Injectable Nanohybrid Scaffold for Biopharmaceuticals Delivery and Soft Tissue Engineering

An injectable nanofibrous hydrogel scaffold integrated with growth factors (GFs) loaded polysaccharide nanoparticles was developed that specifically allows for targeted adipose‐derived stem cells (ASCs) encapsulation and soft tissue engineering. The nanofibrous hydrogel was produced via biological conjugation of biotin‐terminated star‐shaped poly(ethylene glycol) (PEG‐Biotin) and streptavidin‐functionalized hyaluronic acid (HA‐Streptavidin). The polysaccharide nanoparticles were noncovalently assembled via electrostatic interactions between low‐molecular‐weight heparin (LMWH) and N,N,N‐trimethylchitosan chloride (TMC). Vascular endothelial growth factor (VEGF) was entrapped in the LMWH/TMC nanoparticles by affinity interactions with LMWH.     

Biofunctionalization of titanium with PEG and anti-CD34 for hemocompatibility and stimulated endothelialization

Thrombosis and restenosis are the main causes of failure of cardiovascular and other blood-contacting biomedical devices. It is recognized that rapid endothelialization is a promising method for preventing these complications. Convincing evidence in vivo has further emerged that the vascular homing of endothelial progenitor cells (EPCs) contributes to rapid endothelial regeneration. This study deals with improving the hemocompatibility and enhancing EPC colonization of titanium by covalently bonding PEG(600) or PEG(4000), then end-grafting of an anti-CD34 antibody. For this, a chemically hydroxylated titanium surface was aminosilanized, which was further used for covalent grafting of polyethylene glycol and the antibody. The grafting efficiency was verified in each step. In vitro platelet adhesion analysis confirmed superior hemocompatibility of the modified surface over the control. Affinity of EPC to the surface and inhibition of smooth muscle cell adhesion, two prerequisites for endothelialization, are demonstrated in in vitro cell culture. While the coating selectively stimulates EPC adhesion, its antifouling properties prevent formation of an extracellular matrix and proliferation of the cells. Additional affinity for matrix proteins in the coating is considered for further studies. Potent inhibitory effect on macrophage activation and the relative stability of the coating render this technique applicable.     

Effect of radiation on structure and properties of material made from oxpericardium

In order to reduce calcification of artificial cardiac bio-valve-OX-pericardium valve and prolong its life, it is very important to develop a new method for treatment of OX-pericardium valve. As a new processing method OX-pericardium valves were treated by glutaraldehyde (GA) then sterilized by γ-ray with ⁶⁰Co-source. The effects of radiation on the structure and properties of pericardium valve material were studies in terms of stress at one dimensional stretch i. e. strain response, tensile intensity, stress relaxation curve and variation of contraction temperature. The results show that the OX-pericardium valve treated with 0.1% glutaraldehyde (GA) and irradiated with 25 kGy γ-rays is not only sterile but also the mechanical property of which is similar to that of the fresh OX-pericardium and the presetting energy similar to that of OX-pericardium valve treated with 0.625% glutaraldehyde. These improve OX-pericardium valves so as to prevent them from calcification.     

Tissue engineering of heart valves by recellularization of glutaraldehyde-fixed porcine valves using bone marrow-derived cells

To increase the biocompatibility and durability of glutaraldehyde (GA)-fixed valves, a biological coating with viable endothelial cells (ECs) has been proposed. However, stable EC layers have not been formed successfully on GA-fixed valves due to their inability to repopulate. In this study, to improve cellular adhesion and proliferation, the GA-fixed prostheses were detoxified by treatment with citric acid to remove free aldehyde groups. Canine bone marrow mononuclear cells (MNCs) were differentiated into EC-like cells and myofibroblast-like cells in vitro. Detoxified prostheses were seeded and recellularized with differentiated bone marrow- derived cells (BMCs) for seven days. Untreated GA-fixed prostheses were used as controls. Cell attachment, proliferation, metabolic activity, and viability were investigated and cell-seeded leaflets were histologically analyzed. On detoxified GA-fixed prostheses, BMC seeding resulted in uninhibited cell proliferation after seven days. In contrast, on untreated GA-fixed prostheses, cell attachment was poor and no viable cells were observed. Positive staining for smooth muscle a-actin, CD31, and proliferating cell nuclear antigen was observed on the luminal side of the detoxified valve leaflets, indicating differentiation and proliferation of the seeded BMCs. These results demonstrate that the treatment of GA-fixed valves with citric acid established a surface more suitable for cellular attachment and proliferation. Engineering heart valves by seeding detoxified GA-fixed biological valve prostheses with BMCs may increase biocompatibility and durability of the prostheses. This method could be utilized as a new approach for the restoration of heart valve structure and function in the treatment of end-stage heart valve disease.     

Development of injectable high molecular weight hyaluronic acid hydrogels for cartilage regeneration

The study focuses on developing hyaluronic acid (1200 kilo Dalton) hydrogels for cartilage regeneration. In spite of being highly biocompatible; a large amount of water absorption and easily degrading nature restricts the use of hyaluronic acid in the field of tissue regeneration. This can be rectified by crosslinking hyaluronic acid with a crosslinking agent such as divinyl sulfone; which results in a biocompatible hydrogel with superior rheological properties. Different amounts of divinyl sulfone have been used for crosslinking hyaluronic acid to get three types of hydrogels with differing properties. Swelling studies, rheology analysis, enzymatic degradation and scanning electron microscopic analysis were conducted on all the different types of hydrogels prepared. Viscoelastic properties of the hydrogel were analyzed so that a hydrogel with better elastic property and stability is obtained. Scanning electron microscopy was used to study the morphology of the HA hydrogels. The cytotoxicity testing was conducted to prove the non-toxic nature of the hydrogels and cell culture studies using adipose mesenchymal stem cells showed better adhesion and proliferation properties in all the three hydrogels. Thus hyaluronic acid hydrogel makes a promising material for cartilage regeneration.     

Biofunctionalized Electrospun PCL‐PIBMD/SF Vascular Grafts with PEG and Cell‐Adhesive Peptides for Endothelialization

Artificial small‐caliber vascular grafts are still limited in clinical application because of thrombosis, restenosis, and occlusion. Herein, a small‐caliber vascular graft (diameter 2 mm) is fabricated from poly(ε‐caprolactone)‐b‐poly(isobutyl‐morpholine‐2,5‐dione) (PCL‐PIBMD) and silk fibroin (SF) by electrospinning technology and then biofunctionalized with low‐fouling poly(ethylene glycol) (PEG) and two cell‐adhesive peptide sequences (CREDVW and CAGW) with the purpose of enhancing antithrombogenic activity and endothelialization. The successful grafting of PEG and peptide sequences is confirmed by X‐ray photoelectron spectroscopy. The suitable surface wettability of the modified vascular graft is testified by water contact angle analysis. The surface hemocompatibility is verified by platelet adhesion assays and protein adsorption assays, and the results demonstrate that both platelet adhesion and protein adsorption on the biofunctionalized surface are significantly reduced. In vitro studies demonstrate that the biofunctionalized surface with suitable hydrophilicity and cell‐adhesive peptides can selectively promote the adhesion, spreading, and proliferation of human umbilical vein endothelial cells. More importantly, compared with control groups, this biofunctionalized small‐caliber vascular graft shows high long‐term patency and endothelialization after 10 weeks of implantation. The biofunctionalization with PEG and two cell‐adhesive peptide sequences is an effective method to improve the endothelialization and long‐term performance of synthetic vascular grafts.     

含悬挂羧基手臂聚乳酸材料的内皮细胞黏附及其细胞活性

为了考察内皮化材料表面的细胞活性, 在前期工作的基础上, 分别在聚乳酸(PLA)、乳酸-苹果酸共聚物(PLMA), 以及含悬挂羟基或羧基的乳酸-苹果酸共聚物膜(PLMAHE,PLMACA)表面种植人脐静脉内皮细胞(HUVEC), 成功地制备了内皮化表面. 通过测定内皮化材料表面内皮细胞释放的内皮型一氧化氮合酶(eNOS)以及一氧化氮的释放量, 间接考察了内皮细胞的抗凝血活性; 另外, 通过内皮化表面的血小板黏附实验, 直接观察了血小板在内皮细胞上的黏附情况. 实验结果表明, 含羧基材料表面的内皮细胞活性比PLA和PLMAHE的高; 相对其它材料PLMACA能更有效地保留黏附于其表面内皮细胞的活性, 其单位内皮细胞的eNOS以及NO的释放量分别为(41.8±8.1) μmol/104 cells和(0.76±0.16) U/104 cells. 电镜照片(SEM)显示, 各种材料表面的内皮细胞均能有效地减少血小板的黏附与聚集; 在内皮细胞脱落的区域, PLMACA仍能较好地实现其抑制血小板黏附的功能, 有望成为新型血管修复(替代)材料.     

人工细胞外基质对血管内皮细胞生存的影响

通过物理吸附方法, 利用胶原、 聚赖氨酸和融合蛋白VEGF-Fc对聚苯乙烯培养板表面进行改性, 以研究细胞外基质材料对血管内皮细胞的影响. 结果表明, 3种蛋白显著提高了聚苯乙烯表面的亲水性. 内皮细胞的黏附、 增殖、 细胞骨架蛋白染色和血管性血友病因子(vWF)免疫染色实验结果表明, 胶原、 聚赖氨酸和VEGF-Fc基质均能有效提高血管内皮细胞的黏附, 其中胶原可与VEGF协同作用促进内皮细胞分化表型的表达; VEGF-Fc基质兼具了VEGF的生物学活性, 可促进内皮细胞的黏附和增殖以及vWF功能性蛋白的表达. 本研究为诱导材料表面内皮化和血管新生的生物活性材料的设计开发提供了新思路.     

Biocompatible hydrogel for cartilage repair with adjustable properties

Synthesis of hydrogel at mild conditions is considered one most important challenge, especially if the hydrogel will be used for hosting bioactive materials or drugs. The procedure of hydrogel preparation should have no effect on the properties of the hosted materials. Hyaluronic acid (HA) was modified by adding dialdehyde groups to its structure to facilitate formation of hydrogel at very mild conditions. Dialdehyde HA (DHA) was prepared through oxidation of HA using sodium metaperiodate as oxidizing agent. The prepared DHA was characterized by Fourier‐transform infrared (FTIR) spectroscopy and X‐ray diffraction (XRD) and aldehyde content. A hydrogel was prepared using different chitosan/DHA molar ratio and fixed amount of glutaraldehyde at 25°C. The prepared hydrogel has tunable properties and pores size depending on the chitosan/DHA molar ratio. Sodium diclofenac was loaded on the hydrogel as a model drug. The hydrogel was characterized by FTIR spectroscopy, swelling rate, gel fraction, drug release profile, and cytotoxicity. The results obtained indicated that the properties of the prepared hydrogel, including gelling time, gel fraction, swelling, pores size, and drug release profile are highly tuned depending on the chitosan/DHA molar ratio. The drug loading efficiency was in the range of 70% to 85%. The cytotoxicity results reveal that the prepared hydrogel has a very low toxicity in presence and absence of sodium diclofenac.     

Development of a Dual‐Functional Hydrogel Using RGD and Anti‐VEGF Aptamer

Synthetic molecular libraries hold great potential to advance the biomaterial development. However, little effort is made to integrate molecules with molecular recognition abilities selected from different libraries into a single biomolecular material. The purpose of this work is to incorporate peptides and nucleic acid aptamers into a porous hydrogel to develop a dual‐functional biomaterial. The data show that an anti‐integrin peptide can promote the attachment and growth of endothelial cells in a 3D porous poly(ethylene glycol) hydrogel and an antivascular endothelial growth factor aptamer can sequester and release VEGF of high bioactivity. Importantly, the dual‐functional porous hydrogel enhances the growth and survival of endothelial cells. This work demonstrates that molecules selected from different synthetic libraries can be integrated into one system for the development of novel biomaterials.     

Effect of hyaluronic acid encapsulation in a silica hydrogel matrix on drug penetration through the skin

It has been found that the encapsulation of high molecular weight hyaluronic acid in a biologically relevant silica hydrogel matrix provides its accelerated penetration into the skin compared to free acid. The developed hybrid hydrogels, in which high molecular weight hyaluronic acid retains its pronounced anti-inflammatory properties and strong hydrating effect, can become the basis for new, more effective soft formlations for the treatment of inflammatory skin diseases, as well as for products used in the beauty industry. It has been shown that the penetration of hyaluronic acid from the hybrid hydrogels depends on the conditions of their synthesis, the average molecular weight and the loading of the acid.     

Stimuli‐Responsive Biomolecule‐Based Hydrogels and Their Applications

This Review presents polysaccharides, oligosaccharides, nucleic acids, peptides, and proteins as functional stimuli‐responsive polymer scaffolds that yield hydrogels with controlled stiffness. Different physical or chemical triggers can be used to structurally reconfigure the crosslinking units and control the stiffness of the hydrogels. The integration of stimuli‐responsive supramolecular complexes and stimuli‐responsive biomolecular units as crosslinkers leads to hybrid hydrogels undergoing reversible triggered transitions across different stiffness states. Different applications of stimuli‐responsive biomolecule‐based hydrogels are discussed. The assembly of stimuli‐responsive biomolecule‐based hydrogel films on surfaces and their applications are discussed. The coating of drug‐loaded nanoparticles with stimuli‐responsive hydrogels for controlled drug release is also presented.     

Exosome‐Modified Tissue Engineered Blood Vessel for Endothelial Progenitor Cell Capture and Targeted siRNA Delivery

Instability and poor targeting causes the long‐term patency of RNA‐modified tissue engineering blood vessels (TEBVs) remaining unsatisfactory. RNA can be enriched in exosome and then delivered into targeted cells while whether exosome‐modified TEBVs achieve RNA targeted delivery is unclear. Here, to promote the expression of klotho protein on the mesenchymal stem cell (MSC)‐derived exosomes, klotho plasmids are first transfected into MSCs, and adenosine kinase (ADK) siRNA is then loaded into exosome (klotho/ADK siRNA‐exosome) using electrotransfection. Flow chamber results show that klotho/ADK siRNA‐exosome can effectively capture circulating endothelial progenitor cells (EPCs). Besides, the captured EPCs can endocytose this exosome, and then decompose it into klotho protein and ADK siRNA. Moreover, ADK siRNA promotes the paracrine of proangiogenic factors and adenosine from EPCs, which further facilitate proliferation and migration of endothelial cells. Based on polyethyleneimine‐capped gold nanoparticles, exosome‐modified TEBVs are constructed through layer‐by‐layer assembly. Animal experimental results show that klotho/ADK siRNA‐exosome‐modified TEBVs can maintain the patency up to one month, and good endothelialization is observed. In short, one exosome‐modified TEBV is constructed, capture molecules on the surface of exosome capture the circulating EPCs, and the loaded RNA achieves its purpose of accurate treatment depending on the needs of patients.     

Phospholipid Polymer Hydrogel Matrices with Dually Immobilized Cytokines for Accelerating Secretion of the Extracellular Matrix by Encapsulated Cells

Construction of 3D tissues by various types of cells with specific characteristics is an important and fundamental technology in tissue reconstruction medicine and animal‐free diagnosis system. To do so, an excellent extracellular matrix (ECM) is needed for encapsulation of cells and maintaining cell activity. Spontaneously forming hydrogel matrix is used by complexation between two water‐soluble polymers, 2‐methacryloyloxyethyl phosphorylcholine polymer bearing phenylboronic acid groups and poly(vinyl alcohol). Two cytokines for cell proliferation are immobilized in the hydrogel matrix to control the activities of the encapsulated cells. The cytokine‐immobilized hydrogel matrix can encapsulate both L929 fibroblasts and normal human dermal fibroblasts under mild condition. The physical properties of the hydrogel matrix can follow the proliferation process of the encapsulated cells. The encapsulated cells secrete ECM in the polymer hydrogel networks upon 3D culturing for 7 days. Consequently, the tissue‐mimicking ECM hybrid hydrogels are fabricated successfully.     

Surface Modification of Multiple Bioactive Peptides to Improve Endothelialization of Vascular Grafts

Endothelialization is an effective approach to prevent thrombus formation and enhance vascular graft survival. Surface modification of biomolecules has been proved to be effective in regulating endothelial cell behaviors. In this study, several peptides including YIGSR, RGD, and REDV sequences are covalently immobilized on the surface of electrospun silk fibroin scaffolds and the effects of combined application of these peptides on cell behaviors are studied. The results show that, compared with the scaffolds modified with single peptides, the scaffolds modified with dual peptides (YIGSR+RGD) could significantly enhance the proliferation of human umbilical vein endothelial cells (HUVECs). However, the combination of REDV+RGD or YIGSR+REDV does not promote the adhesion or proliferation of HUVECs. Notably, YIGSR‐modified scaffolds improved HUVEC migration significantly in comparison to REDV‐ or RGD‐modified groups. Moreover, its combination with either of these two peptides also presents excellent effect on cell migration. Thus, all the data suggest that the combined application of peptides might be a promising method to enhance the endothelialization of small‐diameter vascular grafts.     

The responses of preosteoblasts to collagen/hyaluronic acid polyelectrolyte multilayer coating on titanium

The purpose of this study was to investigate the effect of Collagen/hyaluronic acid (Col/HA) polyelectrolyte multilayer (PEM) coating on the behaviors and function of pre‐osteoblastic cells. Col/HA PEM coating was fabricated on acid etched Ti disc via layer‐by‐layer technique. The formation and properties of Col/HA PEM coating were investigated using scanning electron microscope (SEM), contact angle and quartz crystal microbalance (QCM). Moreover, mouse preosteoblasts were used to evaluate the effect of Col/HA PEM coating on cell behaviors and function. The results of in vitro evaluations showed that the Col/HA PEM coating could accelerate the proliferation and differentiation of preosteoblasts cells. Copyright © 2011 John Wiley & Sons, Ltd.     

Coating typologies and constrained swelling of hyaluronic acid gels within scaffold pores

A set of elastomeric scaffolds with a well defined porous structure was prepared with a template leaching procedure and coated with hyaluronic acid solutions. Depending on the coating process parameters the hyaluronic acid deposited on the pores had configurations ranging from thin disconnected aggregates to a thick continuous layer on the pore surface. The development of the coating layer was studied by scanning electron microscopy and the materials were subjected to dynamical and equilibrium swelling experiments in a water vapor ambient of fixed activity. The porosity change due to coating and to swelling of the coating layer were determined. The hyaluronic acid coating the pores has a different swelling capacity depending on the type of layer formed, as a consequence of the scaffold constraint and of the layer typology. These factors were investigated analytically by modifying the standard theory of gel swelling. An experimental quantity is introduced which reflects the constrainment build-up on gel swelling.     

Formation of Zirconium(IV)–Heparin Complex Multilayers on Solid Surfaces for Long‐Lasting Antiplatelet Application

A facile approach to enhancing the blood compatibility of solid surfaces based on Zr^IV–heparin complexation is reported. Solid surfaces are pretreated with tannic acid (TA)/Zr^IV complexes. Heparin is then deposited on the surface through a spin‐coating process and fixed by a Zr^IV‐mediated crosslinking reaction. Using this approach, TA/Zr^IV/heparin complex multilayers that are highly resistant to human platelet adhesion are formed on various substrates including metal, metal oxides, ceramics, and synthetic polymers. This approach presents a sustainable way for the immobilization of heparin onto surfaces because it does not require any derivatization of heparin molecule as well as time‐consuming processes.     

无机填料在改性两亲性水凝胶防污材料中的应用研究

水凝胶防污材料因其环保特性成为当前海洋防污领域的研究热点,然而其黏附及力学性能的不足仍是限制其实际应用的技术关键。本研究在油性结构单元增加黏附的两亲性水凝胶的基础上,通过物理共混引入Al₂O₃、 TiO₂、蒙脱土和高岭土赋予两亲性水凝胶涂层更多的黏附机制并考察它们对水凝胶涂层其它性能的影响。研究发现,随着无机填料的引入和含量的增加,水凝胶涂层的黏附性能大幅增加,静态泡板和动态划水的不脱落时间分别提高6倍和2.5倍。其中,TiO₂和高岭土能使水凝胶涂层的应力明显增强。虽然,无机填料的加入使两亲性水凝胶涂层抑制牛血清蛋白及小新月菱形藻吸附的能力下降,但在实海挂板实验中依然表现出相当的防污能力,且具有一定的实际应用价值。本研究为探讨无机填料对水凝胶涂层性能的影响提供了一些参考,并对提高两亲性水凝胶涂层黏附及力学性能提供了一种策略。