Hi everyone, I am trying to induce macrophage polarization using THP-1 cells as follows:
1. Cell seeding
2. Incubation with PMA for 24 hours
3. Change to PMA-free medium and let cells rest for 72 hours
4. Treat with LPS + IFN-gamma 20ng/ml for M1 macrophage and IL-4 & IL-13 20ng/ml for M2 macrophage
By the end of step 3 my cells were alive and healthy and so are M2 macrophage cells. However, in less than 24 hours after step 4, I noticed there were many dead cells in my M1 polarization condition. I have tried to reduce the [LPS] (250ng/ml and 100ng/ml) but still received the same results. Anyone else have had similar problems and how do you improve the survival of M1 macrophage cells? Thanks a lot!
You could try PMA-mediated differentiation (150nM) for 48 hours, and let them rest for 24 hours, followed by differentiation (100 ng/ml LPS) + IFNy (20 ng/ml) for 48 hours. There will still be cell death, but its likely to be less because of the proinflammatory cytokines in the media
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