Peroxynitrite-induced Apoptosis In FaDU Cells Is Correlated With The Activities Of PDCD4, DDR1 And DDR2 Genes

Peroxynitrite (ONOO-), a product of nitric oxide and superoxide anion interaction, is a potent and versatile oxidant implicated in a number of pathophysiological processes. Mounting evidence suggests that toxicity of nitric oxide is mainly attributable to the production of ONOO- rather than itself. In fact, ONOO-can diffuse freely across phospholipid membrane bilayers to affect cell metabolism by inducing lipid peroxidation, interfering with mitochondrial function, damaging DNA, and mediating necrosis and apoptosis in different cell types, including cancer cell. Hypopharyngeal carcinoma, which derives from a subsite of the upper aerodigestive tract, is one of the most common cancers in head and neck. Although numerous treatment approaches have been employed, the long-term survival rates of patients suffering from the disease have been improved only minimally during the past two decades. Therefore, studies of the potential new drug targets of the tumor can not only get insight into the mechanisms underlying the pathogenesis of hypopharyngeal carcinoma, but, in turn, offer effective therapeutics as well. Recently, programmed cell death 4, PDCD4, which is identified as a novel tumor suppressor gene, has generated considerable interest in tumor biology. PDCD4 was originally found as a tumor related gene in humans and mapped to chromosome 10q24. PDCD4 protein is known to bind eukaryotic initiation factor 4A that inhibits translation initiation. PDCD4 is ubiquitously expressed in normal tissues, but, lost or suppressed expression of PDCD4 in several tumors that correlates with tumor progression and poor prognosis. Notably, it has been found that PDCD4 is up-regulated during apoptosis in several cell types. To date, however, the expression of PDCD4 in ONOO- induced apoptosis of the FaDU cells is unknown.Discoidin domain receptors (DDRs), the nonintegrin-type receptors for collagen, consist of two closely related types, i.e., DDR1, which mainly is mainly expressed in tumor cells, and DDR2, which is highly expressed in mesenchymal cells. Therefore, the present study was designed to investigate whether FaDU cells experienced apoptosis following treatment with ONOO-, with special attention given to the induction of PDCD4, DDR1 and DDR2 activities in this process.In vitro cultured FaDu cells were subjected to various concentrations of ONOO-, then, the rate of cell apoptosis was measured by flow cytometry.The cell viability was quantified by MTT assay. The morphological changes were observed by acridine orange cytochemistry staining and Ho33342 and PI double staining to determine whether cell experienced apoptosis under the stress of ONOO-. The protein expressions of PDCD4, DDR1 and DDR2 were examined by western blot in response to ONOO-. The mRNA expressions of PDCD4, DDR1 and DDR2 were analyzed by RT-PCR. We demonstrated that ONOO- decreased the survival rates of FaDu cells in a concentration-dependent manner, implying that ONOO- was able to inhibit the growth of FaDu cells in vitro. Under the stress by ONOO-, the expression of PDCD4 gene was apparently increased at both mRNA and protein levels. The expressions of DDR1 and DDR2 genes were finally decreased at both mRNA and protein levels.In conclusion, these data suggested that ONOO- can effectively suppress proliferation of FaDu cells via the induction of apoptosis. Our findings demonstrate that PDCD4, DDR1 and DDR2 gene may play an important role in ONOO--induced apoptosis in FaDu cells, which may offer a new target for the treatment of hypopharyngeal carcinoma.