纳米磷酸钙和生物玻璃对牙釉质的仿生修复
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摘要
近年来,生物矿化的研究表明,纳米磷酸钙材料在生物体的硬组织形成过程中具有非常重要的作用。作为脊椎动物中最重要的两种矿化组织：骨骼和牙釉质,虽然它们的功能、性质、形成过程和分级结构都有所不同,但是它们的基本无机组成单元都是纳米级别的磷灰石。在生物矿物的形成过程中,纳米尺寸的无机基本单元在生物体的调控下,在有机基质形成的框架中经过有序组装,形成具有特殊功能的生物矿物。这些无机组成单元和有机大分子之间的相互作用是生物矿化研究中一个核心的问题。本论文研究了纳米磷酸钙在牙釉质修复中的作用。研究涉及了纳米磷酸钙颗粒对牙釉质的直接修复作用：利用生物矿化的原理,发展出一种简单有效地直接使牙釉质再生的方法；最后尝试利用应用广泛的生物材料一生物玻璃来实现牙釉质结构的再生。全文共分章：
第一章简要介绍了生物矿化的基本知识,包括磷酸钙体系的生物体硬组织的结构和形成过程以及仿生矿化的概念。总结了牙釉质脱矿形成龋齿的过程和传统的牙修复材料,紧接着回顾了近年来关于牙釉质修复和重构的方法等。通过总结生物矿化的基本原理和牙釉质修复中存在的问题引出我们对这些问题的观点及相关的研究计划。
第二章研究了具有与牙釉质基本组成单元相类似的纳米羟基磷灰石(HAP)对牙釉质的修复情况。相较于普通的磷灰石和无定形磷酸钙,20nm的HAP纳米颗粒可以牢固的吸附在脱矿的牙釉质表面,并且这些颗粒会沿着釉柱的方向排列。先前的研究已经发现牙釉质的基本组成单元是尺寸在20-40nm左右的纳米羟基磷灰石颗粒,并且这些颗粒在欠饱和或酸性溶液中能保持稳定存在,根据激光共聚焦显微镜的结果表明,20nm的HAP还可以保护牙釉质表面,阻止其在酸性溶液中的继续溶解。由此提出20nm HAP颗粒存修复牙釉质和预防龋齿的发生上有很好的效果。该工作说明了磷酸钙材料具有与生物体矿化组织基本组成单元相似的尺寸在其应用上的重要性。
第三章的工作以第二章的结果为基础,利用生物矿化的基本原理,发展了一种简单有效的牙釉质结构重构的方法。我们之前的研究表明,纳米磷酸钙颗粒可以在氨基酸的调控下,自组装形成类牙或者类骨的结构。在这部分研究中,我们将吸附了20nm磷酸钙颗粒的牙釉质放入含有100mM谷氨酸的生物模拟体液中培养72个小时。实验结果表明,在纳米磷灰石颗粒和氨基酸的共同作用下,牙釉质表面可以形成一层具有良好机械性质的修复层,而且修复层具有与牙釉质类似的组装结构。在过去的工作中,人们尝试利用各种生物体蛋白质、表面活性剂或者氟离子对牙釉质进行再矿化,但是由这些添加剂参与的方法并不能提供着-种简单方便、安全、经济的修复方法。我们介绍到了一种新的仿生矿化方法,所需的材料具有非常高的生物亲和性,而且修复效果也非常明显。这一研究强调了在生物体硬组织形成过程中纳米材料和有机物调控共同作用的重要性。
第四章中,我们发展了第三章中的修复方法,利用活性生物玻璃取代之前所用的纳米磷酸钙颗粒,进一步简化了修复方法,增加了其真正的应用前景和可能。由于我们之前所使用的纳米磷酸钙颗粒的制备具有一定的难度,限制了这种方法在实际应用中的发展。而活性生物玻璃较易制备,而且具有良好的生物相容性和矿化能力,所以,我们利用生物玻璃代替纳米磷酸钙颗粒,以模拟口腔唾液为培养液,在谷氨酸的调控作用下,在牙釉质表面生长出了具有类牙结构的修复层,并且修复以后的牙釉质具有与天然牙釉质相媲美的机械性质。此外,我们还发现,谷氨酸还可以通过加快无定形磷酸钙向羟基磷灰石相转化的结晶速率实现快速修复过程,这个特性是在口腔产品中广泛使用的氟离子不具备的。在这部分内容中,我们提出了一种具有应用前景的牙釉质修复和再生的方法,重点强调了仿生方法在硬组织修复和再生中的应用。
第五章,总结了关于利用纳米磷酸钙和生物玻璃仿生修复牙釉质的创新点,还为今后的研究指出需要解决的关键问题。
The development of biomineralization and biomaterials has demonstrated that nano-sized calcium phosphate particles play an essential role in the formation of hard tissues in biomatiarials. There is a general agreement that the basic building blocks in both bone and enamel are nano-sized apatite particles, although their hierarchical structures and functions are different. These nano building blocks can self-assemble into calcified tissue under the control of an organic matrix during biomineralization process. In our previous work, we demonstrated the remarkable role of nano apatite in enamel repair. The research includes:enamel repair by using nano apatite particles with the similar size as natural enamel blocks; a simple and effective biomimetic way to regenerate enamel structure with nano apatite particles; constructing enamel-like apatite layer on enamel surface by using bioglass under near-physiological conditions. The thesis is composed of five chapters:
In chapter1, we introduce the conception of biomineralization, the formation process and structure of the hard tissues formed by of calcium phosphate, the concept of biomimetic mineralization, the development of enamel demineralization and caries. Then we review the common restoration materials and the recent development of enamel regeneration or repair. The basic problems in enamel repair are summarized based on these above knowledge and the research concerning these problems is proposed at the end of this chapter briefly.
In chapter2, we study the role of nano apatite (HAP) particles which have the similar size and morphology with the natural enamel building blocks in enamel repair. Different from the conventional HAP and amorphous calcium phosphate (ACP), the nano HAP particles can tightly absorb on enamel surface and tend to array along the direction of natural enamel rod structure. Our previous studies have already revealed that the natural enamel building blocks are20-40nm calcium phosphate particles. And these building blocks can resist dissolution in acidic conditions. The CLSM (confocal laser scanning microscopy) results indicate that the demineralization of enamel surface was inhibited with the prevention of these nano particles. Thus the caries or second cries can be prevented significantly by the restoration. The results suggest the importance of size effect of calcium phosphate materials for their application in biomedical engineering.
In chapter3, we develop an effective and simple approach to enamel regeneration. In our previous work, we have revealed that the nano calcium phosphate particles can self-assemble into enamel-like and bone-like structure with the control of amino acids. In this study, we immerse the enamel surface pre-absorbed20nm calcium phosphate particles into SBF solutions (simulated body fluid) containing100mM glutamic acid for72h. The results show that a repaired layer with enamel-like structure is formed on enamel surface in the combined effects of nano particles and glutamic acid. Besides, the regenerated layer has comparable mechanical prooerties with natual enamals. In previous works, various addtives are used in enamel remineralization including proteins, polymers, surfactants and fluoride ions. However, they hardly provide a simple, safe and economic strategy to enamel regeneration. Herein we report that a bio-inspired cooperative effect of glutamic acid and nano HAP can result in the effective regeneration of enamel-like structure under physiological conditions. Importantly, the mechanical characteristics of the repaired enamel are similar as the natual enamal. The study emphasizes the importance of co-reactions between the nano basic building blocks and organic molecules in the calcifications process and the study of biomedical biomaterials.
Based on these above achievements, a more simple and practical method to regenerate enamel is shown in chapter4. Althoughthe method by using nano HAP and glutamine together has remarkable effect in enamel repair, the complex synthesis and purification of20nm HAP particles limite on its further application. Thus, a althernative substance which can replace the role of HAP nanoparticles are in great demand. Bio-glass can be appropriate candidate due to its excellent bio-compatibility and mineralization ability. Herein, the similar HAP layer with enamellike structure is well regenerated on enamel surface with the presence of bioglass and glutamic in SOF (simulated oral fluid). Again, the repaired layer formed on enamel surface shares the similar mechanical properties with that of natural enamel. Besides, glutamic acid can also accelerate the crystallization process from amorphous calcium phosphate to HAP. The promotion effect of Glu in enamel repair could not be achieved with fluoride which is now wildly used in dental products. Our current achievment provides a practical application to repair enamel clinically and highlight the biomimetic strategy in the regeneration of hard tissues.
In chapter5, we emphasize the biomimetic strategy for hard tissue repair or regeneration. Our attempts about the enamel reconstruction may contribute to the development of biomaterials for the hard tissues regeneration. Some important but unsolved issues are also suggested in this chapter.
第一章简要介绍了生物矿化的基本知识,包括磷酸钙体系的生物体硬组织的结构和形成过程以及仿生矿化的概念。总结了牙釉质脱矿形成龋齿的过程和传统的牙修复材料,紧接着回顾了近年来关于牙釉质修复和重构的方法等。通过总结生物矿化的基本原理和牙釉质修复中存在的问题引出我们对这些问题的观点及相关的研究计划。
第二章研究了具有与牙釉质基本组成单元相类似的纳米羟基磷灰石(HAP)对牙釉质的修复情况。相较于普通的磷灰石和无定形磷酸钙,20nm的HAP纳米颗粒可以牢固的吸附在脱矿的牙釉质表面,并且这些颗粒会沿着釉柱的方向排列。先前的研究已经发现牙釉质的基本组成单元是尺寸在20-40nm左右的纳米羟基磷灰石颗粒,并且这些颗粒在欠饱和或酸性溶液中能保持稳定存在,根据激光共聚焦显微镜的结果表明,20nm的HAP还可以保护牙釉质表面,阻止其在酸性溶液中的继续溶解。由此提出20nm HAP颗粒存修复牙釉质和预防龋齿的发生上有很好的效果。该工作说明了磷酸钙材料具有与生物体矿化组织基本组成单元相似的尺寸在其应用上的重要性。
第三章的工作以第二章的结果为基础,利用生物矿化的基本原理,发展了一种简单有效的牙釉质结构重构的方法。我们之前的研究表明,纳米磷酸钙颗粒可以在氨基酸的调控下,自组装形成类牙或者类骨的结构。在这部分研究中,我们将吸附了20nm磷酸钙颗粒的牙釉质放入含有100mM谷氨酸的生物模拟体液中培养72个小时。实验结果表明,在纳米磷灰石颗粒和氨基酸的共同作用下,牙釉质表面可以形成一层具有良好机械性质的修复层,而且修复层具有与牙釉质类似的组装结构。在过去的工作中,人们尝试利用各种生物体蛋白质、表面活性剂或者氟离子对牙釉质进行再矿化,但是由这些添加剂参与的方法并不能提供着-种简单方便、安全、经济的修复方法。我们介绍到了一种新的仿生矿化方法,所需的材料具有非常高的生物亲和性,而且修复效果也非常明显。这一研究强调了在生物体硬组织形成过程中纳米材料和有机物调控共同作用的重要性。
第四章中,我们发展了第三章中的修复方法,利用活性生物玻璃取代之前所用的纳米磷酸钙颗粒,进一步简化了修复方法,增加了其真正的应用前景和可能。由于我们之前所使用的纳米磷酸钙颗粒的制备具有一定的难度,限制了这种方法在实际应用中的发展。而活性生物玻璃较易制备,而且具有良好的生物相容性和矿化能力,所以,我们利用生物玻璃代替纳米磷酸钙颗粒,以模拟口腔唾液为培养液,在谷氨酸的调控作用下,在牙釉质表面生长出了具有类牙结构的修复层,并且修复以后的牙釉质具有与天然牙釉质相媲美的机械性质。此外,我们还发现,谷氨酸还可以通过加快无定形磷酸钙向羟基磷灰石相转化的结晶速率实现快速修复过程,这个特性是在口腔产品中广泛使用的氟离子不具备的。在这部分内容中,我们提出了一种具有应用前景的牙釉质修复和再生的方法,重点强调了仿生方法在硬组织修复和再生中的应用。
第五章,总结了关于利用纳米磷酸钙和生物玻璃仿生修复牙釉质的创新点,还为今后的研究指出需要解决的关键问题。
The development of biomineralization and biomaterials has demonstrated that nano-sized calcium phosphate particles play an essential role in the formation of hard tissues in biomatiarials. There is a general agreement that the basic building blocks in both bone and enamel are nano-sized apatite particles, although their hierarchical structures and functions are different. These nano building blocks can self-assemble into calcified tissue under the control of an organic matrix during biomineralization process. In our previous work, we demonstrated the remarkable role of nano apatite in enamel repair. The research includes:enamel repair by using nano apatite particles with the similar size as natural enamel blocks; a simple and effective biomimetic way to regenerate enamel structure with nano apatite particles; constructing enamel-like apatite layer on enamel surface by using bioglass under near-physiological conditions. The thesis is composed of five chapters:
In chapter1, we introduce the conception of biomineralization, the formation process and structure of the hard tissues formed by of calcium phosphate, the concept of biomimetic mineralization, the development of enamel demineralization and caries. Then we review the common restoration materials and the recent development of enamel regeneration or repair. The basic problems in enamel repair are summarized based on these above knowledge and the research concerning these problems is proposed at the end of this chapter briefly.
In chapter2, we study the role of nano apatite (HAP) particles which have the similar size and morphology with the natural enamel building blocks in enamel repair. Different from the conventional HAP and amorphous calcium phosphate (ACP), the nano HAP particles can tightly absorb on enamel surface and tend to array along the direction of natural enamel rod structure. Our previous studies have already revealed that the natural enamel building blocks are20-40nm calcium phosphate particles. And these building blocks can resist dissolution in acidic conditions. The CLSM (confocal laser scanning microscopy) results indicate that the demineralization of enamel surface was inhibited with the prevention of these nano particles. Thus the caries or second cries can be prevented significantly by the restoration. The results suggest the importance of size effect of calcium phosphate materials for their application in biomedical engineering.
In chapter3, we develop an effective and simple approach to enamel regeneration. In our previous work, we have revealed that the nano calcium phosphate particles can self-assemble into enamel-like and bone-like structure with the control of amino acids. In this study, we immerse the enamel surface pre-absorbed20nm calcium phosphate particles into SBF solutions (simulated body fluid) containing100mM glutamic acid for72h. The results show that a repaired layer with enamel-like structure is formed on enamel surface in the combined effects of nano particles and glutamic acid. Besides, the regenerated layer has comparable mechanical prooerties with natual enamals. In previous works, various addtives are used in enamel remineralization including proteins, polymers, surfactants and fluoride ions. However, they hardly provide a simple, safe and economic strategy to enamel regeneration. Herein we report that a bio-inspired cooperative effect of glutamic acid and nano HAP can result in the effective regeneration of enamel-like structure under physiological conditions. Importantly, the mechanical characteristics of the repaired enamel are similar as the natual enamal. The study emphasizes the importance of co-reactions between the nano basic building blocks and organic molecules in the calcifications process and the study of biomedical biomaterials.
Based on these above achievements, a more simple and practical method to regenerate enamel is shown in chapter4. Althoughthe method by using nano HAP and glutamine together has remarkable effect in enamel repair, the complex synthesis and purification of20nm HAP particles limite on its further application. Thus, a althernative substance which can replace the role of HAP nanoparticles are in great demand. Bio-glass can be appropriate candidate due to its excellent bio-compatibility and mineralization ability. Herein, the similar HAP layer with enamellike structure is well regenerated on enamel surface with the presence of bioglass and glutamic in SOF (simulated oral fluid). Again, the repaired layer formed on enamel surface shares the similar mechanical properties with that of natural enamel. Besides, glutamic acid can also accelerate the crystallization process from amorphous calcium phosphate to HAP. The promotion effect of Glu in enamel repair could not be achieved with fluoride which is now wildly used in dental products. Our current achievment provides a practical application to repair enamel clinically and highlight the biomimetic strategy in the regeneration of hard tissues.
In chapter5, we emphasize the biomimetic strategy for hard tissue repair or regeneration. Our attempts about the enamel reconstruction may contribute to the development of biomaterials for the hard tissues regeneration. Some important but unsolved issues are also suggested in this chapter.
引文
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