羟基磷灰石／壳聚糖复合微球的制备及其生物学作用的体外研究

目的：通过探究羟基磷灰石（HAp）壳聚糖（CS）复合微球支架对骨髓间充质干细胞体外生物学行为的影响,评估其作为骨组织工程支架的可行性。方法纳米 HAp 和CS 复合物通过微流体技术自组装成微球支架,显微镜下形态学观察。将 P2代骨髓间充质干细胞（BMSCs）与微球行体外共培养,计算前6 h 黏附率。培养1、3、6、9 d,计算增殖率并用 GraphPad Prism 6软件对数据进行处理;行扫描电镜及共聚焦扫描式显微镜检测,观察细胞黏附及分布。将细胞微球复合物填入自制模具中培养14～21 d,进行形态观察。结果显微镜镜下微球为完整的圆形,大小一致。BMSCs 与微球体外培养6 h,黏附率达90％以上。6 d 时,BMSCs 的增殖率达到最高。扫描电镜结果显示微球上有大量 BMSCs 黏附定植并分泌大量胞外基质将微球连接成整体;共聚焦扫描式显微镜结果可见明显的细胞骨架微丝蛋白。细胞微球体外模具培养18 d 后,形成了结构完整的组织块。结论HAp-CS 微球是一种良好的促BMSCs 种子细胞黏附定植的支架材料,是促进细胞生长的有效支撑载体,与共培养细胞形成的复合组织块有望应用于体内动物实验修复标准缺损。     

A New Generation of Electrospun Fibers Containing Bioactive Glass Particles for Wound Healing

Chitosan fibers blended with polyethylene oxide (CHIT_PEO) and crosslinked with genipin were fabricated by electrospinning technique. Subsequently, CHIT_PEO bioactive glass composite electrospun mats were fabricated with the aim to achieve flexible structures with adequate mechanical properties and improved biological performance respect to CHIT_PEO fibers, for potential applications in wound healing. Three different compositions of bioactive glasses (BG) were selected and investigated: 45S5 BG, a Sr and Mg containing bioactive glass (BGMS10) and a Zn-containing bioactive glass (BGMS_2Zn). Particulate BGs (particles size < 20 μm) were separately added to the starting CHIT_PEO solution before electrospinning. The two recently developed bioactive glasses (BGMS10 and BGMS_2Zn) showed very promising biological properties in terms of bioactivity and cellular viability; thus, such compositions were added for the first time to CHIT_PEO solution to fabricate composite electrospun mats. The incorporation of bioactive glass particles and their distribution into CHIT_PEO fibers were assessed by SEM and FTIR analyses. Furthermore, CHIT_PEO composite electrospun mats showed improved mechanical properties in terms of Young’s Modulus compared to neat CHIT_PEO fibers; on the contrary, the values of tensile strain at break (%) were comparable. Biological performance in terms of cellular viability was investigated by means of WST-8 assay and CHIT_PEO composite electrospun mats showed cytocompatibility and the desired cellular viability.     

Fabrication and characterization of chitosan–collagen crosslinked membranes for corneal tissue engineering

This article describes a chitosan–collagen composite membrane as corneal tissue-engineering biomaterials. The membrane was prepared by dissolving the chitosan into collagen with the weight ratio of 0, 15, 30, 45, 60, and 100%, followed by crosslinked with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide. Mechanical properties, contact angles, and optical transmittance were determined and compared between chitosan membrane and crosslinking composite membrane. As a result, the optical transparency and mechanical strength of the chitosan–collagen membranes were significantly better than that of the sample of chitosan. In addition, in vitro cell culture studies revealed that the collagen has no negative effect on the cell morphology, viability, and proliferation and possess good biocompatibility. Overall, the dendrimer crosslinked chitosan–collagen composite membranes showed promising properties that suggest that these might be suitable biomaterials for corneal tissue-engineering applications.     

壳聚糖/葡甘聚糖混合膜对人牙周膜细胞作用观察

目的:通过壳聚糖/葡甘聚糖膜单独、以及膜加载碱性成纤维细胞生长因子和第4代人牙周膜细胞共同培养,观察壳聚糖/葡甘聚糖膜对人牙周膜细胞增殖分化的影响。方法:观察不同质量比壳聚糖与葡甘聚糖共混膜对第4代细胞HPDLCs增殖影响及其对碱性磷酸酶活性(ALP)影响。结果:不同质量比的壳聚糖/葡甘聚糖混合膜对HPDLCs的增殖有促进作用,加载bFGF后,HPDLCs增殖得到增强,且壳聚糖/葡甘聚糖膜比3:0和1:2的混合膜还可增强ALP的活性水平。结论:壳聚糖/葡甘聚糖膜是一种潜在的牙周组织再生膜。     

温敏型壳聚糖/甘油磷酸钠缓释胰岛素凝胶系统的制备

目的:初步探讨制备温敏性壳聚糖/甘油磷酸钠载胰岛素凝胶系统.方法:应用壳聚糖与甘油磷酸钠制备具有温敏性的载有胰岛素的凝胶体系,从凝胶体系的pH值、粘度测定、胶凝时间研究不同配比、不同pH值对壳聚糖/甘油磷酸钠(CS/β-GP) 体系凝胶化性能的影响.结果:壳聚糖凝胶在室温(25℃)下呈液态,56% 甘油磷酸钠(β-GP)与3%壳聚糖(CS)在pH值6.8-7.2,37℃下凝胶化时间(GT) 从1h缩短到8-12min,凝胶形态良好,并且在负载胰岛素溶液后C/GP凝胶形态未受影响.结论:一定配比CS/GPS 体系在37℃具有快速凝胶化性能,在加入胰岛素后并未改变其温敏特性,为后续的温敏性壳聚糖/甘油磷酸钠载胰岛素凝胶系统的体外释药实验打下了基础.     

磁性阳离子高分子脂质体结合野生型p53投递系统的构建

目的用合成的磁性阳离子高分子脂质体与野生型p53(WTp53)基因质粒结合,构建靶向纳米载体投递系统,为进一步行静脉内皮细胞转染奠定基础。方法应用大肠杆菌重组质粒扩增法扩增WTp53质粒DNA,合成的磁性阳离子高分子脂质体为羧甲基壳聚糖十八烷基季铵盐/胆固醇(OQCMC/Chol)包覆油溶性Fe3O4的载体粒子。在不同的pH值及不同的WTp53质粒DNA和载体粒子质量比的条件下,将WTp53质粒DNA与OQCMC/Chol超微磁性载体粒子进行混合,并分别进行DNA结合实验和沉淀实验。观察电泳结果,并用SPSS 13.0统计软件进行数据分析。在DNaseⅠ模仿体内酶的环境下,将其加入制备好的超微载体投递系统中,电泳后观察投递系统对DNA的保护作用。应用透析系统模拟体内环境,观察超微投递系统载体与基因分离情况,并利用紫外分光光度计测量超微载体投递系统累积缓释曲线。结果应用透射电镜观察OQCMC/Chol超微磁性载体粒子和WTp53结合后的投递粒子。在pH 7条件下,WTp53质粒DNA和载体粒子质量比为1∶0.6,p53质粒DNA与OQCMC/Chol超微磁性载体粒子经过直接静电作用几乎100%结合,并证实该投递系统对DNA具有较好的保护作用。超微载体投递系统在7～12 h左右有明显的突释波峰,到第54小时以后,超微载体释放浓度变化缓慢,并可持续释放9 d以上。结论成功构建磁性阳离子高分子脂质体结合WTp53投递系统,将为进一步行细胞转染奠定可靠的基础,并为构建血管内靶向定位基因投递新方法和新技术提供可靠理论依据。     

Fabrication and characterization of Mg-doped chitosan–gelatin nanocompound coatings for titanium surface functionalization

Titanium and its alloys have been widely used in clinic and achieved great success. Due to the bio-inertness of titanium surface, challenges still exit in some compromised conditions. The present study aimed to functionalize titanium surface with magnesium (Mg)-doped chitosan/gelatin (CS/G) nanocompound coatings via electrophoretic deposition (EPD). CS/G coatings loaded with different amount of magnesium were successfully prepared on titanium substrate via EPD. Physicochemical characterization of the coatings confirmed that magnesium ions were loaded into the coatings in a dose-dependent manner. XRD results demonstrated that co-deposition of magnesium influenced the crystallinity of the coatings, and a new crystalline substance presented, namely hydrated basic magnesium carbonate. Mechanical tests showed improved tensile and shear bond strength of the magnesium-doped coatings, while the excessively high magnesium concentration could eventually decrease the bonding strength. Sustained release of magnesium ion was detected by ICP-OES within 28 days. TEM images also displayed that nanoparticles could be released from the coatings. In vitro cellular response assays demonstrated that the Mg-doped nanocompound coatings could enhance the proliferation and osteogenic differentiation of MC3T3-E1 cells compared to CS/G coatings. Therefore, it could be concluded that Mg-doped CS/G nanocompound coatings were successfully fabricated on titanium substrates via EPD. It would be a promising candidate to functionalize titanium surface with such organic–inorganic nanocompound coatings.     

In vitro release and In vivo biocompatibility studies of biomimetic multilayered alginate-chitosan/β-TCP scaffold for osteochondral tissue

Biomimetic three-layered monolithic scaffold (TLS) intended for the treatment of osteocondral defects was prepared by using alginate, chitosan and β-tricalcium phosphate (β-TCP) to study drug release behavior of the alternative drug delivery system and to investigate the therapeutic efficacy of the scaffold. Dexamethasone sodium phosphate (Dex) as a model drug was incorporated into the scaffold by solvent sorption method and in vitro release studies were conducted. In addition, the scaffold was implanted into the defects formed in the trochlea of Sprague–Dawley rats to assess the healing potential of the TLS on the osteochondral defect against reference Maioregen® comparatively. The release studies showed that after an initial burst at 3rd h, dexamethasone is released slowly during a 72-h period. In vivo studies indicated that the TLS has good tissue biocompatibility and biodegradation rate and showed better results during osteochondral healing process compared to the reference. All results demonstrated that the alginate-chitosan/β-TCP scaffold could be evaluated as a good candidate for osteochondral tissue applications.