Phospho histone h3

All samples were fixed in 10% formalin, paraffin-embedded and 5-μm sections were stained with TUNEL using the ApopTag kit (Calbiochem). IHCfor Mcl-1, phospho-p70-S6K (Cell Signaling Technologies) and phospho-histone H3 (Santa Cruz Biotechnology) were performed on sections as described [45 (link), 46 (link)]. All histological analyses were photographed using Olympus DP2 software (200x).

Whole-cell lysates were prepared in Pierce RIPA Buffer with Halt^TM Protease & Phosphatase Inhibitor Cocktail (Thermo Scientific). Protein concentration was measured using Bradford assay (Bio-Rad). An appropriate volume of Laemmli sample buffer was added to equal amounts of total proteins followed by boiling for 5 min. Cells lysates were resolved by TRIS–glycine sodium dodecyl sulphate–polyacrylamide gel electrophoresis gels (Bio-Rad), transferred onto nitrocellulose membranes and the membranes then incubated with primary antibodies overnight at 4 °C. The immunoblots were subsequently washed and incubated with horseradish peroxidase-conjugated anti-mouse or -rabbit antibodies (Amersham Biosciences) for 1 h before detection by ChemiDoc MP system (Bio-Rad) using ECL substrate (Thermo Scientific). Antibodies used were: Abcam: Lamin B1 (ab65986); IκBα (ab7217), Nup153 (ab171074); Bethyl:BubR1 (A300-386A); Cell Signaling Technology (CST): cleaved caspase-3 (Asp 175) (9664S), cleaved PARP (Asp214) (9546), phospho-histone H3 (Ser10) (9701S), phospho-histone H2A.X (Ser139) (2577), phospho-NF-κB p65 (Ser536) (3033), NF-κB p65 (D14E12) (8242), phospho-IκBα (Ser32/36) (9246), p21 Waf1/Cip1 (12D1) (#2947); Santa Cruz: Actin (sc-47778), cyclin B1 (H-20) (sc-594), p53 (DO-1) (sc-126); Novus Biologicals: CXCL8/IL-8 (6217), IL-1 beta/IL-1F2 (8516).