有關薑黃素引發斑馬魚肌肉病變之機制探討

Underlying mechanism of curcumin-induced myopathy in zebrafish

簡上博(Shang-Po Jiang)

指導教授 ： 黃銓珍

國立臺灣大學/生命科學院/生化科學研究所/碩士(2016年)

https://doi.org/10.6342/NTU201601196

摘要

薑黃素(curcumin)是一種從薑黃根莖中提取得到的黃色色素。早期用來做食品加工的食用色素；而現在醫學研究也發現薑黃素具有抗氧化、抗發炎、降血脂等功能，甚至於能抑制癌細胞增生，促進其走向細胞凋亡而達到抗癌的醫療作用。先前研究指出，用低濃度的薑黃素處理斑馬魚胚胎，會造成不正常的發育問題；而於此篇研究，我們將斑馬魚胚胎以高濃度(54 μM)的薑黃素處理，觀察到會有尾部潰爛的現象，此現象會隨著溫度、濃度以及時間呈現正向相關。利用AO staining也發現尾部的細胞有細胞凋亡的現象發生，此外，延時錄相顯示出這樣的潰爛現象是一個快速、連續的尾部崩解現象。我們接著使用蘇丹黑的染色方法觀察到免疫反應隨著薑黃素的處理而上升，並透過qPCR更證明了在necroptosis裡面扮演重要角色的HSP90基因表現量也隨著薑黃素處理而上升。而Rip1、Rip3以及HSP90目前都具有專一抑制的抑制劑，分別為Necrostatin-1、Dabrafenib及17-DMAG，我們選用這三種藥物去測試是否能延緩斑馬魚的尾部潰爛現象以及此現象的發生是否與Necroptosis相關。我們將受精後一天的斑馬魚浸泡在具有Curcumin藥物處理的養殖水再個別搭配三種抑制藥物處理後發現尾部潰爛現象確實可被延緩，而從中證明了這個薑黃素所造成的尾部潰爛的現象是一個透過rip1以及rip3去作用的necroptosis路徑。
最後，我們透過先讓薑黃素作用，之後再加入抑制藥物的實驗方式去模擬病人的發病模式，發現此薑黃素造成尾部潰爛現象的斑馬魚可作為Necroptosis研究的生物模式以及未來可以應用於治療藥物的篩選。

關鍵字

薑黃素(curcumin) ； Programmed Necrosis/Necroptosis ； Necrostatin-1 ； Dabrafenib

並列摘要

Curcumin has multiple roles in different pathway such as antioxidant, hypoglycemic, anti-inflammatory and anti-cancer function. In previous study, curcumin was proved to have its embryotoxic and teratogenic effects on zebrafish development at low dose. In this study, we indicated that 24 hpf embryos treated with curcumin at high dose (54 μM) displayed severe tail bud damage, which is in a dose- and temperature-dependent manner. By using AO staining, we demonstrated that apoptosis was involved in the tail bud damage of embryos. Moreover, time-lapse recording indicated that curcumin-induced myopathy is a continuous and fast damage phenomenon. We then performed Sudan black B staining and discovered an excessive immune response in curcumin-induced myopathy. Besides, qPCR data showed that higher mRNA level of heat-shock protein 90 (hsp90) was induced during curcumin-induced myopathy. It led us to investigate the roles of rip1/rip3 complex during curcumin-induced myopathy and its correlation with necroptosis. By using necrostatin-1 (rip1 inhibitor), dabrafenib (rip3 inhibitor) and hsp90 inhibitor to treat zebrafish embryos with curcumin treatment, our data indicated that all of these three inhibitors could postpone curcumin-induced myopathy progression, suggesting that curcumin- induced myopathy may be a rip1- and rip3-dependent pathway.
Taken together, in this study we established a curcumin-induced myopathy in zebrafish, which has potential to be a new necroptosis animal model.

並列關鍵字

curcumin ； myopathy ； necroptosis ； rip1 ； rip3

參考文獻

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