Engraftment of human HSCs in nonirradiated newborn NOD-scid IL2rγ null mice is enhanced by transgenic expression of membrane-bound human SCF

Abstract

Immunodeficient mice engrafted with human HSCs support multidisciplinary translational experimentation, including the study of human hematopoiesis. Heightened levels of human HSC engraftment are observed in immunodeficient mice expressing mutations in the IL2-receptor common γ chain (IL2rg) gene, including NOD-scid IL2rγ(null) (NSG) mice. Engraftment of human HSC requires preconditioning of immunodeficient recipients, usually with irradiation. Such preconditioning increases the expression of stem cell factor (SCF), which is critical for HSC engraftment, proliferation, and survival. We hypothesized that transgenic expression of human membrane-bound stem cell factor Tg(hu-mSCF)] would increase levels of human HSC engraftment in nonirradiated NSG mice and eliminate complications associated with irradiation. Surprisingly, detectable levels of human CD45(+) cell chimerism were observed after transplantation of cord blood-derived human HSCs into nonirradiated adult as well as newborn NSG mice. However, transgenic expression of human mSCF enabled heightened levels of human hematopoietic cell chimerism in the absence of irradiation. Moreover, nonirradiated NSG-Tg(hu-mSCF) mice engrafted as newborns with human HSCs rejected human skin grafts from a histoincompatible donor, indicating the development of a functional human immune system. These data provide a new immunodeficient mouse model that does not require irradiation preconditioning for human HSC engraftment and immune system development.

Figure 1

Radiosensitivity of NSG and NSG-Tg(hu-mSCF). NSG and NSG-Tg(hu-mSCF) mice were irradiated with 350 cGy (A), 300 cGy (B), or 250 cGy (C). Mice (5 mice per group) were then monitored for survival.

Figure 2

Weights of mice engrafted with human HSC as newborns. Weights of nonirradiated NSG-Tg(hu-mSCF) and 100-cGy irradiated NSG and NSG-Tg(hu-mSCF) mice engrafted with human HSCs as newborns were obtained over time after weaning. (A) Male mice. (B) Female mice. Each symbol represents an individual data point. The lines represent the average of the individual data points.

Figure 3

Human CD45⁺ cell chimerism and CD3⁺ T-cell development in the blood of newborn-engrafted mice at 12 weeks of age. Nonirradiated NSG and NSG-Tg(hu-mSCF) and 100-cGy irradiated NSG and NSG-Tg(hu-mSCF) newborn mice were engrafted with human HSCs as described in “Engraftment of mice human HSCs.” Mice were analyzed by flow cytometry for the percentage of human CD45⁺ and the percentage of human CD45⁺ cells that were CD3⁺ in the blood at 12 weeks of age. (A) Percentage of human CD45⁺ cells. (B) Percentage of human CD45⁺ cells that were CD3⁺. No significant difference between groups was observed.

Figure 4

Human CD45⁺ cell chimerism in the bone marrow of newborn-engrafted mice at 12 weeks of age. Nonirradiated NSG and NSG-Tg(hu-mSCF) and 100-cGy irradiated NSG and NSG-Tg(hu-mSCF) newborn mice were engrafted with human HSCs as described in “Engraftment of mice human HSCs.” Mice were analyzed by flow cytometry for the percentage of human CD45⁺ cells in the bone marrow at 12 weeks of age. (A) Percentage of human CD45⁺ cells. (B) Total number of human CD45⁺ cells. Each symbol represents an individual animal. Significant differences were observed in the number of human CD45⁺ cells: 0 cGy NSG-Tg(hu-mSCF) vs 0 cGy NSG (P < .01); 0 cGy NSG vs 100 cGy NSG (P < .01).

Figure 5

Skin transplantation on nonirradiated NSG-Tg(hu-mSCF) mice that were non-HSC–engrafted or engrafted with human HSCs as newborns. Nonirradiated NSG-Tg(hu-mSCF) mice were non-HSC–engrafted (left panels) or engrafted with human HSCs as newborns (right panels) as described in “Engraftment of mice with human HSCs.” At 12 to 14 weeks of age, all mice were transplanted and monitored for rejection of human skin allografts. Left side indicates the stain used for each row. (A-B) H&E indicates hematoxylin and eosin. (C-D) CD45 indicates all human hematolymphoid cells. (E-F) CD31 indicates human endothelium. (G-H) INV indicates human involucrin. (I-J) VIM indicates human vimentin.

Figure 6

Human CD45⁺ cell chimerism in the blood of newborn-engrafted mice at 12 weeks of age. Nonirradiated NRG and NRG-Tg(hu-mSCF) mice and 400-cGy NRG and NRG-Tg(hu-mSCF) mice were engrafted as newborns with human HSCs as described in “Engraftment of mice with human HSCs.” Mice were analyzed by flow cytometry for the percentage of human CD45⁺ cells in the blood at 12 weeks of age. Each symbol represents an individual animal. No significant differences were observed among the nonirradiation groups or among the irradiated groups. Engraftment in either of the nonirradiated groups was significantly different from the engraftment observed in either of the irradiated groups (P < .001).