本文選題：前交叉韌帶 + 腱骨愈合��； 參考：《安徽醫(yī)科大學(xué)》2015年碩士論文

【摘要】：目的探討在兔自體肌腱移植物重建前交叉韌帶(anterior cruciate ligament,ACL)模型股骨道內(nèi)注入自體外周血間充質(zhì)干細胞(peripheral blood mesenchymal stem cells,PBMSCs)凝膠對早期腱骨愈合的影響。方法健康3至4月齡新西蘭大白兔32只,隨機分為實驗組與對照組,每組16只,實驗組予以皮下注射粒細胞集落刺激因子(G-CSF)動員6天后于耳緣靜脈采集外周血,密度梯度離心法分離純化PBMSCs,細胞培養(yǎng)箱內(nèi)采用貼壁培養(yǎng)法培養(yǎng)并傳代至第三代,光鏡下觀察細胞形態(tài)后行流式細胞檢測鑒定所培養(yǎng)細胞表面特定抗原(CD11b、CD29、CD34、CD90)的表達。對全部32只新西蘭大白兔采用拋硬幣法隨機選取一側(cè)膝關(guān)節(jié),建立兔自體肌腱移植物重建前交叉韌帶模型,實驗組于動物模型股骨道內(nèi)注入PBMSCs凝膠,對照組僅注入凝膠。分別于術(shù)后第2、4、8、12周每組隨機取4只實驗動物處死取出膝關(guān)節(jié),其中1只做Masson三色染色觀察移植物在股骨道中組織愈合的病理表現(xiàn),3只做股骨道移植物抗牽拉試驗觀察其愈合強度,記錄移植物從骨道內(nèi)脫出時的拉力值,實驗組與對照組所得數(shù)據(jù)采用兩樣本t檢驗,比較兩組數(shù)值有無顯著性差異(選取a=0.01為檢驗水準)。結(jié)果光鏡下觀察待測細胞以集落方式貼壁生長,呈長梭型或多角形。流式細胞學(xué)檢測得出被檢測細胞表面CD29、CD90、CD11b、CD34陽性表達率分別為97.3%、98.4%、1.3%、1.3%,提示被檢測細胞表面表達CD29、CD90,不表達CD11b、CD34。Masson三色染色結(jié)果提示:術(shù)后第2周:兩組在腱骨交界處均有大量壞死組織形成,纖維排列紊亂,成纖維細胞與膠原纖維均無明顯增生;術(shù)后第4周:對照組仍有大量壞死組織,纖維排列不齊,有少量成纖維細胞增生,但未見明顯膠原纖維。PBMSCs組壞死組織較空白組減少,組織排列欠規(guī)則,有少量成纖維細胞及膠原纖維長入;術(shù)后第8周:對照組有大量成纖維細胞長入,纖維排列欠規(guī)則,腱骨愈合界面有少量膠原纖維長入。PBMSCs組有大量成纖維細胞長入,部分纖維排列整齊,部分欠規(guī)則,腱骨愈合界面有大量膠原纖維長入;術(shù)后第12周:對照組成纖維細胞數(shù)量減少,膠原纖維生成增多,腱骨愈合交界處成纖維細胞與膠原纖維數(shù)量比約為1:1。PBMSCs組僅殘余少量成纖維細胞,大量膠原纖維規(guī)則排列在腱骨愈合交界處。在實驗組與對照組中,股骨道移植物抗牽拉力強度均隨時間延長呈上升趨勢,在術(shù)后第2周、第4周,對兩組的抗牽拉強度進行比較,無明顯統(tǒng)計學(xué)意義(P0.01)。在術(shù)后第8周及第12周,對兩組的抗牽拉強度進行比較,PBMSCs組抗牽拉力強度大于對照組,差異有統(tǒng)計學(xué)意義(P0.01)結(jié)論PBMSCs凝膠對兔自體肌腱移植物重建ACL的早期腱骨愈合有促進作用。
[Abstract]:Objective to investigate the effect of autologous peripheral blood mesenchymal stem cells (peripheral blood mesenchymal stem cells, PBMSCs) gel on early tendon bone healing in the femoral canal of anterior cruciate ligament (ACL) model of autogenous tendon graft in rabbits. Methods 32 rabbits were randomly divided into 3 to 4 month old New Zealand white rabbits. Group and control group, with 16 rats in each group, the experimental group was subcutaneously injected with granulocyte colony stimulating factor (G-CSF) to collect peripheral blood in the ear vein for 6 days, and the density gradient centrifugation was used to separate and purify PBMSCs. The cell culture box was cultured and passed to the third generation in the cell culture box, and the cell morphology was observed by the flow cytometry. The expression of the specific antigen (CD11b, CD29, CD34, CD90) on the surface of the cultured cells. All 32 New Zealand white rabbits were randomly selected to select one side of the knee joint with a coin toss method. The rabbit autologous tendon graft was established to reconstruct the anterior cruciate ligament. In the experimental group, the PBMSCs gel was injected into the femoral canal of the animal model and the control group was injected only with the gel. The knee joints were taken out of 4 experimental animals in each group at 2,4,8,12 weeks. 1 of them were stained with Masson Tri Color staining to observe the pathological manifestation of tissue healing in the femoral canal. 3 of the femur graft resistance test was performed to observe the healing strength of the graft, and the tension values of the graft from the bone canal were recorded. The data obtained from the experimental group and the control group were recorded. Two samples t test was used to compare the significant differences between the two groups (select a=0.01 as the test level). Results the cells under the microscope were observed under the light microscope, and the cell surface was adhered to the wall. The flow cytometry showed that the positive expression rates of CD29, CD90, CD11b and CD34 were 97.3%, 98.4%, 1.3%, 1.3%, respectively. The expression of CD29, CD90, and non expression of CD11b on the surface of the cells showed that second weeks after the operation: second weeks after the operation, there were a large number of necrotic tissue in the two groups at the junction of the tendon and bone, the fibrous arrangement was disorderly, and the fibroblasts and the collagen fibers had no obvious hyperplasia; fourth weeks after the operation, there were still a large number of necrotic tissue in the control group. There was no obvious collagen fibrous fibroblast proliferation, but no obvious collagen fibrous.PBMSCs group was less than the blank group, the tissue arrangement was less regular, and a small amount of fibroblasts and collagen fibers grew. Eighth weeks after the operation, there were a large number of fibroblasts in the control group, the fiber arrangement was not regular, and the tendon bone healing interface with a small amount of collagen fibers grew into the.PBMSCs group. For Twelfth weeks after the operation, the number of fibrous cells in the control group decreased, the collagen fibers increased, and the number of fibroblasts and collagen fibers at the junction of tendon bone healing was only a small amount of fibroblasts in group 1:1.PBMSCs, and the number of fibroblasts and collagen fibers at the juncture of tendon bone healing were only a small amount of fibroblasts. A large number of collagen fibers were arranged at the juncture of tendon bone healing. In the experimental group and the control group, the tensile strength of the femur transplants increased with time. In the second week and fourth weeks after the operation, the tensile strength of the two groups was compared. There was no significant statistical significance (P0.01). The anti traction of the two groups at eighth and 12 weeks after the operation. The strength of the PBMSCs group was higher than that of the control group, and the difference was statistically significant (P0.01). Conclusion PBMSCs gel could promote the early tendon bone healing of the rabbit autologous tendon graft for the reconstruction of ACL.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：R687

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