Growth of human embryonic stem cells using derivates of human fibroblasts.
The majority of human embryonic stem cell (hESC) lines have been derived and grown using mouse or human feeder cells, or using Matrigel, an animal derivative rich in extracellular matrix (ECM) proteins. However, reliance on feeder layers and animal products limits the manipulation and clinical application of hESC. Alternatively, human fibroblasts produce an ECM which could be employed to coated plates and be easily sterilized. We have shown that hESC grown on this matrix and in the presence of medium conditioned by fibroblast cells maintain markers of pluripotency, including expression of cell surface proteins (SSEA3, SSEA4, TRA-1-60, TRA-1-81), alkaline phosphatase activity, and specific intracellular markers (NANOG, OCT, REX1). Moreover, hESC cultured on this novel human-derived ECM display a normal karyotype. This growth system reduces exposure of hESC to feeder layers and animal ingredients, thereby limiting the risk of pathogenic contamination and additionally facilitating manipulation of hESCs.
Escobedo-Lucea
Carmen
C
Centro de Investigación Príncipe Felipe, Valencia, Spain.
Stojkovic
Miodrag
M
D001483:Base Sequence; D015415:Biomarkers / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D002460:Cell Line; D049109:Cell Proliferation; D017077:Culture Media, Conditioned; D017931:DNA Primers / Q000235:genetics; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D007621:Karyotyping; D008855:Microscopy, Electron, Scanning; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D015415:Biomarkers; D017077:Culture Media, Conditioned; D017931:DNA Primers