Nanofibrous membrane of collagen–polycaprolactone for cell growth and tissue regeneration

Nanofibrous substrates of synthetic polymers including polycaprolactone (PCL) have shown considerable potential in tissue regeneration. This paper reports the use of PCL/collagen nanofibers to improve the in vitro osteoblastic responses for the applications in bone regeneration area. Collagen and PCL were dissolved in a co-solvent, and the resulting solution was electrospun into a nanofibrous web. Nonwoven fibrous matrices were successfully produced at various compositional ratios (PCL/collagen = 1/3, 1 and 3 by weight). Although the PCL nanofiber was hydrophobic, the presence of collagen significantly improved the water affinity, such as the water contact angle and water uptake capacity. Tensile mechanical tests showed that the collagen–PCL nanofiber had a significantly higher extension rate (approximately 2.8-fold) than the PCL while maintaining the maximum tensile load in a similar range. The osteoblastic cells cultured on the collagen–PCL nanofibrous substrate showed better initial adhesion and a higher level of growth than those cultured on the PCL nanofiber. Furthermore, real-time RT-PCR revealed the expression of a series of bone-associated genes, including osteopontin, collagen type I and alkaline phosphatase. The expression of these genes was significantly higher on the collagen–PCL nanofiber than on the PCL nanofiber. When subcutaneously implanted in mouse the collagen–PCL membrane facilitated tissue cells to well penetrate into the nanofibrous structure at day 7, whilst no such cell penetration was noticed in the pure PCL nanofiber. Overall, the presence of collagen within the PCL nanofiber improves the water affinity, tensile extension rate, and the tissue cell responses, such as initial adhesion, growth, penetration and the expression of bone-associated genes. Therefore, the collagen–PCL nanofibrous membrane may have potential applications in the cell growth and bone tissue regeneration.     

Fabrication of glycidyl methacrylate-modified silk fibroin/poly(L-lactic acid-co-ε-caprolactone)–polyethylene glycol diacrylate hybrid 3D nanofibrous scaffolds for tissue engineering

In order to provide a biomimetic natural extracellular matrix microenvironment with excellent mechanical capacity for tissue regeneration, a novel porous hybrid glycidyl methacrylate-modified silk fibroin/poly(L-lactic acid-co-ε-caprolactone)–polyethylene glycol diacrylate (SFMA/P(LLA-CL)–PEGDA) hybrid three-dimensional (3D) nanofibrous scaffolds was successfully fabricated through the combination of 3D nanofibrous platforms and divinyl PEGDA based photocrosslinking, and then further improved water resistance by ethanol vapor post-treatment. Scanning electron microscopy and micro-computed tomography results demonstrated significant PEGDA hydrogel-like matrices bonded nanofibers, which formed a 3D structure similar to that of “steel bar (nanofibers)‒cement (PEGDA)”, with proper pore size, high porosity, and high pore connectivity density. Meanwhile, the hybrid 3D nanofibrous scaffolds showed outstanding swelling properties as well as improved compressive and tensile properties. Furthermore, these hybrid 3D nanofibrous scaffolds could provide a biocompatible microenvironment, capable of inducing the material‒cell hybrid and regulating human umbilical vein endothelial cells proliferation. They thus present significant potential in tissue regeneration.     

Design and production of sintered β-tricalcium phosphate 3D scaffolds for bone tissue regeneration

The characteristics of sintered β-tricalcium phosphate (β-TCP) scaffolds produced by 3D printing were studied by means of X-ray diffraction, Scanning Electron Microscopy, Fourier transform infrared spectroscopy, uniaxial compression tests and cytotoxicity tests, using human osteoblast cells.The results reported include details of the β-TCP scaffolds' porosity, density, phase stability, mechanical behavior and cytotoxic profile. Collectively, these properties are fundamental for the future application of these scaffolds as bone substitutes for individualized therapy.     

Synthesis of activated poly(α,β-malic acid) using N-hydroxysuccinimide and its gelation with collagen as biomaterials

The development of synthetic biodegradable polymers, such as poly(α-hydroxy acid), is particularly important for constructing medical devices, including scaffolds and sutures, and has attracted growing interest in the biomedical field. Here, we report a synthesis method for activated polymalic acid (Su-PMA) by the reaction between synthesized α,β-PMA and N-hydroxysuccinimide (HOSu) in the presence of 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC), and present an evaluation of the gelation properties of Su-PMA with collagen. The Su content of the resulting Su-PMA is higher than the feed ratio. Su content and chain length of Su-PMA influence gelation properties, as Su-PMA with low Su content formed a gel.     

Crosslinked collagen–gelatin–hyaluronic acid biomimetic film for cornea tissue engineering applications

Cornea disease may lead to blindness and keratoplasty is considered as an effective treatment method. However, there is a severe shortage of donor corneas worldwide. This paper presents the crosslinked collagen (Col)–gelatin (Gel)–hyaluronic acid (HA) films developed by making use of 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) as the crosslinker. The test results on the physical and biological properties indicate that the CGH631 film (the mass ratio of Col:Gel:HA = 6:3:1) has appropriate optical performance, hydrophilicity and mechanical properties. The diffusion properties of the CGH631 film to NaCl and tryptophan are also satisfactory and the measured data are 2.43 × 10^? 6 cm²/s and 7.97 × 10^? 7 cm²/s, respectively. In addition, cell viability studies demonstrate that the CGH631 film has good biocompatibility, on which human corneal epithelial cells attached and proliferated well. This biocompatible film may have potential use in cornea tissue engineering.     

PLLA–collagen and PLLA–gelatin hybrid scaffolds with funnel-like porous structure for skin tissue engineering

In skin tissue engineering, a three-dimensional porous scaffold is necessary to support cell adhesion and proliferation and to guide cells moving into the repair area in the wound healing process. Structurally, the porous scaffold should have an open and interconnected porous architecture to facilitate homogenous cell distribution. Moreover, the scaffolds should be mechanically strong to protect deformation during the formation of new skin. In this study, the hybrid scaffolds were prepared by forming funnel-like collagen or gelatin sponge on a woven poly(l-lactic acid) (PLLA) mesh. The hybrid scaffolds combined the advantages of both collagen or gelatin (good cell-interactions) and PLLA mesh (high mechanical strength). The hybrid scaffolds were used to culture dermal fibroblasts for dermal tissue engineering. The funnel-like porous structure promoted homogeneous cell distribution and extracellular matrix production. The PLLA mesh reinforced the scaffold to avoid deformation. Subcutaneous implantation showed that the PLLA–collagen and PLLA–gelatin scaffolds promoted the regeneration of dermal tissue and epidermis and reduced contraction during the formation of new tissue. These results indicate that funnel-like hybrid scaffolds can be used for skin tissue regeneration.     

PLLA–collagen and PLLA–gelatin hybrid scaffolds with funnel-like porous structure for skin tissue engineering

Abstract

In skin tissue engineering, a three-dimensional porous scaffold is necessary to support cell adhesion and proliferation and to guide cells moving into the repair area in the wound healing process. Structurally, the porous scaffold should have an open and interconnected porous architecture to facilitate homogenous cell distribution. Moreover, the scaffolds should be mechanically strong to protect deformation during the formation of new skin. In this study, the hybrid scaffolds were prepared by forming funnel-like collagen or gelatin sponge on a woven poly(l-lactic acid) (PLLA) mesh. The hybrid scaffolds combined the advantages of both collagen or gelatin (good cell-interactions) and PLLA mesh (high mechanical strength). The hybrid scaffolds were used to culture dermal fibroblasts for dermal tissue engineering. The funnel-like porous structure promoted homogeneous cell distribution and extracellular matrix production. The PLLA mesh reinforced the scaffold to avoid deformation. Subcutaneous implantation showed that the PLLA–collagen and PLLA–gelatin scaffolds promoted the regeneration of dermal tissue and epidermis and reduced contraction during the formation of new tissue. These results indicate that funnel-like hybrid scaffolds can be used for skin tissue regeneration.     

Fabrication and evaluation of biomimetic scaffolds by using collagen–alginate fibrillar gels for potential tissue engineering applications

Pore architecture and its stable functionality under cell culturing of three dimensional (3D) scaffolds are of great importance for tissue engineering purposes. In this study, alginate was incorporated with collagen to fabricate collagen–alginate composite scaffolds with different collagen/alginate ratios by lyophilizing the respective composite gels formed via collagen fibrillogenesis in vitro and then chemically crosslinking. The effects of alginate amount and crosslinking treatment on pore architecture, swelling behavior, enzymatic degradation and tensile property of composite scaffolds were systematically investigated. The relevant results indicated that the present strategy was simple but efficient to fabricate highly interconnected strong biomimetic 3D scaffolds with nanofibrous surface. NIH3T3 cells were used as a model cell to evaluate the cytocompatibility, attachment to the nanofibrous surface and porous architectural stability in terms of cell proliferation and infiltration within the crosslinked scaffolds. Compared with the mechanically weakest crosslinked collagen sponges, the cell-cultured composite scaffolds presented a good porous architecture, thus permitting cell proliferation on the top surface as well as infiltration into the inner part of 3D composite scaffolds. These composite scaffolds with pore size ranging from 150 to 300 μm, over 90% porosity, tuned biodegradability and water-uptake capability are promising for tissue engineering applications.     

Fabrication of three-dimensional poly(ε-caprolactone) scaffolds with hierarchical pore structures for tissue engineering

The physical properties of tissue engineering scaffolds such as microstructures play important roles in controlling cellular behaviors and neotissue formation. Among them, the pore size stands out as a key determinant factor. In the present study, we aimed to fabricate porous scaffolds with pre-defined hierarchical pore sizes, followed by examining cell growth in these scaffolds. This hierarchical porous microstructure was implemented via integrating different pore-generating methodologies, including salt leaching and thermal induced phase separation (TIPS). Specifically, large (L, 200–300 μm), medium (M, 40–50 μm) and small (S, < 10 μm) pores were able to be generated. As such, three kinds of porous scaffolds with a similar porosity of ~ 90% creating pores of either two (LS or MS) or three (LMS) different sizes were successfully prepared. The number fractions of different pores in these scaffolds were determined to confirm the hierarchical organization of pores. It was found that the interconnectivity varied due to the different pore structures. Besides, these scaffolds demonstrated similar compressive moduli under dry and hydrated states. The adhesion, proliferation, and spatial distribution of human fibroblasts within the scaffolds during a 14-day culture were evaluated with MTT assay and fluorescence microscopy. While all three scaffolds well supported the cell attachment and proliferation, the best cell spatial distribution inside scaffolds was achieved with LMS, implicating that such a controlled hierarchical microstructure would be advantageous in tissue engineering applications.     

Preparation and characterization of bimodal porous poly(γ-benzyl-L-glutamate) scaffolds for bone tissue engineering

An ideal scaffold in bone tissue-engineering strategy should provide biomimetic extracellular matrix-like architecture and biological properties. Poly(γ-benzyl-L-glutamate) (PBLG) has been a popular model polypeptide for various potential biomedical applications due to its good biocompatibility and biodegradability. This study developed novel bimodal porous PBLG polypeptide scaffolds via a combination of biotemplating method and in situ ring-opening polymerization of γ-benzyl-L-gIutamate N-carboxyanhydride (BLG-NCA). The PBLG scaffolds were characterized by proton nuclear magnetic resonance spectroscopy, X-ray diffraction, differential scanning calorimetry, scanning electron microscope (SEM) and mechanical test. The results showed that the semi-crystalline PBLG scaffolds exhibited an anisotropic porous structure composed of honeycomb-like channels (100–200 μm in diameter) and micropores (5–20 μm), with a very high porosity of 97.4 ± 1.6%. The compressive modulus and glass transition temperature were 402.8 ± 20.6 kPa and 20.2 °C, respectively. The in vitro biocompatibility evaluation with MC3T3-E1 cells using SEM, fluorescent staining and MTT assay revealed that the PBLG scaffolds had good biocompatibility and favored cell attachment, spread and proliferation. Therefore, the bimodal porous polypeptide scaffolds are promising for bone tissue engineering.     

Poly(ε-caprolactone) and poly(D,L-lactic acid-co-glycolic acid) scaffolds used in bone tissue engineering prepared by melt compression-particulate leaching method

Porous bioresorbable polymers have been widely used as scaffolds in tissue engineering. Most of the bioresorbable scaffolds are aliphatic polyesters and the methods employed to prepare the porous morphology may vary. This work describes and evaluates the in vitro degradation of porous and dense scaffolds of poly(ε-caprolactone) (PCL) and poly(D,L-lactic acid-co-glycolic acid) (50/50) (PLGA50) prepared by particulate leaching-melt compression process. Biological evaluation was carried out using osteoblast cell cultures. The results showed an autocatalytic effect on the dense samples. Osteoblasts presented intermediate adhesion and the cell morphology on the surface of these materials was dispersed, which indicated a good interaction of the cells with the surface and the material.     

Chemical hydrogels based on a hyaluronic acid-graft-α-elastin derivative as potential scaffolds for tissue engineering

In this work hyaluronic acid (HA) functionalized with ethylenediamine (EDA) has been employed to graft α-elastin. In particular a HA-EDA derivative bearing 50 mol% of pendant amino groups has been successfully employed to produce the copolymer HA-EDA-g-α-elastin containing 32% w/w of protein. After grafting with α-elastin, remaining free amino groups reacted with ethylene glycol diglycidyl ether (EGDGE) for producing chemical hydrogels, proposed as scaffolds for tissue engineering. Swelling degree, resistance to chemical and enzymatic hydrolysis, as well as preliminary biological properties of HA-EDA-g-α-elastin/EGDGE scaffold have been evaluated and compared with a HA-EDA/EGDGE scaffold. The presence of α-elastin grafted to HA-EDA improves attachment, viability and proliferation of primary rat dermal fibroblasts and human umbilical artery smooth muscle cells. Biological performance of HA-EDA-g-α-elastin/EGDGE scaffold resulted comparable to that of a commercial collagen type I sponge (Antema®), chosen as a positive control.     

Electrospinning of poly (ε-caprolactone-co-lactide)/Pluronic blended scaffolds for skin tissue engineering

For skin tissue engineering, an ideal scaffold should mimic the natural extracellular matrix of the native skin. In this study, we reported a novel elastic sub-micron fiber scaffold blending poly (ε-caprolactone-co-lactide) (PLCL) and Pluronic at different ratios by electrospinning. PLCL and Pluronic were co-electrospun with the ratio of 100/0, 99/1, 95/5, 90/10, 85/15, and 75/25. These scaffolds were evaluated in terms of fiber morphology, mechanical properties, and hydrophilicity for the purpose of culturing adipose-derived stem cells (ADSCs). Cell attachment and proliferation on the scaffolds were also evaluated to demonstrate the potential of serving as a skin graft. The results indicated that all of the electrospun fibers possessed smooth surface textures and interconnected porous structures with the average diameter ranging from approximately 750–1140 nm. The higher tensile strength was observed in 95/5 and 90/10 PLCL/Pluronic blended membranes, while further incorporation of Pluronic almost has no effect on tensile strength. The water contact angle was 85° for scaffold with the ratio of 99/1, while 0° for 90/10, 85/15, and 75/25. In addition, the elevation of Pluronic content in composition resulted in a corresponding increase in swelling behavior. Compared with PLCL, the better cell adhesion and proliferation potential of ADSCs was exhibited on all PLCL/Pluronic blended scaffolds. ADSCs on the blended scaffolds were highly elongated and well integrated with the surrounding fibers, indicating the good cytocompatibility of PLCL/Pluronic scaffolds. Thus, these blended scaffolds have the potentially high application prospect in the field of skin tissue engineering.     

Tetracycline-encapsulated P(3HB) microsphere-coated 45S5 Bioglass®-based scaffolds for bone tissue engineering

Bioglass^®-based scaffolds for bone tissue engineering have been developed, which can also serve as carriers for drug delivery. For this, P(3HB) microspheres (PMSs) loaded with tetracycline were fabricated and immobilised on the scaffold surfaces by a modified slurry dipping technique. The sustained drug delivery ability in simulated body fluid was confirmed by using UV–Vis absorption spectroscopy measurements. The MTT assay using mouse fibroblast cells provided evidence that the tetracycline loaded microspheres produced in this study show limited cytotoxicity. The scaffolds developed in this work provide mechanical support, adequate 3D surface roughness, bioactivity and controlled drug delivery function, and are thus interesting candidates for bone tissue engineering applications.     

Fabrication of porous poly(L-lactide) (PLLA) scaffolds for tissue engineering using liquid–liquid phase separation and freeze extraction

PLLA scaffolds were successfully fabricated using liquid–liquid phase separation with freeze extraction techniques. The effects of different processing conditions, such as method of cooling (direct quenching and pre-quenching), freezing temperature (−80°C and −196°C) and polymer concentration (3, 5 and 7 wt%) were investigated in relations to the scaffold morphology. SEM micrographs of scaffolds showed interconnected porous network with pore size ranging from 20 to 60 μm. The scaffolds had porosity values ranging from 80 to 90%. Changes to the interconnected network, porosity and pore size were observed when the method of cooling and polymer concentration was changed. Direct quenching to −80°C gave a more porous interconnected microstructure with uniform pore size compared to samples prepared using pre-quenching method. Larger pores were observed for samples quenched at −80°C compared to −196°C. Scaffolds prepared using direct quenching to −196°C had higher elastic modulus and compressive stress compared to those quenched to −80°C. The compressive elastic modulus ranged from 4 to 7 MPa and compressive stress at 10% strain was from 0.13 to 0.18 MPa.     

Fabrication and characterization of injection molded poly (ε-caprolactone) and poly (ε-caprolactone)/hydroxyapatite scaffolds for tissue engineering

In this study, poly(ε-caprolactone) (PCL)/sodium chloride (NaCl), PCL/poly(ethylene oxide) (PEO)/NaCl and PCL/PEO/NaCl/hydroxyapatite (HA) composites were injection molded and characterized. The water soluble and sacrificial polymer, PEO, and NaCl particulates in the composites were leached by deionized water to produce porous and interconnected microstructures. The effect of leaching time on porosity, and residual contents of NaCl and NaCl/HA, as well as the effect of HA addition on mechanical properties was investigated. In addition, the biocompatibility was observed via seeding human mesenchymal stem cells (hMSCs) on PCL and PCL/HA scaffolds.The results showed that the leaching time depends on the spatial distribution of sacrificial PEO phase and NaCl particulates. The addition of HA has significantly improved the elastic (E′) and loss moduli (E″) of PCL/HA scaffolds. Human MSCs were observed to have attached and proliferated on both PCL and PCL/HA scaffolds. Taken together, the molded PCL and PCL/HA scaffolds could be good candidates as tissue engineering scaffolds. Additionally, injection molding would be a potential and high throughput technology to fabricate tissue scaffolds.     

Electrospun fibrous scaffold of hydroxyapatite/poly (ε-caprolactone) for bone regeneration

Development of fibrous scaffold of hydroxyapatite/biopolymer nanocomposite offers great potential in the field of bone regeneration and tissue engineering. Hydroxyapatite (HA)/poly (ε-caprolactone) (PCL) fibrous scaffolds were successfully prepared by electrospinning dopes containing HA and PCL in this work. It was found that pre-treating HA with γ-glycioxypropyltrimethoxysilane (A-187) was effective in improving HA dispersion both in solutions and in a PCL matrix. Mechanical properties of the scaffolds were greatly enhanced by the filling of A187-HA. The bioactivity of PCL was remarkably improved by the addition of HA and A187-HA. Fibroblasts and osteoblasts were seeded on scaffolds to evaluate the effect of A-187 on biocompatibility of HA/PCL composites. Based on this study, good dispersion of HA in PCL matrix was granted by pretreatment of HA with A-187 and A187-HA/PCL fibrous scaffolds were obtained by electrospinning. These results demonstrated that the scaffolds may possess improved mechanical performance and good bioactivity due to A187-HA incorporation.     

Synthesis and Characterization of Cu Nanoparticles Embedded in PAN/β-Cyclodextrin (β-CD) Composite Nanofiber Films

In the study, PAN/β-cyclodextrin (β-CD) one dimensional composite nanofibers were synthesized with sol-gel method and electrospinning, and then, we dipped the composite nanofibers into NaBH₄ aqueous solution acted as reducing agent to fabricate PAN/β-CD/Cu composite nanofibers. We got the PAN/β-CD/Cu composite nanofibers with different concentration of β-CD, and Cu nanoparticles formed on its surface. The final products were characterized using scanning electronic microscope (SEM), fourier transform infrared spectroscopy (FTIR) and transmission electron microscope (TEM). The results of characterization showed that the Cu nanoparticles with small size were well-distributed on the composite nanofibers.