Figure 5.

2A4 promotes phagocytosis of AL amyloidosis heart extracts by a human macrophage cell line in vitro. (A) 2A4 promoted clearance of the extracellular AL heart deposits by THP-1 macrophages. Clearance was measured by area of extracellular LC protein deposits, immunostained for λ LC, remaining after 24 h and was significantly increased by 2A4 treatment (gray bar) compared with an isotype control antibody (black bar). Data are presented as mean ± SEM (n = 3). (B, C) 2A4 induced internalization of pHrodo-conjugated AL heart deposits by THP-1 cells, visualized as an increase in intracellular pHrodo-fluorescence shown in a representative bright-field overlay (20×; B) or quantified as the percentage of THP-1 cells with elevated relative mean fluorescence intensity (C), which represents internalized pHrodo-labeled aggregates. Values represent mean ± SEM of 60 optical fields per group, pooled from three independent wells (Figure 5A) or of three independent flow cytometry runs with approximately 1000 cells per run (Figure 5C). Statistical differences were determined by 2-tailed Student t test. Scale bar = 100 μm.