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論文名稱(中文) 抽菸,喝酒,砷暴露,第I、II相代謝酵素的基因多形性與泌尿道上皮癌之相關性研究
論文名稱(英文) Cigarette smoking, alcohol consumption, arsenic exposure, genetic polymorphisms of phase I、II metabolized-enzymes and urothelial carcinoma
校院名稱 臺北醫學大學
系所名稱(中) 公共衛生學研究所
系所名稱(英) Graduate Institute of Public Health
學年度 97
學期 1
出版年 98
研究生(中文) 王淵宏
學號 D508091002
學位類別 博士
語文別 中文
口試日期 2008-11-13
論文頁數 108頁
口試委員 委員-陳建仁
委員-江漢聲
委員-李德章
委員-蒲永孝
委員-陳彥州
委員-葉劭德
指導教授-邱弘毅
關鍵字(中) 抽菸
喝酒
砷暴露
第I、II相代謝酵素
基因多形性
泌尿道上皮癌
關鍵字(英) Cigarette smoking
Alcohol consumption
Arsenic exposure
Phase I、II metabolized-enzymes
Genetic polymorphism
Urothelial carcinoma
學科別分類
中文摘要 研究背景:抽菸、無機砷及危險職業暴露是泌尿道上皮癌已知的危險因子。近來有研究指出喝酒可能與膀胱癌有關,但可能的機制尚未被詳細探討。人體接觸到環境致癌物質之後,通常先由第I相代謝酵素[例如細胞色素P450酵素, cytochrome P-450 (CYPs)]將有毒化學物質經代謝活化後形成親電子性物質,再由第II相代謝酵素[例如麩胺基硫轉移酵素, Glutathione S-transferases (GSTs)或sulfotransferase (SULT)]將活性中間產物代謝成毒性較低的親水性物質排出體外。若體內的第I、II相代謝酵素的功能失去平衡,長期累積之後會增加罹患癌症的風險。另外,與酒精代謝相關的酵素包括CYP2E1、乙醇去氫酵素(alcohol dehydrohenase, ADH)及乙醛去氫酵素(aldehyde dehydrogenase, ALDH),假若這些酒精代謝相關的酵素產生變異,可能會增加罹患癌症的風險。因此,本論文將抽菸,喝酒,砷暴露及第I、II相代謝酵素基因多形性與泌尿道上皮癌進行合併分析,藉以探討基因與基因及基因與環境危險因子間的交互作用對於罹患泌尿道上皮癌的影響◦
材料與方法:本論文選取540位經病理診斷確定的泌尿道上皮癌(包括腎盂、輸尿管及膀胱部位的癌症)患者作為病例組個案,另外選取540位與病例組個案的年齡頻率匹配且未曾罹患任何癌症的患者作為對照組個案。基因多形性主要是以聚合酵素連鎖反應-限制片段長度多形性方法來判定。第I相代謝酵素包括CYP1A1、CYP2A6及CYP2E1,第II相代謝酵素包括GSTO1、GSTO2及SULT1A1,與酒精代謝相關的酵素包括ADH2、ADH3及ALDH2。利用goodness-of-fit卡方檢定進行哈溫平衡(Hardy-Weinberg Equilibrium)檢定。以非條件式多變項邏輯斯迴歸估計危險對比值(odds ratios, ORs)及95%信賴區間(confidence intervals, CIs)。而基因的單套型(haplotype)是採用Haploview 3.2分析軟體(http://sourceforge.net/projects/ haploview)來進行分析。
研究結果:本研究發現抽菸、無機砷及職業暴露與泌尿上皮癌有顯著相關。同時具有抽菸及喝酒習慣者有較高的危險性(OR=3.0)及顯著的協同作用,而經常使用消炎止痛藥者有2倍危險性。在各別的基因多形性分析中,帶有CYP1A1 A4889G的G/G基因型有顯著較高的危險性(OR=1.9)。CYP1A1危險雙套型有1.8倍顯著的危險性。帶有3個或以上第I相代謝酵素的危險基因型/雙套型者有2.5倍的顯著危險性。同時帶有抽菸/喝酒及職業暴露(≥1)、無機砷暴露(high)及危險基因型(≥3)者有7.7倍顯著的危險性。另外,帶有GSTO2 A424G-G/G、GSTO2 A-183G -G/G及SULT1A1 G638A-G/G基因型有顯著較高的危險性。GSTO1/2的危險雙套型也有1.8倍顯著的危險性。帶1個或以上第II相代謝酵素的危險基因型/雙套型者有2.3倍的顯著危險性。同時帶有抽菸/喝酒及職業暴露(≥1)、無機砷暴露(high)及危險基因型(≥1)者有6.8倍顯著的危險性。將抽菸/喝酒、無機砷暴露及職業暴露合併分析後,結果發現帶有其中1個及帶有其中2個或以上環境暴露者分別有1.5及2.6倍顯著的危險性。最後,在帶有其中2個或以上環境危險因子暴露之下,同時帶有3個或以上第I相代謝酵素的危險基因型/雙套型及1個或以上第II相代謝酵素的危險基因型/雙套型者具有最顯著的危險性 (OR=19.4, 95% CI =4.5-83.1)。
結論:從本論文的研究結果綜合來看,環境危險因子的暴露及第I、II相代謝酵素的危險基因型/雙套型分別對於罹患泌尿道上皮癌有顯著的影響之外,尤其是當環境與基因之間具有協同作用時,影響的程度會更加明顯。因此,未來需要更大的樣本及納入其他具功能性的基因多形性一併分析,以期對泌尿道上皮癌的致病機轉有更完整的瞭解。

英文摘要 Background:Cigarette smoking and exposures of arsenic and risk-occupations are known risk factors for urothelial carcinoma (UC). Recent studies indicated that alcohol consumption is likely to be associated with bladder cancer, but the possible mechanism has not been well investigated. After human exposed to environmental carcinogens, most of these substances are firstly converted into reactive carcinogenic metabolites by phase I enzymes including Cytochrome P-450 (CYPs) and these reactive components were removed by phase II enzymes including Glutathione S- transferases (GSTs) or Sulfotransferase (SULT). The lone-term imbalance of enzyme activity between phase I and phase II enzymes will lead to the development of cancer. Besides, CYP2E1、alcohol dehydrohenase(ADH) and aldehyde dehydrogenase (ALDH) are known alcohol-related enzymes and higher frequencies of variations of these enzymes are also likely to lead to the development of cancer. Therefore, to investigate the joint effects of gene-gene and gene-environment interactions on risk of UC, this study included environmental exposures of cigarette smoking, alcohol consumption, arsenic and risk-occupations and genetic polymorphisms of phase I 、II enzymes in combined analyses.
Materials and Methods:A total of 540 pathologically-confirmed UC cases including cancers of renal pelvis, ureter and bladder were selected as UC cases group. A total of 540 cancer-free controls, frequency-matched on age, were recruited from individuals who admitted to the same hospitals with UC cases for a health examination. Genetic polymorphisms of were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Selected enzymes including phase I enzymes- CYP1A1、CYP2A6 and CYP2E1, phase II enzymes-GSTO1、GSTO2 and SULT1A1, and alcohol-metabolized enzymes-ADH2、ADH3 and ALDH2. A goodness-of-fit X2 test was performed to examine Hardy-Weinberg Equilibrium (HWE). We used a unconditional multivariate logistic regression to estimate odds ratios (ORs) and 95% confidence intervals (CIs). Haplotype analysis was calculated by Haploview 3.2 (http://sourceforge.net/projects/ haploview).
Results:Cigarette smoking, arsenic exposure and risk occupational exposure are significantly associated with UC risk. Study subjects with both of cigarette smoking and alcohol consumption have a significantly higher UC risk of 3.0. Individuals with frequent NSAID usage have a significantly higher UC risk of 2.0. In genotyping analysis, subjects with CYP1A1 A4889G G/G genotype have a significantly higher UC risk (OR=1.9). Those carrying risk diplotypes of CYP1A1 have a significantly higher UC risk of 1.8. Subjects who carried three or more risk genotypes/diplotypes of phase I enzymes have a significantly increased UC risk of 2.5. Individuals with cigarette smoking/alcohol consumption and occupational exposures (≥1), arsenic exposure (high), and risk genotypes/diplotypes of phase I enzymes (≥3) have a significantly increased UC risk (OR=7.7). Significantly increased UC risks of 1.6, 2.5 and 1.8 were found for subjects carrying GSTO2 A424G-G/G, GSTO2 A-183G -G/G and SULT1A1 G638A-G/G genotypes, respectively. Individuals with risk diplotypes of GSTO1/2 also have a significantly higher UC risk of 1.8. Those carrying one or more risk genotypes/diplotypes of phase II enzymes have a significantly increased UC risk (OR=2.3). Individuals who carried cigarette smoking/alcohol consumption and occupational exposures (≥1), arsenic exposure (high), and risk genotypes/diplotypes of phase II enzymes (≥1) have a significantly increased UC risk (OR=6.8). In a combination analysis of cigarette smoking/alcohol consumption, arsenic exposure and occupational exposure, significantly higher UC risks of 1.5 and 2.6 were found for subjects with one risk factor and those with two or more risk factors, respectively. Finally, subjects who exposed to two or more of environmental risk factors carrying three or more risk genotypes/diplotypes of phase I enzymes and one or more risk genotypes/diplotypes of phase II enzymes have the significantly highest risk of UC (OR=19.4, 95% CI =4.5-83.1).
Conclusion:In addition to significant effects from exposures of environmental risk factors and risk genotypes/diplotypes of phase I、II enzymes on UC risk, the effects on the development of UC will be more predominant especially under the existence of gene-environment interaction. Therefore, a larger sample size and other functional polymorphisms of candidate genes should be took into consideration to provide a more comprehensive understanding of UC.
論文目次 目 錄
中文摘要---------------------------------------------I-III
英文摘要--------------------------------------------IV-VII
第一章 前言-----------------------------------------1-3
第二章 文獻探討
1.膀胱癌的描述性流行病學----------------------------4-7
2.膀胱癌的危險因子----------------------------------7-15
3.膀胱癌與易感性基因的多形性------------------------15-19
第三章 抽菸,喝酒,砷暴露及第I、II相代謝酵素基因多形性與
泌尿道上皮癌之相關性研究
第一節 前言-----------------------------------------20-26
第二節 材料與方法-----------------------------------27-34
第三節 結果-----------------------------------------35-73
第四節 討論-----------------------------------------74-81
第四章 結論-----------------------------------------82-83
參考文獻--------------------------------------------84-101
附件一----------------------------------------------102-108

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系統識別號 U0007-0402201015321200
論文名稱(中文) 白朮炮製之研究
論文名稱(英文) The Studies of the Processed Atractylodes Rhizoma
校院名稱 臺北醫學大學
系所名稱(中) 藥學系(博士班)
系所名稱(英)
學年度 98
學期 1
出版年 99
研究生(中文) 王坤謄
學號 D301093008
學位類別 博士
語文別 中文
口試日期 2010-01-12
論文頁數 151頁
口試委員 委員-黃怡超
委員-陳福安
委員-許明照
委員-蔡麗雪
指導教授-王靜瓊
關鍵字(中) 白朮
炮製
atractylon
atractylenolide III
酒精引發胃黏膜損傷
自由基清除
MMPs
關鍵字(英) Atractylodes Rhizoma
processing
atractylon
atractylenolide III
free radical scavenging
MMPs
學科別分類
中文摘要 白朮為經常用於補脾健胃之中藥,且根據傳統醫書記載利用不同輔料進行加工炮製後可減去燥性與增強補脾益氣效用。但炮製後的品質至今仍無標準化規範。因此本研究將針對白朮炮製前後成分及藥理變化製訂出白朮之標準規範。研究方法共分為三個部分: 第一部分先建立白朮指標成分快速分離流程,結果顯示: 利用n-hexane冷浸萃取法再經矽膠管柱層析,可快速分離得到atractylon、atractylenolides I、II、III與biatractylolide等這五個指標成分。第二部分收集白朮市售品,分析其指標成分的含量,並利用自行炮製之白朮樣品建立標準白朮炮製流程,以了解白朮炮製後化學成分之變化。結果顯示: 台灣所販賣的白朮市售品中以紅土及灶心土炒製品比例較高(32/38),而其中指標成分含量相差甚多,差異將近50%。此外,市售白朮生品中atractylon含量較高,而炮製品則是以atractylenolide III含量較高。利用恆溫炫風式滾筒炒鍋進行藥材炮製,結果顯示atractylon隨加熱時間延長而含量下降,atractylenolides II及III則隨加熱時間延長而含量上升。在重金屬檢測中,白朮經輔料炮製後其鉛及銅含量隨之上升。第三部分則利用體內外胃黏膜損傷試驗與抗氧化試驗來評估白朮炮製前後藥理活性之改變,結果顯示: 白朮炮製前後樣品皆無降低胃酸總酸度之功效,而在體外與體內的實驗中皆能顯著的抑制酒精所引發之胃黏膜損傷,但組間無差異。Atractylenolide III為白朮中主要活性成分,其調控機制為促進TIMP-1及TIMP-2蛋白質表現,進而抑制MMP-2及MMP-9酵素活性及蛋白質表現,減緩酒精性胃潰瘍之發生。抗氧化功效上,白朮生品精油具有顯著的DPPH自由基清除、抑制脂質過氧化與提升肝臟過氧化氫酵素功效,但炮製後其抗氧化效用隨之下降,而白朮生品與炮製品精油對H9C2, 3T3及WI-38細胞株皆不具有顯著細胞毒性。綜合以上結果,本研究建議白朮使用於胃黏膜保護功效時以清炒較佳,因白朮清炒後其胃黏膜保護功效無顯著下降,但可降低生品中水分含量,更易於保存。此外,白朮樣品內atractylenolide III含量在1.08-1.84 mg/g之間,即有顯著的胃黏膜保護功效。
英文摘要 Atractylodes Rhizoma, the dried rhizome of Atractylodes ovata De Candolle, is widely used for strengthening the stomach functions and usually underwent the processing procedure before usage. Nowadays, the standard operation procedure (SOP) for using Atractylodes Rhizoma is not well-established. Hence, the SOP of Atractylodes Rhizoma was set up according to the variations of chemical contents and pharmacological effects after processing. The study was divied into 3 parts. Firstly, atractylon, atractylenolides I, II, III and biatractylolide were isolated through silica gel chromatography. Secondly, the commercial Atractylodes Rhizoma was collected for analyzing the contents of atractylon, atractylenolides II and III. Results showed that almost all commercial Atractylodes Rhizoma was processed samples (32/38), including stir-frying with red soil or burnt clay. Besides, contents of these 3 pure components in commercial Atractylodes Rhizoma varied a lot (almost reaching 50%). Contents of atractylon were higher in raw materials, but contents of atractylenolides were higher in processed samples. In addition, following the prolonged processing time, contents of atractylon were decreased, but atractylenolides II and III increased. In the heavy metal analysis, contents of Pb and Cu were increased following stir-frying with assistant substrates. Thirdly, we used in vitro and in vivo gastroprotective assays and anti-oxidative effects to evaluate the pharmacological variations after processing. Results showed that raw and processed Atractylodes Rhizoma did not significantly decrease the total acid of gastric juice. However, raw and processed Atractylodes Rhizoma significantly protected the ethanol-induced gastric mucosal damage in vitro and in vivo, but no significant changes between each group were found. Atractylenolide III was the principal gastroprotective component of Atractylodes Rhizoma and the possible mechanism was through regulating the expressions of MMP-2, MMP-9, TIMP-1 and TIMP-2. In the anti-oxidative effects, raw materials displayed significant DPPH-scavenging, anti-lipid peroxidative and enhanced catalase activities effects. However, the anti-oxidative effects of raw materials were reduced after processing. Both raw and processed Atractylodes Rhizoma essential oils did not show the cytotoxicities in H9C2, 3T3 and WI-38 cell lines, Taken together, we suggested that Atractylodes Rhizoma which was stir-fried without assistant substrates was better for the gastroproective usages. In addition, the atractylenolide III contents in Atractylodes Rhizoma should range from 1.08 to 1.84 mg/g.
論文目次 目錄…………………………………………………………………………………………………………………………1
圖目錄………………………………………………………………………………………………………………………9
表目錄……………………………………………………………………………………………………………………11
中文摘要…………………………………………………………………………………………………………………12
英文摘要…………………………………………………………………………………………………………………14
縮寫表………………………………………………………………………………………………………………………16
緒論……………………………………………………………………………………………………………………………18
白朮之文獻回顧………………………………………………………………………………………………………23
第一部分 白朮指標成分分離 36
壹、前言…………………………………………………………………………………………………………………37
貳、實驗材料與方法………………………………………………………………………………………………41
1. 白朮指標成分分離標準流程制定…………………………………………………………………41
1-1. 實驗材料………………………………………………………………………………………………………41
1-2. 化學試劑………………………………………………………………………………………………………41
1-3. 實驗儀器…………………………………………………………………………………………………………41
1-4. 分離流程…………………………………………………………………………………………………………42
叁、實驗結果………………………………………………………………………………………………………………44
1. Atractylon之圖譜解析………………………………………………………………………………44
2. Atractylenolide I之圖譜解析………………………………………………………………45
3. Atractylenolide II之圖譜解析……………………………………………………………46
4. Atractylenolide III之圖譜解析……………………………………………………………47
5. Biatractylolide之圖譜解析……………………………………………………………………48
肆、討論…………………………………………………………………………………………………………………………49
第二部分 白朮炮製前後成分變化 51
壹、前言…………………………………………………………………………………………………………………………52
貳、實驗材料與方法……………………………………………………………………………………………………55
1. 白朮市售品之收集.………………………………………………………………………………………………55
2. 白朮標準炮製流程建立…………………………………………………………………………………………55
2.1 炮製所需之材料……………………………………………………………………………………………………55
2-1.1. 中藥材白朮………………………………………………………………………………………………………55
2-1.2. 炮製之輔料………………………………………………………………………………………………………55
2-1.3. 炮製之設備………………………………………………………………………………………………………55
2-2. 炮製之方法……………………………………………………………………………………………………………55
3. 白朮炮製樣品之成分變化………………………………………………………………………………………56
3-1. 化學試劑………………………………………………………………………………………………………………56
3-2. 實驗儀器…………………………………………………………………………………………………………………56
3-3. 炮製炮製樣品之品質管制……………………………………………………………………………………57
3-3.1. 白朮炮製樣品外觀顏色分析…………………………………………………………………………57
3-3.1.1. 分析原理…………………………………………………………………………………………………………57
3-3.1.2. 分析方法…………………………………………………………………………………………………………57
3-3.2. 白朮炮製樣品水分含量分析…………………………………………………………………………58
3-3.2.1. 樣品製備…………………………………………………………………………………………………………58
3-3.2.2. 分析方法………………………………………………………………………………………………………58
3-3.3. 白朮炮製樣品重金屬含量分析………………………………………………………………………58
3-3.3.1. 樣品製備…………………………………………………………………………………………………………58
3-3.3.2. 分析方法…………………………………………………………………………………………………………58
3-3.4. 白朮炮製樣品指標成分分析……………………………………………………………………………58
3-3.4.1. 樣品製備…………………………………………………………………………………………………………58
3-3.4.2. 高效液相層析儀分析……………………………………………………………………………………59
3-3.4.3. 標準檢量線製備……………………………………………………………………………………………59
3-3.4.4. 分析方法之確效………………………………………………………………………………………………60
3-3.5. 白朮炮製樣品水萃取物指標成分含量分析……………………………………………………61
3-3.5.1 樣品製備………………………………………………………………………………………………………………61
3-3.5.2. 標準檢量線製備…………………………………………………………………………………………61
3-3.6. Atractylon於受熱後之變化………………………………………………………………………61
3-3.6.1. 樣品製備……………………………………………………………………………………………………………61
3-3.6.2. 標準檢量線製備………………………………………………………………………………………………61
3-3.7. 白朮炮製樣品精油含量變化………………………………………………………………………………61
3-3.7.1. 精油萃取……………………………………………………………………………………………………………61
3-3.7.2. 高效液相層析儀分析………………………………………………………………………………………62
3-3.7.3. 氣相層析質譜儀分析………………………………………………………………………………………63
叁、實驗結果……………………………………………………………………………………………………………………………64
1. 白朮市售品種類整理…………………………………………………………………………………………………64
2. 白朮樣品炮製結果………………………………………………………………………………………………………65
3. 白朮炮製樣品之品質管制……………………………………………………………………………………………65
3-1. 白朮炮製前後樣品之顏色………………………………………………………………………………………65
3-2. 白朮炮製前後樣品之水分含量……………………………………………………………………………66
3-3. 白朮炮製前後樣品之重金屬含量………………………………………………………………………67
3-4. 白朮市售品及炮製樣品指標成分之分析…………………………………………………………67
3-4.1. 檢量線製備…………………………………………………………………………………………………………67
3-4.2. 分析方法之確效性……………………………………………………………………………………………68
3-4.3. 市售品指標成份含量………………………………………………………………………………………71
3-4.4. 白朮炮製前後樣品指標成分之含量………………………………………………………………73
3-4.5. 白朮炮製前後樣品之水萃取物中指標成分含量分析…………………………………74
3-4.6. 白朮炮製前後樣品之精油中指標成分含量 分析……………………………………74
3-4.7. Atractylon受熱後氧化之情形……………………………………………………………………75
3-4.8. 白朮炮製前後樣品精油組成之變化…………………………………………………………………76
肆、討論……………………………………………………………………………………………………………………………80
第三部分 白朮炮製前後藥理活性變化 84
壹、前言………………………………………………………………………………………………………………………………85
貳、實驗材料與方法…………………………………………………………………………………………………………89
1. 胃酸總酸度試驗…………………………………………………………………………………………………………89
1-1. 實驗動物…………………………………………………………………………………………………………………89
1-2. 實驗樣品…………………………………………………………………………………………………………………89
1-3. 實驗材料與儀器……………………………………………………………………………………………………89
1-4. 實驗方法………………………………………………………………………………………………………………89
2. 胃黏膜損傷保護試驗……………………………………………………………………………………………90
2-1. 胃黏膜細胞試驗……………………………………………………………………………………………………90
2-1.1. 胃黏膜細胞分離及培養………………………………………………………………………………90
2-1.1.1. 實驗動物…………………………………………………………………………………………………90
2-1.1.2. 胃黏膜細胞分離與培養所需試劑………………………………………………………90
2-1.1.3. 胃黏膜細胞分離流程……………………………………………………………………………90
2-1.1.4. 胃黏膜細胞鑑定流程……………………………………………………………………………91
2-1.2. 白朮炮製前後樣品與指標成分對胃黏膜細胞之毒性評估…………………91
2-1.2.1. 實驗樣品……………………………………………………………………………………………………91
2-1.2.2. 化學試劑…………………………………………………………………………………………………92
2-1.2.3. 實驗儀器…………………………………………………………………………………………………92
2-1.2.4. 實驗方法……………………………………………………………………………………………………93
2-1.3. 酒精濃度對胃黏膜細胞之影響……………………………………………………………………93
2-1.4. 白朮炮製前後樣品與指標成分保護酒精所引發胃黏膜細胞毒性之作用…………………………………………………………………………………………………………………………………………93
2-1.5. 白朮炮製前後樣品與指標成份保護酒精誘導胃黏膜細胞膜損傷之作用…………………………………………………………………………………………………………………………………………93
2-1.5.1. 實驗原理…………………………………………………………………………………………………94
2-1.5.2. 實驗方法…………………………………………………………………………………………………94
2-2. 大鼠急性胃潰瘍試驗………………………………………………………………………………………94
2-2.1. 實驗樣品…………………………………………………………………………………………………………94
2-2.2. 實驗動物…………………………………………………………………………………………………………94
2-2.3. 化學試劑…………………………………………………………………………………………………………94
2-2.4. 實驗儀器………………………………………………………………………………………………………95
2-2.5. 實驗所需之抗體……………………………………………………………………………………………95
2-2.6. 實驗方法………………………………………………………………………………………………………95
2-2.6.1. 酒精濃度誘導大鼠急性胃潰瘍之試驗………………………………………………95
2-2.6.2. 白朮炮製前後樣品與指標成分保護酒精誘導大鼠急性胃潰瘍之試驗……………………………………………………………………………………………………………………………………96
2-2.6.3. 胃潰瘍之病理切片…………………………………………………………………………………96
2-2.6.4. 受質凝膠電泳法………………………………………………………………………………………96
2-2.6.5. 西方點墨法…………………………………………………………………………………………………97
3. 抗氧化能力試驗…………………………………………………………………………………………………………98
3-1. DPPH自由基清除試驗…………………………………………………………………………………………98
3-1.1. 實驗樣品……………………………………………………………………………………………………………98
3-1.2. 化學試劑……………………………………………………………………………………………………………98
3-1.3. 實驗儀器……………………………………………………………………………………………………………98
3-1.4. 實驗方法……………………………………………………………………………………………………………98
3-2. 電子自旋共振法之自由基偵測法……………………………………………………………………99
3-2.1. 實驗樣品……………………………………………………………………………………………………………99
3-2.2. 化學試劑……………………………………………………………………………………………………………99
3-2.3. 實驗儀器……………………………………………………………………………………………………………99
3-2.4. 實驗方法……………………………………………………………………………………………………………99
3-3. 脂質過氧化之試驗………………………………………………………………………………………………100
3-3.1. 實驗樣品……………………………………………………………………………………………………………100
3-3.2. 實驗動物……………………………………………………………………………………………………………100
3-3.3. 化學試劑……………………………………………………………………………………………………………100
3-3.4. 實驗儀器……………………………………………………………………………………………………………100
3-3.5. 實驗方法……………………………………………………………………………………………………………100
3-4. 過氧化氫酵素試驗………………………………………………………………………………………………100
3-4.1. 實驗樣品……………………………………………………………………………………………………………101
3-4.2. 化學試劑……………………………………………………………………………………………………………101
3-4.3. 實驗儀器……………………………………………………………………………………………………………101
3-4.4. 實驗步驟……………………………………………………………………………………………………………101
4. 細胞毒性試驗………………………………………………………………………………………………………………101
4-1. 實驗樣品…………………………………………………………………………………………………………………101
4-2. 化學試劑…………………………………………………………………………………………………………………101
4-3. 實驗儀器…………………………………………………………………………………………………………………101
4-4. 實驗方法………………………………………………………………………………………………………………102
叁、實驗結果………………………………………………………………………………………………………………………103
1. 白朮炮製前後樣品水萃取物對於胃酸總酸度之影響………………………………………103
2.白朮炮製前後樣品與指標成分保護酒精誘導胃黏膜細胞毒性之作用……………103
2-1. 胃黏膜細胞之分離及鑑定…………………………………………………………………………………103
2-2. 白朮炮製前後樣品與指標成分對大鼠胃黏膜細胞毒性評估……………………104
2-3. 建立酒精誘導大鼠胃黏膜細胞毒性模式………………………………………………………104
2-4. 白朮炮製前後樣品保護酒精誘導大鼠胃黏膜細胞毒性之作用………………105
2-5. 白朮指標成分保護酒精誘導大鼠胃黏膜細胞毒性之作用………………………106
3. 白朮炮製前後樣品與指標成分保護酒精誘導大鼠急性胃潰瘍之作用…………107
3-1. 酒精引發大鼠急性胃潰瘍模式建立…………………………………………………………………108
3-2.白朮炮製前後樣品水萃取物與指標成分保護酒精誘導大鼠急性胃潰瘍之作用……………………………………………………………………………………………………………………………………………108
3-3. 大鼠急性胃潰瘍之病理分析………………………………………………………………………………109
3-4. 潰瘍部位之MMP-2及MMP-9活性分析………………………………………………………………109
4. 白朮炮製前後樣品抗氧化作用…………………………………………………………………………………116
4-1. 白朮炮製前後樣品清除DPPH自由基之作用……………………………………………………116
4-2. ESR偵測白朮炮製前後樣品精油清除自由基之作用………………………………………117
4-3. 白朮炮製前後樣品精油抑制脂質過氧化之作用……………………………………………118
4-4. 白朮炮製前後樣品精油增加過氧化氫酵素活性之作用………………………………119
5. 白朮炮製前後樣品與指標成分對於正常細胞之細胞毒性…………………………………120
肆、討論……………………………………………………………………………………………………………………………122
總結……………………………………………………………………………………………………………………………………127
參考文獻………………………………………………………………………………………………………………………………127
致謝………………………………………………………………………………………………………………………………………140
論文發表………………………………………………………………………………………………………………………………141
附錄一…………………………………………………………………………………………………………………………………142
附錄二…………………………………………………………………………………………………………………………………144
附錄三…………………………………………………………………………………………………………………………………146
附錄四…………………………………………………………………………………………………………………………………148
附錄五…………………………………………………………………………………………………………………………………150
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系統識別號 U0007-0407200710570200
論文名稱(中文) 以家庭參與介入於第二型糖尿病控制不佳患者之成效探討
論文名稱(英文) The Effectiveness of Family Partnership Intervention in Unstable Control Patients with Type 2 Diabetes
校院名稱 臺北醫學大學
系所名稱(中) 護理學研究所
系所名稱(英) Graduate Institute of Nursing
學年度 95
學期 2
出版年 96
研究生(中文) 康春梅
學號 G455092013
學位類別 碩士
語文別 英文
口試日期 2007-06-21
論文頁數 95頁
口試委員 指導教授-張文英
委員-陳品玲
委員-章淑娟
關鍵字(中) 家庭參與
第二型糖尿病控制不佳患者
成效
關鍵字(英) Family participation
Type II diabetes
effectiveness
學科別分類
中文摘要 糖尿病是高死亡率、高醫療耗費之疾病,除例行之篩檢及治療外,是否家屬的參與能獲得較佳之控制結果呢?故本研究目的旨在比較家庭參與組與一般照護組於第二型糖尿病控制不佳之患者,其糖尿病控制指標、家庭支持行為、疾病知識、態度及自我照顧行為之差異。
本研究採前-後測設計,以診斷為第二型糖尿病且糖化血色素(A1C)平均>7%者為研究對象。病患入選後依診斷時間順序,分為家庭參與組及一般照護組。資料收集後以描述性、卡方檢定、獨立樣本t 檢定、曼-惠特尼U考驗及魏氏帶檢定進行分析。
本研究共收集56人;家庭參與組28人,一般照護組28人。家庭參與組病患平均年齡為55.3歲,以男性居多(n=16;57.1 %),教育程度以高中(職)居多(n=17;60.7%),全數(100%)與家人同住,17位(60.7 %)有糖尿病家族史。而一般照護組病患平均年齡為51.7歲,男女各為14人(50 %),教育程度以高中(職)以上居多(n=15;53.5%),27人(96.4%)與家人同住,21位(75.0 %)有糖尿病家族史,以上變項,兩組皆無顯著差異(p > .05)。
在介入六個月後之糖尿病控制指標中,家庭參與組之糖化血色素(A1C)、空腹血糖(FBS)、總膽固醇(TC)、三酸甘油脂(TG)、低密度脂蛋白膽固醇(LDL-C)的改善程度均較一般照護組佳,但在統計上均未達顯著差異(p > .05)。
而家庭參與組在家庭支持行為平均得分由52.14分提高為56.46分,疾病知識由平均10.86提高為16.18分,疾病態度由平均56.39分提高為59.11分,自我照顧行為平均88.14分提高為101.89分。而一般照護組之家庭支持行為得分由平均51.21分下降為49.82分,疾病知識由平均12.32分提高為14.64分,疾病態度由平均56.43分稍微提高至56.57分,自我照顧行為由平均為87.64分提高為96.68分;兩組前三項變項達統計上之顯著差異(p < .05);但自我照顧行為則未達統計上之顯著差異(p > .05)。
本研究發現家庭參與方案對糖尿病病患之糖尿病控制指標、家庭支持行為、疾病知識、態度、自我照顧行為確有改善之效果,故此結果除可提供組織決策者,更瞭解家庭參與介入糖尿病控制之成效外,亦可作為其他慢性病衛教之範例,提升其自我照護成效。
英文摘要 Diabetic Mellitus is a high mortality and costly disease. Besides routine examinations and treatments, whether family participation can have better outcomes than those who do not? Therefore, the aims of this study were to compare the differences in the diabetic metabolic control, family supportive behaviors, patients’ knowledge, attitudes, and self-care behaviors between family partnership intervention care (FPI) group and usual care (UC) group for patients with type II diabetes.
This study was pre-posttest design. Samples included in this study were who had diagnosed as type II diabetes and A1C was great than 7%. After screening, samples were randomly assigned to FPI group or UC group based on the time they were diagnosed. Descriptive statistics, independent t-test, χ2, Mann-Whitney U test and Wilcoxon signed ranks test were used to analyse the data.
There were 56 patients participated in this study, 28 patients in FPI group and 28 in UC group. The mean age of the FPI group was 53.3 years. Of these, 16 (57.1%) were males, 17 (60.7%) had senior high school education, 27 (96.4%) were married, and 28 (100%) were living with their family, and 17 (60.7%) had a history of diabetes. In contrast, the mean age of the UC group was 51.7 years. Of these, 14 were males (50%), 15 (53.5%) had senior high school education, 24 (85.7%) were married, and 27 (96.4%) were living with family, and 21 (75.0%) had a history of diabetes. However, no significant differences were found between groups (p > .05).
After 6 months follow-ups, physical measures such as A1C, BMI, FBS, TC, TG, LDC-C, and HDC-C were better improved than those of patients in the UC group. However, no significant differences were found between groups on above variables (p > .05).
For the FPI group, the mean scores of family supportive behaviors were increased 4.32 points, patients’ knowledge was increased 5.32 points, patients’ attitudes was increased 2.71 points, and patients’ self-care behaviors was increased 2.71 points. For the UC group, the mean scores of family supportive behaviors was decreased 1.39 points, patients’ knowledge was increased 2.32 points, patients’ attitudes was increased 0.14 points, and patients’ self-care behaviors was increased 9.04 points. There was a significant difference in the mean scores of family supportive behaviors, patients’ knowledge, and patients’ attitudes between groups (p< .05); however, no significant difference was found on patients’ self-care behaviors between groups (p > .05).
The findings demonstrate that the DM control was improved for FPI group than that of UC group. The scores of family supportive behaviors, patients’ knowledge, patients’ attitudes, and patients’ self-care behaviors were also improved in FPI group than that of UC group. Thus, the results not only provide information to decision makers to better understanding the effectiveness of DM family participation, but also can be used for other chronic disease to improve the outcomes of self-care.
論文目次 目 錄
頁數
致 謝 ………………………………………………………… Ⅰ
中文摘要 ………………………………………………………… Ⅱ
英文摘要 ………………………………………………………… Ⅳ
目 錄 ………………………………………………………… Ⅵ
圖表目次 ………………………………………………………… Ⅸ
第一章 緒論
第一節 研究背景及動機………………………………………… 1
第二節 研究目的………………………………………………… 4
第三節 研究假設………………………………………………… 5
第四節 名詞界定………………………………………………… 6
第二章 文獻查證
第一節 第二型糖尿病簡介……………………………………… 10
第二節 糖尿病的控制指標與相關研究………………………… 13
第三節 糖尿病與家庭參與的相關研究………………………… 16
第四節 糖尿病之疾病知識、疾病態度的相關研究…………… 18
第五節 糖尿病與自我照顧的相關研究………………………… 21
第六節 研究架構………………………………………………… 23
第三章 研究方法
第一節 研究設計………………………………………………… 24
第二節 研究對象………………………………………………… 27
第三節 研究工具………………………………………………… 28
第四節 研究工具信效度檢定…………………………………… 32
第五節 倫理考量………………………………………………… 34
第六節 資料收集過程…………………………………………… 35
第七節 資料處理與分析………………………………………… 38
第四章 研究結果
第一節 糖尿病病患之基本屬性分布…………………………… 40
第二節 兩組照護模式對於糖尿病控制指標之比較…………… 44
第三節 兩組照護模式對於家庭支持行為、疾病知識、態度、
自我照顧行為之影響……………………………….…… 48
第五章 討論
第一節 兩組照護模式對於糖尿病控制指標之差異………….…... 51
第二節 兩組照護模式對於家庭支持行為、疾病知識、疾病態度
及自我照顧行為之影響………………...………………….. 53
第六章 結論與建議
第一節 結論…………………………………………….………… 56
第二節 研究之貢獻…………………………………………….… 59
第三節 研究限制與未來研究方向之建議……………………….. 61
參考資料
中文部份…………………………………………………………..... 62
英文部份…………………………………………………………..... 66
附錄
附錄一 糖尿病病人基本資料登錄表
附錄二 糖尿病個案家庭衛教及管理記錄表
附錄三 糖尿病個案居家電話追蹤與諮詢表
附錄四 糖尿病家庭支持行為量表
附錄五 糖尿病人知識、態度量表
附錄六 糖尿病自我照顧行為量表
附錄七 專家內容效度專家名冊
附錄八 致專家效度鑑定函
附錄九 專家內容效度檢定CVI結果
附錄十 糖尿病家庭參與照護之研究同意書
附錄十一 糖尿病一般照護之研究同意書


圖表目次
頁數
圖一 以家庭參與介入於第二型糖尿病控制不佳患者之架構概
念圖…………………………………………………………… 23
圖二 以家庭參與介入於控制不佳之第二型糖尿病患者之資料收
集流程圖……………………………………………………… 37
圖三 兩組第二型糖尿病病患A1C平均值之差異比較………….. 46
表一 研究設計……………………………………………………… 25
表二 家庭參與組與一般照護組之照護模式比較………………… 26
表三 各量表之信度分析結果………………………………… 33
表四 資料統計分析方法…………………………………………… 39
表五 兩組第二型糖尿病個案之基本資料分析比較……………… 42
表六 流失與收案之第二型糖尿病個案之基本資料分析比較…… 43
表七 兩組第二型糖尿病病患之糖尿病控制指標之前後差異比較 47
表八 兩組第二型糖尿病病患之家庭支持行為、疾病知識、態度
及自我照顧行為之前後差異比較…………………………… 50
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系統識別號 U0007-0407200923272700
論文名稱(中文) 應用決策樹預測壞死性筋膜炎病人的死亡率
論文名稱(英文) Predicting mortality in patients with necrotizing fasciitis by using decision tree algorithm
校院名稱 臺北醫學大學
系所名稱(中) 醫學資訊研究所
系所名稱(英) Graduate Institute of Biomedical Informatics
學年度 97
學期 2
出版年 98
研究生(中文) 范文誌
學號 G158096009
學位類別 碩士
語文別 英文
口試日期 2009-06-17
論文頁數 48頁
口試委員 指導教授-蔣以仁
委員-李元綺
委員-邱泓文
委員-鄧昭芳
委員-陳日昌
關鍵字(中) 決策樹
壞死性筋膜炎
死亡率
關鍵字(英) mortality
necrotizing fasciitis
decision tree
學科別分類
中文摘要 目的: 試圖從入院臨床特徵或檢查找出簡單的規則,以求能預測死亡率及辨識出高死亡率的壞死性筋膜炎病人。

方法: 此回溯性的病例回顧研究收錄了兩個醫院,出院診斷為壞死性筋膜炎的急診成人病患。運用chi-square檢驗(或Mann-Whitney-U檢驗)和C4.5決策樹來分析23項不同的入院臨床特徵或檢查變項。預測的結果變項為院內死亡。

結果: 共收錄了272名病患。死亡率為17%。在單變量分析方面,肝硬化、癌症、慢性腎病、腎上腺不足、低血壓、白血球數、白血球band form與血紅素,都和死亡有顯著相關。而C4.5決策樹則找出低血壓、白血球數、與白血球band form 3個獨立變項,並衍生出六條規則將壞死性筋膜炎病人分為高死亡率與低死亡率兩群。經交叉驗證後,C4.5決策樹的準確度為84.2% (95% 信賴區間,80.3%-88.1%)。

結論: 藉著常規的量血壓與檢驗白血球,急診醫師可快速地找出高死亡率的壞死性筋膜炎病人。
英文摘要 Objective: To identify simple admission clinical characteristics or laboratory tests to predict mortality but also differentiate between high and low mortality risk groups in patients with necrotizing fasciitis.

Methods: This retrospective chart-review study included adult patients who were admitted to two hospitals through the emergency department with the discharge diagnosis of necrotizing fasciitis. Both the chi-square test (or the Mann-Whitney-U test) and C4.5 decision tree were utilized to analyze 23 variables among clinical characteristics and laboratory tests. The main outcome measure was in-hospital mortality.

Results: 272 patients were included and the overall mortality rate was 17%. On univariate analysis, significant variables associated with mortality included liver cirrhosis, cancer, chronic kidney disease, adrenal insufficiency, hypotension, white blood cell (WBC) count, WBC band form, and hemoglobin. Three independent predictors of mortality - WBC count, WBC band form, and hypotension- were determined by means of the C4.5 decision tree. From these predictors, six decision rules were produced to classify patients with necrotizing fasciitis into high and low mortality risk groups. The accuracy of C4.5 decision tree with cross-validation was 84.2% (95% confidence interval, 80.3%-88.1%).

Conclusions: By using routine blood pressure measurement and simple laboratory test, WBC count and differential, emergency physicians may rapidly identify patients with high mortality.
論文目次 Chapter 1 Introduction
1.1 Background--------------------------------------------1
1.2 Purpose-----------------------------------------------2
Chapter 2 Literature Review
2.1 Necrotizing fasciitis---------------------------------3
2.1.1 Pathophysiology-----------------------------------3
2.1.2 Classification------------------------------------3
2.1.3 Diagnosis-----------------------------------------5
2.1.4 Treatment-----------------------------------------6
2.2 Necrotizing fasciitis studies in Taiwan---------------7
2.3 Prediction models for mortality of necrotizing fasciitis--------------------------------------------------9
Chapter 3 Mathods and Materials
3.1 Study design-----------------------------------------11
3.2 Materials--------------------------------------------13
3.2.1 Study population---------------------------------13
3.2.2 Inclusion criteria-------------------------------13
3.2.3 Exclusion criteria-------------------------------13
3.2.4 Attribute definition-----------------------------14
3.3 Methods----------------------------------------------15
3.3.1 Univariate analysis------------------------------15
3.3.1.1 Chi-square test-----------------------------15
3.3.1.2 Mann-Whitney-U test-------------------------17
3.3.2 Multivariate analysis----------------------------20
3.3.2.1 Prediction model: C 4.5 decision tree-------20
3.3.2.2 Data mining software: IndexMiner------------27
3.3.3 Model accuracy measurement-----------------------30
Chapter 4 Results
4.1 Patient demographics---------------------------------31
4.2 Univariate analysis----------------------------------33
4.3 Multivariate analysis--------------------------------35
4.4 Model accuracy---------------------------------------37
Chapter 5 Discussion
5.1 Mortality predictors---------------------------------38
5.2 Comparison with other multivariate analysis----------40
5.3 Model accuracy---------------------------------------42
Chapter 6 Conclusion
6.1 Conclusion-------------------------------------------43
6.2 Future Prospect--------------------------------------44
Reference-------------------------------------------------45
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系統識別號 U0007-0901200918334800
論文名稱(中文) 在體外模式下利用週期性機械力探討心肌細胞內加壓素 基因表現及其分子調控之機轉
校院名稱 臺北醫學大學
系所名稱(中) 醫學科學研究所
系所名稱(英) Graduate Institute of Medical Sciences
學年度 97
學期 1
出版年 98
研究生(中文) 李育憲
學號 G160095007
學位類別 碩士
語文別 中文
口試日期 2008-12-30
論文頁數 68頁
口試委員 指導教授-林建煌
共同指導教授-徐國基
委員-駱惠銘
委員-蕭哲志
委員-林秋梅
關鍵字(中) 加壓素
關鍵字(英) urotensin II
學科別分類
中文摘要 加壓素是一種血管收縮促進因子,可能參與高血壓和冠狀動脈硬化症的形成。目前研究顯示,在體外實驗模式下,加壓素可作為心肌細胞肥大的指標基因,並可能與心血管疾病上扮演特定連結。在臨床上已有證據顯示,加壓素在心血管疾病上可視為一種獨立潛在危險因子,在心肌細胞肥大及鬱血性心衰竭時,血漿中的加壓素含量會顯著增加。也有一些相關的研究指出在心血管系統中,ROS 也可能刺激加壓素的增加,進而誘發心肌細胞肥大和心肌纖維化,以致可以加速心血管疾病的病程變化,但目前對其分子調節機轉並不清楚。機械力過度負荷是造成心臟疾病原因之一,利用直接的機械展延來研究加壓素在心肌細胞的表現尚未被探討過,而且我們認為加壓素在機械展延下之表現可能與 ROS 的調節作用有關。因此本篇研究的目的:在體外模式下利用週期性機械力探討心肌細胞內加壓素基因表現及其分子調控之機轉。本研究是將心肌細胞接種於一矽膠彈性膜上,再配合機械展延儀器,於每分鐘 60次循環下達到 20% 或 10% 的展延程度。結果發現週期性機械力明顯地會使加壓素蛋白質與其訊息核糖核酸分子表現上升也會透過 STAT1 使的去氧糖核酸-蛋白質轉錄因子的結合活性能力上升。當加入相關抑制劑 ( PD98059, NAC, ERK siRNA, urotensin II siRNA ) 後則又會使的加壓素蛋白質和去氧核糖核酸-蛋白質轉錄因子的結合活性表現下降。最後得到的結論則是:心肌細胞在週期性機械力作用後會經由 ERK 和 STAT1 訊息傳遞路徑影響加壓素上升最後誘發導致心肌纖維化等等的相關心血管疾病。
英文摘要 Urotensin II is a potent vasoconstrictive peptide and involves in hypertension and atherosclerosis. A recent study identified that cardiomyocytes are able to express urotensin II receptor and receive the direct hypertroplic signal from urotensin II. Clinical evidence implicated that urotensin II as a potential independent risk factor for coronary heart disease which increased plasma urotensin II levels are correlated with cardiac hypertrophy and congestive heart failure. Thus, it suggested that urotensin II may play an important role in biological link of cardiovascular pathophysiologies. Several other studies have also indicated that ROS is also able to stimulate the generation of urotensin II in cardiovascular system and leading to cardiac hypertrophy. While urotensin II has been shown increase the degree of cardiovascular diseases but its molecular regulation mechanism remains unclear. Mechanical force overload is both cause and consequences of most heart diseases. How cyclical mechanical stretch affects the regulation of urotensin II in cardiacmyocytes has not been previously characterized. We sought to investigate the cellular and molecular mechanisms of regulation of urotensin II by cyclic stretch in cadiacmyocytes. In the present study, cadiacmyocytes grown on a flexible membrane base were stretched by vacuum to 20% of maximum elongation, at 60 cycles/min. Cyclic stretch significantly increased urotensin II protein and mRNA expression after stretch. Cyclic stretch also significantly increased DNA-protein binding activity of STAT1. Addition of PD98059, NAC, ERK siRNA, urotensin II siRNA 30 minutes before stretch inhibited the induction of urotensin II protein and abolished the DNA-protein binding activity induced by cyclic stretch. In conclusion, cyclic mechanical stretch enhances urotensin II expression in cultured cadiacmyocytes. The stretch-induced urotensin II is mediated by angiotensin II and ROS, at least in part, through ERK and STAT1 pathway.
論文目次 目錄
中文摘要 ……………………………………………………………1
英文摘要 ……………………………………………………………3
一、研究背景 ………………………………………………………5
動脈粥樣硬化 ( Atherosclerosis ) ……………………5
Urotensin II …………………………………………………7
機械性展延(Mechanical Stretch) ……………………8
心臟肥大(Cardiac Hypertrophy) …………………10
二、研究動機 …………………………………………………………13
三、研究方法與步驟…………………………………………………14
心肌細胞分離與培養
( Primary cardiomycytes culture ).14
心肌細胞周期性展延
(In vitro cyclical stretch on culturedcardiomycytes)15
心肌細胞核醣核酸的抽取 ( RNA extraction ……………16
去氧核糖核酸探針之製備( cDNA probe preparation )…17
- 反轉錄 ( Reverse transcription ) ……………17
- Real-time quantitative PCR ……………………17
- 純化PCR 後產物 ( PCR product purification ).18
Total protein 的抽取 ( Total protein extraction ) .19
西方墨點轉印法 ( Western Blotting )…………………20
RNA 干擾 ( RNAinterference )……………………………22
促進子活性分析法 ( Promoter acivity assay )………22
Angiotensin II 酵素免疫分析法
( Enzyme-linked immunoassay , ELISA ) …………………23
免疫螢光染色分析法
( Immunofluorescence staining assay )…………………24
電泳膠移動分析
( Electrophoretic Mobility ShiftAssay )…………… 25
- 核蛋白的抽取 …………………………………………25
- Oligomer DNA primer preparation ………………26
- End labeling probe …………………………………27
- Gel shifting …………………………27
統計分析…………………………………28
四、結果 ……………………………………………………………30
Urotensin II 蛋白質表現 ……………………………30
Urotensin II mRNA 表現 ……………………………30
ERK1/2 (p42/p44 MAP kinase)的活化 ……………31
抑制劑與RNA干擾 …………………………………………33
促進子活性 ………………………………………………34
Gel Shifting 表現 …………………………………………35
五、討論 …………………………………………………………38
六、參考文獻 ……………………………………………………43
圖表 …………………………………………………………………50
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------------------------------------------------------------------------ 第 6 筆 ---------------------------------------------------------------------
系統識別號 U0007-0907200711050400
論文名稱(中文) 台灣地區鮑氏不動桿菌分子分型資料庫及其整合子分子抗藥性機轉之研究
論文名稱(英文) Molecular Fingerprinting Database Establishment of Acinetobacter baumannii in Taiwan and the Study of Antibiotic Resistance of Integron.
校院名稱 臺北醫學大學
系所名稱(中) 醫學科學研究所
系所名稱(英) Graduate Institute of Medical Sciences
學年度 95
學期 2
出版年 96
研究生(中文) 操雅婷
學號 M102094011
學位類別 碩士
語文別 中文
口試日期 2007-07-06
論文頁數 76頁
口試委員 指導教授-吳瑞裕
委員-林俊茂
委員-周正中
關鍵字(中) 鮑氏不動桿菌
多重抗藥性
ERIC PCR
REP PCR
Integron
TEM
SHV
關鍵字(英) Acinetobacter baumannii
multidrug-resistant
ERIC PCR
REP PCR
Integron
TEM
SHV
學科別分類
中文摘要 在抗生素的不當使用之下,已造成抗藥性細菌快速演化並且大量的產生。近來,鮑氏不動桿菌(Acinetobacter baumannii)成為醫院中感染的重要病原菌之一。而具有多重抗藥性的A. baumannii(multidrug-resistant Acinetobacter baumannii; MDR-AB)更是台灣特有並常引起院內的突發性感染,在國內已陸續出現對第三線抗生素imipenem具抗藥性的菌株,是目前院內感染致病菌之中,最難以控制及治療的革蘭氏陰性菌之一。
如何快速並準確鑑定致病菌的類別與分型對於臨床及流行病學是最為迫切的。我們以ERIC PCR及REP PCR兩種分子指紋圖譜技術來進行細菌的分子分型,以完整的數位圖檔建立台灣地區鮑氏不動桿菌標準之分子分型資料庫。
除此,整合子(integron)是近年來被認為是細菌透過水平傳播以獲得新抗藥性基因及抗藥性基因散佈最重要的機制之一。整合子內之基因卡匣(gene cassettes)上可攜帶有各類型的抗藥基因,細菌經由integron主導基因卡匣透過進行特定部位重組作用的方式嵌入或移出integron,造成攜帶之抗藥性基因種類的改變和抗藥性基因的散佈,因此,integron在臨床菌株抗藥性之分佈扮演著重要角色。
廣效性乙內醯胺酶(Extended-spectrum β-lactamase;ESBL)已知對二、三代乙內醯胺具有較強的抗藥性,其中最具代表性的為TEM與SHV兩大類。因此本論文研究亦針對TEM和SHV兩組的基因序列,進行PCR分析,並探討其基因組成。
藉由上述ERIC PCR、REP PCR分子分型的結果、MIC資料庫、integron內抗藥性基因組成、TEM及SHV之變異等完整資料,進行各項比對分析,進而希望能發展出一套能夠提供臨床進行快速準確的菌種鑑別與治療方針的系統平台。
英文摘要 Because of the misuse in antibiotics administration, bacteria with drug resistance have been increased and transformed dramatically. Acinetobacter baumannii has become one of the most important clinical pathogens in these years. Recently multidrug-resistant A. baumannii (MDR-AB) has high occurrence involving in nosocomial outbreak in several Taiwan medical centers, and also been demonstrated to against the third generation β-lactam, such as imipenem. Therefore, A. baumannii now is recognized as one of the most difficult grem-negative bacteria to deal with in clinical treatment and epidemiology.
Rapid and accurate methods to identify the bacterial species and genotype are the most critical issue in clinical treatment and epidemiological study. ERIC (Enterobacterial repetitive intergenic consensus sequence) and REP (repetitive extragenic palidromic sequence) PCR have demonstrated with excellent discriminative capability among bacterial pathogens, and were used for genotyping of A. baumannii to establish full database in our previous research.
Many studies have shown that integron play an important role involving in the horizontal transfer of antibiotic resistance genes among different bacterial species. Gene cassette could carry large varieties of resistance genes within integron through site-specific recombination to move-in or move-out the integron. We first obtained full length of gene cassette through integron PCR, then analyzed each resistance gene using DNA sequencing and NCBI sequence blast.
Extended-spectrum β-lactamase is the enzyme defined as the ability hydrolyze to the second-or third-generation of β-lactam antibiotics. Several common ESBL types are identified with highly occurring in variety of AB or MDR-AB strains such as TEM and SHV, this study can establish a standard platform for accurate bacterial identification system that may provides a novel treatment study to bacterial infection in clinical.
論文目次 縮寫表………………………………………………………………………5
中文摘要……………………………………………………………………7
英文摘要……………………………………………………………………9
第一章 序論………………………………………………………………11
一、A. baumannii鮑氏不動桿菌歷史背景……………………………12
二、抗生素的種類與作用機轉…………………………………………12
三、細菌抗藥性的分子機轉……………………………………………14
四、A. baumannii的抗藥性及其機轉…………………………………16
五、A. baumannii抗藥性基因之傳遞…………………………………17
六、菌種鑑別的分子方法………………………………………………22
第二章 研究動機、目的與實驗設計……………………………………27
一、研究動機、目的……………………………………………………27
二、實驗設計……………………………………………………………27
三、利用分子生物學方法研究之A. baumannii抗藥性分子機轉………29
第三章 實驗材料與方法…………………………………………………30
一、實驗菌種來源及保存………………………………………………30
二、A. baumannii檢體之分子分型……………………………………31
第四章 結果………………………………………………………………35
一、A. baumannii genomic DNA萃取與純化…………………………35
二、ERIC PCR分子定型之結果………………………………………35
三、REP PCR分子定型之結果…………………………………………37
四、Integron存在於A. baumannii中之結果…………………………37
五、TEM gene與SHV gene存在於A. baumannii中之情形…………39
第五章 討論………………………………………………………………40
一、A. baumanni的ERIC PCR分子分型與MIC資料之比對………40
二、A. baumanni的REP PCR分子分型………………………………43
三、A. baumanni的Integron PCR分子分型與MIC資料之比對……44
四、TEM PCR與SHV PCR……………………………………………50
第六章 結論………………………………………………………………51
第七章 研究結果之應用………………………………………………53
參考文獻……………………………………………………………………55
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系統識別號 U0007-1108200804174000
論文名稱(中文) 探討玻尿酸-四環黴素膠體經由關節內注射具協同性治療退化性關節炎
論文名稱(英文) THE COMBINED EFFECT OF HYALURONIC ACID AND DOXYCYCLINE VIA INTRA-ARTICULAR INJECTABLE GELS FOR THE TREATMENT OF OSTEOARTHRITIS
校院名稱 臺北醫學大學
系所名稱(中) 醫學科學研究所
系所名稱(英) Graduate Institute of Medical Sciences
學年度 96
學期 2
出版年 97
研究生(中文) 藍絜
學號 M109095011
學位類別 碩士
語文別 英文
口試日期 2008-07-14
論文頁數 108頁
口試委員 指導教授-陳建和
共同指導教授-許明照
共同指導教授-梁有志
委員-宋信文
委員-王德原
關鍵字(中) 關節炎(OA)
減緩關節炎症狀藥物(Disease-modifying osteoarthritis drugs,DMODs)
改善連接組織結構藥物 (connective tissue structure- modifying agents,CTSMAs)
四環黴素(Doxycycline, DC)
基質金屬蛋白?類(MMPS)
第二型膠原蛋白(type II collagen)
玻尿酸Hyaluronic acid (HA)
關鍵字(英) osteoarthritis (OA)
disease-modifying osteoarthritis drugs (DMODs)
connective tissue structure- modifying agents (CTSMAs)
Doxycycline (DC)
type II collagen
matrix metalloproteinases (MMPs)
Hyaluronic acid (HA)
學科別分類
中文摘要 退化性關節炎是一種緩慢漸進式的退化性疾病,目前治療關節炎藥物的研究重點,在於進一步修飾已知傳統藥物之官能基,使其成為具有改善連結組織結構(CTSMAs)以及減緩關節炎症狀之藥物(DMODs)。國外臨床上四環黴素(DC)已普遍使用於治療關節炎,它能抑制第一型介白素B(IL-1B) 和基質金屬蛋白酶類(MMPs)的生合成,也能改善subchondrobone的組織結構,長期使用能有效的治療關節炎。關節腔直接注射玻尿酸(HA)亦是目前臨床上廣泛用於治療關節炎的方式,可抑制MMPs的表現及促進醣胺聚醣(GAG)生成外,亦具備高黏度與黏彈的特性,給予關節物理性的保護並提供止痛的效果。本篇論文是利用鋅離子連結DC及玻尿酸,於外科手術引發關節炎之紐西蘭白兔做關節腔直接注射法來驗證HA-DC膠體是否具備協同之治療效果。本次研究結果顯示,此HA-DC膠體能提供較佳之黏彈性質,並具有止痛功能;此巨型網狀結構使得HA不易被降解,可延緩並增強藥物保護關節軟骨之效果。而關節軟骨染色與切片的結果更顯示,使用HA-DC膠體之組別具有較正常之關節軟骨分佈,只產生少量的磨損裂縫及骨刺,顯示此藥物產生修復與抑制軟骨降解之效果;且相較於單獨使用HA或DC顯示了更佳的治療效果。這些實驗結果符合我們先前一系列的細胞實驗與膠體備製測試,數據皆顯示此HA-DC膠體結構確實能有效減緩關節炎症狀並提供良好之止痛效果。
英文摘要 Osteoarthritis (OA) is a degenerative disease that disunites the extra cellular matrix (ECM) of articular cartilage and is arduous to cure because cartilage is a complex and nonvascular tissue. Treatment of OA has targeted modifiable reagent to be the connective tissue structure-modifying agents (CTSMAs) and disease- modifying osteoarthritis drugs (DMODs), such as hyaluronic acid (HA) or modified tetracycline. However, the aim of this present study was to examine whether intra-articular injection of the HA-DC gel restores the matrix of arthritic knee joints in rabbit model. The injectable HA-DC gel was composed of HA, doxycycline (DC) and zinc chloride. It was administered five times (at days 0, 3, 6, 9 and 12) into the partial meniscectomy and unilateral fibular ligament transected knee joints of 13-week old New Zealand White rabbits, and the effects of HA-DC gel injection were compared with that of DC, HA or normal saline. Evaluation of OA progression was histopathologically by Alcian blue and eosin staining, by the modified grading method and by scanning electron microscopy at day 14. HA-DC gel group showed smoother cartilage surface, no or minimal signs of ulceration, smaller osteophytes, and less fissure formation than the other groups. Normal distribution of chondrocytes or inflammation was noticed in areas of slight OA changes in the HA-DC gel group indicating regenerative ability. HA-DC gel seemed to inhibit the progression of osteoarthritis by protecting the layers of articular cartilage, decreasing the degree of pain and restoring the elasticviscosity. Both macroscopic and histological data of this study supported those biochemical results and gel preparation of previous studies showing the synergistic therapeutic effect of HA-DC gel on osteoarthritis.
論文目次 INDEX I
GRAPHICS INDEX IV
TABLE INDEX VII
I. 摘要 VIII
II. ABSTRACT IX
1 INTRODUCTION 1
2 MATERIALS AND METHODS 5
2.1 PREPARATION OF HYALURONAN–DOXYCYCLINE GELS 5
2.1.1 Materials 5
2.1.2 Confirmation of precipitation- turbidity 5
2.1.3 Gel preparation 5
2.1.4 Viscosity analysis 7
2.1.5 Elastoviscosity evalution 7
2.1.6 Statistical analysis 7
2.2 IN VITRO CELL EXPERIMENTS 8
2.2.1 Cell Lines and Reagents 8
2.2.2 Human bovine chondrocyte isolation and culture 8
2.2.3 Gel preparation 8
2.2.4 WST-1 assay 8
2.2.5 MTT cell viability assay 9
2.2.6 Reverse transcription-coupled PCR (RT–PCR) Analysis 9
2.2.7 Western Blot Analysis 11
2.2.8 Statistical analysis 11
2.3 IN VIVO ANIMAL EXPERIMENTS 12
2.3.1 Reagents 12
2.3.2 Gel preparation 12
2.3.3 Animals 12
2.3.4 Experimental OA model 13
2.3.5 Experimental design 13
2.3.6 Observation of general status and body weight measurement 14
2.3.7 Pain assessment 14
2.3.8 Specimen collection 15
2.3.9 Macroscopic specimen examination 15
2.3.10 Miroscopic specimen examination 15
2.3.11 Statistical analysis 19
3 RESULTS 20
3.1 PREPARATION OF HA-DC GEL 20
3.1.1 Higher concentration of HA accompanied higher viscosity and pH value. 20
3.1.2 Interaction of HA and DC induced precipitation. 20
3.1.3 pH value and DC concentration influenced the extent of precipitation. 21
3.1.4 Temperature and DC concentration influenced the extent of precipitation. 21
3.1.5 Concentration of HA and DC influenced the extent of precipitation. 21
3.1.6 DC consistency influenced the pH value. 22
3.1.7 The effect of different concentration of DC influenced viscosity and turbidity. 22
3.1.8 Zinc/ DC mole ratio influenced viscosity and pH. 22
3.1.9 Formation of gelatinization: the HA-DC gel was formed with a dose dependence manner. 23
3.1.10 Elastoviscosity property of HA-DC gel and HA-DC structure. 24
3.2 CELL EXPERIMENTS 25
3.2.1 Treatment of higher DC concentration induced cell death. 25
3.2.2 Treatment of DC reduced the gene expressions of MMP-1, MMP-3 and MMP-13 in IL-1β induced inflammatory SW1353. 25
3.2.3 Treatment of DC induced no influence on the gene expression of IL-8 in inflammatory SW1353 induced by IL-1β. 26
3.2.4 Treatment of IL-1β induced type II collagen expression but DC decreased that. 26
3.2.5 Treatment of HA (Artz-Dispo) stimulated chondrocyte cell proliferation. 26
3.2.6 Treatment of HA (Artz-Dispo) reduced the gene expression of MMP-1, MMP-3, MMP-13 in SW1353 induced by IL-1β. 28
3.2.7 Treatment of HA (Artz-Dispo) reduced the gene expression of IL-8 in SW1353 induced by IL-1?? 28
3.2.8 Treatment of HA (Artz-Dispo) increased the gene expression of type II collagen in SW1353. 29
3.2.9 Treatment of HA-DC structure induced serious inflammatory reaction 29
3.2.10 Treatment of HA-DC gels influenced cell survival in SW1353. 29
3.2.11 Treatment of HA-DC gel reduced the gene expression of MMP-1, MMP-3, MMP-13 in SW1353 induced by IL-1β. 30
3.2.12 Treatment of HA-DC gels reduced the gene expression of IL-8 in SW1353 induced by IL-1β. 30
3.3 ANIMAL EXPERIMENTS 32
3.3.1 Observation of general status and body weight measurement. 32
3.3.2 Treatment of DC, HA, gel or NS influenced the percentage distribution on left hind paw. 32
3.3.3 Inhibitory effect of reagents on the cartilage degradation and erosion of macroscopical appearances. 32
3.3.4 Inhibitory effect of reagents on the cartilage degradation and erosion of microscopical appearances. 33
4 DISCUSSION 35
5 REFERENCES 47
6 FIGURES 53
6.1 HYALURONAN–DOXYCYCLINE GELS PREPARATION. 53
6.2 IN VITRO CELL EXPERIMENTS 67
6.3 IN VIVO ANIMAL EXPERIMENTS 86
7 APPENDIX 106

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系統識別號 U0007-1407200816165200
論文名稱(中文) 淫羊藿苷對於LPS誘發的退化性關節炎之作用
論文名稱(英文) Effects of Icariin on LPS-Induced Osteoarthritis
校院名稱 臺北醫學大學
系所名稱(中) 藥學研究所
系所名稱(英) Graduate Institute of Pharmacy
學年度 96
學期 2
出版年 97
研究生(中文) 蔡曉雯
學號 M301095002
學位類別 碩士
語文別 中文
口試日期 2008-06-19
論文頁數 102頁
口試委員 指導教授-許秀蘊
共同指導教授-孫瑞昇
委員-蔡麗雪
委員-邱仁輝
關鍵字(中) 淫羊藿苷
退化性關節炎
關鍵字(英) Icariin
osteoarthritis
學科別分類
中文摘要 隨著老年人囗的成長,關節炎病症的持續惡化會嚴重影響病人的生活起居,並增加社會經濟的負擔。對年長者而言,關節炎是限制行動的首要原因,而且可能是造成殘疾的主要因素。關節軟骨是由水份、軟骨細胞和軟骨細胞分泌的胞外基質(extracellular matrix,ECM)所組成,分布在可動關節的表面,保護骨頭、減輕磨損、幫助關節活動和承載身體重量。當關節軟骨發生病變,日常活動就會受到阻礙;最常見的關節病變是骨關節炎,又稱退化性關節炎,全球約有6%的人(台灣約有15%)罹患此病,是由於老化、遺傳或是過度使用所造成,其最主要病理特徵是軟骨ECM瓦解。而基質金屬蛋白酶 (matrix metalloproteinase,MMP) 則可以分解ECM中的蛋白質成分,造成ECM的瓦解。
淫羊藿苷(icariin)是從植物劍葉淫羊藿(Epimedium sagittatum, Berberidaceae )中所萃取出來具藥理活性的成分。它具有很多的作用,例如:抗氧化、免疫調節和勃起功能…等。除了上述的作用之外,它還能夠刺激骨母細胞 (osteoblasts) 的分化,提高alkaline phosphatase (ALP) 的活性,增加大鼠體內第一型膠原蛋白(Type I collagen)的表現,幫助骨頭的生長。並且,icariin在發炎反應上面,可以降低組織胺的量,因此可以降低毛细血管通透,並且能夠減輕腳腫脹的程度。到目前為止,icariin 還沒有明顯的不良反應。本研究欲探討在軟骨受傷害的情況下,icariin是否有治療或保護的作用,更進一步的說,或許可提供另一個對於軟骨傷害治療的新方向。本實驗取用初生四天之幼鼠為動物模式,取其膝關節軟骨培養軟骨母細胞,並採用三度空間培養法 (3-dimensional cultures) 培養軟骨細胞,然後再加入icariin 或在lipopolysaccharide (LPS)存在的情況下,分別來模擬自然老化退化性關節炎的情形,並檢測軟骨細胞增生情形,再進一步的檢測軟骨細胞內基質glycosaminoglycans (GAG),以及軟骨細胞內基質total collagen的量,與一些會造成軟骨傷害的物質的表現,如nitric oxide (NO),metalloproteinases (MMPs),COX-2..等等。並且利用即時定量反轉錄聚合酶連鎖反應 (Real Time - PCR) 來探討基因MMP-1,MMP-3,MMP-13,COX-2,iNOS對軟骨細胞的影響。或許可提供另一個預防軟骨傷害及治療的新方向。
結果顯示,Icariin在誘導軟骨細胞增生時,在10-9M濃度下時最佳,而在加LPS 情況時,10-9M濃度下icariin對LPS的傷害具有保護作用,LPS可以誘導NO、MMP1、MMP3、MMP13產生,但可以抑制GAG、total collagen合成。Icariin可以部分的減少NO、MMP1、MMP3、MMP13產生。所以Icariin在軟骨細胞受傷害的情況下具有保護作用。所以Icariin可扮演一個對抗軟骨傷害的角色。
英文摘要 With the growth of the population of elder persons, arthritis disease will seriously affect the continued deterioration of the patient's daily life, and to increase the socio-economic burden. To the elderly, arthritis is the first cause of restrictions on movement, and may cause disability is the main reason. Articular cartilage is composed of water, chondrocytes and extracellular matrix (extracellular matrix, ECM). The components are in the distribution of movable joints on the surface, the protection of the bones, reducing wear and tear, to joint activities and help carry the body weight. When the articular cartilage lesion occurs, the daily life will be hindered; the most common joint disease is osteoarthritis, and it is also known as degenerative arthritis, about 6% people of the world suffering from the disease is the result of aging , genetic or caused by the excessive use. Its main feature is the cartilage ECM collapse. The MMP (matrix metalloproteinase) can break down the proteins in ECM, resulting in the collapse of ECM. Icariin is the major pharmacologically active component of Epimedium sagittatum (Berberidaceae) It has a lot of functions, for example: antioxidant, immune regulation and erectile function, et. Al. One of the components, Icariin can reduce the amount of histamine in the inflammatory response. It also could reduce capillary permeability, and reduce the degree of swelling of feet. So far, Icariin has no obvious adverse reactions. The study was to investigate the effects of icariin on lps-induced osteoarthritis. The animal model is four days of newborn ICR mice, using its knee cartilage to culture chondrocytes. To avoid the change of phenotype on chondrocyte, we use three-dimensional culture model . Further detection of glycosaminoglycan (GAG) within the matrix, we use DMB assay. The OHP assay is used to detect the content of total collagen, and DMB assay to glycosaminoglycan (GAG) within the matrix. Real time PCR was used to investigate gene MMP1, MMP3, MMP13, COX-2 and iNOS on cartilage cells. It might provide another treatment for cartilage damage in the new direction. The results show that icariin in the induction of chondrocyte proliferation, at the 10-9 M concentration is the best. In the same concentration, icariin has a protective effect from the injury of LPS. LPS can induce NO, MMP-1, MMP-3, MMP-13 production, but it inhibits the GAG, total collagen synthesis. Icariin can reduce part of NO, MMP-1, MMP-3, MMP-13 production.Therefore, in the situations of cartilage cells injury, icariin may have a protective effect. It showed us that icariin might provide another treatment for cartilage damage in the new direction.
論文目次 第一章 緒論 12
1-1前言 12
1-2 研究目的 14
第二章 理論基礎 16
2-1 硬骨的發育與生長 16
2-1-1膜內骨化(Intramembranous ossification) 16
2-1-2 軟骨內骨化 (Endochondral ossification) 17
2-2 軟骨的發育與生長 (相關參考資料 Wheater’s Functional Histology) 19
2-2-1造成軟骨損傷原因 20
2-3 軟骨相關之病理研究 (相關參考資料 Robbins Pathologic Basis of Disease) 21
2-3-1關節炎 21
2-3-1-1退化性關節炎(Osteoarthritis) 22
2-3-1-2感染性關節炎(Infectious Arthritis) 24
2-3-1-3化膿性關節炎(Suppurative Arthritis) 24
2-3-1-4結核性關節炎(Tuberculous Arthritis) 24
2-3-1-5病毒性關節炎(Viral arthritis) 25
2-3-2治療方法 26
2-4軟骨相關成分分析 28
2-4-1 膠原蛋白 (collagen) 29
2-4-2 醣蛋白 (proteoglycan) 30
2-4-3 前列腺素(prostaglandins) 31
2-4-4金屬蛋白分解酵素Matrix metalloproteinase (MMPs) 33
2-4-5 一氧化氮合成酶Nitric oxide synthases (NOS) 36
2-4-6 Icariin (icarrin,icarin) 38
第三章 材料與方法 40
3-1實驗試薬與耗材 40
3-1-1試薬 40
3-1-2耗材 42
表 3-1-2 1耗材 42
3-2實驗儀器 43
3-3 Icariin試藥配製 44
3-4細胞培養試藥配製 45
3-6初代培養軟骨細胞方 49
3-6-1軟骨細胞3D培養方法 51
3-6-2 Agarose cell specimens溶解方法 54
3-7細胞活性分析 54
3-8 GAG (DMB assay) 57
3-9 NO (Griess reaction) 58
3-10 Hydroxyproline assay (OHP) 59
3-11即時(Real-time)定量反轉錄聚合酶連鎖反應 61
3-11-1 Total RNA之純化 61
3-11-2 Total RNA含量分析 62
3-11-3反轉錄(Reverse transcriptase reaction, RT)合成cDNA 62
3-11-4即時定量聚合酶連鎖反應 63
3-11-5即時定量聚合酶連鎖反應使用之primer 64
第四章 結果與討論 66
4-1 細胞活性分析 ( MTT assay ) 66
4-1-1 Icariin 對 Chondrocyte 體外培養模式整體活性(MTT assay)的影響 66
4-1-2 Icariin 對lipopolysaccharide(LPS) 刺激chondrocyte的影響 70
4-2 Griess Assay 73
4-2-1 Icariin對chondrocyte產生NO的影響 73
4-2-2 Icariin對lipopolysaccharide (LPS)刺激chondrocyte產生NO的影響 75
4-3 Real Time PCR 80
4-3-1 10-9 M Icariin對COX-2 mRNA的表現 80
4-3-2 10-9 M Icariin對iNOS mRNA的表現 81
4-3-3 Icarii對MMP mRNA的表現 84
4-3-3-1 10-9 M Icariin對MMP-1 mRNA的表現 84
4-3-3-2 10-9 M Icariin對MMP-3 mRNA的表現 85
4-3-3-3 10-9 M Icariin對MMP-13 mRNA的表現 86
4-4 Icariin對LPS抑制軟骨細胞產生GAG的影響 88
4-5 Icariin對LPS抑制軟骨細胞產生Total collagen的影響 90
第五章 結論 與 未來展望 92
第六章 參考文獻 93
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系統識別號 U0007-1407201013370500
論文名稱(中文) 淫羊藿苷對骨母細胞及蝕骨細胞之影響
論文名稱(英文) Effects of Icariin on Osteoblasts and Osteoclasts
校院名稱 臺北醫學大學
系所名稱(中) 藥學系(博士班)
系所名稱(英) Pharmacy (PhD)
學年度 98
學期 2
出版年 99
研究生(中文) 謝采蓓
學號 D301096006
學位類別 博士
語文別 中文
口試日期 2010-06-29
論文頁數 156頁
口試委員 指導教授-許秀蘊
共同指導教授-孫瑞昇
委員-蔡東湖
委員-蔡麗雪
委員-劉華昌
關鍵字(中) 淫羊藿苷
骨母細胞
蝕骨細胞
關鍵字(英) Icariin
oteoblast
osteoclast
學科別分類
中文摘要 淫羊藿是一種中國常用於治療骨質疏鬆症的草藥。淫羊藿中主要活性黃酮配糖體為是淫羊藿苷(Icariin),有研究顯示可以提高骨癒合及減少骨質疏鬆症的發生。不過,詳細的分子機制仍不清楚。在本研究中探討淫羊藿苷分子機制,主要通過刺激成年小鼠骨母細胞初代培養,抑制蝕骨細胞分化和骨再吸收作用,透過抑制MAPKs / NF-κB活化表現,調控低氧誘導因子-1α (HIF-1α; Hypoxia-Inducible Factor 1α) 和前列腺素E2 (Prostaglandin E2,PGE2)的合成。
本研究中骨母細胞初代培養從八個月Imprinting Control Region (ICR) 成年母鼠取得。研究中發現Icariin會增加成年母鼠骨母細胞的活性,促進骨母細胞生成及分化。從骨母細胞活性分析發現10−8 M icariin表現最佳。在這個濃度下,Icariin會刺激細胞增生,增加鹼性磷酸酵素活性,增加骨母細胞礦化,促進一氧化氮(NO)生成。Icariin會抑制骨母細胞凋亡,藉由抑制caspase-3活化,延長骨母細胞生命週期。Icariin會增加Bone morphogenetic protein-2 (BMP-2) mRNA、SMAD4 mRNA、Core bonding factor 1 (Cbfa1/Runx2) mRNA、Osteoprotegerin(OPG)、Receptor activator of nuclear factor (NF)-kB ligand (RANKL) mRNA的基因表現。利用BMP受體拮抗劑 Noggin及一氧化氮合成酶抑制劑L-NAME來探討相關的機轉,在本研究中發現給予Nogginr及L-NAME後,可以抑制Icariin之細胞增生、ALP之活性增加、NO之生成,也會調控BMP-2 mRNA、SMAD4 mRNA、Cbfa1/Runx2 mRNA、OPG mRNA、RANKL mRNA表現。本研究經體外實驗發現Icariin為一種骨細胞刺激劑,會誘導骨生成作用,藉由BMP-2及NO生成,調控SMAD4 mRNA、Cbfa1/Runx2 mRNA、OPG mRNA、RANKL mRNA表現。
Icariin在蝕骨細胞與骨母細胞共同培養模式下,可以抑制蝕骨細胞的活性,10−8 M Icariin對於抑制蝕骨細胞特有的分化指標:抗酒石酸性磷酸酵素 (tatrate resistances acid phosphatease;TRAP) 效果最佳。Icariin會縮小細菌脂多醣體 (LPS, Lipopolysaccharide) 誘導的蝕骨細胞大小,同時降低TRAP 及酸性磷酸酵素 (acid phosphatease;ACP) 活性,但不會影響蝕骨細胞存活。Icariin也會抑制骨再吸收作用,降低Interleukin-6 (IL-6) 和Tumor necrosis factor-alpha (TNF-α) mRNA及蛋白質表現。Icariin會增強OPG mRNA 基因表現,而減少RANKL mRNA 基因表現。Icariin減少前列腺素E2 (Prostaglandin E2,PGE2)及COX-2之合成。此外,Icariin會在正常氧氣濃度下減少LPS誘導之蝕骨細胞中低氧誘導因子-1α (HIF-1α; Hypoxia-Inducible Factor 1α) 的生成。在蝕骨細胞中,Icariin減少LPS造成的ERK1/2、I-kappa-B-alpha (IκBα)、p38 及JNK活化;而在骨母細胞中,Icariin則會減少LPS造成的ERK1/2及IκBα活化,反而增加p38活化。Icariin經由p38 及 JNK路徑來抑制LPS誘導之蝕骨作用,。
總而言之,本研究發現Icariin會促進體外骨形成作用,藉由BMP-2及NO生成,抑制骨再吸收作用,預防發炎性骨質流失,減少p38 及 JNK路徑活化。由本研究發現Icariin在骨質疏鬆症的治療上深具潛力。
英文摘要 Epimedii herba is one of the most frequently used herbs in formulas prescribed for the treatment of osteoporosis in China. The main active flavonoid glucoside extracted from Epimedium pubescens Maxim. is icariin, which has been reported to enhance bone healing and reduce osteoporosis occurrence. However, the detailed molecular mechanisms remain unclear. In this present study, we examine the molecular mechanisms of icariin by using primary osteoblast cell cultures obtained from adult mice; and icariin can inhibit osteoclast differentiation and bone resorption by suppressing MAPKs/NF-κB regulated hypoxia inducible factor-1α (HIF-1α) and prostaglandin E2 (PGE2) synthesis.
The osteoblast cells were harvested from 8-month old female Imprinting Control Region (ICR) mice. The effects of icariin stimulation on the proliferation, differentiation and maturation of osteoblasts were examined. The viability of the osteoblasts reached its maximum at 10−8 M icariin. At this concentration, icariin increased the proliferation and matrix mineralization of osteoblasts and promoted nitric oxide (NO) synthesis. Icariin decresed osteoblasts apotosis by inhibiting caspase-3 activation to promote cell life cycle. With icariin treatment, the BMP-2, SMAD4, Cbfa1/Runx2, and OPG gene expressions were up-regulated; the RANKL gene expression was however down-regulated. Concurrent treatment involving the BMP antagonist (Noggin) or the NOS inhibitor (L-NAME) diminished the icariin-induced cell proliferation, ALP activity, NO production, as well as the BMP-2, SMAD4, Cbfa1/Runx2, OPG, RANKL gene expressions. In this study, we demonstrate that in vitro icariin is a bone anabolic agent that may exert its osteogenic effects through the induction of BMP-2 and NO synthesis, subsequently regulating Cbfa1/Runx2, OPG, and RANKL gene expressions.
After treatment with icariin, osteoclast co-culture cells were decreased tatrate resistances acid phosphatease (TRAP) activity significantly at the concentration of 10-8 M. Icariin reduced the size of LPS-induced osteoclasts formation, and diminished their TRAP and acid phosphatease (ACP) activity without inhibition of cell viability. Icariin also inhibited LPS-induced bone resorption and interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) mRNA and protein expression. The gene expression of OPG was up-regulated, while RANKL was down-regulated. Icariin also inhibited the synthesis of cyclo-oxygenase type-2 (COX-2) and PGE2. In addition, icariin had a dominant repression effect on LPS-induced HIF-1α expression of osteoclasts. On osteoclasts, icariin suppresses LPS-mediated activation of the ERK1/2, I-kappa-B-alpha (IκBα), p38 and JNK; while on the osteoblasts, icariin reduced the LPS-induced activation of ERK1/2 and IκBα, but increased the activation of p38. Icariin inhibited LPS-induced osteoclastogenesis program by suppressing the p38 and JNK pathway.

In conclusion, we demonstrated that icariin in vitro has bone formation activity through the induction of BMP-2 and NO synthesis, and inhibitory effects on bone resorption that can prevent inflammatory bone loss by suppressing the p38 and JNK pathway. This study was found that icariin should be a potential compound used for osteoporosis treatment.
論文目次 目 錄
目錄 I
附表目錄 III
附圖目錄 IV
名詞縮寫 VI
中文摘要 1
英文摘要 3
第一章 緒論 5
第二章 理論基礎 8
第一節 骨細胞 8
一、 骨母細胞 8
二、 骨細胞 8
三、 蝕骨細胞 9
四、 骨基質 9
第二節 骨母細胞表現 10
一、骨母細胞生理作用 10
二、骨形態發生蛋白 10
三、一氧化氮NO對骨母細胞之影響 12
第三節 蝕骨細胞表現 15
一、蝕骨細胞生理作用 15
二、RANKL/RANK/OPG 調控蝕骨細胞機制 17
三、蝕骨細胞中RANK的訊息傳遞路徑 18
四、蝕骨細胞的酸性磷酸酶活性 18
五、LPS誘導蝕骨作用 19
第四節 淫羊藿 20
一、淫羊藿 20
二、淫羊藿苷Icariin 21

第三章 材料與方法 23
第一節 材料 23
ㄧ、實驗儀器 23
二、實驗試薬與耗材 24
第二節 試藥配製 26
第三節 骨母細胞培養方法 30
第四節 蝕骨細胞共同培養之培養方法 33
第五節 細胞活性分析 35
第六節 鹼性磷酸酵素分析 36
第七節 細胞礦化Alizarin Red-S 染色分析 37
第八節 細胞一氧化氮分析 38
第九節 細胞凋亡分析 39
第十節 蝕骨細胞共同培養下酸性磷酸酵素分析 40
第十一節 蝕骨細胞共同培養下TRAP stain定性分析 42
第十二節 骨再吸收作用分析 43
第十三節 即時(real-time)定量反轉錄聚合酶連鎖反應 44
第十四節 西方點墨法 Western Blotting 51
第十五節 IL-6、TNF-?悀垵GE2測定方法 54

第四章 結果 55
第一節 Icariin對於骨母細胞之影響 55
一、Icariin 對骨母細胞不同濃度作用下細胞活性分析 55
二、Icariin促進骨母細胞分化及骨礦化 57
三、Icariin促進NO生成 61
四、Icariin會抑制骨母細胞凋亡 63
五、Icariin透過蛋白質合成增加ALP及NO表現 65
六、Icariin對 BMP-2, SMAD4, Cbfa1/Runx2, OPG,及 RANKL mRNA之影響 67
七、Icariin表現BMP-2和NO刺激骨母細胞成熟及分化可被L-NAME或Noggin抑制 74
第二節 Icariin對於蝕骨細胞共同培養之影響 84
一、Icariin減少蝕骨作用 84
二、Icariin抑制LPS誘導蝕骨作用,而不影響細胞存活 88
三、Icariin調控LPS誘導RANKL增加和OPG減少 94
四、Icariin抑制LPS誘導前發炎細胞激素生成 97
五、Icariin減少LPS誘導PGE2藉由抑制COX-2表現 100
六、Icariin抑制LPS活化MAPKs及NF-κB路徑 103
七、Icariin抑制蝕骨作用經由LPS在常氧環境下增加HIF -1α表現 108

第五章 討論 110

第六章 結論 121

參考文獻 122

謝采蓓 Hsieh, Tsai-Pei 已發表之論文 137
附表目錄
表1.實驗儀器 23
表2.實驗試薬 24
表3.細胞培養試藥配製 26
表4. Icariin試藥配置稀釋表: 27
表5. Cycloheximide試藥配置稀釋表: 28
表6. Noggin試藥配製 28
表7. L-NAME試藥配製 28
表8.細胞活性分析試藥配製 29
表9.細胞一氧化氮分析試藥配製 29
表10.細胞礦化Alizarin Red-S 染色試藥配製 29
表11.骨母細胞培養盤種類與細胞密度關係 31
表12.蝕骨細胞共同培養下培養盤種類與細胞密度關係 34
表13.細胞凋亡反應 39
表14.反轉錄反應-1 (RT-1) 45
表15.反轉錄反應-2 (RT-2) 46
表16.實驗使用之引子 47
表17.即時定量聚合酶連鎖反應 48
表18.聚合酶連鎖反應 49
表19.電泳藥品配置 50
表20.一級抗體 52
表21.西方點墨法試劑配置 53


附圖目錄
圖1. 骨母細胞分化過程合成蛋白質影響 10
圖2. BMP-2路徑的訊息傳遞 12
圖3. Icariin結構圖 21
圖4.Cycloheximide結構式 28
圖5.骨母細胞實驗流程 32
圖6.蝕骨細胞共同培養實驗流程 34
圖7.鹼性磷酸酵素(ALP)分析實驗原理 36
圖8. NO作用原理 38
圖9. Icariin 對骨母細胞不同濃度作用細胞活性影響 56
圖10. Icariin對骨母細胞分泌鹼性磷酸酵素之影響 59
圖11. Icariin對骨母細胞之骨礦化影響 60
圖12. Icariin促進NO生成 62
圖13. Icariin會抑制骨母細胞凋亡 64
圖14. Cycloheximide抑制Icariin引起的ALP活性增加及NO產生 66
圖15. Icariin對於BMP-2 mRNA之表現 69
圖16. Icariin對於SMAD4 mRNA之表現 70
圖17. Icariin對於Cbfa1/Runx2 mRNA之表現 71
圖18. Icariin對於OPG mRNA之表現 72
圖19. Icariin對於RANKL mRNA之表現 73
圖20. L-NAME或Noggin對Icariin引起細胞增生之影響 75
圖21. L-NAME或Noggin對Icariin增加ALP之影響 76
圖22. L-NAME或Noggin對Icariin增加NO之影響 77
圖23. L-NAME或Noggin對Icariin調控BMP-2 mRNA之影響 78
圖24. L-NAME或Noggin對Icariin調控SMAD4 mRNA之影響 79
圖25. L-NAME或Noggin對Icariin調控Cbfa1/ Runx2 mRNA之影響 80
圖26. L-NAME或Noggin對Icariin調控OPG mRNA之影響 81
圖27. L-NAME或Noggin對Icariin調控RANKL mRNA之影響 82
圖28. L-NAME或Noggin對Icariin調控mRNA之影響RT-PCR結果 83
圖29.不同濃度Icariin對蝕骨細胞之ACP及TRAP表現 85
圖30.不同濃度Icariin對蝕骨細胞之TRAP染色影響 86
圖31. Icariin對蝕骨細胞之ACP及TRAP時間表現 87
圖32. Icariin對LPS誘導ACP及TRAP之影響 89
圖33. Icariin對LPS誘導TRAP染色之影響 90
圖34. Icariin對LPS誘導骨再吸收作用之影響 92
圖35. Icariin對細胞存活率沒有影響 93
圖36. Icariin對LPS誘導RANKL mRNA之影響 95
圖37. Icariin對LPS減少OPG mRNA之影響 96
圖38. Icariin對LPS誘導IL-6之影響 98
圖39. Icariin對LPS誘導TNF-α之影響 99
圖40. Icariin對LPS誘導COX-2之影響 101
圖41. Icariin對LPS誘導PGE2之影響 102
圖42. Icariin在蝕骨細胞中對LPS活化MAPKs及IκBα之影響 104
圖43. Icariin在骨母細胞中對LPS活化MAPKs及IκBα之影響 105
圖44. Icariin在蝕骨細胞中對LPS活化MAPKs及IκBα第60分鐘影響 106
圖45. Icariin在骨母細胞中活化MAPKs之影響 107
圖46. Icariin在蝕骨及骨母細胞中對LPS誘導HIF-1α之影響 109
圖47. Icariin對於骨母細胞影響之分子機轉 119
圖48. Icariin作用在LPS誘導蝕骨作用之分子機轉 120

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系統識別號 U0007-2207200818171900
論文名稱(中文) 小葉葡萄神經保護功能之研究
論文名稱(英文) Study of the neuroprotective effects of Vitis thunbergii var. taiwaniana in vitro and in vivo
校院名稱 臺北醫學大學
系所名稱(中) 醫學科學研究所
系所名稱(英) Graduate Institute of Medical Sciences
學年度 96
學期 2
出版年 97
研究生(中文) 沈怡君
學號 G160094001
學位類別 碩士
語文別 中文
口試日期 2008-07-02
論文頁數 56頁
口試委員 指導教授-梁有志
委員-梁弘人
委員-侯文琪
關鍵字(中) 小葉葡萄
神經退化性疾病
海人酸
白藜蘆醇
關鍵字(英) Vitis thunbergii var. taiwaniana
Neurodegenerative diseases
Kainic acid
Resveratrol
學科別分類
中文摘要 小葉葡萄(Vitis thunbergii var. taiwaniana,簡稱VTT)是一台灣原生葡萄,與其它Vitis spp.一樣在台灣是被用來當作一種民間傳統藥物或是可食用的植物。它富含多酚類化合物(polyphenols),特別是槲皮素(quercetin)、白藜蘆醇(resveratrol),這些多酚類化合物在很多研究中,被證實具有保護心血管疾病、抗癌、抗衰老及改善腦神經退化性疾病(neurodegenerative diseases)。以白藜蘆醇為例,它具有高度抗氧化、抗發炎的活性,並且被視為是一種植物雌激素(phytoestrogen),這些活性均有助於其保護腦神經細胞。在抗氧化方面,它可直接捕捉活性氧分子(reactive oxygen species, ROS)或經由活化抗氧化酵素,來防止神經元細胞及神經膠質細胞(glial)受到活性氧分子的攻擊;在抗發炎方面,它可抑制經β-amyloid, LPS, cytokines所引起神經膠質細胞(如微小神經膠質細胞)的過度發炎反應;此外過去的研究發現estrogen具有優越的保護神經功能,更年期後的婦女若能長期服用植物性雌激素,可有效降低老人失智症的發生率,它還能對抗多種來源的氧化壓力,而白藜蘆醇具有植物雌激素的活性,我們認為白藜蘆醇也有estrogen 類似的神經保護功能。
在本研究中,我們使用小葉葡萄的根、支幹、葉及主幹的粗萃取物,在體外、體内試驗其抗腦神經退化的活性。首先在有血清培養的BV2微小神經膠質細胞中,發現支幹及葉萃取物顯著抑制iNOS蛋白質表現及NO產生; 而在無血清培養的BV2微小膠細胞中,四種粗萃取物均可顯著抑制iNOS蛋白質表現及NO產生。在初代培養的大鼠神經細胞中,處以海人酸(kainic acid, 簡稱KA) 發現可引起活性氧化物的產生,並會引起神經細胞死亡。使用四種粗萃取物均可顯著抑制活性氧化物的產生,然而只有根及支幹的萃取物可以降低KA所引起的神經細胞死亡。總結體外試驗,小葉葡萄粗萃取物降低微小神經膠質細胞發炎及神經細胞死亡的活性順序分別是支幹≈葉>根>主幹。在動物試驗方面,發現給予小葉葡萄粗萃取物,雖然不能降低小鼠發生癲癇,但顯著延後癲癇發作的時間,其中支幹萃取物及葉萃取物效果較顯著,發作的小鼠也較少。四種小葉葡萄萃取物,均可保護海馬迴神經細胞,其中小葉葡萄支幹萃取物,保護效果較佳。小葉葡萄粗萃取物在動物體上保護神經的活性順序分別是支幹≈葉>根>主幹。
英文摘要 Vitis thunbergii var. taiwaniana (VTT) is rich in quercetin, resveratrol, and other polyphenols that are known to protect against cardiovascular diseases and cancers, as well as to promote anti-aging effects in numerous organisms. Resveratrol has antioxidant and anti-inflammatory activities, and acts as a phytoestrogen. All of the abilities contributes to protect against neurodegenerative diseases. In antioxidant, resveratrol can prevent neuron and glial cells from reactive oxygen species (ROS) attack by directly scavenge ROS or indirectly activate antioxidant enzymes. On the other hand, resveratrol can inhibit inflammation induced by ??-amyloid, LPS or cytokines in microglia. Previous studies have demonstrated that estrogen has an excellent function in protecting neuron, such as Alzheimer's disease and in protecting ROS.
In this plan, we used various kinds of VTT extracts including its root, branch, leaf, and trunk to examine the protective function in neurodegenerative diseases in vivo and in vitro. We found that the branch and leaf extracts of VTT significantly inhibited iNOS protein expression and nitric oxide production in BV2 microglia when cultured in medium with 10% FBS. On the other hand, all four kinds of VTT extracts could inhibit iNOS protein expression and nitric oxide production in BV2 microglia when cultured in medium without FBS. Using primary neurons, we found that kainic acid (KA) could induce ROS production in a dose-dependent manner and finally resulted in neuron cells death. All four kinds of VTT extracts inhibited the ROS production, however only root and branch extracts were able to prevent the cell death induced by KA. The protective activity of various kinds of VTT extracts on the microglia inflammation and neuron cells death could be ranked as follows: branch≈leaf>root>trunk. In animal study, we found that i.p. injecting KA caused seizure, neurons cells death in hippocampus area, furthermore induced mice death in C57BL/6 mice. Administration of all four kinds of VTT extracts could delay the onset time of seizure and decrease the hippocampus neurons cells death. The potency of VTT extracts could be ranked as follows: branch≈leaf>root>trunk. However, all four kinds of VTT extracts could not decrease the number of mice death induced by KA.
論文目次 致謝 1
中文摘要(Abstract in chinese) 2
英文摘要(Abstract in English) 4
緒論(Introduction) 6
壹、 前言 6
貳、 神經退化性疾病 8
1. 腦神經組織簡介 8
2. 腦神經組織發炎反應 8
3. 致病因子 9
3-1. 氧化壓力(Oxidative stress) 10
3-2. 粒線體功能喪失(Mitochondrial dysfunction) 10
3-4. Ab蛋白質的沉積 10
4. 治療方法 11
4-1. 抗氧化劑(Antioxidants) 11
4-3. 神經營養因子與抗细胞凋亡因子(Neurotrophic and antiapoptotic factors) 12
4-4. 疫苗注射(Vaccination therapy) 12
4-5. 蛋白酶抑制劑(b-and g-secretase inhibitors) 12
4-6. 降膽固醇藥物(Cholesterol-lowering drugs) 12
4-7. 鋅/銅金屬離子螯合劑(Copper/zinc chelator) 12
4-8. 非類固醇抗發炎藥物(Nonsteroidal anti-inflammatory drugs) 12
參、 葡萄科植物簡介 13
1. 台灣原生葡萄科植物 13
1-1. 細本葡萄(Vitis thunbergii Sieb. & Zucc. var. adstricta (Hance) Gagnep) 14
1-2. 小葉葡萄組織培養苗 15
1-3. 臺灣原生種葡萄屬相關命名 15
2. 葡萄屬相關研究 15
肆、 興奮性神經傳導物質(Excitatory neurotransmitters) v.s.神經退化性疾病 17
實驗材料與方法(Materials and methods) 25
壹、 實驗材料 (Materials) 25
貳、 小葉葡萄萃取液之製備 (Preparation of VTT extracts) 27
參、 細胞培養 (Cell culture) 27
肆、 西方墨點法 (Western blotting) 27
伍、 細胞存活率測試 (MTT assay) 28
陸、 亞硝酸鹽含量之測定 (Determination of nitrite level) 28
柒、 過氧化氫產生之測定 (Determination of hydrogen peroxide production) 28
捌、 動物與處理 (Animals and treatments) 28
玖、 神經病理學分析 (Neuropathological analysis) 29
壹拾、統計分析 (Statistical analysis) 29
附 錄 30
溶液及試劑配方 30
實驗結果(Results) 32
壹、 使用小葉葡萄 ( VTT ) 4個部位萃取物(根、支幹、葉、主幹)於發炎的微小膠質細胞之結果 32
貳、 小葉葡萄四種萃取物(根、支幹、葉、主幹)作用於KA誘發出的過氧化物生成以及初神經細胞死亡的結果 33
參、 小葉葡萄四種萃取物對於阻止KA誘發老鼠腦部病理性變化的結果 33
討論(Discussion) 35
壹、 在in vitro及in vivo的實驗四種小葉葡萄的萃取物中,支幹與葉的萃取物是最有效的 35
貳、 在vitro及vivo的實驗四種小葉葡萄的萃取物中,支幹與葉的萃取物可預防KA造成的神經細胞壞死 36
參、 KA在老鼠實驗中因不同品種及劑量其臨床表現不同,但都會造成海馬迴CA3區的損害 36
結論 37
參考文獻(References) 38
結果圖表 44
Figure 1 小葉葡萄的萃取物在BV2微小膠細胞中對LPS誘導iNOS產生及NO產生的影響。 45
Figure 2 小葉葡萄的萃取物在BV2微小膠細胞中對LPS誘導iNOS產生及NO產生的影響。 49
Figure 3 小葉葡萄的萃取物在大鼠神經細胞中對KA誘導ROS產生的影響。 51
Figure4 小葉葡萄的萃取物在大鼠神經細胞中對KA誘導細胞凋亡的影響。 53
Figure 5 海馬迴經Nissl’s 染色之代表性染色圖。 55
Table 1小葉葡萄對於由KA誘發小鼠的抽蓄、死亡及病理上變化的結果。 56

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系統識別號 U0007-2207200911250400
論文名稱(中文) 結合觸感技術設計互動式電腦學習系統 -以牙科脫蠟鑄造噴火槍融熔鈀銀合金為例
論文名稱(英文) Using Haptic Technology to Design Computer Assisted Learning Systems for Dental Casting Training – In the Case of melting palladium silver alloy with a dental lost-wax casting blow torch
校院名稱 臺北醫學大學
系所名稱(中) 醫學資訊研究所
系所名稱(英) Graduate Institute of Biomedical Informatics
學年度 97
學期 2
出版年 98
研究生(中文) 楊承益
學號 M110096012
學位類別 碩士
語文別 中文
口試日期 2009-06-23
論文頁數 95頁
口試委員 指導教授-劉建財
委員-徐建業
委員-邱泓文
委員-黃衍文
委員-詹前隆
關鍵字(中) 牙科鑄造
電腦輔助教學
觸感科技
Nintendo Wii Remote
關鍵字(英) Dental Casting
CAI
Haptic Technologies
Nintendo Wii Remote
學科別分類
中文摘要 牙科鑄造學(Dental Casting)對於牙體技術學系學生及牙體技術師而言是非常重要的一門科目,無論是在小型到大型的假牙修復物,都需要使用鑄造技術。目前牙科修復物主要之鑄造方式仍以脫蠟鑄造(Dental Lost-Wax Casting technique)為主,牙科脫蠟鑄造最常利用噴火槍(blow torch或稱blow pipe)融熔合金,但調整噴火槍並利用噴火槍熔融金屬之過程相當複雜、危險與困難,需透過不斷的練習以熟悉調整方式,但初學者無法獨自使用這些高危險性的鑄造器械練習,需有鑄造專業知識的教師陪伴,否則將造成鑄造失敗損失成本,更可能釀成嚴重公安意外。
為了提供一個讓初學者安全學習的環境、減少學習訓練耗材成本,並且能夠在操作練習時給予即時的指導,降低教學者的教學負擔,並記錄學習者的操作過程供教學者與學習者有效掌握學習效果,因此本研究建立一個電腦輔助教學系統(Computer Assisted Instruction ; CAI),透過Flash製作各項視覺化的模擬元件及模擬環境,並使用觸感(Haptic)技術操控模擬環境,藉此模擬噴火槍操作方式,讓使用者透過此系統的使用,熟悉噴火槍火焰調整與融熔金屬的方式,再透過即使語音指導功能的建置,讓學習者獲得系統即時的教導。系統為了有效模擬噴火槍的操作方式與噴火槍進行金屬熔融所產生的現象,遂採用任天堂公司所開發的Nintendo Wii Remote 觸感(Haptic)無線互動裝置進行噴火槍的模擬,再將Haptic裝置的操作訊號透過藍芽傳輸的方式傳入電腦主機,進行操作CAI系統中的模擬環境。
本雛形系統發展出帳號管理、教學指導、自我訓練及報表模組,並由後端Tutoring資料庫支援教學指導及自我訓練模組的操作方式及規則,在此二模組中的訓練方是即使用Haptic裝置模擬噴火槍調整與金屬融熔的操做。Log資料庫記錄學習者操作系統之軌跡,其操作軌跡包括使用者登入、登出、操作時間、操作日期、選擇之學習模組、操作過程中所發生的錯誤事件等等資訊,最後提供報表模組讓使用者及教學者能夠掌握學習狀況,讓學習者了解自我學習成效,也能讓教學者針對學習者之弱點進行強化。本系統之建立不僅能有效模擬噴火槍調整,其直覺性操作、有效監控學習過程以及即時教導功能的鑄造輔助教學環境,不僅能提供學生自我學習、減低教學成本、提昇操作安全、還能有效掌握學生學習狀態並降低教師教學負擔。
英文摘要 Dental casting is a very important discipline for students of dental technology and dental technicians. Small denture fixtures, e.g. inlays, onlays, crowns, endodontic posts and large denture bases all require the casting technique. In the dental field, lost-wax casting technique is one of the favorite methods to fabricate denture for dental technicians.
The lost-wax dental casting process use high temperature to melt metal alloys. The most frequently used instrument in the dental lost-wax casting technique to melt metal alloys for dental technicians is the blow torch (or called blow pipe). However, adjustment of the blow pipe is dangerous and difficult to a certain degree and hence requires repeated practice to do it well. Beginners cannot practice by themselves and need guidance from a teacher. Otherwise it will result in casting failure and cost losses. In a worst scenario, it may also lead to a serious public safety accident.
This study uses the computer assisted instruction (CAI) with the visual interface and Wii remote haptic wireless interactive device developed by Nintendo to effectively simulate blow pipe adjustment and establish an assisted casting training environment that is authentic, intuitive, and can effectively monitor the learning process for students to learn spontaneously, cut down the teaching cost, enhance operation safety, and reduce the teacher’s burden.
論文目次 標題 i
審定書 ii
誌謝 iii
目錄 v
表目錄 vii
圖目錄 viii
公式目錄 ix
中文摘要 x
英文摘要 xiii
第一章 緒論
1.1研究背景 1
1.2研究動機 4
1.3研究目的 6
第二章 文獻探討
2.1 牙科鑄造(Dental Casting) 7
2.1.1牙科脫蠟鑄造(Dental Lose-Wax Casting) 7
2.1.2脫蠟鑄造金屬熔融(Alloy Melting) 8
2.1.3牙科用鈀-銀合金(Palladium-Silver Alloy) 12
2.2電腦輔助教學(Computer Assisted Instruction ; CAI) 14
2.2.1 CAI定義 14
2.2.2 CAI源起 15
2.2.3 CAI分類 17
2.2.3.1 Tutorial 18
2.2.3.2 Drill and Practice 19
2.2.3.3 Simulation 20
2.2.3.4 Game 22
2.2.3.5 Problem Solving 22
2.2.4 CAI於生物醫學應用之分類 23
2.2.5 CAI於學術上之研究 24
2.3觸感科技(Haptic Technologies) 26
2.3.1 Haptic定義 26
2.3.2 Haptic生理學之運作 27
2.3.3 Haptic相關之研究 30
2.4 Nintendo Wii Remote 32
第三章 研究方法與材料
3.1牙科脫蠟鑄造熔融金屬之臨床操作簡介 38
3.2牙科鑄造金屬熔融模擬教學系統功能與規格 41
3.2.1牙科鑄造金屬熔融模擬教學系統功能 41
3.2.2牙科鑄造金屬熔融模擬教學系統規格 42
3.3牙科鑄造金屬熔融模擬教學系統之軟硬體規劃 43
3.3.1牙科鑄造金屬熔融模擬教學系統軟體規畫 43
3.3.2牙科鑄造金屬熔融模擬教學系統硬體規劃 49
3.4軟體開發工具 52
3.4.1牙科鑄造金屬熔融模擬教學系統工作環境建構 52
3.5牙科鑄造金屬熔融模擬教學系統製作 54
3.6學習模組音效製作 59
第四章 研究結果
4.1 系統功能介面與使用 62
4.1.1 硬體操作介面 62
4.1.2 軟體操作介面 63
4.2系統操作個案情境 78
第五章 結果與討論
5.1研究結果 82
5.2討論與未來展望 86
第六章 參考文獻
6.1參考文獻 90
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系統識別號 U0007-2207201016525700
論文名稱(中文) 兩種真菌醱酵液之生物活性成分研究
論文名稱(英文) Chemical investigation of the fermented broths of Theissenia rogersii and Stilbohypoxylon elaeicola
校院名稱 臺北醫學大學
系所名稱(中) 生藥學研究所
系所名稱(英) Graduate Institute of Pharmacognosy
學年度 98
學期 2
出版年 99
研究生(中文) 張雅雯
學號 M303097009
學位類別 碩士
語文別 中文
口試日期 2010-06-24
論文頁數 181頁
口試委員 委員-李水盛
委員-蕭哲志
指導教授-李宗徽
關鍵字(中) 真菌
誘導型一氧化氮合成酶
Theissenia rogersii
Stilbohypoxylon elaeicola
elaeicolaside
elaeicolactone
關鍵字(英) isopimarane diterpene glycoside
elaeicolaside
elaeicolactone
Theissenia rogersii
Stilbohypoxylon elaeicola
iNOS
學科別分類
中文摘要 利用抑制一氧化氮合成酶的活性分析平台來篩選真菌培養株,藉以篩檢出具有抗發炎活性的真菌株,發現某些真菌株的醱酵培養液或菌絲體萃取物,對於RAW264.7細胞的產生一氧化氮具有顯著的抑制作用。遂選定:Theissenia rogersii(#92031201)及Stilbohypoxylon elaeicola(#173)兩株進行其活性成分研究。在選擇以麥芽抽取物(malt extract)為培養基加以擴大培養後,針對醱酵液所含代謝產物進行分析與分離,計獲得12個化合物,分別為: 2,5-dihydroxymethylfuran(1)、pycnidione(2)、maltol(3)、tyrosol(4)、hymatoxin K(5)、hymatoxin L(6)、elaeicolactone(7)、5-hydroxymethylfurfural(8)、elaeicolasdie A(9)elaeicolaside B(10)、elaeicolaside C(11)和 elaeicolaside D(12),其中屬於倍半萜之化合物7,及屬於二萜糖苷的化合物9、10及12,皆為新化合物。在100 μM的濃度下,10對於RAW264.7細胞的產生一氧化氮具有顯著的抑制活性且不具細胞毒性。此外,2及10對於A549肺腺癌細胞具有強的毒殺作用,其GI50分別為5.25 nM及9.79 μM。
英文摘要 The ethyl acetate extracts of the fermented broths of Theissenia rogersii (#92031201) and Stilbohypoxylon elaeicola (#173) were found to exhibit significant activity against inducible nitric oxide synthase (iNOS) in our preliminary screening. Therefore, bioassay-guided fraction and separation of the active component from these two broths were carried out which resulted in the isolation of twelve compounds. Their structures were elucidated to be 2,5-dihydroxymethylfuran (1), pycnidione (2), maltol (3), tyrosol (4), hymatoxin K (5), hymatoxin L (6), elaeicolactone (7), 5-hydroxymethylfurfural (8), elaeicolasdie A (9), elaeicolaside B (10), elaeicolaside C (11) and elaeicolaside D (12). Of these compounds identified, 7, 9, 10 and 12, classified as respective a sesquiterpene and three diterpene glycosides, were novel chemical entities. Compound 10 exhibited significant inhibitory activity on NO production of RAW264.7 cells without any cytotoxicity. In addition, 2 and 10 exerted potent cytotoxicity against an A549 lung cancer cell line with GI50 values of 5.25 nM and 9.79 μM, respectively.
論文目次 摘要 I
Abstract II
總目錄 III
圖目錄 VII
縮寫表 XII
壹、緒論與研究目的 1
貳、炭角菌科真菌之天然物文獻回顧 5
参、實驗結果與討論 34
3.1.1 2,5-Dihydroxymethylfuran(1)之結構解析 39
3.1.2 Pycnidione(2)之結構解析 43
3.1.3 Maltol(3)之結構解析 54
3.1.4 Tyrosol(4)之結構解析 58
3.1.5 Hymatoxin K(5)之結構解析 61
3.1.6 Hymatoxin L(6)之結構解析 73
3.1.7 Elaeicolactone(7)之結構解析 82
3.1.8 5-Hydorxymethylfurfural(8)之結構解析 92
3.1.9 Elaeicolaside A(9)之結構解析 96
3.1.10 Elaeicolaside B(10)之結構解析 106
3.1.11 Elaeicolaside C(11)之結構解析 116
3.1.12 Elaeicolaside D(12)之結構解析 126
3.2 一氧化氮濃度及細胞毒性測定結果 135
3.3 A549細胞生長抑制測試結果 137
3.4 討論 138
肆、實驗部分 141
4.1 儀器設備與試劑 141
4.2 真菌材料 143
4.3 培養基配置 143
4.4.1 #92031201 培養液之成分分離與純化 145
4.4.2 #173培養液之成分分離與純化 148
4.5 單糖組成分析 152
4.6 單糖旋光度測定 153
4.7 一氧化氮(NO)濃度之測定:Griess reagent assay 154
4.8 細胞毒性測試(alamarBlue assay) 157
4.9 細胞生長測試(sulforhodamine B, SRB) 159
4.10 各成分之物理數據 161
參考文獻 165
附錄一 173
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系統識別號 U0007-2501200713254600
論文名稱(中文) 第二型糖尿病患之白袍效應與相關因子
論文名稱(英文) Correlates of White-Coat Effect in Patients with Type II Diabetes.
校院名稱 臺北醫學大學
系所名稱(中) 護理學研究所
系所名稱(英) Graduate Institute of Nursing
學年度 95
學期 1
出版年 96
研究生(中文) 楊惠如
學號 G455092004
學位類別 碩士
語文別 中文
口試日期 2007-01-18
論文頁數 118頁
口試委員 指導教授-蔡佩珊
委員-曹麗英
委員-蘇千田
關鍵字(中) 第二型糖尿病
白袍效應
自主神經功能
心率變異性
關鍵字(英) Type II Diabetes
White-coat effect
Autonomic function
Heart rate variability
學科別分類
中文摘要 論 文 摘 要
論文名稱:第二型糖尿病患之白袍效應與相關因子
研究所名稱:臺北醫學大學護理學研究所
研究生姓名:楊惠如
畢業時間:九十五 學年度 第一學期
指導教授:蔡佩珊 臺北醫學大學護理學研究所 副教授

本研究目的旨在探討第二型糖尿病患之白袍效應與相關因子,及心率變異性測量對於第二型糖尿病患發生白袍效應的預測能力。於北台灣北部某一門診收案,受試者包括34位(23到70歲)規律服用降血糖藥物且未接受過高血壓藥物治療的第二型糖尿病個案。研究變項包括飯前血糖、糖化血色素、總膽固醇、三酸甘油酯數值及體位測量,並在晨間安排心率變異性、三項心血管自主神經功能測試與握力壓力測試值;透過動態血壓測量儀(SpaceLabs 90207, Redmond, WA)取得臨床血壓與居家血壓值,以BioPacMP100訊號擷取系統分析心跳間距與心率變異頻譜,由連續性血壓測量儀(Finometer, TNO Biomedical Instrumentation, Amsterdam, Netherlands)取得心血管參數,所有資料以SPSS 11.0套裝統計軟體進行分析。以臨床收縮壓值減去ABPM日間收縮壓差值分為三組,大於等於5 mmHg為白袍效應組, 小於5 mmHg但大於-5 mmHg為無白袍效應組,小於等於-5 mmHg為反向白袍效應組。
依據本研究對白袍效應之定義,白袍效應組佔38.2 %,無白袍效應組佔32.4 %,反向白袍效應組佔29.4 %。三組間在人口社會學及生化因子之比較以ANOVA分析並以LSD作事後比較,結果顯示:白袍效應組之糖尿病史顯著較他組長;反向白袍效應組比白袍效應組之三酸甘油酯顯著較高;臨床收縮壓值以白袍效應組較反向白袍效應組高;而日夜間血壓差值則以反向白袍效應組較白袍效應組高;無白袍效應組在心率變異性高頻功率與總功率項皆比其他兩顯較高。經線性迴歸檢定臨床收縮壓值、日夜間血壓差值及心率變異性總功率可獨立預測第二型糖尿病患之白袍效應,三者對於白袍效應之變異量的解釋力為39.7 % (p = .001)。
本研究確認了第二型糖尿病患發生白袍效應的相關因素,且證實心率變異性對於第二型糖尿病患發生白袍效應是具有預測性的。



關鍵字:第二型糖尿病、白袍效應、自主神經功能、心率變異性
英文摘要 Abstract

Title of Thesis: Correlates of White-Coat Effect in Patients with Type II Diabetes.
Institution: Graduate Institute of Nursing, Taipei Medical University
Author: Huei-Ju Yang
Thesis directed by: Pei-Shan Tsai, Ph.D., Associate Professor

The purpose of this study was to examine (1) the correlates of white-coat effect in patients with type II diabetes mellitus and (2) the predictive power of heart rate variability (HRV) in predicting the size of the white coat-effect in type II diabetics.
Thirty-four male and female type II diabetics, aged 20 to 70, who had never been treated with antihypertensive medications, were recruited from the Outpatient Department of a hospital located in northern Taiwan. The biochemical variables included fasting blood sugar, HbA1C, cholesterol, and triglyceride. Resting HRV, tests of cardiovascular autonomic neuropathy, and handgrip stress test were employed in a morning session. Clinic blood pressure (BP) was measured by an automatic BP recorder (SpaceLabs 90207, Redmond, WA). Ambulatory BP monitoring (APBM) was carried out using the SpaceLabs BP monitor during two 24-hour periods in a day representing the participant’s typical day of the week. White coat effect (WCE) was calculated by subtracting the ABPM daytime systolic BP (SBP) from the clinic SBP. Participants were grouped according to the size of their WCE. Those who showed a WCE of 5mmHg and above were assigned to the WCE group; those who showed a WCE of between -5 and 5 mmHg were assigned to the no white coat effect (NWCE) group; those who exhibited a WCE of -5mmHg and lower were assigned to the reverse white coat effect (RWCE) group.
Based on the grouping criteria, 38.2 % of the participants were categorized as WCE, 32.4 % as NWCE and 29.4 % as RWCE. A comparison of socio-demographic and biological variables among three groups with ANOVA and post hoc LSD revealed that: the WCE group had significantly longer DM history than other groups; the RWCE group had a significantly higher triglyceride level than the WCE group; the WCE had significantly higher clinic BP than the RWCE group; the NWCE group had higher day-night SBP difference than the WCE group; the NWCE group had significantly higher HRV in high frequency and total power than other groups. The linear regression model showed that clinic SBP, day-night SBP difference, and HRV TP independently predict the size of the WCE in type II diabetics (R2 = 39.7 %, p = .001).
This study identified correlates of the WCE and demonstrated that HRV is useful in predicting the size of the WCE in type II diabetics.






Key words: Type II Diabetes, White-coat effect, Autonomic function, Heart rate variability
論文目次 目 錄
頁 數
致 謝 ……………………………………………………………. Ⅰ
中文摘要 ……………………………………………………………. Ⅲ
英文摘要 ……………………………………………………………. Ⅴ
目 錄 ……………………………………………………………. Ⅶ
圖表目次 ……………………………………………………………. Ⅸ
第一章 緒論
第一節 研究動機與重要性 ………………………………………1
第二節 研究目的 …………………………………………………9
第三節 名詞界定 …………………………………………………10
第二章 文獻查證
第一節 第二型糖尿病 ……………………………………………12
第二節 白袍高血壓 ………………………………………………23
第三節 白袍效應 …………………………………………………30
第四節 自主神經功能 ……………………………………………37
第三章 研究方法
第一節 研究設計 …………………………………………………52
第二節 研究樣本 …………………………………………………52
第三節 研究工具 …………………………………………………54
第四節 研究流程 …………………………………………………56
第五節 統計分析方法 …………………………………………………65
第四章 分析與結果
第一節 個案基本屬性 ……………………………………………65
第二節 動態血壓測量儀測量值分析 ……………………………68
第三節 心率變異性 ………………………………………………71
第四節 標準三項心血管自主神經功能測試 ……………………73
第五節 握力壓力測試 ……………………………………………75
第五章 討論
第一節 具白袍效應第二型糖尿病患之特質 ……………………78
第二節 心率變異性對白袍效應的預測性 ………………………81
第六章 結論與建議 ……………………………………………………83
參考資料
中文部份 …………………………………………………………… 86
英文部份 …………………………………………………………… 87



圖 表 目 次

圖一 收案流程圖 …………………………………………………………64
表一 第二型糖尿病患有白袍效應(WCE)、無白袍效應(NWCE)以及反向白袍效應(RWCE)三組個案人口學、 體位測量及生化檢驗資料 …………………………………………………………………67
表二 第二型糖尿病WCE、NWCE以及RWCE三組,臨床血壓與動態血壓之比較 ………………………………………………………………70
表三 第二型糖尿病患WCE、NWCE以及RWCE三組,於心率變異性之比較 …………………………………………………………………72
表四 第二型糖尿病患WCE、NWCE以及RWCE三組,三項標準心血管自主神經功能指標之比較 …………………………………………74
表五 第二型糖尿病患WCE、NWCE及RWCE三組,於握力壓力前與測試當中,心血管參數差值(壓力反應性)之比較 …………………76
表六 第二型糖尿病患發生白袍效應的預測因子 ……………………….77
參考文獻 參考文獻
中文部份
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陳國群(1979)•161例糖尿病患之腦血管意外及心肌梗塞•中華醫誌,26,260-271。
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蔡孟書、吳英黛、詹曉龍、賴金鑫、戴東原(2003)•糖尿病患者休息時心率變異性的影響因素•臺灣醫學會雜誌,7(1),10-18。




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系統識別號 U0007-2507200909405400
論文名稱(中文) 血管收縮素受體阻斷劑對糖尿病或高血壓病人尿中D-乳酸濃度之影響
論文名稱(英文) The Effect of AngiotensinⅡType 1 Receptor Blocker on Urinary D-Lactate Concentrations in Diabetic or Hypertensive Patients
校院名稱 臺北醫學大學
系所名稱(中) 藥學研究所
系所名稱(英) Graduate Institute of Pharmacy
學年度 97
學期 2
出版年 98
研究生(中文) 陳欣平
學號 M301096006
學位類別 碩士
語文別 中文
口試日期 2009-07-06
論文頁數 126頁
口試委員 指導教授-李仁愛
委員-蔡東湖
委員-黃偉展
關鍵字(中) d-乳酸
微白蛋白尿
糖尿病腎病變
高血壓腎病變
血管收縮素受體阻斷劑
關鍵字(英) D-Lactate
NAG
microalbuminuria
diabetic nephropathy
hypertensive nephropathy
angiotensin II type 1 receptor blocker
ARB
valsartan
學科別分類
中文摘要 根據我們之前的動物實驗結果,尿中D-乳酸濃度(μM/mM creatinine)可能可以作為偵測早期腎臟損傷之指標;人體實驗結果,糖尿病與高血壓病患尿中D-乳酸濃度在腎臟損傷之生化指標微白蛋白尿(microalbuminuria)尚未出現前,即顯著高於正常人(P<0.05)。因此,本研究進一步探討:糖尿病與高血壓病患服用具有腎保護功能及降低蛋白尿之Angiotensin Ⅱ receptor blocker—valsartan,對病患尿中D-乳酸、N-acetyl-β-D-glucosaminidase(NAG)及microalbumin濃度之影響,以及尿中D-乳酸與腎臟病變之相關性。
本試驗採收39位健康成年人之尿液做為對照組,並且將糖尿病(n = 74)與高血壓(n = 81)患者分別分為三組:未服用valsartan,服用valsartan 12週,以及服用valsartan 24週。患者服用valsartan之劑量依照醫師之診斷,其範圍為40-120 mg/day。結果顯示,未服用valsartan之糖尿病患者尿中D-乳酸濃度(31.69 ± 8.83 μM/mM cr)皆顯著高於服用valsartan12週(9.08 ± 2.12 μM/mM cr)或服用valsartan 24週(4.03 ± 0.90 μM/mM cr)之患者;未服用valsartan之高血壓患者尿中D-乳酸濃度(12.57 ± 3.85 μM/mM cr)皆顯著高於服用valsartan12週(8.85 ± 1.50 μM/mM cr)或服用valsartan 24週(3.73 ± 1.11 μM/mM cr)之患者。值得注意的是,糖尿病或高血壓患者尿中D-乳酸、NAG及microalbumin濃度下降之趨勢與病患服用valsartan週數有明顯正相關性,而且服用valsartan 24週之糖尿病或高血壓患者,其尿中D-乳酸、NAG及microalbumin濃度與正常人比較皆無統計之差異。
綜合上述,糖尿病患者尿中D-乳酸與微白蛋白尿具有相同之特性—確實會因患者服用valsartan之影響而下降,而且在微白蛋白尿出現之前,尿中D-乳酸濃度即顯著升高。因此,我們建議可以偵測尿中D-乳酸濃度作為早期腎臟損傷之指標。
英文摘要 According to our previous research, there was a significant increase in urine levels of D-lactate (μM/mM creatinine) in diabetic rats compared to normal rats (P < 0.01). We further demonstrated the urine levels of D-lactate in diabetic or hypertensive patients were prominently elevated before the appearance of microalbuminuria (P < 0.05). In this study, we investigated an angiotensin II receptor blocker, valsartan, with proven effect on microalbuminuria on the clinical effects of urinary D-lactate concentrations in diabetic or hypertensive patients.
We studied three groups in diabetic or hypertensive patients who were not receiving valsartan, receiving 12-week, and 24-week valsartan. Valsartan dosage ranged from 40-120 mg/day according to doctor’s diagnosis. Urine levels of D-lactate, NAG (N-acetyl-β-D-glucosaminidase), and microalbumin were measured in 74 diabetic patients, 81 hypertensive patients, and 39 healthy subjects as controls.
In diabetic patients, the levels of urinary D-lactate were significantly lower in patients receiving either 24-week (4.03 ± 0.90 μM/mM cr) or 12-week (9.08 ± 2.12 μM/mM cr) valsartan compared with those not receiving valsartan (31.69 ± 8.83 μM/mM cr). In hypertensive patients, the levels of urinary D-lactate were significantly lower in patients receiving either 24-week (3.73 ± 1.11 μM/mM cr) or 12-week (8.85 ± 1.50 μM/mM cr) valsartan compared with those not receiving valsartan (12.57 ± 3.85 μM/mM cr). The descending tendency of urinary D-lactate, NAG, and microalbumin all had a positive relationship with the duration of patients receiving valsartan. Notably, there were no difference in urine levels of D-lactate, NAG, and microalbumin between 24-week valsartan group of diabetic or hypertensive patients and healthy subjects.
In conclusion, the significant reduction of urinary D-lactate was similar with the proven effect on microalbuminuria by valsartan in diabetic or hypertensive patients. We suggest that urinary D-lactate may be a useful indicator for early diagnosis of diabetic or hypertensive nephropathy, serving in the assessment of therapeutic effects.
論文目次 目錄 I
中文摘要 VII
Abstract VIII
表目錄 IX
圖目錄 X
縮寫表 XII
第一章 緒論 1
壹、 前言 1
貳、 研究背景 5
一、D-Lactate 5
1. D-Lactate簡介 6
2. D-Lactate來源 7
2.1 腸胃道的製造 7
2.2 飲食攝取或服藥時吸收 7
2.3 甲基乙二醛(Methylglyoxal ; MG) 8
3. D-Lactate藥物動力學 10
3.1 D-Lactate之吸收 10
3.2 D-Lactate之代謝 12
3.3 D-Lactate之排泄 12
4. D-Lactate相關疾病 13
4.1 D-乳酸中毒(D-Lactic acidosis) 13
4.2 短腸症候群(short-bowel syndrome;SBS) 14
4.3 腹瀉 15
4.4 神經毒性(Neurotoxicity) 15
4.5 抑制記憶力 16
5. 尿中D-lactate與糖尿病或高血壓之相關性 17
5.1 糖尿病 17
5.2 高血壓 20
二、Methylglyoxal(MG) 23
1. MG生成途徑 23
2. MG病理特性 25
3. MG相關疾病研究 27
3.1 胰島素阻抗症(Insulin resistance) 27
3.2 高血壓 28
三、Advanced glycation end-products(AGEs) 31
1. AGEs生成途徑 31
2. AGEs病理特性 34
四、Angiotensin II type 1 receptor blocker(ARB) 37
1. Renin-Angiotensin System(RAS)作用機轉 37
2. ARB之作用機轉 40
3. ARB之臨床實證 42
4. ARB對腎臟之影響 44
5. ARB之藥物交互作用 45
6. Valsartan 46
6.1 Valsartan作用機轉 46
6.2 Valsartan劑量 46
6.3 Valsartan之藥物動力學 47
6.4 Valsartan之副作用 48
参、研究目的 49
第二章、實驗材料與方法 50
壹、實驗儀器及材料 50
一、實驗試藥 50
二、實驗儀器 51
貳、分析方法 53
一、D-Lactate分析方法 53
1. 分析條件 54
1.1 生物檢品中(D+L)-Lactate之單離 56
1.2 D-Lactate與L-Lactate之chiral separation 57
2. 尿液檢品之製備 57
3. 檢量線配製方法 58
4. 螢光衍生化方法 58
5. 分析方法再確效 59
5.1 檢量線線性(Linearity) 59
5.2 準確度試驗(Accuracy) 60
5.3 精密度試驗(Precision) 60
二、N-acetyl-β-D-glucosaminidase(NAG)分析方法 61
1. 分析條件 62
2. 尿液檢品之製備 62
3. 檢量線配製方法 62
三、Creatinine分析方法 63
1. 分析條件 63
2. 尿液檢品之製備 64
3. 檢量線配製方法 64
参、人體試驗與研究設計 65
一、 受試對象與納入標準 65
二、 試驗設計與試驗流程 67
1. ARB藥物Valsartan來源 67
2. 試驗設計 67
3. 試驗流程 67
4. 分析項目 67
肆、 實驗數據處理 68
伍、 統計方法 69
第三章、實驗結果與討論 70
壹、D-Lactate分析方法再確效 70
一、 檢量線線性(Linearity) 70
二、 準確度試驗(Accuracy) 71
三、 精密度試驗(Precision) 71
參、 人體實驗結果 74
一、 糖尿病患者 74
1. 未服用valsartan之糖尿病患者尿中D-lactate、NAG及microalbumin濃度分析 74
2. 服用valsartan對糖尿病患者尿中D-lactate濃度之影響... 75
3. 服用valsartan對糖尿病患者尿中NAG濃度之影響 75
4. 服用valsartan對糖尿病患者尿中microalbumin濃度之影響 76
5. 糖尿病患者尿中D-lactate,NAG及microalbumin濃度變化與服用valsartan之相關性 76
二、 高血壓患者 79
1. 未服用valsartan之高血壓患者尿中D-lactate、NAG及microalbumin濃度分析 79
2. 服用valsartan對高血壓患者尿中D-lactate濃度之影響... 80
3. 服用valsartan對高血壓患者尿中NAG濃度之影響 80
4. 服用valsartan對高血壓患者尿中microalbumin濃度之影響 81
5. 高血壓患者尿中D-lactate,NAG及microalbumin濃度變化與服用valsartan之相關性 81
参、討論 84
一、 ARB(valsartan)降低糖尿病或高血壓患者尿中 D-lactate之可能作用機轉 85
二、 Microalbumin與ARB(valsartan)之相關性 93
三、 NAG與ARB(valsartan)之相關性 100
第六章 結論及未來方向 102
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系統識別號 U0007-2607201017454300
論文名稱(中文) 丹參酚酸B 對於纖維蛋白原之作用機轉討論
論文名稱(英文) To study Salvianolic acid B on the fibrinogen level for HepG2-TRα1 and the hemorheology parameter for T3 rats.
校院名稱 臺北醫學大學
系所名稱(中) 生醫材料暨工程研究所
系所名稱(英) Institute of Biomedicac Materials Engineering
學年度 98
學期 2
出版年 99
研究生(中文) 林怡君
學號 M225097017
學位類別 碩士
語文別 中文
口試日期 2010-07-01
論文頁數 76頁
口試委員 指導教授-劉得任
委員-黃義侑
委員-侯文琪
關鍵字(中) 三碘甲狀腺素
纖維蛋白原
丹參酚酸B
過量表現甲狀腺素受體的肝癌細胞株
關鍵字(英) Triiodothyronine (T3)
Fibrinogen
Salvianolic acid B (Sal B)
HepG2-TRα1
學科別分類
中文摘要 甲狀腺素 (Triiodothyronine, T3)是一種可調控細胞生長、發育與分化之重要因子。然而,過量的甲狀腺素會造成血液中纖維蛋白原 (fibrinogen) 上升,促使紅血球聚集度增加,血液黏度提高,造成血液流變異常。丹參 (Salvia miltiorrhiza Bunge) 是一種具有活血化瘀功能的中草藥,被廣泛應用於治療心血管疾病。丹參酚酸B (Salvianolic acid B,Sab) 是丹參水溶性有效成分,因此透過探討丹參酚酸B 對大鼠血液中纖維蛋白原的影響。
利用T3刺激HepG2-TRα1細胞,使其生成纖維蛋白原,再利用不同濃度10、30、50、80、100 μM 丹參酚酸B處理,由Q-PCR及ELISA分別得知纖維蛋白原mRNA及protein生成量,由實驗結果得,以30 μM以上之丹參酚酸B處理,纖維蛋白原mRNA生成量會開始明顯降低;而纖維蛋白原protein層面,則是50 μM以上的丹參酚酸B,可降低纖維蛋白原生成量,因此可推論:丹參酚酸B可抑制纖維蛋白原mRNA生成。
本實驗使用八週齡之Sprague-Dawley 大鼠,給予T4飲用水,於第六週開始,每天分別餵食30、50、80mg/(kg day)丹參酚酸B,持續六週。並抽取其血液檢測血液流變之參數。結果顯示,餵食50、80mg/(kg day)丹參酚酸B組於第四、六週之纖維蛋白原含量較T4組低;於第六週時,紅血球聚集度較T4組低;於低shear rate下,三組餵食丹參酚酸B組相較於T4組其黏度皆有降低之趨勢。纖維蛋白降解產物(FDP)也無明顯差異。由此可知:丹參酚酸B具有降低纖維蛋白原生成之功效。
英文摘要 The thyroid hormone, 3, 3', 5-triiodo-L-thyronine (T3) regulates growth, development and differentiation processes in animals. Nevertheless, excessive T3 will raise the fibrinogen level in the blood established leading to the erythrocyte aggregation increasingly, and enhance the whole blood viscosity, creating the anomaly in hemorheological.
Salvia miltiorrhiza Bunge is widely used for the treatment of atherosclerosis-related disorders and known as Danshen in Chinese traditional medicine is effective in promoting circulation and decreasing stasis in the blood. Salvianolic acid B (Sab) is a potent water-soluble from the roots of Salvia miltiorrhiza Bunge. Therefore, to explore the effects of Sab on the fibrinogen mRNA level in the blood for Rats.
We first adapt HepG2-TRα1 cell culture and T4 rat animal model in this preliminary study.To study the fibrinogen mechanism after T3 treatment in TRa-overexpressed hepatoma cell line (HepG2-TRa1) and then activate HepG2-TRa1 with 10、30、50、80、100 μM Sab. In addition to fibrinogen mRNA levels evaluate by Q-PCR, the fibrinogen protein was evaluate by ELISA. According to the result , the fibrinogen mRNA levels were decrease after the treatment of 30 μM Sab, and in the fibrinogen protein levels, it is significant decrease in the 50 μM Sab. The results indicate that Sab can inhibite the release of fibrinogen mRNA.
In the in vivo study, we selected the 8-week-old Sprague-Dawley rat as the animal experimental models and fed them a diet containing L-thyroxine for 6 weeks and then fed them 30、50、80mg/(kg day) of Sab for 6 weeks, respectively, in order to evaluate the hemorheological parameters.The results showed that the fibrinogen levels of the orally given 50、80mg/(kg day) of Sab groups are lower than the T4 group at fourth and sixth week. At sixth week, the same groups showed that the erythrocyte aggregation of the 50、80mg/(kg day) of Sab groups are lower compared with the T4 group. At the low shear rate, except that all of the Sab groups showed that the whole blood viscosity compared with the T4 group decreased. The FDP values was not significant in all of the groups. The results indicate:Sab might play an important role in the release of fibrinogen mRNA.
論文目次 摘 要 1
ABSTRACT 2
第一章 緒論 4
第二章 文獻回顧 6
2-1 甲狀腺 (Thyroid) 6
2-1-1 甲狀腺素 (Thyroid Hormone) 6
2-1-2 甲狀腺素受體(Thyroid Hormone Receptors,TR) 8
2-1-3 甲狀腺荷爾蒙反應位元(Thyroid Hormone Response Element, TRE) 9
2-1-4 纖維蛋白原(Fibrinogen) 9
2-1-5 T3與Fibrinogen之關係 10
2-1-6 T3調控纖維蛋白原之分子機制 11
2-2 血液流變 13
2-2-1 血液流變學 13
2-2-2 血液黏度 13
2-2-3 血液流變學參數的檢測原理 14
2-2-4 全血黏度之檢測 15
2-2-5 紅血球聚集度(erythrocyte aggregation)的測量 17
2-2-6 血液流變異常與常見循環疾病之相關係 18
2-2-7 循環系統疾病血液流變學變化 19
2-3 中藥丹參 19
2-3-1 丹參簡介 19
2-3-2 丹參成分 20
2-3-3 丹參相關研究 21
2-3-4 丹參酚酸B (Salvianolic acid B) 22
第三章 實驗動機與目的 23
第四章 材料與方法 24
4-2 儀器設備 24
4-3 實驗材料與試劑 28
4-3-1 一般藥品 28
4-3-2 實驗用藥品 28
4-3-3 細胞培養試劑 29
4-3-4 其他 30
4-3-5 丹參酚酸B 的純化與鑑定 31
4-4 實驗方法 32
4-4-1 細胞實驗(In vitro) 32
4-4-1-4 RT-PCR(reverse transcription-PCR) 37
4-4-2 動物實驗(In vivo) 45
(1) 藥物配製 45
(2) 動物 45
(3) 實驗流程 45
4-4-3 實驗方法 47
4-4-4 統計與分析 51
第五章 結果 52
5-1 細胞實驗 52
5-1-1 T3促進Fibrinogen mRNA 表現量上升 52
5-1-2 丹參酚酸B對HepG2-TRα1細胞存活率分析 53
5-1-3 丹參酚酸B對纖維蛋白原mRNA表現量之影響 54
5-1-4 丹參酚酸B對纖維蛋白原蛋白質表現量之影響 55
5-2 動物實驗各項血液參數值之檢測 56
5-2-1血液中纖維蛋白原及FDP之含量 56
5-2-2紅血球聚集度(Erythrocyte Aggregation Index) 57
5-2-3全血黏度(Whole Blood Viscosity) 58
5-2-4 全血球計數(Complete Blood Count, CBC) 59
第六章 討論 60
6-1三碘甲狀腺素調控下游基因之影響 60
6-2丹參酚酸B對纖維蛋白原之調控 61
6-3四碘甲狀腺素對血液流變參數及血液生化值之影響 62
第七章 結論及未來展望 65
7-1 結論 65
7-2 未來展望 66
第八章 參考文獻 67

圖目錄
【圖2.1】甲狀腺素之迴饋作用 6
【圖2.2】影響血液黏度之因素(血液流變學) 14
【圖2.3】黏度(η) 與剪切率(γ) 的函數關係可見下圖 15
【圖2.4】圓錐/平板原理(cone/plate)黏度計示意圖 16
【圖2.5】全血雷射反向散射強度和時間變化之關係圖 18
【圖4.1】Animal Blood Counter 26
【圖4.2】(左)Laser-assisted optical rotational cell analyser(LORCA) 27
【圖4.3】(右)黏度儀(HAAKE Rotation Rheometer RS-1, Germany) 27
【圖4.4】95% 丹參酚酸B 31
【圖4.5】丹參標準品HPLC分析圖譜 32
【圖4.6】心臟採血工具 48
【圖4.7】心臟採血流程 48
【圖4.8】管餵工具 50
【圖5.1】T3促進Fibrinogen mRNA 表現量上升 52
【圖5.2】丹參酚酸B對HepG2-TRα1之毒性分析 53
【圖5.3】不同濃度丹參對細胞受到T3刺激後,纖維蛋白原之表現量 54
【圖5.4】不同濃度丹參對細胞受到T3刺激後,纖維蛋白原之表現量 55
【圖5.5】血液紅血球聚集程度 57
【圖5.6】Control組、實驗組(T4)及丹參酚酸B組之全血黏度比較 58

表目錄
【表5.1】血液中Fibrinogen及FDP之含量 56
【表5.2】Control組、實驗組(T4)及餵食丹參酚酸B之全血球計數 59
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系統識別號 U0007-2607201112132000
論文名稱(中文) 混合式項目推薦排序演算法於健康知識網站之應用
論文名稱(英文) A Hybrid Item-based Recommendation Ranking Algorithm Applied on a Healthcare Website
校院名稱 臺北醫學大學
系所名稱(中) 醫學資訊研究所
系所名稱(英) Graduate Institute of Biomedical Informatics
學年度 99
學期 2
出版年 100
研究生(中文) 胡少杰
學號 M110098011
學位類別 碩士
語文別 中文
口試日期 2011-06-23
論文頁數 85頁
口試委員 指導教授-劉建財
委員-鄭伯壎
委員-許明暉
關鍵字(中) 協同過濾演算法
可及性
排序
使用者訪問行為
網路日誌
關鍵字(英) Collaborative filtering
Accessibility
Ranking
User Access Pattern
IIS log
學科別分類
中文摘要 International Telecommunication Union (ITU) 2006至2009年間對世界各國網際網路人口使用情形的調查,發現使用人數呈現逐年增長的趨勢。除此之外,網際網路上的資源也漸趨豐富與多樣化,其蓬勃發展亦影響了健康產業,有學者發現民眾在網路上搜尋健康相關資訊的數量持續增加且以指數型成長,可看出網際網路已成為民眾接收健康資訊一重要管道。

上網找尋資料的工作隨著資訊過載變得費時且低效率。為了解決問題,兩種普遍的網際網路存取工具,協助使用者提昇查尋結果的可用性:分別為搜尋引擎與提供目錄服務的入口網站系統。使用者從尋找過程中學習健康知識、提升自我健康管理能力、減低醫療成本的花費、提高民眾對醫療服務的滿意度以及醫病關係的改善。但消費者在網路上查找健康資訊時可能面臨,(1)資訊過載(2)紊亂(3)未精煉的資訊(包含相關與不相關)(4)專業化術語成為使用者下達關鍵詞搜尋時的障礙(5)專業的醫療知識與專有名詞增加民眾學習的難度(6)資訊的品質與可信度。

另外,使用者在使用搜尋工具搜尋健康資訊的過程中可能面臨問題(1)過多未精煉的資訊(包含相關與不相關)(2)健康領域專業化術語,造成使用者下達關鍵詞的障礙,為有效的利用搜尋引擎,使用者須明確知道搜尋資料的方向及關鍵字(3)缺乏文字特徵屬性的資料可能搜尋不到或排名順位後面而不易被使用者找到。部分健康宣導性媒體資源 (如:圖檔轉存的pdf),在缺乏文字特徵屬性的情況下,可能增加被搜尋到的困難度。

在目錄服務的入口網站,使用者在網站中由上往下在以階層式分類的網站結構中尋找需要的資訊,雖有網站導覽的協助但階層式的分類、網站結構與異動性等因素降低階層中頁面的可及性(Accessibility)以及增加使用者搜尋上的困難,其因素包含(1)資源異動、網站或網頁消失、網頁功能改變或內容隨著時間被移置到網站中其他位置(2)使用者與網站設計人員對網站結構認知的差異同樣也增加搜尋上的困難(3)階層的增加降低底層網頁被搜尋到的可能性。

研究中我們發現健康九九使用者中有約12.53%用過搜尋引擎,其中有7.86%用過站內搜尋,顯示大部分的使用者未用過搜尋工具。雖然搜尋引擎提供便利迅速的關鍵字搜尋功能,但是使用者為了找到真正想要的資訊必須一一確認所有找到的網頁,導致其精確率(Precision)與目錄服務的入口網站系統相比來得低,而入口網站所收集到的網頁相對來說較少,因此相關的結果會較搜尋引擎少便造成召回率(Recall)較搜尋引擎來得低。為了更有效益且主動的提供資訊與知識,遂有學者提出資訊過濾(Information Filtering),其中代表作就是推薦系統,期望能經由系統分析使用者閱覽紀錄獲取其偏好或興趣,從使用者的興趣中協助找到相關的資源文件,縮短過濾資訊的時間並改善搜尋引擎精確率低的問題。
使用者閱覽紀錄中可能因無意義的回饋評分影響了推薦結果,因此分析前需剔除回饋評分雜訊,再從過濾掉的資訊中找出文章的相關性。可是相關性的強弱會受到網頁可及性(Accessibility)影響,除此之外相關性僅能知道頁面的回饋評分有相似的趨勢,無法得知回饋評分的高低。在本研究方法中,我們考慮相關係數、回饋評分的高低與頁面的可及性,對推薦清單中的頁面重新排序─提出新方法SORS。

為了驗證SORS的效果,使用衛生署國民健康局健康九九網站的網站日誌(IIS log),進行方法的評估。從歷史閱覽資料中根據使用者4比1的比率分成訓練組(Training Data)與測試組(Testing Data),並從訓練組中挑選一篇使用者曾經看過的文章(Target),並訓練組資料中找出與Target相關之其他文章(Top N,N為相關文章數)作為推薦用,從得到的推薦結果利用測試組進行評估。實驗結果根據不同的參數Top N(推薦文章數)、LEN(推薦分析資料的時間區間長度)進行SORS與KNN的Recall、Precision以及系統效能比較。並發現SORS的首N項小於KNN時其Recall與Precision依然有較好的效果。
英文摘要 From International Telecommunication Union (ITU) Internet population 2006 to 2009 observation report, Internet usage is increasing in countries of the world. The richness and diversity of online information resource has made the Internet to flourish. Researchers observed that people searching online health information has shown an exponential growth trend. The impact has influenced the health industry and becoming one of the considerable methods of obtaining health information.

The information overloading has decreased the efficacy and efficiency of online retrieval. To overcome these problems, search engine and hierarchical structured portal system has been emerged. In the search engine (like: Google), users could retrieve related information through keywords search. As to the directory service portal system (like: health99 or Yahoo), users will browse top to down inside the systems’ directories to obtain and learn information which interest them. Health information could bring benefits such as knowledge increment, self-care management capacity increment, clinical cost reduction, patient’s medical services satisfaction increment and doctor-patients relation strengthening. However, users may encounter difficulties while looking for health information on the Internet, such as (1) information overloading (2) disorganization (3) mass of information (related and unrelated) (4) imprecise or inappropriate search term (5) innaccessible or overly technical language (6) quality and reliability.

Users may encountered problems while searching online health information such as (1) unrefined query result (related and unrelated) (2) imprecise or inappropriate keywords decreasing search efficiency (3) lack of words feature (media resources) on the online documents may decreased probability being searched or augment ranking position from retrieving results. On the other hand, user could also find information through system like yahoo portal system, which organized information in a hierarchical structure. Sitemape can provide navigation information across website sections. However, (1) the internet is uncontrolled and unwatched, without notification, information may disappear, functions may changed or contents could be moven one place another from time to time (2) cognitive differences between users and website designers (3) Increase numerber of the hierarchical layers may also decrease the probability of items of being searched.

In our study, 12.53% of user have used search engine, among them 7.86% have used site search, which shows most of users do not use search tool. The conveniency of search engine has provided convenient and rapid way to retrieve information. However, users found their interest by confirming each retrieved result, this leads a lower precision rate compare to hierarchical structure portal system. On the other side, less retrieving result due to collection and classification difficulties, portal system has lower recall rate compare to the search engine. In order to efficiently and activately provide information, researchers started research on Information Filtering, also known for its recommendation system.

System analyzed and recommends information related to user’s interest from their visiting pattern. Furthermore, there might exist meaningless interest inside the visiting pattern and affect the recommendation result, therefore we need to remove the noise before analyzing related interest. We express the relation of related items by using correlation coefficient. However, item’s accessibility might affect the correlation coefficient, besides it didn’t concerned the relevance feedback magnitude. In our research, we have merged a new sorting method concerning the correlation coefficient, relevance feedback magnitude and item accessibility.

We assessed SORS with Health 99 Internet Information Service Log, operated by the Bureau of Health Promotion, Taiwan. The visiting patterns were divided by users into 80% training data and 20% testing data. The recommendation list is obtained by calculating related item to a certain item (Target) from the training data. The comparison between SORS and KNN (K Nearest Neighbor) was made. We discovered a better result performance while Top N is equal or smaller in SORS. Under different experiment parameter (Top N: size of recommendation list, LEN: time length used to recommendation analysis) we compare recall, precision and system performance between SORS and KNN, and discovered a better recall and precision of SORS under smaller or equal condition compare to KNN.
論文目次 標題 I
審定書 II
上網授權書 III
同意公開申請書 IV
保密同意書 V
誌謝 VI
總目錄 VII
圖目錄 VIII
表目錄 XI
論文摘要 XII
Abstract XV
第一章 緒論
1.1 研究背景 1
1.2 研究動機 2
1.3 研究目的 4
第二章 文獻探討
2.1 健康資訊的興盛與過載問題 5
2.1.1 概述 5
2.1.2 目錄服務系統與搜尋引擎 6
2.2 網頁可及性影響因素 8
2.3 網頁使用探勘 10
2.3.1 網頁資料分類 10
2.3.2 使用者閱覽紀錄區段化處理 11
2.4 推薦系統 12
2.4.1 相關回饋 13
2.4.2 回饋評分雜訊 15
2.4.3 協同過濾式推薦系統 16
2.4.4 內容式推薦系統 19
2.4.5 混合式推薦系統 20
2.5 統計抽樣 22
第三章 研究方法
3.1 研究架構 25
3.2 網路日誌檔 26
3.3 網路日誌資料前處理 29
3.3.1 資料清除 29
3.3.2 閱覽紀錄區段化處理 30
3.4 內容頁面的定義 32
3.5 回饋雜訊處理方法 33
3.5.1 資料蒐集 33
3.5.2 閥值建置 35
3.6 方法設計 40
3.7 系統開發工具 47
第四章 SORS與KNN效能評估
4.1 資料描述 48
4.2 評估方法 49
4.3 實驗流程 50
4.4 實驗結果 52
4.4.1 方法評估 55
4.4.2 系統效能評估 67
第五章 討論與結論
5.1 研究討論 77
5.2 未來展望 81
參考文獻
中文文獻 82
英文文獻 82
電子資料 86
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------------------------------------------------------------------------ 第 17 筆 ---------------------------------------------------------------------
系統識別號 U0007-2707200713321100
論文名稱(中文) 台灣1997至2002年高血壓藥物的用藥型態評估及當AIIA導入市場對臨床使用之影響
論文名稱(英文) An evaluation on the pharmaceutical expenditure of antihypertensive agents during 1997 to 2002 and Impact of introduction of Angiotensin II Antagonist on the antihypertensive drug utilization in Taiwan
校院名稱 臺北醫學大學
系所名稱(中) 藥學研究所
系所名稱(英) Graduate Institute of Pharmacy
學年度 95
學期 2
出版年 96
研究生(中文) 林銅祿
學號 D87010011
學位類別 博士
語文別 中文
口試日期 2007-07-20
論文頁數 145頁
口試委員 指導教授-鄭慧文
指導教授-高雅慧
委員-湯藻薰
委員-邱弘毅
委員-許秀藴
委員-高純綉
委員-康照洲
關鍵字(中) 藥物經濟學
藥物使用量
殘餘值
每日劑量
血管張力素II 拮抗劑
市場佔有率
關鍵字(英) Antihypertensive agent
Defined daily dose (DDD)
Drug utilization
Pharmaceutical expenditure
Angiotensin II antagonist (AIIA)
Angiotensin converting enzyme inhibitor (ACEI)
ß
- blocker (BB)
Calcium channel blocker (CCB)
Market share
學科別分類
中文摘要 利用健保局學術研究資料庫(National Health Insurance Academic Research Database;NHIARD),探討從1997年至2002年台灣高血壓藥物的用藥支付費用及型態,並評估當新的高血壓藥物類別血管張力素II 拮抗劑(Angiotensin II Antagonist;AIIA)導入市場後,對原有高血壓藥物使用行為所造成的影響;特別是針對台灣市場佔有率最高的血管收縮素轉換酶抑制劑(Angiotensin Converting Enzyme Inhibitors;ACEI)、乙型腎上腺素阻斷劑(β- Blockers;BB)、鈣離子阻斷劑(Calcium Channel Blockers;CCB)及其他類別Miscellaneous agents(MIS)等,四大類高血壓藥物。
從健保資料庫(NHIARD)取得1997年至2002年高血壓用藥資料,包括門診病例記錄及用藥處方資料,藥物支付費用可分成5個部份,包括相關的藥物價格、病患人數、每位病患的就醫平均數、每位醫師看診病歷處方中的每日劑量及殘餘值;同時並取得病患的病歷檔案(Medication profile)、醫院看診記錄(Hospital visit record)及藥品申報資料(Drug claim data),執行評估高血壓用藥的申報金額、臨床使用量、處方情形與病患市場佔有率分析等等,藉以瞭解當AIIA導入市場後,對高血壓藥物ACEI、BB和CCB及MIS等類別所造成的影響,本研究將以市場佔有率表示各類別高血壓藥物間的市場相對強度,包括:支付金額市場佔有率、臨床市場佔有率、處方數量市場佔有率、病患人數市場佔有率,同時並探討AIIA對不同層級醫院用藥的滲透時間。
1997年至2002年總高血壓用藥增加102%,主要的成因是混合效應-從病患人數增加34%,醫師看診病歷處方中每位病患的每日劑量增加33%,而高血壓用藥總殘餘值因素僅有7%的影響。此結果與瑞典、西班牙兩國經驗有所不同,其藥物支付費用之增加主要是來自於殘餘值。詳細的殘餘值分析顯示:原廠品牌的類別產品有11%的成長,但一般學名藥卻減少12%。至於醫院的殘餘值:醫學中心成長13%、區域醫院成長17%、地區醫院成長10%,但基層醫療診所卻下降14%。此結果顯示,各層級醫院醫師有不同的處方用藥行為,醫院層級醫師的用藥行為偏向新藥或原開發廠品牌藥,基層醫療醫師則傾向使用一般學名藥。至於,高血壓藥物各類別於支付金額市場佔有率、臨床市場佔有率、處方數市場佔有率、病患人數市場佔有率的分析上,當AIIA導入市場後,對其他高血壓藥物ACEI、BB、CCB和MIS所造成的影響。研究結果顯示,AIIA對ACEI影響最大,這是由於藥理性質相類似所造成的結果,AIIA對BB僅有小部份的影響,對CCB無影響且有正面加分作用,MIS則漸被淡忘。AIIA市場上的成長,最主要的原因是來自於取代ACEI及與其他高血壓藥物一起服用的併服療法,尤其是與CCB和BB併服後讓CCB維持成長BB流失不多。AIIA於各層級醫院以臨床市場佔有率及處方數量市場佔有率的成長強度分析依:醫學中心、區域醫院、地區醫院、基層醫療依序為:4.95:3.77:2.77:1及5.28:4.17:2.94:1。可知,醫學中心>區域醫院>地區醫院>基層醫療。
高血壓藥物支付費用成長的重要原因是病患人數及醫師門診處方中DDDS值的增加所致,各層級醫師有不同的處方用藥行為,醫院層級醫師的用藥行為偏向新藥或原開發產的品牌藥,基層醫療醫醫師則傾向使用一般學名藥。CCB是目前高血壓用藥的主流雖有小成長但再成長有限,AIIA呈現持續成長的趨勢,BB雖微下滑但維持平穩,ACEI有被AIIA取代現象僅以大量價格低廉之學名藥維持其數量上之優勢,MIS已漸被淡忘。由於醫院層級醫師用藥行為偏向新藥及品牌藥,所以醫學中心對AIIA的利用與擴散具有關鍵性之角色。
英文摘要 Background and Purpose
Antihypertensive medications have represented a tremendous financial burden to the health care plan globally. This study examined the utilization pattern of the antihypertensive agents to analyze the underlying reasons responsible for the pharmaceutical expenditure in Taiwan during 1997 to 2002 as well as to evaluate the long term impact of a new pharmacological class antihypertensive medicine- angiotensin II antagonist (AIIA) to the clinical utilization of the existing antihypertensive medications in Taiwan.
Methods
The claims data during 1997 to 2002 were obtained from National Health Insurance Academic Research Database (NHIARD), which include ambulatory service record and prescription data of the entire population. Drug expenditure was decomposed into 5 components: relative drug price, number of patients treated, average physician visit per patient, Defined Daily Dose (DDD) per physician visit and a residual. Gross growth of DDD and prescription numbers, market share analysis including monetary market share (MMS), clinical market share (CMS), prescription market share (PrMS) and patient market share (PtMS), market penetration time and DDDs/ prescription were used to assess AIIA's impact on the angiotensin converting enzyme inhibitors (ACEI), β- Blockers (BB), calcium channel blockers (CCB) and other miscellaneous antihypertensive agents (MIS).
Result
Total antihypertensive drug spending increased 102% during this period, mainly due to the compounding effect from the increment of patients treated (34%) and DDD per physician visit (33%). Residual analysis revealed that the aggregate residual for antihypertensive agents only had a 7% effect; the brand- name product had 11% increment and the generic product had a 12% decrement. It also showed that while hospital sector had a positive 11% residual, primary care clinics had an 11% decrement. Detailed analysis on each sub- group revealed that CCB had the most significant gross growth of DDDs and prescription at 117.1 and 3.4 million increments, respectively. CMS results revealed that the introduction of AIIA had the most significant impact to the clinical utilization of MIS (-5.5%) and a moderate impact to both BB (-4.5%) and ACEI (-4.1%). Whereas PMS demonstrated the most significant impact to MIS (-6.5%), a moderate impact to ACEI (-2.3%) and a very minor impact to BB (-0.9%). AIIA, however, had a positive CMS (+3.9%) and PMS (+2.4%) correlation with CCB. AIIA utilization implicated by relative growth strength for the CMS and PMS at Medical Center, Regional Hospital, District Hospital and Primary Care Clinic were 4.95 : 3.77 : 2.77 : 1 and 5.28 : 4.17 : 2.94 : 1, respectively.
Conclusion
The most important factors that contribute to the expenditure surge of antihypertensive agents are number of treated patients and DDD per physician visit. While physicians at the hospital sector adopted more new and innovative medications, their counterpart at the primary care clinics tended to switch some off- patent products to the generics. The introduction of AIIA did not affect the negative clinical utilization drift of MIS and BB since this depressing trend started before AIIA introduction. On the other hand, the preference of using CCB and AIIA to control hypertension among the physicians in Taiwan was on the ascending side. Medial center was the early leading adaptor for AIIA and furthermore played an important role in the utilization diffusion of AIIA.
論文目次 中文摘要……………………………………………………………1
英文摘要……………………………………………………………3
誌 謝……………………………………………………………5
目 錄……………………………………………………………6
表 目 錄……………………………………………………………8
圖 目 錄……………………………………………………………10
第一章 緒論
第一節 研究背景…………………………………………………12
第二節 研究目的…………………………………………………15
第三節 專有名詞縮寫表…………………………………………16
第二章 文獻探討
第一節 前言………………………………………………………18
第二節 處理高血壓之藥物經濟學分析…………………………19
第三節 高血壓藥物使用基準……………………………………22
第四節 高血壓藥物使用型態及趨勢……………………………31
第五節 影響高血壓處方型態及用藥因素之探討………………32
第三章 研究材料與方法
第一節 研究架構…………………………………………………34
第二節 研究對象及期間…………………………………………35
第三節 研究變項…………………………………………………37
第四節 研究流程…………………………………………………40
第五節 資料整理流程……………………………………………44
第六節 統計分析…………………………………………………45
第四章 以藥物經濟學理論評估台灣1997年至2002年高血壓藥物的用藥型態
第一節 前言………………………………………………………51
第二節 分析方法…………………………………………………53
第三節 研究結果…………………………………………………54
第四節 討論………………………………………………………63
第五章 當AIIA導入市場後對高血壓藥物臨床使用上之影響
第一節 前言………………………………………………………88
第二節 分析方法…………………………………………………90
第三節 研究結果…………………………………………………92
第四節 討論………………………………………………………97
第六章 綜合討論
第一節 年與季之異同……………………………………………112
第二節 高血壓藥物單獨或併服療法的處方型態………………113
第三節 高血壓用藥國內外趨勢之比較…………………………115
第四節 本研究的限制條件………………………………………117
第七章 結論………………………………………………………121
第八章 建議
第一節 對中央主管機關之建議…………………………………123
第二節 對未來研究者之建議……………………………………124
參考文獻……………………………………………………………125
投稿論文……………………………………………………………133
附錄一 門診處方及治療明細檔CD檔……………………………134
附錄二 門診處方醫令明細檔OO檔………………………………139
附錄三 醫事機構基本資料檔HOSB檔……………………………140
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系統識別號 U0007-2907200813540600
論文名稱(中文) 臺灣火刺木成分於人類黑色素細胞之活性探討
論文名稱(英文) The related activities of the constituents from Pyracantha koidzumii in human epidermal melanocytes
校院名稱 臺北醫學大學
系所名稱(中) 生藥學研究所
系所名稱(英) Graduate Institute of Pharmacognosy
學年度 96
學期 2
出版年 97
研究生(中文) 劉宴伶
學號 M303095001
學位類別 碩士
語文別 中文
口試日期 2008-07-08
論文頁數 144頁
口試委員 指導教授-李美賢
委員-郭悅雄
委員-徐鳳麟
關鍵字(中) 臺灣火刺木
臺灣特有種
人類黑色素細胞
酪氨酸酵素
關鍵字(英) Pyracantha koidzumii
endemic species in Taiwan
human epidermal melanocytes
tyrosinas
學科別分類
中文摘要 臺灣火刺木(Pyracantha koidzumii (Hayata) Rehder)薔薇科火刺木屬(Pyracantha)常綠灌木,為臺灣特有種,產於東部低海拔河床地區。本實驗室以人類黑色素細胞之酪氨酸酵素抑制活性進行二十六種臺灣特有種篩選,結果顯示臺灣火刺木果實95% 乙醇萃取物之細胞毒性低(100 μg/mL細胞存活率> 80%)且具細胞內酪氨酸酵素抑制活性(IC50=54.8 μg/mL),故本研究乃採集大量臺灣火刺木之果實,以 95% 乙醇萃取,進行成分分離及其活性探討。經管柱層析分離純化得到十三個化合物,由物理和光譜相關數據解析後,發現其中兩個為新化合物,dibenzofuran結構的3,6-dihydroxy-2,4-dimethoxy-dibenzofuran (9) 和 biphenyl 結構的 3,4-dihydroxy-5-methoxybiphenyl-2′-O-β-D-gluco- pyranoside (13),以及11 個已知化合物,分別為 flavonoids 結構quercetin (1), rutin (2), hyperoside (3), isoquecitrin (4) 和 helicioside B (5);diphenyl ketone glycosides 結構garcimangosone D (6) 和 pyrafortunoside B (7);dibenzofuran結構9-hydroxyeriobofuran (8), fortuneanoside L (10), 2,4-dimethoxy-3,6,9-trihydroxy-dibenzofuranyl-6-O- β-D-glucopyranoside (11) 和biphenyl 結構2-hydroxyaucuparin (12),以上十三個化合物均首次由臺灣火刺木中分離所得。此十三個化合物進行人類黑色素細胞活性篩選試驗,於細胞存活率測試中,發現十二種化合物1~7, 9~13之細胞存活率 > 80 %,進一步對細胞內酪氨酸酵素活性測試,結果顯示新化合物3,4-dihydroxy-5-methoxybiphenyl-2′-O-β-D- glucopyranoside (13) 於100 μM濃度下,對酪氨酸酵素抑制活性最佳,進而以酵素活性染色法(Zymography)探討化合物13 對於酪氨酸酵素氧化活性,結果顯示隨劑量增加,其酵素抑制作用增加;在西方墨點法分析中,結果顯示酪氨酸酵素及TRP 2蛋白表現亦隨化合物13劑量增加,酵素蛋白抑制作用增加。於酪氨酸酵素動力學實驗中,對照組的Km為339.91 μM,Vmax為1.22×10-2 ΔA/min,化合物13於100 μM處理時,Km為261.60 μM,Vmax為8.27×10-3 ΔA/min,屬於非競爭型的混合型抑制劑。綜合本實驗結果,臺灣火刺木分離出十三化合物,其中新化合物3,4-dihydroxy-5-methoxybiphenyl-2′-O-β-D-glucopyranoside (13) 對於抑制人類黑色素細胞內酪胺酸酵素最具潛力,其於人類黑色素細胞中之活性機制將更進一步探討。
論文目次 目錄
目錄 I
圖目錄 IV
表目錄 VIII
縮寫表 IX
縮寫表(續) X
中文摘要 XI
英文摘要 XIII
第一章 緒論 1
一、臺灣火刺木與其它同屬植物研究介紹 1
(一)臺灣火刺木基本介紹 1
(二)火刺木屬植物相關介紹 2
(三)火刺木屬植物活性研究回顧 5
(四)火刺木屬植物之化學成份研究 6
二、人類黑色素細胞及其相關介紹 16
(一)人類黑色素細胞(Melanocytes) 16
(二)黑色素(Melanin) 17
(三)酪氨酸酵素(Tyosinase) 17
(四)黑色素之生化合成(Melanogensis) 18
(五)傳統美白保養品作用機轉 19
三、酪氨 酸酵素動力學機制之研究 21
(一)酵素活性中心性質 21
(二)酵素之抑制劑與其反應型態 21
(三)酪氨酸酵素抑制劑之作用機制 25
四、研究動機 26
第二章 實驗材料與方法 27
一、火刺木之萃取與分離部份 27
(一)一般試藥及溶媒 27
(二)色層分析法材料 27
(三)儀器 28
(四)臺灣火刺木萃取及分離流程 29
(五)水解及單醣的組成分析 32
(六)各成分之物理數據 33
二、 臺灣火刺木之活性實驗部份 42
(一)一般藥品與試劑 42
(二)細胞株及細胞培養液 43
(三)實驗抗體 43
(四)活性儀器設備 43
(五)分析軟體 43
(六)人類黑色素細胞培養 44
(七)人類黑色素細胞存活率試驗 44
(八)蛋白定量法 45
(九)人類黑色素細胞內酪氨酸酵素活性試驗 46
(十)酪氨酸酵素氧化活性染色法 46
(十一)西方墨點法 47
(十二)酪氨酸酵素酵素動力學實驗 48
第三章 結果與討論 49
一、臺灣火刺木之化合物及其結構鑑定 49
(一)Quercetin (1) 之結構解析 49
(二)Rutin (2) 之結構解析 52
(三)Hyperoside (3) 之結構解析 55
(四)Isoquecetrin (4) 之結構解析 58
(五)Helicioside B (5) 之結構解析 61
(六)Garcimangosone D (6) 之結構解析 64
(七)Pyrafortunoside B (7) 之結構解析 67
(八)9-hydroxyeriobofuran (8) 之結構解析 71
(九)3, 6-dihydroxy-2,4-dimethoxy-dibenzofuran (9)之結構解析 74
(十)Fortuneanosides L (10) 之結構解析 77
(十一)2,4-dimethoxy-3,6,9-trihydroxy-dibenzofuranyl-6-O-β-D-glucopyranoside (11) 之結構解析 80
(十二)2-hydroxyaucuparin (12) 之結構解析 83
(十三)3,4-dihydroxy-5-methoxybiphenyl-2′-O-β-D-glucopyranoside (13) 之結構解析 86
二、臺灣火刺木成分之活性探討 89
(一)人類黑色素細胞內存活率試驗 89
(二)人類黑色素細胞內酪氨酸酵素活性試驗 90
(三)人類黑色素細胞內酪氨酸酵素之活性染色試驗 92
(四)人類黑色素細胞內酪氨酸酵素之西方墨點法試驗 93
(五)人類黑色素細胞內酪氨酸酵素抑制動力學實驗 94
第四章 結論 95
第五章 參考文獻 100
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