Grb2 silencing decreases phagocytic uptake in RPE cells.

<p>RPE-J cells were transfected with a pool of Grb2 targeting siRNAs or a non-targeting siRNA control, and expression was evaluated 5 days later. (A) Transcript levels for <i>Grb2</i>, <i>Mertk</i>, and <i>β-actin</i> evaluated using RT-PCR. (B) Protein levels of Grb2, Mertk, and β-actin evaluated by western blot analysis of RPE-J cell homogenates. (C-D) Phagocytic activity assays. (C) Confocal images of representative fields of RPE-J cells showing ingested OS labeled in red, and DAPI-stained nuclei shown in blue. Cells were incubated with AlexaFluor 555-labeled bovine rod OS for 4 h and quenched by addition of trypan blue. (D) OS ingestion and binding were quantified for three independent assays and plotted as a percentage of control total OS after 4 h that was set as 100%. Error bars represent mean ± SEM, n = 3. P values were calculated using Student’s <i>t</i> test, and are as shown: p**<0.05, p****<0.00005.