Interspecies Chimeras and the Generation of Humanized Organs
2019; Lippincott Williams & Wilkins; Volume: 124; Issue: 1 Linguagem: Inglês
10.1161/circresaha.118.314189
ISSN1524-4571
AutoresDaniel J. Garry, Mary G. Garry,
Tópico(s)Biomedical Ethics and Regulation
ResumoHomeCirculation ResearchVol. 124, No. 1Interspecies Chimeras and the Generation of Humanized Organs Free AccessArticle CommentaryPDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessArticle CommentaryPDF/EPUBInterspecies Chimeras and the Generation of Humanized Organs Daniel J. Garry and Mary G. Garry Daniel J. GarryDaniel J. Garry Daniel J. Garry, MD, PhD, Paul and Sheila Wellstone Muscular Dystrophy Center, University of Minnesota, 2231 6th St SE, CCRB 4–146, Minneapolis, MN 55455, Email E-mail Address: [email protected] From the Department of Medicine, Paul and Sheila Wellstone Muscular Dystrophy Center, University of Minnesota, Minneapolis. and Mary G. GarryMary G. Garry Correspondence to Mary G. Garry, PhD, Paul and Sheila Wellstone Muscular Dystrophy Center, University of Minnesota, 2231 6th St SE, CCRB 4–146, Minneapolis, MN 55455, Email E-mail Address: [email protected] From the Department of Medicine, Paul and Sheila Wellstone Muscular Dystrophy Center, University of Minnesota, Minneapolis. Originally published3 Jan 2019https://doi.org/10.1161/CIRCRESAHA.118.314189Circulation Research. 2019;124:23–25End-stage or advanced heart failure is a lethal disease. The only curative therapy is solid organ transplantation, but it is limited because of the scarcity of organs. Alternative strategies are being developed including the use of interspecies chimeras to generate humanized hearts in gene-edited pigs. These interspecies chimeras will provide a unique research model and impact the treatment of chronic, end-stage disease such as heart failure.Heart failure remains a deadly epidemic. Despite the advances by pharmacological and device therapies, the mortality of patients with heart failure with reduced ejection fraction remains ≈50% at 5 years after diagnosis. The only curative therapy for advanced heart failure with reduced ejection fraction is orthotopic heart transplantation.1 Although estimates suggest that upward of 100 000 Americans would benefit from heart transplantation, only ≈2200 adults in the United States receive an allograft.1,2 The disparity between those who need this life-saving technology (ie, a heart transplant) and those who receive such a graft is what drives the innovation for new therapies.Xenotransplantation as a strategy to provide an alternative source of organs, such as a heart, has had a rich and long-standing history.1,3 The first cardiac xenotransplant was undertaken by Dr James D. Hardy and his team at the University of Mississippi Medical Center (January 24, 1964) where they transplanted a nonhuman primate (ie, chimpanzee) heart into a 68-year-old patient—although the donor heart was successfully implanted, the patient expired after ≈90 minutes of support.3 Twenty years later, Dr Leonard L. Bailey at Loma Linda University Medical Center transplanted a baboon heart into an infant (AKA Baby Fae) with hypoplastic left heart syndrome. Baby Fae survived for 21 days before her death. The legendary cardiac transplant pioneer, Norman Shumway, often quipped that "The future of transplantation is xenotransplantation and it will always be the future," suggesting that the immunologic barriers to xenotransplantation would never be fully overcome for it to be a broadly used therapy.4 Nevertheless, recent advances in immunobiology and, separately, the ability to gene edit animals suggest that the barriers for xenotransplantation may be waning.5,6An alternative approach relies on the use of technologies such as blastocyst complementation to engineer an interspecies chimera.7 The overall strategy is to establish a host which could be genetically modified that lacks a single or multiple lineage(s), thus providing a permissive niche for a donor stem cell to contribute to the absent lineage in the recipient (host) embryo. This would provide the donor stem cell a competitive advantage as the host would be unable to form the lineage of interest (ie, the lineage that had been deleted).7 The feasibility of this strategy has been amplified with the derivation and use of human induced pluripotent stem cells (hiPSCs), which can be readily obtained from a patient who would desire or need the organ. In this fashion, the patient becomes both the donor and eventually the recipient. Furthermore, hiPSCs lack the ethical issues associated with human embryonic stem cells.8,9 In addition to the hiPSCs, the ability to use gene-editing technologies such as CRISPR/Cas9 has further accelerated these research initiatives.8,9Previous studies have used blastocyst complementation to deliver GFP (green fluorescent protein)-labeled mouse embryonic stem cells (ESCs) into the Pdx1 (the master regulator for the pancreas) deficient rat blastocyst to produce a mouse-rat chimera which resulted in a rat-sized pancreas that was a derivative of the mouse pluripotent stem cells (ie, a GFP expressing pancreas).10 Importantly, these chimeric pancreata were physiologically functional and subsequent transplantation into a diabetic mouse normalized and maintained glycemic control for more than a year in the absence of immunosuppression.10 These studies emphasized the important role of the host environment, which defined the size of the chimeric organ (the mouse is ≈1/10th the size of a rat) in spite of the fact that the chimeric pancreas was produced entirely from the mouse ESCs. Furthermore, these studies provided a platform for interspecies chimeras using hiPSCs.Studies have examined the capacity of hiPSCs to survive and differentiate after their delivery into mouse blastocysts. hiPSCs delivered into the developing mouse embryo (at the gastrulation stage) and differentiated in vitro demonstrated chimeric competency as the hiPSCs proliferated and differentiated or contributed to all 3 germ layer derivatives.11 These studies emphasized the importance of matching similar developmental stages of donor stem cells with the host environment (ie, host embryo). Although human-mouse chimeras provided a platform for the mechanistic dissection of factors that impact cell survival, proliferation, differentiation, and viability of the chimeric embryo, ultimately it will be necessary to have a large animal as a host to generate appropriately sized humanized organs. Sheep and pigs are both possible hosts for the generation of human-sized organs.4 Sheep have been successfully used in biomedical research as it is a readily available large animal that has been extensively evaluated in the study of embryonic development. Pigs have served as a source of insulin and bioprosthetic heart valves for humans for more than a half a century.12 Pigs have a relatively short gestational period (114–120 days), and gene mutations have been achieved using somatic cell nuclear transfer technology or directly using CRISPR/Cas9 gene editing in the zygote. Therefore, genetically mutated pig embryos could be generated that lack an entire lineage(s), cardiac, for example, thereby establishing a niche for the human stem cells to rescue the mutant embryo by producing a humanized heart.7 In our respective laboratories, we have used the pig as our large animal model of choice. The pig is a well-established physiological model, somatic cell nuclear transfer technology has been used extensively in this species, its postnatal growth is rapid, and the swine (pig) industry nationwide and worldwide is extensive.7,13 In our experience, the somatic cell nuclear transfer techniques are relatively efficient in the pig model, and in vitro chimeric embryos can be analyzed pre-gastrulation. Furthermore, in vitro studies of the interspecies chimeras offer the opportunity to promote efficiencies in an attempt to engineer a viable chimeric pig.Download figureDownload PowerPointFigure 1. A schematic overview that highlights the use of human induced pluripotent stem cells (hiPSCs) from patients in combination with blastocyst complementation to generate a humanized heart.In addition to the use for xenotransplantation, these chimeric large animals would also be a novel preclinical research tool used in drug discovery programs, toxicology studies, device development, or as a one-of-a-kind educational model.7 As a proof of principle study, Wu et al14 delivered 8 to 10 human pluripotent stem cells into wild-type porcine blastocysts and following implantation harvested the chimeric embryos at embryonic day 28 and found differentiated human derivative cells but noted that the efficiency was markedly reduced. The major hurdles for the efficient generation of chimeric organs appear to be, in part, because of the evolutionary divergence between donor cells and the host embryonic environment.7,10,14 Other major hurdles include the matching of the developmental stage of the donor cells and the host embryo, the survival of the donor cells once delivered into the host embryo, and the selective contribution of the donor cells to the absent lineage(s) in the host embryo without significant contribution to other lineages.Every major innovation is accompanied by ethical issues, and these human-large animal chimeric initiatives will be no exception. There are several ethical issues associated with this technology, but perhaps one of the major issues is the extent of the hiPSC contribution to other lineages in the large animal host such as the contribution to neural and germ lineages.15 Use of neural-specific suicide inhibitors (to preclude contribution to the developing cerebral cortex) and a moratorium on the reproductive mating of the chimeric animals are possible solutions to these issues. Nevertheless, these technologies have the potential to, 1 day, provide our patients with an unlimited number of organs for transplantation and have the opportunity to democratize the organ donation program—such that every patient who needs a new heart could serve as the donor (hiPSCs) and ultimately the recipient of a humanized heart generated in a gene-edited pig model.Download figureDownload PowerPointFigure 2. Blastocyst complementation to generate chimeric embryos. Porcine embryonic fibroblasts can be gene edited and the nuclei used for somatic cell nuclear transfer (SCNT) of porcine oocytes. GFP (green fluorescent protein)-labeled mouse embryonic stem cells (2 cells) were delivered into the cloned porcine morula and the chimeric embryo developed to the blastocyst stage in vitro.In summary, advanced technologies such as hiPSCs, gene editing, and somatic cell nuclear transfer have the potential to provide revolutionary therapies for chronic end-stage diseases. In our estimation, the pursuit of these human:pig chimeras using blastocyst complementation should be pursued with the goal of engineering humanized organs for research and as a source for organ transplantation. Although issues (research-related and ethical issues) may emerge during the course of these studies, new translational research models and additional sources of transplantable organs are urgently needed for the treatment of chronic, terminal diseases such as advanced heart failure.Sources of FundingThis work was supported by grants from Regenerative Minnesota Medicine, the American Heart Association, and the Department of Defense (11763537).DisclosuresThe authors are cofounders of NorthStar Genomics.FootnotesThe opinions expressed in this article are not necessarily those of the editors or of the American Heart Association.*These authors contributed equally to this article.Correspondence to Mary G. Garry, PhD, Paul and Sheila Wellstone Muscular Dystrophy Center, University of Minnesota, 2231 6th St SE, CCRB 4–146, Minneapolis, MN 55455, Email [email protected]eduDaniel J. Garry, MD, PhD, Paul and Sheila Wellstone Muscular Dystrophy Center, University of Minnesota, 2231 6th St SE, CCRB 4–146, Minneapolis, MN 55455, Email [email protected]eduReferences1. Garry DJ, Goetsch SC, McGrath AJ, Mammen PP. Alternative therapies for orthotopic heart transplantation.Am J Med Sci. 2005; 330:88–101.CrossrefMedlineGoogle Scholar2. Vega JD, Moore J, Murray S, Chen JM, Johnson MR, Dyke DB. Heart transplantation in the United States, 1998-2007.Am J Transplant. 2009; 9:932–941. doi: 10.1111/j.1600-6143.2009.02568.xCrossrefMedlineGoogle Scholar3. Cooper DKC, Ekser B, Tector AJ. A brief history of clinical xenotransplantation.Int J Surg. 2015; 23:205–210. doi: 10.1016/j.ijsu.2015.06.060CrossrefMedlineGoogle Scholar4. Cooper DKC. A brief history of cross-secies organ transplantation.Proc Bayl Univ Med Cent. 2012; 25:49–57.CrossrefMedlineGoogle Scholar5. Ekser B, Li P, Cooper DKC. 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