有序排列聚甲基丙烯酸甲酯电纺纳米纤维作为大鼠原代雪旺细胞负载支架的可行性研究

目的:探讨有序聚甲基丙烯酸甲酯(polymethylmethacrylate,PMMA)电纺纳米纤维作为雪旺细胞负载支架的潜力。方法:构建有序PMMA电纺纳米纤维,以随机PMMA电纺纤维作为对照,纯化大鼠原代雪旺细胞并在纤维结构上进行培养,利用慢病毒技术转染绿色荧光蛋白基因作为显色手段,观察有序PMMA电纺纤维的拓扑线索在定向引导SCs生长上的作用,分析细胞对纤维结构的依从性;并在此基础上,持续动态观察雪旺细胞对有序电纺纤维的依从性,从而评价有序PMMA电纺纳米纤维作为神经损伤后植入性雪旺细胞负载支架的潜力。结果:雪旺细胞在随机及有序PMMA电纺纤维上均能顺利贴壁并生长;较之随机电纺纤维,雪旺细胞对有序纤维具有更好的依从性,能够受其接触引导形成和纤维走行方向一致的定向生长,并能够生成更长的细胞突起(P=0.0079);动态的观察则进而显示SCs对有序电纺纳米纤维的拓扑线索能维持稳定的依从性。结论:有序PMMA电纺纳米纤维具有作为神经损伤后植入性雪旺细胞负载支架的潜力。     

有序聚甲基丙烯酸甲酯电纺纳米纤维作为大鼠背根神经元负载支架的可行性研究

目的探讨具备有序或无序拓扑结构的聚甲基丙烯酸甲酯(polymethylmethacrylate,PMMA)电纺纳米纤维材料作为原代大鼠背根神经元(dorsal root ganglion neurons,DRGn)负载支架的可行性。方法构建具有随机或有序拓扑结构的PMMA电纺纳米纤维;利用PMMA薄膜组作为对照,分离纯化大鼠原代DRGn,利用慢病毒技术转染绿色荧光蛋白基因作为显色手段,分析在随机及有序纤维支架上DRGn神经突的生长能力以及神经突和电纺纤维的依存关系。结果大鼠原代DRGn能够顺利在PMMA材料上贴壁并生长,在培养第2天,较之平面环境,DRGn未见平均神经突数量及神经突长度的明显区别,电纺纤维的拓扑结构对于DRGn神经突的生长具有明显的接触引导作用,在有序电纺纤维上DRGn能够生成和基质纤维延伸方向一致的神经突;通过GFP表达与背景纤维的合成图,发现在随机或有序电纺纤维上,DRGn神经突均能循纤维向前生长,但相比随机纤维,神经突似乎更倾向于接受有序电纺纤维的引导。结论有序PMMA电纺纳米纤维具有作为神经损伤后大鼠DRGn负载支架的潜力。     

大鼠原代背根神经元和雪旺细胞在随机及有序排列聚甲基丙烯酸甲酯电纺纳米纤维上共培养的形态学

目的: 探讨聚甲基丙烯酸甲酯(PMMA)电纺纳米纤维支架的拓扑线索对于大鼠原代背根神经元(DRGn)培养及其与雪旺细胞(SCs)共培养的影响。 方法: 构建具有随机分布和轴向有序排列拓扑结构的PMMA电纺纳米纤维,分别为随机和有序PMMA电纺纳米纤维组,以PMMA薄膜作为对照组;分离纯化大鼠原代DRGn和SCs,与上述各组PMMA电纺纳米纤维共培养;利用慢病毒技术转染荧光蛋白基因作为显色方法,观察PMMA电纺纳米纤维的拓扑线索对于DRGn神经突生长的影响,在共培养实验通过荧光图像的快速傅立叶转换(FFT)及半高全宽值(FWHM)的计算,定量分析电纺纤维对DRGn神经突和SCs细胞突起的接触引导作用。 结果: 大鼠原代DRGn和SCs均能够顺利在PMMA材料上贴壁并生长;与PMMA薄膜组比较,随机和有序PMMA电纺纳米纤维组DRGn平均神经突数量及神经突长度差异无统计学意义(P>0.05);共培养实验中,电纺纤维的拓扑线索对于 DRGn神经突和SCs细胞突起的生长均具有明显的接触引导作用,与PMMA薄膜组和随机PMMA电纺纳米纤维组比较,有序PMMA电纺纳米纤维组DRGn和SCs的FWHM值均明显降低 (P<0.01),有序PMMA电纺纳米纤维能够在空间上促成DRGn神经突和SCs细胞突起建立共定位。 结论: 有序PMMA电纺纳米纤维具有作为脊髓损伤(SCI)后植入性支架材料的潜力,其拓扑线索有可能加速SCs的轴突髓鞘化过程。     

骨髓基质细胞在气电纺丝蛋白纤维支架上早期粘附的研究

目的检测大鼠骨髓基质细胞在气电纺蚕丝蛋白纳米纤维支架上的早期粘附状况,研究其基本生物学性能.方法将具有良好生物性能的蚕丝蛋白通过气流高压静电纺丝工艺制成纳米纤维,并且通过激光共聚焦显微镜对骨髓基质细胞在气电纺蚕丝蛋白纳米纤维支架表面的早期粘附进行观察.结果骨髓基质细胞对气电纺蚕丝蛋白无纺织物表现出良好的亲和性,细胞在材料表面4h即形成了稳定的粘附,并有部分细胞开始在材料表面铺展,铺展细胞数与粘附细胞数的比率为75.9%,而对照组仅为49.63%.结论气电纺蚕丝蛋白纳米纤维支架具有良好的亲和性,提示其作为骨组织工程材料的可能性.     

聚甲基丙烯酸甲酯静电纺丝纳米纤维对大鼠原代星形胶质细胞生长的影响

目的探讨聚甲基丙烯酸甲酯（polymethylmethacrylate, PMMA）电纺纳米纤维的拓扑线索对于大鼠原代星形胶质细胞生
长能力及方式的影响,为脊髓损伤后植入性细胞支架的构建提供前期基础。方法构建具有随机或有序拓扑结构的PMMA电
纺纳米纤维;分离并纯化大鼠原代星形胶质细胞;利用PMMA薄膜作为对照,利用慢病毒技术转染绿色荧光蛋白基因作为显色
手段,分析在星形胶质细胞不同拓扑结构纤维支架上的生长特点。结果随机及有序PMMA电纺纤维均能支持星形胶质细胞
的生长,其拓扑结构能够显著影响星形胶质细胞的生长方式,在有序纤维系统上细胞突起的生长方向能够和基质纤维的延伸方
向保持高度一致;通过绿色荧光蛋白和基质纤维的合成图,发现在两种拓扑结构的纤维系统上,细胞突起均能依附在纤维上向
远处延伸;较之PMMA薄膜,在有序纤维上的星形胶质细胞能生成更长的细胞突起（P<0.01）,而在随机纤维上的细胞则形成更
短的突起（P<0.01）。结论PMMA电纺纳米纤维的拓扑结构能够显著影响大鼠原代星形胶质细胞生长能力及方式,其作为植
入性支架具有减轻脊髓损伤后损伤灶胶质瘢痕形成的潜在价值。
     

聚己内酯电纺纤维支架材料对骨髓基质细胞增殖及分化的影响

目的:观察聚己内酯(PCL)电纺纤维支架材料对骨髓基质细胞(BMSCs)增殖及分化特点的影响,评价其作为骨组织工程的支架材料的应用前景.方法:将兔BMSCs与PCL电纺纤维支架材料在培养板内共培养.采用形态学观察、MTT法及ALP检测等方法检测BMSCs在PCL电纺纤维表面的粘附、增殖和分化能力.结果:体外培养的BMSCs能在PCL电纺纤维表面正常粘附和增殖,并且表达较高的碱性磷酸酶活性.结论:PCL电纺纤维适合作为支架材料应用于BMSCs为种子细胞的组织构建.     

聚己内酯-碳酸亚乙酯［Poly(CL-EC)］和血管内皮生长因子(VEGF)静电混纺支架的构建及其生物学性能

 目的  以聚己内酯-碳酸亚乙酯［Poly(CL-EC)］共聚物混合血管内皮生长因子(vascular endothelial growth factor,VEGF),采用静电纺的方法构建纳米支架并检测其生物学性能。方法  按照EC/CL共聚物比例为1∶9、1∶6、1∶4,Poly(CL-EC)浓度分别为5%、10%、15%电纺纤维膜,分析电纺纤维膜的表征和力学性能。然后将VEGF按照0 ng/g、10 ng/g、100 ng/g、1 μg/g的质量比与Poly (CL-EC)溶液混合,电纺制备纳米支架。对混纺膜进行细胞增殖和黏附试验、乳酸脱氢酶(lactate dehydrogenase,LDH)释放试验、间接溶血试验和皮下植入试验等检测。结果  根据Poly (CL-EC)电纺纤维膜的表征和力学性能,我们选用EC/CL比例为1∶6的10% Poly(CL-EC)与VEGF构建混合电纺膜。细胞增殖和黏附试验证实Poly(CL-EC)/VEGF电纺膜具有良好的细胞相容性,尤其是血管内皮细胞;LDH释放试验、接触溶血试验和体内植入试验显示该材料无细胞毒性、有较好的血液相容性和很低的异物反应。结论  静电纺构建的Poly(CL-EC)/VEGF具有良好的生物学性能,能够作为组织工程支架材料。     

聚己内酯静电纺丝纳米纤维基复合材料在口腔医学中的应用研究进展

静电纺丝可制备出模拟细胞外基质的生物支架材料，以聚己内酯（poly-caprolactone，PCL）作为静电纺丝原料可获得良好的生物相容性，而将PCL电纺纳米纤维与无机材料组合，可以改善支架材料亲水性和机械性能，调控降解性能，增强生物矿化能力，具有良好的应用前景。文章对PCL静电纺丝纳米纤维基复合材料在口腔医学中的应用进行综述。     

卟啉化聚酰亚胺电纺纤维膜用于微量甲醇蒸气的快速检测

目的:制备一种卟啉化聚酰亚胺电纺纤维膜用于微量甲醇蒸气的快速检测.方法:通过化学共聚将卟啉引入聚酰亚胺大分子主链;采用静电纺丝技术制备卟啉化聚酰亚胺电纺纤维膜,通过改变溶液组成、纺丝电压等,调控纳米纤维微结构,从而获得具有微／纳立体结构和大比表面积的电纺纤维膜,应用于微量甲醇蒸气的快速检测.结果:制备的卟啉化聚酰亚胺电纺纤维膜直径分布均匀、形貌良好,且保持了卟啉的基本光谱特性.当甲醇蒸气与此电纺纤维膜作用时,可引起其紫外光谱的红移与荧光强度的减弱,而其它一些常见醇类并无此现象.交替通入150 ppm的甲醇蒸气与氮气后,电纺纤维膜的荧光强度几乎没有改变,显示出良好的可重复使用性.结论:基于卟啉优异的光敏性能和纳米纤维高比表面积的特点,设计并制备了卟啉化聚酰亚胺电纺纤维膜,用于微量甲醇蒸气的检测.该电纺纤维膜具有灵敏度高,选择性好,并可重复使用等优点.     

同轴电纺制备刚性多糖纳米纤维膜

将壳聚糖、海藻酸钠或透明质酸配制成水溶液,聚氧化乙烯(PEO)溶解在二甲基甲酰胺(DMF)/H2O 混合溶剂中,以上述两溶液分别作为内、外纺丝液进行同轴电纺制备成纤维膜,进而利用适当溶剂除去 PEO 外壳,得到纯多糖纳米纤维膜.纤维结构通过 TEM、SEM 进行表征.结果表明:同轴电纺可一步制得外壳为 PEO、内核为刚性多糖的核壳纳米纤维,纤维外壳 PEO 组分可以用氯仿萃取除去;与单轴电纺法制得的刚性多糖纤维相比,同轴电纺可以保持最终纤维的结构完整性.     

电纺PLLA/PVP纳米纤维膜材料用于血管组织工程的前期研究

目的:研究静电纺丝在血管组织工程中的应用。方法:利用静电纺丝技术制备不同质量分数的聚乳酸(PLLA)/聚乙烯吡咯烷酮(PVP)电纺丝膜,并通过差示扫描量热分析(DSC)、扫描电子显微镜(SEM)观察、接触角测试(CA)和力学性能分析进行材料学表征。在不同的电纺丝膜上种植血管平滑肌细胞(VSMCs),并分别在2、4、6 d取样进行SEM、激光共聚焦显微镜(LCMS)观察,以检测VSMCs在材料上铺展的形貌和分布情况。经MTT测试,以了解材料对VSMCs增殖的影响,用血小板黏附实验测试材料的血液相容性。结果:混入PVP的PLLA电纺丝膜具有良好的表面结构,亲水性显著提高,且随着PVP含量的增多PLLA电纺丝膜的亲水性增高,但对力学性能影响不明显。细胞实验结果表明:混有PVP的PLLA电纺丝膜利于VSMCs的黏附生长,具有良好的细胞相容性和血液相容性。结论:成功地将PVP和PLLA进行了静电纺丝,该材料具有良好的细胞相容性和血液相容性,其在血管组织工程应用中具有广阔的前景。     

不同比例聚乳酸-聚己内酯/泊洛沙姆静电纺纳米材料作为皮肤支架的可能性

 目的 探讨不同比例聚乳酸-聚己内酯/泊洛沙姆［poly (ε-caprolactone-co-lactide) /Poloxamer,PLCL/Poloxamer］静电纺材料作为皮肤组织支架的可能性。方法 静电纺丝技术制备纯PLCL及PLCL/Poloxamer比例75/25、85/15、90/10、95/5的纳米纤维材料,测定支架表面形态、机械以及亲水性能。将脂肪干细胞接种至支架表面,第1、3、7、10天行水溶性四唑盐(CCK 8)分析,并于第3天扫描电镜观察细胞形态。结果 90/10、95/5比例支架的断裂强度高于纯PLCL及75/25、85/15比例支架(分别为9.31±0.29,9.27±0.50,7.08±0.21,7.46±0.27和7.47±0.21,MPa),90/10、85/15、75/25比例的水接触角低于纯PLCL及95/5比例(分别为0° ,0°,0°,131.5°±8.9°和7.8°±2.7°)。在第1、3、7、10天时,PLCL/Poloxamer材料脂肪干细胞黏附及增殖能力明显高于纯PLCL。扫描电镜显示90/10比例PLCL/Poloxamer支架上有大量脂肪干细胞生长,细胞形态完整、表面光滑。结论 90/10比例PLCL/Poloxamer支架的机械性能及亲水性能高于其他组,该材料与大鼠脂肪干细胞的相容性良好,是更适合脂肪干细胞黏附及增殖的组织工程材料。     

电纺纳米银/碳纳米纤维敷料的制备及抗菌效果评价

目的：利用静电纺丝技术制备纳米纤维敷料,并研究其抗菌效果。方法：采用聚丙烯腈(PAN)为前驱体高分子,利用静电纺丝技术及高温碳化方法制备碳纳米纤维敷料,并通过银镜反应制备纳米银/碳纳米纤维复合敷料。将2种样本与庆大霉素纸片及优拓（urgotul）敷料采用Kirby-Baucer法进行体外抑菌实验。结果：成功制备出具有纳米级支架结构且负载纳米银的的创面敷料。体外抑菌实验结果显示,纳米银/碳纳米纤维复合敷料和优拓对金黄色葡萄球菌、铜绿假单胞菌和大肠埃希菌标准菌株均有较好的抑菌效果;纳米银/碳纳米纤维复合敷料对于金黄色葡萄球菌及铜绿假单胞菌抑菌效果优于优拓,但对大肠杆菌的抑制效果低于优拓;碳纳米纤维未显示出具有抑菌能力。结论：纳米银/碳纳米纤维复合敷料具有纳米级支架结构,是具有较好抗菌能力的烧伤创面敷料。     

Effect of IKVAV peptide nanofiber on proliferation,adhesion and differentiation into neurocytes of bone marrow stromal cells

This study examined the effect of IKVAV peptide nanofiber on proliferation, adhesion and differentiation into neurocytes of bone marrow stromal cells （BMSCs）. IKVAV Peptide-amphiphile was synthesized and purified. Then, hydrogen chloride was added to the diluted aqueous solutions of PA to induce spontaneous formation of nanofiber in vitro. The resultant samples was observed tmder transmission electron microscope. BMSCs were cultured with IKVAV peptide nanofiber. The effect of IKVAV nanofiber on the proliferation, adhesion and induction differentiation of BMSCs was observed by inverted microscopy, calcein-AM/PI staining, cell counting and immunofluorescence staining. The results demonstrated that IKVAV peptide-amphiphile could self-assemble to form nanofiber gel. BMSCs cultured in combination with IKVAV peptide nanofiber gel grew well and the percentage of live cells was over 90%. IKVAV peptide nanofiber gel exerted no influence on the proliferation of BMSCs and could promote the adhesion of BMSCs and raise the ra- tio of neurons when BMSCs were induced to differentiate into neurocytes. It is concluded that BMSCs could proliferate and adhere well and yield more neurons during when induced to differente into neurocytes on IKVAV peptide nanofiber gel.     

Biocompatibility evaluation of electrospun aligned poly(propylene carbonate) nanofibrous scaffolds with peripheral nerve tissues and cells in vitro

Background  Peripheral nerve regeneration across large gaps is clinically challenging. Scaffold design plays a pivotal role in nerve tissue engineering. Recently, nanofibrous scaffolds have proven a suitable environment for cell attachment and proliferation due to similarities of their physical properties to natural extracellular matrix. Poly(propylene carbonate) (PPC) nanofibrous scaffolds have been investigated for vascular tissue engineering. However, no reports exist of PPC nanofibrous scaffolds for nerve tissue engineering. This study aimed to evaluate the potential role of aligned and random PPC nanofibrous scaffolds as substrates for peripheral nerve tissue and cells in nerve tissue engineering.

Methods  Aligned and random PPC nanofibrous scaffolds were fabricated by electrospinning and their chemical characterization were carried out using scanning electron microscopy (SEM). Dorsal root ganglia (DRG) from Sprague-Dawley rats were cultured on the nanofibrous substrates for 7 days. Neurite outgrowth and Schwann-cell migration from DRG were observed and quantified using immunocytochemistry and SEM. Schwann cells derived from rat sciatic nerves were cultured in electrospun PPC scaffold-extract fluid for 24, 48, 72 hours and 7 days. The viability of Schwann cells was evaluated by 3-[4,5-dimethyl(thiazol-2-yl)-2,5-diphenyl] tetrazolium bromide (MTT) assay.

Results  The diameter of aligned and random fibers ranged between 800 nm and 1200 nm, and the thickness of the films was approximately 10–20 μm. Quantification of aligned fiber films revealed approximately 90% alignment of all fibers along the longitudinal axis. However, with random fiber films, the alignment of fibers was random through all angle bins. Rat DRG explants were grown on PPC nanofiber films for up to 1 week. On the aligned fiber films, the majority of neurite outgrowth and Schwann cell migration from the DRG extended unidirectionally, parallel to the aligned fibers. However, on the random fiber films, neurite outgrowth and Schwann cell migration were randomly distributed. A comparison of cumulative neurite lengths from cultured DRGs indicated that neurites grew faster on aligned PPC films ((2537.6±987.3) μm) than randomly-distributed fibers ((493.5±50.6) μm). The average distance of Schwann cell migration on aligned PPC nanofibrous films ((2803.5±943.6) μm) were significantly greater than those on random fibers ((625.3±47.8) μm). The viability of Schwann cells cultured in aligned PPC scaffold extract fluid was not significantly different from that in the plain DMEM/F12 medium at all time points after seeding.

Conclusions  The aligned PPC nanofibrous film, but not the randomly-oriented fibers, significantly enhanced peripheral nerve regeneration in vitro, indicating the substantial role of topographical cues in stimulating endogenous nerve repair mechanisms. Aligned PPC nanofibrous scaffolds may be a promising biomaterial for nerve regeneration.

     

功能化自组装多肽水凝胶与嗅鞘细胞生物相容性研究

Olfactory ensheathing cell (OEC) transplantation is a promising or potential therapy for spinal cord injury (SCI). However, the effects of injecting OECs directly into SCI site have been limited and unsatisfied due to the complexity of SCI. To improve the outcome, proper biomaterials are thought to be helpful since these materials would allow the cells to grow three-dimensionally and guide cell migration. In this study, we made a new peptide hydrogel scaffold which named GRGDSPmx by mixing the pure RADA16 and designer-peptide RADA16-GRGDSP solution, and we examined the molecular integration of the mixed nanofiber scaffolds using Atomic force microscopy (AFM). In addition, we have studied the behavior of OECs in GRGDSPmx condition as well as on RADA16 scaffold by analyzing their phenotypes such as cell proliferation, apoptosis, survival and morphology. The experimental results showed that GRGDSPmx could be self-assembled to form a hydrogel. Inverted optical microscope and Scanning electron microscope (SEM) analysis showed that OECs are viable and they proliferate within the nanostructured environment of the scaffold. MTT assay demonstrated that OECs proliferation rate was increased on GRGDSPmx scaffold compared to the pure RADA16 scaffold. In addition, OECs on GRGDSPmx scaffolds also showed less apoptosis and maintained the original spindle-shape morphology. Calcein-AM/PI fluorescence staining revealed that OECs cultured on GRGDSPmx grew well and the viable cell count was 95%. These results suggested that this new hydrogel scaffold provided an ideal substrate for OEC 3D culture and suggested its further application for SCI repair.     

Compatibility of olfactory ensheathing cells with functionalized self-assembling peptide scaffold in vitro

Background Olfactory ensheathing cell （OEC） transplantation is a promising or potential therapy for spinal cord injury （SCI）. However, the effects of injecting OECs directly into SCI site have been limited and unsatisfied due to the complexity of SCI. To improve the outcome, proper biomaterials are thought to be helpful since these materials would allow the cells to grow three-dimensionally and guide cell miqration.     

聚丙烯基纳米碳纤维复合材料结晶行为研究

采用示差扫描量热法(DSC)研究了纳米碳纤维对PP／CNF复合材料中PP结晶行为的影响。结果表明：纳米碳纤雏可提高PP的结晶温度，但略降低其结晶速率和结晶度。提高纳米碳纤雏含量或减小纳米碳纤雏直径，PP结晶行为的上述变化更为明显，但结晶速率随纳米碳纤维含量出现先减小后增大的趋势。     

多次交替浸渍法构筑簇状ZnO的PAN复合纳米纤维膜及其光催化性能

以聚丙烯腈（PAN）与氯化锌（ZnCl₂）作为前驱物,采用静电纺丝工艺制备PAN/ZnCl₂复合纳米纤维膜,分别采用多次冷热交替浸渍法和单次冷热静置浸渍法得到簇状PAN/ZnO-1和PAN/ZnO-2复合纳米纤维膜。利用扫描电镜（SEM）、傅里叶红外光谱（FT-IR）、X射线衍射（XRD）、X射线能量色散光谱（XPS）和热重分析仪（TG）对复合纳米纤维膜的表面形貌和微结构进行了表征,并以亚甲基蓝（MB）为污染物模型,评价其光催化降解性能。结果表明：经冷热交替浸渍后,纳米ZnO粒子均匀地附着在PAN纤维表面,尤其在PAN/ZnO-1复合纳米纤维膜表面还出现了花状ZnO粒子;相比单次冷热静置浸渍法处理的PAN/ZnO-2复合纳米纤维膜,经多次冷热交替浸渍的PAN/ZnO-1复合纳米纤维膜循环使用3次后对MB的降解率仍可达到90%以上,具有更好的光催化活性和循环使用性能。同时,MB溶液的初始质量浓度、催化剂用量和染料溶液的pH等因素对样品的的光催化降解率有一定影响。