Effect of culture conditions on diploid to giant-cell transformation in postimplantation mouse trophoblast

Development ◽  
1981 ◽  
Vol 62 (1) ◽  
pp. 217-227
Author(s):  
J. Rossant ◽  
W. Tamura-Lis
Keyword(s):  
Giant Cell ◽  
Cell Transformation ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Formation ◽  
Culture Conditions ◽  
Embryonic Cells ◽  
Giant Cell Formation ◽  
Ectoplacental Cone

Diploid extraembryonic ectoderm and ectoplacental cone from the 7·5-day mouse embryo were grown in vitro under a variety of culture conditions in an attempt to discover conditions which maintain trophoblast in a diploid state and prevent giant-cell formation. It was found that maintenance of tissue integrity was not enough to keep the tissues dividing and diploid, but that the presence of inner-cell-mass derivatives did have some effect. This effect was only apparent when trophoblast cells were entirely enclosed by embryonic tissues. Monolayers of embryonic or embryonal carcinoma cells did not prevent giant-cell formation. Diploid extraembryonic ectoderm and ectoplacental cone responded differently: ectoplacental cells eventually formed trophoblast giant cells even when enclosed by embryonic cells whereas extraembryonic ectoderm cells apparently could be maintained in a diploid condition. This and other differences in properties between extraembryonic ectoderm and ectoplacental cone are discussed with reference to a new model for the postimplantation trophoblast lineage in the mouse.

The development of trophoblast in vitro from blastocysts containing varying amounts of inner cell mass

Development ◽  
1975 ◽  
Vol 33 (1) ◽  
pp. 177-185
Author(s):  
J. D. Ansell ◽  
M. H. L. Snow
Keyword(s):  
Giant Cell ◽  
Cell Transformation ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Calf Serum ◽  
Foetal Calf Serum ◽  
Trophoblast Cells ◽  
Intact Mouse

When intact mouse blastocysts are cultured in vitro in medium supplemented with foetal calf serum, trophoblast cells proliferate and undergo giant cell transformation such as occurs in vivo. If the amount of inner cell mass in the blastocyst is decreased by culture with [3H]-thymidine then giant cell transformation occurs normally but proliferation is reduced. In the absence of inner cell mass no proliferation occurs, and giant cell transformation is more rapid than in undamaged blastocysts.

Berberine alleviates LPS-induced apoptosis, oxidation, and skewed lineages during mouse preimplantation development

2022 ◽  
Author(s):  
Xiaosu Miao ◽  
Wei Cui
Keyword(s):  
Inner Cell Mass ◽  
Polycystic Ovary ◽  
Cell Mass ◽  
Cell Lineage ◽  
Culture Conditions ◽  
Preimplantation Development ◽  
Embryonic Cells ◽  
Preimplantation Embryo Development ◽  
New Treatment ◽  
Induced Apoptosis

Abstract Female infertility is a heterogeneous disorder with a variety of complex causes, including inflammation and oxidative stress, which are also closely associated with the pathogenesis of Polycystic Ovary Syndrome (PCOS). As a new treatment for PCOS, berberine (BER), a natural compound from Berberis, has been clinically applied recently. However, the mechanisms underlying the association between BER and embryogenesis are still largely unknown. In this study, effects of BER on preimplantation development was evaluated by using both normal and inflammatory culture conditions induced by lipopolysaccharide (LPS) in the mouse. Our data first suggest that BER itself (25 nM) does not affect embryo quality or future developmental potency, moreover, it can effectively alleviate LPS-induced embryonic damage by mitigating apoptosis via ROS−/caspase-3-dependent pathways and by suppressing pro-inflammatory cytokines via inhibition of NF-κB signaling pathway during preimplantation embryo development. In addition, skewed cell lineage specification in inner cell mass (ICM) and primitive endoderm (PE) caused by LPS can also be successfully rescued with BER. In summary, these findings for the first time demonstrate the non-toxicity of low doses of BER and its anti-apoptotic and anti-oxidative properties on embryonic cells during mammalian preimplantation development.

53 EFFECTS OF INSULIN-TRANSFERRIN-SELENIUM IN DEFINED AND PORCINE FOLLICULAR FLUID-SUPPLEMENTED MEDIUM ON IN VITRO PRODUCTION OF PORCINE SCNT EMBRYOS

2007 ◽  
Vol 19 (1) ◽  
pp. 144
Author(s):  
Y. U. Kim ◽  
D. P. Bhandari ◽  
M. S. Hossein ◽  
S. M. Park ◽  
E. Lee ◽  
...  
Keyword(s):  
Somatic Cell ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Defined Medium ◽  
Culture Conditions ◽  
Developmental Competence ◽  
Differential Staining ◽  
In Vitro Production ◽  
Fetal Fibroblasts

Insulin promotes the uptake of glucose and amino acids, and is beneficial for maturation of oocytes in vitro. Transferrin is an iron-transport protein and selenium is an essential trace element. Insulin-transferrin-selenium (ITS) together has been used in some in vitro maturation systems. The present study was designed to evaluate the effects of ITS in defined and porcine folicular fluid (pFF)-supplemented IVM medium on the glutathione (GSH) concentration, and on developmental competence after somatic cell nuclear transfer. ITS liquid media supplement (I-3146) was purchased from Sigma-Aldrich (St Louis, MO, USA). Basic IVM medium was TCM-199 supplemented with 10 ng mL-1 epidermal growth factor, 4 IU mL-1 pregnant mare serum gonadotropin (PMSG) and hCG and either 1% PVA (defined medium) or 10% pFF. Ten �g mL-1 insulin, 5.5 �g mL-1 transferrin, and 5 �g mL-1 selenium was used for the entire 44-h culture period. The GSH content of a gruop of 10 to 20 oocytes was determined by the dithionitrobezoic acid-glutathione disulfide (DTNB-GSSG) reductase recycling assay. Fetal fibroblasts were used as somatic cell donors and reconstructed embryos were cultured in mNCSU-23 medium for 168 h. Cleavage and blastocyst formation was observed at 48 h and 168 h, respectively. The quality of blastocysts was assessed by differential staining of the inner cell mass (ICM) and the trophectoderm (TE) cells. Each experiment was replicated for 5 times. The data were analyzed by one-way ANOVA, and Tukey was used as a posthoc test. The level of GSH production significantly varied in different culture conditions. The highest GSH concentration was observed in the pFF + ITS group (8.2 picomol/oocyte). A total of 116, 125, 126, and 120 reconstructed oocytes were cultured, and 10.1, 15.3, 17.2, and 21.8% blastocysts were observed for PVA, PVA + ITS, pFF, and pFF + ITS groups, respectively (P < 0.05). The numbers of inner cell mass, trophrectoderm cells, and total cells were significantly higher in the pFF + ITS group compared with the other groups. The average number of total cells in blastocysts was 31.9 � 1.8, 43.1 � 3.5, 46.7 � 4.9, and 52.3 � 6.7 for PVA, PVA + ITS, pFF, and pFF + ITS groups, respectively (P < 0.05). ITS supplement improved the developmental competence in both the defined and the pFF supplemented groups. We recommend supplementing porcine IVM medium with 10 �g mL-1 insulin, 5.5 �g mL-1 transferrin, and 5 �g mL-1 selenium.

58 ISOLATION OF BOVINE TROPHOBLAST AND ITS REPROGRAMMING BY NUCLEAR TRANSFER

2011 ◽  
Vol 23 (1) ◽  
pp. 134
Author(s):  
I. M. Saadeldin ◽  
B. H. Kim ◽  
B. Roibas da Torre ◽  
O. J. Koo ◽  
G. Jang ◽  
...  
Keyword(s):  
Nuclear Transfer ◽  
Inner Cell Mass ◽  
Quantitative Polymerase Chain Reaction ◽  
Cell Mass ◽  
Donor Cell ◽  
Developmental Competence ◽  
Control Group ◽  
Embryonic Cells ◽  
Blastocyst Development

Nuclear transfer (NT) has been used to produce many cloned offspring using several types of cells, including embryonic cells. Even though inner cell mass cells have been used as donor karyoplast for producing cloned animals, there are few studies using trophoblast. In mice, clones were born by nuclear transfer of trophoblasts from the expanded blastocyst into enucleated oocytes as a trial to show the totipotency of both inner cell mass and trophectoderm cells isolated from blastocysts (Tsunoda and Kato 1998 J. Reprod. Fertil. 113, 181–184). However, bovine trophoblast cell (TC) lines have not been used in NT to date. The purpose of this study was to elucidate whether TC as donor cell can be reprogrammed in bovine enucleated oocyte and determine the relative abundance of interferon tau (IFNτ) expression in the resulting cloned preimplantational embryos. Hatched blastocysts produced by IVF were used to isolate TCs on mouse embryonic fibroblasts treated with mitomycin C as feeder cells. TCs and adult fibroblasts (AF, control group for NT) were microinjected to perivitelline space of in vitro mature enucleated oocytes and electrically fused. Reconstructed embryos were chemically activated and cultured in a 2-step chemically defined medium. Levels of IFNτ expression in IVF-, TC-, and AF-derived blastocysts were analyzed using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). IVF produced embryos were used as reference to analyze the linear progressive expression of IFNτ through mid-, expanded, and hatching blastocysts. As a result, TCs expressing IFNτ were successfully isolated and cultured on feeder layers. It grew as cell sheets of cuboidal epithelium with high proliferation capacity as a single colony originated from a small clump of cells measured 0.5 cm within 7 days of culture. TCs were reprogrammed in the enucleated oocytes to blastocyst with similar efficiency to AF (14.5% and 15.6%, respectively; P ≤ 0.05). RT-qPCR studies showed that IFNτ expression was higher in TC-derived blastocysts than IVF- and AF-derived blastocysts. Both IVF- and TC-derived blastocysts, showed progressive increase of IFNτ expression through the advancement of blastocyst development when it was compared to AF-derived blastocysts. In conclusion, using TCs expressing IFNτ as donor cell for bovine NT could increase the developmental competence of cloned embryos as indicated by progressive linear increase in IFNτ expression. This study was supported by grants from IPET (#109023-05-1-CG000), NRF (#M10625030005-10N250300510), MKE (#2009-67-10033839, #2009-67-10033805), and BK21 program. Saadeldin I. M. is supported by Islamic Development Bank (IDB) merit scholarship, Jeddah, Saudi Arabia.

In vivo and in vitro development of mouse embryos homozygous for the embryonic lethal velvet coat (Ve) mutation

Development ◽  
1981 ◽  
Vol 66 (1) ◽  
pp. 43-55
Author(s):  
J. Rossant ◽  
K. M. Vijh
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Giant Cells ◽  
Blastocyst Stage ◽  
Parietal Endoderm ◽  
Trophoblast Giant Cells ◽  
Vitro Development ◽  
Ectoplacental Cone

Embryos homozygous for the velvet coat mutation, Ve/Ve, were recognized at 6·5 days post coitum by the reduced size of the ectodermal portions of the egg cylinder and the loose, columnar nature of the overlying endoderm. Later in development ectoderm tissues were sometimes entirely absent. Abnormalities appeared in the ectoplacental cone at 8·5 days but trophoblast giant cells and parietal endoderm appeared unaffected. Homozygotes could not be unequivocally identified at 5·5 days nor at the blastocyst stage but were recognized in blastocyst outgrowths by poor development of the inner cell mass derivatives, It has previously been suggested that Ve may exert its action at the blastocyst stage by reducing the size of the inner cell mass, but no evidence for such a reduction was found. Most of the observations on Ve/Ve homozygotes are, however, consistent with the hypothesis that Ve exerts its action primarily on later primitive ectoderm development.

scholarly journals Development of Preimplantation Mouse Embryos in vivo and in vitro

1982 ◽  
Vol 35 (2) ◽  
pp. 187 ◽  
Author(s):  
GM Harlow ◽  
P Quinn
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Culture Conditions ◽  
Cell Stage ◽  
Preimplantation Mouse ◽  
Development In Vitro

The culture conditions for the development in vitro of (C57BL/6 X CBA) F2 hybrid two-cell embryos to the blastocyst stage have been optimized. Commercially available pre-sterile disposable plastic culture dishes supported more reliable development than re-usable washed glass tubes. The presence of an oil layer reduced the variability in development. An average of 85 % of blastocysts developed from hybrid two-cell embryos cultured in drops of Whitten's medium under oil in plastic culture dishes in an atmosphere of 5% O2 : 5% CO2 : 90% N2 ? The time taken for the total cell number to double in embryos developing in vivo was 10 h, and in cultured embryos 17 h. Embryos cultured in vitro from the two-cell stage to blastocyst stage were retarded by 18-24 h in comparison with those remaining in vivo. Day-4 blastocysts in vivo contained 25-70 cells (mean 50) with 7-28 (mean 16) of these in the inner cell mass. Cultured blastocysts contained 19-73 cells (mean 44) with 8-34 (mean 19) of these in the inner cell mass. In the uterine environment, inner-cell-mass blastomeres divided at a faster rate than trophectoderm blastomeres and it is suggested that a long cell cycle is associated with terminal differentiation. Although cultured blastocysts and inner cell masses contained the same number of cells as blastocysts and inner cell masses in vivo, the rate of cell division in cultured inner cell masses was markedly reduced.

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