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研究生:鄭惠敏
研究生(外文):Huimin Cheng
論文名稱:坐骨神經修復後犬腎酸促進廣泛性神經再生之時間特質
論文名稱(外文):Temporal properties for generalized enhancing neuronal regeneration from kynurenic acid after sciatic nerve repair
指導教授:劉念先
指導教授(外文):Nien-Hsien Liou
學位類別:碩士
校院名稱:國防醫學院
系所名稱:生物及解剖學研究所
學門:生命科學學門
學類:生物訊息學類
論文種類:學術論文
論文出版年:2002
畢業學年度:90
語文別:中文
論文頁數:63
中文關鍵詞:神經再生神經修復麩胺酸犬腎酸坐骨神經時間特質
外文關鍵詞:Neuronal regenerationNerve repairGlutamateKynurenic acidSciatic nerveTemporal properties
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根據許多的文獻記載,麩胺酸是影響毒素所導致的神經退化結果和其後進行神經再生過程中的重要因子之一。因此許多學者使用麩胺酸受體的拮抗劑,希望來改善麩胺酸對受損神經所造成更加損傷之程度。所以在我們實驗室已建立的研究模式中,將周邊受損後進行修復手術外,並配合特殊藥物犬腎酸 (kynurenic acid),結果觀察到犬腎酸能更一步促進神經再生程度 (Lee, 1999)。同樣地,承接上述實驗模式,對於坐骨神經修復合併使用犬腎酸模式中發現,在同一時間,對側和術側大白鼠坐骨神經起源部位和目標肌肉腓腸肌間的聯繫有明顯增加的現象;顯示犬腎酸對於調控神經再生扮演一個重要關鍵的角色。因此,在本論文研究,想瞭解在活體實驗中,犬腎酸廣泛性對於坐骨神經修復後促進神經再生的調控機制。在術側方面,大白鼠坐骨神經修復後合併使用犬腎酸 (100mg/kg),分別於術後2週、3週、一個月以及兩個月,將逆行性神經追蹤蔊劑素 (horseradish peroxidase; HRP) 施打至目標肌肉腓腸肌上。結果顯示,隨著時間的延長,在術側L4-L6的背根神經節和脊髓腹角被HRP所標記的神經元有逐漸增加的趨勢。尤其在術後一個月組,開始增加被HRP所標記的神經元數目,表示被截斷神經所造成起源部位和目標肌肉腓腸肌間聯繫中斷開始產生某一程度的改善,而使起源部位和目標肌肉間開始重新恢復聯繫。實驗結果經統計量化分析後顯示,在術後兩個月組中觀察到被HRP所標記的神經元數目與其他三組比較起來有統計意義上的差別。這表示,犬腎酸可能是透過晚期且非同時性機制以促進坐骨神經修復後神經之再生。繼之,承接上述實驗模式,在對側方面正常神經對照組,完整的坐骨神經未進行任何手術,而在術側坐骨神經截斷後縫合以腹腔注射犬腎酸,分別在術後4天、10天、一個月以及兩個月施打HRP至對側目標肌肉腓腸肌。結果顯示,在各組不同時間點上,對側L4-L6的背根神經節和脊髓腹角增加許多被HRP所標記的神經元。實驗結果經統計量化分析後顯示,這四組互相比較起來並沒有具統計意義上之差異。這顯示,犬腎酸可能是透過早期且同時性機制以促進對側正常神經對照組神經再生。綜合以上,在我們設定的實驗模式下,犬腎酸促進兩側性神經再生具有時間差異之特質。

As publicated in the literature, glutamate is one of those key factors affecting outcome from during toxin-induced neurodegeneration and afterwards neuroregeneration process. Hence, ionotropic non-selective glutamate receptor antagonists were utilized as neuroprotectors. Therefore, besides doing nerve repair operation, some endogenous factors of influencing neuroregeneration should be used together to improve the regeneration of damaged nerve. In line with these rationale, we have accomplished enhancing neuronal regeneration from kynurenic acid (KYNA) in sciatic nerve repair rat model (Lee SC, 1999). Extension of those results, connections between the primary origins for sciatic nerve and target muscle gastrocnemius in which the side ipsilateral and contralateral to the nerve repair exhibited dramatic increase; suggesting KYNA playing a critical role in generalized neuroregenerative regulation. To understand the regulatory mechanism of KYNA in vivo, temporal property for enhancing neuronal regeneration after sciatic nerve repair should be taken into consideration. In first set of experiment, rats were divided into four groups such as two weeks, three weeks, one month, and two months after sciatic nerve repair. Two shots of KYNA (100mg/ml/kg) were given under routine procedure. Ipsilateral gastrocnemius were injected with horseradish peroxidase (HRP) during different time points of experiments. Student's t test and ANOVA were used to determine differences among experiments. Results showed progressive escalation in number by HRP-labelled neurons from ipsilateral L4-L6 dorsal root ganglia (DRGs) and spinal ventral horns (SVHs) with increasing time; especially since one month after surgery. Taken together, KYNA can mediate late onset, but not concurrent, enhancing effect for neuronal regeneration after sciatic nerve repair. For second set of experiments, we investigate the temporal property of neuronal regeneration from contralateral side enhanced by KYNA after sciatic nerve repair. Rats were divided into four groups such as four days, ten days, one month and two months after sciatic nerve repair. Prominent increase HRP-labeled neurons from contralateral L4-L6 DRGs and SVHs at the different time points tested. Taken together, KYNA can mediate early onset and concurrent enhancing effect for neuronal regeneration contralaterally after unilateral sciatic nerve repair. These findings indicate there are different patterns of temporal properties for generalized enhancing neuronal regeneration after sciatic nerve repair as compared ipsilateral side with contralateral side.

目錄 ………………………………………………………….………………I
圖次 …………………………………………………………………………IV
中文摘要 …………………………………………………………………V
英文摘要………………………………………………………….……VII
第 一 章 緒言…………………………………………………….………1
第 一 節 前言………………………………….……….…...….1
第 二 節 大白鼠坐骨神經修復實驗模式..……….….…….….1
第 三 節 麩胺酸 (Glutamate)……………….……….…..….7
第 四 節 犬腎酸 (Kynurenic acid )………………….……….12
第 五 節 實驗目的……………………………...…………………14
第 二 章 材料與方法………………………………………….………..15
第 一 節 實驗動物.…………………………..…………….…15
第 二 節 實驗設計組與簡圖………………………..……..….15
壹、實驗分組………………………………………………….15
貳、實驗設計圖……………………………….………………17
第 三 節 手術步驟……………………………..……………...19
壹、對側正常神經對照組………………………….………….…19
貳、術側神經修復後合併使用犬腎酸實驗組……………….20
第 四 節 實驗動物之灌流與固定………………………..….……20
第 五 節 標本製作與切片………………………………………...21
第 六 節 HRP 組織化學反應……………………………………..21
第 七 節 染色、脫水與封片………………………………………22
第 八 節 定量分析與統計方法……………………….…………..22
第 三 章 結果……………………………………………………………24
第 一 節 對側正常神經對照組……………………………………24
第 二 節 術側神經修復後合併使用犬腎酸實驗組…………...24
第 三 節 定量分析…………………………………………………...26
第 四 節 實驗結果…………………………………………………...29
圖 1、實驗結果簡圖…………………………………………………29
圖 2、對側L4-L6背根神經節……………………………………..31
圖 3、對側L4-L6背根神經節統計圖表…………………………33
圖 4、對側L4-L6脊髓腹角…………………………………….…..35
圖 5、對側L4-L6脊髓腹角統計圖表…………………………….37
圖 6、術側L4-L6背根神經節………………………………….…..39
圖 7、術側L4-L6背根神經節統計圖表…………………………41
圖 8、術側L4-L6脊髓腹角…………………………………….…43
圖 9、術側L4-L6脊髓腹角統計圖表……………………….…..45
第 四 章 討論……………………………………………………………47
第 五 章 結論………………………………………………………..…..52
第 六 章 參考資料……………………………………………….….….53

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