LPS-induced production of sMer is independent of protein synthesis and is blocked by metalloproteinase inhibitors. (A) A 6-cm plate of J774 cells was cultured in the presence of 10 μg/mL cycloheximide for 30 minutes, rinsed with serum-free medium, and then incubated in serum-free medium containing 10 μg/mL cycloheximide and 50 ng/mL LPS for 1 hour. Two additional untreated plates were incubated in serum-free medium with or without 50 ng/mL LPS for 1 hour. sMer released into the medium and Mer and p53 in cell lysates were monitored by Western blot. (B) J774 cells were treated with 50 ng/mL LPS to stimulate the cleavage of the Mer extracellular domain. EDTA (5 mM), 200 μM TAPI-0, or DMSO (vehicle used to dissolve TAPI) was added to cell cultures as indicated for 2 hours. Mer remaining in cells and sMer released into the medium after each treatment were detected by Western blot.