Producing primate embryonic stem cells by somatic cell nuclear transfer

Abstract

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.

Taking stem cells to the clinic: Major challenges

“…Nuclear transfer (NT) allows personalizing the hESC-derived tissue to the patient by reprogramming the patient's somatic cells to the pluripotent state using mature oocytes of human or animal origin. This approach although recently accomplished for the macaque [Byrne et al, 2007] and the human [French et al, 2008] is fraught with the problems of low efficiency, faulty faithful epigenesis, the influence of mitochondrial DNA, and implications of any remaining spindle apparatus after oocyte enucleation. Some also consider this approach illogical given the paucity of human oocytes and the concerns of viral transmission if animal oocytes are used.…”

“…Thus far, hESCs generated from surplus human IVF embryos, iPSCs, and non-human primate embryos produce teratomas [Byrne et al, 2007;Aleckovic and Simon, 2008]. It is very likely that hESCs generated via other methods [altered nuclear transfer (ANT), germ cells, parthenogenesis, dead embryos, and blastomeres] will also yield teratomas simply because hESCs are pluripotent.…”

Two-Phase Analysis of Molecular Pathways Underlying Induced Pluripotent Stem Cell Induction

“…Subsequent studies have demonstrated the presence of dominant factors in eggs or embryonic stem cells (ESCs) that can reverse a cell from the differentiated state back to the pluripotent state [4][5][6][7]. Pioneering work by Yamanaka and colleagues (2006) identified the first set of these dominant factors: Oct4, Sox2, Klf4, and c-Myc (OSKM).…”

The ups and downs of somatic cell nucleus transfer (SCNT) in humans

“…Moreover, as noted above, the incidence of genetic and epigenetic instability in human oocytes is disconcerting, further contributing to the overall problem of poor oocyte quality especially as a function of advancing age or environmental factors [15]. Nevertheless, we also do believe that experiments aiming at producing blastocysts by SCNT in humans and primates [28] will certainly accelerate the introduction of some technical improvements, that might be very useful in solving certain problems of assisted human reproduction, into clinical practice. These approaches are now almost exclusively oriented on the elimination of mutated mitochondrial DNA (mtDNA), that can cause some devastating diseases [29].…”

“…For non-human primates, failure to produce cloned offspring by SCNT has been reported. However, blastocysts produced by SCNT have been used to derive embryonic stem cell lines [28]. This now applies for humans too.…”