臺灣博碩士論文加值系統

樟芝 (Antrodia camphorata, AC) 為台灣特有藥用真菌。本試驗使用薇得生技公司所生產的固態發酵樟芝 (solid fermentation A. camphorata, SAC)來檢測其是否具有調節小鼠免疫作用及抗腫瘤之活性。
每天餵食不同劑量50、100、200 mg/Kg SAC，並以生理食鹽水為對照組。結果顯示第4天後餵食SAC小鼠脾臟細胞數目皆較對照組有顯著增加；在餵食後的第11天，除餵食200mg/Kg者之脾臟細胞數目較高外，其他三組無顯著差異。上述增加之脾臟細胞數目以T細胞為主，但是B細胞在整個餵食過程中則無明顯增加。取出之小鼠脾臟淋巴細胞在非特異性抗原Con A的刺激下，刺激指數處理組較對照組有顯著增加。
注射5-flurouracil (5-FU) 之後同時餵食50 mg/Kg SAC及生理食鹽水與正常小鼠之試驗結果顯示：5-FU處理之小鼠，於第4、7天其脾臟細胞數量明顯低於正常小鼠，而餵食生理食鹽水之小鼠脾臟細胞明顯低於餵食SAC之小鼠；在第15天餵食生理食鹽水的小鼠，其脾臟細胞數量大量增生且明顯高於正常小鼠與餵食SAC之小鼠者，而餵食SAC之小鼠脾臟細胞數量與正常小鼠沒有顯著差異。由以上結果推論SAC似乎有保護脾臟細胞之現象。以上所述增加之脾臟細胞仍以T細胞為主，且注射5-FU並餵食50 mg/Kg SAC小鼠之CD4+及CD8+ T細胞比例高於正常及餵食生理食鹽水的老鼠，不過B細胞則無明顯改變。
在抗腫瘤的活性測試，體外試驗結果顯示：Hep 3B、A549及 LL/2 細胞之存活率隨著固態發酵樟芝乙醇萃取液處理濃度及時間之增加而降低。在動物體試驗，固態發酵樟芝能顯著降低老鼠身上的腫瘤大小且延長存活的時間。
綜合以上的結果顯示，固態發酵樟芝具有調節小鼠免疫系統及抗腫瘤的效果。

Antrodia camphorata (A. camphorata) is a medicinal fungus merely found in Taiwan. In this study, solid-state fermentation product of A. camphorata (SAC) from Wei-de biotechnology company was employed to examine the immunomodulatory and antitumor effects on mice.
Treatment of BALB/c male mice with a variety of concentration of SAC (50, 100, and 200 mg/Kg) resulted in a significant proliferation of splenocytes at day 4 as compared with saline treated group. The numbers of splenocytes in mice treated with 200 mg/Kg SAC increase and have significant difference from three other treatments after 11 days. The increased splenocytes are mainly T cells; B cells were not significantly changed. When the splenocytes were stimulated by concanavaliv A in vitro, there was a significant difference between saline treated group and SAC treated group in stimulation index.
BALB/c male mice were injected with 5-FU and fed simultaneously with 50 mg/Kg SAC or saline then compared with naïve. The results are as the following: at day 4 and 7, the number of splenocytes in 5-FU treated mice was significantly less than those of naive and the amount of splenocytes in SAC treated group was much greater than those of saline treated group; at day 15, splenocytes in saline treated group increased significantly as compared to those in non-treated group along with SAC treated group while the number of splenocytes in SAC fed mice was not significantly different from those in non-treated mice. It could be inferred from the above that SAC seems to have the capability to protect the spleen from damage exerted by 5-FU. The increased splenocytes were mainly T cells and the ratio of CD4＋ to CD8＋ in the 50 mg/Kg SAC fed mice was higher than those in saline fed and non-treated mice. The B cell level remains unaltered.
The result of cytotoxicity activity in vitro showed that the survival of Hep 3B, A549 and LL/2 decreased with the increase of treated concentration of ethyl extracted SAC as well as time of treatment. In vivo, SAC could reduce the size of tumor in mice and prolong the survival time as well.
Based on the experiments we have conducted, SAC appears to have the effect of immunomodulatory and antitumor activities against tumors bore in BALB/c male mice.

書名頁………………………………………………………………………………..i
博碩士論文授權書…………………………………………………………………..ii
論文口試委員審定書………………………………………………………………..iii
中文摘要……………………………………………………………………………..iv
英文摘要…………………………………………………………………….……….v
誌謝…………………………………………………………………………………..vi
目錄……………………………………………………………………………….. ..vii
圖目錄……………………………………………………………………………….viii
表目錄……………………………………………………………………………….ix
第一章、引言……………………………………………………………………….1
第二章、前人研究………………………………………………………………….2
第一節、樟芝……………………………………………………………..2
第二節、固態發酵………………………………………………………..6
第三節、菇蕈類及中草藥芝免疫調節…………………………………..7
第四節、癌症…………………………………………………………….13
第五節、研究動機……………………………………………………….17
第三章、實驗材料與方法…………………………………………………………18
第一節、實驗材料………………………………………………….……18
第二節、實驗方法……………………………………………………….20
第四章、結果與討論………………………………………………………………28
第一節、固態發酵樟芝菌絲對BALB/c小鼠免疫系統之影響…….….28
第二節、固態發酵樟芝對B6小鼠皮下原位癌細胞之影響….…..…...34
第五章、結論………………………………………………………………………37
參考文獻……………………………………………………………………….…...38
縮寫指引…………………………………………………………………….….…..68
附錄一、流式細胞儀之工作原理…………………………………………………69
附錄二、MTS分析原理…………………………………………………………..73
作者簡介……………………………………………………………………………74
表目錄
表一、實驗期間各組小鼠體重之變化……………………………………..…..….52
表二、餵食固態發酵樟芝30 天小鼠血清中GOT之變化………………………53
表三、餵食固態發酵樟芝30 天小鼠血清中GPT之變化……………………….54
表四、固態發酵樟芝對BALB/c小鼠之脾臟細胞總數之變化…………………..56
表五、固態發酵樟芝對BALB/c小鼠之脾臟淋巴細胞對非特異性抗原之反應..57
表六、固態發酵樟芝對受5-flurouracil (5-FU) 免疫抑制的BALB/c小鼠脾臟內細胞總數之變化……………………………………………………………….62
表七、固態發酵樟芝乙醇萃取液處理Hep 3B, A549, LL/2及HEL 299細胞的50%之抑制濃度 (IC50)………………………………………………………….71
圖目錄
圖一、BALB/c小鼠在餵食固態發酵樟芝30天的期間，每7天測定小鼠體重之變化。……………………………………………………………………..….52
圖二、BALB/c小鼠餵食固態發酵樟芝30天之肝臟組織切片…………………….55
圖三、BALB/c小鼠在餵食固態發酵樟芝菌絲懸浮液30天的期間，各組間脾臟中T細胞之變化趨勢………………………………………………………..58
圖四、BALB/c小鼠在餵食固態發酵樟芝菌絲懸浮液30天的期間，各組間脾臟中CD4+ T細胞之變化趨勢…………………………………………………59
圖五、BALB/c小鼠在餵食固態發酵樟芝菌絲懸浮液30天的期間，各組間脾臟中CD8+ T細胞之變化趨勢……………………………………………….60
圖六、BALB/c小鼠在餵食固態發酵樟芝菌絲懸浮液30天的期間，各組間脾臟中B細胞之變化趨勢……………………………………………………….61
圖七、固態發酵樟芝對受5-flurouracil (5-FU) 免疫抑制的BALB/c小鼠脾臟內T 細胞之變化…………………………………………………………………..63
圖八、固態發酵樟芝對受5-flurouracil (5-FU) 免疫抑制的BALB/c小鼠脾臟內CD4+ T 細胞之變化………………………………………………………...64
圖九、固態發酵樟芝對受5-flurouracil (5-FU) 免疫抑制的BALB/c小鼠脾臟內CD8+ T 細胞之變化………………………………………………………...65
圖十、固態發酵樟芝對受5-flurouracil (5-FU) 免疫抑制的BALB/c小鼠脾臟內B 細胞之變化…………………………………………………………………..66
圖十一、固態發酵樟芝乙醇萃取液對Hep 3B細胞存活率之影響………………...67
圖十二、固態發酵樟芝乙醇萃取液A549 細胞存活率之影響……………………68
圖十三、固態發酵樟芝乙醇萃取液LL2 細胞存活率之影響……………………..69
圖十四、固態發酵樟芝乙醇萃取液HEL 299 細胞存活率之影響………………...70
圖十五、固態發酵樟芝對帶有腫瘤之B6小鼠腫瘤體積之影響………………….72
圖十六、固態發酵樟芝對帶有腫瘤之B6小鼠存活率之影響…………………….73

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