Exploring the Maturation of a Monocytic Cell Line Using Self-organizing Maps of Single-cell Raman Spectra

Phorbol myristate acetate (PMA)-differentiated THP-1 cells are routinely used in lieu of primary macrophages to study macrophage polarization during host-pathogen interactions and disease progression. The phenotypes of THP-1 macrophages are influenced by the level and duration of PMA stimulation and possibly also by the presence of adhesion factors. Here, we use self-organizing maps (SOMs) of single-cell Raman spectra to probe the effects of PMA stimulation conditions and adhesion factors on THP-1 cell differentiation. Raman spectra encoding for biochemical composition were acquired from individual cells on substrates coated with fibronectin or poly-l-lysine before and after stimulation with 20 or 200 nM PMA for two different time intervals. SOMs constructed from these spectra showed the extent of spectral dissimilarity between different chronological cell populations. For all conditions, the SOMs indicated that the spectra acquired from cells after three-day treatment had diverged from those of untreated cells. The SOMs also showed that the higher PMA concentration produced both fully and partially differentiated cells for both adhesion factors after three days, whereas the outcome of stimulation for three days with the lower PMA concentration depended on the adhesion factor. On poly-l-lysine, treatment with 20 nM PMA for three days induced an intermediate stage of differentiation, but the same treatment produced partially and fully differentiated cells when applied to THP-1 cells on fibronectin. These results are consistent with the modulation of the transition of THP-1 monocytes into macrophage-like cells by integrin-binding interactions. Furthermore, differences in culture and stimulation conditions may confound comparison of results from separate studies.