Abstract
The injurious effects of reactive oxygen species on osteoblasts and the potential protective role played by green tea polyphenols (GtPP) were investigated using primarily cultured rat calvarial osteoblasts. Oxidative stress was induced in cultured osteoblasts, either by adding 100 mmol/L H2O2 or by the action of 40 U/L xanthine oxidase (XO) in the presence of xanthine (250 μmol/L). After incubation, the cellular viability, function and morphology were evaluated. Both treatments produced a significant reduction in osteoblast viability, as assessed by a two-colored fluorescence staining method combined with flow cytometric analysis and MTT assay. A significant reduction in the alkaline phosphatase activity was observed after H2O2 addition, whereas XO did not have the same effect. On the microscopic observations, the morphological changes and intracellular ultrastructural damages were remarkably induced by both treatments. The H2O2-induced alterations were prevented by pre-incubating the osteoblasts with 200 μg/ml GtPP for 1 h. When the oxidative stress was induced by XO, the cellular viability and morphology was also maintained at the same polyphenol concentration. These results demonstrate that GtPP can act as a biological antioxidant in a cell culture experimental model and protect cells from oxidative stress-induced toxicity.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Ahmad N, Feyes DK, Nieminen AL, Agarwal R, Mukhtar H. Green tea constituent epigallocatechin-3-gallate and induction of apoptosis and cell cycle arrest in human carcinoma cells. J Natl Cancer Inst. 1997;89:1881–6.
Ahmad N, Gupta S, Mukhtar H. Green tea polyphenol epigallocatechin-3-gallate differentially modulates nuclear factor κB in cancer cells versus normal cells. Arch Biochem Biophys. 2000;376:338–46.
Aronow MA, Gerstenfeld LC, Owen TA. Factors that promote progressive development of the osteoblast phenotype in cultured fetal rat calvaria cells. J Cell Phys. 1990;143:213–21.
Berridge V, Tan AS. Characterization of the cellular reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT): subcellular localization, substrate dependence, and involvement of mitochondrial electron transport in MTT reduction. Arch Biochem Biophys. 1993;303:474–82.
Block G, Patterson B, Subar A. Fruit, vegetables and cancer prevention: a review of the epidemiological evidence. Nutr Cancer. 1992;18:1–29.
Cao Y, Cao R. Angiogenesis inhibited by drinking tea. Nature. 1999;398:381.
Carmichael J, De Graff WG, Minna JD, Mitchel JB. Evaluation of a tetrazolium based semiautomated colorimetric assay: assessment of radiosensitivity. Cancer Res. 1987;47:943–6.
Chen HW, Huang HC. Effect of curcumin on cell cycle progression and apoptosis in vascular smooth muscle cells. Br J Pharmacol. 1998;124:1029–40.
Decker EA. The role of phenolics, conjugated linoleic acid, carnosine, and pyrroloquinoline quinone as nonessential dietary antioxidants. Nutr Rev. 1995;53:49–58.
Di Mascio P, Murphy ME, Sies H. Antioxidant defense systems: the role of carotenoids, tocopherols and thiols. Am J Clin Nutr. 1991;53(S1):194S–200S.
Gupta S, Ahmad N, Nieminen AL, Mukhtar H. Growth inhibition, cell-cycle dysregulation, and induction of apoptosis by green tea constituent (−)-epigallocatechin-3-gallate in androgen-sensitive and androgen-insensitive human prostate carcinoma cells. Toxicol Appl Pharmacol. 2000;164:82–90.
Halliwell B. Free radicals and antioxidants: a personal view. Nutr Rev. 1994;52:253–65.
Han D-W, Suh H, Park YH, Cho BK, Hyon S-H, Park J-C. Preservation of human saphenous vein against reactive oxygen species-induced oxidative stress by green tea polyphenol pre-treatment. Artif Organs. 2003 (in press).
Harman D. Aging: A theory based on free radical and radiation chemistry. J Gerontol. 1956;11:298–300.
Holian O, Walter RJ. Resveratrol inhibits the proliferation of normal human keratinocytes in vitro. J Cell Biochem. 2001;81:55–62.
Hosoya S, Suzuki H, Yamamoto M, Kobayashi K, Abiko Y. Alkaline phosphatase and type I collagen gene expressions were reduced by hydroxyl radical-treated fibronectin substratum. Mol Genet Metab. 1998;65:31–4.
Hsieh TC, Wu JM. Differential effects on growth, cell cycle arrest, and induction of apoptosis by resveratrol in human prostate cancer cell lines. Exp Cell Res. 1999;25:109–15.
Hyon S-H, Kim D-H. Long-term preservation of rat pancreatic islets under physiological conditions. J Biotechnol. 2001;85:241–6.
Ikeda T, Nagai Y, Yamaguchi A, Yokose S, Yoshiki S. Aged-related reduction in bone matrix protein mRNA expression in rat bone tissue: application of histomorphometry to in situ hybridization. Bone. 1995;19:17–23.
Manna C, Galletti P, Cucciolla V, Moltedo O, Leone A, Zappia V. The protective effect of the olive oil polyphenol (3,4-dihydroxyphenyl)-ethanol counteracts reactive oxygen metabolite-induced cytotoxicity in Caco-2 cells. J Nutr. 1997;127:286–92.
Nakayama M, Suzuki K, Toda M, Okubo S, Hara Y, Shimamura T. Inhibition of the infectivity of influenza virus by tea polyphenols. Antivir Res. 1993;21:289–99.
Pal S, Choudhuri T, Chattopadhyay S, et al. Mechanisms of curcumin-induced apoptosis of Ehrlich's ascites carcinoma cells. Biochem Biophys Res Commun. 2001;288:658–65.
Park J-C, Sung HJ, Lee DH, Park YH, Cho BK, Suh H. Specific determination of endothelial cell viability in the whole cell fraction from cryopreserved canine femoral veins using flow cytometry. Artif Organs. 2000;24:829–33.
Park J-C, Sung HJ, Lee DH, Park YH, Cho BK, Suh H. Viability of cells in cryopreserved canine cardiovascular organs for transplantation. Yonsei Med J. 2000;41:556–62.
Sies H. Oxidative stress, oxidants and antioxidants. New York: Academic Press; 1991.
Suh H, Park J-C, Han D-W, Lee DH, Han CD. A bone replaceable artificial bone substitute: cytotoxicity, cell adhesion, proliferation, and alkaline phosphatase activity. Artif Organs. 2001;25:14–21.
Suzuki H, Hayakawa M, Kobayashi K, Takiguchi H, Abiko Y. H2O2-derived free radicals treated fibronectin substratum reduces the bone nodule formation of rat calvarial osteoblast. Mech Ageing Dev. 1997;98:113–25.
Thang PT, Patrick S, Teik LS, Yung CS. Anti-oxidant effects of the extracts from the leaves of Chromolaena odorata on human dermal fibroblasts and epidermal keratinocytes against hydrogen peroxide and hypoxanthine-xanthine oxidase induced damage. Burns. 2001;27:319–27.
Wlodarski KH, Reddi AH. Alkaline phosphatase as a marker of osteoinductive cells. Calcif Tissue Int. 1986;39:382–5.
Yang CS, Wang Z-Y. Tea and cancer. J Natl Cancer Inst. 1993;85:1038–49.
Yang CS, Yang GY, Landau JM, Kim S, Liao J. Tea and tea polyphenols inhibit cell hyperproliferation, lung tumorigenesis, and tumor progression. Exp Lung Res. 1998;24:629–39.
Yang CS, Prabhu S, Landau J. Prevention of carcinogenesis by tea polyphenols. Drug Metab Rev. 2001;33:237–53.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Park, Y., Han, DW., Suh, H. et al. Protective effects of green tea polyphenol against reactive oxygen species-induced oxidative stress in cultured rat calvarial osteoblast. Cell Biol Toxicol 19, 325–337 (2003). https://doi.org/10.1023/B:CBTO.0000004986.51081.c5
Issue Date:
DOI: https://doi.org/10.1023/B:CBTO.0000004986.51081.c5