The objective of this research project is to establish temporal and causal relationship between glucocorticoid-induced programmed cell death (apoptosis) and the effect of this group of steroids of cholesterol and dolichol synthesis. Two cell lines derived from human acute T-cell leukemia will be used. Although one line (CEM- C7) is glucocorticoid-sensitive and the other line (CEM-Cl) is glucocorticoid-resistant, both lines have specific functioning glucocorticoid receptors. Because steroid resistance is a frequent complication during chemotherapy, changes in lipid composition in membranes of neoplastic lymphoid cells might be a factor in the development of glucocorticoid-induced phenotypic alterations that do not directly involve the receptor system. The hypothesis which will be tested in the course of this study is formulated around the assumption that steroid induced lipid changes increase influx of ionic calcium into the cells thus stimulating the calcium-dependent endonuclease leading to nuclear fragmentation. The methodology to be used in this project will involve cholesterol and dolichol analyses by HPLC combined with radioisotope labeling studies of DNA fragmentation and measurement of Ca2+ influx by means of fluorescent probe Fura 2. Besides its importance in elucidating the mechanism of resistance to glucocorticoids, this study is also relevant to the role of lipid synthesis in the retroviral infection of lymphoid cells including infections caused by HIV.
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