An improved culture medium for mouse blastocysts

Spindle, Akiko

doi:10.1007/bf02619196

Cited by 144 publications

(61 citation statements)

References 16 publications

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“…Time post-hCG was used to measure the developmental age of the embryos which were collected at 18 hr; unfertilized oocytes; 48 hr, 2-cells; 60 hr, 4-cells; 65-68 hr, 8-cells; 80-85 hr, morulae and 90 hr, blastocysts. Days 1 to 3 embryos (zygotes through 8-cell compacted) were flushed from the reproductive tract using flushing medium I while day 4 embryos were flushed with flushing medium II (Spindle, 1980). Flushing medium I consists of calcium lactate (1.71 mM), sodium pyruvate (0.25mM) and bovine serum albumin (3 mg/ml) added to 10X Leibovitz-modified Hank's balanced salt solution (HBSS) and diluted with water to 1 X (Spindle, 1980).…”

Section: Superovulation and Mouse Embryo Collectionmentioning

confidence: 99%

“…-valine, and Lglutamine at 0.1, 0.5, 1.03, 0.2, 1.0, 2.0, 0.25, 0.5, 2.0, 0.1, 0.1,1.0, and 2.0 mM respectively) and 1 X BME vitamins to 10X Leibovitz -modified HBSS diluted with water to 1 X (Spindle, 1980). Embryos were washed 4-5 times in flushing medium and quick-frozen in a small amount of medium using liquid nitrogen, in batches of 30-50 embryos.…”

Section: Superovulation and Mouse Embryo Collectionmentioning

confidence: 99%

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mRNAs encoding aquaporins are present during murine preimplantation development

et al. 2000

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The present study was conducted to investigate the mechanisms underlying fluid movement across the trophectoderm during blastocyst formation by determining whether aquaporins (AQPs) are expressed during early mammalian development. AQPs belong to a family of major intrinsic membrane proteins and function as molecular water channels that allow water to flow rapidly across plasma membranes in the direction of osmotic gradients. Ten different AQPs have been identified to date. Murine preimplantation stage embryos were flushed from the oviducts and uteri of superovulated CD1 mice. Reverse transcription–polymerase chain reaction (RT‐PCR) methods employing primer sets designed to amplify conserved sequences of AQPs (1–9) were applied to murine embryo cDNA samples. PCR reactions were conducted for up to 40 cycles involving denaturation of DNA hybrids at 95°C, primer annealing at 52–60°C and extension at 72°C. PCR products were separated on 2% agarose gels and were stained with ethidium bromide. AQP PCR product identity was confirmed by sequence analysis. mRNAs encoding AQPs 1, 3, 5, 6, 7, and 9 were detected in murine embryos from the one‐cell stage up to the blastocyst stage. AQP 8 mRNAs were not detected in early cleavage stages but were present in morula and blastocyst stage embryos. The results were confirmed in experimental replicates applied to separate embryo pools of each embryo stage. These results demonstrate that transcripts encoding seven AQP gene products are detectable during murine preimplantation development. These findings predict that AQPs may function as conduits for trophectoderm fluid transport during blastocyst formation. Mol. Reprod. Dev. 57:323–330, 2000. © 2000 Wiley‐Liss, Inc.

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Section: Superovulation and Mouse Embryo Collectionmentioning

confidence: 99%

Section: Superovulation and Mouse Embryo Collectionmentioning

confidence: 99%

mRNAs encoding aquaporins are present during murine preimplantation development

et al. 2000

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“…Dissections were performed in FM-II embryo culture medium [21], a modification of embryonic stem cell media [22]. UDS assays were performed using MEM supplemented with Earles' balanced salts solution, 10% fetal calf serum, and 10 μCi mliter −1 [methyl-3 H]thymidine (80 Ci mmol −1 ; Dupont NEN) as labeling medium.…”

Section: Mediamentioning

confidence: 99%

“…One of the advantages of utilizing a functional assay for NER is that this assay can effectively survey the functioning of all the gene products (approximately [20][21][22][23][24][25][26][27][28][29][30] that comprise damage recognition and repair for this pathway. We have evaluated unpassaged embryonic lineages as well as extraembryonic lineages of the visceral or parietal yolk sacs from midgestational embryos and compared them with normal human and mouse skin fibroblasts.…”

Section: Introductionmentioning

confidence: 99%

Elevated DNA Excision Repair Capacity in the Extraembryonic Mesoderm of the Midgestation Mouse Embryo

Latimer

Hultner

Cleaver

et al. 1996

Experimental Cell Research

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In order to determine whether there is differential cell-type-specific DNA repair we measured the nucleotide excision repair capacity of the four distinct cell lineages that comprise the extraembryonic yolk sac using the unscheduled DNA synthesis assay. Yolk sacs from mouse embryos at 11.5-12.5 days gestation were microdissected to yield purified trophoblast, parietal endoderm, mesoderm, and visceral endoderm, as well as fetal skin fibroblasts which were then grown as primary explants. At this midgestational stage of development, the yolk sac provides essential functions for the sustenance of the embryo while the complex process of organogenesis is proceeding in the liver, kidney, and gut. Trophoblast giant cells, parietal endoderm, and visceral endoderm all demonstrated low levels of unscheduled DNA synthesis consistent with levels measured in adult mouse skin fibroblasts. As has previously been documented, embryonic mouse skin fibroblasts were reproducibly 2-to 3-fold higher than adult mouse skin fibroblasts in levels of DNA excision repair. The extraembryonic mesoderm, however, displayed a statistically significant level of unscheduled DNA synthesis 10-fold higher than adult mouse skin fibroblasts or the other lineages of the midgestation yolk sac. Further, the S-indexes of these lineages were also determined to assess the possible relevance of differential repair to the proliferative status of the cells. These data demonstrate that DNA excision repair capacity is lineage-specific during embryogenesis in the mouse. These studies may begin to provide a context for understanding the perplexing developmental aspects such as the characteristic congenital abnormalities associated with the human heritable DNA repair deficiency diseases.

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“…Parthenogenetic activation was performed by a modification of the procedure described by Cuthbertson (1983). Oocytes were collected into flushing medium I (FM-I) (Spindle, 1980), and cumulus cells were removed by addition of 1 mg/ml hyaluronidase in phosphate-buffered saline followed by washes through nine drops of FM-I. Oocytes were activated by incubation in oil-covered 100 pl drops of embryo culture medium containing 6.2% ethanol (prewarmed to 37°C and equilibrated with 5% COz) for exactly 4.5 min.…”

Section: Experimental Procedures Animals and Embryo Culturementioning

confidence: 99%

Abnormal development of embryonic and extraembryonic cell lineages in parthenogenetic mouse embryos

Sturm

Flannery

Pedersen

1994

Developmental Dynamics

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Parthenogeticallyactivated,diploid mouse oocytes can develop to midgestation stages in utero. However, even these advanced parthenogenones appear to die because of much reduced trophoblast and yolk sac development. Previous studies have compared the general features of parthenogenetic and androgenetic development and determined the fate of uniparental cells in chimeras with normal embryos. These studies led to the concept of genomic imprinting as the cause for developmental failure when either the maternal or the paternal genome is duplicated, with the corresponding deficiency of the other. Genomic imprinting appears to arise during gametogenesis and to act through dosage effects in a set of imprinted genes, whose expression depends on their parental origin. In this study we undertook a more detailed morphological analysis of parthenogenetic development in the mouse and established a classification system to quantify the developmental extent of parthenogenones. We found that the failure of parthenogenones occurred at different times during early postimplantation development, generating a spectrum of concepti which had developed to different extents, with only a small fraction of the embryos reaching advanced somite stages. In all parthenogenones differentiation and proliferation of the trophectoderm and primitive endoderm lineages (both extraembryonic) was abnormal, and in all, even the best-developed parthenogenones, we observed similar deficiencies in the embryonic lineages, especially the mesoderm. Common to all abnormally developed lineages was that the proportion of undifferentiated precursor cells was much reduced, while their differentiated descendants were relatively abundant. We propose, therefore, that the failure of parthenogenones to develop to term is due to abnormal regulation of differentiation and proliferation in both embryonic and extraembryonic lineages. In this hypothesis, the apparent tissue specific defects observed in parthenogenones arise as a consequence of the functional importance of certain tissues (like the trophoblast) early in development. The spectrum of parthenogenones thus appears to reflect critical events in early development, whose regulation are affected by genomic imprinting. o 1994 Wiley-Liss, Inc. 0 1994 WILEY-LISS, INC.

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An improved culture medium for mouse blastocysts

Cited by 144 publications

References 16 publications

mRNAs encoding aquaporins are present during murine preimplantation development

mRNAs encoding aquaporins are present during murine preimplantation development

Elevated DNA Excision Repair Capacity in the Extraembryonic Mesoderm of the Midgestation Mouse Embryo

Abnormal development of embryonic and extraembryonic cell lineages in parthenogenetic mouse embryos

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