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Pigs share many physiological, biochemical, and anatomical similarities with humans and have emerged as valuable large animal models for biomedical research. Considering the advantages in immune system resemblance, suitable size, and longevity for clinical practical and monitoring purpose, SCID pigs bearing dysfunctional RAG could serve as important experimental tools for regenerative medicine, allograft and xenograft transplantation, and reconstitution experiments related to the immune system. In this study, we report the generation and phenotypic characterization of RAG1 and RAG2 knockout pigs using transcription activatorlike effector nucleases. Porcine fetal fibroblasts were genetically engineered using transcription activator-like effector nucleases and then used to provide donor nuclei for somatic cell nuclear transfer. We obtained 27 live cloned piglets; among these piglets, 9 were targeted with biallelic mutations in RAG1,3 were targeted with biallelic mutations in RAG2, and 10 were targeted with a monoallelic mutation in RAG2. Piglets with biallelic mutations in either RAG1 or RAG2 exhibited hypoplasia of immune organs, failed to perform V(D)J rearrangement, and lost mature B and T cells. These immunodeficient RAG 1/2 knockout pigs are promising tools for biomedical and translational research. [ABSTRACT FROM AUTHOR]

Neuronal restricted progenitors (NRPs) represent a type of transitional intermediate cells that lie between multipotent neural progenitors and terminal differentiated neurons during neurogenesis. These NRPs have the ability to self-renew and differentiate into neurons, but not into glial cells, which is considered an advantage for cellular therapy of human neurodegenerative diseases. However, difficulty in the extraction of highly purified NRPs from normal nervous tissue prevents further studies and applications. In this study, we report the conversion of human fetal fibroblasts into human induced NRPs (hiNRPs) in 11 days by using just three defined factors: Sox2, c-Myc, and either Brn2 or Brn4. The hiNRPs exhibited distinct neuronal characteristics, including cell morphology, multiple neuronal marker expression, self-renewal capacity, and a genome-wide transcriptional profile. Moreover, hiNRPs were able to differentiate into various terminal neurons with functional membrane properties but not glial cells. Direct generation of hiNRPs from somatic cells will provide a new source of cells for cellular replacement therapy of human neurodegenerative diseases. [ABSTRACT FROM AUTHOR]