OBJECTIVE: To derive new human embryonic stem cell (hESC) lines on pathogen-free human placental fibroblast feeders under serum-free conditions. Because the embryo develops in close contact with extraembryonic membranes, we hypothesized that placental mesenchyme might replicate the stem cell niche in situ. DESIGN: We isolated and characterized human placental fibroblast lines from individual donors and tested their ability to support growth of federally registered hESC lines. Moreover, we performed extensive pathogen testing to ensure their suitability as feeders for the derivation of therapy-grade hESCs. RESULT(S): Human placental fibroblasts were comparable or superior to mouse embryo fibroblasts as hESC feeders. We used these qualified placental fibroblasts to derive two new hESC lines in knockout Dulbecco's modified Eagle's medium with serum-free 20% knockout serum replacement. The cells, which had a normal karyotype, were grown for more than 25 passages, expressed markers of stemness including Oct-3/4, Tra 1-60, Tra 1-80, and SSEA-4, exhibited high telomerase activity, and differentiated in vitro and in vivo into cells derived from all three germ layers, confirming their pluripotency. Additionally, newly derived hESCs were adapted to growth on a human placental laminin substrate in a defined medium. CONCLUSION(S): To our knowledge, this is the first report of hESC derivation in the absence of serum on qualified pathogen-free human feeders.
OBJECTIVE: To derive new human embryonic stem cell (hESC) lines on pathogen-free human placental fibroblast feeders under serum-free conditions. Because the embryo develops in close contact with extraembryonic membranes, we hypothesized that placental mesenchyme might replicate the stem cell niche in situ. DESIGN: We isolated and characterized human placental fibroblast lines from individual donors and tested their ability to support growth of federally registered hESC lines. Moreover, we performed extensive pathogen testing to ensure their suitability as feeders for the derivation of therapy-grade hESCs. RESULT(S): Human placental fibroblasts were comparable or superior to mouse embryo fibroblasts as hESC feeders. We used these qualified placental fibroblasts to derive two new hESC lines in knockout Dulbecco's modified Eagle's medium with serum-free 20% knockout serum replacement. The cells, which had a normal karyotype, were grown for more than 25 passages, expressed markers of stemness including Oct-3/4, Tra 1-60, Tra 1-80, and SSEA-4, exhibited high telomerase activity, and differentiated in vitro and in vivo into cells derived from all three germ layers, confirming their pluripotency. Additionally, newly derived hESCs were adapted to growth on a human placental laminin substrate in a defined medium. CONCLUSION(S): To our knowledge, this is the first report of hESC derivation in the absence of serum on qualified pathogen-free human feeders.
Authors: R Tayfur Tecirlioglu; Linh Nguyen; Karen Koh; Alan O Trounson; Anna E Michalska Journal: In Vitro Cell Dev Biol Anim Date: 2010-02-23 Impact factor: 2.416
Authors: Maria V Camarasa; Robbie W Kerr; Sharon F Sneddon; Nicola Bates; Lisa Shaw; Rachel A Oldershaw; Fiona Small; Melissa A Baxter; Tristan R Mckay; Daniel R Brison; Susan J Kimber Journal: In Vitro Cell Dev Biol Anim Date: 2010-02-23 Impact factor: 2.416
Authors: Kristiina Rajala; Bettina Lindroos; Samer M Hussein; Riikka S Lappalainen; Mari Pekkanen-Mattila; Jose Inzunza; Björn Rozell; Susanna Miettinen; Susanna Narkilahti; Erja Kerkelä; Katriina Aalto-Setälä; Timo Otonkoski; Riitta Suuronen; Outi Hovatta; Heli Skottman Journal: PLoS One Date: 2010-04-19 Impact factor: 3.240
Authors: Somdutta Roy; Philippe Gascard; Nancy Dumont; Jianxin Zhao; Deng Pan; Sarah Petrie; Marta Margeta; Thea D Tlsty Journal: Proc Natl Acad Sci U S A Date: 2013-03-04 Impact factor: 11.205