Framing the Debates on Human Cloning and Human Embryonic Stem Cells: Pluripotent vs. TOTIPOTENT


1 Erik Parens and Lori P. Knowles, "Reprogenetics and Publc Policy: Reflections and Recommendations", Hastings Center Special Supplement (July-August 2003), at: (emphasis added) [Back]

2 See White Paper: Alternative Sources of Pluripotent Stem Cells, The President's Council on Bioethics, Washington, D.C., May 2005 ( [Back]

3 In Carla Garnett, "The Science of Stem Cells", NIH Record, Feb 8, 2002 ( [Back]

4 John C. Fletcher, "Pluripotential stem cell research: Two recent reports", Current Debate on Embryo Research, ASBH Exchange 1999 Archive, American Society for Bioethics and Humanities ( [Back]

5 Margaret Doris, "Human Embryonic Stem Cell Research: An Overview By Margaret Doris, Bioethics Working Group at Boston University on "Stem Cell Research" ( [Back]

6 Embryonic "stem" cells from the inner cell mass (ICM) of the human blastocyst are totipotent: Ronan O'Rahilly and Fabiola Muller, Human Embryology & Teratology (3rd ed.) (New York: Wiley-Liss, 2001): "As soon as a cavity can be detected (by light microscopy) in the cellular mass of the morula, the organism is termed a blastocyst. This occurs when about 16-32 cells are present. The embryo is about 4 days in age and is not yet attached to the uterine mucosa. ... The appearance of the blastocyst demonstrates the differentiation into (1) trophoblast (or trophectoderm), the peripherally situated cells and (under the influence of E-cadherin) in first epithelium formed, and (2) embryonic cells proper. The latter, at first few in number, form the inner cell mass (ICM). The trophoblast at the future site of attachment is sometimes termed polar, the remainder being called mural. The cells of the ICM (inner cell mass) are considered to be totipotent initially. They give rise directly to various lines of embryonic stem cells. (p. 39) [Back]

7  Germ line cells Are totipotent (and therefore can be cloned by "twinning"): Ronan O'Rahilly and Fabiola Muller, Human Embryology & Teratology (New York: Wiley-Liss, 1994); also, O'Rahilly and Muller, ibid. (3rd ed., 2001). [Note: O'Rahilly is one of the originators of the Carnegie Stages of Early Human Embryological Development, and has sat on the international Nomina Embryologica Committee for decades - DNI]: "Primordial germ cells (PGC), or gonocytes, are generally believed to be both extragonadal and extraembryonic in origin. They are difficult to recognize in very young human embryos. Claims for them have been made as early as in the blastocyst, and they are believed to be segregated at latest by 2 1/2 weeks and possibly much earlier. At 4 weeks they can be identified in the umbilical vesicle (yolk sac) and hindgut. A week or so later, they have migrated to the gonads. When the primordial germ cells have settled in the gonad, they become more spherical, stain less intensely with alkaline phosphatase, undergo mitosis, and are then referred to as oogonia or spermatogonia, depending on whether they are situated in an ovary or a testis. ... The unifying feature in the formation of primordial germ cells would seem to be the exemption of those cells from the processes of regional, somatic differentiation. . (pp., 23-24)

... Cells differentiate by the switching off of large portions of their genome. Future somatic cells thereby lose their totipotency and are liable to senescence, whereas germ cells regain their totipotency after meiosis and fertilization. (p. 39)

... Stem cells comprise a small subpopulation of multipotent or pluripotent, ultrastructurally unspecialized, slow-cycling cells that possess the ability of self-renewal and can produce cells that are destined to differentiate. (In contrast, primordial germ cells and those of a morula are totipotent; i.e., they can develop into any type of embryonic tissue and can even form an entirely new embryo) ... In addition to embryonic stem cells from preimplantation blastocysts, stem cells can be obtained from an adult. (p. 136) ... Ethical concerns are intensified by the experimental finding in primates (in contrast to the mouse) that embryonic stem cells are totipotent and can develop into a complete embryo with a primitive streak. ... If the source of the stem cells is a human embryo or fetus, however, ethical and legal issues have to be considered. (pp. 136-137)

Benjamin Lewin, Genes VII [Oxford: Oxford University Press and Cell Press, 2000), p. 605]: A change in methylation pattern occurs during embryogenesis. All allelic differences are lost when priordial germ cells develop in the embryo; irrespective of sex, the previous patterns of methylation are erased, and a typical gene is then unmethylated. The methylation pattern of germ cells is therefore established by a two stage process: first the previous pattern is erased by a genome-wide demethylation; then the pattern specific for each sex is imposed.

Santos F, Dean W, "Epigenetic reprogramming during early development in mammals", Reproduction. 2004 Jun;127(6):643-51 [PMID: 15175501] ( Epigenetic modifications serve as an extension of the information content by which the underlying genetic code may be interpreted. ... In mammals, DNA methylation and the modification of histones account for the major epigenetic alterations. Two cycles of DNA methylation reprogramming have been characterised. During germ cell development, epigenetic reprogramming of DNA methylation resets parent-of-origin based genomic imprints and restores totipotency to gametes. [Back]

8  Germ line cells are diploid (and therefore can be cloned by nuclear transfer): Ronan O'Rahilly and Fabiola Muller, Human Embryology & Teratology (3rd ed.) (New York: Wiley-Liss, 2001): Gametogenesis is the production of germ cells (gametes), i.e., spermatozoa and oocytes. ... The gametes are believed to arise by successive divisions from a distinct line of cells (the germ plasm), and the cells that are not directly concerned with gametogenesis are termed somatic. ... Diploid refers to the presence of two sets of homologous chromosomes: 23 pairs, making a total of 46. This is characteristic of somatic and primordial germ cells alike. (p. 19)

Tom Strachan and Andrew Read, Human Molecular Genetics (2nd ed.) (New York: Wiley-Liss, 1999): A subset of the diploid body cells constitute the germ line. These give rise to specialized diploid cells in the ovary and testis that can divide by meiosis to produce haploid gametes (sperm and egg). ... The other cells of the body, apart from the germ line, are known as somatic cells ... most somatic cells are diploid ... (p. 28)

Keith Moore and T.V.N. Persaud, The Developing Human: Clinically Oriented Embryology (6th ed. only) (Philadelphia: W.B. Saunders Company, 1998): Meiosis is a special type of cell division that involves two meiotic cell divisions; it takes place in germ cells only. Diploid germ cells give rise to haploid gametes (sperms and oocytes). (p. 18)

Bruce M. Carlson, Human Embryology & Developmental Biology (St. Louis, MO: Mosby, 1999): "In a mitotic division, each germ cell produces two diploid progeny that are genetically equal." (p. 2) [Back]

9  See, e.g., the most authoritative site at National Museum of Health and Medicine, Research Collections, Human Developmental Anatomy Center: "Anatomy", Carnegie Stages of Early Human Development (; see pdf files of Developmental Stages in Human Embryos by Ronan O'Rahilly and Fabiola Müler, published by Carnegie Institution of Washington, Publication 637. 1987. Stage One (, [from penetration through zygote formation]: Embryonic life commences with fertilization, and hence the beginning of that process may be taken as the point de depart of stage 1.

Stage Two (, page 1): Stage 2 comprises specimens from 2 cells up to the appearance of the blastocystic (or segmentation) cavity. The more advanced examples (from about 12 cells on) of stage 2 are frequently called morulae ... The organism proceeds along the uterine tube ... It leaves the tube and enters the uterine cavity during the third or fourth day after ovulation, when probably 8-12 cells are present ... It has been shown experimentally ... that a blastomere isolated from the mammalian 2-cell organism is capable of forming a complete embryo. Separation of the early blastomeres is believed to account for about one-third of all cases of monozygotic twinning in the human (Corner, 1955).

Stage Three (, free blastocyst): Stage 3 consists of the free (that is, unattached) blastocyst, a term used as soon as a cavity ... can be recognized by light microscopy. ... The blastocyst is the hollow mass of cells from the initial appearance of the cavity (stage 3) to immediately before the completion of implantation at a subsequent stage. The blastocystic cavity, under the light microscope, begins by the coalescence of intercellular spaces when the organism has acquired about 32 cells. In in vitro studies, a cavity formed in some human embryos at 16-20 cells (Edwards, 1972). ... In stage 3 the zona pellucida may be either present or absent. In vitro, the blastocyst emerges from the zona at about 6-7 days. The emergence is commonly referred to as "hatching."... The embryonic cells proper become surrounded by the trophoblastic cells and form an inner mass. Studies of various mammals have indicated that the inner cell mass represents more than the embryo itself, insofar as it constitutes a germinal mass of various potentialities which continues for a time to add cells to the more precociously developed trophoblast. The inner cell mass gives origin to the hypoblast, and its remainder (the "formative cells") constitutes the epiblast. The epiblastic cells soon become aligned into what was frequently described as the "germ disc." Duplication of the inner cell mass probably accounts for most instances of monozygotic twinning (Corner, 1955; Bulmer, 1970). ... In vitro, "many blastocysts fail to hatch fully from their zona pellucida," and "two separate embryos could form if the inner cell mass was bisected during hatching" (Edwards, Mettler, and Walters, 1986).

Stage 4 (, implanting blastocyst): Stage 4, the onset of implantation, is reserved for the attaching blastocyst, which is probably 5-6 days old.

Stage 5 (, page 1implanted blastocyst): Stage 5 comprises embryos that are implanted to a varying degree but are previllous, i.e., that do not yet show definite chorionic villi. Such embryos are believed to be 7-12 days old. ... An amniotic cavity is found by stage 5. If duplication of the embryo occurs after the differentiation of the amnion, the resulting monozygotic twins should be monochorial and monoamniotic (fig. 5-2). It has been estimated that the frequency of monoamniotic twins among monozygotic twins is about 4 percent (Bulmer, 1970). About once in every 400 monozygotic twin pregnancies, the duplication is incomplete and conjoined ("Siamese") twins (e.g., the second specimen of Shaw, 1932) result. [Back]

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