In vitro differentiation of cultured human pluripotent stem cells (hPSCs) constitutes a promising tool for the generation of specific cell types useful for cell replacement therapies. Despite rapid progress over the last decade, numerous challenges encompassing cell-production quantity, quality and safety remain to be addressed before full-fledged clinical application becomes a reality. Moreover, generation of functional organs from hPSCs, a possible alternative to meet the rising demands for organ donors, remains, to date, improbable. Overall, these shortcomings are due to our limited understanding of molecular and genetic events underlying in vivo tissue/organ generation. Throughout evolution, nature has evolved a sophisticated and robust intrinsic genetic program that works seamlessly with extrinsic developmental niches in a choreographed spatiotemporal manner to enable embryonic cells to commit to specific cell lineages and be organized into higher-order tissue architectures. Blastocyst complementation is based on emptying these developmental niches by altering the expression of gene(s) critical for a specific cell/tissue type and use donor chimeric-competent PSCs to colonize the vacant niche and allow for the generation of tissues/organs of donor origin. With the advent of CRISPR/CAS9-mediated genome engineering technologies, gene knockout animals can be generated with high efficiency by co-injection of Cas9 mRNA and sgRNAs directly into zygotes. Concurrently, chimeric- competent/nave hPSCs have seen several promising developments. By combining the strength of zygote genome editing and chimeric-competent/nave hPSCs, this proposal seeks to establish a novel interspecific blastocyst complementation system for the generation of functional human organs in a large animal host, the pig. This offers an attractive platform for realizing hPSCs' full potenial towards generating clinically relevant cells/tissues/organs. If successful, this approac
Despite substantial progress in the last decade; no human pluripotent cell-based therapies have transitionedfrom experimental to clinical practice due to numerous remaining challenges. We propose to generatefunctional human organs and tissues in vivo in a large animal host; the pig; by using a novel interspecificblastocyst complementation system that combines na ve human iPSCs with recently discovered gene-editingtechnologies. If successful; this approach will lead to a paradigm shift in regenerative medicine and help toovercome the shortage of tissues and organs available for donation.
| Bogliotti, Yanina Soledad; Wu, Jun; Vilarino, Marcela et al. (2018) Efficient derivation of stable primed pluripotent embryonic stem cells from bovine blastocysts. Proc Natl Acad Sci U S A 115:2090-2095 |
| Wu, Jun; Vilarino, Marcela; Suzuki, Keiichiro et al. (2017) CRISPR-Cas9 mediated one-step disabling of pancreatogenesis in pigs. Sci Rep 7:10487 |
