POTENTIATION OF MURINE ASTROCYTE ANTIOXIDANT DEFENSE BY BCL-2 - PROTECTION IN PART REFLECTS ELEVATED GLUTATHIONE LEVELS

Overexpression of the proto-oncogene bcl-2 has been shown to protect a variety of cell types from oxidative and non-oxidative injury, blocki ng apoptotic and necrotic types of cell death, Retroviral vectors were used to stably overexpress bcl-2 in primary murine astrocyte cultures with more than 95% efficiency, Compared to beta-galactosidase-express ing and uninfected control cells, bcl-2 overexpressing astrocytes suff ered < 40% injury after 24 h glucose deprivation, while controls were essentially completely injured. After exposure to 0.2 mM hydrogen pero xide, the bcl-2 overexpressing astrocytes suffered < 40% the injury se en in controls, In contrast, when the cultures were injured by combine d oxygen-glucose deprivation, no difference in the extent or time cour se of injury was found between cells overexpressing bcl-2 and those ex pressing beta-galactosidase, To investigate one possible mechanism of bcl-2 protection, eve measured the levels of glutathione and three ant ioxidant enzymes, Astrocytes overexpressing bcl-2 had elevated glutath ione levels (130-200%), increased superoxide dismutase (170%) and glut athione peroxidase (140%) activities compared with beta-galactosidase- expressing controls. Bcl-2 overexpressing astrocytes suffered less lip id peroxidation after glucose deprivation, as assessed by cis-parinari c acid fluorescence, and demonstrated more rapid removal of hydrogen p eroxide from the medium, When glutathione levels were decreased 80% by pretreatment with buthionine sulfoximine, the extent of protection fr om glucose deprivation of bcl-2 overexpressing cells was decreased by about half, Increased antioxidant defence contributes to protection fr om glucose deprivation in bcl-2 overexpressing astrocytes.