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研究生:莊閔捷
研究生(外文):Min-Jay Chuang
論文名稱:黃岑素在大鼠動物模式中對於視網膜缺血/再灌流傷害的保護作用
論文名稱(外文):The Effect of Baicalain on the Retina Subjected to Ischemia/Reperfusion:In Vivo Studies
指導教授:何禮剛何禮剛引用關係趙效明
指導教授(外文):Li-Kang HoHsiao-Ming Chao
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:藥理學研究所
學門:醫藥衛生學門
學類:藥學學類
論文種類:學術論文
論文出版年:2009
畢業學年度:97
語文別:英文
論文頁數:53
中文關鍵詞:視網膜缺血/再灌流黃岑苷元活性氧族基質金屬酵素第九型
外文關鍵詞:retinaischemia/re[erfisionbaicaleinROSMMP-9
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視網膜缺血在青光眼和視網膜血管阻塞的病人中扮演了重要的角色,所以針對視網膜缺血傷害治療的研究就非常重要。視網膜缺血手術可經由提高眼內壓至120毫米汞柱而誘發,再利用網膜電氣生理圖 (electroretinogram,ERG)、免疫組織染色法 (immunohistochemistry,IHC)、即時定量聚合酶反應 (real time PCR)和活性氧族測量 (dichlorofluorescein assay, DCF assay)去評估缺血傷害程度和藥物治療的效果。研究發現當60分鐘暫時性缺血/再灌流後,視網膜的電生理圖ERG b wave ratio顯著降低、代表無軸突細胞 (amacrine cell) 之乙醯膽鹼轉化酵素 (choline acetyltransferase,ChAT) 的免疫染色表現消失、代表視網膜神經節細胞 (retina ganglion cell)表現的Thy-1 mRNA顯著減少。而這些傷害現象可經由預處理給予10 μM和100 μM的baicalein (黃岑苷元)而獲得改善,且在劑量100 μM下的baicalein療效具有統計意義。此外,研究也發現,在缺血/再灌流後第12和24小時,視網膜內的基質金屬酵素第九型mRNA (metalloproteinase-9,MMP-9)有顯著意義的增加,而此現象可以被預處理100 μM的baicalein所抑制。細胞內活性氧族 (reactive oxygen species,ROS)含量的測定發現,預處理100 μM baicalein可以抑制視網膜細胞在氧化壓力下所產生的大量ROS,具有良好的抗氧化效果。綜合上述實驗結果,baicalein可以保護視網膜受到視網膜缺血/再灌流、甚至於青光眼的傷害,可能是藉由抑制缺血再灌流所誘導的MMP-9 mRNA來達成,而baicalein的抑制MMP-9 mRNA的機制則可能和其減少ROS的抗氧化能力有關。
Ischemia plays an important role in glaucoma and retinal vascular occlusion, which renders investigation vital. Retinal ischemia was induced by raising intraocular pressure to 120 mmHg. Its management and mechanism was evaluated by measuring electroretinogram (ERG) b-wave amplitudes, the amounts of reactive oxygen species (ROS), choline acetyltransferase (ChAT) immunoreactivity, and the mRNA levels of matrix metalloproteinase-9 (MMP-9), and Thy-1. Sixty minutes of ischemia plus reperfusion resulted in retinal changes characterized by decreased ERG b-wave amplitudes, loss of choline acetyltransferase (ChAT) immunolabeled amacrine cell bodies/neuronal processes, and down-regulated Thy-1 m-RNA levels (indexing retinal ganglion cells, RGCs). Of clinical importance, this is the first study showing that ischemic detrimental effects are dose-dependently (with the less effect at 10 μM) and significantly (at 100 μM) blunted when baicalein was applied 15 minutes before retinal ischemia. Moreover, the 60-minute ischemia plus 12 or 24 hours of reperfusion induced significant up-regulated MMP-9 mRNA levels. The former effect was further significantly attenuated by pre-administered baicalein (100 μM). The dichlorofluorescein assay results show that pre-treated 100 μM baicalein can significantly attenuate the significant increased levels of ROS in the retina exposed to an oxidative stress. Conclusively, baicalein protects against retinal ischemia and possibly glaucoma by, at least in part, attenuating I/R-induced MMP-9 mRNA up-regulation, possibly related to an oxidative stress.
Abstract
Contents
Figures
Abbreviations
Chapter 1:Introduction
1.1 Retinal ischemia/reperfusion
1.2 Ischemic animal model
1.3 Matrix metalloproteinases (MMPs)
1.4 Baicalein
1.5 Aims
Chapter 2:Material and Method
2.1 Animal
2.2 Anaesthesia and euthanasia of animals
2.3 Induction of retinal ischemia
2.4 Drug administration
2.5 Electroretinogram(ERG)
2.6 Isolation and cryosection of a rat retina
2.7 Immunofluorescence analysis
2.8 Assessment of retinal Thy-1 and MMP-9 mRNA levels
2.9 ROS assay
2.10 Statistics
Chapter 3:Results
3.1 Electroretinogram(ERG)
3.2 Immunofluorescence analysis
3.3 Assessment of retinal Thy-1 mRNA levels
3.5 Assessment of retinal MMP-9 mRNA levels
3.5 Measurement of the ROS levels
Chapter 4:Discussion
Conclusion
Reference

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