Novel autogenic feeders derived from human embryonic stem cells (hESCs) support an undifferentiated status of hESCs in xeno-free culture conditions

Hsin Fu Chen, Ching Yu Chuang, Yu Kai Shieh, Hao Wei Chang, Hong Nerng Ho, Hung Chih Kuo

Research output: Contribution to journalArticle

38 Citations (Scopus)

Abstract

BACKGROUNDClinical-grade human embryonic stem cells (hESCs) ideally should be derived and maintained in xeno-free culture conditions using defined chemicals or materials of human origin. This will reduce the possibility of xeno-derived pathogenic infection and/or unfavorable immune reaction in clinical application. The present study therefore aimed to derive autogenic feeders from hESCs and evaluate their capability to support the pluripotency of hESCs in xeno-free culture conditions.METHODS AND RESULTSH9 hESCs were cultured in media containing human serum (HS), serum replacement (SR) or KFM combination, to generate autogenic feeders (named HSdF, SRdF and KFMdF, respectively). Reverse transcription polymerase chain reaction, flow cytometry and immunofluorescence analysis using pluripotent stem cell markers, markers of early cell lineages and surface markers revealed that HSdF, SRdF and KFMdF likely belonged to different cellular subpopulations. The efficiency of the autogenic feeders in maintaining pluripotency of H9 hESCs using media containing SR, fetal bovine serum, HS or 1 HS plus various combinations of growth factors was evaluated by flow cytometric analysis of Oct4 expression. All three autogenic feeders were shown to be capable of maintaining the undifferentiated status of H9 hESCs in SR-containing media in long-term culture. When supplemented with bFGF, activin A and noggin, hESCs could also be maintained favorably on KFMdF in a medium containing 1 HS without losing their pluripotent potentials both in vitro and in vivo.CONCLUSIONSNovel autogenic feeders can be derived from hESCs under xeno-free conditions and they can robustly maintain the pluripotent identity of hESCs in xeno-free media containing a low concentration of HS.

Original languageEnglish
Pages (from-to)1114-1125
Number of pages12
JournalHuman Reproduction
Volume24
Issue number5
DOIs
Publication statusPublished - Jan 1 2009
Externally publishedYes

Fingerprint

Embryo Culture Techniques
Activins
Pluripotent Stem Cells
Fibroblast Growth Factors
Xenobiotics
Embryonic Stem Cells
Culture Media
Cell Differentiation
Cultured Cells
Carrier Proteins
Flow Cytometry
Serum
Human Embryonic Stem Cells
Cell Lineage
Reverse Transcription
Fluorescent Antibody Technique
Intercellular Signaling Peptides and Proteins

Keywords

  • Autogenic feeder
  • Embryonic stem cells
  • Human serum
  • Pluripotency
  • Xeno-free culture

ASJC Scopus subject areas

  • Reproductive Medicine
  • Obstetrics and Gynaecology

Cite this

Novel autogenic feeders derived from human embryonic stem cells (hESCs) support an undifferentiated status of hESCs in xeno-free culture conditions. / Chen, Hsin Fu; Chuang, Ching Yu; Shieh, Yu Kai; Chang, Hao Wei; Ho, Hong Nerng; Kuo, Hung Chih.

In: Human Reproduction, Vol. 24, No. 5, 01.01.2009, p. 1114-1125.

Research output: Contribution to journalArticle

Chen, Hsin Fu ; Chuang, Ching Yu ; Shieh, Yu Kai ; Chang, Hao Wei ; Ho, Hong Nerng ; Kuo, Hung Chih. / Novel autogenic feeders derived from human embryonic stem cells (hESCs) support an undifferentiated status of hESCs in xeno-free culture conditions. In: Human Reproduction. 2009 ; Vol. 24, No. 5. pp. 1114-1125.
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T1 - Novel autogenic feeders derived from human embryonic stem cells (hESCs) support an undifferentiated status of hESCs in xeno-free culture conditions

AU - Chen, Hsin Fu

AU - Chuang, Ching Yu

AU - Shieh, Yu Kai

AU - Chang, Hao Wei

AU - Ho, Hong Nerng

AU - Kuo, Hung Chih

PY - 2009/1/1

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N2 - BACKGROUNDClinical-grade human embryonic stem cells (hESCs) ideally should be derived and maintained in xeno-free culture conditions using defined chemicals or materials of human origin. This will reduce the possibility of xeno-derived pathogenic infection and/or unfavorable immune reaction in clinical application. The present study therefore aimed to derive autogenic feeders from hESCs and evaluate their capability to support the pluripotency of hESCs in xeno-free culture conditions.METHODS AND RESULTSH9 hESCs were cultured in media containing human serum (HS), serum replacement (SR) or KFM combination, to generate autogenic feeders (named HSdF, SRdF and KFMdF, respectively). Reverse transcription polymerase chain reaction, flow cytometry and immunofluorescence analysis using pluripotent stem cell markers, markers of early cell lineages and surface markers revealed that HSdF, SRdF and KFMdF likely belonged to different cellular subpopulations. The efficiency of the autogenic feeders in maintaining pluripotency of H9 hESCs using media containing SR, fetal bovine serum, HS or 1 HS plus various combinations of growth factors was evaluated by flow cytometric analysis of Oct4 expression. All three autogenic feeders were shown to be capable of maintaining the undifferentiated status of H9 hESCs in SR-containing media in long-term culture. When supplemented with bFGF, activin A and noggin, hESCs could also be maintained favorably on KFMdF in a medium containing 1 HS without losing their pluripotent potentials both in vitro and in vivo.CONCLUSIONSNovel autogenic feeders can be derived from hESCs under xeno-free conditions and they can robustly maintain the pluripotent identity of hESCs in xeno-free media containing a low concentration of HS.

AB - BACKGROUNDClinical-grade human embryonic stem cells (hESCs) ideally should be derived and maintained in xeno-free culture conditions using defined chemicals or materials of human origin. This will reduce the possibility of xeno-derived pathogenic infection and/or unfavorable immune reaction in clinical application. The present study therefore aimed to derive autogenic feeders from hESCs and evaluate their capability to support the pluripotency of hESCs in xeno-free culture conditions.METHODS AND RESULTSH9 hESCs were cultured in media containing human serum (HS), serum replacement (SR) or KFM combination, to generate autogenic feeders (named HSdF, SRdF and KFMdF, respectively). Reverse transcription polymerase chain reaction, flow cytometry and immunofluorescence analysis using pluripotent stem cell markers, markers of early cell lineages and surface markers revealed that HSdF, SRdF and KFMdF likely belonged to different cellular subpopulations. The efficiency of the autogenic feeders in maintaining pluripotency of H9 hESCs using media containing SR, fetal bovine serum, HS or 1 HS plus various combinations of growth factors was evaluated by flow cytometric analysis of Oct4 expression. All three autogenic feeders were shown to be capable of maintaining the undifferentiated status of H9 hESCs in SR-containing media in long-term culture. When supplemented with bFGF, activin A and noggin, hESCs could also be maintained favorably on KFMdF in a medium containing 1 HS without losing their pluripotent potentials both in vitro and in vivo.CONCLUSIONSNovel autogenic feeders can be derived from hESCs under xeno-free conditions and they can robustly maintain the pluripotent identity of hESCs in xeno-free media containing a low concentration of HS.

KW - Autogenic feeder

KW - Embryonic stem cells

KW - Human serum

KW - Pluripotency

KW - Xeno-free culture

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