【作者】 黄师；

【导师】 侯铁胜；

【作者基本信息】 第二军医大学 ， 外科学， 2007， 博士

【摘要】 自1958年Smith和Robinson首创颈前路椎间盘切除植骨融合术以来,自体髂骨一直是颈前路植骨融合术中选材的金标准,然而取自体髂骨存在如下缺点:供区疼痛、出血、感染、撕脱骨折、神经损伤、取材有限、增加手术时间等。由于上述不利因素,学者们一直努力寻找理想的替代物。同种异体骨或异种骨因具有一定的骨传导作用和相对广泛取材来源已在动物实验和临床实践中成功应用,但存在传染疾病(艾滋病、乙型肝炎、丙型肝炎、等)和免疫排斥反应的风险。将前路减压切除的自体骨充填于钛网或钛笼应用于椎间融合虽可避免取髂骨,但切除的骨质通常较少。有研究表明钛网或钛笼移位率为17%,在上位椎体下沉率为42%,下位椎体下沉率为50%,其推广应用目前尚有争议。随着材料工艺学及生物力学的发展,人工合成的骨移植替代材料的研究不断深入。目前常用的人工骨主要有磷酸钙和硫酸钙两大类,其中磷酸钙主要由羟基磷灰石和β-磷酸钙组成,它们具有如下优点:良好的骨传导作用,材料来源广泛,生物相容性良好等;但也有一些不足之处:首先,人工合成材料一般不具有成骨性及骨诱导性,其次,人工材料生物降解时间各不相同,应根据实际需要进行选择。部分材料吸收较快,如:硫酸钙在植入后6-8周已吸收,此时新骨形成尚未完成,机械支撑性能较弱,易致钢板螺钉松动断裂等。而羟基磷灰石虽然支撑性能虽好,但新骨爬行替代缓慢,材料降解缓慢,甚至不降解。β-TCP生物降解时间在8-12周左右,较为适中。β-TCP和CS作为植骨材料应用于非负重区域骨缺损的修复已有较多报道,但应用于颈前路植骨融合术的报道在国内外较少见。本课题拟对β-TCP和CS在颈椎前路椎间融合的效果进行初步的探索,以期获得理想的自体髂骨的替代物。为了更好地模拟人类的颈椎手术,最好采用非人类的灵长类动物作为模型,然而此类动物价格昂贵,来源有限,饲养困难,故广大学者多采用山羊、绵羊、猪等动物作为模型进行研究,但很少有比较人与山羊颈椎的生物力学特征的研究报告。本课题从生物力学角度出发,对人与山羊体外三维生物力学运动特征进行比较,了解山羊能否成为颈椎前路手术的良好模型。因颈椎椎间融合术后植入物需要承受头部重力作用,需要对硫酸钙和β-磷酸钙在体外的生物力学稳定性进行研究,国内外相关报道也较少。本课题研究硫酸钙和β-磷酸钙的材料力学性能,比较山羊与人颈椎体外三维力学运动特征,建立颈前路融合术的动物模型,观察β-磷酸钙和硫酸钙在山羊颈前路植骨融合术的融合效果,主要研究工作由以下三部分组成:一、山羊与人颈椎生物力学特性的比较目的:比较山羊与人颈椎的生物力学特性方法:取新鲜成年人尸体与纯种崇明白山羊颈椎标本(C0-T1)各8具,在脊柱三维运动测试仪上检测两者屈曲、后伸、左右侧屈、左右轴向旋转等模式下的运动范围和中性区。结果:山羊与人颈椎在前屈运动方式下,C1-2节段的ROM分别为16.9±5.1度和14.3±3.2度,超过其它节段;但山羊在各个节段前屈运动的ROM和人相比均无统计学差异(P>0.05)。而且山羊在各个节段前屈运动的NZ和人相比亦无统计学差异(P>0.05)。在后伸运动方式下,山羊与人C1-2节段的ROM分别为20.6±4.8度和18.7±3.7度,C0-1节段的ROM分别为19.3±4.7和18.4±4.3度,明显超过其它节段;但山羊与人在各个节段后伸运动的ROM相比均无统计学差异(P>0.05)。而且人与山羊在各个节段后伸运动的NZ相比亦无统计学差异(P>0.05)。在轴向旋转运动模式下,人C1-2节段的ROM为56.3±8.9度,约占整个颈椎旋转活动范围的70%;山羊C1-2节段的ROM为48.6±8.6度,约占整个颈椎旋转活动范围的63%;人C1-2节段的NZ为29.6±6.5度,约占整个颈椎旋转运动NZ的75%;山羊C1-2节段的NZ为25.9±7.1度,约占整个颈椎旋转运动NZ的68%。然而山羊在各个节段旋转运动的ROM和NZ和人相比均无统计学差异(P>0.05)。在左右侧屈运动模式下,山羊在各个节段侧屈运动的ROM和人相比均无统计学差异(P>0.05),山羊C6-7节段的NZ和人相比有统计学差异(P<0.05),其它节段的NZ和人相比无统计学差异(P>0.05)。结论:纯种崇明白山羊与人的颈椎在前屈、后伸、左右侧屈和轴向旋转等6种生理运动模式下的ROM及NZ相近,两者的生物力学特征基本一致。从生物力学角度出发,纯种崇明白山羊可作为颈椎融合及生物力学研究的良好动物模型。二、β-TCP、硫酸钙与人冻干异体髂骨的生物力学性能比较目的:体外测定硫酸钙、β-TCP与人冻干髂骨最大抗压缩强度及弹性模量方法:取3cm×1.5cm×1.0cm条块状硫酸钙、β-TCP与人冻干髂骨各8块,固定在材料试验机上,做最大抗压缩试验,加载速度1mm/s,描记载荷-变形曲线,计算材料的最大抗压强度及弹性模量。结果: CS、β-TCP与人冻干髂骨最大抗压缩强度分别为3.26±0.86Mpa,3.16±0.75 Mpa,2.97±0.81 Mpa;三者的弹性模量分别为82.35±10.54 Mpa,76.98±11.78 Mpa,35.35±6.98 Mpa。硫酸钙、β-TCP最大轴向抗压强度与冻干异体髂骨无明显差异(P>0.05),两者的弹性模量均大于冻干异体髂骨(P<0.05)结论:硫酸钙、β-TCP具有良好的结构支撑作用。三、β-TCP、硫酸钙在山羊颈前路椎间融合术的融合评价目的:评价硫酸钙、β-TCP应用于山羊颈前路椎间植骨融合术的效果。方法:将24只纯种崇明山羊随机等分为3组,分别以条块状硫酸钙、β-TCP、自体髂骨为植骨材料,制备正常山羊的前路颈3/4椎间盘切除减压植骨钢板内固定术模型,术后1、6、12、24周拍X线片,术后12、24周行三维CT重建检查,术后24周处死动物,拆除钢板后行体外三维生物力学测试,检测融合节段C3-4的ROM和NZ,并进行脱钙与不脱钙组织切片的组织病理学检查及骨组织形态计量学检测。结果:术后6周平片提示:硫酸钙组开始吸收。术后12周平片和CT矢状位重建检查示CS组仅有1只动物达到完全融合,而β-TCP组及自体骨组分别有4只和5只达到完全融合;术后24周,CS组仅有1只动物达到完全融合,而β-TCP组及自体骨组分别有5只和6只达到完全融合。术后24周生物力学测试表明在颈椎前屈/后伸、左侧屈/右侧屈、左/右轴向旋转等生理运动模式下,β-TCP组和自体骨组C3-4节段的ROM分别比CS组的C3-4节段的ROM减少(P<0.05)。术后24周HE切片染色检查示CS组有少量骨组织从相邻的椎体长人,移植骨内部新骨形成少,有不规则的条索状纤维组织和少量软骨组织充填其中。β-TCP组新生骨分布均匀,形成的骨组织绝大多数是小梁状骨,还有少量的残余β-TCP材料在吸收改建之中,在移植块的中心区域有新生的骨髓组织生成。自体骨组显示出良好的骨性融合,骨组织绝大多数是成熟的小梁骨,新生骨髓组织在融合区域大量形成。骨组织计量学定量分析示CS组、β-TCP组和自体骨组新骨形成率分别为25.8±5.1%、45.5±6.7%、49.2±5.8%,β-TCP组和自体骨组相近(P>0.05),与CS相比有明显差异(P<0.05)。结论:β-TCP和自体髂骨在颈前路椎间融合术中具有相近的融合效果,优于CS,β-TCP可作为自体髂骨良好的替代物。更多还原

【Abstract】 Iliac autografts has been the golden standard of bone grafting in anterior cervical fusion since Smith and Robinson first performed anterior cervical discectomy and fusion in 1958. However, harvesting autografts has some disadvantage such as pain and infection of harvested sites, tear fractures, nerve injuries, limited supply, increased surgical time, and so on. Because of above mentioned disadvantage many scholars have always strived to seek ideal bone substitutes.Allograft or xenograft bone has been successfully applied in experimental studies and clinical trials because of its osteoconductivity and relative widely resources. But it is still a main concern that the risk of immunological rejection and potential transmitted virus such as type B or C hepatitis virus, human immunodeficiency virus.Although it can avoid harvesting iliac bone grafts by the means of using the local bone obtained from decompression combined with the titanium mesh or cage, sometimes there is less local bone available for use. There are some studies reported that the dislocation rate of titanium mesh or cage was about 17 percents and the subsidence rate in upper and caudal vertebrae were 42 percents and 50 percents respectively. So this method has limited use.With the development of materials technology and biomechanics, the research related to synthetic bone substitutes has been more extensive. Now the most used bone substitutes are calcium sulphate (CS) and calcium phosphate. The latter is mainly composed of hydroxyapatite (HA) andβ-tricalcium phosphate (β-TCP). All These synthetic materials enjoy several advantages over other bone graft materials in that they are osteoconductive, nontoxic, biocompatible, easy to sterilize and available in a virtually unlimited supply. However, they have some disadvantages. Firstly, they are not osteoinductive and osteogenic. Secondly, they have different duration of degradation. The resorption of some materials are rapid, such as calcium sulphate which was absorbed four to six weeks after implantation. At that time the new bone formation is incomplete and the fusion mass is weak, this may lead to breakage of plate system. HA is a relatively inert substance that is retained in vivo for prolonged periods of time, and it can provide structural stability.β-TCP typically undergoes biodegradation within 8-12 weeks of its introduction into the area of bone formation.There are many reports about CS andβ-TCP as bone expander for repairing non weight-bearing bone defects, whereas rare are applied in anterior cervical fusion. In order to obtain ideal substitutes for anterior cervical fusion, we planed to investigate the effect of CS andβ-TCP applied in anterior cervical fusion.To mimic human cervical spine surgery more accurately, it has best to use non human primate as models. However these animals are expensive, limited supplying and difficult to handle, many scholars most use goat, sheep and swine as animal models. By far, the comparison of three-dimensional biomechanics between goat and human are rare reported. It still remains concern about whether goat is a optimal model for anterior cervical spine surgery. To solve this problem, it needs to compare the three-dimensional biomechanics of goat and human.In addition, the bone grafts used in anterior cervical interbody fusion require supporting gravity of head, so it should be to investigate biomechanical stability of CS andβ-TCP in vitro. And rare reports have addressed this issue up to now.In general, we planed to investigate biomechanical stability of CS andβ-TCP, compare the three-dimensional biomechanics between goat and human cervical spine, establish optimal model for anterior cervical fusion and observe the fusion effects of CS andβ-TCP as bone substitutes. Our study made up of three parts as follow.Part one: Comparison of Three-dimensional Biomechanical Characteristics Between Goat and Human Cervical SpineObjective. To compare three-dimensional biomechanical characteristics between goat and human cervical spine.Methods. Eight fresh cervical spines were harvested respectively from adult human cadaver and pure breed ChongMing white goat. And the range of motions(ROM) and neutral zone(NZ) of the specimens were tested respectively in three-dimentional testing apparatus under flexion, extension, right or left bending and axial rotational loads.Results. Under flexion loads, ROM of C1-2 of goat and human are 16.9±5.1 and 14.3±3.2 degrees respectively, lager than other levels. But the ROM and NZ of each level of cervical spine of goat has no statistical significance respectively when comparing with human (P>0.05). Under extension loads, ROM of C1-2 of goat and human are 20.6±4.8 and 18.7±3.7 degrees respectively, and ROM of C0-1 of goat and human are 19.3±4.7 and 18.4±4.3 degrees respectively, more superior than other levels. But the ROM and NZ of each level of cervical spine of goat has no statistical significance respectively when comparing with human (P>0.05). Under axial torques, ROM of C1-2 of human is 56.3±8.9 degrees, made up of 70 percents of ROM of the whole cervical spine. And ROM of C1-2 of goat is 56.3±8.9 degrees, made up of 68 percents of ROM of the whole cervical spine. NZ of C1-2 of goat and human are 25.9±7.1 and 29.6±6.5 degrees respectively, made up of most of whole cervical spine. However, the ROM and NZ of each level of cervical spine of goat has no statistical significance respectively when comparing with human (P>0.05). Under right or left bending loads, the ROM of each level of cervical spine of goat has no statistical significance respectively when comparing with human (P>0.05). Except C6-7 level, the NZ of other levels of cervical spine of goat have no statistical significance when comparing with human (P>0.05).Conclusions. Pure breed ChongMing white goat had similar biomechanical characteristics with human under flexion, extension, right or left bending and axial rotational loads. It served as an optimal model for anterior cervical fusion and biomechanical research.Part two: Biomechanics of CS andβ-TCP in Vitro Compare with Human Freeze-drying Iliac AllograftsObjective. To investigate the axial compression strength and elastic modulus of CS,β-TCP and human freeze-drying iliac allografts.Methods. Respective eight pieces of CS,β-TCP and human freeze-drying iliac allografts in same size (3cm×1.5cm×1.0cm) were obtained for mechanical testing. Each specimen was mounted in testing apparatus, loaded axial compression with a velocity of 1mm per second and the load–displacement curves were plotted. Thus the axial compression strength and elastic modulus were calculated.Results. The axial compression strength of CS,β-TCP and human freeze-drying iliac allografts were 3.26±0.86Mpa, 3.16±0.75 Mpa, 2.97±0.81 Mpa respectively, and elastic modulus were 82.35±10.54 Mpa, 76.98±11.78 Mpa, 35.35±6.98 Mpa. The axial compression strength of CS andβ-TCP have no statistical significance when compared with human freeze-drying iliac allografts. However, the elastic modulus of both were lager than human freeze-drying iliac allografts (p<0.05).Conclusions. CS andβ-TCP had optimal structural supporting stability. Part three: Evaluation of Efficacy ofβ-TCP and CS as Bone Substitutes for Goat Anterior Cervical Fusion.Objective. To evaluate efficacy ofβ-TCP and CS as bone substitutes for goat anterior cervical fusion.Methods. twenty four pure breed adult ChongMing white goats were divided into three groups(group CS, groupβ-TCP and group auto) equally and randomly. And the animals were performed with anterior C3-4 discectomy and fusion with different grafts according to preoperative plan. all animals were tested by means of X-ray in 1th, 6th , 12th and 24th week after operation. And the three-dimensinal CT reconstruction were performed in 12th and 24th week. After all animal were euthanized in 24th week, Cervical plate systems were removed and biomechanical testing was performed in non destructively. And then histomorphological and histomorphometrical analysis of decalcified and non decalcified slices were carried out.Results. Radiograph of six weeks postoperative revealed CS began to absorbed. Radiograph and CT reconstruction in 12th week postoperative suggested that there was only one animal obtaining complete fusion in group CS whereas 4 and 5 respective in groupβ-TCP and Auto. In 24th week the number of complete fusion still remained one in group CS, but 5 and 6 respective in groupβ-TCP and Auto. The biomechanical testing revealed that ROM of C3-4 of groupβ-TCP and Auto decreased compared with group CS. It was observed in HE stained slices that less new bone formation, Cartilaginous tissue and massive fibrous tissue in group CS while massive mature trabecular bone and mallow in groupβ-TCP and Auto. There was a small quantity of ceramic Remnants in groupβ-TCP. The new bone formation rate was 25.8±5.1%, 45.5±6.7%, 49.2±5.8% respectively in group CS,β-TCP and Auto. There was no statistical significance between groupβ-TCP and Auto (P>0.05), while statistical significance was seen in group CS when comparing with groupβ-TCP or Auto (P<0.05).Conclusions.β-TCP had similar efficacy with autografts in anterior cervical fusion and was superior to CS.β-TCP is an optimal substitute for iliac autografts.更多还原