Figure 1. EBERs trigger inflammatory response in NPC cell lines.

A. mRNA levels of inflammatory cytokines (TNFα, IL-1α and IL-6) were determined by RT-PCR in HNE2 cells transfected with different doses of EBER1 (left panel) or EBER2 (right panel) expression plasmids for 24 h. Mock is the vector control. B. mRNA levels of inflammatory cytokines were measured by qPCR in HNE2 cells stably expressing single-copy EBER1 or EBER2 maintained in G418 supplemented medium. C. HNE2 cells stably expressing 10 tandem copies of EBER1 and EBER2 were subjected to qPCR for mRNA levels of inflammatory cytokines. D. Inflammatory cytokine expression triggered by in vitro transcribed EBER1 or EBER2. Results were presented as a fold of increase relative to the corresponding EBER plasmids. E. HNE2 cells were stimulated with in vitro transcribed EBER1 or EBER2. TNFα, IL-6 and IL-1α released into the culture supernatant were quantified by ELISA. F. (Left panel), exosomes were isolated from cell culture supernatant of C666–1 or 2 μg in vitro transcribed EBERs treated HNE2 cells, and then approximate equal amount of exosomes determined by TEM were applied to challenge untreated HNE2 cells. TNFα transcripts was determined by q-PCR. (Right panel), representative TEM images of exosomes from C666–1 cells culture supernatant. G. siRNA targeting EBER1 or EBER2 was transfected into C666–1 cells. mRNA levels of EBER1, EBER2 and TNFα were measured 72 h post transfection. All the data are from 3 independent experiments and representative data are expressed as mean ± SD. In (B), (C), (E), (F) and (G), *P < 0.05 versus control or untreated group.