Role of IFN-β and IFN-λ in PolyI:C-mediated anti-HIV activity. (A) Effect of PolyI:C-stimulated hCMEC/D3 culture SN on ISG expression of macrophages. HCMEC/D3 cells were stimulated with 1 µg/mL PolyI:C for 16 hours. Culture SN was collected for treatment of macrophages (10% v/v) for 6 hours. RNA was extracted, and the expression of ISGs (ISG56, MxA, OAS-1, and PKR) was measured by qRT-PCR. The results shown are mean ± SD of triplicate cultures, representative of 3 separate experiments. (B) Effect of neutralization antibodies (Abs) to IFNs or IFN-λ receptor on PolyI:C-stimulated hCMEC/D3 culture SN-mediated anti-HIV activity. PolyI:C-stimulated hCMEC/D3 culture SN was preincubated with anti-IFN-α (10 µg/mL), anti-IFN-β (10 µg/mL), or anti-IFN-γ (10 µg/mL) for 1 hour and then used to treat macrophages 24 hours prior to HIV Jago infection. For IFN-λ receptor pretreatment, the anti–IL-10Rβ neutralization antibody (10 µg/mL) was added to treat macrophages for 1 hour prior to the addition of SN. Eight days postinfection, RNA was extracted, and HIV GAG expression was measured by qRT-PCR. The results shown are mean ± SD of triplicate separate experiments. Asterisks indicate that the differences between the indicated groups are statistically significant (*P < .05; **P < .01). n.s., not significant.