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. 2021 Jul 21;11:14849. doi: 10.1038/s41598-021-94360-9

Figure 5.

Figure 5

Targeting CD276 on macrophages with specific antibodies induces distinct immunoreceptor phosphorylation accompanied with kinase phosphorylation. (A) Monocytes were cultured either in GM-CSF or M-CSF supplemented medium for 7 days. CD276 expression on monocyte-derived macrophages (MDMs) was analyzed by flow cytometry and CD276 MFI values from six paired samples are summarized in the graph. (B-E) After 7 days culture, MDMs were reseeded for Phospho-Immunoreceptor and Phospho-kinase array. After 3 h resting, MDMs were treated with αCD276 antibody or isotype for 10 min for Phospho-immunoreceptor array and 30 min for Phospho-Kinase array. Cell lysates were processed for phosphorylation signatures. In total macrophages from 4–5 donors were tested. Two representative donors are shown for Phospho-Immunoreceptor arrays: (B) M-CSF MDMs and (C) GM-CSF MDMs. Two representative and matching donors are shown for Phospho-Kinase arrays: (D) M-CSF MDMs and (E) GM-CSF MDMs. (F) MDMs (GM-CSF) were left untreated or treated with LPS + IFNγ in the presence of αCD276 antibody or isotype control for 24 h. The graphs summarize the concentration of PAI-1 in the supernatants. Each dot represents one donor.