HIV-1 Tat induces time dependent expression of IL-6 and IL-8 in SVG astrocytes. Seven hundred thousand SVG astrocytes were transfected with 0.3 μg of HIV-1 Tat plasmid using Lipofectamine 2000. (a, d) Cells were harvested at 1, 3, 6, 12, 24, 48 and 72 hours and total RNA was isolated. The expression levels of IL-6 and IL-8 were determined by real time RT-PCR. The values represented are normalized to their mock-transfected controls. (b, e) The concentrations of IL-6 and IL-8 in cell culture supernatants were measured at 6, 12, 24, 48, 72 and 96 hours after transfection by multiplex cytokine assay. Open bars and closed bars represent protein concentrations of mock- and HIV-1 Tat- transfected cells, respectively. Each experiment was done at least in triplicate and each bar represents the ± SE of three individual experiments. (c, f) Primary astrocytes from 2 independent donors were treated with 200 ng/ml of Tat protein for 2 hours and total RNA was isolated. The expression levels of IL-6 and IL-8 were determined by real time RT-PCR. Statistical analyses was performed by Student’s t-test and ** denotes P-value of ≤ 0.01.