of vitrified embryos at all developmental stages were morphologically normal after recovery. The developmental rate of vitrified 1-cell embryos to blastocyst (40.0%) or hatched blastocyst (32.7%) or term (9.3%) was significantly lower than that from other stages (P < 0.05). Vitrified embryos from 2-cell to early blastocyst stage showed similar blastocyst (71.8–89.5%) and hatched blastocyst rates (61.1–69.6%) and could develop to term without a significant loss of survival compared with those of fresh embryos (P > 0.05). Vitrified 2-cell embryos showed the highest survival rate in vivo (50.6%, 88/174), compared with that from other stages (9.3–30.5%, P < 0.05). The data demonstrate that the OPS method is suitable for the cryopreservation of IVF mouse embryos from 2-cell stage to early blastocyst stage without a significant loss of survival. Embryos at the 2-cell stage had the best tolerance for cryopreservation in the present study.

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