Derivation of embryonic stem (ES) cells genetically identical to a patient by somatic cell nuclear transfer (SCNT) holds the potential to cure or alleviate the symptoms of many degenerative diseases while circumventing concerns regarding rejection by the host immune system. However, the concept has only been achieved in the mouse, whereas inefficient reprogramming and poor embryonic development characterizes the results obtained in primates. Here, we used a modified SCNT approach to produce rhesus macaque blastocysts from adult skin fibroblasts, and successfully isolated two ES cell lines from these embryos. DNA analysis confirmed that nuclear DNA was identical to donor somatic cells and that mitochondrial DNA originated from oocytes. Both cell lines exhibited normal ES cell morphology, expressed key stem-cell markers, were transcriptionally similar to control ES cells and differentiated into multiple cell types in vitro and in vivo. Our results represent successful nuclear reprogramming of adult somatic cells into pluripotent ES cells and demonstrate proof-of-concept for therapeutic cloning in primates.
"In rhesus monkey SCNT-ESC lines have been obtained (Byrne et al., 2007), but the live birth of cloned animals has not yet been reported. Compared with normal embryos, the ICM cells of cloned embryos maintain a high level of DNA methylation and this may disturb normal embryo development after SCNT (Yang et al., 2007). "
[Show abstract][Hide abstract] ABSTRACT: During human pre-implantation development the totipotent zygote divides and undergoes a number of changes that lead to the first lineage differentiation in the blastocyst displaying trophectoderm and inner cell mass on day 5. The trophectoderm is a differentiated epithelium needed for implantation and the inner cell mass (ICM) forms the embryo proper and serves as a source for pluripotent embryonic stem cells. The blastocyst implants around day 7. The second lineage differentiation occurs in the ICM after implantation resulting in specification of primitive endoderm and epiblast. Knowledge on human pre-implantation development is limited due to ethical and legal restrictions on embryo research and scarcity of materials. Studies in the human are mainly descriptive and lack functional evidence. Most information on embryo development is obtained from animal models and embryonic stem cell cultures and should be extrapolated with caution. This paper reviews totipotency and the molecular determinants and pathways involved in lineage segregation in the human embryo, as well as the role of embryonic genome activation, cell cycle features and epigenetic modifications.
Molecular Human Reproduction 04/2014; 20(7). DOI:10.1093/molehr/gau027 · 3.75 Impact Factor
"Somatic cells derived from these ntES cells have the same immune antigens as the donors and would not be rejected after transplantation into the same individuals. These ntES cells have been established from not only mice, but also from monkey cells.27,28) However, human somatic-cell-nuclear-transferred cells had arrested at 8 cell stage in development.29,30) "
[Show abstract][Hide abstract] ABSTRACT: The "reversion of cell fate from differentiated states back into totipotent or pluripotent states" has been an interest of many scientists for a long time. With the help of knowledge accumulated by those scientists, we succeeded in converting somatic cells to a pluripotent cell lineage by the forced expression of defined factors. These established induced pluripotent stem (iPS) cells have similar features to embryonic stem (ES) cells, including pluripotency and immortality. The iPS cell technology provides unprecedented opportunities for regenerative medicine and drug discovery.
Proceedings of the Japan Academy Ser B Physical and Biological Sciences 03/2014; 90(3):83-96. DOI:10.2183/pjab.90.83 · 2.65 Impact Factor
"Quantitative Analysis The most common comparison at the gene expression level is done by looking at the transcriptome of the cells and by comparing gene expression profiles. In niche (Byrne et al., 2007). Particularly, it is the existence of a functional hierarchy among signaling molecules, which contribute to either the initiation of large-scale phenotypic change or the maintenance of the current state. "
[Show abstract][Hide abstract] ABSTRACT: Somatic cell nuclear transfer (SCNT) and induced pluripotent stem cell (iPSC) technology, also called "direct" reprogramming can be used to generate pluripotent cells. A direct comparison of the two methods can reveal much about the underlying architecture of the reprogramming process. Unfortunately, existing comparisons are limited. To address this, we conducted two comparisons: a literature review that established what is known about such comparisons and a quantitative analysis of secondary microarray data. Our analysis contributes to the existing literature by using the 8-cell embryo as the standard for comparing a range of embryonic stem cells (ESCs) and iPSCs. The goal of these comparisons was to establish differences between cell types of the same cellular state, pluripotency. Using a criterion independent of previous studies, it was found that iPSCs are transcriptionally closer to 8-cell embryos than are ESCs. However, when comparing both ESCs and 8-cell embryos with iPSCs, the same genes tend to be identified as differentially expressed. Annotation using gene ontology terms revealed about half of these genes were ribosomal proteins. These results were confirmed in two ways. One of these was through a mutual information analysis that revealed elevated mutual information for all gene expression in iPSCs. The other was through an indirectly reconstructed gene network analysis. This information can be used to improve our choice of reprogramming approach (SCNT versus iPSC) in future endeavors, as well as identifying groups of supportive (but not essential) genes that might be used to augment efficiency in the reprogramming process.
Principles of Cloning, 2nd edited by Jose Cibelli, Ian Wilmut, Rudolf Jaenisch, John Gurdon, Robert Lanza, Michael West, Keith Campbell, 10/2013: chapter 37: pages 465-471; Elsevier.
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