臺灣博碩士論文加值系統

牛樟芝為台灣特有藥用真菌，已有許多研究指出其具有相當多功效性，但子實體和菌絲體間於功效和安全性則有些差異，如本實驗室在先前對大鼠進行的28 天連續餵食的安全性試驗中，已知菌絲體及子實體水萃物對腎上腺及卵巢並無影響，子實體酒萃物對腎上腺及卵巢則會造成細胞脂質堆積現象。本實驗目的為探討牛樟芝子實體酒萃物對於倉鼠及大鼠腎上腺及卵巢細胞脂質堆積之差異，並評估其可能的機制。實驗包含兩部分，第一部分為敘利亞倉鼠28 天餵食毒性試驗，評估牛樟芝子實體酒萃物是否亦會在倉鼠腎上腺及卵巢造成細胞脂質堆積現象；第二部分為大鼠7 天餵食短期毒性試驗，評估牛樟芝子實體酒萃物對固醇合成的影響以及可能機制，分別給予大鼠兩種已知的固醇生成抑制劑Tricresyl phosphate (TCP)及Ketoconazole (KET)以及牛樟芝子實體酒萃物(ACE)，比較腎上腺及卵巢脂質堆積差異、血液固醇與中間產物含量變化並進一步評估大鼠及倉鼠腎上腺CYP11A1及荷爾蒙敏感性脂肪酶(Hormone sensitive lipase, HSL)蛋白質表現量。第一部分結果顯示，倉鼠28 天餵食毒性試驗中，牛樟芝子實體酒萃物(2.4 g/kg bw)對倉鼠體重、血液學、血清生化學、尿液學、臟器重量、肉眼及組織學檢查均無明顯差異，同時腎上腺及卵巢並無脂質堆積現象。第二部分結果顯示，大鼠7 天餵食短期毒性試驗中，TCP (0.7 g/kg bw)、KET (0.15 g/kg bw)以及ACE (1及2 g/kg bw)組之腎上腺皮質細胞肥大及脂質堆積病理積分均有顯著性增加，而卵巢間質細胞肥大之病理積分僅ACE組與TCP組有顯著性增加。以ELISA偵測血清中皮質固醇(Corticosterone)以及相關前驅物的變化，ACE組與對照組比較雖無明顯差異，但ACE高劑量組(2 g/kg bw)對雌性大鼠腎上腺HSL表現量具有顯著性降低作用，推測可能對上游區之膽固醇的代謝具有抑制作用。
綜合以上結果，牛樟芝子實體酒萃物於高劑量(2 g/kg bw)對雌性大鼠腎上腺HSL表現量具有抑制作用，推測為引起腎上腺皮質及卵巢間質細胞之細胞肥大及脂質堆積主因，但對倉鼠並無影響，顯示具有種間差異性，其原因可能與倉鼠和大鼠固醇合成細胞獲取膽固醇的途徑差異有關，即倉鼠腎上腺獲取膽固醇方式主要為自行合成路徑，而血液中獲取膽固醇的途徑以低密度脂蛋白為主；而大鼠則以高密度脂蛋白為主，且細胞質內含有較多以脂質滴儲存的膽固醇酯可利用。由於倉鼠獲得膽固醇的來源及代謝途徑和人類較類似，本實驗結果可作為牛樟芝子實體酒萃物於人體食用安全性之參考。

Antrodia cinnamomea is an endemic medicinal mushroom in Taiwan and poses many bioactive compounds. There are some difference about efficacy and safety between fruiting body and mycelia. In our previous 28-day feeding study in rats revealed that hypertrophy and lipidosis in the adrenal glands and ovaries were found at doses higher than 1 g/kg bw Antrodia cinnamomea fruiting body ethanol extract (ACE) treatment but not water extract nor mycelia . The aim of this study is to compare the effect induced by ACE between rat and hamster and elucidate the possible mechanisms. Firstly, hamsters were administrated by oral gavage of 2.4 g/kg bw ACE for 28 days to evaluate the lipidosis in the adrenal glands and ovaries will happen or not. Secondly, a 7-day short-term study in rat was conducted and serum concentrations of steroids, morphological change in adrenal glands and ovaries were measured for the comparison between ACE and other two well-known inhibitors of tricresyl phosphate (TCP) and ketoconazole (KET) on adrenal steroidogenesis. Additionally, CYP11A1 and hormone sensitive lipase (HSL) expression in the adrenal glands were detected by Western blotting. In the 28-day feeding toxicity study of hamster, no significant changes were found in weight, urine and hematological analysis, organs weights, gross and histological examination. Indeed, no significant change of lipidosis in the adrenal glands and ovaries were found in 2.4 g/kg bw ACE treated hamsters. In the 7-day short term feeding study of rats, significant increased scores of lipidosis and hypertrophy in the adrenal cortex were observed in rat by oral gavage of TCP (0.7 g/kg bw), KET (0.15 g/kg bw) or ACE (1 or 2 g/kg bw). Significant increase of ovarian interstitial hypertrophy scores were also found in TCP and ACE treated rats. Although no significant difference between control and ACE group in corticosterone and precursors in serum of rats were found by ELSA, a significant decrease of HSL expression was found in the 2 g/kg bw ACE-treated female rats by Western blotting analysis and was thought to reflect the inhibit effect on cholesterol utilization.
In conclusion, different effects of lipidosis in the adrenal glands and ovaries induced by ACE were found between rats and hamsters. The pathogenic differences may due to different cholesterol utilized system of steroidogenic cells between rats and hamsters. It is considered that hamsters utilize cholesterol via endogenous synthesis and majority of lipoprotein-derived cholesterol utilized is low density lipoprotein while for rats is obtained from high density lipoprotein and rich cellular stored cholesteryl esters (lipid droplet). Due to the metabolic system of cholesterol utilization in human is similar to hamsters; the results in this study can be used to be a reference for the food safety of ACE in human.

第一章 前言 1
第二章 文獻探討 2
第一節 牛樟芝介紹以及成分分析 2
第二節 牛樟芝調節脂質代謝之相關文獻 3
第三節 牛樟芝安全性相關文獻 4
第四節 腎上腺及卵巢之生理介紹 4
第五節 腎上腺及卵巢之毒性反應 6
第六節 Tricresyl phosphate及Ketoconazole對腎上腺及性腺之影響 7
第七節 血液固醇荷爾蒙中間產物監測與腎上腺固醇生成抑制之毒理評估 9
第八節 腎上腺生理及毒性反應的種間差異 9
第九節 荷爾蒙敏感性脂肪酶 10
第三章 研究目的與實驗設計 13
第一節 研究目的 13
第二節 實驗設計 13
第三節 實驗架構 14
第四章 材料與方法 15
第一節 試驗樣品及製備 15
第二節 倉鼠28天餵食毒性試驗 17
第三節 大鼠連續口服7日毒性試驗 19
第四節 以西方墨點法偵測組織酵素表現量 23
第五章 結果 28
第一節 倉鼠28天餵食毒性試驗 28
第二節 大鼠7天餵食短期毒性試驗 58
第三節 以西方墨點法偵測組織酵素表現量 117
第六章 討論與結論 123
第七章 參考文獻 131

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