Nickel Ion Inhibits Nuclear Factor-Kappa B Activity in Human Oral Squamous Cell Carcinoma

Abstract

BackgroundThe spontaneous IL-8 secretion observed in OSCC is partially dependent on the disregulated activity of transcription factor NF-κB. Nickel compounds are well established human carcinogens, however, little is known about the influence of nickel on the spontaneous secretion of IL-8 in oral squamous cell carcinoma (OSCC) cells. The aim of the present study was to investigate whether Ni2+ ions can influence on IL-8 secretion by OSCC.Methods and ResultsThe IL-8 secretion was measured by ELISA. The expression of IL-8 mRNA was examined by real-time PCR. The NF-κB activity was measured by luciferase assay. The phosphorylation status and nuclear localization of NF-κB subunits were examined by Western blotting or Transfactor kit and immunofluorescence staining, respectively. The interaction of NF-κB p50 subunit and Ni2+ ions was examined by Ni2+-column pull down assay. The site-directed mutagenesis was used to generate a series of p50 mutants. Scratch motility assay was used to monitor the cell mobility. Our results demonstrated that, on the contrary to our expectations, Ni2+ ions inhibited the spontaneous secretion of IL-8. As IL-8 reduction was observed in a transcriptional level, we performed the luciferase assay and the data indicated that Ni2+ ions reduced the NF-κB activity. Measurement of p50 subunit in the nucleus and the immunofluorescence staining revealed that the inhibitory effect of Ni2+ ions was attributed to the prevention of p50 subunit accumulation to the nucleus. By Ni2+-column pull down assay, Ni2+ ions were shown to interact directly with His cluster in the N-terminus of p50 subunit. The inhibitory effect of Ni2+ ions was reverted in the transfectant expressing the His cluster-deleted p50 mutant. Moreover, Ni2+ ions inhibited the OSCC mobility in a dose dependent fashion.ConclusionsTaken together, inhibition of NF-κB activity by Ni2+ ion might be a novel therapeutic strategy for the treatment of oral cancer.

Ouabain signaling in oral squamous cell carcinoma cells

“…The human OSCC cell line HSC3 (obtained from the Japanese Collection of Research Bioresources [JCRB] Cell Bank, Ibaraki, Japan) was maintained as described previously (14).…”

“…The luciferase reporter vector encompassing the IL-8 untranslated region (UTR) was described previously (14). This vector contains two NF-κB-binding sites, and the shorter fragment, which contains only the lower binding site, was amplified and subcloned to the pGL4 basic vector (NF-κB-short vector) to eliminate the upper binding site.…”

Nickel chloride administration prevents the growth of oral squamous cell carcinoma

“…NF-κB protein levels increase gradually from premalignant lesions to invasive cancer, indicating its role during the early stages of carcinogenesis [ 4 – 7 ]. In a previous study, we reported the inhibitory effect of Ni 2+ ions on NF-κB activity wherein, the ions directly bind to NF-κB p50 subunit and inhibit its nuclear transport, thereby, inhibiting IL-8 secretion [ 8 ].…”

A static magnetic field inhibits the expression of platelet-derived growth factor-AA in human oral squamous cell carcinoma

“…Based on these results, we investigated the presence of inhibitory sites in order to ascertain the effect of a magnetic field on signal transduction pathways. Shionome et al reported that OSCC constantly activates the transcription factor NF-κB (28). Although PDGF-AA does not have a NF-κB binding site in the regulatory region, the action of PDGF-AA was observed by suppression of NF-κB using TPCK, a specific NF-κB-inhibitor.…”