两种富血小板纤维蛋白的降解特性研究
发布时间：2018-08-25 09:16
【摘要】：目的:从抽取人血液制备两种富血小板纤维蛋白进行的体外降解实验和实验动物口内植入实验两个部分对比分析富血小板纤维蛋白和改良型富血小板纤维蛋白(Platelet-Rich Fibrin,PRF和Advanced Platelet-Rich Fibrin,A-PRF)的降解特性,评估其临床应用价值,为A-PRF膜和PRF膜的临床应用提供相关实验依据。方法:1.体外降解实验:10名志愿者各抽取10ml肘静脉血用以制取A-PRF和PRF膜,A-PRF以Ghanaati法制备(1500rpm,14min),PRF以Choukroun法制备(2700rpm,12min)。制备好的A-PRF、PRF膜浸泡在人工唾液内置于二氧化碳培养箱,每天记录膜的物理状态并测量膜标本的重量。2.兔口内植入实验:36只实验兔随机分为A-PRF组(A组)和PRF组(B组),每只兔抽取10ml血液用以制备A-PRF或PRF膜,A-PRF(2000rpm,24min),PRF(3250rpm,10min)。将制备好的膜修剪成6mm直径的圆形膜片。于每只兔硬腭处制备两处模型:在距上颌内侧门牙4.5mm的腭中线处用环切钻制备直径5mm的软组织缺损,剥离粘骨膜显露骨面,并向周围潜行分离1mm;在软组织缺损下方2mm约一个腭皱襞处用手术刀片做边长5mm的角形切口,翻全层粘骨膜瓣显露骨面。A组在软组织缺损区植入自体A-PRF圆形膜片,边缘缝合固定(A1组,即A-PRF膜暴露组);角形瓣内植入A-PRF圆形膜片并缝合(A2组,即A-PRF膜埋置组)。B组在两处植入自体PRF圆形膜片,软组织缺损区为B1组(即PRF膜暴露组),角形瓣区为B2组(即PRF膜埋置组)。四组每组18个实验样本。在术后3d、5d、7d、10d、14d、21d,A、B两组各随机处死3只动物,取出位于上颚部的膜标本,进行形态学观察并称重计算降解率,扫描电镜观察降解过程,HE染色,观察组织病理变化,进行炎性评分。结果:1.体外降解实验:(1)在人工唾液培养环境下,第14天所有PRF膜标本均完全降解,第17天所有A-PRF膜标本均完全降解。(2)A-PRF和PRF膜的降解速度有差异(P0.05)。2.兔口内植入实验:(1)形态学观察:术后第7d:A2和B2组创口基本愈合;术后第14d,A1和B1组创口基本愈合。术后第7d:B1组膜材料明显吸收较难完整取出;术后第10d:B1组无可见标本,A1组、B2组膜取出困难;术后第14d:A2组膜取出困难,A1组、B2组无可见标本;术后第21d:A2组无法找到植入标本。(2)降解情况:完全降解的时间A1组14天,A2组21天,B1组10天,B2组14天。组间对比:A1、A2两组膜标本的降解率在前5个时间节点有统计学差异(P0.05);B1、B2组的降解率除第三天无统计学差异(P=0.0660.05),其余时间点差异均有统计学意义(P0.05);A1、B1两组膜第5天、7天、10天差异有统计学意义(P0.05);A2、B2组在五个时间节点的降解率均有统计学差异(P0.05)。(3)扫描电镜结果:A-PRF和PRF膜均呈三维网状结构,由三叉结构连接。降解过程中纤维蛋白网状结构逐渐崩解,表现为纤维蛋白条索的断裂,网间孔隙增大,三维网状结构逐渐不规则成形,膜降解为碎片,最终观察不到明显的网状结构。(4)苏木精-伊红染色观察:四组镜下均表现为红染的纤维蛋白条索逐渐减少,可见炎性细胞的浸润,成纤维细胞和新生毛细血管数量逐渐增多。术后第3d、5d、7d,A1组炎性反应都较A2组重(P0.05),B1组炎性反应也较B2组重(P0.05);术后10d,A1与A2间及B1与B2组间炎性反应无统计学差异(P0.05)。结论:1.A-PRF与PRF膜均可在人工唾液中发生降解,PRF膜的完全降解时间为14天,A-PRF膜为17天,PRF膜在人工口腔环境中的降解速度比A-PRF膜快。2.在兔口腔内,暴露条件加快了膜的降解。3.在兔口腔埋置条件下,A-PRF膜的降解速度慢于PRF膜。4.A-PRF和PRF膜不可单独作为屏障膜使用。5.A-PRF和PRF膜的降解为纤维蛋白的降解和三维网状结构的崩塌。6.A-PRF和PRF膜在暴露条件下会加重炎症反应。
[Abstract]:OBJECTIVE: To compare and analyze the degradation characteristics of platelet-rich fibrin (PRF) and modified platelet-rich fibrin (A-PRF) in vitro and in vivo, and to evaluate their clinical significance. METHODS: 1. In vitro degradation experiment: 10 volunteers were used to prepare A-PRF and PRF membranes from elbow vein blood. A-PRF was prepared by Ghanaati method (1500rpm, 14min), PRF by Choukroun method (2700rpm, 12min). The prepared A-PRF and PRF membranes were immersed in artificial saliva. 36 rabbits were randomly divided into A-PRF group (group A) and PRF group (group B). Each rabbit took 10 ml of blood to prepare A-PRF or PRF membranes, A-PRF (2000 rpm, 24 min), PRF (3250 rpm, 10 min). The prepared membranes were trimmed into 6 mm diameters. Two models were made at each rabbit's hard palate: a 5 mm diameter soft tissue defect was made at the middle line of the palate 4.5 mm from the maxillary medial incisor, the mucoperiosteal was stripped to expose the bone surface, and the soft tissue defect was separated laterally for 1 mm; a 5 mm long corner incision was made at a palatal fold 2 mm below the soft tissue defect, and the whole layer was turned over. In group A, autologous A-PRF circular diaphragm was implanted in the soft tissue defect area, and the edge was sutured and fixed (group A1, exposed group of A-PRF membrane); in group A2, exposed group of A-PRF circular diaphragm was implanted and sutured (group A2, implanted group of A-PRF membrane). In group B, autologous PRF circular diaphragm was implanted in two places, and in group B1, exposed group of PRF membrane in the soft tissue defect area. Zone B2 (PRF membrane embedding group). Each of the four groups had 18 experimental samples. Three animals were randomly killed in three days, five days, seven days, ten days, fourteen days, twenty-one days, A and B after operation. The membrane specimens in the upper jaw were taken out for morphological observation and weighing to calculate the degradation rate. The degradation process was observed by scanning electron microscopy, HE staining, histopathological changes were observed and inflammatory scores were made. In vitro degradation test: (1) All PRF membrane specimens were completely degraded on the 14th day and all A-PRF membrane specimens were completely degraded on the 17th day in artificial saliva culture environment. (2) The degradation rate of A-PRF and PFF membrane was different (P 0.05). 2. In rabbit oral implantation test: (1) Morphological observation: wound healing was observed in groups A2 and B2 on the 7th day after operation; wounds in groups A1 and B1 on the 14th day after operation. On the 7th day after operation, the membrane material in group B1 was obviously absorbed and difficult to be completely removed; on the 10th day after operation, there was no visible specimen in group B1, and it was difficult to remove membrane in group A1 and B2; on the 14th day after operation, it was difficult to remove membrane in group A2, and there was no visible specimen in group A1 and B2; on the 21st day after operation, no implanted specimen could be found in group A2. (2) Degradation: The time of complete degradation was 14 days in group A1, 21 days in group A2 and B1. The degradation rates of membrane samples in A1 and A2 groups at the first five time points were statistically different (P 0.05); the degradation rates of B1 and B2 groups were not statistically different except the third day (P = 0.0660.05), the other time points were statistically significant (P 0.05); A1 and B1 groups on the fifth, seventh and tenth day (P 0.05); A2 and B2 groups on the fifth, seventh and seventh day (P 0.05); Scanning electron microscopy (SEM) results showed that both A-PRF and PRF membranes had three-dimensional network structure, which was connected by trigeminal structure. (4) Hematoxylin-eosin staining showed that the number of red-stained fibrin bands decreased gradually, inflammatory cells infiltrated, fibroblasts and new capillaries increased gradually. Inflammatory reactions in group A1 were heavier than those in group A2 (P 0.05) and group B1 (P 0.05) on the 3rd, 5th, 7th day after operation. There was no significant difference in inflammatory reaction between A1 and A2 and between B1 and B2 groups (P 0.05). Conclusion: 1. Both A-PRF and PRF membranes can be degraded in artificial saliva. The complete degradation time of PRF membranes is 14 days, A-PRF membranes are 17 days, and PRF membranes are degraded faster in artificial oral environment than A-PRF membranes. Dew accelerated the degradation of the membrane. 3. The degradation rate of A-PRF membrane was slower than that of PRF membrane. 4. A-PRF and PRF membrane could not be used as barrier membrane alone. 5. A-PRF and PRF membrane could degrade into fibrin degradation and collapse of three-dimensional network structure.
【学位授予单位】：西南医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R783.6
本文编号：2202430
[Abstract]:OBJECTIVE: To compare and analyze the degradation characteristics of platelet-rich fibrin (PRF) and modified platelet-rich fibrin (A-PRF) in vitro and in vivo, and to evaluate their clinical significance. METHODS: 1. In vitro degradation experiment: 10 volunteers were used to prepare A-PRF and PRF membranes from elbow vein blood. A-PRF was prepared by Ghanaati method (1500rpm, 14min), PRF by Choukroun method (2700rpm, 12min). The prepared A-PRF and PRF membranes were immersed in artificial saliva. 36 rabbits were randomly divided into A-PRF group (group A) and PRF group (group B). Each rabbit took 10 ml of blood to prepare A-PRF or PRF membranes, A-PRF (2000 rpm, 24 min), PRF (3250 rpm, 10 min). The prepared membranes were trimmed into 6 mm diameters. Two models were made at each rabbit's hard palate: a 5 mm diameter soft tissue defect was made at the middle line of the palate 4.5 mm from the maxillary medial incisor, the mucoperiosteal was stripped to expose the bone surface, and the soft tissue defect was separated laterally for 1 mm; a 5 mm long corner incision was made at a palatal fold 2 mm below the soft tissue defect, and the whole layer was turned over. In group A, autologous A-PRF circular diaphragm was implanted in the soft tissue defect area, and the edge was sutured and fixed (group A1, exposed group of A-PRF membrane); in group A2, exposed group of A-PRF circular diaphragm was implanted and sutured (group A2, implanted group of A-PRF membrane). In group B, autologous PRF circular diaphragm was implanted in two places, and in group B1, exposed group of PRF membrane in the soft tissue defect area. Zone B2 (PRF membrane embedding group). Each of the four groups had 18 experimental samples. Three animals were randomly killed in three days, five days, seven days, ten days, fourteen days, twenty-one days, A and B after operation. The membrane specimens in the upper jaw were taken out for morphological observation and weighing to calculate the degradation rate. The degradation process was observed by scanning electron microscopy, HE staining, histopathological changes were observed and inflammatory scores were made. In vitro degradation test: (1) All PRF membrane specimens were completely degraded on the 14th day and all A-PRF membrane specimens were completely degraded on the 17th day in artificial saliva culture environment. (2) The degradation rate of A-PRF and PFF membrane was different (P 0.05). 2. In rabbit oral implantation test: (1) Morphological observation: wound healing was observed in groups A2 and B2 on the 7th day after operation; wounds in groups A1 and B1 on the 14th day after operation. On the 7th day after operation, the membrane material in group B1 was obviously absorbed and difficult to be completely removed; on the 10th day after operation, there was no visible specimen in group B1, and it was difficult to remove membrane in group A1 and B2; on the 14th day after operation, it was difficult to remove membrane in group A2, and there was no visible specimen in group A1 and B2; on the 21st day after operation, no implanted specimen could be found in group A2. (2) Degradation: The time of complete degradation was 14 days in group A1, 21 days in group A2 and B1. The degradation rates of membrane samples in A1 and A2 groups at the first five time points were statistically different (P 0.05); the degradation rates of B1 and B2 groups were not statistically different except the third day (P = 0.0660.05), the other time points were statistically significant (P 0.05); A1 and B1 groups on the fifth, seventh and tenth day (P 0.05); A2 and B2 groups on the fifth, seventh and seventh day (P 0.05); Scanning electron microscopy (SEM) results showed that both A-PRF and PRF membranes had three-dimensional network structure, which was connected by trigeminal structure. (4) Hematoxylin-eosin staining showed that the number of red-stained fibrin bands decreased gradually, inflammatory cells infiltrated, fibroblasts and new capillaries increased gradually. Inflammatory reactions in group A1 were heavier than those in group A2 (P 0.05) and group B1 (P 0.05) on the 3rd, 5th, 7th day after operation. There was no significant difference in inflammatory reaction between A1 and A2 and between B1 and B2 groups (P 0.05). Conclusion: 1. Both A-PRF and PRF membranes can be degraded in artificial saliva. The complete degradation time of PRF membranes is 14 days, A-PRF membranes are 17 days, and PRF membranes are degraded faster in artificial oral environment than A-PRF membranes. Dew accelerated the degradation of the membrane. 3. The degradation rate of A-PRF membrane was slower than that of PRF membrane. 4. A-PRF and PRF membrane could not be used as barrier membrane alone. 5. A-PRF and PRF membrane could degrade into fibrin degradation and collapse of three-dimensional network structure.
【学位授予单位】：西南医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R783.6
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