Triploid pluripotent stem cells

Reporter： XueBinghuaDate： 2011.06.15

Human oocytes reprogram somatic cells to a pluripotent state

Scott Noggle & Dieter EgliNature

1 August 2011

Background

• Limit： Legal and social considerations

• Different： iPS and blastocyst-derived stem cells (gene expression,DNA methylation and differentiation potential)

• Compromise: Genomic integrity (de novo mutations and copy number variations)

Results:

1、 Development fails after somatic genome replacement

Schematic of genome exchange in human oocytes

MII oocyte & Donor cell

38/50

43/43 (12 without)

Skin cells (♂T1D and ♂WT )

GFP

CAGGS promoter:(H2B):GFP

Chromosome condensation

35/35

Developmental potential & Development arrests

Normal cleavage :22/31(71%)

Arrests at a stage of 6–10 cells

Parthenotes:13/52 ( 25%)

IVF： 16/21(76%)

Control for the quality of the oocytes

6-8h

32/32

4/7

Results:

2、Development arrests with transcriptional defects

*ZGA starts at the 4–8 cell stage28, or on day 3 of development*Upregulated transcripts incomparison to unfertilized oocytes 761 transcripts upregulated at zygotic genome activation 124 (16%) were upregulated after genome exchange 62 (8%) were upregulated after amanitin treatment 536(70%) were upregulated at parthenotes

*1,406 genes express at higher levels in the skin donor fibroblasts than in MII oocytes GADD45G is involved in stress-induced cell cycle arrest

6-12cells

Results:

3、 The oocyte genome rescues development

Schematic of somatic genome transfer

Developmental progression

Developmental potential

Results:

4、 Derivation of pluripotent stem cells

Karyotype & Sequence of the mitochondrion

Pluripotency

Global gene expression analysis &Up and down

463/1327 670/1327

*463:Lin28a,Oct4,Sox2, Nanog ,Lefty1*670:FAP, PAPPA, MMP3,CTHRC1 SNAI2 *28 genes and 24 genes were expressed at higher levels in soPS2 and soPS1 than NYSCF1 neuropeptide galanin, SRY, rex1 , neuronatin, NODAL, Cerberus 1 , Lefty2

Results:

5、 Epigenetic reprogramming

Methylation of DNA at the Nanog promoter

RT–PCR and SNP & DMR of PEG3

Schematic of genome-wide SNP capture and allelotyping

Use a genome-wide digital allelotyping approach to distinguish gene expression from the somatic cell-derived genome and the oocyte-derived genome in soPS cells

Contribution of the reprogrammed genome

Transcript ratio for the different cell genes