复合纳米银正畸粘结剂抗菌性和粘结强度的研究

发布时间：2018-05-04 08:07

本文选题：纳米银正畸粘结剂 + 抗菌性　；参考：《武汉大学》2014年博士论文

【摘要】：牙釉质龋白斑是固定正畸治疗中经常伴随的并发症。托槽和带环等固定装置的粘结为致龋菌的黏附和定植创造了良好的条件,从而增加了龋白斑的发病几率。临床研究表明,牙釉质脱矿易发部位恰好是临近托槽粘结剂的位置。为了减少正畸治疗过程中牙釉质脱矿的发生,曾有许多抗菌成分添加到粘结剂中,如氟和洗必泰,但是临床应用的效果表明,仍有很多正畸患者出现不同程度的龋白斑。近年来,虽然正畸粘结技术和材料取得了长足的进展,但是正畸过程中伴随牙釉质脱矿的问题仍未解决。银离子对口腔链球菌有很强的抗菌活性,含银离子的粘结剂对细菌生物膜的形成和口腔变形链球菌的生长都有很强的抑制作用。本研究将纳米银添加到改性树脂-玻璃离子粘结剂(GC Fuji ORTHO LC;GC Corporation, Tokyo, Japan)中,获得复合纳米银正畸粘结剂(nano silver-containing cement, NSC)样品,通过体外和动物龋病模型在抗菌能力、粘结强度以及预防釉质脱矿等几方面与目前临床常用的粘结剂进行比较,希望其在满足临床粘结要求的前提下能在抗菌能力方面获得更佳的表现,为进一步的应用研究奠定基础。第一部分体外实验研究实验一复合纳米银正畸粘结剂样本的制备本研究通过机械研磨和无水乙醇悬浮混合两种物理混合的方法将纳米银无机抗菌粉体添加到改性树脂-玻璃离子粘结剂中,获得复合纳米银正畸粘结剂样品,通过透射电镜对纳米银在粘结剂粉体中的分散情况进行了观察。结果表明,机械研磨方法粉体颗粒要更小一些,纳米银颗粒在粉体中分散均匀,粒径大约10-20nm；在无水乙醇悬浮混合法获得的样品中,纳米银颗粒出现团聚现象。实验二琼脂扩散实验本实验通过琼脂扩散法,来确定样本材料中的抗菌成分能否溶解并渗透到周围环境中和材料内部的抗菌成分能否游离到材料表面发挥持续的抗菌活性。结果表明,新鲜材料组在孵育48小时后,在样本的周围可见直径约13mm的抑菌圈,在材料老化2天、1周和2周后的试验中所有粘结剂样品的周围均未见抑菌圈。这说明氟离子的释放量很快就降低了,纳米银离子很可能不具有从粘结剂基质向周围环境扩散的属性。实验三直接接触和老化实验本实验通过待测试材料与变形链球菌在96孔板的直接接触,然后连续测定后续BHI液体培养基的吸光度值来评价新鲜和经过老化处理的样本材料的抗菌能力。实验的结果表明,所有新鲜复合纳米银材料均呈现强抗菌性,但是随着老化时间的延长,其抗菌成分呈逐渐丧失的趋势,最后,所有测试的材料在老化8周后完全丧失抗菌活性。这种方法在某种程度上还是反映出这种复合了纳米银抗菌粉体的粘结剂比对照组粘结剂具有更强、更长时间的抗菌能力,但是作为一种体外评估方法,还不能反映口内的真实情况,另外由于观察的时间比较短,故而无法对其长期抗菌能力做出相对真实的评价。实验四改性树脂-玻璃离子粘结剂和复合纳米银粘结剂粘结强度的体外研究本实验采用因正畸拔除的上颌第一前磨牙进行体外研究,一方面探索树脂修饰化的玻璃离子粘结剂临床应用的最佳条件,另一方面将制备的复合了不同浓度纳米银的正畸粘结剂样品与改性树脂-玻璃离子粘结剂的粘结强度进行比较,来评估这些样品的粘结强度是否能够满足临床的需要。研究结果表明,潮湿的牙面不利于改性树脂-玻璃离子粘结剂材料的粘结,但是牙面经过酸蚀处理后其粘结强度会获得明显增强；所有样本材料的粘结强度均达到了理想粘结强度6-8Mpa的范围,但是随着纳米银无机抗菌粉添加比例的增加,其粘结强度呈现逐渐降低的趋势。第二部分动物实验实验一龋病模型的建立本实验是利用大鼠龋病模型来模拟人的口腔环境,希望利用这样的环境来间接反映样本材料在口内的抗菌情况。通过将变形链球菌植入经过抗生素处理的大鼠口腔中,喂饲致龋饲料Keyes2000,我们成功构建了大鼠龋病模型,这将是对新研究方法很有意义的一次尝试。实验二材料样本的植入手术本实验用粘结材料将托槽底板粘结在上颌第一双尖牙颊侧的半边牙冠表面,制备成材料样本牙,再将其结扎固定在大鼠口腔下前牙舌侧的根部,以保证样本材料能够长期保存在大鼠口内。通过长期的观察发现,材料样本牙放置的位置没有对大鼠正常进食造成影响,材料几乎没有发生脱落现象。实验三动物口内粘结材料表面细菌生长情况的扫描电镜观察局麻下分别在植入手术后的1个月和3个月两个时间段取出样本牙,每次25只,取出的样本干燥后喷金,对粘结材料表面进行扫描电镜观察。结果显示,Transbond XT表面变形链球菌生长活跃,层层堆积；植入1个月后的GC Fuji ORTHO LC表面变链菌零星分布或少量成簇,3个月后数量明显增多并呈链状；在NSC2,NSC3和NSC4组,随着纳米银含量的增加,变链菌的数量呈现减少的趋势,而且形态趋于分散单个存在,说明纳米银的加入在某种程度上增强了GC Fuji ORTHO LC对变链菌的黏附抑制和杀灭作用。实验四动物口内样本材料周围釉质脱矿情况的micro-CT观察在样本植入3个月后,局麻下取出磨牙已经发生明显龋坏的15只大鼠口内的样本牙,清洗后将牙表面的粘结剂和托槽底板取下,以10微米的分辨率进行micro-CT扫描,三维成像后检查原粘结剂周围区域的釉质表面是否有脱矿后所留下的陷窝。结果显示,在3M Transbond XT材料组牙冠侧面可见釉质脱矿后所留下的大量陷窝,与颊面正上方有明显的分界；在GC Fuji ORTHO LC组的一个样本的侧面发现釉质脱矿后所留下的陷窝；在NSC2、NSC3和NSC4各组的所有样本牙冠的颊面颈部和两个侧面都没有发现明显的吸收陷窝。这说明纳米银能够在观察期有效抑制细菌在材料周围的黏附和菌斑的形成,从而防止或降低了釉质脱矿发生的几率。
[Abstract]:Dental enamel leukoplakia is a frequent complication associated with fixed orthodontic treatment. The bonding of brackets and rings and other fixers creates good conditions for the adhesion and colonization of cariogenic bacteria, thus increasing the incidence of leukoplakia. The clinical study shows that the location of the enamel demineralization easily is the location of the adjacent bracket binder. In orthodontic treatment of enamel demineralization, many antibacterial components have been added to the adhesive, such as fluorine and chlorhexidine, but the clinical effect shows that there are still many orthodontic patients with different degrees of dental caries. In recent years, although the orthodontic bonding technology and materials have made great progress, but the orthodontic process is accompanied by teeth. The problem of enamel demineralization remains unsolved. Silver ions have strong antibacterial activity to oral Streptococcus. The silver ion binder has a strong inhibitory effect on the formation of bacterial biofilm and the growth of Streptococcus mutans. This study added silver nanoparticles to the modified resin glass ion binder (GC Fuji ORTHO LC; GC Corporation, Tok). In yo, Japan), the composite nano silver orthodontic adhesive (nano silver-containing cement, NSC) samples were obtained. Through in vitro and animal caries models, the antibacterial ability, bond strength and the prevention of enamel demineralization were compared with the commonly used adhesives at present. It is hoped that it can meet the requirements of clinical adhesion to the antibacterial energy. Better performance is achieved in terms of force, laying the foundation for further application research.
Part one in vitro experimental study
Experiment 1 Preparation of composite nano silver orthodontic adhesive sample
In this study, the nano silver inorganic antibacterial powder was added to the modified resin glass ion binder by two physical mixing methods, mechanical lapping and anhydrous ethanol suspension. The composite nano silver orthodontic adhesive samples were obtained. The dispersion of nano silver in the binder powder was observed by transmission electron microscope. The results showed that the silver nanoparticles were dispersed in the binder powder by transmission electron microscope. The powder particles are smaller, and the nano silver particles are dispersed evenly in the powder, and the particle size is about 10-20nm. In the samples obtained by the suspension of anhydrous ethanol, the agglomeration of nano silver particles. Experiment two agar diffusion test.
In this experiment, the agar diffusion method is used to determine whether the antibacterial components in the sample materials can be dissolved and penetrated into the surrounding environment and the antibacterial components in the materials can be free to the surface of the material to play a sustained antibacterial activity. The results show that the Bacteriostasis of about 13mm in diameter around the sample is seen around the sample after 48 hours of incubation. There was no bacteriostasis around all the binder samples after 2 days of aging, 1 and 2 weeks, which indicated that the release of fluorine ions decreased rapidly, and the nano silver ions were likely to not have the properties of diffusion from the matrix to the surrounding environment.
Experiment three direct contact and aging test
By direct contact with Streptococcus mutans in 96 orifice plates, the experiment was used to determine the absorbance value of the following BHI liquid medium continuously to evaluate the antibacterial ability of fresh and aging samples. The results showed that all fresh composite nano silver material had strong antibacterial properties, but with aging. In the end, all the tested materials lost their antibacterial activity completely after 8 weeks of aging. This method, to some extent, reflects that the binding agent of this composite nano silver antibacterial powder has a stronger, longer time antibacterial ability than the control group, but as a body. The external evaluation method can not reflect the real situation in the mouth, and the time of observation is short, so it can not make a relatively true evaluation of its long-term antibacterial ability.
Experiment four study on bond strength of modified resin glass ionomer binder and composite nano silver binder in vitro
In this experiment, the first premolar tooth extracted by orthodontic extraction was studied in vitro. On the one hand, the best conditions for the clinical application of the resin modified glass ion binder were explored. On the other hand, the bond strength of the orthodontic adhesive with different concentrations of silver nanoparticles was compared with that of the modified resin glass ion binder. To assess whether the bond strength of these samples can meet the needs of the clinic, the results show that the wet tooth surface is not conducive to the bonding of the modified resin glass ion binder material, but the bonding strength of the tooth surface is obviously enhanced after etching, and the bond strength of all the samples is up to 6 of the ideal bond strength. The range of -8Mpa, however, with the increase of the proportion of nano silver inorganic antibacterial powder, the bond strength of them decreased gradually.
Second parts of animal experiments
The establishment of an experimental caries model
This experiment uses the rat caries model to simulate human oral environment. We hope to use this environment to indirectly reflect the antibacterial situation of the sample material in the mouth. By feeding the Streptococcus mutans into the oral cavity of the rats treated with antibiotics and feeding the caries feed Keyes2000, we have successfully constructed the caries model of the rat. This will be the new research. A meaningful experiment. Experiment two. Implantation of material samples.
In this experiment, the plate was bonded to the surface of the half edge of the maxillary first double canine on the surface of the crown of the maxillary first double canine. The sample was prepared and fixed to the root of the tongue side of the anterior teeth of the rat's mouth to ensure that the sample material could be kept in the mouth of the rat for a long time. There was almost no loss of material in rats.
Experiment three observation of the growth of bacteria on the surface of adhesive material in animals by scanning electron microscope
The sample teeth were removed at 1 months and two periods of 3 months after the implantation, 25 of the samples were removed each time, and the samples were dried after drying. The surface of the bonded materials was observed by scanning electron microscope. The results showed that the Transbond XT surface streptococcus was active and stacked, and the GC Fuji ORTHO LC surface streptococcus was implanted after 1 months. In NSC2, NSC3 and NSC4, the number of Streptococcus mutants showed a decreasing trend in NSC2, NSC3 and NSC4 groups, and the morphology tended to disperse and single, indicating that the addition of nano silver enhanced the adhesion inhibition of GC Fuji ORTHO LC to Streptococcus mutans to some extent. Killing effect.
Experiment four micro-CT observation of enamel demineralization in samples of animals
3 months after the implantation of the sample, the sample teeth in the oral cavity of 15 rats with obvious caries have been taken out under local anesthesia. After cleaning, the adhesive and bottom plate of the tooth surface are removed and the micro-CT scan is performed at the resolution of 10 microns. After three-dimensional imaging, whether the enamel surface around the original adhesive zone has the lacunae left after the demineralization is checked. The results showed that a large number of lacunae left after the enamel demineralization was seen on the side of the crown of the 3M Transbond XT group, and there was a distinct boundary between the buccal surface. The lacunae left after the demineralization of the enamel was found in the side of a sample of the GC Fuji ORTHO LC group; the buccal neck and two sides of all the samples of the samples in NSC2, NSC3 and NSC4 groups were in the NSC2, NSC3 and NSC4 groups. No obvious absorption lacunae were found in the surface, which indicated that nano silver could effectively inhibit the formation of bacterial adhesion and plaque around the material during the observation period, thus preventing or reducing the probability of enamel demineralization.

【学位授予单位】：武汉大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R783.1

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