Effects of administration of salubrinal on multi-nucleated osteoclast formation. Using bone marrow-derived cells isolated from the vehicle control and RANKL-treated mice, an osteoclast differentiation assay was performed. The culture medium was exchanged once on day 4 during the 6-day experiments. TRACP-positive multinuclear cells (> 3 nuclei) were identified as osteoclasts. The areas covered by multi-nucleated osteoclasts are quantified in response to 3 doses of salubrinal (1, 2, and 5 μM). The microphotographs represent the two groups of osteoclast cultures (vehicle control and RANKL administration) with TRACP staining. Bar = 200 μm. A: Area covered by multi-nucleated osteoclasts in response to in vitro administration of salubrinal from day 0 to day 6 (6 days). B: Area covered by multi-nucleated osteoclasts in response to in vitro administration of salubrinal from day 4 to day 6 (3 days). C: Fold change in response to 2 μM salubrinal.