rhbFGF、rhIGF-I与人工骨复合促进犬牙周组织再生的实验研究
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摘要
目的：本实验通过人工构建5只杂种犬慢性牙周病模型,继而将重组人碱性成纤维细胞生长因子(Recombinant human basic fibroblast growth factor,rhbFGF)和重组人胰岛素样生长因子(Recombinant human insulin-like growth factor-I,rhIGF-I)分别及共同与人工骨(Bio-oss)复合,植入犬Ⅱ度根分叉牙周病模型,并配合引导性组织再生术(Guided issue regeneration,GTR)的治疗。观察分析动物模型的牙周组织修复和再生情况,并探讨两种生长因子对促进牙周组织再生的作用及效果,为其在牙周病临床治疗中的应用提供理论依据。
方法：
1.动物模型的建立及分组：选用健康、成年杂种犬5只,将形成的慢性Ⅱ度根分叉病变的上下颌第2、3、4前磨牙,共计48颗牙,随机分为5组,包括3个实验组,1个对照组和1个空白对照组,实验组A：rhbFGF/rhIGF-I/Bio-oss/GTR;实验组B：rhbFGF/Bio-oss/GTR;实验组C:rhIGF-I/Bio-oss/GTR;对照组D：单纯GTR手术；空白对照组E：翻瓣术。
2.分别在rhbFGF和／或rhIGF-I复合人工骨的植入术前及术后12周记录颊侧根分叉部位的牙周探诊深度(periodontal depth,PD)和附着水平(attachment level,AL);术后2、4、12周拍摄X光片,观察牙槽骨新生情况。
3.术后12周处死实验犬,制取实验区组织块(含牙、牙槽骨、牙周膜)进行切片,HE、Masson染色和组织学测量,观察牙周组织(牙槽骨、牙骨质和牙周膜)的新生情况。
结果：
1.临床检查：术后12周犬模型病变区的PD和AL均有不同程度改善,依次为A、B/C、D和E组；
2.X线检查：实验组A、B、C牙槽骨几乎充满前磨牙的根分叉,并可见较清晰的骨硬板；对照组D新生骨量较实验组少,而多于对照组E；对照组E由少量稀疏骨小梁充填在缺损底部,骨密度较低,骨硬板模糊；
3.组织学观察：
3.1 HE染色可见,实验组A、B、C缺损区可见新生牙槽骨充填根分叉区,其成骨密度较高,钙化良好；新生的牙周纤维细小稠密、富含血管,其方向与根面垂直或接近垂直；新生牙骨质覆盖根分又大部分区域；实验组均未见骨粘连和根吸收等不良愈合。对照组D有部分新生的牙槽骨,呈层状较成熟,牙骨质覆盖根面并向冠方延伸；在牙周间隙内有纤维穿行。对照组E仅有个别缺损区出现少量的新生牙槽骨,且其上方有较多的上皮细胞和非骨性结缔组织覆盖。
3.2 Masson染色可见,实验组A、B、C呈现明显的红染区,外周有少量蓝色着染区；对照组DMasson染色为红-蓝相间,骨小梁中心为红色区域：空白对照组E呈现明显的蓝色,周围有少量的红染区。
4.组织学测量：实验组新生牙槽骨高度、新生牙骨质高度及新生牙周组织的指标均高于对照组及空白对照组(P<0.05),各实验组相比较,A组的各指标虽在数值上较B、C组略高,但统计学分析,三组均无显著性差异(P>0.05)。
结论：
1.通过人工制备犬根分叉部位的骨缺损并辅以稳定的局部刺激因素的方法,可以制备出可靠的、近似于临床的犬牙周病Ⅱ度根分叉病变动物模型,以用于各种牙周组织再生治疗的动物实验研究。
2.通过对动物模型的临床检查、组织学观察、组织学测量,结果证明rhbFGF和/或rhIGF-I与Bio-oss复合材料具有良好的骨引导性和骨诱导性,用于犬Ⅱ度根分叉缺损的修复可明显促进牙周组织再生(牙槽骨牙骨质的再生及牙周新附着的形成)。
3.本实验的结果证明应用rhbFGF和/或rhIGF-I与人工骨复合材料对犬牙周病Ⅱ度根分叉病变模型的牙周组织再生优越于单纯的引导性牙周组织再生术。
4.本实验结果分析表明单独应用rhbFGF、rhIGF-I和此两种生子因子联合应用对牙周组织再生的质量上无显著性差异。
Objective:In this experiment study, the artificial construct five dogs with chronic periodontal disease model, and then the recombinant human basic fibroblast growth factor (recombinant human basic fibroblast growth factor, rhbFGF) and recombinant human insulin-like growth factor (recombinant human insulin-like growth factor-I, rhIGF-I), either separately or simultaneously with the artificial bone (Bio-oss) composite, implanted into dogs degreeⅡfurcation periodontal disease model, and with guided tissue regeneration (Guided issue regeneration, GTR) treatment, observation of animal models of periodontal tissue repair and regeneration, and to explore two kinds of growth factors to promote periodontal tissue regeneration and the effect of clinical treatment of periodontal disease in itsapplication to provide a theoretical basis.
Methods:
1. The formation of five dogs with chronicⅡfurcation of the mandibular first premolar 2,3,4, for a total of 48 teeth were randomly divided into 5 groups, including the three experimental groups, a control group and a blank control group, experimental group A:rhbFGF/rhIGF-I/ Bio-oss/GTR; experimental group B:rhbFGF/Bio-oss/GTR; experimental group C:rhIGF-I/Bio-oss/GTR; the control group D:GTR surgery alone; the blank control group E:flap operation.
2. Before implant rhbFGF and/or rhIGF-I composite bone graft and after 12 weeks,record buccal side of the bifurcation site of periodontal probing depth (periodontal depth,PD)and attachment level (attachment level,AL);after 2,4,12 weeks taken surgery X-rays to observe the new situation of alveolar bone.
3. Dogs were sacrificed 12 weeks after surgeried, slice the preparation experimental tissue zone (including dental, alveolar bone, periodontal ligament), HE, Masson staining and histological measurements, observe periodontal tissue (alveolar bone, teeth bone and periodontal membrane) of the new situation.
Results:
1. The clinical examination:after 12 weeks the PD and AL of dogs's model were improved to varying degrees, followed by A, B/C, D and E group;
2. X-ray examination:the experimental group A, B, C alveolar bone almost are filling the molar furcation, and the bone can be seen more clearly hard board; control group D were higher than the experimental group, less new bone, but more than the control group E; control group E are filling with a few sparse trabecular bone in the bottom of the low bone mineral density, bone hard plate fuzzy;
3. histological observation:HE staining, the experimental group A, B, C shows that new bone defect filling furcation, which become higher bone mineral density, calcium good; new born periodontal fiber small dense, rich in blood vessels, the direction perpendicular or nearly perpendicular to the root surface; new cementum covering most areas of the root bifurcation; the experimental group showed no bone adhesion and root absorption of malunion. Control group D, some new alveolar bone was layered over mature cementum covering the root surface to the coronal extension; fibers within the periodontal gap crossed. The control group E appears only a small number of individual defect new bone, and it has more epithelial cells and non-bone connective tissue covering. Masson staining in the experimental group A, B, C showed obvious red stained area with peripheral small amount of blue stained area; control group D Masson staining red-blue, red area of trabecular bone center; blank control group E showed obvious blue, surrounded by a small amount of red staining area;
4. tissue measurements can be seen in all experimental groups a high degree of new bone, new cementum height and new indicators of periodontal tissue were higher than that and the control group (P<0.05), compared with the experimental group, A group of the indicator, although numerically higher than B, C group was slightly higher, but the statistical analysis, three groups were not significantly different (P>0.05).
Conclusion:
1. Through artificial canine furcation bone defect site and supported by a stable method of local incentives that can be prepared reliable and similar to clinical periodontal disease in dogs II furcation Animal models, to be used for a variety of periodontal tissue regeneration therapy in animal study.
2. By clinical examination of animal models, histological observation, histological measurements, the results show rhbFGF and/or rhIGF-I and Bio-oss composite material has good bone and bone induction guide for dog II degree of root bifurcation defects can significantly promote alveolar bone, cementum and periodontal regeneration and new attachment formation.
3. The experimental results proved to use rhbFGF and/or rhIGF-I and bone composite to treatment canine periodontal diseaseⅡfurcation periodontal regeneration model is superior to simple guiding periodontal tissue regeneration.
4. The experimental results show that alone rhbFGF, rhIGF-I and these two factors combination on the periodontal tissue regeneration (regeneration of alveolar bone, cementum regeneration, periodontal membrane formation) the quality was no significant difference.
方法：
1.动物模型的建立及分组：选用健康、成年杂种犬5只,将形成的慢性Ⅱ度根分叉病变的上下颌第2、3、4前磨牙,共计48颗牙,随机分为5组,包括3个实验组,1个对照组和1个空白对照组,实验组A：rhbFGF/rhIGF-I/Bio-oss/GTR;实验组B：rhbFGF/Bio-oss/GTR;实验组C:rhIGF-I/Bio-oss/GTR;对照组D：单纯GTR手术；空白对照组E：翻瓣术。
2.分别在rhbFGF和／或rhIGF-I复合人工骨的植入术前及术后12周记录颊侧根分叉部位的牙周探诊深度(periodontal depth,PD)和附着水平(attachment level,AL);术后2、4、12周拍摄X光片,观察牙槽骨新生情况。
3.术后12周处死实验犬,制取实验区组织块(含牙、牙槽骨、牙周膜)进行切片,HE、Masson染色和组织学测量,观察牙周组织(牙槽骨、牙骨质和牙周膜)的新生情况。
结果：
1.临床检查：术后12周犬模型病变区的PD和AL均有不同程度改善,依次为A、B/C、D和E组；
2.X线检查：实验组A、B、C牙槽骨几乎充满前磨牙的根分叉,并可见较清晰的骨硬板；对照组D新生骨量较实验组少,而多于对照组E；对照组E由少量稀疏骨小梁充填在缺损底部,骨密度较低,骨硬板模糊；
3.组织学观察：
3.1 HE染色可见,实验组A、B、C缺损区可见新生牙槽骨充填根分叉区,其成骨密度较高,钙化良好；新生的牙周纤维细小稠密、富含血管,其方向与根面垂直或接近垂直；新生牙骨质覆盖根分又大部分区域；实验组均未见骨粘连和根吸收等不良愈合。对照组D有部分新生的牙槽骨,呈层状较成熟,牙骨质覆盖根面并向冠方延伸；在牙周间隙内有纤维穿行。对照组E仅有个别缺损区出现少量的新生牙槽骨,且其上方有较多的上皮细胞和非骨性结缔组织覆盖。
3.2 Masson染色可见,实验组A、B、C呈现明显的红染区,外周有少量蓝色着染区；对照组DMasson染色为红-蓝相间,骨小梁中心为红色区域：空白对照组E呈现明显的蓝色,周围有少量的红染区。
4.组织学测量：实验组新生牙槽骨高度、新生牙骨质高度及新生牙周组织的指标均高于对照组及空白对照组(P<0.05),各实验组相比较,A组的各指标虽在数值上较B、C组略高,但统计学分析,三组均无显著性差异(P>0.05)。
结论：
1.通过人工制备犬根分叉部位的骨缺损并辅以稳定的局部刺激因素的方法,可以制备出可靠的、近似于临床的犬牙周病Ⅱ度根分叉病变动物模型,以用于各种牙周组织再生治疗的动物实验研究。
2.通过对动物模型的临床检查、组织学观察、组织学测量,结果证明rhbFGF和/或rhIGF-I与Bio-oss复合材料具有良好的骨引导性和骨诱导性,用于犬Ⅱ度根分叉缺损的修复可明显促进牙周组织再生(牙槽骨牙骨质的再生及牙周新附着的形成)。
3.本实验的结果证明应用rhbFGF和/或rhIGF-I与人工骨复合材料对犬牙周病Ⅱ度根分叉病变模型的牙周组织再生优越于单纯的引导性牙周组织再生术。
4.本实验结果分析表明单独应用rhbFGF、rhIGF-I和此两种生子因子联合应用对牙周组织再生的质量上无显著性差异。
Objective:In this experiment study, the artificial construct five dogs with chronic periodontal disease model, and then the recombinant human basic fibroblast growth factor (recombinant human basic fibroblast growth factor, rhbFGF) and recombinant human insulin-like growth factor (recombinant human insulin-like growth factor-I, rhIGF-I), either separately or simultaneously with the artificial bone (Bio-oss) composite, implanted into dogs degreeⅡfurcation periodontal disease model, and with guided tissue regeneration (Guided issue regeneration, GTR) treatment, observation of animal models of periodontal tissue repair and regeneration, and to explore two kinds of growth factors to promote periodontal tissue regeneration and the effect of clinical treatment of periodontal disease in itsapplication to provide a theoretical basis.
Methods:
1. The formation of five dogs with chronicⅡfurcation of the mandibular first premolar 2,3,4, for a total of 48 teeth were randomly divided into 5 groups, including the three experimental groups, a control group and a blank control group, experimental group A:rhbFGF/rhIGF-I/ Bio-oss/GTR; experimental group B:rhbFGF/Bio-oss/GTR; experimental group C:rhIGF-I/Bio-oss/GTR; the control group D:GTR surgery alone; the blank control group E:flap operation.
2. Before implant rhbFGF and/or rhIGF-I composite bone graft and after 12 weeks,record buccal side of the bifurcation site of periodontal probing depth (periodontal depth,PD)and attachment level (attachment level,AL);after 2,4,12 weeks taken surgery X-rays to observe the new situation of alveolar bone.
3. Dogs were sacrificed 12 weeks after surgeried, slice the preparation experimental tissue zone (including dental, alveolar bone, periodontal ligament), HE, Masson staining and histological measurements, observe periodontal tissue (alveolar bone, teeth bone and periodontal membrane) of the new situation.
Results:
1. The clinical examination:after 12 weeks the PD and AL of dogs's model were improved to varying degrees, followed by A, B/C, D and E group;
2. X-ray examination:the experimental group A, B, C alveolar bone almost are filling the molar furcation, and the bone can be seen more clearly hard board; control group D were higher than the experimental group, less new bone, but more than the control group E; control group E are filling with a few sparse trabecular bone in the bottom of the low bone mineral density, bone hard plate fuzzy;
3. histological observation:HE staining, the experimental group A, B, C shows that new bone defect filling furcation, which become higher bone mineral density, calcium good; new born periodontal fiber small dense, rich in blood vessels, the direction perpendicular or nearly perpendicular to the root surface; new cementum covering most areas of the root bifurcation; the experimental group showed no bone adhesion and root absorption of malunion. Control group D, some new alveolar bone was layered over mature cementum covering the root surface to the coronal extension; fibers within the periodontal gap crossed. The control group E appears only a small number of individual defect new bone, and it has more epithelial cells and non-bone connective tissue covering. Masson staining in the experimental group A, B, C showed obvious red stained area with peripheral small amount of blue stained area; control group D Masson staining red-blue, red area of trabecular bone center; blank control group E showed obvious blue, surrounded by a small amount of red staining area;
4. tissue measurements can be seen in all experimental groups a high degree of new bone, new cementum height and new indicators of periodontal tissue were higher than that and the control group (P<0.05), compared with the experimental group, A group of the indicator, although numerically higher than B, C group was slightly higher, but the statistical analysis, three groups were not significantly different (P>0.05).
Conclusion:
1. Through artificial canine furcation bone defect site and supported by a stable method of local incentives that can be prepared reliable and similar to clinical periodontal disease in dogs II furcation Animal models, to be used for a variety of periodontal tissue regeneration therapy in animal study.
2. By clinical examination of animal models, histological observation, histological measurements, the results show rhbFGF and/or rhIGF-I and Bio-oss composite material has good bone and bone induction guide for dog II degree of root bifurcation defects can significantly promote alveolar bone, cementum and periodontal regeneration and new attachment formation.
3. The experimental results proved to use rhbFGF and/or rhIGF-I and bone composite to treatment canine periodontal diseaseⅡfurcation periodontal regeneration model is superior to simple guiding periodontal tissue regeneration.
4. The experimental results show that alone rhbFGF, rhIGF-I and these two factors combination on the periodontal tissue regeneration (regeneration of alveolar bone, cementum regeneration, periodontal membrane formation) the quality was no significant difference.
引文
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