Neutralization of IL-1β and TNF-α protects cell death of human neuroblastoma cell line, SK-N-SH. (A) WNV (MOI-1)- infected SK-N-SH cells were treated with neutralizing antibodies against IL-1β-, -6-, -8-, or TNF-α and cell toxicity was assessed by cell proliferation assay. Percentage cell toxicity was calculated by comparing to control cells. While neutralization of IL-8 and IL-6 did not result in significant reduction in WNV-induced cell death, it was significantly attenuated in the presence of anti-IL1β or -TNF-α at day 2 after infection. Data are expressed as mean ± SD from three independent experiments conducted in triplicates. (B) WNV (MOI-1)-infected SK-N-SH cells were treated with anti-IL-1β or -TNF-α and assessed for apoptosis at day 2 after infection. Abundant TUNEL-positive cells (red, i) co-localized with DAPI (blue; ii) were observed in WNV-infected cells. In contrast, apoptosis induced by WNV was significantly attenuated in the presence of anti-IL-1β (iii) and anti-TNF-α (v). The images depicted are representative results of three independent experiments. (C) Quantitative representation of TUNEL-positive cells in each group from three independent cover slips from three independent experiments. Quantitative analysis indicated that neutralization of IL-1β and TNF-α reduced the number of TUNEL positive cells to less than half compared to WNV-infected cells. *p < 0.05.