spacer gif spacer gif spacer gif spacer gif spacer gif
QUICK SEARCH:   [advanced]


spacer gif
     Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents    

Right arrow Help viewing high resolution images
Right arrow Return to article
(Downloading may take up to 30 seconds.
If the slide opens in your browser, select File -> Save As to save it.)
Click on image to view larger version.


Figure 2


Fig. 2. Rotenone and TTFA induce autophagy. HEK 293 and U87 cells were treated as described in Fig. 1. (A) The percentage of cells with AVOs (autophagosomes and autolysosomes) was determined by flow cytometry. Rates of AVO formation induced by rotenone (R) and TTFA (T) are indicated in (i) HEK 293 and (ii) U87 cells over a 72-hour time course. (iii) Effect of 3-MA (2.0 mM) on the formation of AVOs that were induced by rotenone or TTFA after a treatment of 48 hours in U87 and HEK 293 cells. P values less than 0.05 represent significant difference between conditions, as indicated. (B) Electron-microscopy pictures were taken of HEK 293 cells that were untreated (control) or treated with TTFA (0.5 mM) for 48 hours. Arrows represent double-membrane vacuoles in TTFA-treated cells (enlarged image). N represents the nucleus. (C) Formation of GFP-LC3-labeled vacuoles (dots). The percentage of cells with GFP-LC3-labeled vacuoles (dots) in (i) HEK 293 and (ii) U87 cells following rotenone or TFFA treatment over a 48-hour time course was determined. Error bars represent s.e. of three independent experiments. (iii) Representative pictures from three independent experiments of U87 cells treated with GFP alone; GFP-LC3 alone; GFP-LC3 and rotenone; and GFP-LC3, rotenone and 3-MA (2.0 mM) were captured by an Olympus microscope and coolsnap camera. The nucleolus was stained with DAPI (blue). (iv) HEK 293 and U87 cells were treated with rotenone or TTFA in the presence or absence of 3-MA (2.0 mM). (D) The conversion of LC3-I to LC3-II was determined in (i) HEK 293 and (ii) U87 cells treated with rotenone or TTFA for 6, 16 or 24 hours in the presence or absence of the lysosomal inhibitor NH4Cl (30 mM). Cells were lysed and western blotted for expression of LC3. Blots were stripped and re-probed with anti-actin antibody for equal loading. (E) Beclin 1 expression was determined after rotenone and TTFA treatment in HEK 293 and U87 cells after 48 hours of treatment. The cells were lysed and western blotted for beclin 1 and actin was used as a loading control.





Right arrow Return to article