| 摘要: | 文獻指出白藜蘆醇(resveratrol)在包括糖尿病、阿茲海默和心血管疾病 慢性病中，有改善的研究成果。本研究探討為同分子量的三個白藜蘆醇多聚體在抗氧化活性、抑制糖化反應、細胞模式神經保護、與誘導類似阿茲海默症動物模式的學習與記憶改善功能評估。以牛血清蛋白和5 mM 甲基乙二醛(methylglyoxal)在37C反應5天，生成糖化最終產物(advanced glycation end-products, AGEs)的蛋白質糖基化模式系統中發現，白藜蘆醇多聚體GJ2、GJ3、GJ10 (50和100 μM)有濃度相關抑制BSA的糖基化活性；抑制單胺氧化酶B活性依序為 GJ10 > GJ3 > resveratrol > GJ2；抑制乙醯膽鹼酶活性依序為GJ3 GJ10 > GJ2 > resveratrol；在總抗氧化能力(ORAC)實驗中，白藜蘆醇和白藜蘆醇多聚體GJ2、GJ3、GJ10都具有優於Trolox抗氧化能力，其中ORAC活性依序是GJ2 > GJ3 > resveratrol > GJ10。利用神經膠質瘤SH-SY5Y為模式，以甲基乙二醛誘導SH-SY5Y細胞死亡，在白藜蘆醇和白藜蘆醇多聚體GJ2、GJ3、GJ10不具細胞毒性濃度下 (2.5 到20 M)，GJ3在2.5 μM和GJ10在10 μM即具有明顯SH-SY5Y神經細胞保護作用，而白藜蘆醇與GJ2在測試濃度下並沒有細胞保護作用。在進行以東莨宕鹼(scopolamine)誘導類似阿茲海默動物模型中，可區分為短期餵食一次與長期十四天，每天餵食一次。短期試驗，於開始行為試驗前一小時用管餵方式給予白藜蘆醇多聚體GJ2、GJ3、GJ10 (40 mg/Kg)，藥物給與後半小時i.p注射東莨菪鹼(1 mg/Kg)。結果顯示，以水迷宮評估記憶學習試驗中，在去除平台後，給予GJ3和GJ10的小鼠在平台所在的象限游泳時間和進出該象限的次數高於對照組，並呈現顯著性差異 (P < 0.05)。長期試驗部分，前7天單純管餵給與藥物，第8天後管餵藥物持續到的十四天，且餵藥後半小時i.p注射東莨菪鹼，連續注射7天，實驗總共14天。在被動迴避評估記憶學習試驗中，GJ3組停留於明室的時間增加並與對照組成顯著性差異 (P＜0.001)；而在水迷宮評估記憶學習試驗中，在去除平台後，給予GJ3的小鼠在平台所在的象限游泳時間和進出該象限的次數高於對照組，並呈現顯著性差異 (P < 0.01)。犧牲後，腦組織生化活性測驗方面，未餵藥、i.p注射東莨菪鹼的控制組的乙醯膽鹼酶活性高於正常組及給予GJ3和GJ10餵藥組；在丙二醛含量，GJ3組顯著低於控制組 (P < 0.001)；在總谷胱甘肽上，GJ3組顯著高於控制組 (P < 0.001)；而且在腦中的腦衍生神經滋養因子及其受體的表現量上，GJ3組都顯著高於控制組。以上的研究結果，我們推測白藜蘆醇多聚體GJ3可能是有益於改善神經變性疾病，在阿茲海默症神經保護方面有很大的潛力，而其相關機制需再進一步研究。
It is reported that resveratrol was showed to have improved activities against several chronic diseases, such as diabetes, Alzheimer’s disease, and cardiovascular diseases. In this study, it was aimed to investigate the three resveratrol oligomers (GJ2, GJ3, and GJ10) with same molecular mass in antioxidant and anti-glycation activities, neuroprotective activities in cell models, and the improvement of memory dysfunctions in scopolamine-induced Alzheimer’s mice models. The bovine serum albumin (BSA) was reacted with methylglyoxal at 37C for 5-days, and the advanced glycation end-products (AGEs) formations were decreased dose-dependently with GJ2, GJ3, and GJ10 additions detected by immune stains. The order of resveratrol and resveratrol oligomers for monoamine oxidase B inhibition was GJ10 > GJ3 > resveratrol > GJ2；the order of resveratrol and resveratrol oligomers for acetylcholinesterase inhibition was GJ3 GJ10 > GJ2 > resveratrol；for ORAC activity evaluation, resveratrol and resveratrol oligomers showed better ORAC activity than that of trolox, and the order was GJ2 > GJ3 > resveratrol > GJ10。The SH-SY5Y neuroblastomacells were selected as models for investigating neuroprotective activities of resveratrol and resveratrol oligomers in methylglyoxal-treated ones. Under non-cytotoxic concentrations (2.5 to 20 M) of resveratrol and resveratrol oligomers of GJ2, GJ3, and GJ10, it was found that GJ3 at 2.5 μM and GJ10 at 10 μM exhibited significant protections against methylglyoxal-induced cell death, however, resveratrol and GJ2 showed no neuroprotective activities under tested concentrations. For in vivo animal experiments, two procedures were used in scopolamine-induced Alzheimer’s mice models. One, short-term procedure, resveratrol oligomers of GJ2, GJ3, and GJ10 each was oral administered (40 mg/kg) once a day for 3-days by a gauge 30 min before scopolamine injection (1 mg/kg, i.v.), 30 min later, the mice behaviors were evaluated by Morrison water maze. The first two days were training tests and the third day was probe test (without platform). It was found that mice in the swimming time and crossover in the target quadrant of the probe test were higher in the groups of GJ3 and GJ10 and showed significant difference compared to the control (P < 0.05)。Two, long-term procedure, resveratrol oligomers of GJ3 and GJ10 each was oral administered (40 mg/kg) once a day for 14 successive days, and from the 8th day to the end the scopolamine was injected (i.v.) 30 min after GJ3 and GJ10 oral administrations. The mice behaviors were evaluated by passive step-through and Morrison water maze. It was found that the step-through latency (time for staying in the illuminated chamber, sec) were higher in the GJ3 group and showed significant difference compared to the control (P < 0.001)。For probe test in the Morrison water maze, it was found that mice in the swimming time and crossover in the target quadrant of the probe test were higher in the GJ3 group and showed significant difference compared to the control (P < 0.01)。After being sacrifice, extracts of brain tissues were assayed for acetylcholinesterase activity, MDA contents, and total glutathione contents. It was found that the control group (scopolamine-induced) showed higher acetylcholinesterase activity than that of blank and intervened groups; the GJ3 intervened group had lower MDA contents and higher total glutathione contents and showed significant difference compared to the control (P < 0.001). In addition, brain brain-derived neurotrophic factor and its receptor were also showed significant difference compared to the control. The present results revealed that GJ3 might be beneficial to have neuroprotective activities which might improve neurodegenerative disease and will need further investigations. |
