丝素蛋白/柠檬酸/乙二醇膜的性能研究

用丝素蛋白、柠檬酸和乙二醇制备了一种新型丝素蛋白膜(简称SFCE膜),研究了其力学性能、膜中丝素蛋白的聚集态及对水的pH值的影响．     

CaCO3-丝素蛋白膜的动态粘弹谱

制备了一种含有碳酸钙(CaCO3)的丝素蛋白膜(简称CaFM膜)。结合X射线衍射方法,通过研究其动态粘弹谱,研究了CaCO3的掺入对CaFM膜中丝素蛋白分子聚集态结构的影响。     

丝素蛋白膜上vWf抗体的固定化及其体外抗凝血性能

用凝血因子的抗体对生物材料进行表面改性以提高其抗凝血性能。以丝素蛋白膜为基质，利用等离子体处理辅助的共价交联方法对vWf因子（von Wilebrand factor)抗体进行了固定化。用酶联免疫法和抗体过剩法对固定化效果进行了评价，固定化抗体的活性采用体外凝血时间（APPT，TT和PT）测定进行检测。结果显示，通过这种方法可以有效地将vWf抗体固定化，丝素蛋白膜固定化vWf抗体后，其抗凝血性能有了一定的改善。本研究结果拓宽了抗体固定化技术的应用范围，同时为抗凝血材料的设计提供了一种新的思路。     

柞蚕丝素/壳聚糖共混膜的结构及细胞相容性

用碳化二亚胺(EDC)作为交联剂,流延法制备柞蚕丝素(ASF)/壳聚糖(CS)共混膜。用扫描电镜(SEM)、红外光谱(FT-IR)、热重分析(TG)和四甲基偶氮唑盐比色法(MTT)对膜的结构及细胞相容性进行了研究。结果显示,柞蚕丝素和壳聚糖具有较好的相容性和较强的相互作用,壳聚糖能阻碍共混膜内的丝素蛋白形成β-折叠构象。EDC能分别与柞蚕丝素及壳聚糖反应,从而对共混膜进行有效的交联,并使膜的热稳定性提高。与ASF膜或CS膜相比,共混比为80/20和40/60的ASF/CS共混膜更有利于细胞生长和增殖,作为一种新型的生物材料具有良好的研究和开发应用前景。     

丝素蛋白含量对纳米羟基磷灰石仿生矿化和体外细胞相容性的影响

通过制备含有不同丝素蛋白(SF)含量的丝素蛋白/纳米羟基磷灰石复合材料, 着重研究丝素蛋白含量对纳米羟基磷灰石(n-HA)仿生矿化过程中晶体成核与生长及体外细胞相容性的影响. 结果表明: SF的加入对n-HA晶体的成核和生长具有明显的调控作用; 丝素含量的大小对n-HA晶体成核和生长没有明显的区别, 但对晶粒在有机大分子中的聚集状态有明显的影响: 当SF含量不超过20wt%时, n-HA晶粒呈现放射状团簇, 当SF含量超过20wt%时, n-HA晶粒无序团聚. 体外细胞相容性结果显示, SF的加入可以促进材料与细胞的界面亲和性, 但SF含量对这种亲和性的影响不明显, 20wt%和30wt%的SF含量对增殖能力具有较强的促进作用.     

抗凝血丝素蛋白材料的研究进展

综述了作为抗凝血材料的丝素蛋白的改性方法,重点讨论了用s-羧甲基还原角蛋白、硫酸、二氧化硫和vWf(Von Willebrand fator)因子的抗体等对丝素蛋白的改性,并分析了改性材料的抗凝血机理和抗凝血性能.     

聚乙二醇丙三醇对丝素蛋白膜的改性

以乙二醇和环氧氯丙烷为原料,合成了一种新的高分子化合物:聚乙二醇丙三醇(PEGG),用红外光谱和核磁共振对其进行了表征。用流延法制备了丝素蛋白/PEGG共混膜,用X射线衍射法,红外光谱法对共混膜的结构进行测试分析,结果表明,随着PEGG含量的增加,丝素蛋白(SF)的结晶度有所下降,PEGG与丝素蛋白之间有氢键结合,因此可能抑制了s ilkⅠ结晶。测试了共混膜的力学性能,结果表明,共混膜的断裂伸长率、弹性随着PEGG含量的增加而增大,共混膜的断裂强度均高于24M Pa,而杨氏模量都小于80 M Pa。SF/PEGG共混膜具有良好的柔软性和弹性。     

再生丝素蛋白肽/聚乙烯醇膜材料性能的实验研究

采用不同配比的再生丝素蛋白肽(SFP)与聚乙烯醇(PVA)水溶液共混的方法制备了再生丝素蛋白肽/聚乙烯醇(SFP/PVA)膜材料,对SFP/PVA膜材料进行了红外表征。通过电子拉力试验机进行了不同配比SFP/PVA膜材料的力学性能试验,结果显示配比为30/70的膜最大承受力为10.57N,拉伸强度为35.2MPa,力学性能最佳。膜材料降解性实验在人工体液中进行,结果显示其降解从SFP开始,随后与SFP结合部分的PVA在接触到水介质后逐步溶失。MTT法对HepG2细胞的促生长试验结果表明,各配比SFP/PVA膜材料均具有良好的促HepG2细胞快速黏附和生长性。综合各性能指标,配比为30/70的SFP/PVA膜具有作为组织工程支架载体的应用前景。     

后处理对静电纺丝素纤维膜性能的影响

静电纺丝素纤维膜在载药、组织工程及多种电池的研究中具有广泛的应用.静电纺丝素纤维膜在制备过程中,需要后处理以保持纤维的形状及提升纤维膜的性能.本研究以乙醇和甘油对静电纺丝素纤维膜进行后处理,并通过SEM、FTIR、XRD、TGA等表征手段系统研究了不同后处理条件对静电纺丝素纤维膜的结构及性能的影响,并分析及探讨了其影响机制.结果表明:仅用75％(体积分数)乙醇后处理可使丝素蛋白分子β折叠含量增大,纤维膜断裂强度显著增强;仅用甘油后处理可使丝素蛋白分子链无规卷曲度提高,纤维膜的断裂伸长率显著提高,亲水性大大增强;而用75％乙醇结合不同浓度的甘油对静电纺丝素纤维膜进行后处理可调控其力学性能,使其更能满足在不同领域的应用需求.     

纳米羟基磷灰石丝素蛋白仿生矿化材料的制备研究

以Ca(NO₃)₂与Na₃PO₄为无机相的前驱体,将丝素蛋白直接溶于Ca(NO₃)₂溶液中,不经过脱盐处理,直接滴入Na₃PO₄溶液中反应,在37℃下丝素蛋白和羟基磷灰石晶体之间相互作用,仿生合成了纳米羟基磷灰石(n-HA)丝素蛋白(SF)生物矿化材料.用FTIR、XRD、XPS和SEM进行表征.结果表明,羟基磷灰石和丝素蛋白两相间具有较强的化学键合,矿化材料中无机相包含少量碳酸根,为缺钙类骨羟基磷灰石并且呈现一定的长轴取向性,说明丝素蛋白大分子对羟基磷灰石晶体的成核和生长起着模板和调控作用.矿化物颗粒尺寸在50~200nm之间,其抗压强度为32.21MPa,可作为非承重部位骨组织缺损修复材料.     

Bovine pericardium coated with biopolymeric films as an alternative to prevent calcification: In vitro calcification and cytotoxicity results

Bovine pericardium, for cardiac valve fabrication, was coated with either chitosan or silk fibroin film. In vitro calcification tests of coated and non coated bovine pericardium were performed in simulated body fluid solution in order to investigate potential alternatives to minimize calcification on implanted heart valves. Complementary, morphology was assessed by scanning electron microscopy — SEM; X-ray diffraction (XRD) and infrared spectroscopy (FTIR-ATR) were performed for structural characterization of coatings and biocompatibility of chitosan. Silk fibroin films were assayed by in vitro cytotoxicity and endothelial cell growth tests. Bovine pericardium coated with silk fibroin or chitosan did not present calcification during in vitro calcification tests, indicating that these biopolymeric coatings do not induce bovine pericardium calcification. Chitosan and silk fibroin films were characterized as non cytotoxic and silk fibroin films presented high affinity to endothelial cells. The results indicate that bovine pericardium coated with silk fibroin is a potential candidate for cardiac valve fabrication, since the affinity of silk fibroin to endothelial cells can be explored to induce the tissue endothelization and therefore, increase valve durability by increasing their mechanical resistance and protecting them against calcification.     

Preparation and properties of a chitosan-based carrier of corneal endothelial cells

A novel chitosan-based membrane that was made of hydroxypropyl chitosan, gelatin and chondroitin sulfate was used as a carrier of corneal endothelial cells. The characteristics of the blend membrane, such as transparency, equilibrium water content, permeability, mechanical properties, protein absorption ability, hydrophilicity and surface morphology, were determined. To study the effects of the membrane on cell attachment and growth, rabbit corneal endothelial cells were cultured on this artificial membrane. The biodegradability and biocompatibility of the blend membrane were in vivo evaluated by its implantation into the muscle of the rats. Glucose permeation results demonstrated that the blend membrane had higher glucose permeability than natural human cornea. Scanning electron microscopy (SEM) analysis of the membranes demonstrated that no fibrils were observed. As a result, the optical transparency of the membrane was as good as the natural human cornea. The average value of tensile strength of the membrane was 13.71 MPa for dry membrane and 1.48 MPa for wet membrane. The value of elongation at break of the wet was 45.64%. The cultured rabbit corneal endothelial cells formed a monolayer on the blend membrane which demonstrated that the membrane was suitable for corneal endothelial cells to attach and grow. In addition, the membranes in vivo showed a good bioabsorption property. The mild symptoms of inflammation at sites of treatment could be resolved as the implant was absorbed by the host. The results of this study demonstrated that the hydroxypropyl chitosan-chondroitin sulfate-gelatin blend membrane can potentially be used as a carrier for corneal endothelial cell transplantation.     

生物材料细胞粘附肽仿生修饰的研究进展

RGD(Arg-G1y-Asp)短肽序列是一种细胞粘附肽,能被细胞膜上的整合素识别,参与细胞与基质间的粘附.为改善合成生物材料缺乏细胞识别信号的缺点,可将含RGD序列肽经本体或表面修饰引入材料,使材料具有良好的细胞亲和性.综述了采用含RGD肽对各种合成生物材料进行仿生修饰的研究进展.     

Optimization of the interaction between ethylenevinyl alcohol copolymers and human endothelial cells

To better understand and optimize the fine interactions that occur during adhesion events between human cells and synthetic materials, we seeded human umbilical vein endothelial cells (HUVEC) onto ethylene-vinyl alcohol (EVOH) copolymer films prepared by casting. Different adhesive proteins, e.g. fibronectin and gelatin, and the monoclonal antibody (MoAb) CLB-HEC19 specific for the endothelial cell membrane were used to coat the materials. We used atomic force microscopy (AFM) to analyse the EVOH film structure, to test its planarity and homogeneity, before seeding it with endothelial cells. The metabolic changes induced in the endothelial cells by interactions with the copolymer functional groups and the adhesive proteins were monitored by a micro-electronic pH sensor, positioned close to the HUVEC monolayer. We found that the adhesion of HUVEC onto various substrates was finely modulated by the MoAb CLB-HEC19 and that the endothelial cell metabolic rate was enhanced when cultured onto a CLB-HEC19 coating. Surface roughness seems also to play a role in the interaction with HUVEC. The AFM measurement analysis demonstrated that L6 surface is rougher than R20. These surface characteristics could favour cell adhesion; in fact HUVEC adhesion results on R20 were significantly lower than on L6.     

Three-dimensional mesoporous gold film to enhance the sensitivity of electrochemical detection

Cell-cell and cell-extracellular matrix (ECM) adhesion are fundamental and important in the development of a cell-based chip. In this study, a novel, simple, rapid, and one-step technique was developed for the fabrication of a uniform three-dimensional mesoporous gold thin film (MPGF) onto a gold (Au) coated glass plate based on an electrochemical deposition method. Scanning electron microscopy images demonstrated that the resulting MPGF electrode had uniformly distributed pores with diameters of about 20 nm. The cyclic voltammetric behavior of [Fe(CN)(6)](4-/3-) coupled onto MPGF and Au electrodes demonstrated that the MPGF electrode had a higher electrocatalytic sensitivity and reversibility than the bare Au electrode. The Arg-Gly-Asp (RGD) sequence containing the peptide was immobilized on the MPGF and bare Au substrates. HeLa cancer cells were then cultured on the RGD peptide layer. The successful immobilization of the peptide and cells was confirmed by atomic force microscopy. The cell proliferation and viability were evaluated by cyclic voltammetry and Trypan blue dyeing assay. These results indicated that the RGD/MPGF modified electrodes showed an electrochemical sensitivity in the detection of cancer cells which is approximately three times higher, especially at low cell density, than RGD/Au electrodes. This much improved sensitivity of the MPGF modified electrode demonstrates the potential for the fabrication of a highly sensitive and low-cost cell-based chip for rapid cancer detection.     

Synthesis of selenium nanoparticles in the presence of silk fibroin

This paper describes a solution-phase approach to the synthesis of selenium nanoparticles by reducing selenious acid solution with ascorbic acid in the presence of silk fibroin. The monodispersed spherical selenium colloid particles obtained were very stable in silk fibroin solution and characterized by Atomic force microscopy and X-ray techniques. The influences of temperature and ultrasonication on the morphology of selenium nanoparticles were also discussed. The experiments showed that the selenium nanoparticles with various morphologies could be obtained under different temperatures and the appropriate ultrasonication time was 60 min. This result indicated that the silk fibroin molecules intimately associated with the surface of the selenium particles and controlled the growth particles.     

Tailoring RGD local surface density at the nanoscale toward adult stem cell chondrogenic commitment

Arginine-glycine-aspartic acid (RGD) dendrimer-based nanopatterns on poly(L-lactic acid) were used as bioactive substrates to evaluate the impact of the RGD local surface density on the chondrogenic induction of adult human mesenchymal stem cells.During chondrogenic commitment,active extracellular matrix (ECM) remodeling takes place,playing an instructive role in the differentiation process.Although three-dimensional environments such as pellet or micromass cultures are commonly used for in vitro chondrogenic differentiation,these cultures are rather limited with respect to their ability to interrogate cells in cell-ECM interactions.In the present study,the nanopatterns of the tunable RGD surface density were obtained as a function of the initial dendrimer concentration.The local RGD surface density was quantified through probability contour plots for the minimum interparticle distance,constructed from the corresponding atomic force microscopy images,and correlated with the cell adhesion and differentiation response.The results revealed that the local RGD surface density at the nanoscale acts as a regulator of chondrogenic commitment,and that intermediate adhesiveness of cells to the substrates favors mesenchymal cell condensation and early chondrogenic differentiation.     

角膜内皮细胞载体壳聚糖基共混膜的制备及性质研究

为探讨壳聚糖基共混膜作为组织工程化角膜内皮细胞载体的可行性,制备了羟乙基壳聚糖/明胶/硫酸软骨素共混膜,并评价其性质。结果表明该膜片具有良好的透明度,适宜的含水量;细胞毒性0～1级;体外培养的角膜内皮细胞能够很好地贴附和生长于共混膜上;植入大鼠肌肉内,能够稳定地降解,诱发的炎症反应明显低于不可降解手术缝合线,说明该膜片具有良好的组织相容性,有望作为角膜细胞载体构建组织工程化角膜。     

Fabrication and characters of a corneal endothelial cells scaffold based on chitosan

A novel chitosan-based membrane that made of hydroxyethyl chitosan, gelatin and chondroitin sulfate was used as a carrier of corneal endothelial cells. The characteristics of the blend membrane including transparency, equilibrium water content, ion and glucose permeability were determined. The results showed that the optical transparency of the membrane was as good as the natural human cornea. The water content of this scaffold was 81.32% which was remarkably close to the native cornea. The membrane had a good ion permeability and its glucose permeability was even higher than natural human cornea. The cultured rabbit corneal endothelial cells formed a monolayer on the membrane. The results demonstrated that the membrane was suitable for corneal endothelial cells to attach and grow on it. In addition, the membranes in vivo could be degraded steadily with less inflammation and showed a good histocompatibility. These results demonstrated that the hydroxyethyl chitosan-chondroitin sulfate-gelatin blend membrane can potentially be used as a carrier for corneal endothelial cell transplantation.