臨床上，過量活性氧化物質(Reactive Oxygen Species, ROS)的產生是促進許多炎症疾病進展的關鍵因素，例如骨關節炎等。氫氣(Hydrogen gas, H2)被視為是一種新的醫學氣體，對細胞和器官具有抗氧化、抗發炎和抗凋亡等保護作用，其最大優點為在體內的施用安全性高且無明顯的副作用。目前給予H2的方式主要有直接吸入含有H2的混合氣體、飲用飽和氫水及透過靜脈或腹腔注射含有H2的生理食鹽水等，而在常溫常壓下H2在水中的溶解度很低且其具有容易逸散的特性，導致這些給予方式在病灶處在短時間內不易有效地累積H2濃度，須長期給予H2。在本研究裡，我們將鎂粉包覆於具疏水性的聚乳酸-甘醇酸(poly lactic-co-glycolic acid, PLGA)載體載體內，以原位注射方式將此微球注射至小鼠的骨關節炎處，藉由發炎處所產生的組織液滲透至微球內部後，與鎂粉進行反應產生H2，除了可以保護因鎂粉造成的組織傷害，也可視為一氫氣緩釋系統，其產氫時間可以延長至3天，且產生的H2可以藉由擴散進入組織和細胞中，於病灶處達到有效的治療濃度，進而清除病灶處所產生的ROS，以達到治療骨關節發炎的效果。

Inflammation is associated with numerous diseases including osteoarthritis, which is featured by the overproduction of reactive oxygen species (ROS). Hydrogen (H2) has been reported to have the capability to reduce oxidative stress by selectively scavenging hydroxyl radical (•OH) with no apparent side effects and can thus be served as a therapeutic medical gas for the treatment of inflammatory diseases. Administration of H2-gas (via inhalation with air), H2-water (via oral intake) or H2-saline (via intravenous drip infusion), has been shown promising in treating several inflammatory diseases in animal models. However, the amount of H2 absorbed by the body systematically through these methods may not enough to scavenge the ROS generated in inflamed tissues, because the solubility of H2 in the biological environment is low. To dissolve this problem, a poly lactic-co-glycolic acid (PLGA) microparticle system containing magnesium powders (Mg@PLGA MPs) that can generate H2 gas in a sustained manner is proposed herein. The feasibility of using the as-prepared Mg@PLGA MPs to scavenge ROS, thus improving local inflammation, is studied in a mouse model with experimentally created osteoarthritis, via local injection.

摘要 I
Abstract II
目錄 III
圖目錄 VI
表目錄 VII
第一章 緒論 1
1-1 骨關節炎及其治療 1
1-2活性氧化物質與骨關節炎 1
1-3 生物醫學氣體及氫氣 2
1-4 鎂與鎂合金 4
1-5 聚乳酸-甘醇酸 5
1-6 研究動機與目的 5
第二章 實驗材料和方法 8
2-1 包覆鎂粉之PLGA(Mg@PLGA)微球的製備 8
2-2 Mg@PLGA MPs之型態與粒徑分析 8
2-3 Mg@PLGA MPs包覆之計算 9
2-4 氫氣濃度量測 10
2-5 小鼠巨噬細胞(RAW264.7)的培養與繼代 11
2-6 材料毒性測試 11
2-6-1 細胞存活率分析 11
2-6-2 LIVE/DEAD細胞存活觀察 11
2-7 細胞內ROS的含量分析 12
2-8 蛋白質定量及促炎細胞因子測定 12
2-9 細胞免疫螢光染色 13
2-10 動物實驗 13
2-10-1 鎂粉接觸組織之危害分析 13
2-10-2 關節炎小鼠疾病模型的建立 14
2-10-3 小鼠體內ROS含量偵測 14
2-10-4 體內治療效果評估 14
2-10-5 動物組織病理分析及組織免疫螢光染色 15
2-11 統計分析 15
第三章 實驗結果與討論 16
3-1 載體物化性分析 16
3-1-1 Mg@PLGA MPs之大小與型態 16
3-1-2 Mg@PLGA MPs的包覆及釋放 17
3-2 體外實驗 19
3-2-1 不同材料對細胞培養基pH值的影響 19
3-2-2 材料的毒性測試 20
3-2-3 Mg@PLGA MPs抑制LPS誘導ROS的表現 22
3-2-4 Mg@PLGA MPs降低LPS誘導促炎細胞因子的表達 23
3-3 動物實驗 26
3-3-1 Mg於組織的影響 26
3-3-2 Mg@PLGA MPs顯著抑制ROS的表現 27
3-3-3 Mg@PLGA MPs降低促炎細胞因子的表現 28
3-3-4 Mg@PLGA MPs減少脂質氧化 29
3-3-5 H&E組織病理切片 30
第四章 結論 31
參考文獻 32

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