Hyperglycemia and HIV-1 Nef significantly enhanced lipid oxidation in the CNS in vitro and in vivo. (A). Primary human astrocytes were cultured with 10,15, 20 and 25 mM glucose containing medium for 12 hours. The cells were then transduced HIV-1 nef expressing viral particles. 48 hours later, the Nef transduced astrocytes and cellular supernatants were collected and the lipid oxidation was determined by measuring the production of 8-isoprostaglandin-F2- α using ELISA kit (Stressgen, Victoria, BC, Canada). Astrocytes without any additional glucose (5 mM) treatment were used as control. Our results indicate that hyperglycemia increased the production of 8-isoprostaglandin-F2- α in dose dependent manner and Nef alone also showed a 3-fold increase in 8-isoprostaglandin-F2- α. (B) Hyperglycemia and HIV-1 Nef significantly enhanced the production of 8-isoprostaglandin-F2- α in the brain of mice: 1 × 107 viral particles generated through HIV-1 vectors or SNV vectors were injected into the brain of diabetes-induced mice via the cortex as described before. Age-matched non-diabetic mice injected with an equal volume of citrate buffer served as control. After 8 weeks the mice were sacrificed and lysates from the brain tissues were subjected to ELISA to determine the release of 8-isoprostaglandin-F2- α. The results depicted in this figure indicate that hyperglycemia enhanced the production of 8iso-F2- α in a dose-dependent manner and HIV-1 Nef either alone or in combination with hyperglycemia also enhanced the release of 8-isoprostaglandin-F2- α in CNS causing oxidative stress. The results are the mean value of triplicate samples.