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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 朱瑞民 | |
dc.contributor.author | Cheng-Chi Liu | en |
dc.contributor.author | 劉正吉 | zh_TW |
dc.date.accessioned | 2021-06-13T04:27:45Z | - |
dc.date.available | 2006-07-27 | |
dc.date.copyright | 2006-07-27 | |
dc.date.issued | 2006 | |
dc.date.submitted | 2006-07-20 | |
dc.identifier.citation | 2000, Translational Research: the Role of VEGF in Tumor Angiogenesis. Proceedings of a symposium. Washington, DC, USA. November 16, 1999. Oncologist 5 Suppl 1, 1-57.
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/33171 | - |
dc.description.abstract | 腫瘤經常以樹突狀細胞(dendritic cell, DC) 為目標,進而對DC造成傷害而藉此逃避生物體免疫系統的辨識。在小鼠和人類許多的腫瘤研究中都已經發現腫瘤會造成DC的傷害。一般腫瘤會透過下述幾種機制來影響及改變DC的活性,例如抑制DC前驅物的分化、壓制其成熟化反應的發生、妨礙吞噬及表現抗原能力的表現,或者造成DC的死亡。腫瘤通常會利用上述一種或多種不同的機制來促進腫瘤細胞的生長。
在我們的研究中發現犬傳染性花柳性腫瘤 (canine transmissible venereal tumor, CTVT) 也會透過所有上述的機制對於單核球來源的DC(monocyte-derived DC, Mo-DC)造成嚴重的傷害。我們將正常犬與接種腫瘤後所得到的Mo-DC,利用流式細胞分析儀(flow cyto- meter)及real-time PCR,比較其表面抗原表現,包括CD1a、CD83,共同刺激因子(costimulatory factors,如CD40,CD80和CD86) 以及MHC class I與II分子。由實驗中發現無論是未成熟的DC (immature DC, iDC)或者是透過脂多醣(lipopolysaccride, LPS)刺激活化的DC(LPS-treated mature DC, mDC),其表面抗原的呈現,在腫瘤的影響下,無論是生長期(progression phase,P期)或消退期(regression phase,R期),都比正常犬低。除表面抗原呈現外,DC的功能也受到腫瘤嚴重的破壞。在P期時發現iDC吞噬dextran的活性明顯比正常犬為低。而在同種異體淋巴球反應(Allogeneic mixed lymphocyte reaction, MLR)中也發現mDC刺激淋巴球活化的能力在腫瘤的影響下明顯受到抑制。此外,在CTVT的作用下我們發現犬隻周邊血液單核球細胞(peripheral blood mononuclear cell, PBMC)中單核球(monocytes)的數量減少約40%。而且在單核球分化為iDC或者是每顆iDC轉化成mDC的比例中,腫瘤接種犬都明顯比正常犬為低。有趣的是,在R期,受到抑制的DC活性似乎大部分已恢復。無論是mDC的數量、表面抗原的表現、抗原吞噬能力及MLR的表現都有明顯恢復的現象。我們相信應該是有一種未知的機制在腫瘤進入R期時促使Mo-DC的功能逐漸回復。 於研究中也發現來自P期的腫瘤細胞上清液在與單核球及DC共同培養48小時後,發現單核球及DC發生細胞凋亡(apoptosis)的現象。於現今的文獻中鮮少有研究指出一種腫瘤能如此廣泛的傷害Mo-DC。綜合上述所言,可以確定CTVT會造成DC發生自體衰亡的現象、抑制單核球分化成為iDC、阻止iDC轉化為mDC並且阻礙DC許多功能的表現。這種種對DC的傷害,都促使腫瘤逃避犬隻免疫系統的辨識。這些發現可以提供我們瞭解腫瘤對於免疫系統的影響,並藉此發展有效的腫瘤免疫治療的策略。 再者,近年來根據小鼠及人類NK細胞(nature killer cell, NK cell)及DC的研究中,發現二者之間的互動對於免疫反應的進行有明顯的影響,但對於犬隻相關文獻探討較少。本研究中另一目的是針對犬隻NK細胞及DC間的交互作用進行研究。在觀察NK細胞的細胞毒殺能力中發現,無論是休止型或者是以IL-2(Interleukin 12)活化後的LAK(lymphokine- activated killer cell)細胞,在CTVT的影響下,細胞毒殺能力都明顯降低。雖然在R期NK細胞的毒殺能力較P期為佳,但仍然明顯受到抑制。觀察在CTVT的生長時可以發現,無論在P期或R期,mDC的CD80,CD83及CD86表現會明顯下降。雖然CTVT影響DC的成熟,但DC在與NK細胞共同培養的作用時發現,NK細胞卻能協助DC的成熟。最後觀察細胞激素(cytokine)的變化,本實驗中以單獨未受任何刺激的DC作為對照組,發現在DC與NK細胞共同培養時,產生大量IL-12與IL-18,此種細胞激素會活化NK細胞。當CTVT生長於P期與R期時,除了iDC與mDC本身分泌的細胞激素量就比正常少外,與NK細胞共同培養後,雖然mDC所分泌的IL-12、IL-18、IFN-γ與TNF-α會顯著上升,但總量仍較正常明顯為少。但在R期時IL-12、IL-18、IFN-γ與TNF-α的分泌量都明顯比P期為多。 綜合以上所述,CTVT對DC明顯造成傷害,包括抑制單核球分化成iDC、壓制mDC成熟化反應的發生、妨礙iDC吞噬功能及mDC表現抗原能力的表現,及造成單核球與iDC的細胞凋亡。此外,CTVT亦影響NK細胞其細胞毒殺能力。同時進一步發現因為DC的影響間接刺激NK細胞的活化,反之亦同。因此,NK細胞與DC在腫瘤之間的交互作用,在CTVT的模式中再一次得到證實。 | zh_TW |
dc.description.abstract | Tumors often target dendritic cells (DCs) to evade host immune surveillance. DC injury has been reported in many tumors in rodents and humans. They exert systemic impact by altering the activities of DCs through the following mechanisms: inhibition of differentiation, suppression of maturation, impediment of functions, or killing of the cells. Tumors usually employ one or more of these mechanisms to facilitate their progressive growth.
We reported here that canine transmissible venereal tumor (CTVT) induced devastating effects on monocyte-derived DCs through all of the mechanisms mentioned above. Using flow cytometry and real-time RT-PCR, the expression of the surface markers of monocyte-derived DCs, including CD1a, CD83, costimulatory factors (CD40, CD80, and CD86), and MHC classes I and II were compared between normal dogs and canine transmissible venereal tumor (CTVT) dogs. It was found that the tested surface markers were all or mostly lower in concentrations in both immature DCs (iDCs) and LPS-treated mature DCs (mDCs) from tumor dogs during the progression (P) or the regression (R) phase than they were in normal dogs. The functions of the DCs were also seriously damaged by the tumor. The iDCs from dogs with P-phase tumors performed significantly lower endocytotic activity on dextran uptake than the normal dogs. Allogeneic mixed-lymphocyte reaction (MLR) was much lower in the mDCs from tumor dogs. In addition, the tumor decreased the number of monocytes in the peripheral blood by 40%. The efficiency of DC generation per monocyte or mDC generation per iDC was much lower in tumor dogs. Interestingly, during R phase, the inhibited DC activities were partially recovered, as evidenced by the elevated number of the mDCs and higher expression of the surface markers, plus the better dextran uptake and MLR activities. It is believed that the monocyte-derived DCs re-established by the host through an unknown mechanism contributed to the regression of the tumor. We also found that the 48h cultured P phase CTVT supernatants containing protein components killed both monocytes and DCs by apoptosis. Very few investigation reported tumor-caused damage on monocyte-derived DCs with such a wide spectrum. It is concluded that CTVT killed the DCs through apoptosis, inhibited the differentiation of the cells from monocytes and from iDCs to mDCs, and restrained the functions of the DCs. These caused devastating damage to the DCs and enabled the tumor to evade host immunity. These findings provide further understanding of the tumor effects on the host immune system and are useful in developing strategies for cancer immune therapy. In mouse and humans, the NK cells (nature killer cells, NK cells) and DC interactions play an important role in moderating innate immune responses. In dogs, this interaction has rarely been studied. Thus, another goal of this research was to understand the interactions between NK cells and DCs in dogs. The results showed that the growth of the CTVT significantly hampered the cytotoxicity of the resting and IL-2 activated killer cells. Although the cytotoxicity of the NK cells during R phase was improved, the suppression of the activity was still obvious. The mRNA expression of CD80, CD83, and CD86, the markers for DC maturation, was also lower during the growth of CTVT. These indicated that CTVT also inhibited the maturation of DCs. The NK-DC interaction studies showed that NK cells apparently helped the maturation of mDCs from iDCs in dogs. On the other hand, during normal NK-DC co-culturing in dogs, IL-12 and IL-18 were produced in high concentration that were much higher than those cultured alone. These cytokines were potent cytokines to activate NK cells. In addition, the results also provided evidence that CTVT inhibited the general production of the cytokines. Furthermore, although the production of IL-12, IL-18, IFN-g, and TNF-a were increased during NK-DC co-culturing from CTVT dogs, it was still significantly lower than normal dogs without CTVT. Similarly, R-phase dogs showed some recovery of cytokine production. To summarize, the diversifying injury due to the growth of CTVT caused devastating damage to DCs, including the inhibition of differentiation from monocytes to iDCs and iDCs to mDCs, the suppression of DC maturation, the impairment of DC functions such as antigen uptake and MLR activities, and the apoptosis of monocytes and DCs. In addition, this tumor also suppressed the NK cytotoxicity and hampered the interactions between DC and NK cells. Thus, CTVT is an ideal cancer model for studying the complex interrelationships between tumor growth and host reactions. | en |
dc.description.provenance | Made available in DSpace on 2021-06-13T04:27:45Z (GMT). No. of bitstreams: 1 ntu-95-R91629011-1.pdf: 3053318 bytes, checksum: d606d24cde25d84923be17d80580f74c (MD5) Previous issue date: 2006 | en |
dc.description.tableofcontents | 中文摘要••••••••••••••••••••••••VI
英文摘要•••••••••••••••••••••••••X 圖次••••••••••••••••••••••••••XIV 表次••••••••••••••••••••••••••XVI 縮寫符號•••••••••••••••••••••••XVII 中英對照••••••••••••••••••••••••XIX 第一章 緒言•••••••••••••••••••••••1 第二章 文獻探討•••••••••••••••••••••4 第一節 腫瘤研究模式---犬傳染性花柳性腫瘤之相關探討••4 一、 犬傳染性花柳性腫瘤的病理特徵•••••••••4 二、 犬傳染性花柳性腫瘤的細胞特性•••••••••5 三、 犬傳染性花柳性腫瘤與免疫系統之影響••••••7 第二節 樹突狀細胞在抗腫瘤免疫反應中的角色•••••11 一、 樹突狀細胞的特性••••••••••••••11 二、 樹突狀細胞在生物體內的來源與分佈••••••13 三、 樹突狀細胞在抗腫瘤免疫反應中的表現•••••14 四、 目前犬樹突狀細胞的研究與發現••••••••18 第三節 自然殺手細胞在抗腫瘤免疫反應中的角色••••20 一、 自然殺手細胞的特性•••••••••••••20 二、 自然殺手細胞在生物體內的來源與分佈•••••23 三、 自然殺手細胞在抗腫瘤免疫反應中的表現••••24 四、 目前犬自然殺手細胞的研究與發現•••••••28 第四節 樹突狀細胞與自然殺手細胞間的交互作用對於抗腫瘤免疫反應的影響•••••••••••••••31 一、 樹突狀細胞與自然殺手細胞間的交互作用••••31 二、 對於腫瘤的影響•••••••••••••••34 第三章 實驗目標•••••••••••••••••••••39 第四章 材料與方法••••••••••••••••••••40 第一節 犬傳染性花柳性腫瘤腫瘤模式的建立••••••40 一、 實驗動物••••••••••••••••••40 二、 犬傳染性花柳性腫瘤的分離與接種•••••••40 三、 犬傳染性花柳性腫瘤與腫瘤浸潤淋巴球的純化與收 集•••••••••••••••••••••40 第二節 犬單核球來源的樹突狀細胞(Monocyte derived dendritic cell, Mo-DC)的取得••••••••••••••42 第三節 犬自然殺手細胞的取得••••••••••••43 一、 周邊血液淋巴球 (peripheral blood lymphocytes, PBL) 的分離•••••••••••••••••••43 二、 利用磁珠分離系統純化自然殺手細胞••••••43 第四節 細胞表面抗原分析••••••••••••••45 一、 流式細胞分析儀•••••••••••••••45 二、 Real-Time PCR•••••••••••••••• 47 第五節 犬傳染性花柳性腫瘤對樹突狀細胞及自然殺手細胞的影響••••••••••••••••••••48 一、 犬傳染性花柳性腫瘤細胞培養後之上清液對樹突狀細胞的影響••••••••••••••••••48 二、 樹突狀細胞吞噬抗原能力測試•••••••••49 三、 樹突狀細胞表現抗原能力測試:混合淋巴球反應••50 四、 自然殺手細胞之細胞毒殺作用•••••••••50 五、 樹突狀細胞與自然殺手細胞之互動下樹突狀細胞成熟化的反應••••••••••••••••••52 六、 樹突狀細胞與自然殺手細胞之互動下自然殺手細胞細胞毒殺作用的變化••••••••••••••52 七、 樹突狀細胞與自然殺手細胞之互動下細胞激素的變化•••••••••••••••••••••53 第六節 統計分析••••••••••••••••••55 第五章 結果•••••••••••••••••••••••56 第一節 犬傳染性花柳性腫瘤模式的建立••••••••56 一、 腫瘤細胞的人工接種與採樣••••••••••56 二、 腫瘤細胞表面抗原的表現與腫瘤浸潤淋巴球的細胞族群變化•••••••••••••••••••57 第二節 犬傳染性花柳性腫瘤細胞之上清液對於犬樹突狀細胞的毒殺作用•••••••••••••••••60 第三節 不同腫瘤生長期對周邊血液細胞數量的影響•••62 第四節 PBMC、單核球、與樹突狀細胞的細胞表面抗原表現狀況•••••••••••••••••••••64 第五節 犬傳染性花柳性腫瘤的生長對於樹突狀細胞吞噬作用的影響•••••••••••••••••••68 第六節 犬傳染性花柳性腫瘤的生長影響同種異體之混合淋巴球反應•••••••••••••••••••70 第七節 自然殺手細胞細胞毒殺作用••••••••••72 一、 自然殺手細胞表面抗原的表現•••••••••72 二、 犬傳染性花柳性腫瘤影響自然殺手細胞的毒殺能力•••••••••••••••••••••74 第八節 犬傳染性花柳性腫瘤對樹突狀細胞與自然殺手細胞交互作用之影響••••••••••••••••75 一、 犬傳染性花柳性腫瘤的生長對自然殺手細胞促進樹突狀細胞之成熟的影響•••••••••••••75 二、 樹突狀細胞對於自然殺手細胞在犬傳染性花柳性腫瘤的影響下之細胞毒殺能力的變化••••••••78 三、 NK-DC共同培養下的細胞激素分泌狀況•••••81 第六章 討論•••••••••••••••••••••• 84 第七章 參考文獻•••••••••••••••••••••94 附錄•••••••••••••••••••••••••••A 一、 犬傳染性花柳性腫瘤與腫瘤浸潤淋巴球的純化與收集•A 二、 犬單核球來源的樹突狀細胞(Monocyte derived dendritic cell, Mo-DC)的取得••••••••••••••C 三、 周邊血液淋巴球 (peripheral blood lymphocytes, PBL) 的分 離••••••••••••••••••••••••F 四、 利用磁珠分離系統純化自然殺手細胞••••••••G 五、 自然殺手細胞之細胞毒殺作用••••••••••I | |
dc.language.iso | zh-TW | |
dc.title | 犬傳染性花柳性腫瘤對樹突狀細胞及與自然殺手細胞相互作用的影響 | zh_TW |
dc.title | The effects of canine transmissible venereal tumor on dendritic cells and the interactions between dendritic cells and nature killer cells | en |
dc.type | Thesis | |
dc.date.schoolyear | 94-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 季匡華,陶秘華,張天傑,廖光文 | |
dc.subject.keyword | 犬傳染性花柳性腫瘤,生長期,P期,消退期,R期,周邊血液單核球細胞,單核球來源的DC,未成熟的樹突狀細胞,iDC,成熟的樹突狀細胞,mDC,混合淋巴細胞反應, | zh_TW |
dc.subject.keyword | Canine transmissible venereal tumor, CTVT,progression phase, P phase,regression phase, Rphase,peripheral blood mononuclear cell, PBMC,monocyte-derived DC, Mo-DC,immature dendriric cells, iDC,mature dendritic cell, mDC,mixed lymphocyte reaction, MLR, | en |
dc.relation.page | 147 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2006-07-22 | |
dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
dc.contributor.author-dept | 獸醫學研究所 | zh_TW |
顯示於系所單位: | 獸醫學系 |
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