Inhibitory effect of ethanol extract of Ocimum sanctum on osteopontin mediated metastasis of NCI-H460 non-small cell lung cancer cells

BMC Complementary and Alternative Medicine

Sung-Hoon Kim¹  Oh Sung Kwon¹  Myoungseok Jeong¹  Kwon Jeong¹  Jeong-Un Choi¹  Gunho Won¹  Sunhee Kim¹  Eun Jung Sohn¹  Tae-kyung Kwak¹ 

[1]College of Oriental Medicine, Kyung Hee University, Hoegidong, Dongdaemungu, Seoul 130-701, Republic of Korea

关键词: PI3K;    uPAR;    uPA;    Osteopontin;    Metastasis;    EEOS;   

Others  :  1085753
DOI  :  10.1186/1472-6882-14-419

 received in 2013-12-31, accepted in 2014-06-27,  发布年份 2014

【 摘 要 】

Background

Osteopontin (OPN) is one of important molecular targets in cancer progression, metastasis as a calcium-binding, extracellular-matrix-associated protein of the small integrin-binding ligand and, N-linked glycoprotein. In the present study, anti-metastatic mechanism of ethanol extracts of Ocimum sanctum (EEOS) was elucidated on OPN enhanced metastasis in NCI-H460 non- small cell lung cancer cells.

Methods

Cell viability was measured by MTT assay. Adhesion and invasion assays were carried out to see that EEOS inhibited cell adhesion and invasion in OPN treated and non-treated NCI-H 460 cells. RT-PCR was used to determine the mRNA levels of uPA, uPAR, and EGFR.

Results

EEOS significantly inhibited cell adhesion and invasion in OPN treated and non treated NCI-H460 cells, though EEOS did not show any toxicity up to 200 μg/ml. EEOS effectively attenuated the expression of OPN and CD44 and also OPN activated the expression of CD44 in NCI-H460 cells. In addition, EEOS effectively suppressed the expression of phosphatidylinositide 3-kinases (PI3K) and cyclooxygenase 2 (COX-2) and the phosphorylation of Akt at protein level in OPN treated NCI-H460 cells. Also, EEOS significantly attenuated the expression of urokinase plasminogen activator (uPA), its receptor (uPAR) and epidermal growth factor receptor (EGFR) at mRNA level and reduced vascular endothelial growth factor (VEGF) production and MMP-9 activity in OPN treated NCI-H460 cells. Furthermore, PI3K/Akt inhibitor LY294002 enhanced anti-metastatic potential of EEOS to attenuate the expression of uPA and MMP-9 in OPN treated NCI-H 460 cells.

Conclusion

Overall, our findings suggest that anti-metastatic mechanism of EEOS is mediated by inhibition of PI3K/Akt in OPN treated NCI-H460 non-small cell lung cancer cells.

【 授权许可】

   
2014 Kwak et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Fidler IJ: The pathogenesis of cancer metastasis: the ‘seed and soil’ hypothesis revisited. Nat Rev Cancer 2003, 3(6):453-458.
• [2]Weiss L: Metastasis of cancer: a conceptual history from antiquity to the 1990s. Cancer Metastasis Rev 2000, 19(3–4):193-383. I-XI
• [3]O’Regan A, Berman JS: Osteopontin: a key cytokine in cell-mediated and granulomatous inflammation. Int J Exp Pathol 2000, 81(6):373-390.
• [4]Ramaiah SK, Rittling S: Pathophysiological role of osteopontin in hepatic inflammation, toxicity, and cancer. Toxicol Sci 2008, 103(1):4-13.
• [5]Vernon HJ, Osborne C, Tzortzaki EG, Yang M, Chen J, Rittling SR, Denhardt DT, Buyske S, Bledsoe SB, Evan AP, Fairbanks L, Simmonds HA, Tischfield JA, Sahota A: Aprt/Opn double knockout mice: osteopontin is a modifier of kidney stone disease severity. Kidney Int 2005, 68(3):938-947.
• [6]Jiang XJ, Feng T, Chang LS, Kong XT, Wang G, Zhang ZW, Guo YL: Expression of osteopontin mRNA in normal and stone-forming rat kidney. Urol Res 1998, 26(6):389-394.
• [7]Huang J, Pan C, Hu H, Zheng S, Ding L: Osteopontin-enhanced hepatic metastasis of colorectal cancer cells. PLoS One 2012, 7(10):e47901.
• [8]Nemoto H, Rittling SR, Yoshitake H, Furuya K, Amagasa T, Tsuji K, Nifuji A, Denhardt DT, Noda M: Osteopontin deficiency reduces experimental tumor cell metastasis to bone and soft tissues. J Bone Miner Res 2001, 16(4):652-659.
• [9]Pan HW, Ou YH, Peng SY, Liu SH, Lai PL, Lee PH, Sheu JC, Chen CL, Hsu HC: Overexpression of osteopontin is associated with intrahepatic metastasis, early recurrence, and poorer prognosis of surgically resected hepatocellular carcinoma. Cancer 2003, 98(1):119-127.
• [10]Curran S, Murray GI: Matrix metalloproteinases: molecular aspects of their roles in tumour invasion and metastasis. Eur J Cancer 2000, 36(13 Spec No):1621-1630.
• [11]Pakneshan P, Szyf M, Farias-Eisner R, Rabbani SA: Reversal of the hypomethylation status of urokinase (uPA) promoter blocks breast cancer growth and metastasis. J Biol Chem 2004, 279(30):31735-31744.
• [12]He Y, Liu XD, Chen ZY, Zhu J, Xiong Y, Li K, Dong JH, Li X: Interaction between cancer cells and stromal fibroblasts is required for activation of the uPAR-uPA-MMP-2 cascade in pancreatic cancer metastasis. Clin Cancer Res 2007, 13(11):3115-3124.
• [13]Rabbani SA, Xing RH: Role of urokinase (uPA) and its receptor (uPAR) in invasion and metastasis of hormone-dependent malignancies. Int J Oncol 1998, 12(4):911-920.
• [14]Yothaisong S, Dokduang H, Techasen A, Namwat N, Yongvanit P, Bhudhisawasdi V, Puapairoj A, Riggins GJ, Loilome W: Increased activation of PI3K/AKT signaling pathway is associated with cholangiocarcinoma metastasis and PI3K/mTOR inhibition presents a possible therapeutic strategy. Tumour Biol 2013, 34(6):3637-3648.
• [15]Shi MD, Liao YC, Shih YW, Tsai LY: Nobiletin attenuates metastasis via both ERK and PI3K/Akt pathways in HGF-treated liver cancer HepG2 cells. Phytomedicine 2013, 20(8–9):743-752.
• [16]Wang G, Wang F, Ding W, Wang J, Jing R, Li H, Wang X, Wang Y, Ju S, Wang H: APRIL induces tumorigenesis and metastasis of colorectal cancer cells via activation of the PI3K/Akt pathway. PLoS One 2013, 8(1):e55298.
• [17]Zhu B, Zhou X: The study of PI3K/AKT pathway in lung cancer metastasis and drug resistance. Zhongguo Fei Ai Za Zhi 2011, 14(8):689-694.
• [18]Li B, Tsao SW, Li YY, Wang X, Ling MT, Wong YC, He QY, Cheung AL: Id-1 promotes tumorigenicity and metastasis of human esophageal cancer cells through activation of PI3K/AKT signaling pathway. Int J Cancer 2009, 125(11):2576-2585.
• [19]Hua K, Feng W, Cao Q, Zhou X, Lu X, Feng Y: Estrogen and progestin regulate metastasis through the PI3K/AKT pathway in human ovarian cancer. Int J Oncol 2008, 33(5):959-967.
• [20]Wang XF, Su SB: Review on experimental research of Chinese herbal medicine and its components intervention for tumor metastasis. Zhongguo Zhong Yao Za Zhi 2008, 33(22):2583-2587.
• [21]Lee HJ, Lee EO, Rhee YH, Ahn KS, Li GX, Jiang C, Lu J, Kim SH: An oriental herbal cocktail, ka-mi-kae-kyuk-tang, exerts anti-cancer activities by targeting angiogenesis, apoptosis and metastasis. Carcinogenesis 2006, 27(12):2455-2463.
• [22]Chen CC, Sureshbabul M, Chen HW, Lin YS, Lee JY, Hong QS, Yang YC, Yu SL: Curcumin suppresses metastasis via Sp-1, FAK inhibition, and E-cadherin upregulation in colorectal cancer. Evid Based Complement Alternat Med 2013, 2013:541695.
• [23]Takahashi RU, Takeshita F, Honma K, Ono M, Kato K, Ochiya T: Ribophorin II regulates breast tumor initiation and metastasis through the functional suppression of GSK3beta. Sci Rep 2013, 3:2474.
• [24]Shu G, Mi X, Cai J, Zhang X, Yin W, Yang X, Li Y, Chen L, Deng X: Brucine, an alkaloid from seeds of Strychnos nux-vomica Linn., represses hepatocellular carcinoma cell migration and metastasis: the role of hypoxia inducible factor 1 pathway. Toxicol Lett 2013, 222(2):91-101.
• [25]Singh S, Majumdar DK, Rehan HM: Evaluation of anti-inflammatory potential of fixed oil of Ocimum sanctum (Holybasil) and its possible mechanism of action. J Ethnopharmacol 1996, 54(1):19-26.
• [26]Llovet JM, Bruix J: Molecular targeted therapies in hepatocellular carcinoma. Hepatology 2008, 48(4):1312-1327.
• [27]Godhwani S, Godhwani JL, Vyas DS: Ocimum sanctum: an experimental study evaluating its anti-inflammatory, analgesic and antipyretic activity in animals. J Ethnopharmacol 1987, 21(2):153-163.
• [28]Kelm MA, Nair MG, Strasburg GM, DeWitt DL: Antioxidant and cyclooxygenase inhibitory phenolic compounds from Ocimum sanctum Linn. Phytomedicine 2000, 7(1):7-13.
• [29]Magesh V, Lee JC, Ahn KS, Lee HJ, Lee EO, Shim BS, Jung HJ, Kim JS, Kim DK, Choi SH, Ahn KS, Kim SH: Ocimum sanctum induces apoptosis in A549 lung cancer cells and suppresses the in vivo growth of lewis lung carcinoma cells. Phytother Res 2009, 23(10):1385-1391.
• [30]Hannan JM, Marenah L, Ali L, Rokeya B, Flatt PR, Abdel-Wahab YH: Ocimum sanctum leaf extracts stimulate insulin secretion from perfused pancreas, isolated islets and clonal pancreatic beta-cells. J Endocrinol 2006, 189(1):127-136.
• [31]Venu Prasad MP, Farhath K: Antifatigue activity of ethanolic extract of Ocimum sanctum in Rats. Res J Med Plant 2012, 6(1):37-46.
• [32]Kim SC, Magesh V, Jeong SJ, Lee HJ, Ahn KS, Lee EO, Kim SH, Lee MH, Kim JH: Ethanol extract of Ocimum sanctum exerts anti-metastatic activity through inactivation of matrix metalloproteinase-9 and enhancement of anti-oxidant enzymes. Food Chem Toxicol 2010, 48(6):1478-1482.
• [33]van Rossen ME, Hofland LJ, van den Tol MP, van Koetsveld PM, Jeekel J, Marquet RL, van Eijck CH: Effect of inflammatory cytokines and growth factors on tumour cell adhesion to the peritoneum. J Pathol 2001, 193(4):530-537.
• [34]Koopman JL, Slomp J, de Bart AC, Quax PH, Verheijen JH: Mitogenic effects of urokinase on melanoma cells are independent of high affinity binding to the urokinase receptor. J Biol Chem 1998, 273(50):33267-33272.
• [35]Herron GS, Werb Z, Dwyer K, Banda MJ: Secretion of metalloproteinases by stimulated capillary endothelial cells. I. Production of procollagenase and prostromelysin exceeds expression of proteolytic activity. J Biol Chem 1986, 261(6):2810-2813.
• [36]Stamenkovic I: Extracellular matrix remodelling: the role of matrix metalloproteinases. J Pathol 2003, 200(4):448-464.
• [37]Ahmed M, Behera R, Chakraborty G, Jain S, Kumar V, Sharma P, Bulbule A, Kale S, Kumar S, Mishra R, Raja R, Saraswati S, Kaur R, Soundararajan G, Kumar D, Sanyal M, Ramdashi A, Ghosh P, Kundu GC: Osteopontin: a potentially important therapeutic target in cancer. Expert Opin Ther Targets 2011, 15(9):1113-1126.
• [38]Wang S, Li S, Xie D, Tang Q, Liu J, Chen Y, Yang X: CD44 regulates epithelial-mesenchymal transition and metastasis in nasopharyngeal cancer cells. Lin Chung Er Bi Yan Hou Tou Jing Wai Ke Za Zhi 2013, 27(5):250-254.
• [39]Gvozdenovic A, Arlt MJ, Campanile C, Brennecke P, Husmann K, Li Y, Born W, Muff R, Fuchs B: CD44 enhances tumor formation and lung metastasis in experimental osteosarcoma and is an additional predictor for poor patient outcome. J Bone Miner Res 2013, 28(4):838-847.
• [40]Iguchi H, Yokota M, Fukutomi M, Uchimura K, Yonemasu H, Hachitanda Y, Nakao Y, Tanaka Y, Sumii T, Funakoshi A: A possible role of VEGF in osteolytic bone metastasis of hepatocellular carcinoma. J Exp Clin Cancer Res 2002, 21(3):309-313.
• [41]Arii S, Ishigami S, Mori A, Onodera H, Imamura M: Implication of VEGF and MMPs in hepatic metastasis of human colon cancer. Nihon Geka Gakkai Zasshi 1998, 99(7):436-440.
• [42]Ding Y, Zhang H, Zhong M, Zhou Z, Zhuang Z, Yin H, Wang X, Zhu Z: Clinical significance of the uPA system in gastric cancer with peritoneal metastasis. Eur J Med Res 2013, 18(1):28. BioMed Central Full Text
• [43]Sabe H, Hashimoto S, Morishige M, Ogawa E, Hashimoto A, Nam JM, Miura K, Yano H, Onodera Y: The EGFR-GEP100-Arf6-AMAP1 signaling pathway specific to breast cancer invasion and metastasis. Traffic 2009, 10(8):982-993.
• [44]Pryczynicz A, Guzinska-Ustymowicz K, Kemona A, Czyzewska J: Expression of EGF and EGFR strongly correlates with metastasis of pancreatic ductal carcinoma. Anticancer Res 2008, 28(2B):1399-1404.
• [45]Lee JM, Yanagawa J, Peebles KA, Sharma S, Mao JT, Dubinett SM: Inflammation in lung carcinogenesis: new targets for lung cancer chemoprevention and treatment. Crit Rev Onc Hematol 2008, 66(3):208-217.
• [46]Nonaka Y, Iwagaki H, Kimura T, Fuchimoto S, Orita K: Effect of reactive oxygen intermediates on the in vitro invasive capacity of tumor cells and liver metastasis in mice. Int J Cancer 1993, 54(6):983-986.
• [47]Kozaki K, Koshikawa K, Tatematsu Y, Miyaishi O, Saito H, Hida T, Osada H, Takahashi T: Multi-faceted analyses of a highly metastatic human lung cancer cell line NCI-H460-LNM35 suggest mimicry of inflammatory cells in metastasis. Oncogene 2001, 20(31):4228-4234.
• [48]Chen JS, Wang Q, Fu XH, Huang XH, Chen XL, Cao LQ, Chen LZ, Tan HX, Li W, Bi J, Zhang LJ: Involvement of PI3K/PTEN/AKT/mTOR pathway in invasion and metastasis in hepatocellular carcinoma: association with MMP-9. Hepatol Res 2009, 39(2):177-186.
• [49]Cole GW Jr, Alleva AM, Zuo JT, Sehgal SS, Yeow WS, Schrump DS, Nguyen DM: Suppression of pro-metastasis phenotypes expression in malignant pleural mesothelioma by the PI3K inhibitor LY294002 or the MEK inhibitor UO126. Anticancer Res 2006, 26(2A):809-821.
• [50]Liu H, Yang Y, Xiao J, Lv Y, Liu Y, Yang H, Zhao L: COX-2-mediated regulation of VEGF-C in association with lymphangiogenesis and lymph node metastasis in lung cancer. Anat Rec (Hoboken) 2010, 293(11):1838-1846.
• [51]Singh B, Berry JA, Shoher A, Ayers GD, Wei C, Lucci A: COX-2 involvement in breast cancer metastasis to bone. Oncogene 2007, 26(26):3789-3796.
• [52]Gohji K, Nomi M, Hara I, Arakawa S, Kamidono S: Influence of cytokines and growth factors on matrix metalloproteinase-2 production and invasion of human renal cancer. Urol Res 1998, 26(1):33-37.
• [53]O’Keefe RJ, Guise TA: Molecular mechanisms of bone metastasis and therapeutic implications. Clin Orthop Relat Res 2003, (415 Suppl):S100-S104.