Transplantation of Human Aldehyde Dehydrogenase Expressing Cells Leads to Widespread Tissue Distribution of Donor Cells in the Novel NOD/SCID/MPSVII Xenotransplantation Model.

Beta-glucuronidase (GUSB) is a lysosomal enzyme expressed in virtually all human and murine cell types. Transplantation of GUSB-expressing human hematopoietic stem cells (HSC) into the recently developed GUSB-deficient NOD/SCID/MPSVII mouse allows for the sensitive histochemical detection of xenotransplanted cells in hematopoietic and non-hematopoietic tissues. Most notably, human cells expressing GUSB can be detected without reliance on the expression of human cell surface markers. In addition, we have recently characterized a novel population of reconstituting HSC from human umbilical cord blood (UCB) by lineage depletion (Lin⁻) followed by selection of cells with high aldehyde dehydrogenase (ALDH) activity. We have also demonstrated the robust hematopoietic engraftment of ALDH^hiLin⁻ cells in the NOD/SCID model. Because donor cell surface protein expression may be altered in non-hematopoietic tissues due to cell fusion or other mechanisms, we have used the NOD/SCID/MPSVII model to study the tissue distribution of ALDH^hiLin⁻ or ALDH^loLin⁻ cells in multiple organs. Tail vein injection of 2x10⁵–4x10⁵ ALDH^hiLin⁻ cells into sub-lethally irradiated (300cGy) NOD/SCID/MPSVII mice (n=3) demonstrated high levels of hematopoietic (human CD45⁺ cells) engraftment in the BM (70.5±15.1%), spleen (7.0±2.8%) and peripheral blood (17.0±10.7%). In contrast, injection of 2x10⁵–10⁶ ALDH^loLin⁻ cells (n=3) did produce significant human cell engraftment in these hematopoietic tissues, with only 1 of 3 mice engrafting at 0.3% CD45⁺ cells in the murine BM. By using GUSB-specific histochemical staining, significant human engraftment was detected, with single cell sensitivity, in hematopoietic (BM and spleen) and non-hematopoietic (liver, pancreas, and lung) tissues of mice transplanted with ALDH^hiLin⁻ cells. GUSB activity was colocalized with CD45 immunohistochemical staining in the BM, liver, and pancreas of mice transplanted with ALDH^hiLin⁻ cells. Human cells were not detected in the muscle or brain of these engrafted mice. In contrast, mice transplanted with equal doses of ALDH^loLin⁻ cells showed no GUSB expression in the BM, spleen, liver, pancreas, lung, muscle, and brain. We are currently developing a flow cytometric assay to enumerate the frequency of GUSB expressing cells in the tissues of the transplanted mice. These data indicate that ALDH^hiLin⁻ UCB cells demonstrate previously uncharacterized engraftment in the liver, pancreas, and lung of NOD/SCID MPSVII mice. This novel model provides a unique opportunity to visualize donor human cells after xenotransplantation without reliance upon the expression of cell surface proteins, in situ hybridization, or cell labeling using viral transduction or membrane dyes.