TGFβ enhances levels of MAP1S protein and autophagy flux in different pancreatic cell lines.

<p>A) A real-time RT-PCR analysis of the impact of TGFβ on the expression of <i>MAP1S</i> gene in pancreatic cancer cell line PANC-1 and Capan-2. The mRNA levels in untreated samples were set as 1. B) An immunoblotting analysis of the protein levels of MAP1S and LC3 in untreated or TGFβ-treated PANC-1 and Capan-2 cells in the absence (Ctrl) or presence of lysosomal inhibitor Bafilomycin A1 (BAF). Lysates with the same amount of total proteins were loaded in each lane and β-Actin served as another loading control. C,D) Plots of relative levels of MAP1S (C) or LC3-II (D) as shown as a representative in panel (B). The levels of MAP1S in untreated cells were set as 1. Results are the means ± S.D. of at least three repeats and the differences were compared using Student’s t test. **, P≤0.01; and *, P≤0.05. E) A fluorescent imaging analysis of the impact of TGFβ on PANC-1 cells transiently transfected with a plasmid for 48 hrs to express GFP–LC3 in the absence (Ctrl) or presence of TGFβ and/or BAF for 12 hrs. F) A quantification of GFP–LC3-labelled autophagosomes as shown in panel (E). The data were the average number of GFP-LC3 punctate foci ± S.D. for ten randomly selected images with size of 512 pixels×512 pixels for each that covers about 10 cells on average. The significance of differences was determined by Student’s t test. ***P≤0.001.