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研究生:葉宗樹
研究生(外文):Tsung-Shu Yeh
論文名稱:綠梔子素交聯明膠充填矽膠管於修護截斷大鼠坐骨神經再生影響之評估
論文名稱(外文):Assessment of silicone tube filled with genipin cross-linking gelatin on regeneration of severed rat sciatic nerves
指導教授:陳悅生
指導教授(外文):Yueh-Sheng Chen
學位類別:碩士
校院名稱:中國醫藥學院
系所名稱:中國醫學研究所
學門:醫藥衛生學門
學類:醫學學類
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:94
中文關鍵詞:周圍神經再生矽膠管膠原蛋白明膠綠梔子素
外文關鍵詞:peripheral nerveregenerationsilicone tubecollagengelatingenipin
相關次數:
  • 被引用被引用:9
  • 點閱點閱:234
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周邊神經斷傷後,若沒有適當的處理,可能會造成神經瘤形成,使得神經無法順利再生。而在選用何種神經接合術來修補斷傷的周邊神經時,神經兩斷端的間距是最重要的考慮因素。神經斷端縫合術和神經束縫合術適用於神經較短斷裂間距之修補,但當神經斷傷間距太長時,神經移植術和神經管接合術是較適當的手術方法,但因神經移植段來源取得的困難,於是利用神經管接合術來修補較長的神經缺損成為較佳的選擇。
神經管接合術是將神經兩斷端手術縫合於神經管的兩端,並利用神經管來引導及支持神經纖維的再生。文獻中已經有多種不同的材料用來製作神經管,矽膠管就是常用的一種。另外,在神經管內添加刺激神經再生的物質,使再生神經在較短的時間內,跨過神經間距完成再生。而常用的神經再生刺激物質有神經生長因子、膠原蛋白、laminin 與fibronectin混合物等。
過去臨床上膠原蛋白常被用來充填神經管,作為刺激神經再生的物質,但因其製備過程較繁雜且價格昂貴。所以本研究以矽膠管分別充填明膠(gelatin)與經綠梔子素(genipin)交聯之明膠,並利用此神經管對截斷1 cm的大鼠坐骨神經做一接合,以觀察其對大鼠周圍神經再生的影響,並與矽膠管充填膠原蛋白和空管組做一比較。
實驗結果:一、神經再生成功率:空管組30﹪、膠原蛋白組90﹪、明膠組80﹪、綠梔子素交聯明膠組80﹪。二、再生神經組織切片比較:空管組有較多的軸突數(p<0.001),形態最為成熟;膠原蛋白組、明膠組、與綠梔子素交聯明膠組其再生軸突數雖較少,但軸突平均面積卻都比空管組大。實驗結果顯示明膠與綠梔子素交聯明膠和膠原蛋白一樣,都可促進大鼠坐骨神經再生軸突的成熟,但殘留於神經管內的膠狀物可能會影響軸突再生的過程,使得軸突再生數目相對於空管組減少了許多。
If the neurotmesis is not properly treated, the neuroma can happen. The gap length between the injured nerve ends is the most important factor in deciding the suturing methods of repairing severed peripheral nerve. End-to-end suturing and fascicular suturing methods are suitable to the shorter gaps. As for the large gaps, nerve grafting and nerve bridging methods are suggested. It is difficult to get the original source of nerve grafting method so that it is better choice to use nerve bridging method to repair the large gap of both ends of severed nerve.
Nerve bridging technique is to put both ends of severed nerves in a tube, which can guide and support nerve fiber regeneration. There are many kinds of materials used for nerve guide tubes, such as the silicone tube which is most commonly used. To make it possible for the regenerating nerve across a longer gap in a shorter time, the nerve guide tube filled with nerve stimulants are used. The nerve stimulants commonly used include the mixture of nerve growth factor, collagen, laminini, and fibronectin.
Collagen is another commonly used stimulants, however it also has several disadvantages, such as the complicated preparation procedures and high price. In this study, we therefore used the pure gelatin and the genipin cross-linking gelatin as the fillings in the silicone tubes to repair dissected rat sciatic nerves with 10 mm gaps. Nerves regenerating in both the fillings were then compared to those in the tubes loaded with collagen as well as the empty tubes.
Experiment results: (1) The success rate of nerve regeneration: 30% in empty tube, 90% in tube with collagen, 80% in tube with gelatin, 80% in tube with genipin cross-linking gelatin. (2) Regenerated nerve section comparison: the regenerated nerve in the empty tube was most mature and had more axons (p<0.001). The rest 3 groups had less axon but their average areas are bigger than that of the empty tube. These results show that gelatin, genipin cross-linking gelatin, and collage could enhance the maturation of regenerated rat sciatic nerves. However, their residues in the tubes could impair the regenerating axons, thus decreasing their numbers successfully crossing the nerve gaps.
目錄
中文摘要-------------------------------------------------------------------------------1
1 前言----------------------------------------------------------------------------------1
1.1實驗目的---------------------------------------------------------------------2
2文獻探討-----------------------------------------------------------------------------3
2.1神經系統介紹---------------------------------------------------------------3
2.1.1中樞神經系統------------------------------------------------------3
2.1.2周圍神經系統------------------------------------------------------4
2.1.3神經元---------------------------------------------------------------4
2.1.3.1神經元的類型---------------------------------------------4
2.1.3.2神經元的構造---------------------------------------------5
2.1.3.3神經元的功能---------------------------------------------8
2.1.4神經纖維------------------------------------------------------------9
2.1.4.1神經纖維分類---------------------------------------------9
2.1.4.2神經纖維的傳導-----------------------------------------10
2.2周圍神經損傷-------------------------------------------------------------11
2.2.1周圍神經損傷的原因--------------------------------------------11
2.2.2 Seddon分類法----------------------------------------------------11
2.2.2.1神經失用-------------------------------------------------11
2.2.2.2軸突斷傷-------------------------------------------------12
2.2.2.3神經斷傷-------------------------------------------------12
2.2.3 Sunderland分類法-----------------------------------------------12
2.2.4神經元傷害後之變化--------------------------------------------12
2.2.5神經元傷害後之復原--------------------------------------------13
2.3周圍神經斷傷的修復----------------------------------------------------15
2.3.1各種神經修補技術-----------------------------------------------15
2.3.2以神經管接合術為實驗之模式--------------------------------16
2.3.2.1神經管材料之選擇--------------------------------------17
2.3.2.2神經間距--------------------------------------------------18
2.3.2.3刺激神經再生的物質-----------------------------------18
2.3.2.4 Schwann細胞--------------------------------------------20
2.3.2.5電刺激-----------------------------------------------------20
2.3.3矽膠管內神經再生之細胞學變化-----------------------------20
2.3.3.1液體堆積--------------------------------------------------20
2.3.3.2纖維橋的形成--------------------------------------------21
2.3.3.3纖維母細胞移行-----------------------------------------21
2.3.3.4 Schwann細胞移行--------------------------------------21
2.3.3.5血管芽的形成--------------------------------------------22
2.3.3.6再生單元和Schwann細胞柱-------------------------22
2.3.3.7髓鞘化-----------------------------------------------------22
2.4神經損傷的中醫觀點-----------------------------------------------------23
2.4.1神經系統在中醫學中的定位-----------------------------------23
2.4.2周圍神經與經脈之關係-----------------------------------------26
2.4.3周圍神經斷傷與相關之中醫病証-----------------------------27
2.4.3.1創傷、金瘡-----------------------------------------------27
2.4.3.2骨折、脫位-----------------------------------------------29
2.4.3.3傷筋--------------------------------------------------------30
2.4.3.4痿病與神經斷傷-----------------------------------------31
2.4.3.5小結--------------------------------------------------------32
2.5梔子與綠梔子素(genipin)------------------------------------------------33
2.5.1梔子------------------------------------------------------------------33
2.5.1.1來源--------------------------------------------------------33
2.5.1.2採製--------------------------------------------------------33
2.5.1.3歷代本草記載功效--------------------------------------33
2.5.1.4成分--------------------------------------------------------34
2.5.1.5藥理作用--------------------------------------------------34
2.5.1.6臨床應用--------------------------------------------------35
2.5.1.7臨床使用應注意事項-----------------------------------36
2.5.2綠梔子素(genipin)-------------------------------------------------37
2.5.2.1生物膠-----------------------------------------------------37
2.5.2.2交聯劑-----------------------------------------------------37
2.5.2.3 genipin的相關研究-------------------------------------37
3材料與方法-------------------------------------------------------------------------40
3.1實驗材料-------------------------------------------------------------------40
3.1.1矽膠管---------------------------------------------------------------40
3.1.2 Collagen------------------------------------------------------------40
3.1.3 Gelatin--------------------------------------------------------------40
3.1.4 Genipin--------------------------------------------------------------40
3.1.5 Toluidine blue------------------------------------------------------40
3.2神經管的配製-------------------------------------------------------------41
3.2.1矽膠管---------------------------------------------------------------41
3.2.2配製內填試劑的矽膠管與分組---------------------------------41
3.2.3試劑注入矽膠管內------------------------------------------------42
3.3實驗動物模式--------------------------------------------------------------42
3.3.1實驗動物分組-----------------------------------------------------42
3.3.2麻醉------------------------------------------------------------------42
3.3.3坐骨神經神經管接合術------------------------------------------42
3.3.4動物飼養環境------------------------------------------------------43
3.3.5觀察實驗大鼠外表之變化---------------------------------------43
3.3.6觀察神經再生情形------------------------------------------------43
3.4組織切片與染色----------------------------------------------------------44
3.4.1觀察切片-----------------------------------------------------------44
3.5組織學定量分析----------------------------------------------------------45
3.5.1拍攝照片-----------------------------------------------------------45
3.5.2神經全部面積的計算方式--------------------------------------45
3.5.3神經內膜面積的計算方式--------------------------------------45
3.5.4神經軸突數目的計算方式--------------------------------------45
3.5.5神經軸突平均面積的計算方式--------------------------------46
3.5.6神經軸突密度的計算方式--------------------------------------46
3.5.7神經軸突面積百分比的計算方式------------------------------46
3.5.8再生血管數目的計算方式---------------------------------------46
3.5.9再生血管總面積的計算方式------------------------------------46
3.5.10再生血管所佔百分比的計算方式----------------------------46
3.6統計方法-------------------------------------------------------------------46
4結果----------------------------------------------------------------------------------47
4.1大白鼠外觀----------------------------------------------------------------47
4.1.1行動方面-----------------------------------------------------------47
4.1.2毛色方面-----------------------------------------------------------47
4.1.3傷口方面-----------------------------------------------------------47
4.1.4重量變化-----------------------------------------------------------47
4.1.5足趾自殘-----------------------------------------------------------47
4.1.6足跟傷口-----------------------------------------------------------48
4.2矽膠管觀察與神經再生情形-------------------------------------------51
4.3切片觀察-------------------------------------------------------------------55
4.3.1正常大鼠坐骨神經切面之描述--------------------------------55
4.3.2矽膠管內白色再生管狀物切面之觀察-----------------------56
4.3.2.1無神經組織細胞----------------------------------------56
4.3.2.2髓鞘化軸突形成----------------------------------------57
4.3.2.3各組切片觀察-------------------------------------------57
4.4神經再生的成功率-------------------------------------------------------63
4.5組織學定量分析----------------------------------------------------------63
4.5.1再生神經之基本資料--------------------------------------------63
4.5.2再生神經切片之統計分析---------------------------------------64
4.5.2.1再生神經軸突數目--------------------------------------67
4.5.2.2再生神經軸突平均面積--------------------------------68
4.5.2.3再生神經內膜面積--------------------------------------69
4.5.2.4再生神經全部面積--------------------------------------70
4.5.2.5再生神經軸突密度--------------------------------------71
4.5.2.6再生神經軸突面積百分比-----------------------------72
4.5.2.7再生血管數目--------------------------------------------73
4.5.2.8再生血管總面積-----------------------------------------74
4.5.2.9再生血管面積百分比-----------------------------------75
5討論----------------------------------------------------------------------------------76
6結論----------------------------------------------------------------------------------80
參考文獻------------------------------------------------------------------------------81
英文摘要------------------------------------------------------------------------------92
謝辭------------------------------------------------------------------------------------94
圖目錄
圖2.1 神經元的類型與構造-------------------------------------------------------6
圖2.2 神經幹的結構---------------------------------------------------------------10
圖2.3 Genipin之分子結構式與其交聯方式---------------------------------39
圖3.1 神經斷端與矽膠管縫合的步驟------------------------------------------43
圖3.2 選取組織切片用的再生神經組織---------------------------------------44
圖4.1 大鼠足趾自殘缺損(G+G4)--------------------------------------------50
圖4.2 大鼠足趾缺損與足跟傷口(C3)---------------------------------------50
圖4.3 大鼠嚴重之足趾缺損與足跟傷口---------------------------------------51
圖4.4 神經管內形成再生神經組織(C4)------------------------------------52
圖4.5 神經管中明顯之再生神經組織(C7),兩側有增生之纖維
組織形成--------------------------------------------------------------------52
圖4.6 將神經管剝離後之再生神經組織(C7)------------------------------53
圖4.7 神經管內形成再生神經組織(G+G7)--------------------------------53
圖4.8 神經管中有再生神經組織形成(G+G6)-----------------------------54
圖4.9 神經管中沒有再生組織形成,僅充滿透明狀液體(G+G2)-----54
圖4.10於顯微鏡下觀察正常大鼠坐骨神經橫切面--------------------------55
圖4.11於顯微鏡下觀察正常大鼠坐骨神經橫切面--------------------------56
圖4.12無神經組織細胞的再生組織(G8)-----------------------------------57
圖4.13 C組再生神經組織(C7)-----------------------------------------------58
圖4.14 G9之再生神經組織,其形態較不成熟-------------------------------59
圖4.15 G3之再生神經組織,其軸突較密集----------------------------------59
圖4.16 G+G4之再生神經組織,其形態較不成熟---------------------------60
圖4.17 G+G5之再生神經組織,則頗為成熟---------------------------------61
圖4.18 N組再生神經組織(N7)-----------------------------------------------62
圖4.19 N組再生神經組織(N10)---------------------------------------------62
圖4.20各組神經再生成功率的比較--------------------------------------------63
圖4.21再生神經平均軸突數四組之比較--------------------------------------67
圖4.22再生神經軸突平均面積四組之比較-----------------------------------68
圖4.23再生神經內膜面積四組之比較-----------------------------------------69
圖4.24再生神經的全部面積四組之比較--------------------------------------70
圖4.25再生神經軸突密度四組之比較-----------------------------------------71
圖4.26再生神經軸突面積百分比四組之比較--------------------------------72
圖4.27再生血管數目四組之比較-----------------------------------------------73
圖4.28再生血管面積四組之比較-----------------------------------------------74
圖4.29再生血管面積百分比四組之比較--------------------------------------75
表目錄
表4.1 實驗動物矽膠管內有無再生神經組織、體重變化、足趾缺損
、足跟傷口的情形--------------------------------------------------------49
表4.2 各組再生神經組織之基本資料-----------------------------------------65
表4.3 再生神經之組織資料比較及檢定結果--------
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