本文選題：納米羥基磷灰石 + 聚丙交酯-乙交酯　； 參考：《吉林大學》2012年博士論文

【摘要】：近來，工程學和材料學的眾多進展，推動了骨組織工程學的發(fā)展。支架材料由天然材料向人工合成高分子材料發(fā)展，由單一支架材料向復合材料及表面修飾材料發(fā)展。天然骨基質是一種無機/有機結合的復合材料，包括天然形成的聚合物（膠原蛋白）和生物礦物質（磷灰石）。并且天然骨基質中羥基磷灰石以小于100nm的晶體與膠原纖維緊密結合構成納米羥基磷灰石/膠原復合材料。因此，構建無機/有機結合的納米復合材料能夠模擬天然骨基質，能更好的促進骨組織的修復。 由此，制備了納米復合材料模擬天然骨基質，一方面本研究利用冷凍干燥法制備了不同n-HA含量的n-HA/PLGA多孔復合材料支架，并從孔隙率，力學強度，掃描電鏡，接觸角對其進行了表征，并探討了不同的n-HA對多孔復合材料支架結構的影響，及對成骨細胞增殖的影響。另一方面PLGA與n-HA的復合，在一定程度上可以提高PLGA的生物活性和骨引導、誘導能力，但仍然不能滿足修復大面積骨缺損的需要，本研究利用冷凍干燥法制備了多孔復合材料支架，并引入了具有生物活性的廉價易獲得小分子化合物，二甲基砜（MSM）。探討了MSM含量的不同對多孔復合材料支架結構功能和生物學行為的影響。本研究還合成了介孔納米羥基磷灰石，并初步探討了表面活性劑比率的不同對其結構和介孔結構對蛋白擔載性能的影響。為組織工程支架在臨床的應用提供實驗依據(jù)，并開發(fā)具有生物活性的廉價的組織工程支架材料。 第一部分HA/PLGA多孔復合材料支架冷凍干燥法制備及表征和細胞增殖的研究。課題組前期工作中了，研究了不同的冷凍速率對多孔支架材料的影響。結果顯示4℃凍存制備的多孔支架，具有更大的孔徑，更高的孔隙率。本研究選用4℃作為凍存溫度，探討n-HA含量的不同對多孔復合材料支架結構性能和細胞增殖的影響。利用冷凍干燥法制備了不同n-HA含量的n-HA/PLGA多孔復合材料支架，n-HA含量分別為0wt.%、5wt.%、10wt.%、20wt.%和40wt.%。利用掃描電鏡觀察多孔支架材料的表面形貌，乙醇溶液置換法測試復合材料的孔隙率，電子式萬能試驗機測試復合材料的力學強度。n-HA與PLGA鋪膜接觸角測試儀測量接觸角。小鼠成骨前體細胞（MC3T3-E1）種植到多孔支架上，MTT法檢測不同材料在3d、7d和14d對成骨細胞增殖的影響。結果顯示：支架孔隙分布均勻，且孔隙保持連通。當n-HA含量為10wt.%和20wt.%時，顯示了較好孔隙率，潤濕性和較高的力學強度，能有效的促進成骨細胞的增殖。 第二部分生物活性MSM/HA/PLGA多孔復合材料支架的制備及表征。前期工作中發(fā)現(xiàn)n-HA/PLGA復合材料，在一定程度上雖然可以提高PLGA的生物活性和骨引導、誘導能力，，但仍然不能滿足修復大面積骨缺損的需要。因此，本研究利用冷凍干燥法引入了具有生物活性并且廉價易獲得小分子化合物二甲基砜（MSM），探討了MSM含量的不同對多孔復合材料的影響。利用掃描電鏡觀察多孔支架材料的表面形貌，乙醇溶液置換法測試復合材料的孔隙率，MSM、n-HA與PLGA鋪膜接觸角測試儀測量接觸角。電子式萬能試驗機測試復合材料的力學強度。利用電感耦合等離子光譜發(fā)生儀測試了多孔支架中MSM的實際含量和累積釋放量。結果顯示：制備的支架具有均勻的孔隙分布，及良好的貫通性。MSM含量為0.1%時孔隙率和力學強度較高，冷凍干燥法能有效的將MSM引入組織工程支架，未造成MSM的明顯丟失，并能延長MSM的釋放時間。 第三部分MSM/HA/PLGA多孔復合材料支架的細胞增殖、黏附和成骨研究。前期工作中，制備了MSM/HA/PLGA多孔復合材料支架，本研究探討多孔復合材料支架對成骨細胞增殖、黏附和骨修復的影響。利用MTT法，DAPI染色和堿性磷酸酶活性測定，觀察多孔支架材料對成骨細胞增殖和黏附滲透的影響，將支架材料植入兔橈骨缺損部位，術后4w和12w采用X光片檢測骨形成量，以評價材料的骨修復能力。結果顯示：MSM含量為0.1%時顯示了較好的成骨增殖和黏附能力，能有效的促進骨修復。 第四部分介孔納米羥基磷灰石的制備表征及蛋白擔載的研究。揭示表面活性劑的比率介孔n-HA結構的影響，以及介孔結構對HA吸附和釋放牛血清白蛋白（BSA）的影響。利用濕沉淀法，用CTAB作為模板在室溫下成功合成了介孔n-HA，用X射線衍射儀對合成的n-HA相位和晶體結構進行了表征。用傅里葉紅外變換光譜對合成納米粒子的化學結構基團進行測定。用場發(fā)射掃描電鏡進行表面形貌分析。用透射電鏡觀察納米粒子的尺寸和形狀。用多角度光散射儀測試粒徑。用氮氣吸附測試合成粒子的孔隙結構參數(shù)。用BSA測試樣品對蛋白擔載釋放性能的影響。結果顯示：合成的樣品均為純相納米低結晶度的介孔材料，當CTAB比率為0.5%時，顯示了良好的介孔結構，較大的蛋白吸附和較長的釋放時間。
[Abstract]:Recent advances in engineering and material science have promoted the development of bone tissue engineering. Scaffolds have developed from natural materials to synthetic polymers, from single scaffolds to composite materials and surface modified materials. Natural bone matrix is an inorganic / organic composite, including naturally formed polymers. Collagen and biological minerals (apatite). And hydroxyapatite in the natural bone matrix, with a compact combination of less than 100nm crystals and collagen fibers, constitutes a nano hydroxyapatite / collagen composite. Therefore, the construction of inorganic / organic composite nanocomposites can simulate the natural bone matrix and can better promote the repair of bone tissue.
As a result, nano composites were prepared to simulate natural bone matrix. On the one hand, the porous n-HA/PLGA composite scaffolds with different n-HA content were prepared by freeze-drying method and characterized by porosity, mechanical strength, scanning electron microscope and contact angle, and the influence of different n-HA on the structure of porous composite scaffold was discussed. And the effect on osteoblast proliferation. On the other hand, the combination of PLGA and n-HA, to some extent, can improve the biological activity of PLGA, bone guidance and induction, but still can not meet the needs of repairing large area bone defects. A small molecular compound, two methyl sulfone (MSM) was easily obtained. The effects of different MSM content on the structure and biological behavior of the porous composite scaffold were investigated. The mesoporous nano hydroxyapatite was also synthesized in this study. The effects of the different surface active agent ratio on the structure and the pore structure of the mesoporous structure on the performance of the protein loading were also preliminarily discussed. To provide experimental evidence for clinical application of tissue engineering scaffolds, and develop cheap bioengineering scaffolds with bioactivity.
The first part of the study on the preparation, characterization and cell proliferation of HA/PLGA porous composite scaffolds. In the previous work, the effects of different freezing rates on porous scaffolds were studied. The results showed that the porous scaffolds prepared at 4 degrees centigrade had larger pore diameter and higher porosity. This study was selected at 4 degrees C for this study. The effects of different n-HA content on the structural properties and cell proliferation of porous composite scaffolds were investigated. The n-HA/PLGA porous composite scaffolds with different n-HA content were prepared by freeze-drying method. The n-HA content was 0wt.%, 5wt.%, 10wt.%, 20wt.% and 40wt.%., respectively, to observe the surface morphology of porous scaffold materials by scanning electron microscope. The porosity of the composite was measured by the ethanol solution replacement method. The mechanical strength of the composite material was measured by the electronic universal testing machine (.N-HA) and the contact angle tester of PLGA membrane. The mouse osteogenic precursor cells (MC3T3-E1) were planted on the porous scaffold. The effects of different materials on the proliferation of osteoblasts were detected by different materials in 3D, 7d and 14d. The pore distribution is uniform and the pores remain connected. When the content of n-HA is 10wt.% and 20wt.%, it shows good porosity, wettability and high mechanical strength, which can effectively promote the proliferation of osteoblasts.
The preparation and characterization of the second bioactive MSM/HA/PLGA porous composite scaffold. In the previous work, it was found that n-HA/PLGA composites, to some extent, could improve the biological activity of PLGA, bone guidance and induction, but still could not meet the needs of repairing large area bone defects. Therefore, this study uses the freeze drying method. Two methyl sulfone (MSM), which has biological activity and cheap and easy to obtain small molecular compound (MSM), has been studied. The effect of different MSM content on porous composite material is investigated. The surface morphology of porous scaffold is observed by scanning electron microscope, the pore rate of composite material is measured by the method of ethanol solution replacement, and the measurement connection of MSM, n-HA and PLGA membrane contact angle tester The mechanical strength of the composite was tested by an electronic universal testing machine. The actual content and cumulative release of MSM in the porous scaffold were measured by an inductively coupled plasma spectrometer. The results showed that the prepared scaffolds had a uniform pore distribution and a good porosity and high mechanical strength when the good penetrating.MSM content was 0.1%. Freeze drying can effectively introduce MSM into tissue engineering scaffolds without causing significant loss of MSM and prolonging the release time of MSM.
The third part of the cell proliferation, adhesion and osteogenesis of MSM/HA/PLGA porous composite scaffold. In the early work, the MSM/HA/PLGA porous composite scaffold was prepared. The effects of porous composite scaffold on osteoblast proliferation, adhesion and bone repair were investigated. The MTT method, DAPI staining and alkaline phosphatase activity were used to observe the multi porous composite scaffold. The effect of scaffold material on osteoblast proliferation and adhesion permeability, the scaffold material was implanted into the rabbit radius defect site. 4W and 12W were used to measure the bone shape to evaluate the bone repair ability after 4W and 12W. The results showed that when the content of MSM was 0.1%, the bone proliferation and adhesion ability was better, and the bone repair could be promoted effectively.
The fourth part of the preparation of mesoporous nano hydroxyapatite and the study of protein loading. The effect of the ratio of mesoporous n-HA on the ratio of surfactant and the effect of mesoporous structure on HA adsorption and release of bovine serum albumin (BSA). The mesoporous n-HA was successfully synthesized with CTAB as a template at room temperature and was diffracted by X ray diffraction. The phase and crystal structure of the synthesized n-HA were characterized. The chemical structural groups of the synthesized nanoparticles were measured by Fourier transform infrared spectroscopy. The surface morphology was analyzed by field emission scanning electron microscopy. The size and shape of the nanoparticles were observed by transmission electron microscopy. The particle size was measured by the multi angle light scattering instrument. The nitrogen gas was used to measure the particle size. The pore structure parameters of the particles were tested. The effect of samples on the release performance of protein loaded with BSA showed that the synthesized samples were mesoporous materials with low crystallinity in pure phase. When the ratio of CTAB was 0.5%, the good mesoporous structure, large protein adsorption and longer release time were shown.
【學位授予單位】：吉林大學
【學位級別】：博士
【學位授予年份】：2012
【分類號】：R318.08

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相關期刊論文 前1條

1 傅德皓;楊述華;馬德彰;李鯤;郜勇;;鼠胚成骨細胞的原代培養(yǎng)與鑒定[J];創(chuàng)傷外科雜志;2006年02期

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