Figure 1.

Graphic representation of the kinetic development of resolving and persistent inflammation in the human monocyte-based in vitro models. (A) Freshly isolated human blood monocytes were first exposed to the chemokine CCL2 for 2 h at 37°C with 1% serum in normoxic conditions, then, after washing, to GMMA (from 2 h), TNF-α (from 3 h, without washing), and IFN-γ (from 7 h, without washing) at 39°C with 5% serum in hypoxic conditions. At 14 h, the inflammatory stimuli were washed off, temperature and serum concentration brought back to 37°C and 1%, respectively, and fresh medium containing IL-10 added. At 24 h, IL-10 was washed off and monocytes were exposed to TGF-β until the end of the experiment. (B) Freshly isolated human blood monocytes were first exposed to the chemokine CCL2 for 2 h at 37°C with 1% serum in normoxic conditions, then, after washing to LPS, PDG, poly(I:C), and CpG (from 2 to 7 h) at 39°C with 5% serum in hypoxic conditions. At 7 h, the inflammatory stimuli were washed off, and fresh medium containing ACPA complexes, GM-CSF, M-CSF, Survivin, and IFN-γ was added. The temperature was kept to 39°C, serum at 5% and oxygen tension at hypoxic levels until the end of the experiment. Cells were harvested at 0, 2, 4, 14, 24, and 48 h for the resolving model, and at 0, 2, 4, 14, 24, 72, and 96 h in the persistent model. Supernatants were collected at the same time points, except for time 0.