自體骨髓提取物Buffy Coat與透明質(zhì)酸復合物對軟骨再生的實驗研究
發(fā)布時間：2019-04-02 03:16
【摘要】：目的:探討自體骨髓提取物Buffy Coat和透明質(zhì)酸(Hyaluronic Acid,HA)復合物對兔膝關(guān)節(jié)軟骨的再生的影響,為Buffy Coat修復軟骨的臨床應(yīng)用提供實驗依據(jù)。方法:將成年新西蘭大白兔36只(16周齡,體重3.0kg-3.5kg)隨機分為三組(A組、B組和C組),每組12只大白兔,A組軟骨缺損處只注射透明質(zhì)酸;B組只注射Buffy Coat;C組注射Buffy Coat與透明質(zhì)酸復合物。Buffy Coat提取在大白兔髂前上棘處提取骨髓進行分離獲得Buffy Coat,制備軟骨缺損模型選擇大白兔右膝關(guān)節(jié)的股骨髁滑車處造軟骨缺損直徑約5mm,深于2mm。術(shù)后第4周和第8周進行取標本,肉眼觀察軟骨缺損區(qū)外觀并拍照保存資料。病理學觀察進行常規(guī)脫鈣、脫水、蠟塊包埋、病理切片、潘紅O染色、封片,在顯微鏡下觀察觀察再生軟骨細胞形態(tài)學、染色現(xiàn)象、結(jié)構(gòu)完整性、表面整齊性、軟骨厚度、再生的軟骨下骨、再生軟骨和周圍正常組織整合情況并拍照保存資料以及請三名相關(guān)實驗人員進行ICRS評分并對三組標本評分結(jié)果統(tǒng)計學比較分析。結(jié)果:1.肉眼觀察:術(shù)后第4周見A組缺損區(qū)保持缺陷,B組中的缺損區(qū)可見修復組織部分填充缺陷區(qū),C組中的缺陷的填充程度比B組較寬,仍未填充滿。術(shù)后第8周三組的缺損區(qū)已被填充滿,A組的修復組織明顯發(fā)白與周圍正常軟骨明顯分開,而B組和C組表面光滑接近于周圍正常軟骨,但有少量發(fā)白。2.鏡下觀察:術(shù)后第4周C組潘紅O染色觀察染色現(xiàn)象輕微減少、表面中等與B組和A組比較好,A組大部分無修復軟骨、軟骨下骨未重建,B組和C組大部分纖維軟骨修復,軟骨下骨部分重建。術(shù)后第8周A組纖維組織修復,表面層較粗糙,軟骨下骨部分重建。B組中的修復組織表面較整齊但仍不規(guī)則,并發(fā)現(xiàn)大量的膠原纖維,細胞分布由具有透明性質(zhì)的柱狀和簇狀細胞組成,但細胞的排列較亂,重建的軟骨下骨大部分重建而不規(guī)則的,也未觀察到軟骨組織和周圍軟骨之間的整合。C組中的缺陷區(qū)比B組整齊接近周圍正常軟骨表面,并發(fā)現(xiàn)大量的膠原纖維與正常軟骨類似的膠原纖維的良好定向排列,細胞的排列大部分類似正常軟骨細胞排列,觀察到有兩邊的修復組織與周圍組織也未能融合,重建軟骨下骨大部分重建。ICRS評分及統(tǒng)計學分析..第4周三組比較明顯有差異(P0.05),組間比較C組軟骨修復效果優(yōu)于B組和A組,顯著統(tǒng)計學意義,有明顯差異(P0.05),第8周ICRS評分三組比較明顯有差異(P0.05),組間比較B組和C組無明顯差異(P0.05),通過第4周和第8周ICRS評分可以看到每組的評分隨時間的推移評分越高。結(jié)論:Buffy Coat與透明質(zhì)酸復合物對兔關(guān)節(jié)軟骨缺損再生有明顯的效果,透明質(zhì)酸凝膠狀物可以與Buffy Coat混合用于解決Buffy Coat液體狀難移植和固定到軟骨損傷部位的難題,為Buffy Coat在臨床應(yīng)用提供了實驗依據(jù)。
[Abstract]:Aim: to investigate the effect of autogenous bone marrow extract (Buffy Coat) and hyaluronic acid (Hyaluronic Acid,HA) complex on cartilage regeneration of rabbit knee joint in order to provide experimental basis for the clinical application of Buffy Coat in repairing cartilage. Methods: thirty-six adult New Zealand white rabbits (16 weeks old, weight 3.0kg-3.5kg) were randomly divided into three groups (group A, group B and group C) with 12 rabbits in each group. Hyaluronic acid was injected into cartilage defect of group A and Buffy Coat; was injected into group B only. Group C was injected with Buffy Coat and hyaluronic acid complex. Buffy Coat to extract bone marrow from anterior superior iliac spine of rabbits. Buffy Coat, was used to prepare cartilage defect model. The diameter of cartilage defect at trochanter of femoral condyle of rabbits was about 5 mm. Deeper than 2 mm. The specimens were taken from the 4th and 8th week after operation. The appearance of cartilage defect area was observed by naked eye and the data were photographed. Pathological observation was carried out by routine decalcification, dehydration, wax embedding, pathological section, dipyridol O staining and sealing. The morphology, staining phenomena, structural integrity, surface uniformity and cartilage thickness of regenerated chondrocytes were observed under microscope. The integration of regenerated subchondral bone, regenerated cartilage and surrounding normal tissue was recorded and the data were taken and kept. Three relevant researchers were asked to evaluate the ICRS score and compare the results of the three groups of specimens statistically. Results: 1. At the 4th week after operation, the defect area in group A remained defect, the defect area in group B was partly filled with defect area, and the filling degree of defect in group C was wider than that in group B, and the defect in group C was still not filled. At the 8th week after operation, the defect area of the three groups had been filled, and the repair tissue of group A was obviously white from the surrounding normal cartilage, while the surface of groups B and C was smooth close to that of the surrounding normal cartilage, but a small amount of white was observed. Microscopic observation: at the 4th week after operation, the staining phenomenon of dipyridamole O in group C decreased slightly, the surface of group C was better than that of group B and group A, most of the cartilage in group A had no repair, the subchondral bone had not been reconstructed, and most of fibrocartilage in group B and C had been repaired. The subchondral bone was partially reconstructed. At the 8th week after operation, the fibrous tissue of group A was repaired with rough surface layer and partial reconstruction of subchondral bone. In group B, the surface of the repaired tissue was neat but still irregular, and a large number of collagen fibers were found. The distribution of cells is composed of transparent columnar and clustered cells, but the arrangement of cells is chaotic, and the reconstructed subchondral bone is mostly reconstructed and irregular. No integration between cartilage tissue and surrounding cartilage was observed. The defect areas in group C were nearer to the surface of surrounding normal cartilage than in group B, and a large number of collagen fibers were well aligned with those similar to normal cartilage. Most of the cells were similar to those of normal chondrocytes, and no fusion was observed between the repair tissue and the surrounding tissue. ICRS score and statistical analysis were used to reconstruct the subchondral bone. At the fourth week, there was significant difference among the three groups (P0.05), and the cartilage repair effect of group C was better than that of group B and A (P0.05), and there was significant difference between group C and group A (P0.05). There was significant difference in ICRS score among the three groups at the 8th week (P0.05), but there was no significant difference between the two groups (P0.05). The ICRS scores in the 4th week and 8th week showed that the score of each group was higher with the passage of time. Conclusion the complex of: Buffy Coat and hyaluronic acid has obvious effect on the regeneration of articular cartilage defect in rabbits. Hyaluronic acid gel can be mixed with Buffy Coat to solve the difficult problem of Buffy Coat being difficult to transplant and immobilize to the injury site of cartilage. It provides experimental basis for the clinical application of Buffy Coat.
【學位授予單位】：延邊大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R687
本文編號：2452182
[Abstract]:Aim: to investigate the effect of autogenous bone marrow extract (Buffy Coat) and hyaluronic acid (Hyaluronic Acid,HA) complex on cartilage regeneration of rabbit knee joint in order to provide experimental basis for the clinical application of Buffy Coat in repairing cartilage. Methods: thirty-six adult New Zealand white rabbits (16 weeks old, weight 3.0kg-3.5kg) were randomly divided into three groups (group A, group B and group C) with 12 rabbits in each group. Hyaluronic acid was injected into cartilage defect of group A and Buffy Coat; was injected into group B only. Group C was injected with Buffy Coat and hyaluronic acid complex. Buffy Coat to extract bone marrow from anterior superior iliac spine of rabbits. Buffy Coat, was used to prepare cartilage defect model. The diameter of cartilage defect at trochanter of femoral condyle of rabbits was about 5 mm. Deeper than 2 mm. The specimens were taken from the 4th and 8th week after operation. The appearance of cartilage defect area was observed by naked eye and the data were photographed. Pathological observation was carried out by routine decalcification, dehydration, wax embedding, pathological section, dipyridol O staining and sealing. The morphology, staining phenomena, structural integrity, surface uniformity and cartilage thickness of regenerated chondrocytes were observed under microscope. The integration of regenerated subchondral bone, regenerated cartilage and surrounding normal tissue was recorded and the data were taken and kept. Three relevant researchers were asked to evaluate the ICRS score and compare the results of the three groups of specimens statistically. Results: 1. At the 4th week after operation, the defect area in group A remained defect, the defect area in group B was partly filled with defect area, and the filling degree of defect in group C was wider than that in group B, and the defect in group C was still not filled. At the 8th week after operation, the defect area of the three groups had been filled, and the repair tissue of group A was obviously white from the surrounding normal cartilage, while the surface of groups B and C was smooth close to that of the surrounding normal cartilage, but a small amount of white was observed. Microscopic observation: at the 4th week after operation, the staining phenomenon of dipyridamole O in group C decreased slightly, the surface of group C was better than that of group B and group A, most of the cartilage in group A had no repair, the subchondral bone had not been reconstructed, and most of fibrocartilage in group B and C had been repaired. The subchondral bone was partially reconstructed. At the 8th week after operation, the fibrous tissue of group A was repaired with rough surface layer and partial reconstruction of subchondral bone. In group B, the surface of the repaired tissue was neat but still irregular, and a large number of collagen fibers were found. The distribution of cells is composed of transparent columnar and clustered cells, but the arrangement of cells is chaotic, and the reconstructed subchondral bone is mostly reconstructed and irregular. No integration between cartilage tissue and surrounding cartilage was observed. The defect areas in group C were nearer to the surface of surrounding normal cartilage than in group B, and a large number of collagen fibers were well aligned with those similar to normal cartilage. Most of the cells were similar to those of normal chondrocytes, and no fusion was observed between the repair tissue and the surrounding tissue. ICRS score and statistical analysis were used to reconstruct the subchondral bone. At the fourth week, there was significant difference among the three groups (P0.05), and the cartilage repair effect of group C was better than that of group B and A (P0.05), and there was significant difference between group C and group A (P0.05). There was significant difference in ICRS score among the three groups at the 8th week (P0.05), but there was no significant difference between the two groups (P0.05). The ICRS scores in the 4th week and 8th week showed that the score of each group was higher with the passage of time. Conclusion the complex of: Buffy Coat and hyaluronic acid has obvious effect on the regeneration of articular cartilage defect in rabbits. Hyaluronic acid gel can be mixed with Buffy Coat to solve the difficult problem of Buffy Coat being difficult to transplant and immobilize to the injury site of cartilage. It provides experimental basis for the clinical application of Buffy Coat.
【學位授予單位】：延邊大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R687
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