Analysis of long-term culture properties and pluripotent character of two sibling human embryonic stem cell lines derived from discarded embryos

Abstract

We had earlier reported the derivation and characterization of two new sibling human embryonic stem cell lines BJNhem19 and BJNhem20, from discarded grade III embryos of Indian origin. We report here the characteristics of the two sibling cell lines after long-term continuous culture for over 2 yr during which they have been passaged over 200 times. We show that both cell lines adapt well to culture on various mouse and human feeders as well as in feeder-free conditions. The cells show normal diploid karyotype and continue to express all pluripotency markers. Both cell lines differentiate to derivatives of all three germ layers in vitro. However as reported earlier, BJNhem19 is unable to generate teratomas in nude or SCID mice or differentiate to beating cardiomyocytes when tested over several passages during long-term stable culture. On the other hand, the cardiac differentiation capacity of BJNhem20 is greatly increased, and it can generate beating cardiomyocytes that proliferate when isolated and cultured further. In conclusion, the two cell lines have maintained a stable phenotype for over 2 yr and are indeed immortal. Their derivation from grade III embryos does not seem to have any adverse effect on their long-term phenotype. The cells can be obtained for research purposes from the UK Stem Cell Bank and from the authors.

The Influence of Early Embryo Traits on Human Embryonic Stem Cell Derivation Efficiency

“…These poor-quality embryos (PQEs) are typically discarded following treatment or donated for research after patient consent. Despite impaired implantation and developmental potential, PQEs have been shown to be a viable source for the derivation of new human embryonic stem cell (hESC) lines [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16]. It is estimated that hundreds of thousands of embryos per year are discarded because of poor quality and the majority of infertility patients are in favor of donating them for stem cell research [7,17,18].…”

Heterogeneity in Human Embryonic Stem Cells May Prevent Endodermal Guided Differentiation

“…In contrast, culture in a commercially available BEGM (with tri-iodothyronin and retinoic acid but without BSA) promoted differentiation to predominantly bronchiolar alveolar cell (i.e., Clara cell) phenotype. Lung lineage-specific cell differentiation was achieved in a relatively shorter span of time (22 days) in contrast to other reported lung lineage culture conditions [22][23][24][25][26][27][28][29][30]. These culture media, normally used to maintain and grow mature cells, could successfully induce differentiation of pluripotent embryonic stem cells into three types of mature lung lineage-specific non-ciliated cells.…”

“…These cells were derived from the inner cell mass (ICM) of grade III poor quality blastocysts that were not suitable for in vitro fertility treatment. Both the lines as claimed by JNCASR are pluripotent and have been extensively characterized and cultured continuously for over 250 passages [28][29][30][31][32][33][34][35].…”

“…Previous work [22][23][24][25][26][27][28][29][30] has demonstrated differentiation steps to AEII cells from murine ES cells and the hES H7 cell line.…”

Evaluating differentiation propensity of in-house derived human embryonic stem cell lines KIND-1 and KIND-2

“…Two studies on hES cell lines derived from the Indian subcontinent have also described the differentiation bias existing amongst cell lines. Of the two cell lines derived by Inamdar and group (2009), BJNhem20 has increased tendency towards cardiac differentiation whereas BJNhem19 lacks this ability (Venu et al 2010). Mehta et al (2010) have shown a clear trend for RelicellhES1 to differentiate into neuronal lineage.…”