Enhancement of Radiation-induced Apoptosis by 6-Formylpterin

Abstract

Radiation-induced apoptosis and its possible enhancement in the presence of 6-formylpterin (6-FP), a metabolite of folic acid, were examined in human myelomonocytic lymphoma U937 cells. When cells were treated with 6-FP at a nontoxic concentration of 300 μM, and then exposed to X-rays at a dose of 10 Gy, significant enhancement of radiation-induced apoptosis as determined by nuclear morphological change, phosphatidylserine (PS) externalization and DNA fragmentation were observed. Flow cytometry for the detection of intracellular hydrogen peroxide (H²O²) revealed that 6-FP increased the formation of intracellular H²O², which further increased when the cells were irradiated. Decrease of mitochondria trans-membrane potential (MMP), release of cytochrome c from mitochondria, and activation of caspase-3 were enhanced after the combined treatment. Remarkable activation of protein kinase C δ (PKC δ) and its translocation from cytosol to mitochondria were detected in combined treatment. Increase of intracellular Ca²⁺ concentrations ([Ca²⁺]i) was also observed, however, neither calpain I nor calpain II could inhibit the apoptosis. In addition, c-Jun NH²-terminal kinase ( JNK) activation was not enhanced in the combined treatment. A protein involved in a caspase-independent apoptosis pathway, apoptosis inducing factor (AIF), remained unchanged even 3 h after treatment. These results indicate that intracellular H²O² generated by 6-FP enhances radiation-induced apoptosis via the mitochondria-mediated caspase-dependent pathway, with the active involvement of PKC δ.

• Radiation
• Apoptosis
• 6-Formylpterin
• ROS
• PKC δ