Figure 2. Apoptosis increases the expression of functional TREM2-L on neuronal and non-neuronal cells.

(A) TREM2-L expression was quantified by flow cytometry before (left) and after (right) induction of apoptosis in Neuro2A cells by staurosporine (sts) or MPP⁺. The median fluorescence intensity (MFI) of TREM2-Fc or TREM1-Fc binding of each gate is shown. Apoptotic Neuro2A cells (Annexin V^hi) express 3-10 fold greater levels of TREM2-L compared to Annexin V^lo Neuro2A cells, but do not express TREM1-L (n=7). (B) TREM2-L expression is increased on non-neuronal cells during apoptosis. BWZ, WEHI-231, and P388D1 cells bind to TREM2-Fc, but not TREM1-Fc. Treatment with staurosporine boosts TREM2-Fc binding to Annexin V^hi cells by 7-10 fold (n=3). (C) Neuronal cells activate TREM2/DAP12 signal transduction as assessed by BWZ.TREM2/DAP12 reporter cells. Healthy Neuro2A cells induce a modest level of cellular activation in the BWZ.TREM2/DAP12 cell line, and this is increased to near-maximal levels when the Neuro2A are treated with MPP⁺ to induce apoptosis (top panel). Pretreating the BV2 cells with a blocking TREM2 mAb (black bars), but not with a control rat IgG1 mAb (gray bars), significantly reduces cellular activation in response to both untreated and apoptotic Neuro2A. Ventral midbrain neurons (VMN) elicit similar responses (third panel), while cortical neurons (CN) demonstrate little stimulation unless they are apoptotic (second panel). BWZ.TREM2/DAP12 reporter cell activation in response to primary neurons was effectively blocked with the TREM2 mAb. None of the neuronal cells activate the parental BWZ reporter cell line (representative results for VMN are shown in the bottom panel). *p-value <0.05, **p-value < 0.005, ***p-value < 0.0005.