Silibinin strongly inhibits growth and survival of human endothelial cells via cell cycle arrest and downregulation of survivin, Akt and NF-kappaB: implications for angioprevention and antiangiogenic therapy.
Recently, we observed that suppression of tumor xenograft growth by silibinin was associated with reduction in tumor vasculature and an increased apoptosis. Here, we provide evidence for molecular events associated with antiangiogenic efficacy of pharmacologically achievable doses of silibinin in endothelial cell culture system. Our data show that silibinin almost completely (P<0.001) inhibits growth of human umbilical vein endothelial cells (HUVEC) and human microvascular endothelial cells (HMVEC-dermal origin) together with induction of cell death in a dose- and time-dependent manner. Growth inhibition was associated with a strong induction of G1 arrest accompanied by an increase in Kip1/p27, Cip1/p21 and p53. Apoptosis induction (up to 14- to 17-fold in both cell lines, P<0.001) was an underlying mechanism in silibinin-induced death of endothelial cells. In the studies elucidating the molecular events involved in apoptosis, silibinin caused loss of mitochondrial membrane potential and an increase in cytochrome c release from mitochondria. An increase in Bax and a decrease in Mcl-1 proteins were also observed. Silibinin-induced apoptosis involved both caspase-dependent and -independent mechanisms. Silibinin also decreased survivin level and inhibited Akt and NF-kappaB signaling. Two different PI-3K inhibitors, wortmannin and LY294002, showed Akt-independent activation of NF-kappaB. Further, silibinin showed a concentration-dependent strong inhibition of capillary tube formation on matrigel, retraction and disintegration of preformed capillary network, inhibition of matrigel invasion and migration, and a decrease in matrix metalloproteinase-2 secretion by HUVEC. Together, these findings identify pleiotropic mechanisms for antiangiogenic efficacy of silibinin, and suggest its usefulness in angioprevention and antiangiogenic therapy.
Singh
Rana P
RP
Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Health Sciences Center, Denver, CO 80262, USA.
Dhanalakshmi
Sivanandhan
S
Agarwal
Chapla
C
Agarwal
Rajesh
R
D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D017209:Apoptosis; D001681:Biological Assay; D002453:Cell Cycle / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D003094:Collagen / Q000737:chemistry; D015536:Down-Regulation; D004338:Drug Combinations; D004730:Endothelium, Vascular / Q000737:chemistry / Q000187:drug effects*; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D007797:Laminin / Q000737:chemistry; D008564:Membrane Potentials / Q000187:drug effects; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008928:Mitochondria / Q000187:drug effects; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D009363:Neoplasm Proteins; D009389:Neovascularization, Pathologic / Q000628:therapy; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors*; D011509:Proteoglycans / Q000737:chemistry; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors*; D051057:Proto-Oncogene Proteins c-akt; D000077385:Silybin; D012838:Silymarin / Q000494:pharmacology*; D000077022:Survivin
D020533:Angiogenesis Inhibitors; C107489:BIRC5 protein, human; D004338:Drug Combinations; D051034:Inhibitor of Apoptosis Proteins; D007797:Laminin; D008869:Microtubule-Associated Proteins; D016328:NF-kappa B; D009363:Neoplasm Proteins; D011509:Proteoglycans; D011518:Proto-Oncogene Proteins; D012838:Silymarin; D000077022:Survivin; C059485:matrigel; D000077385:Silybin; D003094:Collagen; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt