Fig 2.

Inhibition of gemcitabine-induced autophagy by verteporfin. (A-C) MCF-7 EGFP-LC3 cells were exposed to 500nM gemcitabine (Gem) for 24h in the presence or absence of 10µM verteporfin, or to 0.1% DMSO vehicle control. Cells were fixed and stained with Hoechst 33342 and punctate EGFP-LC3 fluorescence was (A) visualized and (B) quantified using a Cellomics ArrayScan VTI automated fluorescence microscope. (* p < 0.01, Student's t-test)(mean ± S.D (error bars), n=3). (C) MCF-7 EGFP-LC3 cells were exposed to 0.1% DMSO, 500nM gemcitabine, or 30nM rapamycin, a known stimulator of autophagy, for 24h in the absence or presence of 10µM verteporfin. Lysates were collected and immunoblotted for EGFP. (D) BxPC-3 cells were exposed to 500nM gemcitabine for 24h in the presence or absence of 10µM verteporfin, 100nM bafilomycin A1, or 0.1% DMSO vehicle control. Lysates were collected and immunoblotted for LC3 and p62. β-tubulin was monitored as a loading control.