本文選題：種植體　切入點(diǎn)：氧化石墨烯　出處：《第四軍醫(yī)大學(xué)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：鈦及鈦合金因其良好的材料學(xué)性能和生物學(xué)性能被廣泛應(yīng)用于牙科領(lǐng)域,但因其上部結(jié)構(gòu)暴露在口腔環(huán)境中,種植體-軟組織結(jié)合較薄弱部位容易受細(xì)菌侵襲;術(shù)中淤血及受損種植位點(diǎn)為細(xì)菌繁殖提供理想環(huán)境;植體下段表面粗糙程度不僅易于形成骨結(jié)合,也方便細(xì)菌黏附和定植;鈦及鈦合金的生物惰性性質(zhì),這些因素均會誘導(dǎo)種植體周圍炎的發(fā)生。因此,如何使種植體材料表面修飾具有生物活性和抗菌性能以提高種植成功率是眾多學(xué)者一直研究的重點(diǎn)和難點(diǎn)。本研究擬采用電鍍法和紫外還原法使純鈦表面獲得膜狀和顆粒狀改性,以口腔常見菌為研究對象,就改性后純鈦表面抗菌性能進(jìn)行研究,并對其生物活性進(jìn)行初步評價,從抗菌和生物活性角度優(yōu)化種植體材料表面修飾。第一部分種植體表面Ti-GO-Ag多相復(fù)合材料的構(gòu)建及性能分析[目的]在純鈦(Titanium,Ti)表面構(gòu)建不同濃度的氧化石墨烯(Graphene oxide,GO)膜和銀(Silver,Ag)顆粒,并對其材料學(xué)性能進(jìn)行分析。[方法]通過正交實(shí)驗(yàn)設(shè)計設(shè)計工藝參數(shù);電壓為20 V,時間為5 min,濃度分別為20μg/ml、50μg/ml、80μg/ml、100μg/ml條件下通過電鍍法對Ti進(jìn)行修飾;采用紫外還原法在濃度為1 mol/L的硝酸銀溶液中對Ti-GO進(jìn)行加載;利用場發(fā)射掃描電子顯微鏡對表面形貌進(jìn)行觀察;使用拉曼光儀對GO的性狀進(jìn)行分析;通過X射線光電子能譜對Ag進(jìn)行表征;采用原子力顯微鏡對材料表面輪廓和粗糙度進(jìn)行研究;利用接觸角測試儀對材料表面親水性進(jìn)行研究;使用納米壓痕和納米劃痕對材料表面彈性模量和摩擦力進(jìn)行探討;通過電感耦合等離子體質(zhì)譜儀對Ag離子釋放進(jìn)行研究。[結(jié)果]正交實(shí)驗(yàn)優(yōu)化的工藝參數(shù):電壓:20 V,時間:5 min;Ti表面經(jīng)過改性以后表面有類膜物質(zhì)和類顆粒狀物質(zhì),經(jīng)分析可知膜狀物質(zhì)為GO,且濃度不同,GO在Ti表面的形貌和量也不一樣;顆粒狀物質(zhì)為Ag單質(zhì),與GO的濃度成正比;表面輪廓高低起伏,粗糙度與濃度成正比;Ti-GO與濃度成反比,Ti-GO-Ag與濃度成反比;彈性模量與維氏硬度隨GO濃度升高而升高,摩擦力與濃度沒有正反比關(guān)系;各組在第四天Ag離子釋放量出現(xiàn)相似情況。[結(jié)論]正交實(shí)驗(yàn)設(shè)計可以優(yōu)化實(shí)驗(yàn)過程,并使Ti表面獲得修飾;改性后的Ti,其表面形貌、輪廓、粗糙度、親水性能、彈性模量、維氏硬度、摩擦力等均與改性前有所不同,使Ti表面獲得生物涂層(如抗菌涂層);四組實(shí)驗(yàn)組Ag離子釋放總體規(guī)律趨向于降低,指導(dǎo)抗菌過程。第二部分種植體表面Ti-GO-Ag多相復(fù)合材料的抗菌性能研究[目的]研究不同濃度的GO和Ag涂層表面對三種口腔常見菌的抗菌作用。[方法]以金黃色葡萄球菌、變形鏈球菌和牙齦卟啉單胞菌為研究對象,評價20μg/ml、50μg/ml、80μg/ml、100μg/ml四種不同濃度Ti-GO-Ag涂層表面形成細(xì)菌生物膜的能力,并對細(xì)菌表面形貌進(jìn)行觀察;使用激光共聚焦顯微鏡對細(xì)菌胞膜完整性進(jìn)行檢測,并用Image J軟件對熒光強(qiáng)度值和涂層表面細(xì)菌三維形貌進(jìn)行研究;利用流式細(xì)胞儀對細(xì)菌凋亡情況進(jìn)行評價;采用透射電子顯微鏡對細(xì)菌內(nèi)部結(jié)構(gòu)進(jìn)行觀察;通過時實(shí)定量熒光PCR技術(shù)檢測細(xì)菌相對基因表達(dá)情況;對改性后Ti的抗菌機(jī)理進(jìn)行初步探討。[結(jié)果]20μg/ml、50μg/ml、80μg/ml、100μg/ml四種不同濃度Ti-GO-Ag涂層表面對金黃色葡萄球菌、變形鏈球菌和牙齦卟啉單胞菌的抗菌效果均達(dá)到或超過89%,明顯優(yōu)于對照組;三種細(xì)菌表面形貌發(fā)生明顯改變;涂層表面細(xì)菌胞膜完整性受損,紅色熒光強(qiáng)度值高于綠色熒光強(qiáng)度值,細(xì)菌活性均低于對照組,細(xì)菌的三維形貌也比對照組量少、薄和松;細(xì)菌凋亡情況明顯不同于對照組;通過內(nèi)部結(jié)構(gòu)觀察可發(fā)現(xiàn)細(xì)菌破壞主要是銀離子接觸并穿過胞膜進(jìn)入到胞內(nèi)而使細(xì)菌結(jié)構(gòu)破壞;三種細(xì)菌的相對基因表達(dá)量均低于對照組。[結(jié)論]不同濃度的GO和Ag涂層對三種口腔常見菌均有抗菌作用,且對革蘭氏陰性菌的效果優(yōu)于革蘭氏陽性菌,其抗菌機(jī)制是銀離子接觸攻擊細(xì)菌胞膜,使胞膜結(jié)構(gòu)穿孔,胞質(zhì)外泄,從而使細(xì)菌裂解死亡。第三部分種植體表面Ti-GO-Ag多相復(fù)合材料的生物活性評價[目的]評價不同濃度的GO和Ag涂層表面生物活性。[方法]通過人體模擬液浸泡實(shí)驗(yàn)研究不同濃度的GO和Ag涂層表面鈣磷沉積情況;利用CCK-8實(shí)驗(yàn)評價不同濃度的GO和Ag涂層表面鼠骨髓間充質(zhì)干細(xì)胞活性情況。[結(jié)果]不同濃度的GO和Ag涂層表面均可沉積鈣磷等元素,沉積效果較好的是對照組、G20組和G80組,其中對照組和G20組、G20組和G80組之間無統(tǒng)計學(xué)差異;四種不同濃度的GO和Ag涂層表面對鼠骨髓間充質(zhì)干細(xì)胞的存活狀況較對照組差,G20和G50組優(yōu)于G80和G100組。[結(jié)論]四種不同濃度的GO和Ag涂層表面擁有不同程度的生物礦化能力,且對鼠骨髓間充質(zhì)干細(xì)胞的存活狀況均有影響。
[Abstract]:Titanium and titanium alloy due to its good properties and biological properties of materials are widely used in the field of Dentistry, but because of the upper structure is exposed to the oral environment, combined with the weak parts of the implant soft tissue susceptible to bacterial invasion; intraoperative congestion and damaged implant sites provide an ideal environment for bacterial reproduction; implant under a rough surface is not only easy to form osseointegration, also facilitate adhesion and colonization of bacteria; biological inert properties of titanium and titanium alloy, these factors have induced peri implantitis. Therefore, how to make the implant material with surface modification and antibacterial properties of biological activity in order to improve the success rate of planting is the key and difficult point in many scholars have been studied. This research adopts the electroplating method and UV reduction method to the surface of pure titanium film and granular modified to obtain oral common bacteria as the research object, the modified titanium surface antibacterial Study of properties, and to evaluate its biological activity, antibacterial and biological activity from the angle optimization of implant material surface modification. The first part of the implant surface Ti-GO-Ag composite construction and performance analysis [Objective] in pure titanium (Titanium, Ti) the surface of graphene oxide with different concentrations of Graphene (oxide, GO) film and silver (Silver, Ag) on the properties of the particles, and materials are analyzed. Through the orthogonal experiment design method of process parameters; the voltage is 20 V, time 5 min, the concentration were 20 g/ml, 50 g/ml, 80 g/ml, 100 g/ml conditions by electroplating method of Ti was modified by UV; reduction method for loading Ti-GO in the concentration of silver nitrate was 1 mol/L; on the surface morphology were observed by field emission scanning electron microscope; analyzed using Raman spectrometer on the GO characters; by X ray photoelectron spectroscopy The Ag was characterized by atomic force microscope; and the roughness of the surface profile of materials; using the instrument of material surface hydrophilic contact angle measurement; using nanoindentation and nano scratch on the surface of the material of elastic modulus and friction force; optimize the process parameters. The results of orthogonal experiment on the release of Ag ions through. Inductively coupled plasma mass spectrometry: voltage: 20 V, time: 5 min; Ti surface modified after surface class membrane materials and class of granular material. The analysis shows that membrane material was GO, and the different concentration of GO on the morphology and quantity of Ti surface is not the same; granular material as Ag elements, a is proportional to the concentration of GO; surface profile undulating, roughness and is proportional to the concentration of Ti-GO; and is inversely proportional to the concentration, Ti-GO-Ag and concentration is inversely proportional to the elastic modulus and hardness; Vivtorinox with GO concentration increased and increased friction No positive relationship with the ratio of concentration; in each group fourth days Ag ion release appeared similar to the situation. We can optimize the process of experiment conclusion] orthogonal experimental design, and the Ti surface modification; modified Ti, the surface morphology, profile, roughness, hydrophilicity, elastic modulus, hardness and friction force of Vivtorinox. Before modification is different, the Ti surface coating (such as antibacterial coating); four experimental groups, Ag ion release tends to reduce the overall law, guide the antibacterial process. The second part of the implant surface Ti-GO-Ag composite antibacterial antibacterial effect. Methods can study [Objective] to study the effects of different concentrations of GO and Ag coating on the surface of three kinds of common oral bacteria Staphylococcus aureus, Streptococcus mutans and Porphyromonas gingivalis as the research object, the evaluation of 20 g/ml, 50 g/ml, 80 g/ml, 100 g/ml formed four different concentrations of Ti-GO-Ag coating The ability of bacterial biofilm, and the bacterial surface morphology were observed; detection of bacterial cell membrane integrity was performed using laser confocal microscopy, and the fluorescence intensity values of bacteria and coating the surface topography by using Image J software; the evaluation of bacterial apoptosis by flow cytometry was observed by transmission electron microscopy; the internal structure of bacteria; by real-time fluorescence PCR detection of bacterial gene expression relative to preliminary study results.;]20 g/ml of the antibacterial mechanism of Ti after 50 g/ml, 80 g/ml, 100 g/ ml four different concentration of Ti-GO-Ag coating on Staphylococcus aureus, antibacterial effect Streptococcus mutans and Porphyromonas gingivalis were reached or exceeded 89%, significantly better than the control group; three kinds of bacterial surface morphology changed obviously; the integrity of the damaged coating on the surface of the bacterial membrane, strong red fluorescence The degree is higher than the value of green fluorescence intensity values, bacterial activity was lower than the control group, the three-dimensional morphology of bacteria than the control group less, Bo Hesong; bacterial apoptosis is significantly different from the control group; the observation of the internal structure of bacteria found damage is mainly silver ions and contact through the membrane into the intracellular bacteria caused structural damage; the relative gene expression of three kinds of bacteria were lower than the control group. Conclusion: different concentrations of GO and Ag coating of common bacteria had antimicrobial effects on three kinds of oral cavity, and the effect is better than the gram positive bacteria of gram negative bacteria, the antibacterial mechanism of silver ions is to attack the bacterial membrane contact, the membrane perforation, cytoplasm leak, causing bacterial lysis and death. GO biological surface Ag coating. The third part activity method of implant surface Ti-GO-Ag multiphase biological activity evaluation of composites [Objective] evaluation of different concentrations through the body Simulation experiment of Soaking with different concentration of GO and Ag coating on the surface of calcium and phosphorus deposition; evaluation of different concentration using CCK-8 experimental GO and Ag coating surface of rat bone marrow mesenchymal stem cells. The activity of the different concentrations of GO and Ag coating could be deposited calcium phosphate precipitation and other elements, better control group, G20 group and G80 group, the control group and G20 group, there was no significant difference between G20 group and G80 group; four different concentrations of GO and Ag coating on the surface of bone marrow mesenchymal stem cell survival situation worse than that of control group, G20 group and G50 group is better than that of G80 and G100. [Conclusion] four different concentrations of GO and Ag coatings have different degrees of biological mineralization, and the influence of bone marrow mesenchymal stem cell survival in both.

【學(xué)位授予單位】：第四軍醫(yī)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：R783.1
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本文編號：1596429