Generation of human endothelium in pig embryos deficient in ETV2

Satyabrata Das; Naoko Koyano-Nakagawa; Ohad Gafni; Geunho Maeng; Bhairab N Singh; Tara Rasmussen; Xiaoyan Pan; Kyung-Dal Choi; Daniel Mickelson; Wuming Gong; Pruthvi Pota; Cyprian V Weaver; Stefan Kren; Jacob H Hanna; Demetris Yannopoulos; Mary G Garry; Daniel J Garry

doi:10.1038/s41587-019-0373-y

Generation of human endothelium in pig embryos deficient in ETV2

Satyabrata Das, Naoko Koyano-Nakagawa, Ohad Gafni, Geunho Maeng, Bhairab N Singh, Tara Rasmussen, Xiaoyan Pan, Kyung-Dal Choi, Daniel Mickelson, Wuming Gong, Pruthvi Pota, Cyprian V Weaver, Stefan Kren, Jacob H Hanna, Demetris Yannopoulos, Mary G Garry, Daniel J Garry

Research output: Contribution to journal › Letter › peer-review

73 Scopus citations

Abstract

The scarcity of donor organs may be addressed in the future by using pigs to grow humanized organs with lower potential for immunological rejection after transplantation in humans. Previous studies have demonstrated that interspecies complementation of rodent blastocysts lacking a developmental regulatory gene can generate xenogeneic pancreas and kidney1,2. However, such organs contain host endothelium, a source of immune rejection. We used gene editing and somatic cell nuclear transfer to engineer porcine embryos deficient in ETV2, a master regulator of hematoendothelial lineages3-7. ETV2-null pig embryos lacked hematoendothelial lineages and were embryonic lethal. Blastocyst complementation with wild-type porcine blastomeres generated viable chimeric embryos whose hematoendothelial cells were entirely donor-derived. ETV2-null blastocysts were injected with human induced pluripotent stem cells (hiPSCs) or hiPSCs overexpressing the antiapoptotic factor BCL2, transferred to synchronized gilts and analyzed between embryonic day 17 and embryonic day 18. In these embryos, all endothelial cells were of human origin.

Original language	English (US)
Pages (from-to)	297-302
Number of pages	6
Journal	Nature biotechnology
Volume	38
Issue number	3
DOIs	https://doi.org/10.1038/s41587-019-0373-y
State	Published - Mar 2020

Bibliographical note

Funding Information:
This work was supported by a grant from the Department of Defense (grant no. 11763537 to D.J.G.) and Minnesota Regenerative Medicine (to D.J.G.). We thank J. Dutton (University of Minnesota) and G. Daley (Harvard Medical School) for providing hiPSC lines. We thank C. Chapman for assisting with the hiPSC culture studies, E. Skie for genetic analyses, Y. Ren and C. Walter for the FACS analysis, and B. Coffin, A. Arnason, L. Meisner and D. Ly for morphological analyses. We thank A. Caplan for critical comments during the course of these studies. We acknowledge the Mouse Genetics Laboratory, Veterinary Diagnostic Laboratory and the University Imaging Center at the University of Minnesota, NorthStar Genomics, Recombinetics, DeSoto Biosciences, grafikalabs (http://grafikalabs.com/) and MOFA Global for their technical assistance. We also thank R. Prather and the National Swine Resource and Research Center at the University of Missouri for providing technical training and assistance (U42 OD011140).

Publisher Copyright:
© 2020, The Author(s), under exclusive licence to Springer Nature America, Inc.

PubMed: MeSH publication types

Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Journal Article
Research Support, N.I.H., Extramural

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Das, S., Koyano-Nakagawa, N., Gafni, O., Maeng, G., Singh, B. N., Rasmussen, T., Pan, X., Choi, K.-D., Mickelson, D., Gong, W., Pota, P., Weaver, C. V., Kren, S., Hanna, J. H., Yannopoulos, D., Garry, M. G., & Garry, D. J. (2020). Generation of human endothelium in pig embryos deficient in ETV2. Nature biotechnology, 38(3), 297-302. https://doi.org/10.1038/s41587-019-0373-y

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abstract = "The scarcity of donor organs may be addressed in the future by using pigs to grow humanized organs with lower potential for immunological rejection after transplantation in humans. Previous studies have demonstrated that interspecies complementation of rodent blastocysts lacking a developmental regulatory gene can generate xenogeneic pancreas and kidney1,2. However, such organs contain host endothelium, a source of immune rejection. We used gene editing and somatic cell nuclear transfer to engineer porcine embryos deficient in ETV2, a master regulator of hematoendothelial lineages3-7. ETV2-null pig embryos lacked hematoendothelial lineages and were embryonic lethal. Blastocyst complementation with wild-type porcine blastomeres generated viable chimeric embryos whose hematoendothelial cells were entirely donor-derived. ETV2-null blastocysts were injected with human induced pluripotent stem cells (hiPSCs) or hiPSCs overexpressing the antiapoptotic factor BCL2, transferred to synchronized gilts and analyzed between embryonic day 17 and embryonic day 18. In these embryos, all endothelial cells were of human origin.",

author = "Satyabrata Das and Naoko Koyano-Nakagawa and Ohad Gafni and Geunho Maeng and Singh, {Bhairab N} and Tara Rasmussen and Xiaoyan Pan and Kyung-Dal Choi and Daniel Mickelson and Wuming Gong and Pruthvi Pota and Weaver, {Cyprian V} and Stefan Kren and Hanna, {Jacob H} and Demetris Yannopoulos and Garry, {Mary G} and Garry, {Daniel J}",

note = "Funding Information: This work was supported by a grant from the Department of Defense (grant no. 11763537 to D.J.G.) and Minnesota Regenerative Medicine (to D.J.G.). We thank J. Dutton (University of Minnesota) and G. Daley (Harvard Medical School) for providing hiPSC lines. We thank C. Chapman for assisting with the hiPSC culture studies, E. Skie for genetic analyses, Y. Ren and C. Walter for the FACS analysis, and B. Coffin, A. Arnason, L. Meisner and D. Ly for morphological analyses. We thank A. Caplan for critical comments during the course of these studies. We acknowledge the Mouse Genetics Laboratory, Veterinary Diagnostic Laboratory and the University Imaging Center at the University of Minnesota, NorthStar Genomics, Recombinetics, DeSoto Biosciences, grafikalabs (http://grafikalabs.com/) and MOFA Global for their technical assistance. We also thank R. Prather and the National Swine Resource and Research Center at the University of Missouri for providing technical training and assistance (U42 OD011140). Publisher Copyright: {\textcopyright} 2020, The Author(s), under exclusive licence to Springer Nature America, Inc.",

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AU - Das, Satyabrata

AU - Koyano-Nakagawa, Naoko

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AU - Maeng, Geunho

AU - Singh, Bhairab N

AU - Rasmussen, Tara

AU - Pan, Xiaoyan

AU - Choi, Kyung-Dal

AU - Mickelson, Daniel

AU - Gong, Wuming

AU - Pota, Pruthvi

AU - Weaver, Cyprian V

AU - Kren, Stefan

AU - Hanna, Jacob H

AU - Yannopoulos, Demetris

AU - Garry, Mary G

AU - Garry, Daniel J

N1 - Funding Information: This work was supported by a grant from the Department of Defense (grant no. 11763537 to D.J.G.) and Minnesota Regenerative Medicine (to D.J.G.). We thank J. Dutton (University of Minnesota) and G. Daley (Harvard Medical School) for providing hiPSC lines. We thank C. Chapman for assisting with the hiPSC culture studies, E. Skie for genetic analyses, Y. Ren and C. Walter for the FACS analysis, and B. Coffin, A. Arnason, L. Meisner and D. Ly for morphological analyses. We thank A. Caplan for critical comments during the course of these studies. We acknowledge the Mouse Genetics Laboratory, Veterinary Diagnostic Laboratory and the University Imaging Center at the University of Minnesota, NorthStar Genomics, Recombinetics, DeSoto Biosciences, grafikalabs (http://grafikalabs.com/) and MOFA Global for their technical assistance. We also thank R. Prather and the National Swine Resource and Research Center at the University of Missouri for providing technical training and assistance (U42 OD011140). Publisher Copyright: © 2020, The Author(s), under exclusive licence to Springer Nature America, Inc.

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N2 - The scarcity of donor organs may be addressed in the future by using pigs to grow humanized organs with lower potential for immunological rejection after transplantation in humans. Previous studies have demonstrated that interspecies complementation of rodent blastocysts lacking a developmental regulatory gene can generate xenogeneic pancreas and kidney1,2. However, such organs contain host endothelium, a source of immune rejection. We used gene editing and somatic cell nuclear transfer to engineer porcine embryos deficient in ETV2, a master regulator of hematoendothelial lineages3-7. ETV2-null pig embryos lacked hematoendothelial lineages and were embryonic lethal. Blastocyst complementation with wild-type porcine blastomeres generated viable chimeric embryos whose hematoendothelial cells were entirely donor-derived. ETV2-null blastocysts were injected with human induced pluripotent stem cells (hiPSCs) or hiPSCs overexpressing the antiapoptotic factor BCL2, transferred to synchronized gilts and analyzed between embryonic day 17 and embryonic day 18. In these embryos, all endothelial cells were of human origin.

AB - The scarcity of donor organs may be addressed in the future by using pigs to grow humanized organs with lower potential for immunological rejection after transplantation in humans. Previous studies have demonstrated that interspecies complementation of rodent blastocysts lacking a developmental regulatory gene can generate xenogeneic pancreas and kidney1,2. However, such organs contain host endothelium, a source of immune rejection. We used gene editing and somatic cell nuclear transfer to engineer porcine embryos deficient in ETV2, a master regulator of hematoendothelial lineages3-7. ETV2-null pig embryos lacked hematoendothelial lineages and were embryonic lethal. Blastocyst complementation with wild-type porcine blastomeres generated viable chimeric embryos whose hematoendothelial cells were entirely donor-derived. ETV2-null blastocysts were injected with human induced pluripotent stem cells (hiPSCs) or hiPSCs overexpressing the antiapoptotic factor BCL2, transferred to synchronized gilts and analyzed between embryonic day 17 and embryonic day 18. In these embryos, all endothelial cells were of human origin.

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JF - Nature biotechnology

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ER -

Generation of human endothelium in pig embryos deficient in ETV2

Abstract

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Generation of human endothelium in pig embryos deficient in ETV2

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