Proteomics characterisation of the L929 cell supernatant and its role in BMDM differentiation

• Published in: Life science alliance Vol. 4; no. 6; p. e202000957
• Main Authors: Heap, Rachel E, Marín-Rubio, José Luis, Peltier, Julien, Heunis, Tiaan, Dannoura, Abeer, Moore, Adam, Trost, Matthias
• Format: Journal Article
• Language: English
• Published: United States Life Science Alliance LLC 01.06.2021
• Subjects: Animals; Cell Culture Techniques - methods; Cell Differentiation - drug effects; Culture Media, Conditioned - chemistry; Culture Media, Conditioned - pharmacology; Humans; Macrophage Colony-Stimulating Factor - metabolism; Macrophage Colony-Stimulating Factor - pharmacology; Macrophage Migration-Inhibitory Factors - metabolism; Macrophages - drug effects; Macrophages - metabolism; Macrophages - physiology; Proteomics - methods; Resource; Resources
• ISSN: 2575-1077; 2575-1077
• DOI: 10.26508/lsa.202000957

BMDMs are a key model system to study macrophage biology in vitro. Commonly used methods to differentiate macrophages from BM are treatment with either recombinant M-CSF or the supernatant of L929 cells, which secrete M-CSF. However, little is known about the composition of L929 cell-conditioned media (LCCM) and how it affects the BMDM phenotype. Here, we used quantitative mass spectrometry to characterise the kinetics of protein secretion from L929 cells over a 2-wk period, identifying 2,193 proteins. Whereas M-CSF is very abundant in LCCM, we identified several other immune-regulatory proteins such as macrophage migration inhibitory factor (MIF), osteopontin, and chemokines such as Ccl2 and Ccl7 at surprisingly high abundance levels. We therefore further characterised the proteomes of BMDMs after differentiation with M-CSF, M-CSF + MIF, or LCCM, respectively. Interestingly, macrophages differentiated with LCCM induced a stronger anti-inflammatory M1 phenotype that those differentiated with M-CSF. This resource will be valuable to all researchers using LCCM for the differentiation of BMDMs.