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研究生:張亞如
研究生(外文):Ya-Ju Chang
論文名稱:活菌體與熱致死菌體 Enterococcus faecalis YM-73 對於卵白蛋白致敏小鼠之抗過敏影響
論文名稱(外文):Anti-allergic effects of viable and heat-killed Enterococcus faecalis YM-73 on ovalbumin-sensitized BALB/c mice
指導教授:林美吟林美吟引用關係
指導教授(外文):Meei-Yn Lin
口試委員:胡宏熙吳思敬
口試日期:2011-07-20
學位類別:碩士
校院名稱:國立中興大學
系所名稱:食品暨應用生物科技學系所
學門:農業科學學門
學類:食品科學類
論文種類:學術論文
論文出版年:2011
畢業學年度:99
語文別:中文
論文頁數:98
中文關鍵詞:活菌體乳酸菌熱致死乳酸菌過敏性鼻炎IgEIgG1IgG2aIFN-γIL-5
外文關鍵詞:viable lactic acid bacteriaheat-killed treatment lactic acid bacteriaallergic rhinitisIgEIgG1IgG2aIFN-γIL-5
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乳酸菌已被認定為有助於宿主身體健康的益生菌,在免疫調節及抗過敏上也具有相當的功效。現今,乳酸菌的益生功效已經不侷限於活菌型態,熱致死的乳酸菌也具有免疫調節的效果,並且有著保存期限較長、運輸便利及容易儲存等優點。根據先前的研究,我們從 11 株經過熱致死處理的乳酸菌,於體外試驗中挑選出調節免疫功能最具潛力的乳酸菌菌株,其能夠藉由刺激人類周邊免疫單核球細胞 (hPBMCs) 產生 IFN-γ、IL-12p70 及抑制 IL-13,進而誘導較強的 Th1 免疫反應並改善 Th1/Th2 免疫反應的平衡;因此我們選擇 Enterococcus faecalis YM-73 菌株,以活菌及熱致死型態菌體餵食經卵白蛋白致敏 (ovalbumin-sensitized) 的小鼠,持續兩個月,探討其抗過敏的影響。餵食期間檢測小鼠血清中總免疫球蛋白 IgE、IgG1、IgG2a 及特異性免疫球蛋白 OVA-specific IgE、IgG1、IgG2a 的含量;另一方面,將小鼠脾臟細胞,使用 ConA、PHA、LPS、OVA 刺激,培養 48 小時後,檢測脾臟細胞的增生反應及分泌 IFN-γ、IL-5的能力;並且探討餵食 Enterococcus faecalis YM-73 對於過敏性鼻炎的影響。實驗結果顯示,熱致死型態乳酸菌較活菌體型態更能夠降低 OVA 致敏小鼠血清中 IgE、IgG1 以及增加 IgG2a 含量,並誘發脾臟細胞分泌更高的 IFN-γ 及抑制 IL-5。而小鼠過敏性鼻炎的模式中,以活菌型態乳酸菌降低過敏的現象較為顯著,熱致死型態乳酸菌則為其次。因此,熱致死菌 Enterococcus faecalis YM-73 可以藉由改善 Th1/Th2 免疫反應的平衡,而達到抗過敏的效果。

Lactic acid bacteria (LAB) are identified as probiotics which can exert a beneficial health effect on the host, including immunomodulation and anti-allergy effects. Currently, their benefits is not only viable cells but also heat-killed treatment LAB exhibit immunomodulatory properties. Heat-killed treatment LAB have the advantages of allowing a longer shelf-life, easier storage, and transportation. According to previous research, we selected most potential immunomodulatory properties of lactic acid bacteria, which were selected for their ability to induce a strong Th1 immune response and improved the Th1/Th2 balance by enhancing IFN-γ and IL-12p70 and inhibiting IL-13 production by human peripheral blood mononuclear cells (hPBMCs) from 11 strains of heat-killed treatment LAB. Based on the results of this screening, we selected Enterococcus faecalis YM-73 as a potent inhibitor of Th2 cytokines production, and investigated the anti-allergic effect of oral adiministration of viable and heat-killed treatment Enterococcus faecalis YM-73 in ovalbumin-sensitized BALB/c mice for two consecutive months. The serum of total IgE, IgG1, IgG2a and OVA-specific IgE, IgG1, IgG2a were determined by sandwich-ELISA assay. We also investigated the effect of the mice splenocyte treated with ConA, PHA, LPS and OVA by measure the vability of splenocyte and production of IFN-γ and IL-5. Furthermore, we investigate the effects of allergic rhinitis on OVA-sensitized BALB/c mice fed with viable and heat-killed treatment Enterococcus faecalis YM-73. The results showed that heat-killed treatment LAB can reduce the IgE, IgG1 and increase IgG2a from serm, enhance the production of IFN-γ and reduce IL-5 from splenocyte more than viable LAB. Viable Enterococcus faecalis YM-73 can reduce the allergic rhinitis symptom more than heat-killed treatment LAB. Thus, heat-killed treatment Enterococcus faecalis YM-73 could be have anti-allergic ability by improve the Th1/Th2 balance.

中文摘要 i
Abstract ii
目次 iv
表目次 vii
圖目次 viii
第一章 前言 1
第二章 文獻探討 3
一、 乳酸菌之保健功效 3
(一) 抑制病原菌入侵 3
(二) 增強腸道黏膜屏障功能 4
(三) 抗腫瘤及抗癌症 4
(四) 降低膽固醇 5
(五) 免疫調節功能 6
二、 活菌與死菌型態乳酸菌 6
三、 過敏性疾病 7
四、 乳酸菌調節免疫的機制 9
第三章 實驗目的 13
第四章 材料與方法 14
一、 材料 14
(一) 實驗菌株 14
(二) 實驗動物 14
(三) 實驗設備 14
(四) 實驗藥品 15
(五) 實驗相關試劑之配製 16
二、 方法 20
(一) 動物試驗流程圖 21
(二) 血液採集及抗體測定 23
(三) 初代脾臟免疫細胞製備及培養 25
(四) 細胞激素之測定(IL-5, IFN-γ) 27
(五) 小鼠過敏性鼻炎模式 28
(六) 統計分析 28
第五章 結果與討論 29
一、 結果 29
(一) 餵食活菌體及熱致死處理 Enterococcus faecalis YM-73對於 OVA 致敏小鼠體重變化之影響 29
(二) 餵食活菌體及熱致死處理 Enterococcus faecalis YM-73對於 OVA 致敏小鼠血清中抗體含量之變化 30
(三) 餵食活菌體及熱致死處理 Enterococcus faecalis YM-73對於 OVA 致敏小鼠臟器重量之影響 36
(四) 餵食活菌體及熱致死處理 Enterococcus faecalis YM-73對於 OVA 致敏小鼠脾臟細胞之增生反應 37
(五) 餵食活菌體及熱致死處理 Enterococcus faecalis YM-73對於 OVA 致敏小鼠脾臟細胞分泌細胞激素之影響 40
(六) 誘導小鼠產生過敏性鼻炎模式之探討 47
二、 討論 48
(一) 活菌低劑量組 49
(二) 活菌中劑量組 50
(三) 活菌高劑量組 51
(四) 熱致死菌低劑量組 52
(五) 熱致死菌中劑量組 53
(六) 熱致死菌高劑量組 55
(七) 比較活菌型態以及熱致死型態減緩過敏的能力 56
第六章 參考文獻 89



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