Inhibitory effect of Curcumin on phosphorylation NFκB-p65 induced by hydrogen peroxide in Bovine Endothelial Cells

Background & Objective: NFκB is a dimeric transcription factor with multiple subunits. Phosphorylation of  p65 (one of NFκB subunits) by oxidative stress leads to the activation of NFκB. Imbalance between oxidative stress and cellular antioxidant capacity is the pathogenesis of many diseases. Consuming antioxidants in daily meal can be considered as a preventive strategy for inflammatory diseases. Among antioxidant components, curcumin is a natural polyphenol which is extracted from Tumeric. Curcumin can protect the human body from oxidative stress by neutralizing the free radicals. Material & Methods: Bovine Endothelial Cells (BECs) were treated with different concentration of H2O2 (10, 20, 80, 200 μM). To investigate the inhibitory effect of curcumin on H2O2 mediated phosphorylation of p65, BECs were incubated with 5 and 10μM concentration of curcumin. Protein concentration was measured by Bradford method and phosphorylation of p65 was assessed by western blotting. Results: Our finding indicated that p65 phosphorylation was increased two fold in presence of H2O2 (200 μM) in comparison with control. The enhancing effect of H2O2 on p65 phosphorylation decreased at 30 min (P: 0.03) and 2 hours (P:0.015) after treatment with 10 μM dose of curcumin. Conclusion: The result of this study indicates that one of anti-inflammatory mechanisms of curcumin is through NFκB pathway by inhibition of p65 subunit phosphorylation.   &nbsp

Endothelin-1 (ET-1) stimulates carboxy terminal Smad2 phosphorylation in vascular endothelial cells by a mechanism dependent on ET receptors and de novo protein synthesis

Objective: G protein-coupled receptor (GPCR) agonists through their receptors can transactivate protein tyrosine kinase receptors such as epidermal growth factor receptor and serine/threonine kinase receptors most notably transforming growth factor (TGF)-β receptor (TβRI). This signalling mechanism represents a major expansion in the cellular outcomes attributable to GPCR signalling. This study addressed the role and mechanisms involved in GPCR agonist, endothelin-1 (ET-1)-mediated transactivation of the TβRI in bovine aortic endothelial cells (BAECs). Method: The in-vitro model used BAECs. Signalling intermediate phospho-Smad2 in the carboxy terminal was detected and quantified by Western blotting. Key finding: ET-1 treatment of BAECs resulted in a time and concentration-dependent increase in pSmad2C. Peak phosphorylation was evident with 100 nm treatment of ET-1 at 4–6 h. TβRI antagonist, SB431542 inhibited ET-1-mediated pSmad2C. In the presence of bosentan, a mixed ET and ET receptor antagonist ET-1-mediated pSmad2C levels were inhibited. The ET-mediated pSmad2C was blocked by the protein synthesis inhibitor, cycloheximide. Conclusion: In BAECs, ET-1 via the ETB receptor is involved in transactivation of the TβRI. The transactivation-dependent response is dependent upon de novo protein synthesis