AUTHOR=Boettcher Adeline N. , Li Yunsheng , Ahrens Amanda P. , Kiupel Matti , Byrne Kristen A. , Loving Crystal L. , Cino-Ozuna A. Giselle , Wiarda Jayne E. , Adur Malavika , Schultz Blythe , Swanson Jack J. , Snella Elizabeth M. , Ho Chak-Sum (Sam) , Charley Sara E. , Kiefer Zoe E. , Cunnick Joan E. , Putz Ellie J. , Dell'Anna Giuseppe , Jens Jackie , Sathe Swanand , Goldman Frederick , Westin Erik R. , Dekkers Jack C. M. , Ross Jason W. , Tuggle Christopher K. 

TITLE=Novel Engraftment and T Cell Differentiation of Human Hematopoietic Cells in ART−/−IL2RG−/Y SCID Pigs

JOURNAL=Frontiers in Immunology

VOLUME=11

YEAR=2020

URL=https://www.frontiersin.org/journals/immunology/articles/10.3389/fimmu.2020.00100

DOI=10.3389/fimmu.2020.00100

ISSN=1664-3224

ABSTRACT=<p>Pigs with severe combined immunodeficiency (SCID) are an emerging biomedical animal model. Swine are anatomically and physiologically more similar to humans than mice, making them an invaluable tool for preclinical regenerative medicine and cancer research. One essential step in further developing this model is the immunological humanization of SCID pigs. In this work we have generated T<sup>−</sup> B<sup>−</sup> NK<sup>−</sup> SCID pigs through site directed CRISPR/Cas9 mutagenesis of <italic>IL2RG</italic> within a naturally occurring <italic>DCLRE1C (ARTEMIS)</italic><sup>−/−</sup> genetic background. We confirmed <italic>ART</italic><sup>−/−</sup><italic>IL2RG</italic><sup>−/<italic>Y</italic></sup> pigs lacked T, B, and NK cells in both peripheral blood and lymphoid tissues. Additionally, we successfully performed a bone marrow transplant on one <italic>ART</italic><sup>−/−</sup><italic>IL2RG</italic><sup>−/<italic>Y</italic></sup> male SCID pig with bone marrow from a complete swine leukocyte antigen (SLA) matched donor without conditioning to reconstitute porcine T and NK cells. Next, we performed <italic>in utero</italic> injections of cultured human CD34<sup>+</sup> selected cord blood cells into the fetal <italic>ART</italic><sup>−/−</sup><italic>IL2RG</italic><sup>−/<italic>Y</italic></sup> SCID pigs. At birth, human CD45<sup>+</sup> CD3ε<sup>+</sup> cells were detected in cord and peripheral blood of <italic>in utero</italic> injected SCID piglets. Human leukocytes were also detected within the bone marrow, spleen, liver, thymus, and mesenteric lymph nodes of these animals. Taken together, we describe critical steps forwards the development of an immunologically humanized SCID pig model.</p>