| Development of the human/rat chimera model forthe study of the in vivo potential of humanhematopoietic stem cells (HSC)Hematopoiesis is characterized by a continuous, dynamic process of cell turnover. This cell renewal process is supported by a small number of hematopoietic stem cells (HSCs) that are capable of self-renewal and differentiation into multiple hematopoietic Lineages. In the mouse, the ability to assay HSCs in vivo has allowed relatively extensive characterization of these cells. In contrast to murine studies, studies of human HSCs have relied on a number of in vitro assays that identify cells with the capacity to generate hematopoietic progenitors in long-term culture and to differentiation into multiple hematopoietic Lineages. Recently, two elegant animal models that enable the in vivo measurement of human HSCs marrow repopulating potential have been developed. One is the sheep in utero transplantation system in which human hematopoietic progenitor cells are transferred into the sheep fetus in utero before the development of the ovine immune system. The presence or absence of human blood cells is then followed after the sheep is born. Another is the NOD/SCID mouse assay developed by Dick's group. This two models make assay human HSCs in vivo become a possibility. But to us, the first model is not easily accessible, nor the NOD/SCID mouse available. On theconsideration of these problems and the fact that the rat immune system is not fully developed until birth, we tried to transplant neonatal rat with human HSCs to test the possibility of this xenograft model.CD34 used to be used as a prerequisite marker for HSCs, but recently, seve 1 laboratory have proved the existence of CD34-negative HSCs independently. In order not to discard a population of potential stem cells, we purified HSCs from human cord blood (CB) using negative selection strategy based on 10 different Lineage-specific antigens. The so called Lin- cells was injected into the liver of neonatal rat using a micro-injector at an average of 0.5-2 X 106cells each. Peripheral blood (PB) was collected 2> 4> 8 weeks after injection. FACS was used to quantity human cells in the rat PB, PCR and immunohistochemisty were used to detect human cells in the PB as well as in other tissues, such as liver, spleen, pancreas, kidney, et al. As a result, human cells were detected in the rat liver, spleen and pancreas.To date, HSCs have been found more potential in their ability to differentiate into more than hematopoietic Lineage. They can be induced to differentiate into muscle, liver, neuron, et al. In our experiments, human cells were also detected in liver, spleen and pancreas after HSCs injection. These one more time proved the multipotential of HSCs. And we identified that HSCs can differentiate into pancreas in vivo for the first time. Theresults suggested that human HSCs can engraft into rat and differentiate into multiLineage under certain microenvironment.Thus, the human/rat xenograft model may provide a useful and convenient method for assay human HSCs in vivo. Compared with the existed two models, our model is a little crude in that human cells can' t survival long enough to prove HSCs' long-term repopulating ability. But the following strategies may be useful in prolonging human cells' survival in rat: 1) enlarge the cell number injected every neonatal rat; 2) use human stromal cells for co-transplantation; 3) use human-specific cytokines; 4) transplant for more than one times. |
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