【作者】 黄美娜；

【导师】 王远亮； 罗彦凤；

【作者基本信息】 重庆大学 ， 生物医学工程， 2010， 博士

【摘要】 骨是自然界中一种复杂的生物材料,当骨组织因创伤、感染等原因需要实施手术剔除病变骨组织后,会造成大块骨缺损,仅仅依靠骨自身的修复能力已经无法愈合,需要通过骨移植手术,将合适的骨材料填充于缺损部位,以便新骨生成。在骨移植修复过程中,骨不连是热点问题之一。从骨不连发生的原因来讲,在植入体与机体之间存在的间隙,或不规则骨产生的不规则界面,导致骨修复过程中成骨细胞迁移能垒的存在,是其中一个重要的因素。本研究将形状记忆聚氨酯脲(SMPUU)用于骨修复过程中,探索形状记忆聚氨酯脲在骨不连预防方面的效果,目的是利用SMPUU的形状记忆性能,即SMPUU在体温作用下可以恢复至预设的形状,实现植入体与机体之间的空隙填充,降低上述成骨细胞迁移能垒的存在,从而降低了骨不连发生的几率。形状记忆聚氨酯/聚脲是能感知外界温度变化并发生形状改变的新型智能材料,因其良好的形状记忆功能而成为功能材料研究的热点之一,在生物医学领域,尤其是微创手术中具有重要的应用价值。本文基于SMPUU预聚体(HO-P(LA-co-PDO)-OH)合成了一系列的可生物降解SMPUU,并对SMPUU进行化学结构的表征,力学性能、形状记忆性能、体外生物降解性、体外细胞相容性、体内安全性的评价,最终采用兔下颌骨建立骨不连模型,对SMPUU在骨修复中的骨不连预防效果进行评价。主要研究内容和结论如下:1.以D,L-丙交酯、对二氧环己酮为原料,Sn(Oct)2为引发剂,乙二醇为助引发剂,制备了SMPUU预聚体(HO-P(LA-co-PDO)-OH),考察了D,L-丙交酯/对二氧环己酮的比例、助引发剂用量、制备方法对SMPUU预聚体的热性能及分子量的影响,并对助引发剂作用下两种单体的反应机理进行深入阐述。①红外与核磁共振结果表明,本研究中一步法与两步法均可制得SMPUU预聚体(HO-P(LA-co-PDO)-OH);其分子量可以通过控制合成方法、两种单体的比例、助引发剂的用量加以调节;②DSC结果表明,SMPUU预聚体的玻璃化转变温度具有单体比例依赖性,与其分子量也具有重要关系;一步法与两步法分别制得无规与嵌段的SMPUU预聚体。2.以SMPUU预聚体为软段,1,6-六亚甲基二异氰酸酯(HDI)和丁二胺(BDA)为硬段,制备一系列的SMPUU,并深入研究了SMPUU合成的影响因素。①FTIR、1HNMR结果表明,SMPUU预聚体与异氰酸酯反应,丁二胺扩链后成功制得SMPUU;②改变软硬段比例、软段结构可以制得不同理化性能的SMPUU;③DSC分析结果表明,SMPUU的玻璃化转变温度、热塑性与其结构有重要关系,通过调节软/硬段比例、软段结构、软段的分子量,可以控制SMPUU的结构,从而调控其玻璃化转变温度,使其可以控制在合适的应用温度,包括体温附近;④SMPUU理化性能测试结果表明,采用亲水性溶剂和疏水性所合成的溶剂进行SMPUU的合成,所得的SMPUU化学结构基本相同,但力学性能、形状记忆功能完全不同;相对于常用的DMF体系,使用本研究中新型的甲苯-异丙醇体系,制得了具有优良性能的SMPUU。3.通过拉伸试验、压缩试验,考察了分子组成、软段分子量、软硬段的比例、变形量、变形温度、回复温度对于形状记忆性能的影响。①SMPUU具有良好的力学特性。硬段含量增加时,模量增加、断裂伸长率下降;②SMPUU呈现良好的形状记忆性能。在合适的操作条件下,固定率、回复率可稳定在95%以上;软段分子量提高,固定率下降;硬段含量提高,固形率上升,回复率下降;③形状记忆性能受变形量、变形温度、回复温度的影响。形变量为50%-200%时,回复率稳定在95%以上,但形变量为400%,回复率明显下降;变形温度高于玻璃化转变温度15℃时,回复率较高,但是当变形温度高于玻璃化转变温度25℃时,回复率显著下降;形状回复时间随着回复温度的增加而缩短;4.考察了SMPUU的亲/疏水性和降解性。在降解性考察过程中,本研究创新性的通过核磁共振氢谱法考察了SMPUU的降解位点。①SMPUU的静态水接触角、吸水率测试结果表明其亲水性优于PDLLA,并且随着硬段含量(酰胺基和脲基含量的增加)呈上升趋势;②体外降解实验结果表明,与PDLLA相比,SMPUU在降解前期,失重率、分子量变化、pH变化大,但是随后降解速率减缓,主要是由于降解后期SMPUU释放的碱性物质能够中和酸性物质,从而降低了酸致自催化作用,说明SMPUU的降解过程受到亲水性与降解产物两方面的影响;③SMPUU降解材料表面形貌表明,降解8周后,SMPUU膜材表面出现孔洞,且随着硬段含量的增加,孔洞的尺寸及数量呈下降趋势;④核磁共振氢谱分析结果显示,SMPUU的降解最先发生在硬段的酰胺基和脲基部位,然后发生在SMPUU的软段部分,或同时发生在SMPUU的硬段和软段部分,但软段的降解速率明显低于硬段。5.本研究从细胞生长行为和功能行为角度,对大鼠成骨细胞与PDLLA及SMPUU的细胞相容性进行了系统比较。并针对SMPUU的形状记忆性能,研究了SMPUU形状记忆变形过程对成骨细胞生长行为的影响。①与PDLLA相比,SMPUU不利于早期黏附与铺展,成骨细胞对SMPUU表面的适应期较长,因此进入分裂增殖期的时间也较晚;②SMPUU更能促进成骨细胞的分化、矿化,且并不缩短细胞的生长分化周期;③通过拉伸变形、固形、复形的方法制作具有形状记忆变形的SMPUU膜材,并通过细胞生长行为对形状记忆变形的响应进行考察,结果表明具有规则微结构的基底材料促进了细胞的黏附与增殖,并使细胞骨架重组,使细胞的生长具有一定的取向性,细胞尺寸均匀,这一结果为SMPUU在组织工程中的应用提供又一有利基础。6.通过全身急性毒性试验和皮肤致敏实验考察了SMPUU的体内安全性,并通过肌肉植入试验研究了SMPUU的体内生物相容性,最后在确定其体内安全性及生物相容性的基础上,通过在兔下颌骨建立的骨不连模型进行骨修复研究,从而确定具有形状记忆性能的SMPUU在骨修复中预防骨不连的效果。更多还原

【Abstract】 Bone is one of the most complicated composite in the nature. Bone, not only provides mechanical support but also elegantly serves as a reservoir for minerals, particularly calcium and phosphate. It is a good example of a dynamic tissue, since it has a unique capability of self-regenerating or self-remodeling to a certain extent throughout the life. However, many circumstances call for bone grafting owing to bone defects either from traumatic or from non-traumatic destruction. In the case of severe defects and loss of volume, bone can’t heal by itself and grafting is required to restore function without damaging living tissues. In the repair, non-union is one of the hot problems. The one most reason of non-union is that the common bone repair materials result in gap between body and graft, or the irregular interfaces in the irregular defect, which will result in the energy barrier, and non-union. In my study, SMPUU is used in the bone repair for its shape memory property.Shape memory polyurethane is a novel intelligent material that is able to response to external temperature change and exhibit shape memory property. It has received wide attentions in biomedical fields, especially in minimally invasive surgery because of its shape memory property.In this paper, several kinds of biodegradable poly (urethane urea) (SMPUU) based on HO-P(LA-co-PDO)-OH is synthesized. The mechanical property, shape memory property, degradation, compatibility, biological safety and medical application in jaw bone of the rabbit were investigated. The main works and conclusions are included as follows:1. HO-P(LA-co-PDO)-OH was synthesized by melt ring-opening polymerization of D, L-lactide and PDO using Sn(Oct)2 as initiator and EG as coinitiator. Then, an extensive investigation effort was expended in understanding the effects of ratio of D, L-lactide to PDO, dosage of coinitiator on the molecular weight, preparation methods on the Mw and thermal properties were characterized. Otherwise, the reaction mechanism of D, L-lactide and PDO was studied furtherly.①FTIR, NMR showed that SMPUU prepolymer was successfully prepared by one-step method and two-step method. Hydroxyl value, GPC-MALLS analysis indicated that SMPUU prepolymer with different molecular could synthesized by varying the dosage of coinitiator, ratio of D, L-lactide to PDO, and preparation method; ②The results of DSC indicated that the glass transition temperature (Tg) molecular weight of SMPUU prepolymer are affected by the ratio of D, L-lactide to PDO; random copolymer and block copolymer were prepared by one-step method and two-step method, respectively.2. SMPUU was synthesized by polymerizing of SMPUU prepolymer, hexamethylene diisocyanate (HDI) and butanediamine (BDA). The effects of solvent kinds, content of catalyst and the reactive temperature, ratio of hard segment/soft segment on the reactive activity of SMPUU prepolymer and HDI were discussed, and the structure and thermal properties of SMPUU were characterized.①FTIR, 1HNMR exhibited that the SMPUU can be successfully synthesized; By varying the molecular weight of soft segment and the ratio of soft segment/hard segment, the SMPUU with different physicochemical properties could be synthesized.②The result of physical property of SMPUU indificated that the chemical structure of SMPUU from the hydrophilic solvent or hydrophobic solvent are similar, however, the shape memory property, mechanical property are different. The SMPUU with suitable properties can be synthesized in the toluene-IPA solvent.③The results of thermal analysis showed that Tg, thermoplasticity were affected by the structure, Mw of SMPUU, and can be regulated by the ratio of hard segment/soft segment, structure of soft segment, Mw of soft segment and SMPUU.3. The mechanical and shape memory properties of SMPUU were characterized by mechanical tensile tests, compression test and bending test. The paper primarily investigated the influences of molecular weight of soft segment, the ratio of soft segment/hard segment, shape deformation and temperature on the shape memory fixation and recovery ratio.①The results indicated that SMPUU have nice mechanical properties. The modulus and maximum stress increased with the increasing of the hard segment content, while the elongation at break had the opposite trend.②SMPUU had good shape memory properties. Shape recovery ratio and fixation ratio were more than 95%. Meanwhile, the shape memory properties were affected by the components and the hard segment content. The fixation ratio of SMPPU showed slightly downward trend with the increasing of the molecular weight of soft segment; the shape recovery ratio decreased with the increasing of the hard segment content, however, the shape fixation ratio increased.③The deformation strain, deformation/recovery temperature had important effects on the shape memory properties. With the deformation strain increasing, the shape recovery ratio decreased. When the deformation range was from 50 to 200%, the shape memory properties are nice, however, when the deformation increased to 400%, the shape recovery ratio decreased to 85%; SMPUU showed good shape memory behavior when the deformation temperature was Tg+15℃, while the shape recovery ratio had apparent decrease, when the deformation temperature was above Tg+25℃; Shape recovery time shortened with the increasing of the recovery temperature.4. The surface wettability and biodegradation of SMPUU were investigated. Moreover, the degradation positions were confirmed by 1H NMR in my study.①The wettability of SMPUU was smaller than PDLLA controls. Meanwhile, the wettability had a uptrend with the increasing of hard segment content.②Comparing with the PDLLA, the weight loss ratio, Mw, and pH value changes of SMPUU decreased fast at prophase, and then, the changes ratio of SMPUU decreased, which can be attributed to the alkaline substance during the degradation of SMPUU could eliminate or weaken the acid induced auto-catalysis.③During the degradation, the SEM showed that the surface of SMPUU appeared the pores after 8 weeks. The amount of pores decreased with the increasing of content of hard segment, which can generate the alkaline group.④The results of 1H NMR indicated that, the degradation of SMPUU started from the hard segment, or in the hard segment and soft segment simultaneously, however, the degradation rate in soft segment was lower than in the hard segment, so the -NH2 in degradation product can release the acidity during degradation.5. In addition, the cytocompatibility of SMPUU was studied by employing primary SD rat osteoblasts as the model cells and poly (DL-lactic acid) (PDLLA) as the control. The growth behavior and function behavior were detected to indicate the cytocompatibility of SMPUU. Furthermore, the effects of stretching-shape recovering process of SMPUU on ostoblasts behaviors were also examined.①Initial morphology, adhesion, spreading and proliferation of osteoblasts on all SMPUU films is no better than those on PDLLA film. Despite this, the cell doubling rate and migration rate on SMPUU elevated with the increasing culture time. Furthermore, with the increased amount of hard segments in SMPUU, both cell doubling rate and migration rate correspondingly increased, which is mostly possibly attributed to the improved hydrophilicity.②Osteoblast on different SMPUU films all exhibited lower physiological functions compared to those on PDLLA films within the first 10 days after seeding. Thereafter, however, the osteoblasts on SMPUU demonstrated better differentiation and mineralization than those on PDLLA films, and SMPUU3 > SMPUU2 > SMPUU1.③The AFM results showed that there were obvious and regular phase separation resulted from soft segments and hard segments in SMPUU, and some groove-ridge architectures within a scale of micrometers were produced by the stretching-shape recovering process. These special micropatterned structures promoted osteoblasts adhesion and proliferation, and also resulted in partially oriented cell growth along the grooves. Stretching-shape recovering process could change the surface morphology of SMPUU films and contribute to enhanced cell adhesion and proliferation, and even produce oriented arrangement of osteoblasts.6. The safety of SMPUU in vivo was evaluated by sensitization test, acute toxicity tests; the biocompatibility in vivo was evaluated by the implantation experiment in muscle; at last, the bone repair function for preventing non-union was confirmed by implanting in the jaw bone of rabbit.更多还原