A cis-regulatory element controls HDAC9 expression to fine-tune inflammasome-dependent chronic inflammation
Transcriptomic profiling of BMDMs from HDAC9 mutant mice. Overall design: Bone marrow-derived macrophages (BMDM) were generated as established by flushing the bone marrow from femurs and tibiae with ice-cold PBS, resuspending in PBS by repeated pipetting and filtering through a 40 µm cell strainer (BD Biosciences). After centrifugation of the cell suspension at 500 g for 10 min, the pellet was resuspended in culture medium (RPMI 1640 containing 10% FCS, 15% L929-cell-conditioned medium (LCM) and 100 µl/ml gentamycin) and plated on 15 cm untreated culture plates (Greiner). 15 % fresh LCM was added again on day 1 and 2 of culturing. For the stimulation experiments, differentiated macrophages were harvested by gentle scraping, transferred onto untreated 6-well or 12-well plates (Greiner) on Day 7 of culturing and left in LCM-free medium for 24 hours, allowing cells to adhere. Cells were then stimulated in FCS-free medium with LPS or left untreated.