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vol. 14
Basic research

Inhibitory effect of PDGF-BB and serum-stimulated responses in vascular smooth muscle cell proliferation by hinokitiol via up-regulation of p21 and p53

Jiun-Yi Li, Chun-Ping Liu, Wei-Cheng Shiao, Thanasekaran Jayakumar, Yi-Shin Li, Nen-Chung Chang, Shih-Yi Huang, Cheng-Ying Hsieh

Arch Med Sci 2018; 14, 3: 579–587
Online publish date: 2018/04/16
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Vascular smooth muscle cell (VSMC) proliferation plays a major role in the progression of vascular diseases. In the present study, we established the efficacy and the mechanisms of action of hinokitiol, a tropolone derivative found in Chamaecyparis taiwanensis, Cupressaceae, in relation to platelet-derived growth factor-BB (PDGF-BB) and serum-dependent VSMC proliferation.

Material and methods
Primary cultured rat VSMCs were pre-treated with hinokitiol and then stimulated by PDGF-BB (10 ng/ml) or serum (10% fetal bovine serum). Cell proliferation and cytotoxicity were determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay and lactose dehydrogenase assay, respectively. The degree of DNA synthesis was evaluated by BrdU-incorporation measurements and observed using confocal microscopy. Immunoblotting was utilized to determine the protein level of p-extracellular signal-regulated kinase (ERK) 1/2, p-Akt, p-phosphoinositide 3-kinase (PI3K), p-Janus kinase 2 (JAK2), p-p53, and p21Cip1. The promoter activity of p21 and p53 activity were measured by dual luciferase reporter assay.

Treatment with hinokitiol (1–10 µM) inhibited PDGF-BB and serum-induced VSMC proliferation and DNA synthesis in a concentration-dependent manner. Cytotoxicity was not observed in hinokitiol-treated VSMCs at the studied concentrations. Pre-incubation of VSMCs with hinokitiol did not alter PDGF-BB-induced phosphorylation of ERK1/2, Akt, PI3K or JAK2. Interestingly, hinokitiol induced promoter activity of p21 and p21 protein expression in VSMCs. Furthermore, hinokitiol augmented p53 protein phosphorylation and subsequently led to enhanced p53 activity.

These data suggest that the anti-proliferative effects of hinokitiol in VSMCs may be mediated by activation of p21 and p53 signaling pathways, and it may contribute to the prevention of vascular diseases associated with VSMC proliferation.


vascular smooth muscle cell proliferation, hinokitiol, p21, p53

Doran AC, Meller N, McNamara CA. Role of smooth muscle cells in the initiation and early progression of atherosclerosis. Arterioscler Thromb Vasc Biol 2008; 28: 812-9.
Owens GK, Kumar MS, Wamhoff BR. Molecular regulation of vascular smooth muscle cell differentiation in development and disease. Physiol Rev 2004; 84: 767-801.
Dzau VJ, Braun-Dullaeus RC, Sedding DG. Vascular proliferation and atherosclerosis: new perspectives and therapeutic strategies. Nat Med 2002; 8: 1249-56.
Braun-Dullaeus RC, Mann MJ, Sedding DG, et al. Cell cycle-dependent regulation of smooth muscle cell activation. Arterioscler Thromb Vasc Biol 2004; 24: 845-50.
Xiong Y, Hannon GJ, Zhang H, et al. p21 is a universal inhibitor of cyclin kinases. Nature 1993; 366: 701-4.
Coqueret O. New roles for p21 and p27 cell-cycle inhibitors: a function for each cell compartment? Trends Cell Biol 2003; 13: 65-70.
Ortega S, Malumbres M, Barbacid M, et al. Cyclin D-dependent kinases, INK4 inhibitors and cancer. Biochim Biophys Acta 2002; 1602: 73-87.
Sherr CJ, Roberts JM. CDK inhibitors: positive and negative regulators of G1-phase progression. Genes Dev 1999; 13: 1501-12.
Macleod KF, Sherry N, Hannon G, et al. p53-dependent and independent expression of p21 during cell growth, differentiation, and DNA damage. Genes Dev 1995; 9: 935-44.
Chan KC, Ho HH, Peng CH, et al. Polyphenol-rich extract from mulberry leaf inhibits vascular smooth muscle cell proliferation involving upregulation of p53 and inhibition of cyclin-dependent kinase. J Agric Food Chem 2010; 58: 2536-42.
Pellegata NS, Antoniono RJ, Redpath JL, et al. DNA damage and p53-mediated cell cycle arrest: a reevaluation. Proc Natl Acad Sci USA 1996; 93: 15209-14.
Chaudhary LR, Hruska KA. The cell survival signal Akt is differentially activated by PDGF-BB, EGF, and FGF-2 in osteoblastic cells. J Cell Biochem 2001; 81: 304-11.
Bhanoori M, Yellaturu CR, Ghosh SK, et al. Thiol alkylation inhibits the mitogenic effects of platelet-derived growth factor and renders it proapoptotic via activation of STATs and p53 and induction of expression of caspase1 and p21waf1/cip1. Oncogene 2003; 22: 117-30.
Levitzki A. PDGF receptor kinase inhibitors for the treatment of restenosis. Cardiovasc Res 2005; 65: 581-6.
Zhao J, Fujita K, Sakai K. Reactive oxygen species, nitric oxide, and their interactions play different roles in Cupressus lusitanica cell death and phytoalexin biosynthesis. New Phytol 2007; 175: 215-29.
Saeki Y, Ito Y, Shibata M, et al. Antimicrobial action of natural substances on oral bacteria. Bull Tokyo Dent Coll 1989; 30: 129-35.
Arima Y, Nakai Y, Hayakawa R, et al. Antibacterial effect of beta-thujaplicin on staphylococci isolated from atopic dermatitis: relationship between changes in the number of viable bacterial cells and clinical improvement in an eczematous lesion of atopic dermatitis. J Antimicrob Chemother 2003; 51: 113-22.
Morita Y, Matsumura E, Okabe T, et al. Biological activity of beta-dolabrin, gamma-thujaplicin, and 4-acetyltropolone, hinokitiol-related compounds. Biol Pharm Bull 2004; 27: 1666-9.
Koufaki M, Theodorou E, Alexi X, et al. Synthesis of tropolone derivatives and evaluation of their in vitro neuroprotective activity. Eur J Med Chem 2010; 45: 1107-12.
Liu S, Yamauchi H. p27-Associated G1 arrest induced by hinokitiol in human malignant melanoma cells is mediated via down-regulation of pRb, Skp2 ubiquitin ligase, and impairment of Cdk2 function. Cancer Lett 2009; 286: 240-9.
Li LH, Wu P, Lee JY, et al. Hinokitiol induces DNA damage and autophagy followed by cell cycle arrest and senescence in gefitinib-resistant lung adenocarcinoma cells. PLoS One 2014; 9: e104203.
Wang WK, Lin ST, Chang WW, et al. Hinokitiol induces autophagy in murine breast and colorectal cancer cells. Environ Toxicol 2016; 31: 77-84.
Lin KH, Kuo JR, Lu WJ, et al. Hinokitiol inhibits platelet activation ex vivo and thrombus formation in vivo. Biochem Pharmacol 2013; 85: 1478-85.
Jayakumar T, Hsu WH, Yen TL, et al. Hinokitiol, a natural tropolone derivative, offers neuroprotection from thromboembolic stroke in vivo. Evid Based Complement Alternat Med 2013; 2013: 840487.
Huang CH, Chang CC, Thomas PA, et al. Hinokitiol, a tropolone derivative, inhibits mouse melanoma (B16-F10) cell migration and in vivo tumor formation. Eur J Pharmacol 2015; 746: 148-57.
Chang Y, Uen YH, Chen CC, et al. Platonin inhibited PDGF-BB-induced proliferation of rat vascular smooth muscle cells via JNK1/2-dependent signaling. Acta Pharmacol Sin 2011; 32: 1337-44.
Hsieh CY, Liu CL, Hsu MJ, et al. Inhibition of vascular smooth muscle cell proliferation by the vitamin E derivative pentamethylhydroxychromane in an in vitro and in vivo study: pivotal role of hydroxyl radical-mediated PLCgamma1 and JAK2 phosphorylation. Free Radic Biol Med 2010; 49: 881-93.
Wang JJ, Huan SK, Hsieh KH, et al. Inhibitory effect of midazolam on MMP-9, MMP-1 and MMP-13 expression in PMA-stimulated human chondrocytes via recovery of NF-kappaB signaling. Arch Med Sci 2013; 9: 332-9.
Lim Y, Kwon JS, Kim DW, et al. Obovatol from Magnolia obovate inhibits vascular smooth muscle cell proliferation and intimal hyperplasia by inducing p21Cip1. Atherosclerosis 2010; 210: 372-80.
Philipp-Staheli J, Kim KH, Liggitt D, et al. Distinct roles for p53, p27Kip1, and p21Cip1 during tumor development. Oncogene 2004; 23: 905-13.
Harper JW, Adami GR, Wei N, et al. The p21 Cdk-interacting protein Cip1 is a potent inhibitor of G1 cyclin-dependent kinases. Cell 1993; 75: 805-16.
Stewart ZA, Leach SD, Pietenpol JA. p21(Waf1/Cip1) inhibition of cyclin E/Cdk2 activity prevents endoreduplication after mitotic spindle disruption. Mol Cell Biol 1999; 19: 205-15.
Gartel AL, Tyner AL. Transcriptional regulation of the p21 (WAF1/CIP1) gene. Exp Cell Res 1999; 246: 280-9.
Majesky MW, Reidy MA, Bowen-Pope DF, et al. PDGF ligand and receptor gene expression during repair of arterial injury. J Cell Biol 1990; 111: 2149-58.
Kim JH, Jin YR, Park BS, et al. Luteolin prevents PDGF-BB-induced proliferation of vascular smooth muscle cells by inhibition of PDGF beta-receptor phosphorylation. Biochem Pharmacol 2005; 69: 1715-21.
Parcellier A, Tintignac LA, Zhuravleva E, et al. PKB and the mitochondria: AKTing on apoptosis. Cell Signal 2008; 20: 21-30.
Eisenreich A, Rauch U. PI3K inhibitors in cardiovascular disease. Cardiovasc Ther 2011; 29: 29-36.
Cantley LC. The phosphoinositide 3-kinase pathway. Science 2002; 296: 1655-7.
Maira SM, Stauffer F, Schnell C, et al. PI3K inhibitors for cancer treatment: where do we stand? Biochem Soc Trans 2009; 37: 265-72.
Fougerat A, Gayral S, Gourdy P, et al. Genetic and pharmacological targeting of phosphoinositide 3-kinase- gamma reduces atherosclerosis and favors plaque stability by modulating inflammatory processes. Circulation 2008; 117: 1310-7.
Nakashio A, Fujita N, Tsuruo T. Topotecan inhibits VEGF- and bFGF-induced vascular endothelial cell migration via down-regulation of the PI3K-Akt signaling pathway. Int J Cancer 2002; 98: 36-41.
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