聚乳酸接枝改性纳米生物玻璃/PLGA复合材料的制备、表面性质及生物活性

以丙交酯开环聚合原位接枝改性的纳米生物玻璃(PLLA-g-BG)与聚丙交酯-乙交酯(PLGA)复合材料为研究对象, 采用TGA, ESEM和EDX分析其接枝率, 粒子分散性和表面元素分布, 通过将兔成骨细胞种植于材料膜表面进行体外培养, 采用荧光染色法、NIH Image J图像分析软件、MTT法和流式细胞术等手段检测细胞在材料表面的平均黏附数量、扩展面积比、增殖能力和细胞周期的变化, 综合评价新型改性纳米复合材料的生物相容性和生物活性. 结果表明, 聚乳酸表面接枝改性可明显改善纳米生物玻璃粒子的团聚; PLGA中掺入一定比例的改性PLLA-g-BG可明显促进兔成骨细胞的黏附、扩展与增殖; 改性纳米生物玻璃的应用可提高生物可降解聚酯材料的生物相容性和生物活性.     

电活性可生物降解纳米复合材料PAP/op-HA/PLGA的制备及成骨活性

以苯胺五聚体与聚乳酸（PLA）的三嵌段共聚物（PAP）与表面接枝低聚乳酸的纳米羟基磷灰石（op-HA）和聚丙交酯-乙交酯（PLGA）的复合物共混,制备的电活性可降解纳米复合材料PAP/op-HA/PLGA为研究对象,采用紫外可见光谱、循环伏安扫描和标准四探针法分析其电化学特性及电导率。ESEM观察其膜表面形貌,接触角评价其亲水性。通过在材料膜表面接种兔成骨细胞进行体外培养,采用荧光染色、NIH Image J图像分析和Real-time PCR综合评价细胞在材料表面的粘附、扩展（细胞面积比）和成骨相关基因的表达水平,以此评价新型电活性纳米复合骨修复材料PAP/op-HA/PLGA的表面性质和生物活性。结果表明,PAP/op-HA/PLGA的电导率较低（~5?10?6 S/cm）但具有良好的电化学氧化还原性能,PAP含量为0.1%时材料的亲水性明显改善,成骨细胞的粘附和扩展明显增强。培养7天时骨形态蛋白- 2（BMP-2）和骨连接蛋白（Osteonectin）的基因表达水平明显提高,而对Ⅰ型胶原蛋白的基因表达没有明显影响。提示PAP/op-HA/PLGA具有良好的细胞相容性和成骨活性。     

通过正离子聚合原位制备聚谷氨酸苄酯-g-(聚四氢呋喃-b- 聚异丁烯)/银纳米复合材料及其性能研究

采用烯丙基溴官能化聚异丁烯/高氯酸银体系引发四氢呋喃活性正离子聚合制备聚谷氨酸苄酯-g-(聚四氢呋喃-b-聚异丁烯)/银纳米复合材料(PBLG-g-(PTHF-b-PIB)/Ag).研究接枝密度对该纳米复合材料表面组成、形貌及自组装行为的影响,进一步探究纳米复合材料中银的含量、分布、晶型及存在形态,以布洛芬(IBU)作为模拟药物,研究接枝共聚物中接枝密度和平均支链长度对纳米复合材料载药释药行为的影响,通过抑菌圈法和MTT法研究纳米复合材料的抗菌性和细胞毒性.结果表明:通过烯丙基溴官能化聚异丁烯大分子引发四氢呋喃活性正离子开环聚合,可以原位制备不同接枝密度和不同平均支链长度的PBLG-g-(PTHF-b-PIB)/Ag纳米复合材料,其中银的质量含量在0.25%~3.9%之间,与其理论含量基本相吻合,银颗粒以聚集体形态存在,尺寸为5~10 nm,晶型为面心立方结构;该纳米复合材料在四氢呋喃/正己烷(4/1,V/V)混合介质中自组装形成胶束,胶束数目随接枝密度增加而减少,但尺寸增大;随接枝共聚物中接枝密度和纳米银含量增加,纳米复合材料的疏水性增加;随接枝共聚物中接枝密度增加,纳米复合材料表面形貌可由球形结构逐渐转化为双连续相结构;纳米复合材料的载药微球可以通过接枝共聚物中主链PBLG的空心螺旋结构、酰胺键及PTHF支链的醚键结构显示三重载药特性,载药量和累积释药量均随着接枝共聚物中接枝密度或PTHF链段长度增加而增加,且在37oC下的释药率是25oC下释药率的3倍左右.该纳米复合材料的抗菌性能随纳米银含量增加而增加,当纳米银含量为1.48%时,该纳米复合材料1周后细胞存活率为97.7%,即无细胞毒性.     

表面接枝嵌段共聚物刷实现内皮细胞的选择性黏附

利用表面引发原子转移自由基聚合(SI-ATRP)技术将聚(甲基丙烯酸寡聚乙二醇酯)和聚(甲基丙烯酸缩水甘油酯)的双嵌段共聚物刷(POEGMA-b-PGMA)接枝在材料表面,并通过PGMA中丰富的环氧基团开环固定可特异性黏附内皮细胞的多肽GREDVY.静态水接触角、接枝层厚度、X射线光电子能谱(XPS)以及原子力显微镜(AFM)的测试结果证明了各步接枝反应的成功性.细胞培养研究表明这种连接有GREDVY的双嵌段共聚物表面能够有效的促进血管内皮细胞的黏附,同时排斥成纤维细胞的黏附,从而实现了内皮细胞的选择性黏附.     

新型介孔羟基磷灰石/壳聚糖-万古霉素药物释放系统复合材料的制备及体外抗菌和成骨能力

采用冷冻干燥法合成了介孔羟基磷灰石(HA)/壳聚糖(CS)-万古霉素(VCM)药物释放系统复合材料, 利用SEM, XRD和FTIR等方法对材料进行了表征. 结果证实CS与HA混合复合材料具有良好的孔径和孔隙率, 万古霉素吸附于复合材料的表面和内部. 细胞毒性实验[噻唑蓝(MTT)比色法]结果表明, 材料可以促进成骨细胞增殖且具有良好的细胞相容性. 体外抑菌实验结果证实此材料可长时间抑制耐甲氧西林金葡菌(MRSA)的生长, 具有良好的抑菌和杀菌能力. 细胞黏附实验结果表明, 成骨细胞附着于材料表面增殖并通过孔道延伸. 实时聚合酶链式反应(RT-PCR)实验结果表明, 在成骨相关标志产物胶原蛋白-1(COL-1)及骨形态发生蛋白-2(BMP-2)基因上均有较高的表达, 表明材料在体外可以促进成骨细胞生长, 具有良好的成骨能力.     

聚L-乳酸/二氧化硅纳米复合材料及其表面诱导生成类骨磷灰石的制备

为改善纳米SiO2粒子在聚L-乳酸基体中的分散性, 将乳酸齐聚物接枝到纳米SiO2粒子表面, 通过IR, 29Si MAS NMR和TGA对改性SiO2进行表征. 以聚L-乳酸(PLLA)为基体, 加入乳酸齐聚物接枝改性的二氧化硅(g-SiO2)粒子, 采用溶液浇铸法制备PLLA/g-SiO2纳米复合材料, 测试其在模拟体液(SBF)中的生物活性. 通过XRD, IR, SEM和EDS表征手段, 考察材料表面类骨磷灰石形成能力. 结果表明, 乳酸齐聚物成功地接枝到SiO2表面, 当反应36 h时, g-SiO2接枝率最大(9.22%). 随着g-SiO2含量增加和浸泡时间的延长, 材料表面最初形成的无定形沉积物矿化成碳酸羟基磷灰石(Carbonated hydroxyapatite, CHA), 钙磷比为1.72, 类似于人骨无机质, 表明g-SiO2的引入能明显加速复合材料表面CHA沉积, 该复合材料有望成为骨修复填充材料和组织工程支架材料.     

聚(丙交酯-co-乙交酯)/β-磷酸三钙复合物的电纺研究

对生物可吸收聚(丙交酯-co-乙交酯)(poly(lactide-co-glycolide),PLGA)与β-磷酸三钙(-βTCP)复合物体系进行了电纺.研究了PLGA的浓度,-βTCP与PLGA比例,加料速度,电压,喷头与接收体之间的距离等因素对电纺过程的影响,制备出纳米纤维膜,并用扫描电镜(SEM)等对纤维膜进行表征.结果表明,电纺溶液浓度越高,或者加料速度越快,纳米纤维的直径越粗.力学实验显示,复合物中-βTCP的含量增加使纳米纤维膜的拉伸强度和杨氏模量下降.     

肾上腺髓质素微球复合PLGA/纳米羟基磷灰石支架材料的制备及生物活性评价

利用离子乳化交联法制备了负载肾上腺髓质素的壳聚糖微球,应用热致相分离法制备了乳酸和乙醇酸共聚物/纳米羟基磷灰石(PLGA/nHA)支架材料并在其中包覆载药微球.通过扫描电子显微镜、体外释放行为、材料溶血行为、碱性磷酸酶(ALP)活性的测定、支架材料表面细胞荧光染色和MTT[3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐]比色法等手段综合评价载药支架材料的性能及生物活性.结果表明,微球直径均匀,载药支架孔径大小合适并相互穿通.支架材料的溶血率小于5%,符合医用材料的溶血实验要求.载药支架及支架材料本身对成骨细胞及血管内皮细胞的增殖以及成骨细胞的分化均有一定的促进作用.     

与胶原特异性结合的BMP2模拟肽/PLGA 3D打印复合支架的制备及成骨诱导活性

将胶原绑定结构域(CBD)多肽序列与骨形态发生蛋白2模拟肽(BMP2-MP)序列连接制备具有胶原绑定能力的CBD-BMP2-MP, 再将CBD-BMP2-MP与聚丙交酯-乙交酯/胶原(PLGA/COL)3D打印支架相结合, 以支架表面的胶原成分为媒介, 将CBD-BMP2-MP更有效地固定于骨修复材料上, 达到对其进行改性的目的. 利用扫描电子显微镜(SEM)、 电子万能试验机和接触角测量仪对复合支架表面形貌、 力学强度和亲水性等材料学性能进行评价. 用荧光成像法评测 CBD-BMP2-MP及BMP2-MP与支架材料的结合能力. 在各组支架材料表面接种MC3T3-E1细胞进行体外培养, 采用CCK-8、 鬼笔环肽荧光染色、 茜素红染色及qPCR综合评价细胞在材料表面的黏附、 增殖和成骨分化等细胞行为, 研究CBD-BMP2-MP修饰的3D多孔PLGA/COL复合支架的生物学性能. 研究结果表明, 利用3D打印技术制备的多孔支架具有形貌可控的孔隙结构, 为细胞生长创造更有利的细胞微环境, 支架表面胶原成分的加入提高了支架材料的亲水性, 同时对支架材料本身的力学性能无任何影响, 提高了复合支架本身的生物相容性. 与普通BMP2-MP相比, CBD-BMP2-MP具有更好的胶原绑定能力, 与复合支架的结合更稳定, 提高了PLGA/COL复合支架对BMP2-MP的负载能力. 支架表面负载CBD-BMP2-MP后具有极强的促细胞成骨分化能力. MC3T3-E1细胞表现出更高的钙沉积能力, 并且成骨分化相关基因Runx2, ALP, COL-I及OPN等水平也有了明显提升. 表明CBD-BMP2-MP多孔复合支架具有良好的生物相容性和成骨诱导活性, 在骨组织修复领域具有良好的应用前景.     

纳米羟基磷灰石表面接枝聚合左旋丙交酯

为了改善HA纳米粒子与有机PLGA的相容性,分别采用六亚甲基二异氰酸酯加乙二醇、左旋乳酸改性纳米粒子表面后或直接原位接枝聚合左旋丙交酯等3种不同方法,制备了表面修饰聚乳酸的纳米羟基磷灰石(PLLA-g-HA).FTIR、XPS、TEM、TGA测试表明PLLA成功接枝到HA的表面.其中六亚甲基二异氰酸酯加乙二醇改性HA纳米粒子所获得的PLLA接枝率远高于其它两种方法达25%,调整有机相和无机相的比例对PLLA接枝率的影响较小,其在氯仿中可以稳定分散2天以上.共混电纺丝后的拉伸测试表明PLLA-g-HA/PLAG复合纤维膜的力学性能高于HA/PLGA膜,当两者之间的比例为5%拉伸性能达到最大值.     

聚(L-谷氨酸)水凝胶介导羟基磷灰石的生物矿化

以1-乙基-3-(3-二甲基氨基丙基)碳二亚胺盐酸盐(EDC·HCl)为羧基活化剂, 己二酸二酰肼(ADH)为交联剂, 制备了生物活性聚(L-谷氨酸)(PLGA)水凝胶. 通过X射线衍射和扫描电子显微镜等表征了在不同浓度模拟体液(SBF)中羟基磷灰石(HA)的形成和生长. PLGA水凝胶的表面和内部均可观察到HA的形成和生长. 同时探讨了PLGA水凝胶矿化前后的力学性能. 将矿化前后PLGA水凝胶用于脂肪干细胞(ASCs)的培养, 研究其细胞相容性.     

Preparation of porous nanocomposite scaffolds with honeycomb monolith structure by one phase solution freezedrying method

Biodegradable porous nanocomposite scaffolds of poly（lactide-co-glycolide）（PLGA） and L-lactic acid（LAc） oligomer surface-grafted hydroxyapatite nanoparticles（op-HA） with a honeycomb monolith structure were fabricated with the single-phase solution freeze-drying method.The effects of different freezing temperatures on the properties of the scaffolds,such as microstructures,compressive strength,cell penetration and cell proliferation were studied.The highly porous and well interconnected scaffolds with a tunable pore structure were obtained.The effect of different freezing temperature（4℃,-20℃,-80℃and -196℃） was investigated in relation to the scaffold morphology,the porosity varied from 91.2%to 83.0%and the average pore diameter varied from（167.2±62.6）μm to（11.9±4.2）μm while theσ₁₀ increased significantly.The cell proliferation were decreased and associated with the above-mentioned properties.Uniform distribution of op-HA particles and homogeneous roughness of pore wall surfaces were found in the 4℃frozen scaffold.The 4℃frozen scaffold exhibited better cell penetration and increased cell proliferation because of its larger pore size,higher porosity and interconnection.The microstructures described here provide a new approach for the design and fabrication of op-HA/PLGA based scaffold materials with potentially broad applicability for replacement of bone defects.     

DOPA-IGF-1 Coated HA/PLGA Microspheres Promoting Proliferation and Osteoclastic Differentiation of Rabbit Bone Mesenchymal Stem Cells

To explore the ability of dihydroxyphenylalanine-insulin-like growth factor-1 (DOPA-IGF-1) coated hydroxyapatite/poly(lactic-co-glycolic acid)(HA/PLGA) microspheres to promote the proliferation and osteoclastic differentiation of rabbit bone mesenchymal stem cells(rBMSCs), HA/PLGA microspheres with different HA content (10%, 30%, 50%, mass fraction) were prepared by electrospinning method and HA/PLGA microspheres with 50% HA were coated with IGF-1 and DOPA-IGF-1, respectively. They were co-cultured with rBMSCs, respctively. Cell counting kit-8(CCK-8) detection, confocal laser scanning microscopy(CLSM), alkaline phosphatase(ALP) detection and osteogenesis related genes COL IA1, Runx2 and bone morphogenetic protein-2(BMP-2) detection were conducted to detect the proliferation activity, cell morphology, differentiation ability and the expression level of osteogenesis-related genes of cells cultured on all microspheres groups. The results showed that rBMSCs proliferation increased in an HA content dependent manner, and cells proliferated more in the IGF-1 coated and DOPA-IGF-1 coated groups, in particular in DOPA-IGF-1 coated group, and the differences were more remarkable over time (P<0.05). HA/PLGA microspheres promoted the proliferation and osteogenic differentiation of rBMSCs, and DOPA-IGF-1 coating enhanced the proliferation and osteogenic differentiation of rBMSCs.     

Fabrication of poly(lactide‐co‐glycolide) scaffold embedded spatially with hydroxyapatite particles on pore walls for bone tissue engineering

Poly(lactide‐co‐glycolide) (PLGA) scaffolds embedded spatially with hydroxyapatite (HA) particles on the pore walls (PLGA/HA‐S) were fabricated by using HA‐coated paraffin spheres as porogens, which were prepared by Pickering emulsion. For comparisons, PLGA scaffolds loaded with same amount of HA particles (2%) in the matrix (PLGA/HA‐M) and pure PLGA scaffolds were prepared by using pure paraffin spheres as porogens. Although the three types of scaffolds had same pore size (450–600 µm) and similar porosity (90%–93%), the PLGA/HA‐S showed the highest compression modulus. The embedment of the HA particles on the pore walls endow the PLGA/HA‐S scaffold with a stronger ability of protein adsorption and mineralization as well as a larger mechanical strength against compression. In vitro culture of rat bone marrow stem cells revealed that cell morphology and proliferation ability were similar on all the scaffolds. However, the alkaline phosphatase activity was significantly improved for the cells cultured on the PLGA/HA‐S scaffolds. Therefore, the method for fabricating scaffolds with spatially embedded nanoparticles provides a new way to obtain the bioactive scaffolds for tissue engineering. Copyright © 2011 John Wiley & Sons, Ltd.     

A quaternary composite fiber membrane for guided tissue regeneration

This work reported a novel composite fiber membrane containing poly(1actic‐co‐glycolic acid)/six‐armed poly(ε‐caprolactone)/hydroxylapatite‐grafted poly(l ‐lactide)/Pluronic F‐108 (PLGA/SAPCL/HA‐g‐PLLA/PF‐108) produced by electrospinning. SAPCL, HA‐g‐PLLA, and PF‐108 could be well blended with PLGA to make fibers. Fibrous surface and diameter had little difference in morphology with the change of fibrous component. Compared with simple PLGA, PLGA/SAPCL, or PLGA/SAPCL/HA‐g‐PLLA fiber membrane, the quaternary composites not only showed the improved mechanics stability but also possessed better hydrophilicity. The quaternary membrane was conductive to accelerate degradation and maintained nice bioactivity. It was not cytotoxicity. These results suggest that the composite fiber membrane may be qualified as guided bone regeneration substrate. Copyright © 2015 John Wiley & Sons, Ltd.     

Polyelectrolyte multilayers and degradable polymer layers as multicompartment films

Polyelectrolyte multilayers are now a well established concept with numerous potential applications in particular as biomaterial coatings. To timely control the biological activity of cells in contact with a substrate, multicompartment films made of different polyelectrolyte multilayers deposited sequentially on the solid substrate constitute a promising new approach. In a first paper (Langmuir 2004, 20, 7298) we showed that such multicompartment films can be designed by alternating exponentially growing polyelectrolyte multilayers acting as reservoirs and linearly growing ones acting as barriers. In the present study, we first demonstrate however that these barriers composed of synthetic polyelectrolytes are not degraded despite the presence of phagocytic cells. We propose an alternative approach where exponentially growing poly(L-lysine)/hyaluronic acid (PLL/HA) multilayers, used as reservoirs, are alternated with biodegradable polymer layers consisting in poly(lactic-co-glycolic acid) (PLGA) and acting as barriers for PLL chains that diffuse within the PLL/HA reservoirs. We first show that these PLGA layers can be deposited alternatively with PLL/HA multilayers leading to polyelectrolyte multilayer/hydrolyzable polymeric layer films and acting as a reservoirs/barriers system. Bone marrow cells seeded on these films ending by a PLL/HA reservoir rapidly degrade it and internalize the PLL chains confined in this reservoir. Then the cells degraded locally the PLGA barrier and internalize the PLL localized in a lower (PLL/HA) compartment after 5 days of seeding. By changing the thickness of the PLGA layer, we hope to be able to tune the time delay of degradation. Such mixed architectures made of polyelectrolyte multilayers and hydrolyzable polymeric layers could act as coatings allowing us to induce a time scheduled cascade of biological activities. We are currently working on the use of comparable films with compartments filled by proteins or peptides and in which the degradation of the barriers results from a hydrolysis over tunable time scales.     

Loading efficiency and surface conductance of heparin-modified poly(lactide-co-glycolide) nanoparticles

Poly(lactide-co-glycolide) (PLGA) nanoparticles (NPs) with surface modification of heparin were fabricated by microemulsion–diffusion method. These novel colloidal particles were stabilized by lecithin and Tween 80. The effects of lecithin on the loading of heparin onto PLGA NPs and on the surface conductance were analyzed. The electronic micrographs revealed that spherical colloids were prepared and the incorporation of heparin caused a slight coalescence of the particles. In addition, the average diameter of heparin-modified PLGA NPs was between 70 and 220 nm. An increase in the weight percentage of lecithin or in the concentration of heparin enlarged the average diameter. Based on constant amount of surfactants, the loading efficiency of heparin on the particle surfaces reached a maximum when the weight percentage of lecithin was 50%. Moreover, the surface conductance of heparin-modified PLGA NPs was improved by an increased weight percentage of lecithin. A high concentration of heparin in microemulsion also promoted the loading efficiency and surface conductance of heparin-modified PLGA NPs.     

The influence of UV-radiation on hyaluronic acid and its blends with addition of collagen and chitosan

In this paper, the results regarding the influence of UV-irradiation with 254?nm wavelength on the surface and mechanical properties of hyaluronic acid, hyaluronic acid/collagen and hyaluronic acid/collagen/chitosan mixtures are presented. For this study, thin films were prepared by solvent evaporation from solution of HA and mixtures made from HA/Coll and HA/Coll with 30% addition of chitosan. The surface properties of films were investigated by AFM and using contact angle measurements, allowing the calculation of surface free energy and its components. Mechanical properties of films made of biopolymeric blends before and after UV-irradiation have been investigated by mechanical testing machine.