Key Engineering Materials Vol. 587

Abstract: Collagen for biomedical applications is mainly isolated from animal tissues (bovine or porcine skin and bovine or equine Achilles tendons). Type I collagen has been also extracted from skin, bone, fins and scales of fresh water and marine fishes. Fish scales are composed of collagen covered with calcium salts. In the present study we report the preparation of collagen from fish scales for potential cosmetic, pharmaceutical and implant applications. In our laboratory collagen was isolated from scales of Esox lucius. It was the first time that this species were used as sources of collagen. Extraction of collagen from fish scales was done in two steps. In the first step, fish scales were demineralized using EDTA. Energy dispersive X-ray analysis of demineralized scale was carried out for quantitative estimation of inorganic content. Then, demineralized fish scales were dissolved in acetic acid. Collagen isolated from Esox Lucius may serve as an attractive and safe source of collagen for biomedical and pharmaceutical applications. Fish collagen can be processed in sheet, sponges foams, injectable viscous solution, and dispersions.

Abstract: Composites based on calcium phosphates (CP) and mixtures of biopolymers (chitosan and hyaluronic acid) have been prepared by a biomimetic co-precipitation method and tested as scaffolds for bone tissue engineering. The biomimetic strategy is inspired by natural mineralization processes, where the synthesized minerals are usually combined with proteins, polysaccharides or other mineral forms to form composite, in physiological conditions of temperature and pH. Fourier transformed infrared spectroscopy, scanning electron microscopy, X-ray diffraction and XPS analyses confirmed the porous morphology of the scaffolds and formation of various forms of calcium phosphates with amorphous nature. The in vitro degradation studies showed a slow degradation process for CP-biopolymers composites and limited swelling in simulated body fluids. The scaffolds compositions have no negative effect on osteoblasts cell, emphasizing a good biocompatibility.

Abstract: Biomaterial is a non-drug substance suitable for inclusion in systems which augment or replace the function of body tissues or organs. The nano-hydroxyapatite (HAp) is one of the most applied biomaterial, a popular bone substitute, for coatings and other filler materials due to their ability to promote mineralization. Hydroxyapatite [Ca₁₀(PO₄)₆(OH)₂] is one of the most stable forms of calcium phosphate and it occurs in bones as major component (60 to 65%), along with other materials including collagen, chondroitin sulfate, keratin sulfate and lipids. In order to improve the bioactivity and mechanical properties of hydroxyapatite, some synthetic and natural polymers (e.g., cellulose, collagen, chitosan, chitine) have been used as excellent candidates for bone/cartilage tissue engineering applications. In this paper we propose to synthesize novel three-component composite materials from HAp and cellulose (carboxymethyl cellulose, CMC) with one of the following natural polymers (chitosan (CS), chitine (CT), collagen (CLG), all of them named CX), biocompatible (from CMC) and with excellent antimicrobial activity (from HAp). Similarly to CS, CMC has an extensive network of intra- and inter-molecular hydrogen bonds which makes it insoluble in water or in common organic solvents. To prepare this new composite material, ionic liquids (ILs) have been used as potential green solvents for biopolymers, knowing that 1-butyl-3-methylimidazolium chloride [BMImCl] is able to dissolve up to 10% (w/w) of CMC, and other polysaccharides such as CS. In the studied systems, HAp particles were dispersed uniformly in organic phase, and are present strong chemical interactions between the three phases. In our case, a highly porous film was prepared and its bioactivity was investigated by in vitro tests in a simulated body fluid (SBF) for 6 months. By ICP-OES, was found a migration of analyzed metals (Fe, Mn, Cr and Ni) in simulated physiological fluids (SBF) analyzed after a period of 6 months, but only in the ppb concentrations range. Different system (HAp/CX/CMC) composites with weight ratios of 70/10/20, 70/15/15 and 70/20/10 were prepared. The new composites were characterized by infrared spectroscopy Fourier transformed (FT-IR), scanning electron microscopy (SEM) and ICP-OES. Their antimicrobial activity has been tested on Candida albicans, Candida parapsilosis and Staphylococcus aureus, proofing an excellent antimicrobial activity.

Abstract: The properties of new materials based on the blends of collagen and chitosan were studied along with their biological compatibility. Mechanical properties, thermal analysis, FTIR spectra and SEM images were obtained for different blends of chitosan/collagen in weight ratios 75/25, 50/50, 25/75. The materials in the form of 2D sheets were then subjected to cultures of human mesenchymal stem cells (hMSC) directed toward osteogenesis with ascorbate-2-phosphate and dexamethasone. The results showed that specified amounts of components influence the mechanical properties of obtained materials. Proliferation of hMSC decreased with increasing amounts of chitosan in collagen/chitosan films. However, the cellular activity of alkaline phosphatase (ALP), a marker of preosteoblasts and active osteoblasts, for plain collagen films ALP was the highest, whereas different ratios of collagen in chitosan/collagen composites had no effect on overall good ALP activity of hMSC. Based on the presented data, we believe the obtained materials are suitable for bone tissue engineering strategies.

Abstract: The low crystalline hydroxyl carbonate apatite forming capacity of a poly (70lactic-co-30glycolic acid)/15CaO-85SiO₂ composite, which had a dual pore structure, was newly examined in simulated body fluid. The bioactive 15CaO-85SiO₂ particles were synthesized by a sol-gel method using tetraethyl orthosilicate (TEOS) and calcium nitrate tetrahydrate under acidic condition followed by the heat treatment at 600°C for 3h. The poly (70lactic-co-30glycolic acid)/15CaO-85SiO₂ composite was then prepared by a solvent casting using dimethylformide as a solvent. The composite was loaded into a high pressure chamber and then carbon dioxide gas was introduced achieving a final pressure of 20 MPa. After 3 days, carbon dioxide gas was released quickly and resultantly the dual pore structure was obtained. The samples were observed by FE-SEM and its bioactivity was tested in simulated body fluid.

Abstract: The effect of Bioglass addition on mechanical and physical properties of photoactive UDMA-TEGDMA resin composites Laura Nicolae, William Palin, Richard Shelton, Paul Cooper Aim: To determine the effect of Bioglass incorporation on the polymerisation rate (Rp), degree of conversion (DC), flexural strength and modulus (FS/FM) of light-curable filled resin composites (FRCs). Methods: Experimental photoactive resins containing various UDMA-TEGDMAratios and filled with non-silanised irregular or spherical 45S5-Bioglass (50μm; 5-40mass%) and/or silanised silicate glass filler particulates (0.7μm; 50-70mass%) weretested. Rp and DC wereanalysed by real-time near-infrared spectroscopy. FS/FM of resins and FRCs were determined using three-point flexural strength tests. Results: Addition of up to 20mass%45S5-Bioglass filler significantly increased DC and FS(p<0.05), although >20mass% had deleterious effects on mechanical properties. The addition of spherical Bioglass particles resulted in an increased DC compared with that of irregular filler morphology although no significant differences mechanical properties were observed (p>0.05). Conclusion: Addition of 45S5-Bioglass with specific filler content, size and morphology resulted in enhanced mechanical and physical properties compared with conventional silicate glass FRCs.

Abstract: The main objective of this work was to produce membranes of chitosan and collagen type I and check their ability to undergo “in vitro” calcification. The membranes of chitosan-collagen blends were characterized by TGA, infra-red spectroscopy and DSC. Samples of dense and porous membranes were immersed in solution SBF (Simulated Body Fluid) in order to verify their “in vitro” calcification. The membranes were observed by SEM. The production of chitosan-collagen membranes is possible, in dense and porous versions. We can conclude that the blend is less resistant to high temperatures, in comparison to pristine chitosan membranes shown in literature. Through the initial assays of calcification, we observe that it is possible to induce the calcium deposition on a chitosan-collagen membrane, as seen by SEM. Microscopy of fracture surfaces showed fibril structures, probably formed by collagen.

Abstract: During the last two decades learning from nature has given us new directions for the use of natural organic and inorganic skeletons, drug delivery devices, new medical treatment methods initiating unique designs and devices ranging from nanoto macro scale. These materials and designs have been instrumental to introduce the simplest remedies to vital problems in regenerative medicine, providing frameworks and highly accessible sources of osteopromotive analogues, naofibres, micro and macrospheres and mineralising proteins. This is exemplified by the biological effectiveness of marine structures such as corals and shells and sponge skeletons to house self-sustaining musculoskeletal tissues and to the promotion of bone formation by extracts of spongin and nacre seashells. Molecules pivotal to the regulation and guidance of bone morphogenesis and particularly the events in mineral metabolism and deposition similarly exist in the earliest marine organisms because they represent the first molecular components established for calcification, morphogenesis and wound healing. It emerges that bone morphogenic protein (BMP) molecules-the main cluster of bone growth factors for human bone morphogenesis-are secreted by endodermal cells into the developing skeleton. Signalling proteins, TGF and Wnt-prime targets in bone therapeutics-are present in early marine sponge development. Furthermore, ready-made organic and inorganic marine skeletons possess a habitat suitable for proliferating added mesenchymal stem cell populations and promoting clinically acceptable bone formation. In this paper we review the nature, morphology and extent of this association and use of these structures for bone grafts, drug delivery and extracts such as proteins for regenerative medicine. As an example, in human biology a study of matrix vesicles will teach us valuable lessons on how proteins are captured and coated; and how the vesicle is able to dock and fuse with their target. We will describe significant technological trends aimed at producing delivery vehicles using natural-origin soft and hard organized matter; fabricated into capsules and cell-delineated assemblies.Therole model for this specific biomimicry is the filtering microskeleton of Foraminifera. We will outline new selected strategies based on our and others works for the engineering of new bone, based on biomimicry themes using these bioceramics building blocks.

Abstract: Precursors for the preparation of Si-CHA (silicon and carbonate substituted hydroxyl-apatite) were added in stoichiometric composition in amounts either 30 or 50 solid wt. % to a solution of gelatin to form hybrid nano-composite scaffolds (B(G/Si-CHA)). Temperature was maintained at 40°C and pH at 8 during the reaction. Glutaraldehyde was added to complete the cross-linking then washed. Thereafter, glycine was added to the product to get rid of any glutaraldehyde remnants. Both, 30 and 50% obtained scaffolds were compared with ones containing the same proportions from previously prepared nano powders of the Si-CHA named (C(G/Si-CHA)). The four types and a control one made of gelatin alone (G) were examined by X-ray diffraction (XRD), infrared spectroscopy (FTIR), transmission and scanning electron microscopy (TEM & SEM) and mercury porosimeter. In vitro study was carried out on the prepared scaffolds by immersing in simulated body fluid (SBF) solution for different periods between 1and 28 days. The change in the percentage of the different ions with time in the solutions was followed. Results of IR showed the reaction of the carboxylic groups of gelatin with the added ingredients giving a homogeneous distribution of the inorganic phase formed (Si-CHA). High porosity reaching 94% was achieved with macro and micro pores without the addition of a porogen material.

Abstract: In the present work Collagen/Hydroxyapatite microsphere (Col/mHA) scaffold with a multiscale porosity was prepared. Col/mHA composite scaffold was prepared by freeze-drying/dehydrothermal crosslinking method. The HA microspheres (mHA) were obtained by spray drying of nanohydroxyapatite slurry prepared by precipitation technique. XRD analysis revealed that the microspheres were composed only of pure HA phase and EDS analysis revealed that Ca/P ratio was 1.69. The obtained microspheres had an average diameter 6 microns, specific surface area of 40 m2/g by BET analysis and BJH analysis shows meso porous structure having an average pore diameter 16nm. SEM analysis shows that the obtained Col/mHA scaffold had a macro porosity ranging from micron to 200 microns with meso porous mHA embedded in the collagen matrix.