原核表达重组人 OP-1促进拔牙创愈合的实验研究

目的：检测原核表达基因工程重组人OP-1(rhop—1)促进拔牙创愈合，减少牙槽嵴吸收的的效能方法：采用大白兔建立拔牙创的动物模型，以明胶海绵作为裁体，将大肠杆菌原核表达的重组人OP—1与之复合后植入即刻拔除牙齿的牙槽窝内进行干预治疗，通过观察其组织学改变、钙含量及碱性磷酸酶活性测定的变化，探讨原核表达的基因工程重组产品rhOP—1是否促进拔牙创的愈合修复。结果：大体观察结果显示：两组牙槽嵴高度的吸收差异有显著性；组织学HE切片显示：实验组的骨创愈合比对照组大约提前4～6周：碱性磷酸酶(AIP)活性及钙含量测定均显示实验组均明显高于对照组，差异有统计学意义：结论：原核表达基因工程重组人骨形态发生蛋白具有良好的促进骨创愈合、减少或防止牙槽骨吸收的作用     

重组人骨形成蛋白牙槽嵴内诱导成骨的实验研究

目的:探讨原核表达重组人骨形成蛋白-7牙槽嵴内诱导成骨的作用效果。方法:拔除大白兔的切牙建立动物模型,以明胶海绵作为载体,将原核表达的重组人骨形成蛋白-7与之复合后植入即刻拔除牙齿的牙槽窝内进行干预治疗,通过扫描电镜观察及钙含量测定,探讨原核表达重组人骨形成蛋白-7牙槽嵴内诱导成骨的作用效果。结果:扫描电镜观察显示:实验组的骨创愈合比对照组大约提前4~6周;钙含量测定显示实验组明显高于对照组,差异有统计学意义。结论:重组人骨形成蛋白-7具有良好的诱导牙槽嵴内成骨的效果。     

NPP/C-HA/rhBMP-2复合支架修复兔股骨缺损的实验研究

目的: 探讨NPP/C-HA/rhBMP-2人工骨在实验兔体内的成骨效果。方法: 在兔股骨远端做直径7 mm、深10 mm的骨缺损模型,根据缺损形状,制作含有rhBMP-2的NPP/C-HA人工骨,缺损处不植入材料作为空白组,植入NPP/C-HA人工骨为对照组,植入NPP/C-HA/rhBMP-2人工骨为实验组。第4、8、12周时,通过锥形束CT（CBCT）、H-E染色及Masson染色,观察各组成骨效应;检测各组血清中碱性磷酸酶（ALP）活性及组织中OCN阳性染色,确定缺损处成骨分化及骨成熟程度。采用SPSS 18.0软件包对数据进行统计学分析。结果: 大体标本显示,实验组12周时缺损处修复完全,未见炎症、排斥等免疫反应。CBCT显示,12周时,对照组从骨缺损边缘到中心处骨密度增高,但边界仍在;加入rhBMP-2后的实验组效果更加明显,新生骨与周围宿主骨基本一致,边界消失。第8、12周时,实验组HU值显著高于对照组及空白组（P<0.05）。H-E染色及Masson染色显示,第8、12周时,实验组较对照组及空白组有明显的新生骨形成。第8周时,各组血清ALP活性升高,实验组ALP水平显著高于其他组（P<0.05）。OCN免疫组织化学IHC阳性染色显示,实验组较对照组、空白组更明显（P<0.05）。结论: NPP/C-HA/rhBMP-2具有良好的组织相容性、骨诱导性、骨传导性和成骨性,有望成为骨缺损修复良好材料。     

骨形成蛋白-2基因促进犬牙周组织再生的体内研究

目的：观察重组人骨形成蛋白-2（rhBMP-2）质粒对犬牙周骨缺损再生作用的影响.方法：选用6只成年beagle犬,将其下颌两侧的第2、3、4前磨牙（P2、P3、P4）作为实验牙.在每颗实验牙的近中根颊侧制作＂U＂型骨缺损,并随机设计为3组,即pIRES-rhBMP-2组、rhBMP-2组（阳性对照组）和PBS组（空白对照组）.术后第4与第8周分别处死动物.光镜下观察组织学变化,采用Image-Pro-Express系统测量新生牙槽骨和牙骨质的高度、新生结缔组织的附着量.测量结果以SAS 6.12软件包进行Newman-Keulsq检验.结果：实验第8周时,组织学观察可见2个实验组新生牙槽骨接近于正常骨结构,新生牙周纤维稠密,富含血管;空白对照组胶原纤维稀疏,排列不规则.新生组织测量结果显示：2个实验组新生牙槽骨和牙骨质的高度、新生结缔组织的附着量较空白对照组显著增加（P＜0.05,P＜0.01）;但2个实验组间相比无显著性差异（P＞0.05）.结论：pIRES-rhBMP-2有较强的骨诱导活性,能有效促进犬牙周骨缺损的组织再生.     

复方骨形成蛋白生物膜植入牙槽窝的实验研究

目的:以骨形成蛋白、固定化凝血酶、头孢哌酮钠为主药,水溶性壳聚糖为主要膜材,制备复方骨形成蛋白牙槽生物黏附膜,植入大鼠牙槽窝观察其对拔牙后牙槽骨的影响。方法:选用大鼠54只,随机分为3组(实验组、单纯壳聚糖组、对照组),在拔除大鼠下颌切牙后牙槽窝内即刻植入复方骨形成蛋白生物膜、单纯壳聚糖及空白对照。3组动物于植入后3、6、9周时处死,切取下颌骨,用软X线机检查剩余牙槽嵴相对剩余高度、双能X线骨密度分析仪测定骨密度值、常规病理切片观察牙槽窝成骨情况。结果:植入后3、6、9周时实验组剩余牙槽嵴相对剩余高度和骨密度值均高于单纯壳聚糖组及对照组,差异有统计学意义(P<0.05);病理切片显示实验组牙槽窝成骨情况优于单纯壳聚糖组及对照组。结论:复方骨形成蛋白生物膜处方组成合理,是预防剩余牙槽骨吸收的有效方法之一,有一定的临床意义。     

对过大牙槽窝行自体牙移植的动物实验研究

目的研究对受区过大牙槽窝行自体牙移植的方法。方法3只Beagle犬的18颗切牙作为供体牙,分为3组,分别植入到前磨牙区已预备好,近远中向比牙根宽2mm的牙槽窝中。再生组提前7d预备受区,利用再生组织充填;骨材料组使用瑞安生骨替代物充填;对照组不做处理。术后3月大体观察、拍X片和组织学检测比较3组的预后。结果所有移植牙均健康存活,无松动脱落。再生组示牙周膜再生,牙骨质和牙槽骨表面无吸收。骨材料组牙根发生炎症性吸收,有新骨生成。对照组牙根发生表面吸收。结论再生组织可以促进牙周再生,并减少牙根表面吸收;瑞安生骨替代物的使用有助于移植牙的稳定和存活,但其对牙周再生的作用尚待进一步研究。     

rhBMP-2、胶原、珊瑚复合人工骨植入智齿牙槽窝的临床研究

目的：研究重组人骨形成蛋白（rhBMP-2)，胶原，珊瑚复合人工骨在修复口腔颌面部骨缺损和整复颌面畸形的作用。方法：将rhBMP-2，胶原，珊瑚复合人工骨植入阻生智齿拔除后的牙槽窝内，并设单纯珊瑚和空白对照组，术后进行临床和X线检查，记录邻近的第二磨牙远中牙槽嵴的高度。结果：术后12周复合骨组牙槽窝骨缺损和牙槽嵴高度均得到良好的恢复。珊瑚骨组得到较好的恢复，而空白对照组结果不明显，结论：rhBMP-2胶原，珊瑚复合人工骨植入智齿牙槽窝可较好地修复牙槽窝骨缺损及邻近的第二磨牙远中牙槽骨缺损。     

狗牙周缺损处移植自体骨髓干细胞的骨化实验观察

目的 对应用自体骨髓干细胞移植的动物骨化实验的观察进行评价。方法 将6只成年狗的36颗牙，分为实验组和对照组（各18颗牙）。有人工制造的牙周缺损中进行体外培养的自体骨髓干细胞移植，结合引导组织再生（guided tissue regeneration，GTR）方法（实验组）和单纯GTR方法（对照组）。6周后对切片行牙周组织学观察。结果 实验组新牙槽骨骨化效果明显好于对照组。实验组建立了正常钙化骨结构，Masson染色为红色；对照组虽然建立了正常骨结构，但钙化程度极低，基本为胶原性骨结构，Masson染色为蓝绿色。结论 应用自体骨髓干细胞移植结合聚四氟乙烯膜引导牙周组织再生可促进牙周组织的再生、加快正常骨组织结构的建立，并缩短修复再生时间。     

活性纳米羟基磷灰石复合胶原/聚乳酸材料对拔牙创早期愈合及牙槽嵴吸收影响的实验研究

目的 观察活性纳米羟基磷灰石复合胶原/聚乳酸材料(AnHAC/PLA)促进犬拔牙创早期愈合及减少牙槽嵴吸收的效果。方法 将小块状纳米羟基磷灰石复合胶原/聚乳酸材料(nHAC/PLA)、AnHAC/PLA、自体牙槽松质骨植入犬拔牙创中,并设空白对照。术后4周、8周、12周通过大体观察测量、HE染色和M asson′s三色法染色来观察比较拔牙创骨缺损的修复情况及牙槽嵴的吸收情况。结果 AnHAC/PLA材料修复拔牙创骨缺损及减少牙槽骨吸收的能力强,与自体牙槽松质骨组类似,nHAC/PLA材料的能力较弱,但明显优于空白对照组。结论 AnHAC/PLA可早期修复拔牙创,减少牙槽嵴的吸收。     

聚乳酸、rhBMP-2复合体在牙槽骨修复中作用的实验研究

目的研究聚乳酸与rhBMP-2的复合体在牙槽骨修复中作用，寻找一种适合临床应用促进牙槽骨再生的新型的生物材料。方法杂种犬6只，随机分为2组，拔除每只犬的上下颌前磨牙，右侧拔牙创填入聚乳酸与rhBMP-2的复合体，左侧拔牙创自然愈合作对照，于2周、6周和10周行X线检查，于6周和10周分别处死2组犬，取拔牙创标本行光镜和扫描电镜观察对比两组成骨状况及植入材料在组织内的反应。结果X线检查及病理结果显示右侧拔牙创愈合快，成骨速度及成骨质量明显优于左侧自然愈合的拔牙创。电镜显示该材料组织相容性好，无炎性刺激反应，可降解。结论聚乳酸与rhBMP-2复合体是一种促牙槽骨再生的新型生物复合材料．可望成为临床实际应用的修复牙槽骨缺损的新型材料。     

Experimental study of secondary bone graft of alveolar clefts using bone morphogenetic protein (BMP)

The purpose of this study was to evaluate the utility of recombinant human bone morphogenetic protein-2 (rhBMP-2) as a bone substitute on artificial alveolar clefts of adult Macaca fuscata monkeys. First, we created simulated alveolar clefts to represent human bilateral alveolar clefts, and then we implanted a mixture of rhBMP-2 and autogeneous particulate marrow cancellous bone (PMCB) into these models, as experimental groups. The mixture ratio was varied as follows (mixture ratio = rhBMP-2: PMCB). Group 1: rhBMP-2 alone. Group 2: Mixture of rhBMP-2 and PMCB (1:1). Group 3: Mixture of rhBMP-2 and PMCB (3:1). Group 4: Mixture of rhBMP-2 and PMCB (4:1). Positive control group: PMCB alone. Negative control group: PLGA (poly[L-lactide-co-glycolide] copolymer/gelatin sponge complex (PGS) alone. All animals were sacrificed 12 weeks after implantation and evaluated radiographically and histologically. The results were as follows. Bone bridge formation was not observed in Group 1, using rhBMP-2 alone, but it was observed in Groups 2, 3, and 4 using mixture of rhBMP-2 and PMCB. The vertical height of bone bridge formation in Group 4 was equal to that in the positive control group, roughly. The histological finding showed that the bone density of Group 4 was higher than in the positive control group, and the structure of cancellous bone turned into mature type. Bone formation and remodeling was active in Group 4. These results indicated that rhBMP-2 may be an effective substitute for autogenous PMCB in bone grafting into alveolar clefts.     

一种新型复合人工骨修复家兔下颌骨骨缺损的扫描电镜观察

目的探讨珊瑚转化型羟基磷灰石-胶原-重组人骨形成蛋白-2(CHA-Co-rhBMP-2)复合人工骨在修复颌骨缺损上的应用价值,为临床应用提供理论依据。方法健康家兔18只,随机分为实验组和两组对照组,每组6只。在双侧下颌骨体部制造1.0cm×1.0cm贯通性骨缺损,实验组用CHA-Co-rhBMP-2复合人工骨充填修复,对照组分别用CHA-Co复合骨和单纯CHA充填修复。每组动物各取3只分别在术后6周、12周处死,进行组织学观察及统计学分析。结果与对照组相比实验组材料具有明显的骨诱导活性,6周可见明显新骨形成,12周材料大部分降解,降解区域被大量成熟骨组织所取代,骨材料界面结合紧密,无纤维结缔组织。骨密度分析结果认为骨缺损区的骨密度随时间延长有逐渐增高趋势,统计学分析实验组与对照组之间骨密度有显著性差异。较对照组具有明显优势。实验动物均无感染和不良反应出现。结论CHA-Co-rhBMP-2复合人工骨具有良好的生物相容性、降解性及骨诱导活性,是一种理想的颌骨缺损修复材料。     

Effect of recombinant human bone morphogenetic protein-4 dose on bone formation in a rat calvarial defect model

BACKGROUND: Bone morphogenetic proteins (BMPs) are being evaluated for periodontal and bone regenerative therapy. The objective of this study was to evaluate the effect of recombinant human bone morphogenetic protein-4 (rhBMP-4) dose on local bone formation in a rat calvaria defect model. METHODS: Calvarial, 8 mm diameter, critical-size osteotomy defects were created in 140 male Sprague-Dawley rats. Seven groups of 20 animals each received either 1) rhBMP-4 (2.5 microg) in an absorbable collagen sponge (ACS) carrier, 2) rhBMP-4 (5 microg)/ACS, 3) rhBMP-4 (2.5 microg) in a beta-tricalcium phosphate (beta-TCP) carrier, 4) rhBMP-4 (5 microg)/beta-TCP, 5) ACS or 6) beta-TCP carrier controls, or 7) a sham-surgery control, and were evaluated by histologic and histometric parameters following a 2- or 8-week healing interval (10 animals/group/healing interval). RESULTS: Surgical implantation of rhBMP-4/ACS and rhBMP-4/beta-TCP resulted in enhanced local bone formation at both 2 and 8 weeks. Within the dose range examined, rhBMP-4 did not exhibit an appreciable dose-dependent response. Defect closure was not significantly different between the rhBMP-4/ACS and rhBMP-4/beta-TCP groups. New bone area of the rhBMP-4/ beta-TCP group was significantly greater than that of the rhBMP-4/ ACS group; however, bone density in the rhBMP-4/ACS group was significantly greater than that in the rhBMP-4/beta-TCP group at 8 weeks (P < 0.05). CONCLUSIONS: rhBMP-4 combined with ACS or beta-TCP has a significant potential to induce bone formation in the rat calvaria defect model. Within the selected rhBMP-4 dose range and observation interval, there appeared to be no meaningful differences in bone formation.     

A histological study on experimental tooth movement into bone induced by recombinant human bone morphogenetic protein-2 in beagle dogs.

OBJECTIVE: The purpose of this preliminary study was to examine experimental tooth movement into newly generated bone induced by recombinant human bone morphogenetic protein-2 (rhBMP-2). METHOD: After extraction of the maxillary first premolars, bone defects were surgically created in eight adult beagle dogs using a 5-mm-diameter trepan bar. According to which material was grafted into the bone defects, animals were divided into the following four groups: (1) the rhBMP-2 group in which rhBMP-2 with a poly[ D,L-(lactide-co-glycolide)]/gelatin sponge complex was implanted; (2) the spongiosa group in which spongiosa from the tibia was grafted; (3) the nongrafted group in which no material was embedded; and (4) the control group in which only tooth extraction was performed. The osteoinductive activity of rhBMP-2 and tooth movement into the newly generated bone were examined by histological and morphometric comparisons of each group. RESULTS: Considerable new bone formation was observed at the grafted site both in the rhBMP-2 and in the spongiosa groups. The area of generated bone in the rhBMP-2 group was significantly greater than that in the spongiosa group. Newly generated bone, in both the rhBMP-2 and spongisosa groups, showed a similar histological response to orthodontic force as in normal alveolar bone in the control group. However, root resorption occurred on the pressure side in the rhBMP-2 group. CONCLUSION: These results indicated that rhBMP-2 might constitute an alternative material to autogeneous bone grafting for alveolar cleft defects. Further studies regarding tooth movement into generated bone induced by rhBMP-2 are suggested.     

Effect of recombinant human bone morphogenetic protein-2, -4, and -7 on bone formation in rat calvarial defects

BACKGROUND: Currently, more than 20 bone morphogenetic proteins (BMPs) have been identified, and many trials have been carried out using recombinant human BMPs (rhBMPs) for bone tissue engineering. However, comparative analyses on bone formative activities of rhBMP using a preclinical model have been limited. Therefore, the aim of this study was to evaluate and compare the osteogenic potential of rhBMP-2, -4, and -7 delivered with absorbable collagen sponge (ACS) upon early (2 weeks) and complete (8 weeks) wound healing phases in a critical sized rat calvarial defect model. METHODS: Eight-millimeter critical sized calvarial defects were created in 30 male Sprague-Dawley rats. The animals were divided into three groups of 10 animals each. The defects were treated with 0.025 mg/ml rhBMP-2/ACS, rhBMP-4/ACS, or rhBMP-7/ACS. The rats were sacrificed at either 2 (five rats) or 8 (five rats) weeks after surgery, and the results were evaluated histologically, histomorphometrically, and immunohistometrically. RESULTS: The surgical implantation of rhBMP-2/ACS, rhBMP-4/ACS, or rhBMP-7/ACS resulted in enhanced local bone formation in the rat calvarial defect model at both 2 and 8 weeks. The amount of defect closure, new bone area, and bone density were similar in the three groups at each time point (P > 0.05). In terms of bone density and new bone area, there were statistically significant differences between results obtained at 2 weeks and those obtained at 8 weeks in all groups (P < 0.05). Two-way analysis of variance (ANOVA) revealed that there was no correlation between the time and conditions (P > 0.05), but time was found to have a strong influence on defect closure, new bone area, and bone density (P < 0.05). Irrespective of rhBMP type, positive immunoreactions of osteopontin (OPN) and osteocalcin (OCN) were evident at 2 and 8 weeks. Intense OPN and OCN staining was observed near the newly formed bone as well as in some cells within the new bone. CONCLUSIONS: Within the rhBMP types used, rhBMP concentration, and the observation interval, there appears to be no specific differences in bone regenerative potential. All rhBMPs used in this study may be considered effective factors for inducing bone formation.     

Effect of recombinant human bone morphogenetic protein-2 in an absorbable collagen sponge with space-providing biomaterials on the augmentation of chronic alveolar ridge defects

BACKGROUND: Recombinant human bone morphogenetic protein-2 (rhBMP-2) in an absorbable collagen sponge (ACS) carrier has been shown to support significant bone formation in the craniofacial skeleton. When used as an onlay, however, rhBMP-2/ACS may become compressed with limited resulting bone formation. The objective of this study was to evaluate the effect of two space-providing biomaterials, bioactive glass (BG) and demineralized/mineralized bone matrix (DMB), on rhBMP-2/ACS induced alveolar ridge augmentation. METHODS: Bilateral alveolar ridge defects were produced in the mandible in six mongrel dogs. rhBMP-2/ACS with biomaterials was surgically implanted into contralateral defects in four animals. Treatments were alternated between jaw quadrants in consecutive animals. Two animals received rhBMP-2/ACS or sham-surgery in contralateral defects. The animals were injected with fluorescent bone labels to monitor bone formation. Clinical evaluations were made at ridge augmentation and 12 weeks post-implantation when the animals were euthanized and block biopsies collected for histopathologic evaluation. RESULTS: Sham-surgery produced limited horizontal alveolar augmentation (0.1 +/- 0.6 mm). Implantation of rhBMP-2/ACS resulted in alveolar augmentation amounting to 2.2 +/- 1.8 mm. Alveolar augmentation in sites receiving rhBMP-2/ACS with DMB or BG was 2-fold greater compared to rhBMP-2/ACS alone averaging 4.4 +/- 1.3 and 4.6 +/- 1.5 mm, respectively. The DMB biomaterial appeared substituted by newly formed bone. The BG particles were observed imbedded in bone or encapsulated in dense connective tissue without associated bone metabolic activity. Fluorescent light microscopy suggested that the new bone was formed within 4 weeks. CONCLUSION: The bioglass and demineralized/mineralized bone matrix biomaterials utilized in this study in combination with rhBMP-2/ACS supported clinical and histological ridge augmentation.     

Enhanced vertical alveolar bone augmentation by recombinant human bone morphogenetic protein-2 with a carrier in rats

This study aimed to evaluate the effect of recombinant human bone morphogenetic protein-2 (rhBMP-2) on vertical bone regeneration of edentulous ridge. Bilateral upper first and second molars of 8-week-old Wistar rats were extracted and the ridges were allowed to heal for 3 weeks. Compressed poly(lactic-co-glycolic acid) copolymer/gelatin sponge (PGS) was used as a carrier of rhBMP-2. PGS alone (control group) or PGS with 5 mug rhBMP-2 (test group) was implanted at the top part of alveolar ridge. The sham group received no implantation. The rats were killed at 1, 2, 4, 8 and 12 weeks after implantation and examined histologically and histomorphometrically. In the test group, significant bone augmentation was evident on the alveolar ridge throughout the experimental period. Histomorphometric analysis revealed greater tissue volume and height of alveolar bone in the test group compared with the control and sham groups (P < 0.05) from 4 weeks onward and the augmented tissues (5 mm3 in tissue volume and 1.5 mm in bone height) were maintained until 12 weeks. Osteoblast surface increased at 2 and 4 weeks and osteoid thickness reached a peak (25 microm) at 2 weeks. Dynamic variables, which represented calcification, were higher in the test group than the control and sham groups at 4 and 8 weeks (P < 0.05). These results suggest that use of rhBMP-2/PGS may achieve vertical bone augmentation, and stabilizes denture prosthesis or makes up for inadequate bone mass for implant prosthesis.     

Ectopic bone formation after implantation of a slow release system of polylactid acid and rhBMP-2

Objectives: The present study was conducted to test the hypothesis that preshaped polylactic acid (PLA) implants loaded with recombinant human bone morphogenic protein 2 (rhBMP-2) can induce bone formation in a rat ectopic model.
Materials and methods: Two groups of porous cylindrical poly- dl -lactic acid implants of 8-mm diameter were produced by gas foaming with CO₂, incorporating 48 and 96 μg rhBMP-2, respectively, into each implant. Blank PLA implants were used as controls. The release of BMPs and the induction of alkaline phosphatase were assessed in vitro . Osteoinduction in vivo was tested by insertion of 15 implants from each group into the gluteal muscles of Wistar rats. Five implants from each group were retrieved after 6, 13 and 26 weeks and assessed using flat panel volume detector computed tomography and light microscopy.
Results: Both groups of implants showed increased release of rhBMP-2 during the first 24–48 h, with a slightly higher amount being released from the implants with 48 μg. Release during subsequent intervals was <100 ng/72 h in the low-concentration group and >100 ng in the group with 96 μg rhBMP-2. Implants with 95 μg rhBMP-2 exhibited bone formation in vivo on the outside of the implants across the observation period of 26 weeks with invasion of bone into the pores, whereas implants with 48 μg rhBMP-2 failed to induce the formation of bone tissue. No bone formation was found in the control implants.
Conclusions: The results suggest that release rates of rhBMP-2 for ectopic bone induction have to be >100 ng/72 h to maintain the osteoinductive activity of the tested porous PLA implants. This slow release system may have impact on alveolar bone augmentation procedures when used as individually preformed osteoinductive implants.