Effect of HIV-1 infection on ApoE expression in MDMs.

(A) Heat map showing the up-regulation of ApoE expression (indicated in red) among Apo genes in HIV-1-infected MDMs were generated from the microarray analysis. The color bar indicates gene expression in the log2 scale. MDMs prepared from a healthy donor were infected with HIV-1 JR-FL (100 ng/mL p24) by using the supernatants of HEK293A cells transfected with the HIV-1 molecular clone as a source of viruses, and cultured for 3 days prior to total RNA isolation (right). The control uninfected MDMs were prepared by culturing with media for 3 days (left). dpi, days post-infection.(B) MDMs (3 donors) were infected with HIV-1 JR-FL as in (A), cultured for 3, 6 or 8 days, lysed, and subjected to western blot to analyze the expression of ApoE. HIV-1 p24 levels were also analyzed to verify the viral replication. Anti-β-actin blot was used as a loading control. The total cell lysates of MDMs were also prepared immediately before HIV-1 infection as a control (“0 dpi”). The arrowheads indicate the high and low molecular weight ApoE. (C) Heat maps showing the up-regulation of various IFN-stimulated genes (ISGs) including HIV-1 restriction factors such as MX2, APOBEC3G (A3G), BST-2 and IFITMs were generated from the microarray analysis as in (A). (D, E) MDMs (3 donors) were left untreated or treated with 1,000 U/mL IFN-, cultured for 24 h, and subjected to qRT-PCR to analyze the mRNA level of MX2 (panel D) or ApoE (panel E) followed by the normalization to the mRNA level of β-actin. Each mRNA level was calculated relative to untreated MDMs (fold). Error bars in these panels indicate standard deviation of triplicate PCR assays.