|Budget Amount *help
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2004: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 2003: ¥2,100,000 (Direct Cost: ¥2,100,000)
To measure the ability of human hematopoietic stem cells (HSCs), SCID-repopulating cell (SRC) assay has been widely used, in which human HSCs are transplanted into NOD/SCID mouse. However, when human cells are transferred from a tail vein, cells must go through various obstacles until they reach the mouse marrow environment, which could explain the generally low homing efficiency in this system. Thus, the capability of HSCs may be underestimated by this intravenous (iv) transplantation method. In our attempt to reveal true SRC potential to self-renew and differentiate into multi-lineage mature cells in bone marrow, we introduced cells into mouse marrow directly (intra-bone marrow [iBM]) to minimize the effect of factors, which may prevent homing of HSCs, and compared the frequency of SRCs measured by the two method. When cord blood CD34^+CD38^- cells were transplanted in NOD/SCID mice by iBM, a 15-fold higher frequency of SRC, 1. in 44 CD34^+CD38^- cells, was achieved compared with 1 i
n 660 by iv. Furthermore, the iBM transplant showed high levels of engraftment in the secondary transplantation. Our results indicate that the iBM injection strategy is more sensitive and direct way to measure the capability of human SRCs and is useful to investigate the interaction of HSCs and marrow environment in vivo.
Bone marrow (BM) cells are reported to contribute to the process of regeneration following myocardial infarction. However, the responsible BM cells have not been fully identified. Here, we used two independent clonal studies to determine the origin of bone marrow (BM)-derived cardiomyocytes. First, we transplanted single CD34^-c-kit^+Sca-1^+lineage^- side population (CD34^-KSL-SP) cells or whole BM cells from mice ubiquitously expressing enhanced green fluorescent protein (EGFP) into lethally-irradiated mice, induced myocardial infarction (MI), and treated the animals with G-CSF to mobilize stem cells to the damaged myocardium. At 8 weeks post-MI, from 100 specimens we counted only 3 EGFP^+actinin^+ cells in myocardium of CD34^-KSL-SP cells-transplanted mice, but more than 5,000 EGFP^+ actinin^+ cells in whole BM cell-transplanted mice, suggesting that most of EGFP^+actinin^+ cells derived from non-hematopoietic BM cells. Next, clonally purified non-hematopoietic mesenchymal stem cells (MSC), CMG cells, that expressed EGFP in the cardiomyocyte-specific manner were transplanted directly into BM of lethally-irradiated mice, MI was induced, and they were treated with G-CSF. EGFP^+actinin^+ cells were observed in the ischemic myocardium, indicating that CMG cells had been mobilized and differentiated into cardiomyocytes. Together, these results suggest that the origin of the vast majority of BM-derived cardiomyocytes is MSC. Less