剔除ASIC3或TRPV1基因會縮短經發炎產生的長期機械性痛覺敏感現象

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黃郁雰(Yu-Fen Huang) 查詢紙本館藏

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生命科學系

論文名稱

剔除ASIC3或TRPV1基因會縮短經發炎產生的長期機械性痛覺敏感現象
(Deletion of ASIC3 or TRPV1 gene shortens chronic mechanical hyperalgesia induced by inflammation)

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摘要(中)

當組織受傷，受損細胞釋放出多種化學物質，如氫離子、前列腺素E2、生長因子、細胞因子、激肽等。這些發炎介質會透過活化蛋白質激酶A （PKA）或蛋白質激酶Cε （PKCε）使痛覺相關的傷害感受器敏感，進而導致發炎性疼痛。並由之前的研究結果證實急性轉為慢性發炎性疼痛與PKA和PKCε有關。但目前還不清楚是由哪些基因參與調節。也由於氫離子在過去的實驗中有發現是誘發發炎性疼痛很重要的因素，因此認為酸敏感受體於PKA與PKCε轉換之間扮演著重要的角色。為了解決這個問題，我們使用基因剔除或過度表現酸敏感受體的小鼠進行實驗。結果發現無論ASIC3和TRPV1基因剔除小鼠都會縮短CFA引起的機械性痛覺敏感現象。此外，由TRPV1拮抗劑阻斷TRPV1基因也可縮短了CFA誘發的慢性機械性痛覺敏感現象。並於早期階段G2A的過度表現可以透過Gαi路徑降低由
CFA引起的機械性痛覺敏感現象。

摘要(英)

When tissues are injured, damaged cells release multiple chemical mediators, such as protons, PGE2, growth factors, cytokines, kinins and so on. These inflammatory mediators activate protein kinase A (PKA) or protein kinase Cε (PKCε) to sensitize pain-related nociceptor afferents, leading to inflammatory pain. Previous studies have been demonstrated that the transition from acute to chronic inflammatory pain requires the switch of PKA and PKCε-dependency. However, it remains unclear which genes regulate the switch of kinase dependence. Since proton is the decisive factor to induce inflammatory pain, proton-sensing receptors could play an important role in the switch of PKA and PKCε dependency. To address this question, we generated knockout or overexpression mice for proton-sensing receptors. I found that both ASIC3 and TRPV1 knockout mice had shortened CFA-induced mechanical hyperalgesia. Moreover, blocking of TRPV1 by TRPV1 antagonist also shortened CFA-induced chronic mechanical hyperalgesia. Overexpression of G2A reduced CFA-induced mechanical hyperalgesia in early phase through Gαi pathway.

關鍵字(中)

★ 辣椒素受體
★ 酸敏性受體
★ 完全弗氏佐劑

關鍵字(英)

★ TRPV1
★ ASIC3
★ G2A
★ CFA

論文目次

目錄
中文摘要 …………………………………………………………….…………………………... I
英文摘要 ……………………………………………………………………………..…………. II
致謝 ………………………………………………………………………………….………… III
目錄…………………………………………………………………………………………….. IV
圖目錄…………………………………………………………………………………………..VII
表目錄………………………………………………………………………………………….VIII
第一章緒論 1
1.1 慢性疼痛 2
1.2發炎性疼痛 (Inflammatory pain) 3
1.3預發炎模式 (Hyperalgesic priming) 4
1.4離子通道 5
1.4.1 酸敏感離子通道 (Acid-sensing ion channel, ASICs) 5
1.4.2 辣椒素受體 (Transient receptor potential vanilloid channel 1, TRPV1) 6
1.4.3 酸敏感G蛋白偶合受體- OGR1家族 (Proton-sensing G-protein coupled receptor) 7
1.5研究動機 10
第二章實驗材料與方法 11
2.1 實驗材料 12
2.1.1 實驗菌株 12
2.1.2 實驗細胞株 12
2.1.3 實驗動物 12
2.1.4 實驗藥物 12
2.2 實驗方法 13
2.2.1 基因剔除小鼠基因型之辨別 (Genotyping of knockout mice) 13
2.2.2 動物疼痛模式 14
2.2.3 質體於背根神經節表現模式 14
2.2.4 機械性痛覺行為實驗 15
2.2.5 腳掌腫脹程度測量及蘇木素-伊紅染色 (H&E染色) 15
2.2.6 胞內與DRG cAMP累積量分析 16
2.2.7 組織包埋及冷凍組織切片與固定 17
2.2.8 基因分析實驗 18
2.2.9 數據統計分析 19
第三章結果 20
3.1 小鼠皮下注射CFA引發長期發炎性疼痛模式 21
3.2 於野生型小鼠引發發炎反應後，TRPV1與ASIC3基因表現量會持續增加 22
3.3 剔除ASIC3 基因會縮短機械性痛覺持續時間，但發炎情形更加劇烈 22
3.5 剔除TRPV1 基因會縮短機械性痛覺持續時間，但發炎情形並無差異 24
3.6 抑制 TRPV1可以降低野生型小鼠的發炎性疼痛 25
3.7 小鼠腳掌皮下過度表現G2A可以降低pH 5.0 的酸所引起的機械性痛覺敏感現象 26
3.8 小鼠周邊皮下過度表現G2A可以降低CFA所引起的早期機械性痛覺敏感現象 27
3.9 於背根神經節中，在CFA誘發發炎反應中過度表現G2A 27
3.10 於小鼠過度表現G2A會使cAMP累積量下降 28
3.11 小鼠腳掌皮下過度表現G2A可以降低CFA所引起的早期機械性痛覺敏感現象是由Gαi路徑所調控 29
3.12 小鼠周邊皮下過度表現G2A可以降低CFA所引起的早期機械性痛覺敏感現象可能是經由抑制Gαs的路徑而產生的 29
第四章討論 31
第五章參考文獻 38
附錄 71

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指導教授

孫維欣(Wei-Hsin Sun)

審核日期

2015-6-23

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