We propose to continue studies characterizing the non-hemopoietic stromal cells responsible for stem cell maintenance in long term in-vitro liquid marrow cultures. The two candidate stromal regulator cells which have been isolated by combination of radiation and enzyme digestion will be characterized with regard to antigen constitution, collagen biosynthesis, capacity to produce hemopoietic regulatory factors and to support stem cells in long term liquid culture. Non-irradiated stromal cells will also be evaluated initially utilizing high dose lithium which appears to selectively deplete these marrow cultures of all hemopoietic elements. The mechanism whereby lithium stimulates hemopoiesis will continue to be studied in in vitro liquid culture focusing on the data suggesting that lithium acts via an adherent stromal cell. We will evaluate the effect of lithium on hemopoietic regulatory factor production by irradiated and non-irradiated stromal cells, the cell type on which lithium is acting, and the mechanism underlying the stimulation of stem cells. We will continue studies on the stem cell level at which lithium is acting, concentrating initially on the high proliferative potential stem cell (HPP-CFC) and the day 10 and 14 CFU-S. In a similar manner we will continue investigations of the mechanisms underlying the stimulation of in-vitro Dexter culture hemopoiesis by 5 inches adenosine monophosphate and adenosine. We also plan studies on the mechanism by which lithium causes a late decline in in-vitro hemopoiesis and the effect of lithium on S1/SLD and W/Wv liquid marrow cultures. Lastly, we will continue our efforts to establish a more suitable human (porcine) liquid marrow culture, in order to evaluate features outlined above in these systems.
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