RUNX3 confers sensitivity to pheophorbide a-photodynamic therapy in human oral squamous cell carcinoma cell lines.

Photodynamic therapy (PDT) with photosensitizer is one of the promising modalities for cancer treatment. For clinical use of PDT, screening process should be preceded to enhance sensitivity to PDT. Thus, we investigated a molecular biomarker to determine the sensitivity to pheophorbide a (Pa)-PDT in immortalized human oral keratinocytes (IHOK) and oral squamous cell carcinoma (OSCC) cell lines. Two IHOK and several OSCC cell lines were used. After Pa-PDT, cell viability was reduced by more than 50%, and reactive oxygen species were generated in IHOK and OSCC cell lines. Additionally, apoptosis occurred in PDT-treated cells. IHOK(S) and IHOK(P), the two IHOK cell lines derived from the same source, showed a difference in cytotoxicity after Pa-PDT. To explain this difference in cytotoxicity, we looked at the expression of Wnt signaling-related genes in these two cell lines, for the morphology of IHOK(S) which was spindle like and elongated and distinct from IHOK(P) and the parent cell. Among the relevant genes, runt-related transcription factor 3 (RUNX3), an apoptosis-related gene, was selected as a potential marker that confers sensitivity to PDT. We found that the cytotoxicity by Pa-PDT was proportional to RUNX3 expression in OSCC cell lines. Additionally, knockdown of RUNX3 expression reduced cytotoxicity by Pa-PDT, suggesting that RUNX3 might be a biomarker to determine sensitivity to Pa-PDT. This was the first study to find a new target molecule that enhances Pa-PDT effects in IHOK and OSCC cell lines. Hence, the development of a PDT-dependent biomarker could provide a novel approach to improve the effects of PDT on oral precancerous and cancerous lesions.

Helicobacter pylori CagA promotes Snail-mediated epithelial-mesenchymal transition by reducing GSK-3 activity.

Cytotoxin-associated gene A (CagA) is an oncoprotein and a major virulence factor of H. pylori. CagA is delivered into gastric epithelial cells via a type IV secretion system and causes cellular transformation. The loss of epithelial adhesion that accompanies the epithelial-mesenchymal transition (EMT) is a hallmark of gastric cancer. Although CagA is a causal factor in gastric cancer, the link between CagA and the associated EMT has not been elucidated. Here, we show that CagA induces the EMT by stabilizing Snail, a transcriptional repressor of E-cadherin expression. Mechanistically we show that CagA binds GSK-3 in a manner similar to Axin and causes it to shift to an insoluble fraction, resulting in reduced GSK-3 activity. We also find that the level of Snail protein is increased in H. pylori infected epithelium in clinical samples. These results suggest that H. pylori CagA acts as a pathogenic scaffold protein that induces a Snail-mediated EMT via the depletion of GSK-3.

p53 regulates nuclear GSK-3 levels through miR-34-mediated Axin2 suppression in colorectal cancer cells.

p53 is a bona fide tumor suppressor gene whose loss of function marks the most common genetic alteration in human malignancy. Although the causal link between loss of p53 function and tumorigenesis has been clearly demonstrated, the mechanistic links by which loss of p53 potentiates oncogenic signaling are not fully understood. Recent evidence indicates that the microRNA-34 (miR-34) family, a transcriptional target of the p53, directly suppresses a set of canonical Wnt genes and Snail, resulting in p53-mediated suppression of Wnt signaling and the EMT process. In this study, we report that p53 regulates GSK-3ß nuclear localization via miR-34-mediated suppression of Axin2 in colorectal cancer. Exogenous miR-34a decreases Axin2 UTR-reporter activity through multiple binding sites within the 5' and 3' UTR of Axin2. Suppression of Axin2 by p53 or miR-34 increases nuclear GSK-3ß abundance and leads to decreased Snail expression in colorectal cancer cells. Conversely, expression of the non-coding UTR of Axin2 causes depletion of endogenous miR-34 via the miR-sponge effect together with increased Axin2 function, supporting that the RNA-RNA interactions with Axin2 transcripts act as an endogenous decoy for miR-34. Further, RNA transcripts of miR-34 target were correlated with Axin2 in clinical data set of colorectal cancer patients. Although the biological relevance of nuclear GSK-3 level has not been fully studied, our results demonstrate that the tumor suppressor p53/miR-34 axis plays a role in regulating nuclear GSK-3 levels and Wnt signaling through the non-coding UTR of Axin2 in colorectal cancer.

The effects of cadmium on VEGF-mediated angiogenesis in HUVECs.

Cadmium (Cd) is a highly toxic element that causes morphologic alterations and dysfunction in blood vessels. The altered vascular function caused by cadmium has been implicated in a range of chronic diseases, including hypertension. The effects of cadmium are a multisystem phenomenon involving inflammation, hypertrophy, apoptosis, angiogenesis and important processes involved in vascular remodeling systems. Vascular endothelial growth factor (VEGF) plays a major role in cell growth and angiogenesis under pathologic conditions. VEGF secretion is related to anti-apoptosis protein expression and attenuates apoptosis in endothelial cells. This study examined the VEGF-dependent mechanisms of angiogenesis and apoptosis in cadmium-treated endothelial cells (HUVECs). The effects and mechanisms of cadmium in endothelial cells (HUVECs) were examined by exposing the cells to different doses of cadmium chloride (2.5-40 µ m). After the cadmium treatment, the angiogenesis and apoptosis mechanisms related to VEGF in cadmium-treated HUVECs were examined. As a result, the low concentration of cadmium increased the tube formation in HUVECs. In addition, cadmium at concentrations of 5 and 10 µ m increased VEGF secretion and VEGFR2 activity, which suggest that cadmium affects the growth of blood vessels. All three MAPK pathways, namely ERK, JNK and p38, were activated by cadmium in HUVECs. However, high concentrations of cadmium caused cell damage, disrupted tube formation and inhibited VEGF expression and the activities of VEGFR2 and MAPK in HUVECs. Cadmium has dual functions through VEGF-dependent mechanisms in a dose-dependent manner. In this study, the dual effects of cadmium might alter angiogenesis and induce apoptosis through VEGF pathways in HUVECs.