Stem Cell Reports Repor t Nonirradiated NOD,B6.SCID Il2rg / Kit W41/W41 (NBSGW) Mice Suppor t Multilineage Engraftment of Human Hematopoietic Cells Brian E. McIntosh, 1 Matthew E. Brown, 2 Bret M. Duffin, 1 John P. Maufort, 1 David T. Vereide, 1,3 Igor I. Slukvin, 4,5 and James A. Thomson 1,6,7, * 1 Morgridge Institute for Research, Madison, WI 53715, USA 2 Department of Surgery, University of Wisconsin, Madison, WI 53715, USA 3 Biotechnology Center, University of Wisconsin, Madison, WI 53706, USA 4 Wisconsin National Primate Research Center, University of Wisconsin, Madison, WI 53715, USA 5 Department of Pathology and Laboratory Medicine, University of Wisconsin, Madison, WI 53715, USA 6 Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706, USA 7 Department of Molecular, Cellular, and Developmental Biology, University of California, Santa Barbara, Santa Barbara, CA 93106, USA *Correspondence: jthomson@morgridgeinstitute.org http://dx.doi.org/10.1016/j.stemcr.2014.12.005 This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/). SUMMARY In this study, we demonstrate a newly derived mouse model that supports engraftment of human hematopoietic stem cells (HSCs) in the absence of irradiation. We cross the NOD.Cg-Prkdc scid Il2rg tm1Wjl /SzJ (NSG) strain with the C57BL/6J-Kit W-41J /J (C57BL/6.Kit W41 ) strain and engraft, without irradiation, the resulting NBSGW strain with human cord blood CD34+ cells. At 12-weeks postengraftment in NBSGW mice, we observe human cell chimerism in marrow (97% ± 0.4%), peripheral blood (61% ± 2%), and spleen (94% ± 2%) at levels observed with irradiation in NSG mice. We also detected a significant number of glycophorin-A-positive expressing cells in the developing NBSGW marrow. Further, the observed levels of human hematopoietic chimerism mimic those reported for both irradiated NSG and NSG-trans- genic strains. This mouse model permits HSC engraftment while avoiding the complicating hematopoietic, gastrointestinal, and neurological side effects associated with irradiation and allows investigators without access to radiation to pursue engraftment studies with human HSCs. INTRODUCTION The NOD.SCID Il2rg / (NSG) mouse strain dramatically improved investigators’ ability to study human hematopoi- etic stem cell (HSC) engraftment (Rongvaux et al., 2013; Shultz et al., 2005). The NSG strain combines a series of mu- tations that inhibit the host’s immune system by different mechanisms (Bosma et al., 1983; Cao et al., 1995; Greiner et al., 1995; Ohbo et al., 1996; Shultz et al., 1995). The NSG and other host strains require myeloablative con- ditioning (i.e., radiation) to achieve high levels of human chimerism (McDermott et al., 2010; Shultz et al., 2005). In addition to the effects of irradiation on the hematopoiet- ic system that can promote engraftment (Broudy, 1997), confounding side effects of irradiation, including necrosis and apoptosis of gastrointestinal, neural, and muscle tis- sues, can lead to wasting, infection, and even death (Li et al., 2004; Qiu et al., 2010). Two recent studies report limited success in engrafting human HSCs into nonmye- loablated NSG hosts, but the chimerism achieved in the pe- ripheral blood was modest (an average of 3% ± 3% and 18.3% ± 13%) (Brehm et al., 2012; Bueno et al., 2010). Viable mutant Kit oncogene (Kit) murine strains support the engraftment of mouse hematopoietic cells without host irra- diation (Wang and Bunting, 2008). The Kit (i.e., c-Kit, stem cell factor [SCF] receptor) encodes a type I membrane protein in the type III tyrosine kinase growth factor receptor family (Yarden and Ullrich, 1988), which is expressed on hemato- poietic, melanocyte, neural, and germ cells (Mintz and Rus- sell, 1957; Poole and Silvers, 1979; Russell, 1979). When its ligand, SCF, binds to c-Kit, it induces receptor homodimeriza- tion and signal transduction (Hsu et al., 1997). The c-Kit is required for normal hematopoiesis, and viable mutants most closely resemble aplastic anemia (Geissler et al., 1981). Mouse hosts with mutations in Kit thus provide a competitive advantage for WT donor cells and allow the engraftment of HSCs with reduced or no irradiation (Fleish- man, 1996; Waskow et al., 2009). Until recently, these strains have been short-lifespan heterozygotes (e.g., Kit +/Wv ), causing a burden on producing experimental hosts (Cosgun et al., 2014; Waskow et al., 2009). We demonstrate that nonirradiated NBSGW mice are similar in chimerism to that achieved in irradiated NSG (irNSG) mice and that NBSGW mice also support the serial transplantation of HSCs. RESULTS Nonirradiated Xenograft NBSGW Mice Show Increased Chimerism in the Peripheral Blood when Compared with Non-irNSG Mice and Are Similar to Irradiated NSG Mice We outcrossed mice homozygous for the Kit W41 allele with the NSG strain. The resultant F1 triple-heterozygotes Stem Cell Reports j Vol. 4 j 1–10 j February 10, 2015 j ª2015 The Authors 1 Please cite this article in press as: McIntosh et al., Nonirradiated NOD,B6.SCID Il2rg / Kit W41/W41 (NBSGW) Mice Support Multilineage Engraftment of Human Hematopoietic Cells, Stem Cell Reports (2015), http://dx.doi.org/10.1016/j.stemcr.2014.12.005