Abstract
Human pluripotent stem cells (hPSCs), including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), are capable of differentiating into any cell type in the human body and thus can be used in studies of early human development, as cell models for different diseases and eventually also in regenerative medicine applications. Since the first derivation of hESCs in 1998, a variety of culture conditions have been described for the undifferentiated growth of hPSCs. In this study, we cultured both hESCs and hiPSCs in three different culture conditions: on mouse embryonic fibroblast (MEF) and SNL feeder cell layers together with conventional stem cell culture medium containing knockout serum replacement and basic fibroblast growth factor (bFGF), as well as on a Matrigel matrix in mTeSR1 medium. hPSC lines were subjected to cardiac differentiation in mouse visceral endodermal-like (END-2) co-cultures and the cardiac differentiation efficiency was determined by counting both the beating areas and Troponin T positive cells, as well as studying the expression of OCT-3/4, mesodermal Brachyury T and NKX2.5 and endodermal SOX-17 at various time points during END-2 differentiation by q-RT-PCR analysis. The most efficient cardiac differentiation was observed with hPSCs cultured on MEF or SNL feeder cell layers in stem cell culture medium and the least efficient cardiac differentiation was observed on a Matrigel matrix in mTeSR1 medium. Further, hPSCs cultured on a Matrigel matrix in mTeSR1 medium were found to be more committed to neural lineage than hPSCs cultured on MEF or SNL feeder cell layers. In conclusion, culture conditions have a major impact on the propensity of the hPSCs to differentiate into a cardiac lineage.
Highlights
Human pluripotent stem cells include human embryonic stem cells and human induced pluripotent stem cells. hPSCs are able to self-renew and to differentiate into any human cell type; they can be used as a cell model to study embryology and disease pathophysiology. hPSCs have additional utility in drug screening applications and as a cell source for regenerative medicine in the future
The impact of three different culture methods for cardiac differentiation of hPSCs were compared: mouse embryonic fibroblast (MEF) and SNL feeder cell layers combined with conventional stem cell culture medium containing knockout serum replacement and basic fibroblast growth factor, and a Matrigel matrix combined with commercial mTeSR1 medium
The morphology of the pluripotent stem cell colonies varies with the culture conditions In this study, a single human embryonic stem cells (hESCs) line (H7) and four human induced pluripotent stem cells (hiPSCs) lines (UTA.00112.human foreskin fibroblast (hFF), UTA.04602.WT, UTA.00525.LQT2 and UTA.00106.hFF) were cultured with three different culture methods: MEF and SNL feeder cell layers combined with conventional stem cell culture medium and Matrigel matrix combined with mTeSR1 medium, and subjected into cardiac differentiation in mouse visceral endodermal-like cell (END-2) cocultures
Summary
Human pluripotent stem cells (hPSCs) include human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs). hPSCs are able to self-renew and to differentiate into any human cell type; they can be used as a cell model to study embryology and disease pathophysiology. hPSCs have additional utility in drug screening applications and as a cell source for regenerative medicine in the future. Since the first derivation of a hESC line in 1998 on a mouse embryonic fibroblast (MEF) feeder cell layer [1], many hPSC culture methods based on different human feeder cell layers [2,3], autologous feeder cells [4,5], feeder cell-free [6,7,8,9,10] and suspension culture techniques [11] have been developed and described. Because the ultimate aim of hPSC research is to use the cells in regenerative medicine applications, culture conditions are being optimized in the xenofree direction. Culturing feeder cells is very laborious and time-consuming, and for the regenerative medicine applications, a large number of hPSCs are needed. Despite the tremendous effort made to optimize hPSC culture conditions, a universal and reliable, xenoand feeder-free culture method remains to be discovered
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
