Self-renewal and differentiation capabilities are variable between human embryonic stem cell lines I3, I6 and BG01V.
BACKGROUND
A unique and essential property of embryonic stem cells is the ability to self-renew and differentiate into multiple cell lineages. However, the possible differences in proliferation and differentiation capabilities among independently-derived human embryonic stem cells (hESCs) are not well known because of insufficient characterization. To address this question, a side-by-side comparison of 1) the ability to maintain an undifferentiated state and to self-renew under standard conditions; 2) the ability to spontaneously differentiate into three primary embryonic germ lineages in differentiating embryoid bodies; and 3) the responses to directed neural differentiation was made between three NIH registered hES cell lines I3 (TE03), I6 (TE06) and BG01V. Lines I3 and I6 possess normal XX and a normal XY karyotype while BG01V is a variant cell line with an abnormal karyotype derived from the karyotypically normal cell line BG01.


RESULTS
Using immunocytochemistry, flow cytometry, qRT-PCR and MPSS, we found that all three cell lines actively proliferated and expressed similar "stemness" markers including transcription factors POU5F1/Oct3/4 and NANOG, glycolipids SSEA4 and TRA-1-81, and alkaline phosphatase activity. All cell lines differentiated into three embryonic germ lineages in embryoid bodies and into neural cell lineages when cultured in neural differentiation medium. However, a profound variation in colony morphology, growth rate, BrdU incorporation, and relative abundance of gene expression in undifferentiated and differentiated states of the cell lines was observed. Undifferentiated I3 cells grew significantly slower but their differentiation potential was greater than I6 and BG01V. Under the same neural differentiation-promoting conditions, the ability of each cell line to differentiate into neural progenitors varied.


CONCLUSIONS
Our comparative analysis provides further evidence for similarities and differences between three hESC lines in self-renewal, and spontaneous and directed differentiation. These differences may be associated with inherited variation in the sex, stage, quality and genetic background of embryos used for hESC line derivation, and/or changes acquired during passaging in culture.
Tavakoli
Tahereh
T
Stem Cell Center, Developmental Biology, American Type Culture Collection, Manassas, VA, USA. ttavakoli@atcc.org
Xu
Xiangru
X
Derby
Eric
E
Serebryakova
Yevgeniya
Y
Reid
Yvonne
Y
Rao
Mahendra S
MS
Mattson
Mark P
MP
Ma
Wu
W
D000469:Alkaline Phosphatase / Q000378:metabolism; D000954:Antigens, Surface / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism; D001973:Bromodeoxyuridine / Q000378:metabolism; D018929:Cell Culture Techniques / Q000592:standards; D002454:Cell Differentiation; D002460:Cell Line; D049109:Cell Proliferation; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000502:physiology*; D005434:Flow Cytometry; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D000071317:Nanog Homeobox Protein; D009457:Neuroglia / Q000166:cytology / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D050814:Octamer Transcription Factor-3 / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055746:Stage-Specific Embryonic Antigens / Q000378:metabolism
D000954:Antigens, Surface; D015415:Biomarkers; D018398:Homeodomain Proteins; C486780:NANOG protein, human; D000071317:Nanog Homeobox Protein; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; D055746:Stage-Specific Embryonic Antigens; C519803:TRA-1-81 antigen, human; C066584:stage-specific embryonic antigen-4; D000469:Alkaline Phosphatase; D001973:Bromodeoxyuridine