Hello, I am looking to test some materials that may or may not have an effect on how the polarization of marcophage happens. My question is whether to take the Thp-1 cells into M0 phase before testing the materials or just test the material with the THP-1 cells as such.
try to make your cells different groups, treated groups with the materials that you are going to use and compare with a known polarizing agent then compare both with non- treated cells, i was using PMA at concentration of 50ng/ml of 5x105 cells/ml for 48h at 37c
Dear Vignesh,
I have worked for the past four years with the THP-1 cell line, and although I consider it to be a good model for M0/M1 polarization, it might not be the best model for the study of M2-like phenotypes (for instance, have a look at polarization markers and cytokines obtained in my laboratory upon incubation of resting macrophages with IL-4 and IL-13 http://onlinelibrary.wiley.com/doi/10.1002/adhm.201601012/abstract). Obviously, many aspects of the polarization will depend on the original M0 differentiation protocol and importantly on cell culture timing, but do have a look at different papers online and try and screen for similarities and discrepancies. Having said that, I used myself 50 ng/mL PMA for 24h and then polarized cells with either a M1 or M2 polarization cocktail for a further 24h (if you want to focus on M2 polarization, I would use a longer differentiation time with a lower concentration of PMA). If you are testing the effect of different materials on the M1 or M2 polarization, then I would suggest first differentiating the premonocytes into M0 cells, then polarizing in the presence or absence of your materials of choice to check for their effect on cell phenotypes in terms of morphology, cell markers and cytokines. If you would like to use your materials as growth substrates instead of soluble or colloidal materials, then I would spin said materials in culture slides and try and proceed with the differentiation and polarization on these materials as suggested, comparing all conditions with conventional plastic culture dishes/well plates. However, bear in mind that the PMA differentiation might be affected by the presence of different substrates upon which THP-1 cells adhere to, and also by the presence of specific molecules or molecule scavengers in medium.
Let us know if you need any further help and good luck with the THP-1 cell line - definitely not the easiest cell type to work with!
Best,
Julio.
Dear Julio,
Thank you for the infromation. I just had a look at your paper this morning.
Cheers
Vignesh
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