Project Title: 16, Molecular and Cellular Radiobiology Effects of Radiation and Chemicals on Control of Hemopoiesis Technical Progress in FY 1973: RX-03-02-(b) Fe-55 suicide of differentiated erythropoietic cells was demonstrated and the relationship of Fe-55 dose to suppression of erythropoiesis was established. In addition, it was shown that suppression of erythropoiesis resulted in a diminution of the number of CFUs's, suggesting that the flow of stem cells into erythropoiesis had been increased, partially depleting the CFUs's in the marrow. Since depletion results in increased mitotic race it may be possible to test whether the HSC has limited or unlimited capacity to divide. Initial studies on the determination of the Do of human stem cells irradiated in vitro and grown in diffusion chambers were completed. The Do determined for all granulocytic cells is about 115 rads and for proliferating granulocytic cells approximately 70 rads. ,* In studies on growth of murine bone marrow cells in diffusion chambers it was shown that a humoral factor diffuses into the chambers stimulating the proliferation of the HSC as indicated by the increase in number of CFUg's and yield of total granulocytic series. Also, growth was shown to be greater in the irradiated than in the non-irradiated host. These observations of FY 1972 were confirmed in continued experiments, but since there may be xenogeneic effects (human cultures in the mouse) and allogeneic effects (non-inbred cultures in the mouse), an autologous system of diffusion . chamber culture of hemopoietic cells was developed using goats. Bone marrow is removed from the goat, prepared and introduced into diffusion chambers. ‘ After the goat is irradiated in the Medical Research Reactor gamma chamber the diffusion chambers are implanted into its peritoneal cavity and subsequently harvested at pre-determined intervals, There is excellent growth of the granulocytic series and macrophages, but lesser growth of lymphopotetic and erythropoietic cells. This system eliminates histocompatability problems on cell proliferation. Since large amounts of plasma and peritoneal washings are obtainable in the goat, production and isolation of the factor that diffuses from the irradiated animal into diffusion chambers and stimulates hemopoiesis is being studied. Upon demonstration that renal glomerular cultures of human and goat kidneys would produce EP, a grant was obtained from NIH to further these .studies, All cultures of goat glomeruli produce EP, However, in three cultures made from separate human kidneys, one produces EP and the other t uce colony-stimulating factor (CSF), an agent that increases the yie 3 granulocytic colonies in in vitro bone marrow culture, Glomerular cu iiiieres continue to produce EP forover one year even though the monolayers have been replaced by thick layers of cells. When trypsinized, resuspended, and sub-cultured in almost the total absence of the original glomeruli, these cultures of apithelial cells with rare glomeruli continue to produce EP. Dr. Havran, Mount Sinai University School of Medicine, has commenced column separation and purification of EP from supernates of glomerular cultures. (See Continuation Sheet) 44192849 RX-229