Implication of mitochondria-derived reactive oxygen species, cytochrome C and caspase-3 in N-(4-Hydroxyphenyl)retinamide-induced apoptosis in cervical carcinoma cells

Abstract

N-(4-Hydroxyphenyl)retinamide (4HPR) is currently used in cancer prevention and therapy trials. It is thought that its e ects result from induction of apoptosis. 4HPR-induced apoptosis in human cervical carcinoma C33A cells involves enhanced generation of reactive oxygen species (ROS). In this study we explored the mechanism by which 4HPR increases ROS and induces apoptosis in these cells. 4HPR induced cytochrome c release from mitochondria to cytoplasm, activated caspase-3, and caused a membrane permeability transition (MPT). All these 4HPR's e ects, as well as the induction of apoptosis, were inhibited by antioxidants, which decrease ROS. Thenoyltri¯uoroacetone, a mitochondrial respiratory chain (MRC) complex II inhibitor, and carbonylcyanide m-chlorophenyl hydrazone, which uncouples electron transfer and ATP synthesis and inhibits ROS generation by MRC, inhibited 4HPR-induced ROS generation very e ectively. Rotenone, an MRC complex I inhibitor was less e ective and azide, an MRC complex IV inhibitor, exhibited a marginal e ect. In contrast, antimycin A, an MRC complex III inhibitor, enhanced 4HPR-induced ROS generation. These ®ndings suggest that 4HPR enhances ROS generation by a ecting a target between complex II and complex III, presumably coenzyme Q. This e ect is followed by release of cytochrome c, increased caspase-3 activity, induction of MPT and eventual DNA fragmentation and cell death.

N-(4-hydroxyphenyl) retinamide (4HPR) enhances TRAIL-mediated apoptosis through enhancement of a mitochondrial-dependent amplification loop in ovarian cancer cell lines

“…4HPR has been described to induce MPT through different mechanisms, which included both ROS and ceramide generation. [46][47][48] However, inhibition of ROS or ceramide production in our cells did not block the apoptosis induced by 4HPR and TRAIL.…”

“…[46][47][48] TRAIL induces changes in mitochondrial permeability through a different mechanism which involves translocation of the cleaved p15 form of BID into the mitochondrial membrane. As a marker of ROS generation, measurement of DCFH-DA by flow cytometry was used.…”

All-trans-retinoic acid induces manganese superoxide dismutase in human neuroblastoma through NF-κB

“…Retinoids have been reported to increase ROS including superoxide and peroxides in AML cell lines [4,[27][28]. The specific source of ROS in response to ATRA is yet to be determined however, studies by Suzuki et al [29] using the synthetic retinoid, N-(4-hydroxyphenyl) retinamide, showed mitochondrial-mediated ROS generation in cervical carcinoma cells. Also, recent reports have suggested that ROS can modulate signal transduction and activate the NFκB pathway, thereby regulating the process of cell survival among others [30].…”

pRb and Cdk regulation by N-(4-hydroxyphenyl)retinamide

“…Accordingly, while apoptosis by HPR appears receptor-independent (Delia et al, 1993;Kitareewan et al, 1999), and actually correlates with the ability of HPR to elicit intracellular free radicals and acidi®cation (Delia et al, 1997a;Angoli et al, 1996;Maurer et al, 1999;Suzuki et al, 1999), transcriptional regulation of target genes (e.g. AP1) is retinoid-receptor dependent, according to data showing that HPR selectively activates the transcription by RAR-g, to a less extent by RAR-b, but not by RAR-a (Fanjul et al, 1996;Kazmi et al, 1996).…”