Derivation, culture and retinal pigment epithelial differentiation of human embryonic stem cells using human fibroblast feeder cells.
OBJECTIVE
Retinal pigment epithelium cells derived from human embryonic stem cells (ESCs) could be useful for restoring retinal function in age-related macular degeneration. However the use of non-human feeder cells to support the growth of ESCs for clinical applications raises the concern of possible contamination because of direct contact between animal and human cells.


METHODS
In this study, we produced human ESCs using human fibroblast feeder layers isolated from foreskin and abdominal tissues. Using this system, human ESCs differentiated into retinal pigment epithelium cells in differentiation medium.


RESULTS
Seven human ESC lines were established from 18 blastocysts. These human ESCs showed normal morphology, expressed all expected cell surface markers, had the ability to form embryoid bodies upon culture in vitro and teratomas after injection into SCID mice, and differentiated further into derivatives of all three germ layers. Under conditions of committed differentiation, these human ESCs could differentiate into retinal pigment epithelium cells after 2 months in culture.


CONCLUSIONS
The results of this study demonstrated that human foreskin/abdominal fibroblasts have the potential to support the derivation and long-term culture of human ESCs, which can then be used to generate retinal pigment epithelium cells with characteristic morphology and molecular markers. This technique avoids the concerns of contamination from animal feeder layers during human ESC derivation, culture and differentiation, and will thus facilitate the development of retinal pigment epithelium cell transplantation therapy.
Center for Reproductive Medicine, Tianjin Central Hospital of Obstetrics and Gynecology, Tianjin, 300100, China. tjzys@hotmail.com
Yun-Shan
YS
Zhang
Zhen-Yu
ZY
Lu
Yang
Y
Yu
Xiao-Rong
XR
Li
Wen-Bo
WB
Li
Yi-Na
YN
Wang
Ying
Y
Geng
D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D001755:Blastocyst / Q000166:cytology; D002454:Cell Differentiation; D002460:Cell Line; D002648:Child; D002675:Child, Preschool; D003470:Culture Media / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D061252:Feeder Cells / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism*; D052816:Foreskin / Q000166:cytology; D005855:Germ Layers / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008297:Male; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D061251:Primary Cell Culture / Q000379:methods*; D055213:Retinal Pigment Epithelium / Q000166:cytology* / Q000378:metabolism; D012867:Skin / Q000166:cytology / Q000378:metabolism; D013724:Teratoma / Q000378:metabolism / Q000473:pathology
D015415:Biomarkers; D003470:Culture Media