ZIKV induces ER stress, DNA damage and cell death in human astrocytes.

(A) Astrocytes viability was determined by trypan blue dye exclusion method at different time point after infection with ZIKV. (B) Human astrocytes were infected with ZIKV (MR766, R103451 and PRVABC59) at an MOI of 0.1. After 48 hpi, RNA was extracted from cell lysates and RT-PCR arrays were performed to determine RNA expression of genes involved in human unfolded protein response pathway. Genes significantly altered by ZIKV infection are presented in the graph. (C-H) Western blot analysis from cell lysates (collected at 48 hpi in RIPA buffer) from uninfected and ZIKV infected human astrocytes showed expression of proteins involved in the unfolded protein response, including, PERK, IRE-1α, phosphorylated PERK, phosphorylated IRE-1α, full length and cleaved ATF6α, and DDIT3. (I and J) JNK and NF-κB protein expression by the African and the Asian strain respectively as measured by western blot with cell lysates of human astrocytes collected after 48 hpi. (K-L) Human astrocytes were transfected with siRNA against PERK and infected with ZIKV for 48 hours. Viral titer was measured by RT-PCR (K) and IP-10 levels were measured by ELISA (L). Mock (PBS) infected human astrocytes were used as control and the infection dose of ZIKV was at an MOI of 0.1. Data are presented as the mean ± SEM from three independent experiments. (*p#p<0.05 Vs ZIKV alone).