Feijoa F3 does not activate autophagy or inhibit NF-κB activity in autophagy defective HCT116 cells.
2015-06-25T03:37:33Z (GMT) by
<p>Cells were treated with media, Torin1 (1μM/ml), solvent and feijoa F3 (10mg/ml), in the presence and absence of 100ng/ml PAM3CSK4 stimulation. <b>A and B</b> show the number of autophagosomes in each cell. The puncta were quantified using the spot function in Imaris. Each experimental condition for solvent, media, and feijoa F3 had 3 biological replicates with two technical replicates (giving a maximum of 6 data points) and Torin1 had 2 biological replicates with two technical replicates (giving a maximum of 4 data points). Horizontal line represents median. <b>B</b> is an image representation of <b>A</b>. The spots in the cells are the spheres pseudo colored by the Imaris spot function representing the autophagosome puncta that were stained for by LC3B. <b>C.</b> shows western blot analysis of autophagy. The westerns show LC3B I (16kDa) and II (18kDa) protein bands. β-actin (42 kDa) was used as a control. The bottom graph is the semi quantification of the LC3 I and II band intensity scores, adjusted against β-actin band intensity scores. <b>D</b> shows the normalized percentage NF-κB activation. Percentage inflammation was calculated using <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0130910#pone.0130910.e003" target="_blank">Eq 3</a>. A percentage of greater than 100 shows increased inflammatory response in the presence of the ligand when compared to media (no treatment); and a percentage of less than 100 suggests a decrease in the inflammatory response in the presence of the ligand when compared to media (no treatment) meaning that a condition has inhibited NF-κB signaling. Error bars represent standard deviation of (n = 6).</p>