Fig. 7.

ORC6 depletion inhibits double-stranded RNA- or high mobility group box 1-induced NFκB activation in macrophages/monocytes. Stable THP-1 human macrophages with the specific ORC6 shRNA (“shORC6-6”), the Cas9-expressing THP-1 cells with the lentiviral CRISPR-ORC6-KO construct (koORC6-Clo1), or control THP-1 human macrophages with a scramble control non-sense shRNA plus CRISPR-Cas9 control construct (“shC + koC”), as well as THP-1 macrophages with the ORC6-expressing lentiviral construct (oeORC6-Slc1 or oeORC6-Slc2, two stable clones) or the empty vector (“Vec”) were established, cells were treated with or without HMGB1 (5 µg/mL) or dsRNA [poly(I: C), 10 µg/mL] for designated durations, NFκB activation was tested by p65 DNA-binding assay (A and E), mRNA expression and protein secretion (to the medium) of IL-1βwere evaluated using qPCR (B and F) and ELISA (C and G) assays, respectively, with cell viability analyzed by CCK-8 assay (D and H). Data were presented as mean ± standard deviation (SD, n = 5), with statistical significance indicated by ^#P < 0.05 compared to LPS treatment in “shC + koC”/“Vec” cells. Non-statistically significant differences (P > 0.05) were denoted as “N. S.“. These experiments were repeated five times, yielding consistent results across replicates