珊瑚转化多孔羟基磷灰石复合rhBMP-2人工骨的动物实验研究

目的：寻找新的骨代用材料。方法：采用珊瑚转化多孔羟基磷石（ＣＨＡ）作为移植物支架,用Ｉ型胶原作为ｒｈＢＭＰ－２的缓释载体及赋形剂,将三者复合制成新型人工骨,进行了大鼠标准颅骨缺损骨移植实验。结果：ＣＨＡ／胶原具有良好的生物相容性和骨引导作用。ｒｈＢＭＰ－２／ＣＨＡ／胶原具有骨引导和骨诱导双重特性,其成骨效应明显优于单纯ＣＨＡ／胶原植入。结论：该复合人工骨有可能成为临床修复骨缺损的有效材料。     

重组人骨形成蛋白-2/珊瑚复合人工骨的动物实验研究

本研究将重组人骨形成蛋白－２（ｒｈＢＭＰ－２）和珊瑚以一定的方式复合后,植入小鼠股部肌袋和兔颅骨标准大小缺损,以单纯珊瑚植入作对照,术后不同时间取材,通过组织学方法检测其骨诱导活性和骨修复能力．结果显示：ｒｈＢＭＰ－２／珊瑚复合人工骨植入小鼠肌袋１周,诱导软骨形成,３周,形成编织骨,６周,形成含骨髓的板层骨,同时,珊瑚被部分降解吸收;复合人工骨植入兔颅骨缺损后,以引导成骨和诱导成骨双重机制完成骨修复过程,术后１２周,植入物完全被成熟的骨组织取代,其骨修复效果明显优于单纯的珊瑚．此复合人工骨具有骨传导和骨诱导活性,骨修复能力较强,是一种较理想的新型生物性植骨材料     

Implant system for guiding a new layer of bone. Computed microtomography and histomorphometric analysis in the rabbit mandible

Objective: To prove the concept that an implant system with osteoconductive surface characteristics and an osteoinductive scaffold material has the capacity to guide vertical supracrestal bone growth in a rabbit mandible onlay model.
Material and methods: Thirteen adult white New Zealand rabbits each received custom-designed dental implants. All implants had sandblasted, acid-etched (SLA^™) surfaces, with the coronal aspect (3 mm) of each implant was left outside the lateral aspect of posterior mandibular bone, but covered by periosteum, muscle, subcutaneous tissue, and skin. Bone formation around implants placed adjacent to osteoinductive demineralized bone matrix (DBM) scaffolds were compared with contralateral implants without scaffolds in six rabbits using micro-CT imaging. Bone formation around implants with scaffolds from seven additional rabbits was measured using both micro-CT imaging and quantitative histology.
Results: At 8 weeks, new supracrestal bone was seen adjacent to all implants placed with DBM and two implants without DBM. The mean supracrestal bone heights achieved for implants with and without DBM scaffolds as measured by micro-CT was 2.1±0.9 and 0.8±0.9 mm, respectively ( P =0.008). Histomorphometric analysis illustrated that supracrestal bone-to-implant contact for implants with DBM scaffolds was 58.1±14% and that mean supracrestal bone height was 2.4±0.6 mm.
Conclusions: Successful implant-guided supracrestal osteogenesis has been demonstrated in a rabbit model with the combined use of osteoconductive implant surfaces, an osteoinductive scaffold, and a device that prevents soft tissue downgrowth and provides scaffold stabilization.     

Bone augmentation by recombinant human BMP-2 and collagen on adult rat parietal bone.

A composite of recombinant human bone morphogenetic protein-2 (rhBMP-2) and collagen was implanted beneath the cranial periosteum of 10-month-old rats to observe bone development and absorbent change of carrier collagen. The rhBMP-2/collagen onlay implant resulted in active bone formation and the augmented bone was connected directly with the original bone, whereas the collagen alone resulted in neither bone nor cartilage. The ossification process in the rhBMP-2/collagen occurred directly through bone formation, similar to intramembranous ossification. The carrier collagen fibers were found in the woven bone and were completely absorbed at 8 weeks in the presence of rhBMP-2, while the collagen alone implant remained encapsulated by a thin, fibrous connective tissue. Our results indicate that rhBMP-2/collagen is an effective material as a biological onlay implant, showing osteoinductive properties and being completely replaced by new bone. Carrier collagen not only plays a role in rhBMP-2 delivery, but also provides a cell anchorage for cell differentiation and remains as an artificial matrix in woven bone.     

The effect of Emdogain and platelet-derived growth factor on the osteoinductive potential of hydroxyapatite tricalcium phosphate

The aim of this study was to determine whether hydroxyapatite β-tricalcium phosphate (HA-TCP) either alone or coated with Emdogain (EMD) or recombinant human platelet-derived growth factor-BB (rhPDGF-BB) becomes osteoinductive in the murine thigh muscle model for osteoinduction. Twenty CD1 adult male mice had gelatin capsules implanted into the thigh muscle of both hind limbs. The capsules were either empty or contained one of the following: uncoated particulate HA-TCP, EMD-coated HA-TCP or rhPDGF-BB-coated HA-TCP. The implant sites were assessed histologically at 4 and 8 weeks. A semi-quantitative histological examination was performed to assess the inflammatory changes, reparative processes and osteoinduction within the graft site. At both 4 and 8 weeks, histological analysis failed to demonstrate any osteoinductive activity in any of the specimens from the experimental groups. A minimal chronic inflammatory response and foreign body reaction around the implanted materials was seen which reduced over time. The HA-TCP particles were embedded within fibrous connective tissue and were encapsulated by a dense cellular layer consisting of active fibroblasts and occasional macrophages with the thickness of this layer decreasing over time. The results of this study suggest that the use of commercially available HA-TCP alone or in combination with EMD or rhPDGF-BB is biocompatible but not osteoinductive in the murine thigh muscle model of osteoinduction. Coating HA-TCP with EMD or rhPDGF-BB does not enhance its osteoinductive potential.     

Hydroxyapatite cranioplasty: I. Experimental results from a new quick-setting material

The biophysical characteristics of a new formulation of hydroxyapatite (HA) for craniofacial applications having a rapid set time was evaluated in animals. Using a mature rabbit model, the material was applied in both cranial defects and as a geometrically shaped cranial onlay. Visual and histological assessment was done after 1 year of implantation. The material remained morphologically stable after 1 year, with complete preservation of its onlay shape and stable fill of the cranial defects. A significant osteoconductive response was seen with bone growth across the inner and outer surfaces of the inlays and along the sides of the onlays. No fibrovascular ingrowth or bony replacement of the material itself was evident as a result of the age of the study animals or the compact density of the material. This rapid-setting HA formulation offers similar favorable tissue responses as those of other longer setting HA materials.     

Rat subcutaneous tissue response to macrogranular porous anorganic bovine bone graft

The ideal bone graft must present biocompatibility, osteoconductive and osteoinductive properties, resistance and plasticity. Xenogenic grafts of bovine cancellous bone origin are particularly interesting due to their biologically designed porous structure that enhance both cellular and vascular invasion. The purpose of this study was to evaluate the tissue response induced by bovine macrogranular porous anorganic bone implanted in rat subcutaneous tissue. Forty rats were assigned to 2 groups, as follows: the control group received empty collagen capsules and the test group received subcutaneous implants of the test material. Samples were collected after 10, 20, 30 and 60 days and processed histologically. Histological analysis showed at 10 days a granulomatous inflammatory infiltrate, rich in multinucleated giant cells and free of lymphocytes or plasma cells, similarly to mineralized allograft implanted in rat subcutaneous. In later periods, there was a significant decrease in the inflammatory infiltrate and an increase in fibrosis around graft particles. In conclusion, the test material induced a foreign body-type granuloma with subsequent fibrosis around the graft particles implanted in rat subcutaneous and did not elicit any immune response, thus being considered biocompatible.     

Alveolar ridge augmentation in rats by combined hydroxylapatite and osteoinductive material

Abstract— The purpose of this study was to examine if increased bony ingrowth developed when hydroxylapatite was implanted together with an osteoinductive substrate. Dense hydroxylapatite granules (HA) (Calcitite, Calcitek, San Diego, CA, USA) were mixed with equal volumes of allogenic, demineralized, lyophilized dentin or bone and implanted subperiosteally for alveolar ridge augmentation purposes and heterotopically in the abdominal muscles in rats. Light microscopic evaluation revealed that HA was surrounded by fibrous connective tissue containing foreign body giant cells and it had neither an osteoinductive nor an osteoconductive effect. The newly formed bone induced from the implanted allogenic, demineralized, lyophilized dentin or bone was never found in close contact with the HA and did not incorporate the implant.     

Accelerated endochondral osteoinduction in the absence of bone matrix particles in a rat model system.

Ethanol-precipitated proteins obtained from demineralized rat bone powder (DBP) by 4M guanidine-HCl extraction have been shown to reproducibly induce ectopic endochondral bone formation when subcutaneously implanted in rats in the absence of bone matrix particles. Histologic and biochemical analysis revealed a temporal sequence of chondrocyte differentiation, calcified cartilage formation, neovascularization, osteoblast differentiation, bone formation, osteoclastic bone remodeling, and hematopoietic marrow development that is complete by 21 days. In contrast to previous reports, these results clearly show an osteoinductive response independent of the presence of insoluble extracellular bone matrix. Compared with conventional DBP implants, the guanidine-extractable protein (GE) produces an accelerated and more robust osteoinductive response. Histologically, the initial chondrogenic response at days 6 to 9 is greatly amplified. Alkaline phosphatase specific activity peaks at day 9, several days earlier than for DBP, and is sixfold higher. Calcium accumulation in GE implants at day 12 is fivefold greater than with DBP, and all mineral is localized within the matrix of newly calcified cartilage and new bone. Osteoclasts are up to ninefold more abundant in the rapidly remodeling GE ossicle, making space for hematopoietic marrow. Delivery of GE coprecipitated with inert bone matrix particles was also more effective than DBP, although the response was somewhat attenuated compared with GE alone. Bony filling of 4-mm defects in rat mandibular rami was elicited by 10 mg of GE and followed an endochondral process with increased neovascularization compared with DBP and unimplanted controls. This guanidine-extractable protein fraction should prove useful for inducing quantities of chondrocytes and osteoclasts for in vitro study, and for analysis of osteoinductive requirements.(ABSTRACT TRUNCATED AT 250 WORDS)     

Alveolar ridge augmentation in rats by combined hydroxylapatite and osteoinductive material.

The purpose of this study was to examine if increased bony ingrowth developed when hydroxylapatite was implanted together with an osteoinductive substrate. Dense hydroxylapatite granules (HA) (Calcitite, Calcitek, San Diego, CA, USA) were mixed with equal volumes of allogenic, demineralized, lyophilized dentin or bone and implanted subperiosteally for alveolar ridge augmentation purposes and heterotopically in the abdominal muscles in rats. Light microscopic evaluation revealed that HA was surrounded by fibrous connective tissue containing foreign body giant cells and it had neither an osteoinductive nor an osteoconductive effect. The newly formed bone induced from the implanted allogenic, demineralized, lyophilized dentin or bone was never found in close contact with the HA and did not incorporate the implant.     

Maxillary sinus floor elevation using a combination of DFDBA and bovine-derived porous hydroxyapatite: a preliminary histologic and histomorphometric report

The objective of the study was to determine the osteoconductive potential of bovine-derived porous hydroxyapatite (HA) in combination with demineralized freeze-dried bone allograft (DFDBA) as an alternative to autogenous grafting in the maxillary sinus. The study involved 5 patients treated with 2-stage sinus elevation procedures using a combination of DFDBA and Osteograf/N 300 and 700. The healing time before implant placement ranged from 6 to 13 months. At the time of reentry, a bone core was harvested from each patient and processed for histologic and histomorphometric analysis. Woven and lamellar bone formation was evident in all specimens. Mean trabecular bone volume was 27.92%. The amount of newly formed bone was positively correlated with healing time. The range of new bone formation was 5.36% (6 mo) to 43.68% (12 mo). Residual HA graft particles were evident in all specimens, and the amount was inversely correlated with time. HA particles were often surrounded by an intense inflammatory infiltrate. DFDBA particles, largely present in the 6-month biopsy, were not recognizable in the 10-, 12-, and 13-month specimens, suggesting complete replacement. The combination of Osteograf/N and DFDBA appears to be osteoconductive and may be considered a valid alternative to autogenous bone grafts in sinus lift procedures. Histomorphometric and histologic evaluation may also be used to monitor the status of the future implant site.     

rhBMP-2、PRF和自体骨分别复合珊瑚羟基磷灰石成骨效能的比较研究

目的：通过建立动物骨缺损模型,比较重组人骨形成蛋白-2（rhBMP-2）与珊瑚羟基磷灰石（CHA）复合物、富血小板纤维蛋白（PRF）与珊瑚羟基磷灰石复合物、自体骨与珊瑚羟基磷灰石复合物以及单纯珊瑚羟基磷灰石这四种骨移植材料在骨缺损中的成骨效能。方法：在比格犬双侧胫骨干骺端制备四个相同的骨缺损区,在缺损区分别植入rhBMP-2/CHA、 PRF/CHA、自体骨/CHA及CHA （对照）;3个月后处死动物,行大体标本观察;拍牙科CT,观察各植骨区骨密度情况;制作石蜡切片、 HE染色,比较各植骨区骨组织学特点及新骨形成量。结果：大体标本见四组骨缺损间隙均完全关闭。 X线示自体骨/CHA组和PRF/CHA组骨密度较致密, rhBMP-2/CHA组致密性低于前两者, CHA组未见明显骨致密影。 HE切片见四组新生骨与宿主骨连接紧密,新生骨小梁不规则,粗细不一,排列无序;复合型骨移植材料的新生骨小梁比对照组更密集、粗大,连续性更好;四组植骨区成骨量比较：自体骨/CHA组〉PRF/CHA组〉rhBMP-2/CHA组〉CHA组。结论：复合型骨移植材料成骨效应明显优于单纯珊瑚羟基磷灰石;三种复合型材料中自体骨/CHA成骨效应最好,其次为PRF/CHA, rhBMP-2/CHA最差。     

Experience with freeze-dried PGLA/HA/rhBMP-2 as a bone graft substitute.

We investigated bone induction by recombinant human bone morphogenetic protein-2 in rodents. The purpose of this study was to evaluate the osteoinductive potential of a resorbable bone substitute fabricated from freeze-dried poly(glycolic acid-co-lactic acid) (PGLA) mixed with hydroxyapatite particles incorporated with bone morphogenetic protein-2 in skull defects of rats (FD-PGLA/HA/rhBMP-2). The FD-PGLA/HA/rhBMP-2 composite or as a control, the FD-PGLA/HA by itself were implanted in skull defects (psi 8 mm) of rats. The samples were harvested at 2 or 4 weeks postoperatively and were studied radiographically and histologically. Four weeks after implantation, the FD-PGLA/HA/rhBMP-2 discs were completely replaced by newly-formed bone possessing bone marrow. In contrast, the defects implanted with FD-PGLA/HA were filled only with fibrous connective tissue. The results suggest that the FD-PGLA/HA/rhBMP-2 composite could be an optimum bone substitute with osteoinductive potential and could function as an alternative bone graft material for autogenous bone in humans.     

Osteogenesis in composite grafts of allogenic demineralized bone powder and porous hydroxylapatite

This study evaluated the osteoinductive capabilities of porous hydroxylapatite (PH) and the tissue response following intramuscular implantation of PH alone or in combination with demineralized bone powder (DBP). Six rhesus monkeys each received the implants in three separate soft tissue pockets created in the thoracic region. The implants consisted of 2 cc of either PH alone or DBP alone, or a 1:1 combination of DBP and PH. Two animals were killed at 2 weeks, two at 4 weeks, and two at 12 weeks postimplantation. Histologically, bone formation was seen in the DBP mixed with PH and in the DBP group as early as 4 weeks postimplantation; bone was also occasionally observed within the pores of the PH particles. The PH alone showed no evidence of formation of bone and the material was surrounded by a thick layer of fibrous tissue. It was concluded that PH is not osteoinductive, but can act as a scaffold for growth of bone, and that when mixed with an osteoinductive material, it allows for formation of bone within the implant.     

Bone repair analysis in a novel biodegradable hydroxyapatite/collagen composite implanted in bone

The purpose of this study was to evaluate a biodegradable hydroxyapatite/collagen composite and to examine the use of the calcium ion contained for bone formation and growth. Surgical holes were prepared in the femora and tibiae of beagle dogs, and were filled with the hydroxyapatite/collagen composite labeled with alizarin red. After 4 weeks, calcein was administered to the experimental dogs. After 1 additional week, the femora and tibiae were removed surgically and fixed in formalin. Light microscopy and confocal laser scanning microscopy were used to examine the surgical holes with their implanted materials and the surrounding bone. There were only a few inflammatory cells adjacent to the hydroxyapatite/collagen composite. The newly formed bone in the cortical bone was stained with calcein, which binds to serum calcium, and new bone near the hydroxyapatite/collagen composite in the holes was stained positive for alizarin red, which binds to the calcium in the hydroxyapatite/collagen composite. In addition, osteoblasts near the hydroxyapatite/collagen composite as well as newly formed bone adjacent to the osteoblasts showed alizarin red staining, but the new bone at a distance from the hydroxyapatite/collagen implant reacted only to calcein staining. These results, using the tissue labeling method with calcein and alizarin red, suggested that the calcium bound to the alizarin red released from the hydroxyapatite/collagen composite materials might have been translocated to sites of new bone formation. The present experiment showed that the novel hydroxyapatite/collagen composite is a useful implant material for bone augmentation and that the calcium in the newly formed bone might have been released from the implant.     

Onlay bone augmentation on mouse calvarial bone using a hydroxyapatite/collagen composite material with total blood or platelet-rich plasma

ObjectiveThe aim of this study was to assess newly formed onlay bone on mouse calvarial bone using a new artificial bone material, a hydroxyapatite/collagen composite, with total blood or platelet-rich plasma.DesignThe hydroxyapatite/collagen composite material with normal saline, total blood or platelet-rich plasma was transplanted on mouse calvarial bone. The mice were sacrificed and the specimens were harvested four weeks after surgery. The newly formed bone area was measured on hematoxylin and eosin stained specimens using Image J software.ResultsThe hydroxyapatite/collagen composite materials with total blood or platelet-rich plasma induced a significantly greater amount of newly formed bone than that with normal saline. Moreover, bone marrow was observed four weeks after surgery in the transplanted materials with total blood or platelet-rich plasma but not with normal saline. However, there were no significant differences in the amount of newly formed bone between materials used with total blood versus platelet-rich plasma.ConclusionsThe hydroxyapatite/collagen composite material was valid for onlay bone augmentation and this material should be soaked in total blood or platelet-rich plasma prior to transplantation.     

rhBMP-2在种植体周围骨缺损修复中应用的组织学观察

目的:研究rhBMP-2及不同载体在种植体周围骨缺损修复中的应用。方法:在beagle犬下颌骨植入种植体,颊侧形成裂开性骨缺损,置入复合了不同浓度rhBMP-2的珊瑚羟基磷灰石人造骨(CHA)或可吸收胶原海绵(ACS)。种植体植入后2、4、8、12周,获取含种植体骨标本,进行组织学观察。结果:2周时,rhBMP-2组可见极少量的新生骨组织。4周时,rhBMP-2/ACS组新骨组织由牙槽骨顶端向缺损区中心方向生长;rhBMP-2/CHA组人造骨颗粒内部和周围出现呈岛状生长的新生骨组织。8周时,rhBMP-2/ACS组的新骨形成大片状结构;rhBMP-2/CHA组人造骨颗粒周围较多骨岛形成。12周时,rhBMP-2组的缺损区内骨量和骨高度进一步增加,与种植体形成骨性结合。浓度为0.05 mg/ml和0.2 mg/ml,载体为CHA或ACS促进骨再生作用差异无统计学意义。结论:以CHA或ACS为载体rhBMP-2能促进种植体周围骨缺损区内的骨组织再生并与种植体表面较好地结合。     

Enhancement of bone ingrowth into collagen/HA composite implants using e-PTFE membranes

The objective of this investigation was to study the effectiveness of expanded polytetrafluoroethylene (e-PTFE) membranes for enhancement of bone ingrowth through subperiosteally implanted collagen/HA composite blocks. Twelve rabbits aged 12-15 months served as the experimental animals in this study. Two compressed Collagen/HA composite blocks in the shape of two attached cylinders of different diameters were inserted into two defects of each rabbit calvarium, the smaller cylinder being intrabony, the larger subperiosteal in location. One of the two implants was covered with non-resorbable e-PTFE membrane. The other implant was left uncovered. Specimens were obtained at 4, 8, and 12 weeks. While the implant specimens on the membrane side showed progressive bone formation between and around HA particles at the subperiosteal extrabony locations, the HA particles on the non-membrane side were surrounded and separated by dense fibrous tissue. At intraosseous sites, HA particles were surrounded by new bone throughout the defect on the membrane side, but new bone formation occurred only along the periphery on the non-membrane side. It appears that guided tissue regeneration may be used to enhance new bone formation around and between subperiosteally implanted HA particles.     

天然羟基磷灰石／壳聚糖复合材料修复种植体周围骨缺损的实验研究

目的：探讨天然羟基磷灰石／壳聚糖复合材料修复种植体周围骨缺损的可行性。方法：预成直径为2．0mm的螺纹纯钛种植体48颗,新西兰白兔12只,每只在双侧胫骨各钻2孔,实验组孔径4．5mm,对照组孔径为2．0mm,实验组同期植入复合材料＋纯钛种植体,对照组只植人种植体,分别于术后4、8、12、16周随机处死三只动物,制作硬组织标本、扫描电镜和能谱分析标本,观察移植材料及种植体一新骨界面变化情况,并分析每一时期两组Ca、P、S元素的含量。结果：实验组复合材料植入后未见明显的异物反应。实验组种植体植入后未发生明显移位,16周时能与周围骨组织形成较好的骨结合。能谱分析显示,16周时两组Ca、P、S元素差异无显著性。结论：天然羟基磷灰石／壳聚糖复合材料可以作为种植体周围骨缺损的修复材料,并能促进种植体一骨界面形成较为完善的骨结合。     

Bone and biologically compatible materials in dentistry.

This review focuses on compatible materials that are either implanted in bone tissue to serve as prosthetic devices or used as bone substitutes. It attempts to evaluate the significance of studies of combined osteoconductive materials with osteoinductive factors, like demineralized bone or its purified components, and osteogenic cells. It is important to note that the term osseointegration, which is still widely used by clinicians to indicate the bioactivity of implants in bone healing, is not substantiated by any controlled basic study. Even pure titanium and hydroxyapatite can interfere with the normal healing of a surgical bone wound. This review emphasizes the three-dimensional structure of bone, intercellular communication, and the response of this system to bone damage and materials.