Mitochondria- not death receptor–mediated caspase activation is essential for triptolide-induced cell death. (A) MEFs, MEFs deficient in caspase-9 (MEF-caspase-9–/–), Jurkat cells, and Jurkat cells deficient in caspase-8 (JurkatI2.1) were treated with the indicated concentrations of triptolide for 24 hours. Cell death was determined by annexin V staining with PI. (B) Measurement of cytosolic cytochrome C by Western blot of OCI-AML3 cells treated with triptolide for 24 hours. (C) Determination of MMP by CMXRos-MitoTracker Green staining and flow cytometry analysis of OCI-AML3 cells treated with triptolide for 24 hours. Area I represents the cells with intact mitochondria; area II; cells with partial loss of MMP; and area III, complete loss of MMP. (D) OCI-AML3vec and OCI-AML3Bcl-2 cells at a density of 0.2 × 106/mL were treated for 24 hours with various concentrations of triptolide. Cell death was determined by annexin V staining with PI. (E) Mcl-1 levels in OCI-AML3, U937, and HL-60 cells and cells from patients with AML (n = 6) treated with triptolide for 24 hours determined by Western blot.
