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標題: | 退化性關節炎軟骨之醣胺聚醣分析 Glycosaminoglycan Analysis of Human Osteoarthritis Cartilage |
作者: | Tzung-Sheng Lin 林宗聖 |
指導教授: | 梁碧惠 |
關鍵字: | 醣胺聚糖,退化性關節炎, Glycosaminoglycan,Osteoarthritis, |
出版年 : | 2016 |
學位: | 博士 |
摘要: | 在六十歲以上的人中,超過50%的人會患退化性關節炎,即使得到此病機率如此高,但其病理機轉還尚不清楚,如果可以了解退化的過程與原因那對退化性關節炎的治療可能出現曙光。由於退化性關節炎的每個階段都不同,我們研究退化性關節炎軟骨中可能造成醣胺聚醣(GAG: glycosaminoglycan)降解的酵素活性,以及分析醣胺聚醣組成。我們分析在退化性關節炎磨損部位與非磨損部位的組成且發現,第一,在磨損部位的軟骨醣胺聚醣含量比較少,第二,分析硫酸軟骨素(CS: chondroitin sulfate)發現,除了硫酸軟骨素的量變少之外,其組成也發生改變,CS-A (chondroitin-4-sulfate)變少。將磨損與非磨損部位軟骨分成上中深層,我們也分析硫酸軟骨素在不同層間的分布,發現硫酸軟骨素的含量CS-A、CS-C (chondroitin-6-sulfate)與total CS由上層往深層遞增,磨損部位的CS-A的含量較非磨損部位減少,且發現在磨損部位的CS-A會隨著年紀增長而下降。硫酸軟骨素的組成發生改變,推測其原因可能為硫酸軟骨素的生成與代謝的酵素失衡,硫酸軟骨素生合成降解酵素的分析正在進行中,我們目的是獲得在退化性關節炎發展過程中的關鍵原因。
另一方面,Sulf1與Sulf2 (HS 6-O endosulfatase 1、2)在退化性關節炎的軟骨中過度表現,我們想了解Sulf1與Sulf2是否會影響軟骨中硫酸乙醯肝素(HS: heparan sulfate)組成,我們成功開發HPLC、FL-HPLC與LC-MS的方法,由於可能超出儀器的偵測極限,導致在軟骨並沒有發現硫酸乙醯肝素的存在。由於硫酸乙醯肝素對於細胞存活分化相當重要,過度表現的Sulf1與Sulf2可能會影響硫酸乙醯肝素組成,而影響軟骨細胞生長因子的活化與訊息傳遞,我們也開發Sulfatase的抑制劑,且對比之前報導過的11b (glucosamine-6-sulfamate analogue) (IC50 = 11.3 Osteoarthritis affects more than 50% of people over the age of 60 years old. Even though so populated this disease is, the pathophysiologic progression of this disease remains unclear. A better understanding for the pathway of disease progression would shed light on curative treatment. Because progressive speed of osteoarthritis varies in every phase, GAG (glycosaminoglycan) composition and the degradation enzymatic activity in the cartilage of human osteoarthritis was investigated. We analyzed the GAG composition between remote and lesion cartilage and had several findings. Frist, total GAG quantity in lesion group was lower than in the remote group. Second, CS (chondroitin sulfate) quantity and its subunit CS-A (chondroitin-4-sulfate) also showed significantly decrease in lesion cartilage. Since cartilage was stratified to superficial, middle and deep zone, we also analyzed zonal distribution of CS. CS-A, CS-C (chondroitin-6-sulfate), total CS were gradually increased from superficial to deep zone. There was more CS-A in deep zone of remote cartilage than remote cartilages. In lesion cartilage, age had a significant inverse correlation with the CS-A concentration, but CS-0, CS-C and total CS not. CS composition was reasoned for that catabolic and anabolic enzymes were loss of homeostasis in lesion cartilage. The mRNA expresson of catabolic and anabolic enzymes are analyzed, our purpose was to clarify the key point of osteoarthritis progression. Otherwise, Sulf1 and Sulf2 (HS 6-O endosulfatase 1, 2) were overexpressed in lesion cartilage, we investigated HS (heparin sulfate) composition in the cartilage, since it was modified by Sulf1 and Sulf2. HS analysis were conducted by HPLC, FL-HPLC and LC-MS methods, however, HS was not found in cartilage, we reasoned that the concentration of HS was out of detection limit. Because HS was important for cell survival, overexpression of Sulf1 and Sulf2 should have great influence on HS composition, and cause chondrocyte to loss of homeostasis. We designed and synthesized a series of the sulfatase inhibitors. In comparison to 11b (IC50 = 11.3 |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/19053 |
DOI: | 10.6342/NTU201602936 |
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顯示於系所單位: | 藥學系 |
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