Effect of TNF-α on HIV-1 transcript expression in infected astrocytes. HFA and Jurkat cells (positive control) were infected with HIV-1 NLENG1 (50 ng/ml) (YFP-expressing HIV-1 to track infection in live cells) and treated with TNF-α (10 ng/ml) or left untreated. (A) HIV-1 infection was assessed by the number of GFP-expressing cells. Representative pictures of Jurkat and HFA showing GFP-positive cells. (B) Quantification of HIV-1 infection was done by counting GFP-positive cells in 10 random fields and plotted as mean ± standard error of the mean (SEM). (C) HFA and Jurkat cells were treated with TNF-α (10 ng/ml) in combination with either HIV-1 wt or HIV-1 integrase- defective mutant (ΔINT) as a negative control for viral replication. Cells were harvested after 48 h of treatment for total RNA and monitored for early and late viral mRNA transcripts (tat, nef, and gag) using reverse-transcription coupled real-time PCR. UD = undetectable (n = 2). HFA, human fetal astrocytes; GFP, green fluorescent protein; wt, wild type.