本文關鍵詞：兔自體外周血間充質干細胞對腱—骨界面愈合影響的實驗研究　出處：《安徽醫(yī)科大學》2012年碩士論文　論文類型：學位論文

  更多相關文章： 外周血間充質干細胞 腱—骨界面愈合 前交叉韌帶 交叉韌帶重建

【摘要】：目的探討兔自體外周血間充質干細胞（MSCs）對前交叉韌帶（ACL）重建后腱-骨界面愈合的影響。 方法以粒細胞集落刺激因子作為外周血MSCs動員劑，體外分離、培養(yǎng)、擴增獲得外周血MSCs。48只4~6月齡新西蘭白兔按1-48編號，隨機均分為A、B、C三組，以后肢右膝關節(jié)作為研究對象，采用趾長屈肌腱作為ACL重建移植物。A組兔將外周血間充質干細胞-生物蛋白膠（FG-MSCs）混合注入兔右膝ACL重建模型股骨道頂端間隙內；B組兔僅注射生物蛋白膠；C組兔交叉韌帶重建后不做其他處理。分別在2周、4周、8周、12周各時間點每組各處死4只兔子進行解剖學觀察，取材制成石蠟切片行蘇木素-伊紅染色(Hematoxylin-Eosin Staining,HE)、甲苯胺藍染色(Toluidine Blue Staining,TB)、Masson三色染色（Masson trichrome staining）進行組織形態(tài)學觀察，并進行腱-骨界面各時間點成纖維細胞計數(shù)及組織形態(tài)學Buark評分，以了解腱-骨界面愈合情況。 結果（1）①A組：術后2周可見腱-骨界面間隙由疏松結締組織填充，可見少量成纖維細胞，移植物與骨道之間分離；術后4周可見大量疏松結締組織、成纖維細胞和少量類軟骨細胞及膠原纖維，新生骨形成，腱-骨間隙已不可見；術后8周在腱-骨界面可見大量類軟骨細胞，出現(xiàn)Sharpey纖維集結成束，膠原纖維合成增多,排列較規(guī)則；術后12周腱-骨間形成致密結締組織，可見大量排列有序的Sharpey纖維以及纖維軟骨、鈣化軟骨形成，未出現(xiàn)潮線。②B、C組：術后2周腱-骨界面僅見少量疏松結締組織，少量成纖維細胞增生,未見膠原纖維，同時腱-骨界面明顯分離；術后4周腱-骨間可見少量疏松結締組織及少量新生骨細胞長入，散在分布的類軟骨細胞和膠原纖維，且未見與骨道垂直連接；術后8周腱-骨界面為較多類軟骨細胞及膠原纖維，Sharpey纖維散在分布；術后12周有較多排列無序的類軟骨細胞出現(xiàn)，，膠原纖維密集排列不規(guī)則，Sharpey纖維排列紊亂。（2）術后各時間點三組腱-骨界面高倍鏡下（HE染色）成纖維細胞計數(shù)資料采用單向方差分析及SNK法檢驗：A組*與B組**和A組*與C組***在各時間點比較均有統(tǒng)計學意義（P＜0.05），而B組**與C組***在各時間點比較無統(tǒng)計學意義。（3）基于高倍鏡下腱-骨界面的纖維血管組織和Sharpey纖維評定的組織形態(tài)學Buark評分統(tǒng)計學資料采用Wilcoxon秩和檢驗：12周內A組與B組組織形態(tài)學Buark評分中位數(shù)間的差異有統(tǒng)計學意義（P＜0.01），即A組組織形態(tài)學Buark評分較B組高。 結論兔自體外周血MSCs通過頂端注射的方式能夠促進腱-骨界面的愈合。
[Abstract]:Objective to investigate the effect of autologous peripheral blood mesenchymal stem cells (MSCs) on the healing of tendon bone interface after the reconstruction of anterior cruciate ligament (ACL) in rabbits.
Methods with granulocyte colony-stimulating factor for peripheral blood MSCs mobilization agent, in vitro isolation, culture, amplification of peripheral blood MSCs.48 4~6 month old New Zealand white rabbits by 1-48 number, were randomly divided into A, B, C three groups, after limb knee joint as the research object, as ACL reconstruction graft in group.A the peripheral blood mesenchymal stem cells and fibrin glue with flexor digitorum longus tendon (FG-MSCs) mixed into ACL model of rabbit femur reconstruction of right knee top clearance; B group were only injected with fibrin glue; no other treatment in group C after anterior cruciate ligament reconstruction. In 2 weeks, 4 weeks, 8 week, 12 weeks each time point in each group were sacrificed 4 rabbits were observed in paraffin section, hematoxylin eosin staining (Hematoxylin-Eosin, Staining, HE), toluidine blue staining (Toluidine Blue Staining, TB), Masson trichrome staining (Masson trichrome staining) organization form In order to understand the healing of the tendon bone interface, the fibrous cell count and the histomorphology Buark score at the time point of the tendon bone interface were observed.
Results (1) A group: 2 weeks after surgery, the tendon bone interface by loose connective tissue filled the gap, a small amount of fibroblasts, separation between graft and bone; 4 weeks after operation showed a large amount of loose connective tissue, fibroblasts and a small amount of cartilage cells and collagen fibers, new bone formation that tendon bone gap is not visible; 8 weeks after operation in the tendon bone interface showed a large number of chondrocytes, Sharpey fiber bundles of collagen synthesis increased, a regular arrangement; 12 weeks after operation, the tendon bone formation of dense connective tissue, showing a large number of ordered Sharpey fiber and fiber cartilage. Calcified cartilage formation, without tidal line. The B, C group: after 2 weeks of tendon bone interface but only a small amount of loose connective tissue, a small amount of fibroblast proliferation, collagen fibers were at the same time, the tendon bone interface were separated; after 4 weeks of tendon bone can be seen between the small amount of loose connective tissue And a small amount of new bone cells, scattered cartilage cells and collagen fibers, and no bone and vertical connection; 8 weeks after operation, the tendon bone interface into chondrocytes and collagen fibers, Sharpey fibers were distributed; 12 weeks after surgery more cartilage cell disorder arrangement, dense collagen fibers arranged in irregular, disordered arrangement of Sharpey fiber. (2) at each time point after operation three groups of tendon bone interface at high magnification (HE staining) fibroblast count data using one-way analysis of variance and SNK test: A group and B group and A * * * * * * * in the group and C group at each time point were statistically significant (P < 0.05), B group and C group * * * * * in each time point is not significant. (3) fibrovascular tissue and Sharpey fibers evaluated at high magnification, the tendon bone interface morphology Buark Score statistical data using the Wilcoxon rank test based on: 12 weeks There was significant difference in the median of Buark score between group A and group B (P < 0.01), that is, the histomorphological Buark score of group A was higher than that of the B group.
Conclusion the autologous peripheral blood MSCs can promote the healing of the tendon bone interface by injection of the apical injection.

【學位授予單位】：安徽醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2012
【分類號】：R681;R-332

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