Current Stem Cell Research & Therapy, 2009, 4, 81-86 81 1574-888X/09 $55.00+.00 © 2009 Bentham Science Publishers Ltd. Alternative Strategies for the Derivation of Human Embryonic Stem Cell Lines and the Role of Dead Embryos Svetlana Gavrilov 1 , Virginia E. Papaioannou 1 and Donald W. Landry* ,2 1 Department of Genetics and Development, College of Physicians and Surgeons of Columbia University, 2 Department of Medicine, Division of Experimental Therapeutics, College of Physicians and Surgeons of Columbia University, NY, USA Abstract: The therapeutic potential for human embryonic stem cells (hESC) drives intense public and scientific interest. However, the classical approach for derivation of hESC entails the destruction of human embryos. Controversial ethical issues and correspondingly restrictive federal policies in many countries have prompted the development of alternative approaches for the isolation of hESC. Here, several different strategies are discussed with a focus on the harvesting of live hESC from dead embryos. Keywords: Arrested human embryos, ES cell derivation, alternative approaches, human embryonic stem cells, hESC. INTRODUCTION Since their first isolation a decade ago [1] human embry- onic stem cells (hESC) have drawn intense attention. Through their dual capacity for self-renewal and for differen- tiation into lineages of all three embryonic germ layers, hESC pose as excellent candidates for cell replacement ther- apy and a valuable platform for the discovery of drugs and the study of human development and disease [2-5]. How- ever, as hESC lines are conventionally derived from the in- ner cell mass (ICM) of human preimplantation embryos (see Fig. 1), the arena surrounding their derivation is charged with controversy [6-9]. The ethical, moral and political status of nascent human life is a matter of active debate and many regard the preim- plantation embryo as requiring protection. As a result, some countries have opted to prohibit aspects of hESC research [9- 11]. In the US, federal policy on hESC research is qualifi- edly restrictive, i.e. the use of federal funding for the crea- tion of new hESC lines by means that destroy human em- bryos is forbidden (see Box 1) [12, 13]. Furthermore, US scientists share the same restrictions with scientists in Italy, Austria, Ireland, Poland, Slovakia, Lithuania, Norway and Germany, among European countries [10]. Such ethical re- strictions are characterized by some as impediments to pro- gress. With the moratorium on derivation of new hESC im- posed by some governments, the quality and potential of existing ‘approved’ hESC has come under closer scrutiny. The existing hESC lines, including those available to US scientists (derived before 2001- see Box 1) exhibit variable phenotypic and growth characteristics as they were derived and propagated under different culture conditions using mostly animal feeder layers for support (predominantly mouse embryonic fibroblasts) [14, 15]. Because the utility of existing lines varies significantly, new lines, stable and fully characterized, would be useful. Until recently, extensive *Address correspondence to this author at the Division of Experimental Therapeutics, Collage of Physicians and Surgeons of Columbia University, New York, NY, 10032, USA; Tel: (212) 305-5838; Fax: (212) 305-3574; E-mail: dwl1@columbia.edu characterization of existing hESC lines was lacking. A com- parative study initiated by the International Stem Cell Initia- tive attempted to create a consensus for full characterization by examining many hESC lines, established and maintained in different laboratories worldwide [16]. It was proposed that expression of the genes NANOG, TDGF, POU5F1/OCT4, GABRB3, GDF3 and DNMT3B should be used as a core of markers to define undifferentiated hESC and establish hESC identity [16]. Few of the existing NIH-approved lines meet the highest standards for quality. Another consideration arguing for the creation of new lines is the need for clinical grade hESC lines that could eventually bring stem cell therapy to the bedside. None of the hESC lines available to US researchers match clinical grade criteria as they have been directly or indirectly exposed to animal material during their derivation or propagation in vitro [17]. These potentially xeno- contaminated lines are judged unsuitable for transplantation because of the risk of zoonosis transmitted by animal pathogens and potential activation of animal retroviruses. For the generation of clinical grade hESC, all steps for derivation must avoid the use of animal products and/or components, i.e. derivation must proceed under xeno-free conditions [14, 17]. Only recently human feeder or feeder free cultures have been employed [18, 19]. It is currently acceptable to derive novel hESC lines for possible therapeu- tic purpose using human feeder cells and media with sup- plements of human origin [17]. It remains the ultimate goal to develop clinical grade, xeno-free lines under stringent and controllable conditions. Thus, additional hESC lines are needed and their deriva- tion is an inevitable precondition for the use of hESC in re- generative medicine. Does the promise of a cure justify fur- ther destruction of human embryos? Does the end justify the means? These questions pose a problem not just for re- searchers and regulators but also the general public and, most specifically, the patients who would be offered thera- pies of potentially troubling origins. In the contest between the imperative to respect human life and the mandate to re- duce human suffering, alternative strategies have been pro- posed to derive hESC lines or their equivalent while sparing human embryos. The major alternative approaches are as-