Synchronized waves of osteoclast formation and death observed in long-term cultures.

<p>A, B) RAW 264.7 cells were treated with RANKL 50 ng/ml during 15 days. Samples were fixed, and either stained for TRAP to assess the numbers of TRAP-positive multinucleated osteoclasts, or stained with DAPI to assess nuclear morphology. A) Micrographs of TRAP-stained samples taken at indicated days. Calibration bar applies to all images. B) <i>Black circles:</i> changes in osteoclast numbers during 15 days of culture; <i>open circles:</i> percentage of osteoclasts exhibiting nuclear fragmentation (apoptotic cells), normalized to the total number of osteoclasts. C) Mouse bone marrow cells were treated with MCSF (20 mg/ml) and RANKL (50 ng/ml) during 15 days. Samples were fixed, stained for TRAP, and numbers of TRAP-positive multinucleated osteoclasts were counted. Arrow indicates days when RANKL was added.