IFNα-induced increase of PML NB number is suppressed by TSA. (A) Confocal images of HeLa cells untreated (CON) or treated with 1000 U/mL interferon-α (IFN), 500 ng/mL trichostatin A (TSA), or their combination (IFN + TSA) for 24 hours, immunostained with Sp100 or PML antibody; nuclei were costained with Sytox. Images were acquired by a Leica TCSSP confocal laser-scanning microscope equipped with a Leica HCX PL APO 63×/1.32 NA oil-immersion objective lens, and were processed by Leica Confocal Software version 2.5 (Leica Microsystems Heidelberg GmbH, Mannheim, Germany) and Adobe Photoshop 5.5 (Adobe Systems, San Jose, CA). Bar represents 10 μm. (B-C) Quantification of PML NBs. HeLa cells were treated with IFN, TSA, or IFN + TSA for the indicated time; were incubated first with one drug for a period of 12 hours and then incubated with the second drug after washing with PBS; or were maintained in the fresh medium (fm) for the next 12 hours as indicated for each column. The number of PML NBs per nucleus detected by PML antibody was counted on a total projection of confocal sections (1 section per 0.4 μm) through entire cell nucleus. n indicates the number of counted cells for each condition. Error bars represent standard error. Statistically significant differences to the control (untreated cells) are indicated by ***P < .001.