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. 2019 Feb 12;10(1):e02510-18. doi: 10.1128/mBio.02510-18

FIG 4.

FIG 4

Infection of THP-1 cells with live HCMV inhibits IL-1β secretion in response to UV-HCMV. (A) Schematic depicting five experimental treatments of differentiated THP-1 cells, including mock treatment (condition M), UV-inactivated HCMV for 24 h (condition 1), live HCMV for 24 h (condition 2), UV-inactivated HCMV for 12 h and then retreatment with UV-inactivated virus for 12 h (condition 3), or live HCMV for 12 h and then UV-HCMV virus for 12 h (condition 4). (B) IL-1β secretion measured by ELISA following indicated treatments. (C, D) mRNA levels for IE86 (C) and pro-IL-1β (D) measured by qPCR following indicated treatments. Transcript fold changes are expressed as relative to levels in untreated cells (mock). Values are averages ± SEM from three biological replicates. One-way ANOVA with Bonferroni correction was performed. *, P < 0.05; ***, P < 0.001. (E) Differentiated wild-type (WT) or ΔCASP-1 THP-1 cells were infected with HCMV-TB40/E (MOI of 3) for 6 days, and titers in supernatants from these cells were determined on human fibroblasts. Values are averages ± SEM from three biological replicates. ffu, focus-forming units. (F) Differentiated ΔHUMCYC or ΔIL-1β THP-1 cells were infected with HCMV-TB40/E (MOI of 3) for 6 days, and titers in supernatants from these cells were determined on human fibroblasts. Values are averages ± SEM from three biological replicates. Immunoblot assay shows successful knockout of IL-1β in ΔIL-1β cells. (G) WT or ΔCASP-1 THP-1 cells were infected with HCMV-TB40/E (MOI of 10) for 5 days and then treated with PMA to induce HCMV reactivation. Supernatants were collected 7 days post-PMA treatment, and titers determined on human fibroblasts. Values are averages ± SEM from three biological replicates. (H) Telomerized human fibroblasts (THF) were treated with 2 ng/ml of recombinant IL-1β or with 1,000 U of IFN-β overnight and then infected with HCMV-TB40/E (MOI of 3) for 5 days. Titers in supernatants from THF were next determined on human fibroblasts. Values are averages ± SEM from three biological replicates. One-way ANOVA with Bonferroni correction was performed. *, P < 0.05.