Immortalized Fibroblast‐Like Cells Derived from Human Embryonic Stem Cells Support Undifferentiated Cell Growth

Abstract

Human embryonic stem cells (hESCs) have the potential to generate multiple cell types and hold promise for future therapeutic applications. Although undifferentiated hESCs can proliferate indefinitely, hESC derivatives significantly downregulate telomerase and have limited replication potential. In this study we examine whether the replicative lifespan of hESC derivatives can be extended by ectopic expression of human telomerase reverse transcriptase (hTERT), the catalytic component of the telomerase complex. To this end, we have derived HEF1 cells, a fibroblast-like cell type, differentiated from hESCs. Infection of HEF1 cells with a retrovirus expressing hTERT extends their replicative capacity, resulting in immortal human HEF1-hTERT cells. HEF1-hTERT cells can be used to produce conditioned medium (CM) capable of supporting hESC growth under feeder-free conditions. Cultures maintained in HEF1-CM show characteristics similar to mouse embryonic fibroblast CM control cultures, including morphology, surface marker and transcription factor expression, telomerase activity, differentiation, and karyotypic stability. In addition, HEF1-hTERT cells have the capacity to differentiate into cells of the osteogenic lineage. These results suggest that immortalized cell lines can be generated from hESCs and that cells derived from hESCs can be used to support their own growth, creating a genotypically homogeneous system for the culture of hESCs. Stem Cells 2004;22: 972-980 STEM CELLS 2004;22:972-980 www.StemCells.com

Derivation and immunological characterization of mesenchymal stromal cells from human embryonic stem cells

“…Although the generation of fibroblast-looking cells from human ESCs was originally reported a few years ago, those cells were not fully characterized as MSCs [23][24][25]. Barberi et al were the first to derive MSCs from human ESCs through co-culturing them with the OP9 murine BM stromal cell line for 40 days and characterize them according to accepted criteria for MSCs [20].…”

Engineering Musculoskeletal Tissues with Human Embryonic Germ Cell Derivatives

“…Attempts at producing functional cartilage from human embryonic cell derivatives under chondrogenic conditions have not always been successful, although these cells readily undergo osteogenic and adipogenic differentiations in standard culture conditions [13,40]. Our tailored coculture system, on the other hand, has enabled LVEC cells to undergo chondrogenic differentiation to produce a tissue with a morphology indicative of hyaline cartilage in a 3D environment.…”

Toward xeno-free culture of human embryonic stem cells

“…Fibroblast-like cells have been derived from H1 cells (WA01), denoted HEF1, and immortalized by infection with a retrovirus expressing human telomerase reverse transcriptase (TERT) (Xu et al, 2004). It was found that conditioned medium from these cells supported the growth of both H7 and H9 cells (WA07 and WA09) on Matrigel.…”