An in-Vivo Model of T Cell-Mediated Rejection of Human Hematopoietic CD34+ Stem Cells Using NOD/SCID γ^null (NOG) Mice.

Abstract 4474

We have previously demonstrated that human CD34+ cells include subsets of antigen presenting cells capable of stimulating anti-stem cell T cell alloreactivity in-vitro. In this study we transplanted human CD34+ cells and allogeneic T cells in a NOD/SCID γ^null (NOG) mouse model and evaluated the occurrence of stem cell rejection as well as xenogeneic graft-versus-host disease (GVHD) following the infusion of different doses of T cells. After sublethal irradiation NOG mice were cotransplanted with 2×10⁵ CD34+ cells and HLA mismatched CD4+CD25- T cells at 1:0 (control), 1:2 or 1:10 CD34+ cell: T cell ratio (n=5-10 mice per group). Hematopoietic stem cell and T cell engraftment was assessed in the bone marrow and in the spleen 6 weeks following transplantation or earlier in case the animals died. Control mice transplanted with CD34+ cells alone showed a high level of stem cell engraftment (huCD45+ cells: 60±10%) in the bone marrow, encompassing CD19+ B cells (64±4%), CD34+ cells (18±1%), CD33+ myeloid cells (7±1%), CD14+ monocytes (3±1%), and no T cells within huCD45+ cells. In contrast, mice that were transplanted with CD34+ cells and 4×10⁵ (1:2 ratio) or 2×10⁶ (1:10 ratio) T cells had only 9±2% and 3±1% huCD45+ cells, respectively, in the bone marrow (p=0.01). Moreover, marrow samples of mice cotransplanted with CD34+ cells and T cells at 1:2 or 1:10 ratio included >98% huCD3+ T cells and no CD34+ cells. Spleen engraftment of huCD45+ cells was lower (25±8%) in control mice (1:0 ratio) as compared to 66±10% and 36±11% in 1:2 and 1:10 groups, respectively (p=0.05). As observed in the marrow, also the spleen of animals receiving CD34+ and T cells included >98% CD3+ T cells. Among the T cells, both in the marrow and in the spleen of mice in the 1:2 and 1:10 ratio groups, 60-70% were CD4+CD8- cells, 22-25% CD8+CD4- cells, 1-3% CD56+ cells, and 2-5% CD4+CD25+ cells. In mice receiving 4 ×10⁵ T cells (1:2 ratio), on average 12±6% of the T cells in the bone marrow and spleen were CD4+CD8+. Only mice receiving 2×10⁶ T cells (1:10 ratio) showed GVHD. This was demonstrated by fur changes, reduced survival (p=0.02) and weight loss (p=0.0001) compared to control mice or mice receiving a lower dose of T cells (1:2 ratio). The marrow engraftment of CD3+ cells with disappearance of CD34+ cells in mice receiving low doses of allogeneic T cells, in the absence of evident xenogeneic GVHD, suggests that NOG mouse model represents a useful tool to study human stem cell rejection. This model will be also utilized to investigate new strategies of immunosuppressive cell therapy applied to stem cell transplantation in an HLA mismatched setting.