scholarly journals Differences in embryonic development in sensitive and resistant matings to pregnancy block stimuli in mice

Reproduction ◽  
2003 ◽  
pp. 327-335 ◽  
Author(s):  
HJ Chung ◽  
JK Pak ◽  
BK Kim ◽  
YK Lee ◽  
SK Im ◽  
...  
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Normal Structure ◽  
Histological Observation ◽  
Primitive Endoderm ◽  
Male Pheromone ◽  
Male And Female ◽  
Inner Surface ◽  
The Difference ◽  
Ectoplacental Cone

Pregnancy block from exposure to foreign male mouse pheromones is sensitive to both male and female mating strain, as well as the foreign male pheromone-producing strain. Incidence of pregnancy block by male pheromones in mice is different depending on the combination of females, stud males and stimulus males. BALB/cA females mated with BALB/cA males showed a 100% pregnancy block when exposed to males of the DDK strain (Chung et al., 1997). In contrast, BALB/cA females mated with males of dissimilar strain show high rates of pregnancy even if they are exposed to DDK males; this difference is thought to be due to the difference in viability of embryos (Chung et al., 1999). The present study investigated how development of BALB/cA and F1 embryos differ under the influence of pregnancy block stimuli. F1 embryos had significantly higher numbers of cells than did the BALB/cA embryos (P<0.05) at day 3 of pregnancy after exposure to DDK males or after bromocriptine (dopamine agonist, 4 mg kg(-1), i.p.) treatment. Histological observation after bromocriptine treatment revealed that: (i) on day 4 of pregnancy, BALB/cA embryos tended to form a large blastocoel, but showed abnormalities such as degeneration of primitive endoderm and depression of the outer trophoblast-distal endoderm layer at the periphery of the inner cell mass (ICM) or detachment of the ICM from the outer layer. In contrast, 60-70% of F1 embryos were normal late blastocysts and incipient egg cylinders, but 28-40% of early blastocysts were degenerating; and (ii) day 5 BALB/cA embryos were in the range from incipient egg cylinder with a large proamniotic cavity to ectoplacental cone only, but their proximal endoderm and trophoblast-distal endoderm layer were degenerating. In contrast, the F1 embryos were mostly at the egg cylinder stage and maintained normal structure except for occasional enlargement of the developing yolk sac cavity. These results indicate that the lining of the inner surface of trophoblast by distal endoderm layer may be more firmly established and that the inner environment for development of F1 embryos may be more effectively maintained, thereby making them more resistant to deleterious influences due to pregnancy block stimuli than are BALB/cA embryos.

scholarly journals Totipotency of mouse zygotes extends to single blastomeres of embryos at the four-cell stage

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Marino Maemura ◽  
Hiroaki Taketsuru ◽  
Yuki Nakajima ◽  
Ruiqi Shao ◽  
Ayaka Kakihara ◽  
...  
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Mouse Embryos ◽  
Primitive Endoderm ◽  
Cell Stage ◽  
Supportive Environment ◽  
Mouse Zygotes ◽  
Single Blastomeres ◽  
Multicellular Organisms

AbstractIn multicellular organisms, oocytes and sperm undergo fusion during fertilization and the resulting zygote gives rise to a new individual. The ability of zygotes to produce a fully formed individual from a single cell when placed in a supportive environment is known as totipotency. Given that totipotent cells are the source of all multicellular organisms, a better understanding of totipotency may have a wide-ranging impact on biology. The precise delineation of totipotent cells in mammals has remained elusive, however, although zygotes and single blastomeres of embryos at the two-cell stage have been thought to be the only totipotent cells in mice. We now show that a single blastomere of two- or four-cell mouse embryos can give rise to a fertile adult when placed in a uterus, even though blastomere isolation disturbs the transcriptome of derived embryos. Single blastomeres isolated from embryos at the eight-cell or morula stages and cultured in vitro manifested pronounced defects in the formation of epiblast and primitive endoderm by the inner cell mass and in the development of blastocysts, respectively. Our results thus indicate that totipotency of mouse zygotes extends to single blastomeres of embryos at the four-cell stage.

scholarly journals Interleukin-6 promotes primitive endoderm development in bovine blastocysts

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Lydia K. Wooldridge ◽  
Alan D. Ealy
Keyword(s):  
Interleukin 6 ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Types ◽  
Janus Kinase ◽  
Primitive Endoderm ◽  
Cell Numbers ◽  
Downstream Target ◽  
Dependent Signal ◽  
Endoderm Development

Abstract Background Interleukin-6 (IL6) was recently identified as an embryotrophic factor in bovine embryos, where it acts primarily to mediate inner cell mass (ICM) size. This work explored whether IL6 affects epiblast (EPI) and primitive endoderm (PE) development, the two embryonic lineages generated from the ICM after its formation. Nuclear markers for EPI (NANOG) and PE (GATA6) were used to differentiate the two cell types. Results Increases (P < 0.05) in total ICM cell numbers and PE cell numbers were detected in bovine blastocysts at day 8 and 9 post-fertilization after exposure to 100 ng/ml recombinant bovine IL6. Also, IL6 increased (P < 0.05) the number of undifferentiated ICM cells (cells containing both PE and EPI markers). The effects of IL6 on EPI cell numbers were inconsistent. Studies were also completed to explore the importance of Janus kinase 2 (JAK2)-dependent signaling in bovine PE cells. Definitive activation of STAT3, a downstream target for JAK2, was observed in PE cells. Also, pharmacological inhibition of JAK2 decreased (P < 0.05) PE cell numbers. Conclusions To conclude, IL6 manipulates ICM development after EPI/PE cell fates are established. The PE cells are the target for IL6, where a JAK-dependent signal is used to regulate PE numbers.

Interactions of blastomeres suggest changes in cell surface adhesiveness during the formation of inner cell mass and trophectoderm in the preimplantation mouse embryo

Development ◽  
1982 ◽  
Vol 70 (1) ◽  
pp. 133-152
Author(s):  
Susan J. Kimber ◽  
M. Azim ◽  
H. Surani ◽  
Sheila C. Barton
Keyword(s):  
Inner Cell Mass ◽  
Single Cells ◽  
Cell Mass ◽  
Cell Stage ◽  
Trophectoderm Cells ◽  
Adhesive Surface ◽  
Preimplantation Mouse Embryo ◽  
Divergent Differentiation ◽  
Preimplantation Mouse ◽  
The Difference

Whole 8-cell morulae can be aggregated with isolated inner cell masses from blastocysts. On examining semithin light microscope sections of such aggregates we found that cells of the morula changed shape and spread over the surface of the ICM, thus translocating it to the inside of the aggregate. Using single cells from 8-cell embryos in combination with single cells from other stage embryos or isolated ICMs we show that 1/8 blastomeres spread over other cells providing a suitably adhesive surface. The incidence of spreading is high with inner cells from 16-cell embryos (56 %) and 32-cell embryos (62%) and isolated inner cell masses (64%). In contrast, the incidence of spreading of 1/8 blastomeres is low over outer cells from 16-cell embryos (26%) and 32-cell embryos (13%). Blastomeres from 8-cell embryos do not spread over unfertilized 1-cell eggs, 1/2 or 1/4 cells or trophectoderm cells contaminating isolated ICMs. When 1/8 cells are aggregated in pairs they flatten on one another (equal spreading) as occurs at compaction in whole 8-cell embryos. However, if 1/8 is allowed to divide to 2/16 in culture one of the cells engulfs the other (51-62/ pairs). Based on the ideas of Holtfreter (1943) and Steinberg (1964,1978) these results are interpreted to indicate an increase in adhesiveness at the 8-cell stage as well as cytoskeletal mobilization. Following the 8-cell stage there is an increase in adhesiveness of inside cells while the outside cells decrease in adhesiveness. The difference in adhesiveness between inside and outside cells in late morulae is probably central to the divergent differentiation of (inner) ICM and (outer) trophectoderm cell populations.

scholarly journals Cell lineage allocation in equine blastocysts produced in vitro under varying glucose concentrations

Reproduction ◽  
2015 ◽  
Vol 150 (1) ◽  
pp. 31-41 ◽  
Author(s):  
Young-Ho Choi ◽  
Pablo Ross ◽  
Isabel C Velez ◽  
B Macías-García ◽  
Fernando L Riera ◽  
...  
Keyword(s):  
High Glucose ◽  
Culture Medium ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Lineage ◽  
Primitive Endoderm ◽  
Blastocyst Development ◽  
Embryo Culture Medium ◽  
Effect Of Glucose

Equine embryos developin vitroin the presence of high glucose concentrations, but little is known about their requirements for development. We evaluated the effect of glucose concentrations in medium on blastocyst development after ICSI. In experiment 1, there were no significant differences in rates of blastocyst formation among embryos cultured in our standard medium (DMEM/F-12), which contained >16 mM glucose, and those cultured in a minimal-glucose embryo culture medium (<1 mM; Global medium, GB), with either 0 added glucose for the first 5 days, then 20 mM (0-20) or 20 mM for the entire culture period (20-20). In experiment 2, there were no significant differences in the rates of blastocyst development (31–46%) for embryos cultured in four glucose treatments in GB (0-10, 0-20, 5-10, or 5-20). Blastocysts were evaluated by immunofluorescence for lineage-specific markers. All cells stained positively forPOU5F1. An inner cluster of cells was identified that included presumptive primitive endoderm cells (GATA6-positive) and presumptive epiblast (EPI) cells. The 5-20 treatment resulted in a significantly lower number of presumptive EPI-lineage cells than the 0-20 treatment did.GATA6-positive cells appeared to be allocated to the primitive endoderm independent of the formation of an inner cell mass, as was previously hypothesized for equine embryos. These data demonstrate that equine blastocyst development is not dependent on high glucose concentrations during early culture; rather, environmental glucose may affect cell allocation. They also present the first analysis of cell lineage allocation inin vitro-fertilized equine blastocysts. These findings expand our understanding of the factors that affect embryo development in the horse.

Investigation of cell lineage and differentiation in the extraembryonic endoderm of the mouse embryo

Development ◽  
1982 ◽  
Vol 68 (1) ◽  
pp. 175-198
Author(s):  
R. L. Gardner
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Lineage ◽  
Early Embryo ◽  
Visceral Endoderm ◽  
Primitive Endoderm ◽  
Developmental Potential ◽  
Extraembryonic Endoderm ◽  
Parietal Endoderm ◽  
Endodermal Cells

The technique of injecting genetically labelled cells into blastocysts was used in an attempt to determine whether the parietal and visceral endoderm originate from the same or different cell populations in the early embryo. When the developmental potential of 5th day primitive ectoderm and primitive endoderm cells was compared thus, only the latter were found to colonize the extraembryonic endoderm. Furthermore, single primitive endoderm cells yielded unequivocal colonization of both the parietal and the visceral endoderm in a proportion of chimaeras. However, in the majority of primitive endodermal chimaeras, donor cells were detected in the parietal endoderm only, cases of exclusively visceral colonization being rare. Visceral endoderm cells from 6th and 7th day post-implantation embryos also exhibited a striking tendency to contribute exclusively to the parietal endoderm following blastocyst injection. The above findings lend no support to a recent proposal that parietal and visceral endoderm are derived from different populations of inner cell mass cells. Rather, they suggest that the two extraembryonic endoderm layers originate from a common pool of primitive endoderm cells whose direction of differentiation depends on their interactions with non-endodermal cells.

2 BOVINE EMBRYONIC STEM-LIKE CELLS DERIVED FROM IN VITRO-PRODUCED BLASTOCYSTS

2017 ◽  
Vol 29 (1) ◽  
pp. 108
Author(s):  
Y. S. Bogliotti ◽  
J. Wu ◽  
M. Vilariño ◽  
K. Suzuki ◽  
J. C. Belmonte ◽  
...  
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Embryonic Stem ◽  
Basal Medium ◽  
Primitive Endoderm ◽  
Culture Dish ◽  
Rock Inhibitor ◽  
Immunodeficient Mice

Embryonic stem cells (ESC) are derived from the inner cell mass (ICM) of preimplantation blastocysts. To date, it has been challenging to establish pluripotent ESC lines for domestic animals, which could be important for biotechnological applications, such as genetic engineering and SCNT, and biomedical research. The aim of this work was to derive and characterise bovine embryonic stem-like cells (bESC) from in vitro-produced bovine blastocysts. Embryos were produced by in vitro fertilization of in vitro-matured oocytes aspirated from abattoir ovaries and cultured in groups of 25 in 50-μL drops of KSOM (Evolve, Zenith Biotech) with 4 mg mL−1 BSA for 7 days until they reached the blastocyst stage (Ross et al., 2009 Reproduction 137, 427–437). At that point, the zona pellucida (ZP) was removed using 1 mg mL−1 Pronase (Sigma, St. Louis, MO), and ZP-free blastocysts were washed 6 times in SOF-HEPES. Three derivation approaches were tested: ZP-free whole blastocysts, mechanically isolated ICM, and immunosurgery-derived ICM. In each case, individual blastocysts/ICM were placed in 1 well of a 12-well dish seeded with a monolayer of mouse embryo fibroblasts (MEF) and cultured in mTeSR1 basal medium (without growth factors) supplemented with 20 ng mL−1 FGF2 and 2.5 μM IWR1 (CTFR) (Wu et al. 2015 Nature 521, 316–321). After 48 h, blastocysts/ICM that failed to adhere were physically pressed against the bottom of the culture dish with a 22-gauge needle under a stereoscope to aid attachment. Thereafter, the media was changed daily. Outgrowths (after 6–7 days in culture) were dissociated and passaged using TrypLE and re-seeded in the presence of ROCK inhibitor (Y-27632, 10 μM) onto newly prepared wells containing MEF. Established bESC lines were cultured on MEF and passaged every 4 to 5 days at a 1:10 split ratio. The bESC lines were characterised by immunofluorescence (IF), RNA-seq, and teratoma formation. The efficiency of cell line derivation (evaluated at passage 3) was similar for the 3 approaches: whole blastocysts (9/16, 56.3%), mechanical ICM isolation (7/12, 58.3%), and immunosurgical ICM isolation (7/16, 43.8%). The bESC were passaged and cultured long-term (more than 15 passages) and were subjected to several rounds of freezing and thawing while retaining their morphology and characteristics. IF analysis showed that long-term cultured bESC expressed the markers SOX2 and OCT4 (pluripotency), but did not express CDX2 (trophectoderm) or GATA6 (primitive endoderm). RNAseq analysis of 2 bESC lines showed that ICM markers (POU5F1, NANOG, SOX2, LIN28B, DNAMT3B, UTF1, SALL4) were expressed (RPKM > 0.4), while trophectoderm markers (CDX2, GATA2, GATA3, FGF4, TFAP2A) and primitive endoderm markers (GATA6, HNF4A) were not expressed (RPKM < 0.4). Finally, bESC lines (n = 2) were able to form teratomas in immunodeficient mice. The teratomas contained tissues representative of the 3 germ lineages and expressed lineage-specific markers (ectoderm: TUJ1, endoderm: FOXA2, and mesoderm: ASM). In conclusion, the culture condition used in this work (CTFR) enables robust derivation and long-term in vitro propagation of pluripotent bESC.

scholarly journals ECM-integrin signalling instructs cellular position-sensing to pattern the early mouse embryo

Development ◽  
2021 ◽  
Author(s):  
Esther Jeong Yoon Kim ◽  
Lydia Sorokin ◽  
Takashi Hiiragi
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Spatial Arrangement ◽  
Cell Types ◽  
Embryonic Cells ◽  
Primitive Endoderm ◽  
Integrin Β1 ◽  
Early Mouse Embryo ◽  
Position Sensing ◽  
Early Mouse

Development entails patterned emergence of diverse cell types within the embryo. In mammals, cells positioned inside the embryo give rise to the inner cell mass (ICM) that eventually forms the embryo proper. Yet the molecular basis of how these cells recognise their ‘inside’ position to instruct their fate is unknown. Here we show that provision of extracellular matrix (ECM) to isolated embryonic cells induces ICM specification and alters subsequent spatial arrangement between epiblast (EPI) and primitive endoderm (PrE) cells that emerge within the ICM. Notably, this effect is dependent on integrin β1 activity and involves apical to basal conversion of cell polarity. We demonstrate that ECM-integrin activity is sufficient for ‘inside’ positional signalling and it is required for proper EPI/PrE patterning. Our findings thus highlight the significance of ECM-integrin adhesion in enabling position-sensing by cells to achieve tissue patterning.

scholarly journals Leukemia Inhibitory Factor Stimulates Primitive Endoderm Expansion in the Bovine Inner Cell Mass

2021 ◽  
Vol 2 ◽  
Author(s):  
Lydia K. Wooldridge ◽  
Alan D. Ealy
Keyword(s):  
Leukemia Inhibitory Factor ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Number ◽  
Inhibitory Factor ◽  
Primitive Endoderm ◽  
Total Cell Number ◽  
Cell Numbers ◽  
Bovine Blastocyst

Previous work determined that bovine interleukin-6 (IL6) increases inner cell mass (ICM), primitive endoderm (PE), and total cell number in in vitro produced (IVP) bovine blastocysts. Another IL6 family member, leukemia inhibitory factor (LIF), has the potential to produce the same effects of IL6 due to the presence of its receptor in bovine blastocysts. We compared the abilities of LIF and IL6 to increase ICM cell numbers in day 7, 8, and 9 IVP bovine blastocysts. Supplementation with 100 ng/ml LIF from day 5 onward improved blastocyst formation rates on days 7 and 8 similar to what was observed when supplementing 100 ng/ml IL6. However, LIF supplementation did not cause an increase in ICM numbers like was observed after supplementing IL6. On day 9, increases in PE cell numbers were detected after LIF supplementation, but 300 ng/ml LIF was required to achieve the same effect on PE numbers that was observed by providing 100 ng/ml IL6. Collectively, these results show that LIF can mimic at least some of the effects of IL6 in bovine blastocyst.

An investigation of inner cell mass and trophoblast tissues following their isolation from the mouse blastocyst

Development ◽  
1972 ◽  
Vol 28 (2) ◽  
pp. 279-312
Author(s):  
R. L. Gardner ◽  
M. H. Johnson
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Mouse Blastocyst ◽  
Or Groups ◽  
Ectoplacental Cone

1. Inner cell mass (ICM) and trophoblast were isolated from 3½-day post-coitum mouse blastocysts by microsurgery. 2. Trophoblastic fragments formed vesicles in culture but did not aggregate with other such fragments. They proved as effective as intact blastocysts in inducing decidua in recipient uteri, but thereafter failed to proliferate. 3. Isolated ICMs remained as solid balls of cells that readily aggregated in pairs or groups in culture but failed to induce implantation changes in receptive uteri. 4. Possible explanations for the failure of isolated trophoblast to proliferate after implantation are discussed. It is argued that presence of ICM tissue is necessary for trophoblast proliferation, and suggested that the ICM exerts its effect by controlling development of the ectoplacental cone.

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.

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