The long range goals of this project are: (a) to elucidate at the molecular level pathogenetic mechanisms involved in the uncontrolled growth of human gliomas; (b) to find means for manipulating these mechanisms in ways that could be applied clinically. One of the important factors responsible for the aggressive behavior of gliomas may be their resistance to natural killer cell (NK) cytolysis. We have demonstrated a correlation between the glycolipid composition of cultured human glioma and fetal brain cells and their resistance to NK lysis. This supports our working hypothesis that the glycoconjugate (ie. glycolipid and glycoprotein) composition of glioma cells is an important determinant of their susceptibility to NK attack. The experiments in this proposal have been designed to define the importance of several glycoconjugates and cellular differentiation in determining the sensitivity or resistance of glioma cells to NK. To do this we will subject cultured glioma and fetal brain cells to environmental alterations to change either their glycoconjugate composition or degree of differentiation, and determine their degree of binding to and lysis by NK. Glycoconjugate contents of target cells will be decreased by glycosidases and cycloserine, and increased by non-enzymatic glycosylation or serum-free medium. The degree of target cell differentiation will be altered by phorbol myristate acetate, and glial maturation factor, and analyzed using flow cytometry (glial fibrillary acidic protein, fibronectin, DNA, low angle forward light scatter). Neutral glycolipid and ganglioside compositions of target cells will be chemically quantiated using HPLC, HPTLC and scanning densitometry. Cell surface glycoconjugates will be analysed using galactose oxidase and NaB(3H)4. Membrane fluidity, cholesterol and phospholipids will also be quantitated. Results of chemical analyses and immunological assays will be statistically analysed to determine if glycoconjugates on target cells correlate with levels of target binding and lysis by NK. This information should identify which target cell glycoconjugates are involved in binding, and/or lysis of human gliomas by NK, and provide insight into the molecular mechanisms through which they operate.
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