壹、使用蛋白體學研究技術建立Cu/Zn過氧化物歧化? (SOD1) 突變之肌萎縮性側索硬化症 (ALS) 的治療策略
肌萎縮性側索硬化症 (Amyotrophic lateral sclerosis；ALS) 是一種在中樞神經系統內控制隨意肌的運動神經元 (motor neuron) 退化的疾病。目前為止尚無有效治療方法，研究發現ALS的病人主要是由於Cu/Zn過氧化物歧化? (Superoxide dismutase 1；SOD1) 特殊的錯誤折疊構形所造成。這種結構可能是遺傳性的基因突變或後天的轉譯後錯誤修飾所導致。大部分這種不正常的構形並不會明顯影響酵素本身的功能，卻會對細胞造成毒性，而這種影響主要是因為錯誤摺疊的SOD1蛋白質與原本不相關的特定蛋白質發生特殊的交互作用，導致該蛋白質能力喪失所導致。因此本研究中希望可以藉由酵母菌雙雜合篩選技術來尋找與A4V和G93A突變之SOD1蛋白質不正常交互作用之蛋白質，希望藉由篩選出來的蛋白質來推測ALS可能的致病機轉及建立有效的治療策略。
貳、研究楊梅素 (Myricetin) 對脂多醣體刺激活化之小鼠樹突細胞的免疫調節效應
楊梅素 (Myricetin) 是一種自然界中的黃酮類化合物，存在於許多的水果、蔬菜、茶及藥材中。楊梅素具有抗發炎的特性，但到目前為止還沒有楊梅素對於樹突細胞的免疫調節功能相關的研究。本研究的目的則是要來評估楊梅素調控脂多醣體刺激活化源自於老鼠骨髓樹突細胞的可能性。我們的實驗數據顯示使用高達 10 ug/ml 劑量的楊梅素，在不會引起細胞毒性的情形下，楊梅素明顯的降低由脂多醣體刺激樹突細胞所產生的腫瘤壞死因子-α、間白素-6和間白素-12p70 的分泌量。脂多醣體所誘導在樹突細胞表面的第二型主要組織相容性複合物及CD40、 CD86分子的表現均會被楊梅素所抑制，且楊梅素也會抑制樹突細胞的內吞作用和脂多醣體刺激樹突細胞的遷移能力。此外，脂多醣體刺激樹突細胞引起同種異體 T 淋巴細胞增生也會藉由楊梅素來降低。而且我們的實驗結果證實楊梅素利用抑制 I?羠激?/核因子-?羠和有絲分裂原活化蛋白激?的路徑來降低由脂多醣體刺激活化的樹突細胞。因此，利用楊梅素來調節樹突細胞活性可能是一具有吸引力的策略來治療發炎、自體免疫疾病以及組織移植排斥。
I.Developing therapeutic strategies for amyotrophic lateral sclerosis (ALS) using proteomic diversity of Cu/Zn superoxide dismutase (SOD1) mutant
Amyotrophic lateral sclerosis (ALS) is a degenerative disease of motor neuron that controls voluntary muscle in central nervous system. At present, there is no effective treatment in ALS. A research finds that patients of ALS is mainly caused by specific misfolded Cu/Zn (superoxide dismutase 1；SOD1). This misfolded structure may be influenced by inherited mutations of the gene or wrong modifications of post-translation. Generally this abnormal configuration will not significantly affect activity of enzyme itself, but it can cause toxicity to cells. This toxic effect may be caused by the special interaction with unrelated proteins. Therefore, this research hope to seek the protein that abnormally interacts with SOD1 mutant of A4V and G93A by using yeast-two hybride screening. Moreover, we want to address possible pathogenic mechanism of ALS and then to develop effective therapeutic strategies.
II.The study of immunomodulatory effect of myricetin on lipopolysaccharide-stimulated activation of mouse bone marrow-derived dendritic cells
Myricetin is a naturally occurring flavonoid that is found in many fruits, vegetables, teas and medicinal herbs. It has been demonstrated to have anti-inflammatory properties, but, to date, no studies have described the immunomodulatory effects of myricetin on the functions of dendritic cells (DCs). The aim of this study was to evaluate the potential for myricetin to modulate lipopolysaccharide (LPS)-stimulated activation of mouse bone marrow-derived DCs. Our experimental data showed that treatment with myricetin up to 10 μg/mL dose not cause cytotoxicity in cells. Myricetin significantly decreased the secretion of tumor necrosis factor-α, Interleukin-6 and interleukin-12p70 by LPS-stimulated DCs. The expression of LPS-induced major histocompatibility class II, CD40 and CD86 on DCs was also inhibited by myricetin, and the endocytic and migratory capacity of LPS-stimulated DCs was blocked by myricetin. In addition, LPS-stimulated DC-elicited allogeneic T-cell proliferation was reduced by myricetin. Moreover, our results confirmed that myricetin attenuates the responses of LPS-stimulated activation of DCs via suppression of I?羠 kinase/nuclear factor-?羠 and mitogen-activated protein kinase-dependent pathways. Myricetin has novel immuno-pharmacological activity, and modulation of DCs by myricetin may be an attractive strategy for the treatment of inflammatory, autoimmune disorders and for transplantation.
