Trifolin acetate-induced cell death in human leukemia cells is dependent on caspase-6 and activates the MAPK pathway

Abstract

In the present study we demonstrated that the flavonoid derivative trifolin acetate (TA), obtained by acetylation of naturally occurring trifolin, induces apoptosis. Associated downstream signaling events were also investigated. TA-induced cell death was prevented by the non-specific caspase inhibitor z-VAD-fmk and reduced by the presence of the selective caspase inhibitors z-LEHD-fmk (caspase-9), z-DEVD-fmk (caspase-3) and z-VEID-fmk (caspase-6). The apoptotic effect of TA was associated with (i) the release of cytochrome c from mitochondria which was not accompanied by dissipation of the mitochondrial membrane potential (DeltaPsi(m)), (ii) the activation of the mitogen-activated protein kinases (MAPKs) pathway and (iii) abrogated by the over-expression of Bcl-2 or Bcl-x(L). TA-induced cell death was attenuated by inhibition of extracellular signal-regulated kinases (ERK) 1/2 with U0126 and inhibition of p38(MAPK) with SB203580. In contrast, inhibition of c-Jun NH(2)-terminal kinase (JNK) by SP600125 significantly enhanced apoptosis. Although reactive oxygen species (ROS) increased in response to TA, this did not seem to play a pivotal role in the apoptotic process since different anti-oxidants were unable to provide cell protection. The present study demonstrates that TA-induced cell death is mediated by an intrinsic-dependent apoptotic event involving mitochondria and MAPK, and through a mechanism independent of ROS generation.

Reactive oxygen species: A double-edged sword in oncogenesis

“…However, in p38-related apoptosis that is independent of ROS generation, JNK seems to execute a reverse function. Inhibition of JNK by SP600125 significantly enhanced apoptosis [57] . Via regulation of MAPKs, the protein kinase C (PKC)δ-mediated pathway also involves ROS related apoptosis.…”

Compound A398, a Novel Podophyllotoxin Analogue: Cytotoxicity and Induction of Apoptosis in Human Leukemia Cells

“…Since the HL-60 cells are p53-deficient [54], [55], A398 operates independently of the activation of this protein. So, this compound may also be effective in other tumor cells that have no functional p53, which is very important since the mutation and inactivation of this protein occurs in more than half of all types of cancer, and is correlated with the degree of aggressiveness of some tumors [56], [57].…”

Eupatorin-Induced Cell Death in Human Leukemia Cells Is Dependent on Caspases and Activates the Mitogen-Activated Protein Kinase Pathway

“…Fluorescent microscopy, flow cytometric analysis of propidium iodide-stained nuclei and DNA fragmentation assay were performed as described [13]. Apoptosis was also determined by translocation of phosphatidylserine to the cell surface using the annexin V-fluoresceine isothiocyanate (FITC) apoptosis detection kit (BD Pharmingen) according to the manufacturer's protocol.…”

“…Immunoblot analysis of Bcl-2 family members, caspases, cytochrome c , MAPKs and PARP was performed as previously described [13]. Purity of mitochondrial and cytosolic fractions was determined by probing with antibodies to cytochrome c oxidase and β-actin, respectively.…”