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Daphnetin inhibits TNF ‐α and VEGF ‐induced angiogenesis through inhibition of the IKK s/IκBα/ NF ‐κB, Src/ FAK / ERK 1/2 and Akt signalling pathways
Author(s) -
Kumar Abhishek,
Sunita Priyashree,
Jha Shivesh,
Pattanayak Shakti Prasad
Publication year - 2016
Publication title -
clinical and experimental pharmacology and physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.752
H-Index - 103
eISSN - 1440-1681
pISSN - 0305-1870
DOI - 10.1111/1440-1681.12608
Subject(s) - angiogenesis , mapk/erk pathway , chemistry , iκbα , protein kinase b , phosphorylation , umbilical vein , kinase , vascular endothelial growth factor , microbiology and biotechnology , blot , iκb kinase , human umbilical vein endothelial cell , signal transduction , nf κb , biology , cancer research , in vitro , biochemistry , vegf receptors , gene
Summary Coumarins, identified as plant secondary metabolites possess diverse biological activities including anti‐angiogenic properties. Daphnetin ( DAP ), a plant derived dihydroxylated derivative of coumarin has shown significant pharmacological properties such as anticancer, anti‐arthritic and anti‐inflammatory. The present study was performed to investigate the anti‐angiogenic potential of DAP , focusing on the mechanism of action. The in vivo anti‐angiogenic potential of DAP was evaluated by vascular endothelial growth factor ( VEGF )‐induced rat aortic ring ( RAR ) assay and chick chorioallantoic membrane ( CAM ) assay. For in vitro evaluation, wounding migration, transwell invasion, tube formation and apoptosis assays were performed on VEGF (8 ng/ mL )‐induced human umbilical vein endothelial cells ( HUVEC s). The cellular mechanism of DAP was examined on TNF α (10 ng/mL) and VEGF ‐induced HUVEC s by extracting the mRNA and protein levels using RT ‐ qPCR and western blotting. Our data demonstrated that DAP inhibited the in vivo angiogenesis in the RAR and CAM assay. DAP also inhibited the different steps of angiogenesis, such as migration, invasion, and tube formation in HUVEC s. DAP inhibited nuclear factor‐κB signalling together including TNF ‐α induced IκBα degradation; phosphorylation of IκB kinase ( IKK α/β) and translocation of the NF ‐κB‐p65 protein. Furthermore, western blotting revealed that DAP significantly down‐regulated the VEGF ‐induced signalling such as c‐Src, FAK , ERK 1/2 and the related phosphorylation of protein kinase B (Akt) and VEGFR 2 expressions. DAP reduced the elevated mRNA expression of iNOS , MMP 2 and also, induced apoptosis in VEGF ‐stimulated HUVEC s by the caspase‐3 dependent pathway. Taken together, this study reveals that DAP may have novel prospective as a new multi‐targeted medication for the anti‐angiogenesis and cancer therapy.