Recent studies demonstrated that stem cells in a number of organs/tissues have the potential to differentiate into tissues other than those specified in their origins. The most striking evidence for this """"""""transdifferentiation"""""""" has been between bone marrow and muscular, hepatic, and CNS systems. Since all of the transplantation studies have been conducted using populations of cells, it is possible that the apparent bipotentiality of the stem cells is from engraftment by two separate populations of monopotential stem cells. Our proposal seeks definitive information on the tissue/organ engrafting capabilities of the hematopoietic and stromal stem cells by using single stem cell transplantation. We will use transgenic CD45.2 C57BL/6 (B6) mice that ubiquitously express enhanced green fluorescent protein (EGFP) as the source for donor stem cells and congenic CD45.1 B6 mice as recipients. The contribution of bone marrow EGFP-labeled donor cells to various organs/tissues of the recipient mice will be assessed using laser scanning confocal microscopy (LSCM) and conventional epifluorescence microscopy in combination with differential interference contrast (DIC) microscopy.
Two Specific Aims are proposed: (1) To define the potentials of individual hematopoietic stem cells by single stem cell transplantation to lethally irradiated recipient mice. (2) To define the potentials of stromal stem cells. We have developed a highly efficient technique to create mice with clonal hematopoietic engraftment by combining FACS cell sorting and short-term suspension culture of bone marrow cells. Collaboration with the laboratory of Dr. Christopher J. Drake of Department of Cell Biology and Anatomy, who has considerable expertise in the use of LSCM and DIC and epifluorescence microscopy, has already resulted in exciting preliminary information about the potentiality of a single EGFP-labeled stem cell.
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