FIGURE 9:

Rac1 activation during POS phagocytosis by RPE cells in culture is independent of TAM receptor ligands. RPE-J cells or MerTK-deficient RCS rat primary RPE received POS suspended in DMEM without additives (/) or supplemented with Gas6 (G), MFG-E8 (M), a mix of Gas6 and MFG-E8 (M+G), or FBS (FBS) for 3.5 h or 1.5 h, respectively, before harvest. (A) Immunoblotting of whole-cell lysates showed no differences in TAM receptor or Rac1 protein levels upon challenge with different additives. (B and C) G-LISAs showed that GTP-Rac1 levels of RPE-J (B) and RCS RPE (C) increased to the same extent relative to untreated cells after POS challenge without additives (white bars) or with Gas6 (black bar, G). POS with MFG-E8 alone (light gray bars) raised GTP-Rac1 to the same extent as MFG-E8 plus Gas6 (black bar, M+G). FBS also increased GTP-Rac1 by both cell types (dark gray bar). Bars represent mean ± SD of three independent experiments, each testing duplicate samples; n. s., not significant (p > 0.1).