I want to study the differentiation of macrophages into M1 and M2 macrophages populations against a bacterial protein. I'm using THP1 monocytes as a model. So, is it necessary to differentiate the cells with PMA or LPS. PMA differentiates monocytes to macrophages. So, if I would further treat them with a bacterial protein, will they differentiate into M1 or M2 further? or it will be ok, if I stimulate them with LPS and then treat with bacterial protein and analyze for M1 and M2 differentiation?
To look at macrophages rather than monocytes you would definitely need to treat first with PMA. This treatment will give you "M0" macrophages. Following PMA, LPS/IFNg treatment will give you "M1" macrophages, while treatment with IL-4/IL-13 will polarize to "M2" macrophages. Given all of that, if you want to look at how the bacterial protein polarizes macrophages, I would differentiate the monocytes to macrophages with PMA, then treat with the protein to look at markers of "M1" or "M2" (and use LPS/IFNg treatment as a control for "M1" polarization and IL-4/IL-13 treatment as a control for "M2" polarization. If you want to look at the effect of the bacterial protein on repolarizing "M1" or "M2" macrophages, you would need to differentiate with PMA and then differentiate to "M1" or "M2" with LPS/IFNg or IL-4/IL-13, respectively before treating with the bacterial protein. Good luck!
Dear Sandeep Kumar
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Hello Sandeep Kumar
I work in monocyte-derived of macrophage and THP-1 too in the investigation of control of innate inmunity. For THP-1 cell line, you should need use PMA as stimulus of cell differentiation and after polarizing into M1 or M2. However, you should consider, PMA actives some inflammatory pathwways and maybe when you want to study M1 profile, some pathways or genes could be already activated and over stress your cell model.
I advice to you a period of arrest without PMA. Previously, assess differentiation and inflammatory markers with or without PMA during follow-up period of in vitro experiment.
Best regards
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