Research Paper Volume 2, Issue 12 pp 924—935

DNA damaging agents and p53 do not cause senescence in quiescent cells, while consecutive re-activation of mTOR is associated with conversion to senescence


Figure 3. Effects of rapamycin and serum starvation on etoposide-induced senescence in WI-38t cells. A-C. WI-38t cells were plated at 5000/well in 12 well plates, and the next day either treated with 10 nM rapamycin in complete medium (R), or placed in serum-free medium (no serum or 0), or left in complete medium (control). The next day, 1 μg/ml etoposide (Et) was added, as indicated.

A. After 4 days, cells were stained for beta-Gal and microphotographed (bar - 50 micron).

B. After 4 days, cells were counted: control (C), rapamycin (R), no serum (0).

C. Proliferative potential. In replicate plates, cells were washed and incubated in complete, drug-free medium for 6 days and then counted (black bars). Note: red bars correspond to red bars in panel B. Fold (f) increase in a cell number after drug removal.

D. Immunoblot. Cells were plated in 6 well plates. The next day, cells were treated with 1 μg/ml etoposide (Et) for 24 hrs: control -C, rapamycin - R, no serum −0.