臺灣博碩士論文加值系統

中文摘要
薑黃素 (curcumin，CUR) 是薑黃中最具有活性的成分。許多研究指出，CUR是一種強力抗氧化和抗發炎的天然化合物。四氫薑黃素 (tetrahydrocurcumin，THC) 是CUR的主要生理代謝產物之一，具有許多與CUR相同的生理和藥理活性，並在某些系統中，可能會發揮大於CUR的抗氧化活性。然而，THC抑制肝纖維化的機制，還不是很清楚。本研究主要探討，四氫薑黃素抑制二甲基亞硝胺 (dimethylnitrosamine，DMN) 誘發肝纖維化的功效與機制。在細胞實驗中，我們使用TGF-β1誘發星狀細胞-T6，實驗設計為五組分別為控制組、TGF-β1組、THC 5μM組、THC 10μM組、THC 25μM組，分析α-SMA m-RNA與蛋白質表現量，實驗結果顯示THC明顯抑制TGF-β1誘發星狀細胞活化，α-SMA m-RNA與蛋白質表現量均明顯被抑制下來並呈現Dose dependent現象。接著再使用SD品系雄性大鼠管餵四氫薑黃素(10 mg/kg/day, p.o.)共28天，並同時給予DMN注射到大鼠腹腔3次/週(10 mg/kg, i.p.)。三組動物分別為控制組、DMN組、THC組，28天後犧牲。由實驗結果顯示，給予老鼠THC 4週後，發現血清中的GOT、GPT有顯著的下降，肝纖維化的指標也顯著的下降。另外發現參與肝纖維化的重要分子機制α-SMA與膠原蛋白均呈現抑制現象，主要是因為THC抑制了Smad與TGF-β1訊號途徑，因此而干擾了DMN誘發肝纖維化。綜合以上實驗結果，我們發現餵食THC可以顯著的抑制DMN誘發大鼠肝纖維化。本實驗研究證明THC在體內和體外實驗中，具有抑制肝纖維化作用。因此建議 THC可做為保健肝損傷的食品依據。

關鍵詞：Curcumin、Tetrahydrocurcumin、THC、肝纖維化、二甲基亞硝胺、Dimethylnitrosamine、DMN、膠原蛋白、α-SMA

Abstract
Curcumin (CUR) is the most active component of turmeric. Numerous studies have demonstrated the ability of CUR is a potent antioxidant and anti-inflammatory agent. Tetrahydrocurcumin (THC) is one of the major metabolites of curcumin, and exhibits many of the same physiological and pharmacological activities as curcumin and, in some systems, may exert greater antioxidant activity than curcumin. However, the mechanisms of THC in inhibited hepatic fibrosis (HF) remain unknown. This study explored mechanisms and effective of the THC inhibited by DMN-induced hepatic fibrosis. In order to study the mechanisms and effective of the THC inhibited by DMN-induced hepatic fibrosis, we design in vitro study to show that THC inhibited TGF-β1-induce α-smooth muscle actin (α-SMA) deposition in HSC-T6 cells by blocking Smad 2/3 phosphorylation and nuclear translocation. The anti-hepatic fibrosis effect of THC was also performed in vitro. In vivo study that male SD rats were gavaged with THC (10 mg/kg/day, p.o.) for 28 days and simultaneous injection of DMN into rats abdominal for 3 times/week (10 mg/kg, i.p.). Three groups of animals (control, DMN, DMN+THC 10 mg) were sacrificed after 28 days. The experimental results show that we fed rats after 4 weeks of THC, serum GOT, GPT has a significant decline in the indicators of liver fibrosis was also significantly decline. Another important molecular mechanism of α-SMA found to be involved in liver fibrosis and collagen showed a rejection phenomenon, mainly because of the THC inhibition of Smad and TGF-β1 signal pathway, thus interfering with the DMN-induced hepatic fibrosis. Based on the above experimental results, we found that feeding 10 mg/kg of THC can significantly inhibit the DMN-induced hepatic fibrosis in rats. This study suggested that THC exhibit anti-hepatic fibrosis effects in vivo and in vitro. Therefore, we found that THC can be used as the basis for food health liver injury.

Key words： Curcumin, Tetrahydrocurcumin, THC, hepatic fibrosis, Dimethylnitrosamine, DMN, collagen protein, α-SMA

目錄
誌謝 ................................................................................................................................... I
目錄 ............................................................................................................................... II
圖表目錄 ............................................................................................................................ IV
附圖、表目錄.......................................................................................................................... V
縮寫表 ............................................................................................................................. VII
中文摘要 ............................................................................................................................ IX
Abstract ........................................................................................................................... XI
第壹章 研究背景與目的................................................................................................................... 1
第貳章 文獻回顧........................................................................................................................ 5
第一節、薑黃素簡介 ..................................................................................................................... 5
2-1.1. 結構與活性的相關性 .............................................................................................................. 5
2-1.2. 薑黃中天然類似物及其代謝產物 ...................................................................................................... 9
2-1.3. 薑黃素抗發炎與抑制癌的能力 ...................................................................................................... 16
第二節、四氫薑黃素簡介 ................................................................................................................. 17
第三節、肝纖維化相關的細胞與分子機制 ..................................................................................................... 19
2-3.1. 肝星狀細胞 (Hepatic sellate cells，HSCs) ...................................................................................... 22
2-3.2. 庫氏細胞 (Kupffer Cells) ..................................................................................................... 24
2-3.3. 細胞外基質( Extracellular matrix， ECM ) ...................................................................................... 26
2-3.4. 轉型成長因子-β1 (Transforming growth factor-β1，TGF-β1) ....................................................................... 28
2-3.5. 肝纖維化的分子基礎 ............................................................................................................. 29
第四節、生化指標異常和組織病理學的變化過程 ................................................................................................ 31
第五節、抗纖維化策略 .................................................................................................................. 33
第六節、肝纖維化動物模式 ............................................................................................................... 35
第参章 研究目的與實驗架構............................................................................................................... 42
第一節 研究目的 ...................................................................................................................... 42
第二節 實驗架構 ...................................................................................................................... 43
第肆章 研究材料與方法.................................................................................................................. 45
第一節 實驗材料 ...................................................................................................................... 45
4-1.1. 儀器與廠牌 ................................................................................................................... 45
4-1.2. 藥品來源 ..................................................................................................................... 46
第二節 實驗方法 ...................................................................................................................... 48
4-2.1. 細胞實驗 ..................................................................................................................... 48
4-2.2. 肝纖維化動物實驗 ............................................................................................................... 58
4-2.3. 統計分析 ..................................................................................................................... 70
第伍章 實驗結果與討論.................................................................................................................. 71
第一節 結果與討論 ..................................................................................................................... 71
5-1.1. 以細胞實驗纖維化蛋白質表現量 ..................................................................................................... 71
5-1.2. 以RT-PCR分析THC減少纖維化基因之表現 ............................................................................................. 71
5-1.3. 以肝毒性藥物誘導肝纖維模式，動物之體重差異與變化 ..................................................................................... 71
5-1.4. 以肝毒性藥物誘導肝纖維模式，動物之臟器差異與變化 ..................................................................................... 72
5-1.5. 血清生化檢測 .................................................................................................................. 73
5-1.6. 以SR染色法分析THC對肝纖維化病理狀況之改善情形 ...................................................................................... 76
5-1.7. 以H&E染色法分析THC對肝纖維化病理狀況之改善情形 ..................................................................................... 77
5-1.8. -SMA蛋白質表現量變化 .......................................................................................................... 77
5-1.9. THC 阻礙TGF-β1訊息傳遞的影響 ................................................................................................... 78
5-1.10. THC 阻礙p-Smad訊息傳遞的影響 .................................................................................................. 78
第二節 實驗數據 ...................................................................................................................... 80
第陸章 結論.......................................................................................................................... 87

Reference List

Abdel-Rahman, M. K., (2011). Can apricot kernels fatty acids delay the atrophied hepatocytes from progression to fibrosis in dimethylnitrosamine (DMN)-induced liver injury in rats? Lipids Health Dis. 10, 114.

Adler, M., Verset, C., Moreno, G., (2007). [How to prevent complications of live.r cirrhosis?]. Rev.Med.Brux. 28, 270-275.

Aggarwal, B. B., Harikumar, K. B., (2009). Potential therapeutic effects of curcumin, the anti-inflammatory agent, against neurodegenerative, cardiovascular, pulmonary, metabolic, autoimmune and neoplastic diseases. Int.J.Biochem.Cell Biol. 41, 40-59.

Aggarwal, B. B., Kumar, A., Bharti, A. C., (2003). Anticancer potential of curcumin: preclinical and clinical studies. Anticancer Res. 23, 363-398.

Aggarwal, B. B., Sundaram, C., Malani, N., Ichikawa, H., (2007). Curcumin: the Indian solid gold. Adv.Exp.Med.Biol. 595, 1-75.l

Aggarwal, B. B., Sung, B., (2009). Pharmacological basis for the role of curcumin in chronic diseases: an age-old spice with modern targets. Trends Pharmacol.Sci. 30, 85-94.

Ahmad, A., Ahmad, R., (2012). Understanding the mechanism of hepatic fibrosis and potential therapeutic approaches. Saudi.J.Gastroenterol. 18, 155-167.

Ahmad, A., Fatima, R., Maheshwari, V., Ahmad, R., (2011). Effect of N'-nitrosodimethylamine on red blood cell rheology and proteomic profiles of brain in male albino rats. Interdiscip.Toxicol. 4, 125-131.

Ahmad, R., Ahmed, S., Khan, N. U., Hasnain, A. U., (2009). Operculina turpethum attenuates N-nitrosodimethylamine induced toxic liver injury and clastogenicity in rats. Chem.Biol.Interact. 181, 145-153.

Ahsan, H., Parveen, N., Khan, N. U., Hadi, S. M., (1999). Pro-oxidant, anti-oxidant and cleavage activities on DNA of curcumin and its derivatives demethoxycurcumin and bisdemethoxycurcumin. Chem.Biol.Interact. 121, 161-175.

Al-Attar, A. M., (2012). Attenuating effect of Ginkgo biloba leaves extract on liver fibrosis induced by thioacetamide in mice. J.Biomed.Biotechnol. 2012, 761450.

Albanis, E., Friedman, S. L., (2001). Hepatic fibrosis. Pathogenesis and principles of therapy. Clin.Liver Dis. 5, 315-3vi.

Anand, P., Kunnumakkara, A. B., Newman, R. A., Aggarwal, B. B., (2007). Bioavailability of curcumin: problems and promises. Mol.Pharm. 4, 807-818.

Anand, P., Sundaram, C., Jhurani, S., Kunnumakkara, A. B., Aggarwal, B. B., (2008a). Curcumin and cancer: an "old-age" disease with an "age-old" solution. Cancer Lett. 267, 133-164.

Anand, P., Thomas, S. G., Kunnumakkara, A. B., Sundaram, C., Harikumar, K. B., Sung, B., Tharakan, S. T., Misra, K., Priyadarsini, I. K., Rajasekharan, K. N., Aggarwal, B. B., (2008b). Biological activities of curcumin and its analogues (Congeners) made by man and Mother Nature. Biochem.Pharmacol. 76, 1590-1611.

Annoni, G., Weiner, F. R., Zern, M. A., (1992). Increased transforming growth factor-beta 1 gene expression in human liver disease. J.Hepatol. 14, 259-264.

Arendt, E., Ueberham, U., Bittner, R., Gebhardt, R., Ueberham, E., (2005). Enhanced matrix degradation after withdrawal of TGF-beta1 triggers hepatocytes from apoptosis to proliferation and regeneration. Cell Prolif. 38, 287-299.

Arora, R. B., Kapoor, V., Basu, N., Jain, A. P., (1971). Anti-inflammatory studies on Curcuma longa (turmeric). Indian J.Med.Res. 59, 1289-1295.

Arthur, M. J., (2000). Fibrogenesis II. Metalloproteinases and their inhibitors in liver fibrosis. Am.J.Physiol Gastrointest.Liver Physiol 279, G245-G249.

Baars, A. J., Jansen, M., Breimer, D. D., (1979). Xenobiotica-metabolizing enzymes in Drosophila melanogaster: activities of epoxide hydratase and glutathione S-transferase compared with similar activities in rat liver. Mutat.Res. 62, 279-291.

Barry-Hamilton, V., Spangler, R., Marshall, D., McCauley, S., Rodriguez, H. M., Oyasu, M., Mikels, A., Vaysberg, M., Ghermazien, H., Wai, C., Garcia, C. A., Velayo, A. C., Jorgensen, B., Biermann, D., Tsai, D., Green, J., Zaffryar-Eilot, S., Holzer, A., Ogg, S., Thai, D., Neufeld, G., Van, V. P., Smith, V., (2010). Allosteric inhibition of lysyl oxidase-like-2 impedes the development of a pathologic microenvironment. Nat.Med. 16, 1009-1017.

Bataller, R., Brenner, D. A., (2005). Liver fibrosis. J.Clin.Invest 115, 209-218.

Bataller, R., Gines, P., Nicolas, J. M., Gorbig, M. N., Garcia-Ramallo, E., Gasull, X., Bosch, J., Arroyo, V., Rodes, J., (2000).Angiotensin II induces contraction and proliferation of human hepatic stellate cells. Gastroenterology 118, 1149-1156.

Benyon, R. C., Arthur, M. J., (2001). Extracellular matrix degradation and the role of hepatic stellate cells. Semin.Liver Dis. 21, 373-384.

Benyon, R. C., Iredale, J. P., (2000). Is liver fibrosis reversible? Gut 46, 443-446.

Benyon, R. C., Iredale, J. P., Goddard, S., Winwood, P. J., Arthur, M. J., (1996). Expression of tissue inhibitor of metalloproteinases 1 and 2 is increased in fibrotic human liver. Gastroenterology 110, 821-831.

Bonte, F., Noel-Hudson, M. S., Wepierre, J., Meybeck, A., (1997). Protective effect of curcuminoids on epidermal skin cells under free oxygen radical stress. Planta Med. 63, 265-266.

Borkham-Kamphorst, E., van Roeyen, C. R., Ostendorf, T., Floege, J., Gressner, A. M., Weiskirchen, R., (2007). Pro-fibrogenic potential of PDGF-D in liver fibrosis. J.Hepatol. 46, 1064-1074.

Chen, H. W., Yu, S. L., Chen, J. J., Li, H. N., Lin, Y. C., Yao, P. L., Chou, H. Y., Chien, C. T., Chen, W. J., Lee, Y. T., Yang, P. C., (2004). Anti-invasive gene expression profile of curcumin in lung adenocarcinoma based on a high throughput microarray analysis. Mol.Pharmacol. 65, 99-110.

Choi, J. H., Jin, S. W., Kim, H. G., Khanal, T., Hwang, Y. P., Lee, K. J., Choi, C. Y., Chung, Y. C., Lee, Y. C., Jeong, H. G., (2013). Platycodi Radix attenuates dimethylnitrosamine-induced liver fibrosis in rats by inducing Nrf2-mediated antioxidant enzymes. Food Chem.Toxicol. 56, 231-239.

Cleeland, C. S., Bennett, G. J., Dantzer, R., Dougherty, P. M., Dunn, A. J., Meyers, C. A., Miller, A. H., Payne, R., Reuben, J. M., Wang, X. S., Lee, B. N., (2003). Are the symptoms of cancer and cancer treatment due to a shared biologic mechanism? A cytokine-immunologic model of cancer symptoms. Cancer 97, 2919-2925.

Cohen-Naftaly, M., Friedman, S. L., (2011). Current status of novel antifibrotic therapies in patients with chronic liver disease. Therap.Adv.Gastroenterol. 4, 391-417.

Dantzer, R., Kelley, K. W., (2007). Twenty years of research on cytokine-induced sickness behavior. Brain Behav.Immun. 21, 153-160.

De Bleser, P. J., Niki, T., Rogiers, V., Geerts, A., (1997). Transforming growth factor-beta gene expression in normal and fibrotic rat liver. J.Hepatol. 26, 886-893.

Desmouliere, A., Darby, I., Costa, A. M., Raccurt, M., Tuchweber, B., Sommer, P., Gabbiani, G., (1997). Extracellular matrix deposition, lysyl oxidase expression, and myofibroblastic differentiation during the initial stages of cholestatic fibrosis in the rat. Lab Invest 76, 765-778.

Fallowfield, J. A., Iredale, J. P., (2004). Reversal of liver fibrosis and cirrhosis--an emerging reality. Scott.Med.J. 49, 3-6.

Fan, S., Chen, H. N., Wang, C. J., Tseng, W. C., Hsu, H. K., Weng, C. F., (2007). Toona sinensis Roem (Meliaceae) leaf extract alleviates liver fibrosis via reducing TGFbeta1 and collagen. Food Chem.Toxicol. 45, 2228-2236.

Forbes, S. J., Russo, F. P., Rey, V., Burra, P., Rugge, M., Wright, N. A., Alison, M. R., (2004). A significant proportion of myofibroblasts are of bone marrow origin in human liver fibrosis. Gastroenterology 126, 955-963.

Friedman, S. L., (2003). Liver fibrosis -- from bench to bedside. J.Hepatol. 38 Suppl 1, S38-S53.

Friedman, S. L., (2004). Mechanisms of disease: Mechanisms of hepatic fibrosis and therapeutic implications. Nat.Clin.Pract.Gastroenterol.Hepatol. 1, 98-105.

Friedman, S. L., (2008). Mechanisms of hepatic fibrogenesis. Gastroenterology 134, 1655-1669.

Galli, A., Crabb, D., Price, D., Ceni, E., Salzano, R., Surrenti, C., Casini, A., (2000). Peroxisome proliferator-activated receptor gamma transcriptional regulation is involved in platelet-derived growth factor-induced proliferation of human hepatic stellate cells. Hepatology 31, 101-108.

Gasull, X., Bataller, R., Gines, P., Sancho-Bru, P., Nicolas, J. M., Gorbig, M. N., Ferrer, E., Badia, E., Gual, A., Arroyo, V., Rodes, J., (2001). Human myofibroblastic hepatic stellate cells express Ca(2+)-activated K(+) channels that modulate the effects of endothelin-1 and nitric oxide. J.Hepatol. 35, 739-748.

George J, Suguna L, Jayalakshmi R, Chandrakasan G, (2006). Efficacy of silymarin and curcumin on dimethylnitrosamine induced liver fibrosis in rats.

George, J., Chandrakasan, G., (1996). Glycoprotein metabolism in dimethylnitrosamine induced hepatic fibrosis in rats. Int.J.Biochem.Cell Biol. 28, 353-361.

George, J., Rao, K. R., Stern, R., Chandrakasan, G., (2001). Dimethylnitrosamine-induced liver injury in rats: the early deposition of collagen. Toxicology 156, 129-138.

Goel, A., Jhurani, S., Aggarwal, B. B., (2008a). Multi-targeted therapy by curcumin: how spicy is it? Mol.Nutr.Food Res. 52, 1010-1030.

Goel, A., Kunnumakkara, A. B., Aggarwal, B. B., (2008b). Curcumin as "Curecumin": from kitchen to clinic. Biochem.Pharmacol. 75, 787-809.

Grenard, P., Bresson-Hadni, S., El, A. S., Chevallier, M., Vuitton, D. A., Ricard-Blum, S., (2001). Transglutaminase-mediated cross-linking is involved in the stabilization of extracellular matrix in human liver fibrosis. J.Hepatol. 35, 367-375.

Gressner, A. M., Weiskirchen, R., Breitkopf, K., Dooley, S., (2002). Roles of TGF-beta in hepatic fibrosis. Front Biosci. 7, d793-d807.

Han, J., Hajjar, D. P., Tauras, J. M., Feng, J., Gotto, A. M., Jr., Nicholson, A. C., (2000). Transforming growth factor-beta1 (TGF-beta1) and TGF-beta2 decrease expression of CD36, the type B scavenger receptor, through mitogen-activated protein kinase phosphorylation of peroxisome proliferator-activated receptor-gamma. J.Biol.Chem. 275, 1241-1246.

Hassaninasab, A., Hashimoto, Y., Tomita-Yokotani, K., Kobayashi, M., (2011). Discovery of the curcumin metabolic pathway involving a unique enzyme in an intestinal microorganism. Proc.Natl.Acad.Sci.U.S.A 108, 6615-6620.

Heidelbaugh, J. J., Bruderly, M., (2006). Cirrhosis and chronic liver failure: part I. Diagnosis and evaluation. Am.Fam.Physician 74, 756-762.

Hoehle, S. I., Pfeiffer, E., Solyom, A. M., Metzler, M., (2006). Metabolism of curcuminoids in tissue slices and subcellular fractions from rat liver. J.Agric.Food Chem. 54, 756-764.

Hong, S. W., Jung, K. H., Zheng, H. M., Lee, H. S., Suh, J. K., Park, I. S., Lee, D. H., Hong, S. S., (2010). The protective effect of resveratrol on dimethylnitrosamine-induced liver fibrosis in rats. Arch.Pharm.Res. 33, 601-609.

Hyon, M. K., Kwon, E., Choi, H. J., Kang, B. C., (2011). Dimethylnitrosamine-induced liver fibrosis and recovery in NOD/SCID mice. J.Vet.Med.Sci. 73, 739-745.

Iimuro, Y., Nishio, T., Morimoto, T., Nitta, T., Stefanovic, B., Choi, S. K., Brenner, D. A., Yamaoka, Y., (2003). Delivery of matrix metalloproteinase-1 attenuates established liver fibrosis in the rat. Gastroenterology 124, 445-458.

Inchem, (2010). N-Nitrosodimethylamine (CICADS 38, 2002).

Iredale, J. P., Benyon, R. C., Pickering, J., McCullen, M., Northrop, M., Pawley, S., Hovell, C., Arthur, M. J., (1998). Mechanisms of spontaneous resolution of rat liver fibrosis. Hepatic stellate cell apoptosis and reduced hepatic expression of metalloproteinase inhibitors. J.Clin.Invest 102, 538-549.

Ireson, C., Orr, S., Jones, D. J., Verschoyle, R., Lim, C. K., Luo, J. L., Howells, L., Plummer, S., Jukes, R., Williams, M., Steward, W. P., Gescher, A., (2001). Characterization of metabolites of the chemopreventive agent curcumin in human and rat hepatocytes and in the rat in vivo, and evaluation of their ability to inhibit phorbol ester-induced prostaglandin E2 production. Cancer Res. 61, 1058-1064.

Ireson, C. R., Jones, D. J., Orr, S., Coughtrie, M. W., Boocock, D. J., Williams, M. L., Farmer, P. B., Steward, W. P., Gescher, A. J., (2002). Metabolism of the cancer chemopreventive agent curcumin in human and rat intestine. Cancer Epidemiol.Biomarkers Prev. 11, 105-111.

Issa, R., Zhou, X., Constandinou, C. M., Fallowfield, J., Millward-Sadler, H., Gaca, M. D., Sands, E., Suliman, I., Trim, N., Knorr, A., Arthur, M. J., Benyon, R. C., Iredale, J. P., (2004). Spontaneous recovery from micronodular cirrhosis: evidence for incomplete resolution associated with matrix cross-linking. Gastroenterology 126, 1795-1808.

Jagetia, G. C., Aggarwal, B. B., (2007). "Spicing up" of the immune system by curcumin. J.Clin.Immunol. 27, 19-35.

Jang, J. H., Kang, K. J., Kim, Y. H., Kang, Y. N., Lee, I. S., (2008). Reevaluation of experimental model of hepatic fibrosis induced by hepatotoxic drugs: an easy, applicable, and reproducible model. Transplant.Proc. 40, 2700-2703.

Ju, H. K., Chung, H. W., Lee, H. S., Lim, J., Park, J. H., Lim, S. C., Kim, J. M., Hong, S. S., Kwon, S. W., (2013). Investigation of metabolite alteration in dimethylnitrosamine-induced liver fibrosis by GC-MS. Bioanalysis. 5, 41-51.

Kadiiska, M. B., Gladen, B. C., Baird, D. D., Dikalova, A. E., Sohal, R. S., Hatch, G. E., Jones, D. P., Mason, R. P., Barrett, J. C., (2000). Biomarkers of oxidative stress study: are plasma antioxidants markers of CCl(4) poisoning? Free Radic.Biol.Med. 28, 838-845.

Kalluri, R., Neilson, E. G., (2003). Epithelial-mesenchymal transition and its implications for fibrosis. J.Clin.Invest 112, 1776-1784.

Khopde, S. M., Priyadarsini, K. I., Guha, S. N., Satav, J. G., Venkatesan, P., Rao, M. N., (2000). Inhibition of radiation-induced lipid peroxidation by tetrahydrocurcumin: possible mechanisms by pulse radiolysis. Biosci.Biotechnol.Biochem. 64, 503-509.

Kim, J. E., Kim, A. R., Chung, H. Y., Han, S. Y., Kim, B. S., Choi, J. S., (2003). In vitro peroxynitrite scavenging activity of diarylheptanoids from Curcuma longa. Phytother.Res. 17, 481-484.

Kim, J. M., Araki, S., Kim, D. J., Park, C. B., Takasuka, N., Baba-Toriyama, H., Ota, T., Nir, Z., Khachik, F., Shimidzu, N., Tanaka, Y., Osawa, T., Uraji, T., Murakoshi, M., Nishino, H., Tsuda, H., (1998). Chemopreventive effects of carotenoids and curcumins on mouse colon carcinogenesis after 1,2-dimethylhydrazine initiation. Carcinogenesis 19, 81-85.

Kim, M. R., Kim, H. S., Lee, M. S., Lee, M. J., Jang, J. J., (2005). Cell cycle protein profile of the hepatic stellate cells(HSCs)in dimethylnitrosamine-induced rat hepatic fibrosis. Exp.Mol.Med. 37, 335-342.

Kinnman, N., Goria, O., Wendum, D., Gendron, M. C., Rey, C., Poupon, R., Housset, C., (2001). Hepatic stellate cell proliferation is an early platelet-derived growth factor-mediated cellular event in rat cholestatic liver injury. Lab Invest 81, 1709-1716.

Kitamura, K., Nakamoto, Y., Akiyama, M., Fujii, C., Kondo, T., Kobayashi, K., Kaneko, S., Mukaida, N., (2002). Pathogenic roles of tumor necrosis factor receptor p55-mediated signals in dimethylnitrosamine-induced murine liver fibrosis. Lab Invest 82, 571-583.

Kiuchi, F., Goto, Y., Sugimoto, N., Akao, N., Kondo, K., Tsuda, Y., (1993). Nematocidal activity of turmeric: synergistic action of curcuminoids. Chem.Pharm.Bull.(Tokyo) 41, 1640-1643.

Knolle, P. A., Loser, E., Protzer, U., Duchmann, R., Schmitt, E., zum Buschenfelde, K. H., Rose-John, S., Gerken, G., (1997). Regulation of endotoxin-induced IL-6 production in liver sinusoidal endothelial cells and Kupffer cells by IL-10. Clin.Exp.Immunol. 107, 555-561.

Kobayashi, H., Li, Z. X., Yamataka, A., Lane, G. J., Miyano, T., (2002). Clinical evaluation of serum levels of matrix metalloproteinases and tissue inhibitors of metalloproteinases as predictors of progressive fibrosis in postoperative biliary atresia patients. J.Pediatr.Surg. 37, 1030-1033.

Konsman, J. P., Parnet, P., Dantzer, R., (2002). Cytokine-induced sickness behaviour: mechanisms and implications. Trends Neurosci. 25, 154-159.

Kunnumakkara, A. B., Anand, P., Aggarwal, B. B., (2008a). Curcumin inhibits proliferation, invasion, angiogenesis and metastasis of different cancers through interaction with multiple cell signaling proteins. Cancer Lett. 269, 199-225.

Kunnumakkara, A. B., Diagaradjane, P., Guha, S., Deorukhkar, A., Shentu, S., Aggarwal, B. B., Krishnan, S., (2008b). Curcumin sensitizes human colorectal cancer xenografts in nude mice to gamma-radiation by targeting nuclear factor-kappaB-regulated gene products. Clin.Cancer Res. 14, 2128-2136.

la-Kokko, L., Pihlajaniemi, T., Myers, J. C., Kivirikko, K. I., Savolainen, E. R., (1987). Gene expression of type I, III and IV collagens in hepatic fibrosis induced by dimethylnitrosamine in the rat. Biochem.J. 244, 75-79.

Lai, C. S., Wu, J. C., Yu, S. F., Badmaev, V., Nagabhushanam, K., Ho, C. T., Pan, M. H., (2011). Tetrahydrocurcumin is more effective than curcumin in preventing azoxymethane-induced colon carcinogenesis. Mol.Nutr.Food Res. 55, 1819-1828.

Leandro, G., Mangia, A., Hui, J., Fabris, P., Rubbia-Brandt, L., Colloredo, G., Adinolfi, L. E., Asselah, T., Jonsson, J. R., Smedile, A., Terrault, N., Pazienza, V., Giordani, M. T., Giostra, E., Sonzogni, A., Ruggiero, G., Marcellin, P., Powell, E. E., George, J., Negro, F., (2006). Relationship between steatosis, inflammation, and fibrosis in chronic hepatitis C: a meta-analysis of individual patient data. Gastroenterology 130, 1636-1642.

Lee, M. F., Liu, M. L., Cheng, A. C., Tsai, M. L., Ho, C. T., Liou, W. S., Pan, M. H., (2013a). Pterostilbene inhibits dimethylnitrosamine-induced liver fibrosis in rats. Food Chem. 138, 802-807.

Lee, M. F., Tsai, M. L., Sun, P. P., Chien, L. L., Cheng, A. C., Ma, N. J., Ho, C. T., Pan, M. H., (2013b). Phyto-power dietary supplement potently inhibits dimethylnitrosamine-induced liver fibrosis in rats. Food Funct. 4, 470-475.

Liu, C., Gaca, M. D., Swenson, E. S., Vellucci, V. F., Reiss, M., Wells, R. G., (2003). Smads 2 and 3 are differentially activated by transforming growth factor-beta (TGF-beta ) in quiescent and activated hepatic stellate cells. Constitutive nuclear localization of Smads in activated cells is TGF-beta-independent. J.Biol.Chem. 278, 11721-11728.

Liu, M. L., Mars, W. M., Zarnegar, R., Michalopoulos, G. K., (1994). Collagenase pretreatment and the mitogenic effects of hepatocyte growth factor and transforming growth factor-alpha in adult rat liver. Hepatology 19, 1521-1527.

Lotersztajn, S., Julien, B., Teixeira-Clerc, F., Grenard, P., Mallat, A., (2005). Hepatic fibrosis: molecular mechanisms and drug targets. Annu.Rev.Pharmacol.Toxicol. 45, 605-628.

Lu, X., Liu, P., Xu, G. F., Liu, C. H., Li, F. H., Liu, C., (2003). [The role of hepatic sinusoid capillarization during the formation of portal hypertension in fibrotic rats induced by dimethylnitrosamine]. Zhonghua Gan Zang.Bing.Za Zhi. 11, 595-598.

Luster, M. I., Germolec, D. R., Yoshida, T., Kayama, F., Thompson, M., (1994). Endotoxin-induced cytokine gene expression and excretion in the liver. Hepatology 19, 480-488.

Maher, J. J., Lozier, J. S., Scott, M. K., (1998). Rat hepatic stellate cells produce cytokine-induced neutrophil chemoattractant in culture and in vivo. Am.J.Physiol 275, G847-G853.

Mathurin, P., Xiong, S., Kharbanda, K. K., Veal, N., Miyahara, T., Motomura, K., Rippe, R. A., Bachem, M. G., Tsukamoto, H., (2002). IL-10 receptor and coreceptor expression in quiescent and activated hepatic stellate cells. Am.J.Physiol Gastrointest.Liver Physiol 282, G981-G990.

Matsui, Y., Okuda, Y., Nakagawa, M., Kwon, A. H., Minoura, T., Hiramatsu, Y., Uetsuji, S., Kamiyama, Y., (1994). Effect of hepatocyte volume on energy status in the cirrhotic rat liver. J.Gastroenterol.Hepatol. 9, 613-619.

Matsumoto, K., Nakamura, T., (1996). Emerging multipotent aspects of hepatocyte growth factor. J.Biochem. 119, 591-600.

Mir, A. I., Kumar, B., Tasduq, S. A., Gupta, D. K., Bhardwaj, S., Johri, R. K., (2007). Reversal of hepatotoxin-induced
pre-fibrogenic events by Emblica officinalis--a histological study. Indian J.Exp.Biol. 45, 626-629.

Mohammed, F. F., Pennington, C. J., Kassiri, Z., Rubin, J. S., Soloway, P. D., Ruther, U., Edwards, D. R., Khokha, R., (2005). Metalloproteinase inhibitor TIMP-1 affects hepatocyte cell cycle via HGF activation in murine liver regeneration. Hepatology 41, 857-867.

Mokashi, V., Li, L., Porter, T. D., (2003). Cytochrome b5 reductase and cytochrome b5 support the CYP2E1-mediated activation of nitrosamines in a recombinant Ames test. Arch.Biochem.Biophys. 412, 147-152.

Muhammed Majeed, Vladimir Badmaev, Uma Shivakumar, R.Rajendran, (1995). Curcuminoids: antioxidant Phytonutrients.

Mukhopadhyay, A., Basu, N., Ghatak, N., Gujral, P. K., (1982). Anti-inflammatory and irritant activities of curcumin analogues in rats. Agents Actions 12, 508-515.

Murugan, P., Pari, L., (2006a). Antioxidant effect of tetrahydrocurcumin in streptozotocin-nicotinamide induced diabetic rats. Life Sci. 79, 1720-1728.

Murugan, P., Pari, L., (2006b). Effect of tetrahydrocurcumin on lipid peroxidation and lipids in streptozotocin-nicotinamide-induced diabetic rats. Basic Clin.Pharmacol.Toxicol. 99, 122-127.

Murugan, P., Pari, L., (2006c). Effect of tetrahydrocurcumin on plasma antioxidants in streptozotocin-nicotinamide experimental diabetes. J.Basic Clin.Physiol Pharmacol. 17, 231-244.

Murugan, P., Pari, L., (2005). Effect of tetrahydrocurcumin on erythromycin estolate-induced lipid peroxidation in rats. J.Basic Clin.Physiol Pharmacol. 16, 1-15.

Naito, M., Wu, X., Nomura, H., Kodama, M., Kato, Y., Kato, Y., Osawa, T., (2002). The protective effects of tetrahydrocurcumin on oxidative stress in cholesterol-fed rabbits. J.Atheroscler.Thromb. 9, 243-250.

Nie, Q. H., Duan, G. R., Luo, X. D., Xie, Y. M., Luo, H., Zhou, Y. X., Pan, B. R., (2004). Expression of TIMP-1 and TIMP-2 in rats with hepatic fibrosis. World J.Gastroenterol. 10, 86-90.

Okada, K., Wangpoengtrakul, C., Tanaka, T., Toyokuni, S., Uchida, K., Osawa, T., (2001). Curcumin and especially tetrahydrocurcumin ameliorate oxidative stress-induced renal injury in mice. J.Nutr. 131, 2090-2095.

Osawa, T., Sugiyama, Y., Inayoshi, M., Kawakishi, S., (1995). Antioxidative activity of tetrahydrocurcuminoids. Biosci.Biotechnol.Biochem. 59, 1609-1612.

Pan, M. H., (2000). Studies on the metabolism and bioactivity of curcumin and the molecular mechanism of anti-inflammation and apoptosis by polyphenols.

Pan, M. H., Huang, T. M., Lin, J. K., (1999). Biotransformation of curcumin through reduction and glucuronidation in mice. Drug Metab Dispos. 27, 486-494.

Pan, M. H., Lin-Shiau, S. Y., Lin, J. K., (2000). Comparative studies on the suppression of nitric oxide synthase by curcumin and its hydrogenated metabolites through down-regulation of IkappaB kinase and NFkappaB activation in macrophages. Biochem.Pharmacol. 60, 1665-1676.

Pari, L., Amali, D. R., (2005). Protective role of tetrahydrocurcumin (THC) an active principle of turmeric on chloroquine induced hepatotoxicity in rats. J.Pharm.Pharm.Sci. 8, 115-123.

Pari, L., Murugan, P., (2004). Protective role of tetrahydrocurcumin against erythromycin estolate-induced hepatotoxicity. Pharmacol.Res. 49, 481-486.

Pari, L., Murugan, P., (2006). Tetrahydrocurcumin: effect on chloroquine-mediated oxidative damage in rat kidney. Basic Clin.Pharmacol.Toxicol. 99, 329-334.

Pari, L., Murugan, P., (2007d). Antihyperlipidemic effect of curcumin and tetrahydrocurcumin in experimental type 2 diabetic rats. Ren Fail. 29, 881-889.

Pari, L., Murugan, P., (2007c). Changes in glycoprotein components in streptozotocin--nicotinamide induced type 2 diabetes: influence of tetrahydrocurcumin from Curcuma longa. Plant Foods Hum.Nutr. 62, 25-29.

Pari, L., Murugan, P., (2007b). Influence of tetrahydrocurcumin on tail tendon collagen contents and its properties in rats with streptozotocin-nicotinamide-induced type 2 diabetes. Fundam.Clin.Pharmacol. 21, 665-671.

Pari, L., Murugan, P., (2007a). Tetrahydrocurcumin prevents brain lipid peroxidation in streptozotocin-induced diabetic rats. J.Med.Food 10, 323-329.

Pari, L., Murugan, P., (2005). Effect of tetrahydrocurcumin on blood glucose, plasma insulin and hepatic key enzymes in streptozotocin induced diabetic rats. J.Basic Clin.Physiol Pharmacol. 16, 257-274.

Patro, B. S., Rele, S., Chintalwar, G. J., Chattopadhyay, S., Adhikari, S., Mukherjee, T., (2002). Protective activities of some phenolic 1,3-diketones against lipid peroxidation: possible involvement of the 1,3-diketone moiety. Chembiochem. 3, 364-370.

Perz, J. F., Armstrong, G. L., Farrington, L. A., Hutin, Y. J., Bell, B. P., (2006). The contributions of hepatitis B virus and hepatitis Cvirus infections to cirrhosis and primary liver cancer worldwide. J.Hepatol. 45, 529-538.

Pinzani, M., (2002). PDGF and signal transduction in hepatic stellate cells. Front Biosci. 7, d1720-d1726.

Pinzani, M., Gesualdo, L., Sabbah, G. M., Abboud, H. E., (1989). Effects of platelet-derived growth factor and other polypeptide mitogens on DNA synthesis and growth of cultured rat liver fat-storing cells. J.Clin.Invest 84, 1786-1793.

Pinzani, M., Marra, F., (2001). Cytokine receptors and signaling in hepatic stellate cells. Semin.Liver Dis. 21, 397-416.

Ravindranath, V., Chandrasekhara, N., (1981). In vitro studies on the intestinal absorption of curcumin in rats. Toxicology 20, 251-257.

Reagan-Shaw, S., Nihal, M., Ahmad, N., (2008). Dose translation from animal to human studies revisited. FASEB J. 22, 659-661.

Reimann, T., Hempel, U., Krautwald, S., Axmann, A., Scheibe, R., Seidel, D., Wenzel, K. W., (1997). Transforming growth factor-beta1 induces activation of Ras, Raf-1, MEK and MAPK in rat hepatic stellate cells. FEBS Lett. 403, 57-60.

Rockey, D. C., Chung, J. J., (1996). Endothelin antagonism in experimental hepatic fibrosis. Implications for endothelin in the pathogenesis of wound healing. J.Clin.Invest 98, 1381-1388.

Rojkind M, Mourelle M, (1988). Modification during regeneration and repair in collagen. The liver as bioecological system. pp. 137-59.

Rojkind, M., Giambrone, M. A., Biempica, L., (1979). Collagen types in normal and cirrhotic liver. Gastroenterology 76, 710-719.

S.Vogel, M.V.Gamble, W.S.Blaner, (1999). Biosynthesis, Absorption, Metabolism and Transport of Retinoids In Retinoids: The biochemical and molecular basis of vitamin A and retinoid action. pp. 31-95.

Saile, B., DiRocco, P., Dudas, J., El-Armouche, H., Sebb, H., Eisenbach, C., Neubauer, K., Ramadori, G., (2004). IGF-I induces DNA synthesis and apoptosis in rat liver hepatic stellate cells (HSC) but DNA synthesis and proliferation in rat liver myofibroblasts (rMF). Lab Invest 84, 1037-1049.

Sandur, S. K., Pandey, M. K., Sung, B., Ahn, K. S., Murakami, A., Sethi, G., Limtrakul, P., Badmaev, V., Aggarwal, B. B., (2007). Curcumin, demethoxycurcumin, bisdemethoxycurcumin, tetrahydrocurcumin and turmerones differentially regulate anti-inflammatory and anti-proliferative responses through a ROS-independent mechanism. Carcinogenesis 28, 1765-1773.

Schnur, J., Olah, J., Szepesi, A., Nagy, P., Thorgeirsson, S. S., (2004). Thioacetamide-induced hepatic fibrosis in transforming growth factor beta-1 transgenic mice. Eur.J.Gastroenterol.Hepatol. 16, 127-133.

Shiba, M., Shimizu, I., Yasuda, M., Ii, K., Ito, S., (1998). Expression of type I and type III collagens during the course of dimethylnitrosamine-induced hepatic fibrosis in rats. Liver 18, 196-204.

Shishodia, S., Chaturvedi, M. M., Aggarwal, B. B., (2007). Role of curcumin in cancer therapy. Curr.Probl.Cancer 31, 243-305.

Shishodia, S., Sethi, G., Aggarwal, B. B., (2005). Curcumin: getting back to the roots. Ann.N.Y.Acad.Sci. 1056, 206-217.

Simon, A., Allais, D. P., Duroux, J. L., Basly, J. P., Durand-Fontanier, S., Delage, C., (1998). Inhibitory effect of curcuminoids on MCF-7 cell proliferation and structure-activity relationships. Cancer Lett. 129, 111-116.

Somparn, P., Phisalaphong, C., Nakornchai, S., Unchern, S., Morales, N. P., (2007). Comparative antioxidant activities of curcumin and its demethoxy and hydrogenated derivatives. Biol.Pharm.Bull. 30, 74-78.

Sreejayan, N., Rao, M. N., (1996). Free radical scavenging activity of curcuminoids. Arzneimittelforschung. 46, 169-171.

Subramanian, M., Sreejayan, Rao, M. N., Devasagayam, T. P., Singh, B. B., (1994). Diminution of singlet oxygen-induced DNA damage by curcumin and related antioxidants. Mutat.Res. 311, 249-255.

Sugiyama, Y., Kawakishi, S., Osawa, T., (1996). Involvement of the beta-diketone moiety in the antioxidative mechanism of tetrahydrocurcumin. Biochem.Pharmacol. 52, 519-525.

Sumiyoshi, K., Nakao, A., Setoguchi, Y., Okumura, K., Tsuboi, R., Ogawa, H., (2003). Smads regulate collagen gel contraction by human dermal fibroblasts. Br.J.Dermatol. 149, 464-470.

Sun, K., Wang, Q., Huang, X. H., (2006). PPAR gamma inhibits growth of rat hepatic stellate cells and TGF beta-induced connective tissue growth factor expression. Acta Pharmacol.Sin. 27, 715-723.

Takahara, T., Furui, K., Yata, Y., Jin, B., Zhang, L. P., Nambu, S., Sato, H., Seiki, M., Watanabe, A., (1997). Dual expression of matrix metalloproteinase-2 and membrane-type 1-matrix metalloproteinase in fibrotic human livers. Hepatology 26, 1521-1529.

Tokudome, S., Imaeda, N., Tokudome, Y., Kuriki, K., Cheng, J., Fujii, T., Goto, C., Nagaya, T., Ichikawa, H., Okuyama, H., (2002). Correspondence re: Terry et al., No association between fat and fatty acids intake and risk of colorectal cancer. Cancer Epidemiol. Biomark. Prev., 10: 913-914, 2001. Cancer Epidemiol.Biomarkers Prev. 11, 217-218.

Toyama, T., Nakamura, H., Harano, Y., Yamauchi, N., Morita, A., Kirishima, T., Minami, M., Itoh, Y., Okanoue, T., (2004). PPARalpha ligands activate antioxidant enzymes and suppress hepatic fibrosis in rats.Biochem.Biophys.Res.Commun. 324, 697-704.

Uyama, N., Zhao, L., Van, R. E., Hirako, Y., Reynaert, H., Adams, D. H., Xue, Z., Li, Z., Robson, R., Pekny, M., Geerts, A., (2006). Hepatic stellate cells express synemin, a protein bridging intermediate filaments to focal adhesions. Gut 55, 1276-1289.

Vendemiale, G., Grattagliano, I., Caruso, M. L., Serviddio, G., Valentini, A. M., Pirrelli, M., Altomare, E., (2001). Increased oxidative stress in dimethylnitrosamine-induced liver fibrosis in the rat: effect of N-acetylcysteine and interferon-alpha.Toxicol.Appl.Pharmacol. 175, 130-139.

Wells, R. G., (2008). The role of matrix stiffness in regulating cell behavior. Hepatology 47, 1394-1400.

Wells, R. G., Kruglov, E., Dranoff, J. A., (2004). Autocrine release of TGF-beta by portal fibroblasts regulates cell growth. FEBS Lett. 559, 107-110.

Wheeler, M. D., (2003). Endotoxin and Kupffer cell activation in alcoholic liver disease. Alcohol Res.Health 27, 300-306.

Yasuda, H., Imai, E., Shiota, A., Fujise, N., Morinaga, T., Higashio, K., (1996). Antifibrogenic effect of a deletion variant of hepatocyte growth factor on liver fibrosis in rats. Hepatology 24, 636-642.

Yi-Pin L, (2006). Alcohol- and carbon tetrachloride-induced liver injury as animal models for hepatoprotection evaluation.

Yoshiji, H., Kuriyama, S., Miyamoto, Y., Thorgeirsson, U. P., Gomez, D. E., Kawata, M., Yoshii, J., Ikenaka, Y., Noguchi, R., Tsujinoue, H., Nakatani, T., Thorgeirsson, S. S., Fukui, H., (2000). Tissue inhibitor of metalloproteinases-1 promotes liver fibrosis development in a transgenic mouse model. Hepatology 32, 1248-1254.

Zhou, Y., Zheng, S., Lin, J., Zhang, Q. J., Chen, A., (2007). The interruption of the PDGF and EGF signaling pathways by curcumin stimulates gene expression of PPARgamma in rat activated hepatic stellate cell in vitro. Lab Invest 87, 488-498.

吳昭新, (1996). 脂質檢查－膽固醇和三酸甘油脂（中性脂肪）.

林杰樑, (2003). 生活中的毒.

曾春典, (1993). 中老年人的心臟血管疾病.