Triploid pluripotent stem cells
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Triploid pluripotent stem cells. R eporter : XueBinghua Date : 2011.06.15. Human oocytes reprogram somatic cells to a pluripotent state. Scott Noggle & Dieter Egli Nature 1 August 2011. Background. Limit : Legal and social considerations Different : iPS and blastocyst-derived stem cells

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Triploid pluripotent stem cells

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Triploid pluripotent stem cells

Triploidpluripotent stem cells

Reporter:XueBinghua

Date:2011.06.15


Human oocytes reprogram somatic cells to a pluripotent state

Human oocytes reprogram somatic cells to a pluripotent state

Scott Noggle & Dieter Egli

Nature

1 August 2011


Background

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)


R esults

Results:

1、Development fails after somatic

genome replacement


Schematic of genome exchange in human oocytes

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

MII oocyte & Donor cell


C hromosome condensation

35/35

Chromosome condensation


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%)

Developmentalpotential & Development arrests


C ontrol for the quality of the oocytes

6-8h

Control for the quality of the oocytes


Triploid pluripotent stem cells

32/32

4/7


Triploid pluripotent stem cells

Results:

2、Development arrests with

transcriptional defects


Triploid pluripotent stem cells

6-12cells

*ZGA starts at the 4–8 cell stage28, or on day 3 of development*Upregulatedtranscripts 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 expressat higher levels in the skin donor fibroblasts than in MIIoocytesGADD45G is involved in stress-induced cell cycle arrest


Triploid pluripotent stem cells

Results:

3、The oocyte genome rescues development


Schematic of somatic genome transfer

Schematic of somatic genome transfer


Developmental progression

Developmental progression


Developmental potential

Developmental potential


Triploid pluripotent stem cells

Results:

4、Derivation of pluripotent stem cells


Karyotype sequence of the mitochondrion

Karyotype & Sequence of the mitochondrion


Pluripotency

Pluripotency


G lobal gene expression analysis up and down

463/1327

670/1327

Global gene expression analysis &Up and down

*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


Triploid pluripotent stem cells

Results:

5、Epigenetic reprogramming


M ethylation of dna at the nanog promoter

Methylation of DNA at the Nanog promoter


Rt pcr and snp dmr of peg3

RT–PCR andSNP & DMR of PEG3


Schematic of genome wide snp capture and allelotyping

Schematic of genome-wideSNP 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

Contribution of the reprogrammed genome


T ranscript ratio for the different cell genes

Transcript ratio for the different cell genes


Triploid pluripotent stem cells

Thank you!


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