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470 A ROLE FOR PLATELET-DERIVED GROWTH FACTOR RECEPTOR ALPHA IN HEMATOPOIESIS.

Constitutively active mutations in platelet-derived growth factor receptor alpha (PDGFRα) are known to play a role in the development of hematopoietic neoplasias, but the role of PDGFRα in normal hematopoiesis is unclear. The Soriano lab has identified several transcriptional targets of PDGFRα and found that mice with mutations in these genes have substantially elevated white blood cell counts. My project attempts to further characterize the hematopoietic phenotype of four of the mutant lines as well as link the phenotypes to defective PDGF signaling. In our attempt to characterize the hematopoietic phenotypes of the mutant mice, thymus glands and spleens were harvested, fixed, sectioned, and stained with hematoxylin and eosin. Mutant and wild-type spleens were somewhat different morphologically, with an apparent surplus of lymphocytes seen in the mutants. No morphological differences between mutant and wild-type thymus glands were observed. Flow cytometry performed on the lymphocyte fractions of dissociated spleens showed no differences in the ratio of B to T lymphocytes in wild-type versus mutant mice at either 2 weeks or 6 weeks. To link the hematopoietic phenotype to PDGFRα signaling we attempted to perform similar sorting experiments on PDGFRα mutant embryos but were unable to obtain living embryos. We did, however, assess PDGFRα expression in hematopoietic tissue by analyzing green fluorescent protein (GFP) expression from a knock in PDGFRα line. GFP was expressed in embryonic day 12.5 liver, as well as in adult bone marrow, thymus, and spleen. Three lines of evidence suggest expression in the stromal compartment rather than hematopoietic cells themselves. First, GFP was not expressed in the lymphocyte fraction of spleens or thymus glands as determined by flow cytometry. Second, in antibody-stained cryosections, GFP-positive cells did not overlap with CD4+ cells in thymus or spleen. Finally, embryonic day 12.5 thymus glands expressed GFP strongly, and at this point, the thymus gland contains few hematopoietic cells. Although the role of PDGF signaling in hematopoiesis remains unclear, one may speculate that its expression in the stromal compartment plays a negative regulatory role in hematopoiesis.