Anti-cancer effect of tectochrysin in NSCLC cells through overexpression of death receptor and inactivation of STAT3.

Phenolic compounds (flavonoids and phenolic acid derivatives) are the most important pharmacologically active ingredients, and these compounds could inhibit proliferation of human cancer cells by inducing of apoptotic cell death. Here we focused on the anticancer effects of tectochrysin on human non-small-cell lung cancer (NSCLC) cells and its mechanism of action. We analysed the activity of tectochrysin on NSCLC cells (A549 and NCI-H460) by use of Western blot analysis for major apoptotic proteins and death receptor expression. We also used EMSA for effects on STAT3 DNA binding activity. Tectochrysin (0-80 µM) suppressed the growth of A549 and NCI-H460 lung cancer cells by inducing of apoptotic cell death in a concentration dependent manner. Expression of DR3 and Fas as well as DR downstream pro-apoptotic proteins including cleaved caspase-3, cleaved caspase-8, cleaved caspase-9 and Bax were concomitantly increased, but the expression of anti-apoptotic proteins; Bcl-2 was decreased in both cancer cells. In addition, tectochrysin treatment also inhibited phosphorylation of STAT3 in A549 and NCI-H460 cells. However, deletion of DR3 and Fas by small interfering RNA significantly reversed tectochrysin-induced cell growth inhibitory effect as well as down regulation of STAT3 in A549 and NCI-H460 lung cancer cells. Pull down assay and docking model showed interaction of tectochrysin with STAT3. We propose that tectochrysin leads to apoptotic cell death in NSCLC cells through activation of DR3 and Fas expression via inhibition of STAT3 phosphorylation.

Inhibitory effect of ent-Sauchinone on amyloidogenesis via inhibition of STAT3-mediated NF-κB activation in cultured astrocytes and microglial BV-2 cells.

BACKGROUND: ent-Sauchinone is a polyphenolic compound found in plants belonging to the lignan family. ent-Sauchinone has been shown to modulate the expression of inflammatory factors through the nuclear factor-kappa B (NF-κB) signaling pathway. It is well known that neuroinflammation is associated with amyloidogenesis. Thus, in the present study, we investigated whether ent-Sauchinone could have anti-amyloidogenic effects through the inhibition of NF-κB pathways via its anti-inflammatory property. METHODS: To investigate the potential effect of ent-Sauchinone on anti-neuroinflammation and anti-amyloidogenesis in in vitro studies, we used microglial BV-2 cells and cultured astrocytes treated with ent-Sauchinone (1, 5, and 10 µM) for 24 hours. For the detection of anti-neuro-inflammatory responses, reative oxygen species (ROS) and Nitric oxide (NO) generation and inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression were measured with assay kits and western blotting. ß-secretase and ß-secretase activities and ß-amyloid levels were determined for measuring the anti-amyloidogenic effects of ent-Sauchinone by enzyme assay kits. NF-κB and STAT3 signals were detected with electromobility shift assay (EMSA) to study the related signaling pathways. The binding of ent-Sauchinone to STAT3 was evaluated by a pull-down assay and by a docking model using Autodock VINA software (Hoover's Inc., Texas, United states). RESULTS: ent-Sauchinone (1, 5, and 10 µM) effectively decreased lipopolysaccharide (LPS)-(1 µg/ml) induced inflammatory responses through the reduction of ROS and NO generations and iNOS and COX-2 expressions in cultured astrocytes and microglial BV-2 cells. ent-Sauchinone also inhibited LPS-induced amyloidogenesis through the inhibition of ß-secretase and ß-secretase activity. NF- κB amyloid and STAT3, critical transcriptional factors regulating not only inflammation but also amyloidogenesis, were also inhibited in a concentration dependent manner by ent-Sauchinone by blocking the phosphorylation of I κB and STAT3 in cultured astrocytes and microglial BV-2 cells. The docking model approach showed that ent-Sauchinone binds to STAT3, and the employment of a STAT3 inhibitor and siRNA reversed ent-Sauchinone-induced inhibition NF-κB activation and Aß generation. CONCLUSIONS: These results indicated that ent-Sauchinone inhibited neuroinflammation and amyloidogenesis through the inhibition of STAT3-mediated NF-κB activity, and thus could be applied in the treatment of neuro-inflammatory diseases, including Alzheimer's disease.