"Any Human Cell - iPS, Direct Programmed, Embryonic, Fetal or Adult - Can Be Genetically Engineered to Asexually Reproduce New Human Embryos for Purposes of Reproduction ('Infertility')"

Dianne N. Irving
Copyright November, 2011
Reproduced with Permission

[Note: This article is copyrighted and thus must be acknowledged when using its original ideas and resources or quoting from it.]

"It is quite possible that the advances in human biology in the remainder of the twentieth century will be remembered as the most significant scientific achievement of the animal species known as Homo sapiens. But in order to become a part of medical history, parahuman reproduction and human genetic engineering must circumvent the recalcitrance of an antiquated culture. ... A philosophy of reason will define a human being as one which demonstrates self-awareness volition and rationality. Thus it should be recognized that not all men are human. ... A clear definition of humanity in terms of mental acuity, rather than physical appearance, should be encouraged. (Winston L. Duke, "The new biology", in Reason Magazine, "Parahuman Reproduction, Android Cloning, The New Biology, Artificial Synthesis, Genetic Engineering, Brain Transfers", August 1972)1

Any Human Cell - iPS, Direct Programmed, Embryonic, Fetal or Adult - Can Be Genetically Engineered to Asexually Reproduce New Human Embryos for Purposes of Reproduction ("Infertility")

I. Introduction:

Winston Duke wrote those prescient words back in 1972. He was an early California engineer, libertarian, transhumanist and futurist (all international organizations), all seeking "immortality" of one sort or another, and it is well worth reading his article, if only for historical reasons. It explains in good measure "how we got here". This article will address simply "where we are" today.

The use of various types of stem cells for research purposes to make disease "models" in the lab for regenerative medicine and for "therapies" to cure sick patients for diseases is constantly in the news. But what is not getting such wide reporting is the use of pluripotent stem cells (as well as many other types of cells and genetic engineering techniques) for reproductive purposes. It does not seem to be fully realized or evaluated that what can be done for purposes of "therapies" and "research" can also be done for purposes of "reproduction".

A recent UNESCO draft document, although rather vague and deficient in itself, probably does the best job of at least initially identifying and describing some of these new reproductive technologies in relatively simple form, with a few generalized helpful sketches online. As stated up front in that document, the UNESCO International Bioethics Committee (IBC), at least, is aware of various genetic engineering techniques that are already being used for reproductive purposes and sees a need to address their concerns:

"On the basis of reflection and debate on human cloning" held in 2008‐2009, UNESCO's International BIOETHICS Committee met again this past May to June, 2011, to 'explore whether there is any scientific, social or political change that would justify a new initiative [regarding human cloning] at the international level'".2

The IBC identified six such "concerns" in their current draft document:3

Note especially their concerns about "the terminology used in the bioethical debates", that it "is misleading and does not adequately describe the technical procedures used (or potentially to be used) today. ... An in-depth analysis aiming at re-defining this terminology according to the new developments in human embryo research would be highly beneficial." It is precisely the redefining and deconstruction of the accurate science involved that has gone on for decades now that is the subject of the second part of this article. Will UNESCO continue such scientific deconstruction when they redefine the relevant scientific terminology "according to the new developments in human embryo research", or will they finally acknowledge the accurate scientific facts of human embryology? Will the real human embryo, who is a real living human being, disappear further into even more abstract and false definitions such as "pre-embryo", "fertilized egg", "just a bunch of cells", etc.? It is also of concern that laws, regulations and "guidelines" involving only governmental or "public" money might be a step forward - assuming the accurate scientific terms are used -- but obviously those efforts do not affect the use of private funds and facilities for the same research experiments, as will be noted later.

The UNESCO document then proceeds to list some of the kinds of reproductive cloning research techniques already available that they are particularly concerned about. Note that "reproductive cloning" implies implanting an embryo derived by these following genetic engineering techniques into a woman's uterus. None of these human embryos are derived sexually by the process of fertilization, but rather asexually (without the fusion of sperm and oocytes). The techniques that UNESCO lists includes one such technique that is more familiar, somatic cell nuclear transfer (SCNT) (either by "fusion" of an enucleated egg and a somatic cell, or by "direct injection" of a nucleus into an enucleated egg). But they also list three reproductive technologies that are far less familiar, and that can be used in reproductive cloning that supposedly "do not require the destruction of any embryos": (1) tetraploid complementation, (2) direct derivation of sperm and egg cells, and (3) embryo splitting ("twinning).

The purpose of this article, then, is to briefly identify just these last three (of many) genetic engineering techniques that UNESCO itself has identified that can asexually turn mere human "cells" into new whole human embryos for reproductive purposes "without destroying embryos". (Yet it does stretch the imagination to believe that, in the course of this "ethical" research, millions of living human embryos will not be required to be destroyed as the desired "means" to an "ethical" end.) The main issues here are not whether a human embryo is a "person", but whether scientifically the human embryo is a human organism per se, or just a "bunch of cells" - and whether human embryos can be asexually reproduced for reproductive purposes? What follows is simply a brief listing of representative research studies from around the world over the last decade, each one likewise similar to dozens or hundreds that can be found in the scientific literature. This article is especially written so that people can be more aware of "where we really are". The long-documented, constantly updated, and internationally acknowledged accurate scientific facts of human embryology are given towards the end of this article.

II. Scientific Documentation of Recent Stem Cell Research for Reproductive Purposes

A. Pre-history

The current surge in stem cell research really started back in 1993, when researchers Robert Stillman and Jerry Hall of the George Washington University Medical Center in Washington, D.C., cloned human embryos for the first time. They reported their results at a conference of the American Fertility Society.4 The cloning technique they used was "twinning" (see Section B(3) below), and the researchers proposed its use to boost the efficiency of in vitro fertilization (IVF). Stillman noted that, "Twinning could help women who produce very few eggs and thus have trouble getting pregnant, even with the aid of in vitro fertilization". By splitting an early embryo into its constituent totipotent cells, and then allowing the cells to revert to new embryos, doctors could then transfer more than one embryo to the woman's uterus, thus increasing the odds of a successful pregnancy. "Our research is one small step in that direction," Stillman stated.5

Stillman explained the method they used. They took abnormal human embryos left over from IVF, and used a chemical solution to strip the young embryos of their tough outer coating, called the zona pellucida. The shell-like zona pelucida protects the embryo who at this stage has started to divide and consists of from two to eight cells. Next, the researchers carefully separate the individual cells and coat each with an artificial shell. The team created 48 embryos using this technique. The embryos were discarded after developing for 6 days.6 Although the researchers claimed that their research had been approved by the university's IRB and the Ethics Committee of the Medical Center, a university probe denied this, claiming that this cloning research "was attained without full disclosure of facts".7

In 1998, researcher James Thomson, a developmental biologist, and his research team in Wisconsin, were making history. They are best known as the first to report the successful isolation of human embryonic stem cells derived from the inner cell layer of human embryos left over from IVF "infertility" treatments,8 and later for deriving human induced pluripotent stem (iPS) cells by somatic cell nuclear transfer (SCNT) of human somatic cells in 2007.9

Also in 1998, another research team, headed by another developmental biologist, John Gearhart, derived stem cells from cultured human primitive germ cells (these cells are totipotent and eventually differentiate into sperms or oocytes) retrieved from human "fetuses" (5-9 weeks post-fertilization).10 [Note, however that the "embryonic period" is from the beginning of fertilization through 8 weeks; the "fetal period" is from 9 weeks to birth! However, the 1981 federal regulations on the use of human subjects in research mis-defined "fetus" as beginning at "implantation" (which is 5-7 days post-fertilization) and mis-defined "pregnancy" as also beginning at "implantation"].

In 2006, Yamanaka and his team generated induced pluripotent stem cells (iPS cells) from adult mouse fibroblasts,11 and in 2007 he and his team were able to generate iPS cells from human adult fibroblasts.12

The human stem cell roller coaster went into full gear. These human "pluripotent" stem cells, both embryonic and iPS, are now available from various human sources, using various genetic engineering techniques, and can be used for pure research studies (e.g., models of normal and diseased humans), in regenerative medicine for "therapies" to cure diseases in sick patients - and, for reproductive cloning.

But although these cells are capable of differentiating into all three germ layers of an adult, researchers began to identify potential problems. For example, both human embryonic (HE) cells and iPS cells were discovered to have serious genetic abnormalities, as well as changes in the copy number of genes during reprogramming and time in culture.13 iPS cells were found to induce immune rejection responses when injected,14 and seemed to "hold onto their genetic past" more than HE cells.15 Further, iPS cells were found to cause more tumors than HE cells.16 Thus the question remains, how safe are these "stem cells" for human "therapies" to cure diseases in sick patients, much less for women undergoing human reproductive cloning? Even Ian Wilmut, of Dolly the cloned sheep fame, recently noted that human embryonic stem cells and iPS cells, "tend to lead to tumors", thus scientists should spend their time on non-embryonic forms of research, "particularly on a new method called direct reprogramming."17 The jury is still out as to similar problems with even direct programmed cells.18

B. Current Genetic Engineering Techniques That Turn "Cells" Into New Embryos for Reproductive Purposes

One of the genetic engineering techniques for reproductive cloning that the UNESCO document identified was "tetraploid complementation. Examples of the use of tetraploid complementation are documented in the studies that follow.

1. Triploid and Tetraploid Complementation

It is commonly, but mistakenly, thought that "pluripotent" stem cell can't develop into new embryos, even though they do have the capacity to eventually develop into the three major human tissues of an adult. That is because, e.g., such "stem cells" lack the ability to make the outer layer of cells that normal embryos have which eventually form the placenta, umbilical cord, etc.

But what if you take a "pluripotent" stem cell - human embryonic (HE) or reprogrammed iPS cell -- insert it into or fuse it with an artificial "shell" that will function as and substitute for an embryo's outer layer of cells, and then implant it as you would any normal embryo? That is what has recently been accomplished, and it works. It is now possible to genetically engineer such new embryos for reproductive purposes by means of "tetraploid complementation" (TC) which provides a substitute for the outer layer of cells of an embryo that is normally responsible for producing the later placenta, umbilical cord, etc.

"Tetraploid complementation" started off as an "assay" used to test a pluripotent stem cell's "potency" - the ability of a stem cell to differentiate into the three germ layers and tissues of an adult human being. It has now also become a scientific method of reproducing new embryos per se. There are already several variations on the theme. For example, according to NIH:

Scientists studying mouse chimeras (mixing cells of two different animals) noted that fusing two 8-cell embryos produces cells with 4 sets of chromosomes (tetraploid cells) that are biased toward developing into extra-embryonic tissues such as the placenta. The tetraploid cells do not generate the embryo itself; the embryo proper develops from injected diploid stem cells. This tendency has been exploited to test the potency of a stem cell. Scientists begin with a tetraploid embryo. Next, they inject the stem cells to be tested. If the injected cells are pluripotent, then an embryo develops. If no embryo develops, or if the resultant embryo cannot survive until birth, the scientists conclude that the cells were not truly pluripotent.19

Thus what is involved here is that in tetraploid complementation the cells of two mammalian embryos can be combined to form a new multi-ploid embryo. The technique is now used to construct genetically modified organisms, to study the consequences of certain mutations on embryonic development, and in the study of pluripotent stem cells.

To explain further, the tetraploid complement assay can start with producing a tetraploid embryo in which every chromosome exists fourfold. This is done by taking an embryo at the two-cell stage and fusing the two cells of the embryo by applying an electrical current. The resulting tetraploid embryo will continue to divide, and all daughter cells will also be tetraploid. Such a tetraploid embryo can develop normally to the blastocyst stage and will implant into the wall of the uterus. The tetraploid cells can form the extra-embryonic tissue (placenta etc.), but a proper fetus will rarely develop. However, one can now combine such a tetraploid embryo (either at the morula or blastocyst stage) with normal diploid embryonic stem cells (ES) or iPS cells from a different organism. The resulting embryo will then develop normally. The fetus is exclusively derived from the ES cell, while the extra-embryonic tissues are exclusively derived from the tetraploid cells.

Foreign genes or mutations can also be introduced into ES or iPS cells rather easily, and these ES and iPS cells can then be grown into whole animals using the tetraploid complementation "assay". Also, by introducing targeted mutations into the tetraploid cells and/or into the ES or iPS cells, one can study which genes are important for embryonic and fetal development and which ones are important for development of the extra-embryonic tissues. Finally, tetraploid complementation is also used to test whether induced pluripotent stem cells (stem cells artificially produced from differentiated cells, e.g. from skin cells) are as competent as normal embryonic stem cells, or if a viable animal can be produced from an induced pluripotent stem cell using the tetraploid complementation.

** Thus in Nagy et al's 1973 article, ""Derivation of completely cell culture-derived mice from early-passage embryonic stem cells", it was demonstrated that several newly generated mouse embryonic stem (ES) cell lines were able to produce completely ES cell-derived mice by using tetraploid complementation.20

** In 2003, Tam and Rossant's study, "Mouse embryonic chimeras: tools for studying mammalian development", also documented that, "By combining embryonic stem cell technology, molecularly tagged mutations and sensitive cell lineage markers, chimeras can provide invaluable insights into the tissue-specific requirement and the mode of action of many mouse genes."21

** Lan Kang et al confirm in their 2009 report, "iPS cells can support full-term development of tetraploid blastocyst-complemented embryos", that, "This work serves as a proof of principle that iPSCs can in fact generate full-term embryos by tetraploid complementation."22

** Also in 2009 Xiao-yang Zhao et al confirm in their study, "iPS cells produce viable mice through tetraploid complementation" that IPS cells produce viable mice through tetraploid complementation.23

** Naumann describes in his report, ""Production Of tetraploid mouse embryos by electrofusion", the use of tetraploid complementation as a means of producing new embryos that are chimeras (have genomes from different species). For example, combining tetraploid blastocysts with either embryonic stem (ES) cells or diploid embryos has become an established technique for creating mouse chimeras. In both methods the tetraploid cells are not able to contribute to the embryo itself, but instead create the primitive endoderm derivatives and the trophectoderm. He also explains how complex phenotypes can be produced by generating embryos carrying mutations of multiple genes.24

** Bertolotti praises these techniques in his study, "Fully pluripotent iPS cells: Mouse tetraploid complementation, ethical human ES-like cells and reproductive cloning ban". However, he begrudgingly acknowledges some concerns: "Such a genesis of fertile adult "iPS" mice could revive the roguish temptation of banned human reproductive cloning currently doomed by the failure of human cell reprogramming by somatic cell nuclear transfer (SCNT)."25

** Denker also acknowledges concerns in his study, "Induced pluripotent stem cells: how to deal with the developmental potential", but seems to indicate that it is not a problem:

"Recent developments in research on embryonic stem cells and induced pluripotent stem cells suggest that potentiality of cells should be a new focus in stem cell research ethics and policy. Successful reconstitution of viable embryos from induced pluripotent stem cells using tetraploid complementation has been reported and indicates a way for direct cloning of individuals from these cells. This together with recent observations on gastrulation and pattern formation processes in cultures of embryonic stem cells has considerable ethical relevance after the advent and worldwide spread of induced pluripotent stem cell technology. Available knowledge of the molecular basis of mammalian embryology now makes it possible to envisage ways to deal technically with the ethical dilemma of stem cell potentiality."26

On the other hand, some have tried to alert the public about the ethical and scientific implications of producing new embryos by means of tetraploid complementation.

** For example, in 2009 the Chinese researchers Shi V. Liu, Ke-Ming Cui, and Wei-Kang Zhang attempted to voice their concerns by letter to the journals Nature and Science. The publication of both letters was denied, so these researchers published them elsewhere as "Open Letters".

In their first letter to Nature, "Why Do We Need "Fully Pluripotent" iPS Cells?",27 these researchers questioned the scientific and ethical value of obtaining "fully pluripotent" iPSCs (induced pluripotent stem cells) "using an embryo-sacrificing reproductive cloning technique involving tetraploid complementation, as presented already in some studies published in Nature. Scientifically. For example, they referred to such research as "over-reprogramming" and "over-extension of iPS technology", questioning why researchers should rush into reproductive cloning using some incorrectly programmed stem cells or "man-made cancer stem cells", and pointing out that studies have shown that iPS cells are already more aggressively tumorgenic than HECS's. The researchers also noted that such research would violate NIH Guidelines for funding. Further, they cautioned against following "an extreme point of regenerative medicine" that thinks it ideal to find a "fully pluripotent" stem cell so that a damaged or aged body can be regenerated completely from just one such cell of the same genetic identity - calling such "ideals" a form of "extremely selfish immortality". They were also concerned about the effect that such "extremes" would have on the reputation of good researchers, as well as concerned that once this success in non-human animals is translated into human studies, these dangerous techniques would be applied to humans in clinical trials or abused as biological weapons.

In their second letter to Science, "iPS mice: Passing acid test or breaking basic ethics?",28 the researchers reiterated many of the same concerns as in their letter to Nature. They also took issue with the whole concept of tetraploid complementation as "the gold standard" or "acid test" for "full pluripotency", i.e., determining whether a "pluripotent" cell is capable of "forming an entire viable organism. Quite interestingly, they point out one of the common points of erroneous scientific disinformation long used in most of this research, i.e., the insistence that embryonic cells at the very early two-cell stage of an embryo, would not develop into any embryos on their own, that they were only "pluripotent". (Such a claim directly contradicts the research identified by UNESCO as "embryo splitting", and that is addressed later in this article.)

The researchers point out that "it is an enigma why a "full pluripotency" cell would even require two "disabled" embryos to support their "full" development. It is also puzzling why successful complementation from "full pluripotency" cells often yields only less than 100% chimaerism." They suggest that the "gold standard" is not 100% gold, asking why a non-destructive and simple and transparent "gold standard" - implanting noting but just the testing cell(s) - has not been used for testing the "full pluripotency" of iPSCs:

Ethically, it is difficult to imagine that a method requiring destruction of at least two embryos per standard or original complementation assay would be considered as more "ethical" than a method such as SCNT which utilizes just one egg per nuclear transfer. Considering an egg is just half of a potential future life, we simply cannot accept the fact that damaging two living embryos for generating a potential life is any more ethical practice, regardless whether or not the cells contributing the "full pluripotency" are ethical or not. If iPSCs are already ethical cells aimed for therapeutic cloning, please do not pass them through an unethical reproductive cloning to demonstrate their "full pluripotency" which is not even needed for normal partial body regenerative medicine. We should not destroy more whole lives for obtaining some parts for treating diseases or extending lifespan!29

They conclude by asking, "why should we jump over this mice embryo-killing 'success' in passing an 'acid test' which, proved in principle, basic ethics can be easily broken intentionally or accidentally".30

** Indeed, others have also recently reaffirmed the increased tendency of iPS cells to form tumors, as Tong et al describe in their study, "Mice generated from tetraploid complementation competent iPS cells show similar developmental features as those from ES cells but are prone to tumorigenesis". As the authors noted, "Previously, we and others have demonstrated that fully reprogrammed iPSCs were capable of producing full-term mice via the tetraploid complementation method, yet a thorough postnatal development evaluation of iPS mice is still lacking."31

** Another verification of the tumorgenicity of iPS cells comes from Shi-Lung Lin in his article, "Concise Review: Deciphering the Mechanism Behind Induced Pluripotent Stem Cell Generation". Hopefully, regenerative medicine using pluripotent or multipotent stem cells holds a great promise in developing therapies for treating developmental abnormalities, degenerative disorders, and aging-related illness. However, supply and safety of the stem cells are two major problems with today's regenerative medicine. Recent development of induced pluripotent stem cells (iPSCs) has overcome the supply shortages by allowing the reprogramming of patients' body cells to embryonic stem cell (ESC)-like pluripotent cells. Still, the potential tumorigenicity of iPSCs remains as an obstacle.32

** David Cyranoski also expressed serious concerns about the mice studies using iPS cells and tetraploid complementation to reproduce whole new embryos. In his 2009 article, "Mice made from induced stem cells: Technical feat shows that the different route to stem cells can indeed make a full mammal body",33 Cyranoski states the obvious, that the recent studies do answer " a lingering question about the developmental potential of the cells." Since Shinya Yamanaka of Kyoto University in Japan created the first iPS cells in 2006, "researchers have wondered whether they could generate an entire mammalian body from iPS cells, as they have from true embryonic stem cells. ... Experiments reported online this week suggest that, at least for mice, the answer is yes." And he also cautions that this would essentially be a new way to clone adult mammals - reprogramming DNA from an adult and generating a genetically identical individual. As a potentially easier method that produces fewer abnormalities than conventional cloning, he cautions that "it might evoke interest among mavericks as a tool for human cloning."

** Some secular bioethicists have also voiced concern, writing a lengthy and detailed report that includes several charts of relevant legislation within the United States as well as internationally. The point of their article is best summed up with their statement: "The use of iPSCs and tetraploid complementation for human reproductive cloning would raise profound ethical objections. Professional standards and laws that ban human reproductive cloning by somatic cell nuclear transfer should be revised to also forbid it by other methods, such as iPSCs via tetraploid complementation."34 (emphases added)

** More recently Andrew Pollack again cautions on the use of genetic engineering techniques to derive stem cells for therapeutic use with sick patients, and their application to reproductive cloning: "But the potential problem with iPS cells might make some scientists take another look at making patient-specific tissues by creating an embryo from a patient's cell through so-called therapeutic cloning. That approach... is controversial because the same technique might also be used to create a baby."35

Next Page: 2. Artificial Sex Gametes Derived From iPS, Human Embryonic, and Adult Cells
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