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  • Original Paper
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Silibinin strongly inhibits growth and survival of human endothelial cells via cell cycle arrest and downregulation of survivin, Akt and NF-κB: implications for angioprevention and antiangiogenic therapy

Oncogene volume 24, pages 1188–1202 (2005)Cite this article

Abstract

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-κB signaling. Two different PI-3K inhibitors, wortmannin and LY294002, showed Akt-independent activation of NF-κB. 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.

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Abbreviations

HUVEC:

human umbilical vein endothelial cells

HMVEC:

human microvascular endothelial cells

VEGF:

vascular endothelial cell growth factor

ΔΨm:

mitochondrial membrane potential

JC-1:

5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethyl-benzamidazolocarbocyanin iodide

PARP:

poly(ADP-ribose) polymerase

z-VAD:

benzyloxycarbonyl-Val-Ala-Asp

FMK:

fluoromethyl-ketone

DEVD-pNA:

Asp-Glu-Val-Asp-p-nitroanilide

NF-κB:

nuclear factor kappa B

MMP-2:

matrix metalloproteinase-2

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Acknowledgements

This work was supported in part by DOD prostate cancer fellowship #PC020718 (to RPS) and USPHS Grants CA91883 and CA102514.

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Authors and Affiliations

  1. Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Health Sciences Center, Denver, CO, 80262, USA

    Rana P Singh, Sivanandhan Dhanalakshmi, Chapla Agarwal & Rajesh Agarwal

  2. University of Colorado Cancer Center, University of Colorado Health Sciences Center, Denver, CO, 80262, USA

    Chapla Agarwal & Rajesh Agarwal

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  1. Rana P Singh
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  2. Sivanandhan Dhanalakshmi
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Correspondence to Rajesh Agarwal.

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Singh, R., Dhanalakshmi, S., Agarwal, C. et al. Silibinin strongly inhibits growth and survival of human endothelial cells via cell cycle arrest and downregulation of survivin, Akt and NF-κB: implications for angioprevention and antiangiogenic therapy. Oncogene 24, 1188–1202 (2005). https://doi.org/10.1038/sj.onc.1208276

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