Establishment of a human embryonic stem cell line using...

C - Chemistry – Metallurgy – 12 – N

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C12N 5/0735 (2010.01)

Patent

CA 2608833

Purified preparations of human embryonic stem cells with certain population- specific characteristics are disclosed. This preparation is characterized by the positive expression of the following pluripotent cell surface markers: SSEA-1 (-); SSEA-4 (+); TRA-1-60 (+); TRA-1-81 (+); alkaline phosphatase (+), as well as a set of ES cell markers including Oct-4, Nanog, Rexl, Sox2, Thyl, FGF4, ABCG2, Dppa5, UTFl, Criptol, hTERT, Connexin-43 and Connexin-45. The cells of the preparation are negative for lineage specific markers like Keratin 8, Sox-1, NFH (ectoderm), MyoD, brachyury, cardiac-actin (mesoderm), HNF-3 beta, albumin, and PDXl (endoderm). The cells of the preparation are human embryonic stem cells, have normal karyotypes, exhibit high telomerase activity and continue to proliferate in an undifferentiated state after continuous culture for over 40 passages. The embryonic stem cell line RelicellTM Hes1 also retains the ability, throughout the culture, to differentiate into cell and tissue types derived from all three embryonic germ layers (endoderm, mesoderm and ectoderm). Methods for isolating a human embryonic stem cell line are also disclosed.

L'invention concerne des préparations purifiées de cellules souches embryonnaires humaines possédant certaines caractéristiques spécifiques d'une population. Ladite préparation est caractérisée par l'expression positive des marqueurs de surface cellulaire pluripotents : SSEA-1 (-), SSEA-4 (+), TRA-1-60 (+), TRA-1-81 (+), la phosphatase alkaline (+), ainsi que d'un ensemble de marqueurs cellulaires ES comprenant Oct-4, Nanog, Rexl, Sox2, Thyl, FGF4, ABCG2, Dppa5, UTFl, Criptol, hTERT, Connexin-43 et Connexin-45. Les cellules de la préparation sont négatives pour des marqueurs spécifiques de lignée, comme la kératine 8, Sox-1, NFH (ectoderme), MyoD, brachyury, l'actine cardiaque (mésoderme), HNF-3 bêta, l'albumine, et PDXl (endoderme). Les cellules de la préparation sont des cellules souches embryonnaires humaines, elles possèdent des caryotypes normaux, une activité de la télomérase élevée et elles continuent à proliférer dans un état non différencié, après une culture continue pour plus de 40 passages. La lignée de cellules souches embryonnaires RelicellTM Hes1 possède la capacité, tout au long de la culture, de se différenciser en types cellulaires et tissulaires dérivés des trois feuillets cellulaires (endoderme, mésoderme et ectoderme). Cette invention a aussi pour objet des méthodes d'isolation d'une lignée de cellules souches embryonnaires humaines.

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