臺灣博碩士論文加值系統

眼睛黃斑部位於眼球後正中心，是一層非常精密的組織層。在已開發國家中，老年黃斑病變是50歲以上老年人視力退化的主要原因之一。最近有文獻指出老年性黃斑部的病人其Bruch’s membrane有膽固醇的聚集，且可以發現視網膜色素上皮細胞會內噬低密度脂蛋白和氧化型低密度脂蛋白。氧化型低密度脂蛋白的攝入可能會造成視網膜細胞的凋亡。於學術研究中證實，桑椹水萃物和花青素具有調控脂質代謝，減少細胞攝入膽固醇的功能。因此，在本研究使用肥胖模式誘導眼病變，以證實桑椹水萃物和花青素，其功效機轉為何。
　　在動物模式中，將倉鼠餵食高油脂飲食(0.2%膽固醇及10%玉米油)誘導其肥胖，共為期八週，並觀察各組體重，肝臟重，血清和肝臟中三酸甘油酯、膽固醇、低密度脂蛋白和高密度脂蛋白，以及眼睛組織H&E stain和IHC stain的觀察。結果發現，桑椹水萃物可降低倉鼠中肝臟脂肪肝的現象，以及降低眼球中脂防泡的形成和oxLDL 被吞噬到視網膜細胞中。在細胞模式中，觀察桑椹花青素對於ARPE-19視網膜色素上層細胞以oxLDL誘導凋亡的影響。結果顯示，桑椹花青素可以有效抑制oxLDL造成ARPE-19的氧化傷害，降低oxLDL receptor的表現，和減少細胞凋亡的現象。以上的結果可以得知桑椹水萃物和花青素具有降低脂質在視網膜累積的情形和減少oxLDL對於視網膜所造成的傷害。

The macula is the central posterior portion of the retina, a quite fine organization level. In the developed countries, age-related macular degeneration(AMD) is the leading cause of irreversible blindness in people about 50 years of age or older. Recent scientific evidences showed that a huge of cholesterol accumulate in Bruch’s membrane of the patients with age-related macular degeneration. It is also found that LDL and oxLDL can be uptaken by retinal pigmental epithelial cells. Although uptaking oxLDL would lead to foam cell formation or cell death in macrophage, the effect of oxLDL is still remained unclear in retinal cell.
Mulberry had been demonstrated previously to possess antioxidative and antitumor promoting effects. In this study, we aimed to examine the effect of mulberry water effects (MWE) and mulberry anthocyanin extract (MAE) in AMD. In animal model, hamsters were fed a high fat diet (0.2% of cholesterol and 10% of corn oil) for 8 weeks accompanying by various concentrations of MWE. The results showed that MWE reduced the lipid accumulation, body weight , fatty liver, and the levels of triglyceride and total cholesterol in obese hamsters. Furthermore, we examined the effect of MAE in oxLDL treated retinal pigment epithelium cell (ARPE-19). The results showed that 100µg/mL LDL oxidized by 10µM CuSO4 can induce ARPE cell apoptosis and lipid peroxidation. It can be significantly improved after pretreating with MAE for 24 hours,. The results also showed that scavenger receptors (CD36, SREBP-1) , oxLDL, apoptotic-related protein (caspase3, AIF) and pro-survival proteins in mitochondrial membrane (Bcl-2, Bcl-xL) were involved in regulating the cell apoptosis.
In conclusion, MWE can decelerate lipid and oxLDL accumulating in retinal, and one of its constituents, MAE, can improve oxLDL-induced ARPE cell apoptosis. Base on these evidences, MWE possess the ability to protect retina from age-related macular degeneration caused by obesity.

壹、 中文摘要……………………………………………………1
貳、 英文摘要……………………………………………………2
參、 縮寫檢索表…………………………………………………3
肆、 序論…………………………………………………………4
伍、 研究動機……………………………………………………18
陸、 實驗架構……………………………………………………19
柒、 實驗材料與方法……………………………………………21
捌、 實驗藥品與儀器……………………………………………35
玖、 實驗結果……………………………………………………39
壹拾、 討論…………………………………………………………46
壹拾壹、 圖表……………………………………………………50
壹拾貳、 參考文獻………………………………………………68
壹拾參、 附圖……………………………………………………72

劉力鋼 (2009). "桑椹萃取物降低高油脂高膽固醇飲食引起之高血脂與體脂肪形成之作用與分子機制" 中山醫學大學生化暨生物科技研究所.
徐嘉君 (2009). "多酚類化合物對氧化型低密度脂蛋白引起視網膜色素上皮細胞傷害的抑制作用" 中山醫學大學生化暨生物科技研究所.
林佩穎 (2009). "蘋果多酚延緩脂肪細胞分化並在實驗性肥胖中降低脂肪儲存"中山醫學大學生化暨生物科技研究所.
許珊菁 (2005). "鼠模式中高脂飲食，肥胖與脂質調控基因之表現"
台灣大學微生物與生化學研究所
盧宏民 (2003). 中藥大辭典。五洲出版社，台北，p.3016∼3018。

Aaby, K., D. Ekeberg, et al. (2007). "Characterization of phenolic compounds in strawberry (Fragaria x ananassa) fruits by different HPLC detectors and contribution of individual compounds to total antioxidant capacity." J Agric Food Chem 55(11): 4395-406.
Acton, S., A. Rigotti, et al. (1996). "Identification of scavenger receptor SR-BI as a high density lipoprotein receptor." Science 271(5248): 518-20.
Acton, S. L., P. E. Scherer, et al. (1994). "Expression cloning of SR-BI, a CD36-related class B scavenger receptor." J Biol Chem 269(33): 21003-9.
Aviram, M. (1993). "Modified forms of low density lipoprotein and atherosclerosis." Atherosclerosis 98(1): 1-9.
Bustamante, M., F. Diaz, et al. (2007). "Oxidized low density lipoproteins induce apoptosis in human lymphocytes: involvement of mitogen-activated protein kinases." Cell Mol Biol (Noisy-le-grand) 53 Suppl: OL954-64.
Catapano, A. L., F. M. Maggi, et al. (2000). "Low density lipoprotein oxidation, antioxidants, and atherosclerosis." Curr Opin Cardiol 15(5): 355-63.
Clemons, T. E., R. C. Milton, et al. (2005). "Risk factors for the incidence of Advanced Age-Related Macular Degeneration in the Age-Related Eye Disease Study (AREDS) AREDS report no. 19." Ophthalmology 112(4): 533-9.
During, A., S. Doraiswamy, et al. (2008). "Xanthophylls are preferentially taken up compared with beta-carotene by retinal cells via a SRBI-dependent mechanism." J Lipid Res 49(8): 1715-24.
Duthie, S. J. (2007). "Berry phytochemicals, genomic stability and cancer: evidence for chemoprotection at several stages in the carcinogenic process." Mol Nutr Food Res 51(6): 665-74.
Elner, V. M. (2002). "Retinal pigment epithelial acid lipase activity and lipoprotein receptors: effects of dietary omega-3 fatty acids." Trans Am Ophthalmol Soc 100: 301-38.
Febbraio, M., N. A. Abumrad, et al. (1999). "A null mutation in murine CD36 reveals an important role in fatty acid and lipoprotein metabolism." J Biol Chem 274(27): 19055-62.
Gaut, J. P. and J. W. Heinecke (2001). "Mechanisms for oxidizing low-density lipoprotein. Insights from patterns of oxidation products in the artery wall and from mouse models of atherosclerosis." Trends Cardiovasc Med 11(3-4): 103-12.
Gordiyenko, N., M. Campos, et al. (2004). "RPE cells internalize low-density lipoprotein (LDL) and oxidized LDL (oxLDL) in large quantities in vitro and in vivo." Invest Ophthalmol Vis Sci 45(8): 2822-9.
Hassimotto, N. M., M. I. Genovese, et al. (2008). "Absorption and metabolism of cyanidin-3-glucoside and cyanidin-3-rutinoside extracted from wild mulberry (Morus nigra L.) in rats." Nutr Res 28(3): 198-207.
Heinonen, M. (2007). "Antioxidant activity and antimicrobial effect of berry phenolics--a Finnish perspective." Mol Nutr Food Res 51(6): 684-91.
Hou, D. X. (2003). "Potential mechanisms of cancer chemoprevention by anthocyanins." Curr Mol Med 3(2): 149-59.
Isabelle, M., B. L. Lee, et al. (2008). "Peroxyl radical scavenging capacity, polyphenolics, and lipophilic antioxidant profiles of mulberry fruits cultivated in southern China." J Agric Food Chem 56(20): 9410-6.
Jager, R. D., W. F. Mieler, et al. (2008). "Age-related macular degeneration." N Engl J Med 358(24): 2606-17.
Jeon, H. and S. C. Blacklow (2005). "Structure and physiologic function of the low-density lipoprotein receptor." Annu Rev Biochem 74: 535-62.
Jialal, I. and S. Devaraj (1996). "The role of oxidized low density lipoprotein in atherogenesis." J Nutr 126(4 Suppl): 1053S-7S.
Katsube, N., K. Iwashita, et al. (2003). "Induction of apoptosis in cancer cells by Bilberry (Vaccinium myrtillus) and the anthocyanins." J Agric Food Chem 51(1): 68-75.
Klein, B. E., R. Klein, et al. (1992). "Prevalence of age-related lens opacities in a population. The Beaver Dam Eye Study." Ophthalmology 99(4): 546-52.
Kuhnau, J. (1976). "The flavonoids. A class of semi-essential food components: their role in human nutrition." World Rev Nutr Diet 24: 117-91.
Li, J. and J. Yuan (2008). "Caspases in apoptosis and beyond." Oncogene 27(48): 6194-206.
Liu, L. K., H. J. Lee, et al. (2008). "Mulberry anthocyanin extracts inhibit LDL oxidation and macrophage-derived foam cell formation induced by oxidative LDL." J Food Sci 73(6): H113-21.
Mallat, Z. and A. Tedgui (2000). "Apoptosis in the vasculature: mechanisms and functional importance." Br J Pharmacol 130(5): 947-62.
Matsuura, E., K. Kobayashi, et al. (2006). "Oxidative modification of low-density lipoprotein and immune regulation of atherosclerosis." Prog Lipid Res 45(6): 466-86.
Moore, K. J. and M. W. Freeman (2006). "Scavenger receptors in atherosclerosis: beyond lipid uptake." Arterioscler Thromb Vasc Biol 26(8): 1702-11.
Nature Medicine - 12, 511 - 512 (2006)
Nakajima, J. I., I. Tanaka, et al. (2004). "LC/PDA/ESI-MS Profiling and Radical Scavenging Activity of Anthocyanins in Various Berries." J Biomed Biotechnol 2004(5): 241-247.
Ogawa, K., H. Sakakibara, et al. (2008). "Anthocyanin composition and antioxidant activity of the Crowberry (Empetrum nigrum) and other berries." J Agric Food Chem 56(12): 4457-62.
Pawlowska, A. M., W. Oleszek, et al. (2008). "Quali-quantitative analyses of Flavonoids of Morus nigra L. and Morus alba L. (Moraceae) fruits." J Agric Food Chem 56(9): 3377-80.
Prior, R. L. and X. Wu (2006). "Anthocyanins: structural characteristics that result in unique metabolic patterns and biological activities." Free Radic Res 40(10): 1014-28.
Rheaume, E., L. Y. Cohen, et al. (1997). "The large subunit of replication factor C is a substrate for caspase-3 in vitro and is cleaved by a caspase-3-like protease during Fas-mediated apoptosis." EMBO J 16(21): 6346-54.
SanGiovanni, JP (2009-01-21). "Age-Related Eye Disease Study (AREDS)". ClinicalTrials.gov. Retrieved 2009-06-24.
Sasaki, R., N. Nishimura, et al. (2007). "Cyanidin 3-glucoside ameliorates hyperglycemia and insulin sensitivity due to downregulation of retinol binding protein 4 expression in diabetic mice." Biochem Pharmacol 74(11): 1619-27.
Schweitzer, D., G. E. Lang, et al. (2000). "[Age-related maculopathy. Comparative studies of patients, their children and healthy controls]." Ophthalmologe 97(2): 84-90.
Seddon, J. M., U. A. Ajani, et al. (1994). "Dietary carotenoids, vitamins A, C, and E, and advanced age-related macular degeneration. Eye Disease Case-Control Study Group." JAMA 272(18): 1413-20.
Song, W., H. J. Wang, et al. (2009). "Phytochemical profiles of different mulberry (Morus sp.) species from China." J Agric Food Chem 57(19): 9133-40.
Strauss, O. (2005). "The retinal pigment epithelium in visual function." Physiol Rev 85(3): 845-81.
Szajdek, A. and E. J. Borowska (2008). "Bioactive compounds and health-promoting properties of berry fruits: a review." Plant Foods Hum Nutr 63(4): 147-56.
Tan, J. S., J. J. Wang, et al. (2008). "Dietary antioxidants and the long-term incidence of age-related macular degeneration: the Blue Mountains Eye Study." Ophthalmology 115(2): 334-41.
Toscano, L., E. Seghetti, et al. (1975). "Synthesis and central nervous system effects of 8.9-dihydro(1)benzazepino(3,2.1-ak)(1,4)benzodiazepin-1(2H)-ones and (1) binzazepino(3,2,1-jk)(1,4)benzodiazepin-1(2H)-ones." J Med Chem 18(10): 976-81.
Tserentsoodol, N., J. Sztein, et al. (2006). "Uptake of cholesterol by the retina occurs primarily via a low density lipoprotein receptor-mediated process." Mol Vis 12: 1306-18.
Tsuda, T., F. Horio, et al. (2003). "Dietary cyanidin 3-O-beta-D-glucoside-rich purple corn color prevents obesity and ameliorates hyperglycemia in mice." J Nutr 133(7): 2125-30.
Wang, Y. W. and P. J. Jones (2004). "Conjugated linoleic acid and obesity control: efficacy and mechanisms." Int J Obes Relat Metab Disord 28(8): 941-55.
Winkler, B. S., M. E. Boulton, et al. (1999). "Oxidative damage and age-related macular degeneration." Mol Vis 5: 32.
Wu, X., G. R. Beecher, et al. (2006). "Concentrations of anthocyanins in common foods in the United States and estimation of normal consumption." J Agric Food Chem 54(11): 4069-75.
Yamada, Y., T. Doi, et al. (1998). "Scavenger receptor family proteins: roles for atherosclerosis, host defence and disorders of the central nervous system." Cell Mol Life Sci 54(7): 628-40.
Yin, L., X. Wu, et al. "OX-LDL up-regulates the vascular endothelial growth factor-to-pigment epithelium-derived factor ratio in human retinal pigment epithelial cells." Curr Eye Res 36(4): 379-85.
Youle, R. J. and A. Strasser (2008). "The BCL-2 protein family: opposing activities that mediate cell death." Nat Rev Mol Cell Biol 9(1): 47-59.
Young, J. S. (2008). "Age-related eye diseases: a review of current treatment and recommendations for low-vision aids." Home Healthc Nurse 26(8): 464-71; quiz 472-3.
Zafra-Stone, S., T. Yasmin, et al. (2007). "Berry anthocyanins as novel antioxidants in human health and disease prevention." Mol Nutr Food Res 51(6): 675-83.