Abstract
Conventionally, mouse embryonic fibroblasts (MEFs) inactivated by mitomycin C or irradiation were applied to support the self-renew and proliferation of human embryonic stem cells (hESCs). To avoid the disadvangtages of mitomycin C and irradiation, here MEFs were treated by ethanol (ET). Our data showed that 10% ET-inactivated MEFs (eiMEFs) could well maintain the self-renew and proliferation of hESCs. hESCs grown on eiMEFs expressed stem cell markers of NANOG, octamer-binding protein 4 (OCT4), stage-specific embryonic antigen-4 (SSEA4) and tumour related antigen-1-81 (TRA-1-81), meanwhile maintained normal karyotype after long time culture. Also, hESCs cocultured with eiMEFs were able to form embryoid body (EB) in vitro and develop teratoma in vivo. Moreover, eiMEFs could keep their nutrient functions after long time cryopreservation. Our results indicate that the application of eiMEF in hESCs culture is safe, economical and convenient, thus is a better choice.
Highlights
Human embryonic stem cells are pluripotent, can give rise to all tissue and cell types of the human body [1], have offered great promise for regenerative medicine, gene and cell therapies, and disease modeling[2]
There are reports demonstrated that low concentration of ET can inhibit cell proliferation by impeding DNA synthesis[32, 33], which remind us that mitomycin C (MC) inhibits cell growth through preventing spindle formation
Here we test if low concentration of ET can inactivate mouse embryonic fibroblasts (MEFs), and if the ET-inactivated MEFs can maintain Human embryonic stem cells (hESCs) self-renew like the MC has done
Summary
Human embryonic stem cells (hESCs) are pluripotent, can give rise to all tissue and cell types of the human body [1], have offered great promise for regenerative medicine, gene and cell therapies, and disease modeling[2]. Different concentrations of ethanol were applied to treat MEFs. We found that 10% ethanol-inactivated MEFs (eiMEFs) were capable of supporting the long-term self-renew and proliferation of hESCs. the pluripotency and normal karyotype of hESCs growing on eiMEFs were kept.
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