Spontaneous caspase 3 and 9 activation by estradiol and progesterone at 24 hours using fluorescent caspase substrates. Cell lysates were prepared from 10 × 106 cells using caspase isolation and incubation buffers. Aliquots of the lysates were diluted in caspase incubation buffer and Ac-DEVD-AMC (caspase 3; panel A) or Ac-LEHD-AMC (caspase 9; panel B). The release of AMC fluorescent tag was measured using a cytofluorometer at 0 and 1 hour and specific activity was measured as the difference between 0 and 1 hour results and expressed as caspase activity per microgram protein. Each graph represents at least 5 separate experiments performed in duplicate (mean ± SEM). *P < .05 versus control at 0 hours; **P < .05 versus control at 24 hours. (A) Men, n = 2; women, n = 3. (B) Men, n = 3; women, n = 3.
