STEM CELLS AND AGING

描述（由申请人提供）：骨髓干细胞是衰老决定因素的假设将在小鼠早衰模型中进行测试。首先，在体外和体内检测野生型p53+/+和早衰型p53+/m C57BL/6J小鼠骨髓干细胞的数量和归巢能力。然后，将确定野生型骨髓干细胞在p53+/m突变小鼠中纠正过早衰老的能力，以及来自过早衰老p53+/m突变小鼠的骨髓干细胞在野生型受体中诱导过早衰老的能力。在Specific Aim 1中，野生型p53+/+、p53+/-和p53+/m小鼠的干细胞数量将通过流式细胞术和体外集落形成来确定，并通过体外迁移和体内骨髓再生来确定归巢。在Specific Aim 2中，将来自GFP+雄性C57BI6 p53+/+小鼠的骨髓细胞移植到p53突变型（p53+/m）和野生型（p53+/ +）雌性小鼠身上，并测定其对衰老表型发展的影响。供体细胞对受体组织的贡献将通过GFP和y染色体的存在来识别。如果在这个模型中，衰老是由于骨髓干细胞替代正常组织的能力下降，那么将正常小鼠的骨髓细胞移植到p53突变小鼠身上应该可以恢复正常的表型和正常的寿命。在Specific Aim 3中，将来自GFP+ p53+/m雄性突变小鼠的骨髓细胞移植到辐照的正常野生型雌性小鼠和雌性突变小鼠中，以确定衰老表型是否可以转移突变骨髓干细胞。最后，将确定供体骨髓细胞与受体小鼠组织细胞融合的可能性以及融合细胞在肿瘤中的存在。这些实验的结果应该为成人干细胞最重要的假设特性之一提供关键证据。

DESCRIPTION (provided by applicant): The hypothesis that bone marrow stem cells are a determining factor in ageing will be tested in a mouse model of premature ageing. First, the number and homing capacity of bone marrow stem cells from wild type p53+/+ and premature ageing p53+/m C57BL/6J mice will be assayed in vitro and in vivo. Then, both the ability of wild-type bone marrow stem cells to correct premature ageing in p53+/m mutant mice, and the ability of bone marrow stem cells from premature ageing p53+/m mutant mice to induce premature ageing, in wild-type recipients will be determined. In Specific Aim 1, the number of stem cells in wild-type p53+/+, p53+/- and p53+/m mice will be determined by flow cytometry and by in vitro colony formation, and homing determined by in vitro migration and marrow repopulation in vivo. In Specific Aim 2 bone marrow cells from GFP+ male C57BI6 p53+/+ mice will be transplanted to p53 mutant (p53+/m) and to wild-type (p53 +/+) female mice and the effect on the development of the aging phenotype determined. The contribution of donor cells to recipient tissues will be identified by the presence of GFP and the Y-chromosome. If ageing is due to a decreased ability of bone marrow stem cells to contribute to replacement of normal tissues in this model, transplantation of bone marrow cells from normal mice to p53 mutant mice should restore normal phenotype and normal life span. In Specific Aim 3 bone marrow cells from GFP+ p53+/m male mutant mice will be transplanted to irradiated normal wild-type female mice and to female mutant mice to determine if the aging phenotype can be transferred mutant bone marrow stem cells. Finally the possibility of fusion of donor bone marrow cells with tissue cells of recipient mice and the presence of fused cells in tumors will be determined. The results of these experiments should provide critical proof for one of the most important hypothetical properties of adult stem cells.