E-GEOD-54876 - Human oocytes reprogram adult somatic nuclei to diploid pluripotent stem cells

Status
Released on 27 April 2014, last updated on 6 May 2014
Organism
Homo sapiens
Samples (5)
Array (1)
Protocols (5)
Description
The transfer of somatic cell nuclei into oocytes can give rise to pluripotent stem cells, holding promise for autologous cell replacement therapy. Though reprogramming of somatic cells by nuclear transfer was first demonstrated more than 60 years ago, only recently have human diploid embryonic stem cells been derived after nuclear transfer of fetal and neonatal fibroblasts. Because of the therapeutic potential of developing diploid embryonic stem cell lines from adult cells of normal and diseased human subjects, we have systematically investigated the parameters affecting efficiency and developmental potential in their derivation. We found that improvements to the oocyte activation protocol, including the use of both a kinase and a translation inhibitor, and cell culture in the presence of histone deacetylase inhibitors enable development of diploid cells to the blastocyst stage. Developmental efficiency varied significantly between oocyte donors, and was inversely related to the number of days of hormonal stimulation required to reach mature oocytes, while the daily dose of gonadotropin or the total number of MII oocytes retrieved did not affect developmental outcome. The use of diluted Sendai virus in calcium-free medium during nuclear transfer improved developmental potential, while the use of concentrated Sendai virus induced an increase in intracellular calcium and caused premature oocyte activation. Using these modifications to the nuclear transfer protocol, we successfully derived diploid pluripotent stem cell lines from both postnatal and adult somatic cells of a type 1 diabetic subject. Gene expression analysis was performed on a total of 5 human cell lines, including an isogenic set of 3 nuclear-transfer embryonic stem cell lines and their parental neonatal fibroblast cell line, as well as a fourth nuclear-transfer embryonic stem cell line, which was derived from adult fibroblasts from a type 1 diabetic subject.
Experiment type
transcription profiling by array 
Contacts
Dieter Egli, Ido Sagi, Nissim Benvenisty
MIAME
PlatformsProtocolsVariablesProcessedRaw
Files
Investigation descriptionE-GEOD-54876.idf.txt
Sample and data relationshipE-GEOD-54876.sdrf.txt
Raw data (1)E-GEOD-54876.raw.1.zip
Processed data (1)E-GEOD-54876.processed.1.zip
Array designA-AFFY-141.adf.txt
Links