以電刺激或辣椒素 (capsaicin) 刺激神經可誘發呼吸道神經性發炎。當感覺神經受到刺激造成神經軸突反射可釋放出非膽鹼性神經傳遞物質速激肽 (tachykinin) 而引起發炎。雖然目前相當多文獻指出速激肽會造成呼吸道的神經性發炎包括血漿外洩及平滑肌收縮等現象，但以電刺激胸迷走神經或辣椒素刺激釋放出之非膽鹼性神經傳遞物質所產生之活性氧(reactive oxygen species, 簡稱ROS) 與活性氮 (reactive nitrogen species, 簡稱RNS) 對於呼吸道血漿外洩及平滑肌收縮的機制仍不明確。本研究以電刺激及辣椒素等兩種處理方式探討物質P、ROS與RNS對於呼吸道血漿外洩及平滑肌收縮的機轉。
本研究分成兩個部份，第一部份主要是利用縱膈腔手術進行以阿托品前處理後電刺激右胸迷走神經 (thoracic vagus nerve) 呼吸道之非膽鹼性血漿外洩之調控，並進一步探討電刺激胸迷走神經 (thoracic vagus nerve stimulation, 簡稱TVNS) 是否會經由NK受體，進而使κB核因子 (nuclear factor-kappa B, 簡稱NF-κB) 活化並促進ROS (O2-, H2O2) 產生呼吸道神經性發炎的反應機制；第二部份主要是探討辣椒素可以誘導大鼠體內何種一一氧化氮酶 (nitric oxide synthase, NOS)產生一氧化氮 (nitric oxide, NO)，以及NO在呼吸道血漿外洩及拮抗平滑肌收縮的反應機制。
以前的文獻指出刺激右胸迷走神經會造成氣管下半部、右側主支氣管以及右側次級支氣管大量血漿外洩；第一部份的實驗結果亦驗證此發現。同時結果亦顯示刺激右胸迷走神經會引起血液中物質P釋放增加、血壓降低、氣管收縮、印地安染劑標示氣管血漿滲漏增加等反應。這些反應與增加電刺激頻率(frequency-dependent, 1-10 Hz)成正相關。於電刺激右胸迷走神 (TVNS) 經過30分鐘後血液中超氧離子 (O2-) 和過氧化氫 (H2O2) 等活性氧分子含量有增加的趨勢直到90分鐘為最高。TVNS可活化NF-κB的核因子表現，進而向上調節呼吸道組織之細胞內黏附分子-1 (intercellular adhesion molecule-1, 簡稱ICAM-1) 的總蛋白表現。組織病理學證明TVNS後，右下側支氣管銀染色法產生內皮隙之銀點 (silver dot)、發炎細胞聚集等情形。L-732138 (NK1受體拮抗劑)、SR-48968 (NK2受體拮抗劑)、二甲基硫尿素 (dimethylthiourea, 過氧化氫清除劑) 或兒茶素 (catechins, 氧離子和過氧化氫清除劑) 前處理會降低因TVNS所產生低血壓、氣管收縮以及血漿滲漏情形。NK受體拮抗劑與抗氧化劑可以減弱NF-κB及ICAM-1的向上調節作用 (upregulation)。
第二部份的實驗結果顯示注射辣椒素後會引起血壓降低、氣管收縮以及血漿外滲等反應，同時血漿中一氧化氮濃度隨辣椒素劑量的增加而增加。以L-732138 (NK1受體拮抗劑) 或SR-48968 (NK2受體拮抗) 前處理， 能舒緩辣椒素所引起的低血壓、氣管收縮及氣管血漿滲漏的情形，也能抑制血漿中一氧化氮的濃度。L-NAME (非選擇性一氧化氮合成酶抑制劑) 或aminoguanidine (選擇性的誘導性一氧化氮合成酶 iNOS抑制劑)，兩者能降低由辣椒素引發的血漿一氧化氮濃度升高的現象，並且能防止辣椒素誘發的血漿外滲。L-Arginine (一氧化氮前驅物) 能提高辣椒素誘發的血漿一氧化氮濃度，並增加血漿外滲的情形。以L-Arginine前處理，能改善辣椒素所誘發的氣管收縮，而L-NAME及Aminoguanidine則會加劇辣椒素誘發氣管收縮。
總結以上結果得到下列結論，(1) 電刺激胸迷走神經可促進物質P釋放，增加呼吸道平滑肌的作用、呼吸道阻力、內皮細胞間隙血漿滲漏、發炎細胞聚集，以及產生超氧離子與過氧化氫作用所引起之NF-κB與ICAM-1蛋白質表現；(2) 辣椒素刺激呼吸道感覺神經釋出速激肽，這些物質會作用在呼吸道平滑肌及血管的NK受體，釋出的速激肽會透過 iNOS 增加NO產量，而iNOS合成的NO能減緩速激肽主導的氣管收縮，但會加劇血漿外滲。

Neurogenic inflammatory responses can be induced by antidromic electrical stimulation or intravenous capsaicin injection. These responses were thought to be caused by neuropeptides released from the sensory axon of C-fiber nerve endings. The relation of tachykinins, reactive oxygen species (ROS) and reactive nitrogen species (RNS) on electrical stimulation of thoracic vagus nerve (TVNS) or capsaicin-evoked neurogenic inflammation in respiratory tract of atropine-treated rats was not clear. In the present studies, the role of ROS and RNS on neurogenic inflammation were investigated in TVNS and capsaicin injected rats.
The experiments were divided into two parts. In the first part, TVNS was performed by thoracotomy, non-cholinergic regulation of neurogenic plasma extravasation in the trachea and bronchi were examined, and whether TVNS via NK receptor facilitates neurogenic inflammation by nuclear factor-kappaB (NF-κB) activation and ROS production were expored. Our results in this part showed that TVNS evoked substance P release, hypotension, bronchoconstriction (as shown by increases in smooth muscle electromyographic activity and total pulmonary resistance), trachea plasma extravasation as well as increases in blood O2- and H2O2 ROS amount in a frequency-dependent manner. Histopathological examination demonstrated silver-stained leaky venules, India-ink labeled plasma extravasation, and accumulations of inflammatory cells in the right lower trachea after TVNS. L-732138 (NK1 receptor antagonist), SR-48968 (NK2 receptor antagonist), dimethylthiourea (H2O2 scavenger) or catechins (O2- and H2O2 scavenger) pretreatment reduced TVNS-enhanced hypotension, bronchoconstriction, and plasma extravasation. TVNS upregulated the expression of NF-κB in nuclear protein and intercellular adhesion molecule-1 (ICAM-1) in total protein of the lower respiratory tract tissue in a frequency-dependent manner. The upregulation of NF-κB and ICAM-1 was attenuated by NK receptor antagonist and antioxidants. In the second part, the contribution of nitric oxide (NO) to capsaicin-evoked airway responses was investigated in rats. The measurement of plasma NO level, airway dynamics, airway smooth muscle electromyogram, and plasma extravasation by India ink and Evans blue leakage technique was adapted. Our results in this part showed that capsaicin injection evoked hypotension, bronchoconstriction, trachea plasma extravasation as well as increases in plasma NO level in a dose-dependent manner. L-732138 or SR-48968 pretreatment reduced capsaicin-enhanced hypotension, bronchoconstriction, plasma extravasation, and plasma NO level. Inhibition of a non-selective NO synthase (NOS) inhibitor (NG-nitro-L-Arginine methyl ester, L-NAME), or a selective inducible NO synthase (iNOS) inhibitor (aminoguanidine), reduced capsaicin-induced increases in plasma NO level and protected against capsaicin-induced plasma extravasation, whereas L-arginine (a NO precursor), enhances capsaicin-evoked plasma NO level and plasma extravasation. L-Arginine pretreatment ameliorated capsaicin-induced bronchoconstriction, whereas L-NAME and aminoguanidine exaggerated capsaicin-induced bronchoconstriction.
In summary, both TVNS and capsaicin injection may increase oxidative stress responses. TVNS enhances proinflammatory NF-κB and ICAM-1 expression, increases the production of O2- and H2O2 activity in the respiratory tract of atropine-treated rats. Pretreatment with antioxidants and selective NK receptor antagonists attenuate TVNS evoked airway hyperactivity, proinflammatory response, and oxidative stress. Capsaicin injection stimulates the release of tachykinins, which act on NK1 and NK2 receptors located on the smooth muscles of airways and blood vessels. The interaction of NK receptors with tachykinin enhances furtherly the NO formation, bronchoconstriction, vasodilation, and plasma extravasation in the trachea. The released tachykinins also increase the production of NO via iNOS, and iNOS -evoked NO counteracts tachykinin-mediated bronchoconstriction, but exacerbates tachykinin-mediated plasma extravasation.

【目錄】
頁次
致謝……………………………………………………………………Ⅰ
中文摘要 ………………………………………………………………Ⅱ
英文摘要………………………………………………………………Ⅳ
目錄 ……………………………………………………………………Ⅵ
表次 ……………………………………………………………………Ⅷ
圖次 ……………………………………………………………………Ⅸ
文獻回顧
一、前言 ………………………………………………………………1
二、呼吸道之神經生理系統 …………………………………………1
三、誘發神經性發炎之物質與方法 …………………………………2
四、神經性發炎誘發與抑制之機轉 …………………………………6
五、誘發呼吸道神經性發炎的兩種動物實驗架構 …………………7
六、呼吸道之神經性發炎病理變化 …………………………………9
七、活性氧自由基對於呼吸系統之影響……………………………11
八、活性氮自由基對於呼吸系統之影響……………………………16
九、抗氧化酵素………………………………………………………20
研究目的
一、探討電刺激胸迷走神經對於呼吸系統產生氧化壓力的機制……24
二、探討一氧化氮在辣椒素誘發呼吸道發炎反應的過程中所扮演的角色………………………………………………………24
材料方法
一、實驗動物 …………………………………………………………25
二、藥物配置 ………………………………………………………25
三、各類插管 ………………………………………………………26
四、呼吸道生理學之計量 …………………………………………27
五、呼吸道平滑肌信號測定 ………………………………………27
六、資料分析 ………………………………………………………28
七、電刺激胸迷走神經處理………………………………………29
八、電刺激及藥物前處理誘發神經性發炎程度之測定…………30
九、物質P之測量…………………………………………………32
十、血液中自由基含量檢測定…………………………………32
十一、NFκB與ICAM-1之生化測定…………………………………33
十二、統計方法……………………………………………………35

結果
一、電刺激大鼠胸迷走神經引起呼吸道神經激素受體依賴型神經性發炎反應及氧化壓力 ……………………………………………………36
二、誘導性一氧化氮合成酶合成的一氧化氮可中和辣椒素誘發的呼吸道平滑肌收縮及血漿滲漏的現象 ……………………………………38
討論
一、電刺激胸迷走神經系統對於呼吸道血漿外洩之調控與機轉40
二、辣椒素誘發呼吸道神經性發炎產生一氧化氮拮抗作用之機制………………44
結論…………………………………………………………………… 48
參考文獻 ………………………………………………………………49
表次 …………………………………………………………………66
圖次 ……………………………………………………………………72

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