Development of single mouse blastomeres into blastocysts, outgrowths and the establishment of embryonic stem cells.
The recently developed technique of establishing embryonic stem (ES) cell lines from single blastomeres (BTMs) of early mouse and human embryos has created significant interest in this source of ES cells. However, sister BTMs of an early embryo might not have equal competence for the development of different lineages or the derivation of ES cells. Therefore, single BTMs from two- and four-cell embryos of outbred mice were individually placed in sequential cultures to enhance the formation of the inner cell mass (ICM) and the establishment of embryonic outgrowth. The outgrowths were then used for the derivation of ES cell lines. Based on the expression of ICM (Sox2) and trophectoderm (Cdx2) markers, it was determined that ICM marker was lacking in blastocysts derived from 12% of BTMs from two-cell stage and 20% from four-cell stage. Four ES cell lines (5.6%; 4/72) were established ater culture of single BTMs from two-cell embryos, and their pluripotency was demonstrated by their differentiation into neuronal cell types. Our results demonstrate that sister BTMs of an early embryo are not equally competent for ICM marker expression. However, we demonstrated the feasibility of establishing ES cells from a single BTM of outbred mice.
Lorthongpanich
Chanchao
C
Yerkes National Primate Research Center, Department of Human Genetics and Genetics and Molecular Biology Program, Emory University School of Medicine, North East Atlanta, Georgia 30329, USA.
Yang
Shang-Hsun
SH
Piotrowska-Nitsche
Karolina
K
Parnpai
Rangsun
R
Chan
Anthony W S
AW
D000818:Animals; D015415:Biomarkers / Q000032:analysis; D001755:Blastocyst / Q000166:cytology*; D053624:Blastocyst Inner Cell Mass / Q000166:cytology; D001757:Blastomeres / Q000166:cytology*; D000071616:CDX2 Transcription Factor; D002452:Cell Count; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002478:Cells, Cultured; D002970:Cleavage Stage, Ovum; D053595:Embryonic Stem Cells / Q000166:cytology*; D005260:Female; D018398:Homeodomain Proteins / Q000032:analysis; D007150:Immunohistochemistry; D051379:Mice; D055748:SOXB1 Transcription Factors / Q000032:analysis; D014746:Vimentin / Q000032:analysis
D015415:Biomarkers; D000071616:CDX2 Transcription Factor; C491270:CDX2 protein, human; D018398:Homeodomain Proteins; D055748:SOXB1 Transcription Factors; C527783:Sox2 protein, mouse; D014746:Vimentin