A Euploid Line of Human Embryonic Stem Cells Derived from a 43,XX,dup(9q),+12,-14,- 15,-18,-21 Embryo
Aparecida Siqueira Fonseca S., Montero Costas R., Morato-Marques M., Costa S., Roberto Alegretti J., Rosenberg C., Leme E., da Motta A., Serafini P.C., Pereira L.V.
PLoS One, 2015
Aneuploid embryos (Array-CGH analysis) cultured until day-5 or 6 after in vitro fertilization. To derive new hESC lines under defined xeno-free culture condition, the authors used the CloneStem kit that contains recombinant laminin-521 and E-cadherin as matrix and E8 medium supplemented with 10% human albumin serum for 48h. The first passage was made mechanically after 15 days and the fragments were transferred to laminin-521 and e-cadherin, in E8 medium supplemented with 5 uM of Rock Inhibitor. The next three passages were made mechanically and the fragments were transferred to plates coated only with Laminin-521 in E8 medium, in a split ratio of 1:2. After passage five, the hESCs were maintained in Geltrex and E8 medium. Analyzed pluripotency, G-banding karyotype analysis, SNP genotyping, differentiation capacity (EB). The authors show that, despite the complex chromosomal abnormality, the corresponding hESC line BR-6 is euploid (46,XX). Single nucleotide polymorphism analysis showed that the embryo ́s missing chromosomes were not duplicated in BR-6, suggesting the existence of extensive mosaicism in the trophectoderm lineage. Two embryos out of ten embryos attached to the culture plate and presented cell growth. From these only one embryo gave rise to a new line of hESC. This rate of derivation is relatively low and is similar to those obtained on Matrigel with euploid embryos.
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