臺灣博碩士論文加值系統

青光眼是造成人類致盲之第二大因素,其罹患率逐年上升。其中,急 性高眼壓的傷害因應現代人之生活型態而值得探討。中醫藥古籍中記 載杭菊具有抗發炎、抗氧化、促進血液循環之功效。先前研究顯示杭 菊在乾眼症所引發之角膜損傷與高眼壓引發之視網膜傷害具有緩解作 用,而杭菊對於急性高眼壓所造成的視網膜損傷之緩解功效則是本研 究的重點。本實驗小鼠模式之生理機能與病理分析為主,共分為四組 小鼠:生理對照組、傷害組、低濃度組(100mg/kg)與高濃度組 (500mg/kg),除對照組外皆進境籍性高眼壓傷害;杭菊水萃取物只 餵食低濃度與高濃度組。於實驗第二、九天進行視網膜生理電位測 量;實驗第三、十天進行瞳孔收縮試驗。待實驗結束後犧牲小鼠摘取 眼球進行病理分析。於生理機能上發現:視網膜生理電位之對照組與 高濃度組的電位差皆在0.15mV左右、瞳孔收縮時間皆在2.8秒左右,而 傷害組與低濃度組有則電位差較低、瞳孔收縮時間高於對照組。在病 理分析上,神經節細胞層與視網膜總厚度有隨著餵食濃度增加而增厚 但內核層與外核層無明顯差異。在細胞族群與細胞凋亡部份,視網膜 色素上皮細胞數與神經節細胞數皆隨餵食濃度上升、血管內皮生成因 子、白血球、細胞凋亡訊號隨餵食濃度上升而下降。研究結果顯示餵 食500mg/kg杭菊萃取物能夠有效緩解因急性高眼壓所引發之視網膜傷 害。

Glaucoma is the second leading cause of human blindness, with prevalence roaring up year by year. Among many types of glaucoma, acute ocular hypertension leads to damages to retinal ganglion cells and has attracted much attention due to modern life style tend to over-loading eye . According to ancient Chinese herb medicine, Chrysanthemum morifolium flower is regarded as an effective remedy against glaucomatous injuries. Recent studies have confirmed that it has anti-inflammatory and anti- oxidant activities as well as a circulation-promoting effect. Previous studies also demonstrated its ameliorating effects against dry eye induced corneal injuries and ocular hypertension induced retina damages.This study investigated the underlying mechanisms of using Chrysanthemum morifolium flower extract (CMFE) to mitigate the injuries caused by acute ocular hypertension (AOH) on both physiological and pathological aspects. This study based on mouse model including physiological and pathological aspects. The experimental animals randomly divided into 4 groups as control, damage,low dosage(100mg/kg) and high dosage(500mg/ kg).Electroretinogram was performed in day 2,9 and pupil examination was performed in day 3,10. After finish experiment in day 15, the mice were sacrificed and eyes extracted for pathological analyses. In physiological aspects, electroretinogram shows both of blank and high dose in physiological potential difference are around 0.15mV, pupil response time are around 2.8 seconds while damage and low dose group shows lower potential difference and over longer pupil response time. In pathological aspects, retinal ganglion cell layer(GCL) thickness and total retina thickness result shows dose dependence toward blank group while inner nuclear layer(INL) and outer nuclear layer(ONL) shows no significant difference. In cell population and cell apoptosis analysis result, retinal pigment epithelium(RPE) cell number, retinal ganglion cell (RGC) number shows dose dependence toward blank group and vascular endothelial growth factor , white blood cell number and apoptosis signal shows down when CMFE dosage raise. Base on this result, feeding 500mg/kg CMFE prevents retinal ganglia cell and ischemic damages in an AOH glaucoma mouse model.

中文摘要 i
Abstract ii
壹、緒論 1
1.1眼球結構 3
1.2視網膜結構 5
1.3眼部疾病 8
1.4 青光眼 9
1.4.1青光眼分類與機制 9
1.4.2青光眼之傷害與視力缺損 12
1.4.3青光眼之診斷 13
1.4.4青光眼之治療 17
1.4.5青光眼研究之動物模式 19
1.5青光眼與傳統中醫藥 21
1.5.1傳統中醫藥之青光眼分類 22
1.5.2傳統中醫藥之青光眼成因 24
1.5.3傳統中醫藥之青光眼治療 25
1.6杭菊 26
1.6.1杭菊品種與來源 26
1.6.2杭菊之主要成份 26
1.6.3杭菊與漢醫藥理 28
1.6.4杭菊之古籍用藥 28
1.6.5杭菊之毒性分析 29
1.6.6杭菊與現代保健 29
1.6.7杭菊與眼部疾病 30
貳、研究動機 31
參、材料與方法 34
3.1實驗設計與流程 35
3.2小鼠品系與來源 36
3.3杭菊萃取物之準備與餵食 36
3.3.1 杭菊萃取物之準備 36
3.3.2 杭菊萃取物之配置 36
3.4 小鼠麻醉 37
3.5視網膜生理電位測量 38
3.5.1 視網膜生理電位測量操作環境設定 38
3.5.2 視網膜生理電位測量操作 38
3.5.3 視網膜生理電位測量數據收集 39
3.6瞳孔收縮測試 39
3.6.1瞳孔收縮測試環境設定 39
3.6.2瞳孔收縮測試操作 39
3.6.3瞳孔收縮測試數據收集 40
3.7 小鼠眼球加壓 40
3.7.1眼球加壓設備 40
3.7.2 小鼠眼球加壓操作 40
3.8 小鼠眼球摘取與固定 41
3.9 小鼠眼球包埋與切片 41
3.10 蘇木紫—伊紅染色 42
3.11 免疫組織化學染色 42
3.12 細胞凋亡分析 43
3.13 視網膜結構、病理綜合分析 44
3.13.1視網膜整體厚度與各層細胞厚度綜合分析 44
3.13.2視網膜色素上皮細胞、神經節細胞定量綜合分析 44
3.13.3血管上皮生長因子、白球浸潤分析 44
3.14 統計方法 45
肆、實驗結果 46
4.1杭菊萃取物對視網膜生理電位之效益 47
4.1.1杭菊萃取物對視網膜生理電位圖之生理電位波型圖之效益 47
4.1.2杭菊萃取物對視網膜生理電位之電位差之效益 48
4.2杭菊萃取物對瞳孔收縮之效益 49
4.2.1杭菊萃取物對瞳孔收縮時間之效益 49
4.2.2杭菊萃取物對瞳孔收縮大小之效益 49
4.3杭菊萃取物對視網膜厚度之效益 50
4.3.1杭菊萃取物對視網膜神經節細胞與神經纖維層厚度之效益 50
4.3.2杭菊萃取物對視網膜內核層厚度之效益 51
4.3.3杭菊萃取物對視網膜外核層厚度之效益 51
4.3.4杭菊萃取物對視網膜整體厚度之效益 52
4.4杭菊萃取物對視網膜細胞族群之效益 52
4.4.1杭菊萃取物對視網膜色素上皮細胞之效益 53
4.4.2杭菊萃取物對視網膜神經節細胞之效益 53
4.5杭菊萃取物對視網膜血管新生之效益 54
4.6杭菊萃取物對視網膜白血球浸潤之效益 54
4.7杭菊萃取物對急性高眼壓造成視網膜細胞凋亡之效益 55
伍、討論 56
5.1杭菊萃取物對急性高眼壓之視網膜生理功能之探討 57
5.1.1杭菊萃取物對急性高眼壓之視網膜電位之探討 57
5.1.2杭菊萃取物對急性高眼壓之瞳孔反應之探討 58
5.2杭菊萃取物對急性高眼壓之視網膜結構之探討 59
5.2.1杭菊萃取物對急性高眼壓之視網膜神經節細胞與纖維層結構探討 59
5.2.2杭菊萃取物對急性高眼壓之視網膜內核層結構之探討 60
5.2.3杭菊萃取物對急性高眼壓之視網膜外核層結構之探討 60
5.2.4杭菊萃取物對急性高眼壓之視網膜總厚度之探討 61
5.3杭菊萃取物對急性高眼壓之視網膜細胞族群之探討 62
5.3.1杭菊萃取物對急性高眼壓之視網膜色素上皮細胞族群之探討 62
5.3.2杭菊萃取物對急性高眼壓之視網膜神經節細胞族群之探討 63
5.4杭菊萃取物對急性高眼壓之視網膜發炎型態之探討 64
5.4.1 杭菊萃取物對急性高眼壓之視網膜血管新生之探討 64
5.4.2 杭菊萃取物對急性高眼壓之視網膜白血球浸潤之探討 65
5.5杭菊萃取物對急性高眼壓之視網膜細胞凋亡之探討 66
陸、結論 68
柒、圖次 71
捌、表次 101
玖、參考文獻 108
拾、附錄 115

1.Mainster, M. A., & Turner, P. L. (2010). Ultraviolet-B phototoxicity and hypothetical photomelanomagenesis: intraocular and crystalline lens photoprotection. American journal of ophthalmology, 149(4), 543-549.

2.Riordan-Eva, P., & Whitcher, J. (Eds.). (2008). Vaughan & Asbury''s general ophthalmology. Lange Medical Books/McGraw-Hill Medical Pub. Division.

3.Patel, S., Marshall, J., & Fitzke, F. W. (1995). Refractive index of the human corneal epithelium and stroma. Journal of refractive surgery, 11(2), 100-141.

4.Streilein, J. W., & Cousins, S. W. (1990). Aqueous humor factors and their effect on the immune response in the anterior chamber. Current eye research, 9(sup1), 175-182.

5.Jones, C. E., Atchison, D. A., Meder, R., & Pope, J. M. (2005). Refractive index distribution and optical properties of the isolated human lens measured using magnetic resonance imaging (MRI). Vision Research, 45(18), 2352-2366.

6.Jakobiec, F. A. (Ed.). (1982). Ocular anatomy, embryology, and teratology. Harpercollins.

7.Bergonzo, P., Bongrain, A., Scorsone, E., Bendali, A., Rousseau, L., Lissorgues, G., ... & Yvert, B. (2011). 3D shaped mechanically flexible diamond microelectrode arrays for eye implant applications: The MEDINAS project. Irbm, 32(2), 91-94.

8.Perry, V. H. (1981). Evidence for an amacrine cell system in the ganglion cell layer of the rat retina. Neuroscience, 6(5), 931-944.

9.Kolb H, Fernandez E, Nelson R, editors. SourceWebvision: The Organization of the Retina and Visual System [Internet]. Salt Lake City (UT): University of Utah Health Sciences Center; 1995-2005 May 01 [updated 2007 May 01].

10.Ryan SJ: Retina 4thed,St. Louis: Mosby, 2005.

11..Alfaro DV: Age-Related Macular Degeneration: A comprehensive 
Textbook,2005.

12.Leffler, C. T., Schwartz, S. G., Giliberti, F. M., Young, M. T., & Bermudez, D. (2015). What was Glaucoma Called Before the 20th Century?. Ophthalmology and eye diseases, 7, 21.

13.Stamper, R. L., Lieberman, M. F., Drake, M. V., & Becker, B. (2009). Becker-Shaffer''s diagnosis and therapy of the glaucomas. Elsevier Health Sciences.

14.Crowston, J. (2004). Clinical Guide to Glaucoma Management.

15.Barkan, O. (1938). Glaucoma: Classification, Causes, and Surgical Control*: Results of Microgonioscopic Research. American Journal of Ophthalmology, 21(10), 1099-1117.


16.Sarwat Salim, MD, FACS, and M. Bruce Shields, MD,Iridocorneal Endothelial Syndrome and Glaucoma,https://www.aao.org/eyenet/article/iridocorneal-endothelial-syndrome-glaucoma, 2011

17.McKinnon, S. J. (1997). Glaucoma: A Color Manual of Diagnosis and Treatment. American Journal of Ophthalmology, 123(5), 719.

18.Nanyc B. Carlson and Daniel Kurtz, Clinical Procedure for Ocular Examination,3rd ed,Boston,McGraw-Hill,2004

19.Nanyc B. Carlson and Daniel Kurtz, Clinical Procedure for Ocular Examination,3rd ed,Boston,McGraw-Hill,P61~63,2004

20.Nanyc B. Carlson and Daniel Kurtz, Clinical Procedure for Ocular Examination,3rd ed,Boston,McGraw-Hill,P64~66,2004

21.Quigley, H. A., Dunkelberger, G. R., & Green, W. R. (1989). Retinal ganglion cell atrophy correlated with automated perimetry in human eyes with glaucoma. American journal of ophthalmology, 107(5), 453-464.

22.Nanyc B. Carlson and Daniel Kurtz, Clinical Procedure for Ocular Examination,3rd ed,Boston,McGraw-Hill,P301~303,2004

23.Moseley, M. J. (1995). Non-contact tonometry. Ophthalmic and Physiological Optics, 15, S35-S37.

24.Huang, D., Swanson, E. A., Lin, C. P., Schuman, J. S., Stinson, W. G., Chang, W., ... & Fujimoto, J. G. (1991). Optical coherence tomography. Science (New York, NY), 254(5035), 1178.
25..Nanyc B. Carlson and Daniel Kurtz, Clinical Procedure for Ocular Examination,3rd ed,Boston,McGraw-Hill,P269~270,2004

26.Lai, N. (2004). Clinical ophthalmology: A systematic approach.

27.Nolan, W. P., Foster, P. J., Devereux, J. G., Uranchimeg, D., Johnson, G. J., & Baasanhu, J. (2000). YAG laser iridotomy treatment for primary angle closure in east Asian eyes. British journal of ophthalmology, 84(11), 1255-1259.

28.Beckman, H. (1995). U.S. Patent No. 5,472,440. Washington, DC: U.S. Patent and Trademark Office.

29.A Bouhenni, R., Dunmire, J., Sewell, A., & Edward, D. P. (2012). Animal models of glaucoma. BioMed Research International, 2012.

30.林明育，青光眼看中醫—案例實際探討，1版，台北：商訊文化事業，頁76~84，2008

31.香港浸會大學圖書館，http://libproject.hkbu.edu.hk/was40/detail?channelid=47953&lang=cht&searchword=pid=B00306 ，2017

32.蕭孟衿，行政院農委會茶業改良場，http://www.tres.gov.tw/view.php?catid=1481，201

33.姚莉,靳龍文。2009。藥用菊花的化學成份研究及轉基因進展。J Huaihai Med, Vol. 27, No. 4 : 374-376
34.Lii, C. K., Lei, Y. P., Yao, H. T., Hsieh, Y. S., Tsai, C. W., Liu, K. L., & Chen, H. W. (2010). Chrysanthemum morifolium Ramat. reduces the oxidized LDL-induced expression of intercellular adhesion molecule-1 and E-selectin in human umbilical vein endothelial cells. Journal of ethnopharmacology, 128(1), 213-220.

35.Li, L., Gu, L., Chen, Z., Wang, R., Ye, J., & Jiang, H. (2010). Toxicity study of ethanolic extract of Chrysanthemum morifolium in rats. Journal of food science, 75(6), T105-T109.

36.Mi, X. S., Feng, Q., Lo, A. C. Y., Chang, R. C. C., Lin, B., Chung, S. K., & So, K. F. (2012). Protection of retinal ganglion cells and retinal vasculature by Lycium barbarum polysaccharides in a mouse model of acute ocular hypertension. PLoS One, 7(10), e45469.

37.Li, S. Y., Yang, D., Yeung, C. M., Yu, W. Y., Chang, R. C. C., So, K. F., ... & Lo, A. C. (2011). Lycium barbarum polysaccharides reduce neuronal damage, blood-retinal barrier disruption and oxidative stress in retinal ischemia/reperfusion injury. PloS one, 6(1), e16380.

38.唐于珺（2013）。利用動物實驗探討杭菊萃取物預防紫外線眼表傷害之分析。 中山醫學大學醫學檢驗暨生物技術學系碩士論文。

39.陳明輝（2014）。 杭菊水萃取物預防小鼠急性高眼壓青光眼模式導致視網膜損傷之功效分析。中山醫學大學生化暨生物科技研究所學位論文。

40.Nanyc B. Carlson and Daniel Kurtz, Clinical Procedure for Ocular Examination,3rd ed,Boston,McGraw-Hill,P59~60,2004

41.Hess, E. H., & Polt, J. M. (1960). Pupil size as related to interest value of visual stimuli. Science, 132(3423), 349-350.

42.McKinnon, S. J., Lehman, D. M., Kerrigan-Baumrind, L. A., Merges, C. A., Pease, M. E., Kerrigan, D. F., ... & Quigley, H. A. (2002). Caspase activation and amyloid precursor protein cleavage in rat ocular hypertension. Investigative ophthalmology & visual science, 43(4), 1077-1087.

43.Cunha-Vaz, J., Bernardes, R., & Lobo, C. (2010). Blood-retinal barrier. European journal of ophthalmology, 21, S3-9.