聚乳酸多孔支架制备及细胞实验

以冰粒子作为致孔剂，采用冷冻干燥-粒子滤出复合法制备了块状聚乳酸多孔支架。将聚乳酸溶于氯仿溶液后加入冰粒子，在液氮中冷冻后冷冻干燥获得多孔支架。对支架孔隙结构分析表明，该工艺制备的多孔支架无致孔剂残留，其孔隙大小由加入的冰粒子大小决定。细胞实验表明该多孔支架具有较好的生物相容性并且无细胞毒性。     

Enzyme-degradable phosphorylcholine porous hydrogels cross-linked with polyphosphoesters for cell matrices

Biodegradable highly porous hydrogels composed of poly [2-methacryloyloxyethyl phosphorylcholine (MPC)] cross-linked with polyphosphoesters have been prepared as novel cellular matrices. Well-controlled porous hydrogels were fabricated by using potassium hydrogen carbonate as a porogen salt for forming gas. This process enabled the homogeneous expansion of pores within the polymer hydrogel matrices, leading to well-interconnected high porosity. The mechanical properties of the hydrogels were influenced by the cross-linking density and porous structure. Hydrolysis and enzymatic digestion of the hydrogels were determined under basic conditions. The cross-linking density and porosity influenced the rate of degradation of the hydrogels. Acceleration of the degradation with alkaline phosphatase was also observed. Cultivation of mouse osteoblastic cell (MC3T3-E1) was performed in the highly porous hydrogels and cell viability was well maintained. The rate of cell proliferation also was relatively increased with an increase in the amount of polyphosphoesters in the hydrogel. Basic fibroblast growth factor (bFGF) was physically absorbed by the hydrogels and effectively induced cell proliferation. In conclusion, the porous hydrogels prepared in this study contributed a suitable environment for three-dimensional cell cultivation and may be useful for cell and tissue matrices.     

Preparation of interconnected highly porous polymeric structures by a replication and freeze-drying process

Three-dimensional degradable porous polymeric structures with high porosities (93-98%) and well-interconnected pore networks have been prepared by freeze-drying polymer solutions in the presence of a leachable template followed by leaching of the template. Templates of the pore network were prepared by fusing sugar or salt particles to form a well-connected structure. The interstices of the template were then filled with a polymer solution (5-15% w/v) in 1,4-dioxane, followed by freeze-drying of the solvent. Subsequent leaching of the sugar template ensures the connectivity of the pore network. The scaffold architecture consists of relatively large interconnected pores modeled after the template and smaller pores resulting from the freeze-drying process. The total porosity of the resultant porous structures is determined by the interstitial space of the leachable template and by the polymer concentration in the freeze-drying solution. The freezing temperature also has an effect on the final morphology of the porous structures. Compared with freeze-drying and combination of freeze-drying /particulate leaching techniques, this method facilitates higher interconnectivity of the scaffolds. Porous structures have been prepared from several relevant polymers in the biomedical and tissue-engineering field: poly(D,L-lactide) (PDLLA), 1000PEOT70PBT30, a segmented poly(ether ester) based on polyethylene oxide and polybutylene terephthalate, and poly(epsilon-caprolactone) (PCL). The mechanical properties of the porous structures prepared by this technique depend on the nature of the polymer, porosity, and the freezing temperature. With porosities in the range of 95-97%, the compression moduli of scaffolds prepared from the different polymers could be varied between 13.0 and 301.5 kPa.     

Formation of defined microporous 3D structures starting from cross-linked hydrogels

A new and simple technique was developed to obtain polysaccharide (hyaluronane, alginate and carboxymethylcellulose) -based hydrogels with a defined porous morphology. The technique consists of stratifying a cross-linked hydrogel on a filter with known pore diameter. CO(2) bubbles, produced by the addition of HCl to a porogen salt NaHCO(3), are forced to pass through the filter, and they induce the hydrogel to assume a porous morphology. The presence and distribution of pores was confirmed by scanning-electron microscopy (SEM). A strict correspondence was found between the porosity of the filter and the pore diameter in the hydrogels. Water uptake measurements showed a decreased amount of water taken up by the porous hydrogels compared with the native hydrogels, due to a compacting of the material. An explanation of the porous material properties of Hyal hydrogel was given on the basis of FTIR spectra.     

Pore structure analysis of swollen dextran-methacrylate hydrogels by SEM and mercury intrusion porosimetry

The objectives of this study were to visually examine the surface and interior morphology of a new class of biodegradable hydrogels based on dextran-methacrylate and to quantify the porous structure of these hydrogels in swollen state. Two techniques (scanning electron microscopy and mercury intrusion porosimetry) were used to analyze pore structure of dextran-methacrylate hydrogels as a function of the degree of methacrylate substitution. SEM was used to observe 3-dimensional network structure of cryofixed and fractured swollen dextran-methacrylate hydrogels. Image analysis of the SEM data revealed different pore shapes and sizes, depending on the degree of substitution and the location within the hydrogel. A higher methacrylate-substituted dextran hydrogel showed a compact and rigid pore structure, while a lower substituted hydrogel showed a delicate and fragile pore structure. Mercury intrusion porosimetry analysis of the pore structure of dextran-methacrylate hydrogels provided useful and more reliable quantitative data of pore characteristics, such as total pore area, average pore diameter, their distribution, and bulk density. The total pore area and average pore diameter of swollen dextran-methacrylate hydrogels decreased with an increase in degree of methacrylate substitution, while bulk density increased with an increase in degree of substitution.     

Micro-structured smart hydrogels with enhanced protein loading and release efficiency

A series of temperature-responsive poly(N-isopropylacrylamide) (PNIPAAm) hydrogels with highly porous microstructures were successfully prepared by using hydrophobic polydimethylsiloxane (PDMS) and sodium dodecyl sulfate as liquid template and stabilizer, respectively. These newly prepared hydrogels possess highly porous structures. In contrast to the conventional PNIPAAm hydrogel, the swelling ratios of the porous gels at room temperature were higher, and their response rates were significantly faster as the temperature was raised above the lower critical solution temperature. For example, the novel hydrogel prepared with 40% PDMS template lost over 95% water within 5 min, while the conventional PNIPAAm gel only lost approximately 14% water in the same time. The improved properties are achieved due to the presence of liquid PDMS templates in the reaction solutions, which lead to the formation of porous structures during the polymerization/crosslinking. Lysozyme and bovine serum albumin (BSA) as protein models were for the first time loaded into these micro-structured smart hydrogels through a physical absorption method. The experimental results show that the loading efficiency of BSA with a higher molecular weight is lower than that of lysozyme due to the size exclusion effect, and the loading efficiencies of both proteins in the porous hydrogel are much higher than those in the conventional PNIPAAm hydrogel. For example, the loading efficiency of BSA in porous hydrogel is 0.114, approximately 200% higher than that in conventional hydrogel (0.035). Both lysozyme and BSA were completely released from the porous hydrogel at 22 °C. Furthermore, the release kinetics of the proteins from the porous hydrogel could be modulated by tuning the environmental temperature. These newly prepared porous materials provide an avenue to increase the loading efficiency and to control the release patterns of macromolecular drugs from hydrogels, and show great promise for application in protein or gene delivery.     

Synthesis and characterization of dextran-methacrylate hydrogels and structural study by SEM

The objectives of this study were to develop a simple and reproducible method for the preparation of the hydrogel precursor dextran-methacrylate and to conduct a visual observation of the interior structure of the swollen dextran-methacrylate hydrogel with minimum artifacts. A dextran-methacrylate hydrogel precursor was synthesized by reacting dextran with methacrylic anhydride in the presence of triethylamine as a catalyst. The effects of reaction time, temperature, concentration, and catalyst amount were studied to obtain a wide range of degree of substitution (DS) in dextran by methacrylate. The dextran-methacrylate synthesized showed an enhanced solubility in water and common organic solvents. UV irradiation of dextran-methacrylate by a long-wave UV lamp (365 nm) generated a photocrosslinked hydrogel. This dextran-methacrylate hydrogel showed a range of swelling ratio from 67 to 227% and exhibited an increase in swelling ratio with a decrease in methacrylate substitution. The pH of the swelling media did not affect the swelling behavior of the dextran-methacrylate hydrogels at all the degrees of substitution used. Special cryofixation and cryofracturing techniques were used to prepare aqueous swollen dextran-methacrylate hydrogel samples for SEM observation of their surface and interior structures. A unique three-dimensional porous structure was observed in the swollen hydrogel but was absent in the unswollen hydrogel. Different pore sizes and morphologies between the surface and the interior of swollen hydrogels also were observed.     

Swelling and mechanical properties of glycol chitosan/poly(vinyl alcohol) IPN-type superporous hydrogels

Glycol chitosan/poly(vinyl alcohol) interpenetrating polymer network type superporous hydrogels were prepared using a gas foaming/freeze-drying method. The effect of the molecular weight of the strengthener, poly(vinyl alcohol) (PVA), on the swelling and mechanical behavior of the superporous hydrogels was investigated. The introduction of a small amount of high molecular weight PVA significantly enhanced the mechanical strength but slightly reduced the swelling capacity. The freezing/thawing (F/T) drying process had a significant effect on the physical properties of the glycol chitosan/PVA superporous hydrogels, because hydrogen bonds were formed between the PVA molecules as a result of the number of F/T cycles. The swelling ratio decreased but the mechanical strength increased with increasing freezing time. However, this effect was not as strong as the number of F/T cycles. The differential scanning calorimetry was used to examine how the thermal behavior associated with the hydrogen bond-induced crystalline structure was affected by the F/T process.     

Morphology of poly(methacrylic acid)/poly(N-isopropyl acrylamide) interpenetrating polymeric networks

The morphology of interpenetrating polymeric networks (IPNs) composed of the temperature-sensitive poly(N-isopropyl acrylamide) (PNIPAAm) and the pH-sensitive poly(methacrylic acid) (PMAA) were investigated by scanning electron microscopy (SEM). The IPN hydrogels were prepared by a sequential UV polymerization method. SEM studies were conducted on IPN hydrogel samples dried by different methods, and the influence on the IPN structure was discussed. The environmental conditions induced morphological changes for these dual sensitive IPN hydrogels which were studied by cryogenic SEM, when the gels were analyzed in their wet state. The results showed that the porous size in the IPN was strongly influenced by the environmental pH and temperature. Decrease in pH and increase in temperature resulted in significant pore size decrease for the swollen IPNs hydrogels.     

Structure and properties of bilayer chitosan-gelatin scaffolds

Chitosan-gelatin hybrid polymer network scaffolds were prepared via the freeze-drying technique by using the ice microparticle as a porogen. Monolayer and bilayer scaffolds were obtained by using different pre-freezing methods. The novel bilayer scaffolds were prepared via contact with -56 degrees C lyophilizing plate directly, then lyophilized. The properties of chitosan-gelatin scaffolds, such as microstructure, physical and mechanical and degradable properties, were studied. These results suggested that the porosity and pore size of the scaffolds could be modulated with thermodynamic and kinetic parameters of ice formation. The scaffolds prepared from chitosan and gelatin can be utilized as a promising matrix for tissue engineering.     

Biodegradable nanocomposite hydrogel structures with enhanced mechanical properties prepared by photo-crosslinking solutions of poly(trimethylene carbonate)–poly(ethylene glycol)–poly(trimethylene carbonate) macromonomers and nanoclay particles

Soft hydrogels with elasticity modulus values lower than 100 kPa that are tough and biodegradable are of great interest in medicine and in tissue engineering applications. We have developed a series of soft hydrogel structures from different methacrylate-functionalized triblock copolymers of poly(ethylene glycol) (PEG) with poly(trimethylene carbonate) (PTMC) by photo-crosslinking aqueous solutions of the macromonomers in 2.5 and 5 wt.% colloidal dispersions of clay nanoparticles (Laponite XLG). The length of the PTMC blocks of the macromonomers and the clay content determined the physicomechanical properties of the obtained hydrogels. While an increase in the PTMC block length in the macromonomers from 0.2 to 5 kg/mol resulted in a decrease in the gel content, the addition of 5 wt.% Laponite nanoclay to the crosslinking solution lead to very high gel contents of the hydrogels of more than 95%. The effect of PTMC block length on the mechanical properties of the hydrogels was not as pronounced, and soft gels with a compressive modulus of less than 15 kPa and toughness values of 25 kJ m^?3 were obtained. However, the addition of 5 wt.% Laponite nanoclay to the formulations considerably increased the compressive modulus and resilience of the hydrogels; swollen nanocomposite networks with compressive modulus and toughness values of up to 67 kPa and 200 kJ m^?3, respectively, could then be obtained. The prepared hydrogels were shown to be enzymatically degradable by cholesterol esterase and by the action of macrophages. With an increase in PTMC block length in the hydrogels, the rates of mass loss increased, while the incorporated Laponite nanoclay suppressed degradation. Nanocomposite hydrogel structures with a designed gyroid pore network architecture were prepared by stereolithography. Furthermore, in the swollen state the porous gyroid structures were mechanically stable and the pore network remained fully open and interconnected.     

Morphology of poly(methacrylic acid)/poly(N-isopropyl acrylamide) interpenetrating polymeric networks

The morphology of interpenetrating polymeric networks (IPNs) composed of the temperature-sensitive poly(N-isopropyl acrylamide) (PNIPAAm) and the pH-sensitive poly(methacrylic acid) (PMAA) were investigated by scanning electron microscopy (SEM). The IPN hydrogels were prepared by a sequential UV polymerization method. SEM studies were conducted on IPN hydrogel samples dried by different methods, and the influence on the IPN structure was discussed. The environmental conditions induced morphological changes for these dual sensitive IPN hydrogels which were studied by cryogenic SEM, when the gels were analyzed in their wet state. The results showed that the porous size in the IPN was strongly influenced by the environmental pH and temperature. Decrease in pH and increase in temperature resulted in significant pore size decrease for the swollen IPNs hydrogels.     

Fabrication of porous chitosan scaffolds for soft tissue engineering using dense gas CO2

The aim of this study was to investigate the feasibility of fabricating porous crosslinked chitosan hydrogels in an aqueous phase using dense gas CO(2) as a foaming agent. Highly porous chitosan hydrogels were formed by using glutaraldehyde and genipin as crosslinkers. The method developed here eliminates the formation of a skin layer, and does not require the use of surfactants or other toxic reagents to generate porosity. The chitosan hydrogel scaffolds had an average pore diameter of 30-40 μm. The operating pressure had a negligible effect on the pore characteristics of chitosan hydrogels. Temperature, reaction period, type of biopolymer and crosslinker had a significant impact on the pore size and characteristics of the hydrogel produced by dense gas CO(2). Scanning electron microscopy and histological analysis confirmed that the resulting porous structures allowed fibroblasts seeded on these scaffolds to proliferate into the three-dimensional (3-D) structure of these chitosan hydrogels. Live/dead staining and MTS analysis demonstrated that fibroblast cells proliferated over 7 days. The fabricated hydrogels exhibited comparable mechanical strength and swelling ratio and are potentially useful for soft tissue engineering applications such as skin and cartilage regeneration.     

Iodine-responsive poly (HEMA–PVP) hydrogel for self-regulating burst-free extended release

Swollen hydrogels with extended iodine release kinetics is highly desirable for burn and scald treatment. In this paper semi-interpenetrating poly (HEMA–PVP) hydrogels were prepared by radical polymerization followed by thermo-treatment to crosslink its PVP component. Incorporation of PVP component endows the hydrogels responsive to loaded iodine undergoing a reversible shrunken/swollen volume transition. This resulted in a self-regulating iodine release model, in which shrunken hydrogel at high iodine loading decreased drug diffusion thereby reducing burst release, and then gradually swollen hydrogel as drug release ensures rapid release of dissociated drug from strong affinity sites on hydrogel backbone, achieving a burst-free extended release. The hydrogels demonstrated 11.5-fold higher iodine loading than pure pHEMA hydrogel and showed a highest 40% volume shrink. Initial burst release of iodine was efficiently decreased from 12,894 μg/day of pure pHEMA hydrogel to 2570 μg/day of pHEMA/PVP hydrogel with 37% PVP content. Iodine-loaded hydrogels showed zero-order release at three time periods of 0–15 h, 15 h–3.5 days and 3.5–23.5 days corresponding to release rate of 2570, 776 and 493 μg/day. The work gained a new insight into swollen hydrogel drug delivery system with burst-free extended drug release kinetics.     

Nanocellulose/PEGDA aerogel scaffolds with tunable modulus prepared by stereolithography for three-dimensional cell culture

Three-dimensional (3D) porous scaffolds made of biopolymers have attracted significant attention in tissue engineering applications. In this study, cellulose-nanofibers/polyethylene glycol diacrylate (CNFs/PEGDA) mixture, a novelty 3D material, was prepared by physical mixing the CNFs with a waterborne photopolymerizable acrylic resin (PEGDA). Then the CNFs/PEGDA mixture was used to fabricate 3D cytocompatibility CNFs/PEGDA hydrogel scaffold by stereolithograph（SLA）process. The CNFs/PEGDA hydrogels were shaped by SLA, and then the aerogel scaffolds were prepared by the freeze-drying of hydrogels. The results showed that the CNFs/PEGDA mixtures with different CNFs contents are all transparent, homogeneous and with obvious shear-thinning property. The SLA fabricated CNFs/PEGDA aerogel scaffolds possess high and tunable compressive modulus and high porosity of approximately 90%. It is found that CNFs in the composite scaffolds played a significant role in structural shape integrity, porous structure and mechanical strength. In addition, the NIH 3T3 cells tightly adhere on the CNFs/PEGDA materials and spread on the scaffolds with good differentiation and viability. These results have revealed a superior method to prepare tissue engineering scaffolds which possesses suitable mechanical strength and biocompatibility for 3D cell cultivation.     

Superporous hydrogels for cartilage repair: Evaluation of the morphological and mechanical properties

Despite extensive research in the design of biomaterials for articular cartilage repair, there remains a need for the development of materials with the mechanical compliance to function synergistically with healthy cartilage, but porous enough to allow for tissue integration. In this study, superporous hydrogels of poly(vinyl alcohol) and poly(vinyl pyrrolidone) were prepared using a novel technique consisting of a double emulsion process. The hydrogel emulsions were physically cross-linked by freeze-thaw cycling. The hydrogels had a high degree of porosity, determined using environmental scanning electron microscopy, a technique superior to any method that involves dehydrating the samples. Increasing the volume of organic solvent increased porosity, due to cross-linking of the hydrogel solution around the droplets in the emulsion, leaving pores where the organic solvent was present. Poly(lactic-co-glyclic acid) microparticles formed and were embedded in the matrix. The mechanical properties, measured in confined creep and in unconfined, uniaxial compression, were similar to native articular cartilage. The permeability of the samples was unaffected by changing solvent content, despite changes in porosity. These materials are good candidates for tissue engineering of cartilage because they can mimic mature cartilage mechanically while providing a porous matrix through which cells can migrate and proliferate.     

Preparation of porous hydrolyzable polyrotaxane hydrogels and their erosion behavior

We have prepared porous polyrotaxane hydrogels by using the salt leaching technique. Porous hydrogels were found to have a uniform and highly porous structure. The size of pores in each hydrogel was directly proportional to the size of the sodium chloride particle used. Structural uniformity of the hydrogels is useful not only for uniform cell distribution, but also for well-controlled material properties. Uniform pore size and distribution may ensure the diffusion of nutrients throughout of the gel and the removal of metabolic wastes from the system. The results of an erosion study in phosphate-buffered saline showed that the erosion time of porous polyrotaxane hydrogels was controlled by the poly(ethylene glycol) (PEG) content in the hydrogels. The erosion time of the porous polyrotaxane hydrogel was observed to be almost the same with the non-porous polyrotaxane hydrogel with the same PEG content. From the erosion study, the erosion time of the polyrotaxane hydrogel may be independent of its morphology. Easy control of the erosion time in the polyrotaxane hydrogels is useful in the preparation of scaffolds for tissue engineering.     

Preparation of porous hydrolyzable polyrotaxane hydrogels and their erosion behavior

—We have prepared porous polyrotaxane hydrogels by using the salt leaching technique. Porous hydrogels were found to have a uniform and highly porous structure. The size of pores in each hydrogel was directly proportional to the size of the sodium chloride particle used. Structural uniformity of the hydrogels is useful not only for uniform cell distribution, but also for wellcontrolled material properties. Uniform pore size and distribution may ensure the diffusion of nutrients throughout of the gel and the removal of metabolic wastes from the system. The results of an erosion study in phosphate-buffered saline showed that the erosion time of porous polyrotaxane hydrogels was controlled by the poly(ethylene glycol) (PEG) content in the hydrogels. The erosion time of the porous polyrotaxane hydrogel was observed to be almost the same with the non-porous polyrotaxane hydrogel with the same PEG content. From the erosion study, the erosion time of the polyrotaxane hydrogel may be independent of its morphology. Easy control of the erosion time in the polyrotaxane hydrogels is useful in the preparation of scaffolds for tissue engineering.     

基因重组蛛丝蛋白-聚乙烯醇复合支架材料的制备

目的探讨致孔剂NaCl粒径和比例、变性剂和聚乙烯醇(PVA)等因素对基因重组蛛丝蛋白-PVA复合支架材料形态及性能的影响.方法基因重组蛛丝蛋白溶解于98%甲酸,采用冷冻干燥粒子滤沥法制备重组蛛丝蛋白-PVA复合多孔支架;采用扫描电子显微镜观察支架的形态;采用单纤维强力试验机测试支架机械性能.结果乙醇作变性剂制得的多孔支架力学性能较好,支架的断裂应力、断裂比强度均提高5倍以上,断裂伸长率可达12.21%.以粒径<500μm的NaCl为致孔剂制得的多孔支架力学性能较好.高分子材料PVA能明显改善重组蛛丝蛋白多孔支架的件能.结论重组蛛丝蛋白-PVA复合支架材料有望在组织工程领域得以应用.     

In Situ Gelable Glycation-Resistant Hydrogels Composed of Gelatin and Oxidized Alginate

An in situ gelable glycation-resistant hydrogel has been prepared from oxidized alginate (Oalg) and gelatin. Aminoguanidine, an effective inhibitor of the glycation reaction, was first encapsulated in gelatin microspheres followed by incorporation into the hydrogel. The gelation process was monitored rheologically, and the results showed that the AMG-loaded Oalg/gelatin system solidified quickly at body temperature. Moreover, the hydrogels were highly porous, and the AMG-loaded microspheres dispersed in the hydrogels remained intact. Hydrogels' AMG loadings did not appear to change their degradation behaviors. AMG could be released from the hydrogels in a sustainable manner for a relatively short duration. Incorporation of AMG into the hydrogels resulted in imparting a glycation-resistant capability. Lastly, long-term in vitro incubation of all hydrogel formulations with fibroblasts did not reveal any cytotoxic potential.