Differentiation Potential of Histocompatible Parthenogenetic Embryonic Stem Cells

Harvard University, Cambridge, Massachusetts, United States
Annals of the New York Academy of Sciences (Impact Factor: 4.38). 07/2007; 1106(1):209-18. DOI: 10.1196/annals.1392.011
Source: PubMed


Embryonic stem cells (ESCs) hold unique promise for the development of cell replacement therapies, but derivation of therapeutic products from ESCs is hampered by immunological barriers. Creation of HLA-typed ESC banks, or derivation of customized ESC lines by somatic cell nuclear transfer, have been envisioned for engineering histocompatible ESC-derived products. Proof of principle experiments in the mouse have demonstrated that autologous ESCs can be obtained via nuclear transfer and differentiated into transplantable tissues, yet nuclear transfer remains a technology with low efficiency. Parthenogenesis provides an additional means for deriving ESC lines. In parthenogenesis, artificial oocyte activation initiates development without sperm contribution and no viable offspring are produced in the absence of paternal gene expression. Development proceeds readily to the blastocyst stage, from which parthenogenetic ESC (pESC) lines can be derived with high efficiency. We have recently shown that when pESC lines are derived from hybrid mice, early recombination events produce heterozygosity at the major histocompatibility complex (MHC) loci in some of these lines, enabling the generation of histocompatible differentiated cells that can engraft immunocompetent MHC-matched mouse recipients. Here, we explore the differentiation potential of murine pESCs derived in our laboratory.

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    • "pES cells also escape immuno-rejection when used for autologous cell transplantation (Kim et al., 2007a), because when generated from the patient's own oocytes, they are histocompatible with the oocyte donor. Moreover, pES cells exhibit both homozygous and heterozygous major histocompatibility complex protein because of chromosomal recombination occurring during oocyte activation and embryonic development (Kim et al., 2007a,b; Lengerke et al., 2007; Dighe et al., 2008; Lampton et al., 2008; Revazova et al., 2008). pES cells, therefore, should be considered as an important potential source of histocompatible cell lines for women (De Sousa & Wilmut, 2007; Drukker, 2008). "
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    ABSTRACT: Oocytes can reprogram genomes to form embryonic stem (ES) cells. Although ES cells largely escape senescence, oocytes themselves do senesce in the ovaries of most mammals. It remains to be determined whether ES cells can be established using eggs from old females, which exhibit reproductive senescence. We attempted to produce pluripotent stem cell lines from artificial activation of eggs (also called pES) from reproductive aged mice, to determine whether maternal aging affects pES cell production and pluripotency. We show that pES cell lines were generated with high efficiency from reproductive aged (old) mice, although parthenogenetic embryos from these mice produced fewer ES clones by initial two passages. Further, pES cell lines generated from old mice showed telomere length, expression of pluripotency molecular markers (Oct4, Nanog, SSEA1), alkaline phosphatase activity, teratoma formation and chimera production similar to young mice. Notably, DNA damage was reduced in pES cells from old mice compared to their progenitor parthenogenetic blastocysts, and did not differ from that of pES cells from young mice. Also, global gene expression differed only minimally between pES cells from young and old mice, in contrast to marked differences in gene expression in eggs from young and old mice. These data demonstrate that eggs from old mice can generate pluripotent stem cells, and suggest that the isolation and in vitro culture of ES cells must select cells with high levels of DNA and telomere integrity, and/or with capacity to repair DNA and telomeres.
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    ABSTRACT: This review aims to introduce current and future uses of human embryonic stem cells derived from in vitro-fertilized embryos. These and stem cells derived from parthenogenetic and nuclear transfer embryos could be used for cell therapy, as in vitro cell models for drug discovery/screening, and for studying early human development and pathogenesis of human diseases. However, development of therapeutic and screening applications and products from embryonic stem cells is hampered by several barriers. Therefore, gaps in our current understanding of the basic science of stem cells need to be filled before either application can move forward with confidence.
    No preview · Article · Oct 2007 · Clinical Pharmacology &#38 Therapeutics
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    ABSTRACT: Homozygous human embryonic stem cells (hESCs) are thought to be better cell sources for hESC banking because their human leukocyte antigen (HLA) haplotype would strongly increase the degree of matching for certain populations with relatively smaller cohorts of cell lines. Homozygous hESCs can be generated from parthenogenetic embryos, but only heterozygous hESCs have been established using the current strategy to artificially activate the oocyte without second polar body extrusion. Here we report the first successful derivation of a human homozygous ESC line (chHES-32) from a one-pronuclear oocyte following routine in vitro fertilization treatment. chHES-32 cells express common markers and genes with normal hESCs. They have been propagated in an undifferentiated state for more than a year (>P50) and have maintained a stable karyotype of 46, XX. When differentiated in vivo and in vitro, chHES-32 cells can form derivatives from all three embryonic germ layers. The almost undetectable expression of five paternally expressed imprinted genes and their HLA genotype identical to the oocyte donor indicated their parthenogenetic origin. Using genome-wide single-nucleotide polymorphism analysis and DNA fingerprinting, the homozygosity of chHES-32 cells was further confirmed. The results indicated that 'unwanted' one-pronuclear oocytes might be a potential source for human homozygous and parthenogenetic ESCs, and suggested an alternative strategy for obtaining homozygous hESC lines from parthenogenetic haploid oocytes.
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