Poly-Left-Lactic Acid tubular scaffolds via Diffusion Induced Phase Separation (DIPS): control of morphology

n this work, tubular poly-left-lactic acid scaffolds for vascular tissue engineering applications were produced by an innovative two-step method. The scaffolds were obtained by performing a dip-coating around a nylon fiber, followed by a diffusion induced phase separation process. Morphological analysis revealed that the internal lumen of the as-obtained scaffold is equal to the diameter of the fiber utilized; the internal surface is homogeneous with micropores 1–2 μm large. Moreover, a porous open structure was detected across the thickness of the walls of the scaffold. An accurate analysis of the preparation process revealed that it is possible to tune up the morphology of the scaffold (wall thickness, porosity, and average pore dimension), simply by varying some experimental parameters. Preliminary in vitro cell culture tests were carried out inside the scaffold. The results showed that cells are able to grow within the internal surface of the scaffolds and after 3 weeks they begin to form a “primordial” vessel-like structure. Modeling predictions of the dip-coating process display always an underestimate of experimental data (dependence of wall thickness upon extraction rate).In this work, tubular poly-left-lactic acid scaffolds for vascular tissue engineering applications were produced by an innovative two-step method. The scaffolds were obtained by performing a dip-coating around a nylon fiber, followed by a diffusion induced phase separation process. Morphological analysis revealed that the internal lumen of the as-obtained scaffold is equal to the diameter of the fiber utilized; the internal surface is homogeneous with micropores 1–2 lm large. Moreover, a porous open structure was detected across the thickness of the walls of the scaffold. An accurate analysis of the preparation process revealed that it is possible to tune up the morphology of the scaffold (wall thickness, porosity, and average pore dimension), simply by varying some experimental parameters. Preliminary in vitro cell culture tests were carried out inside the scaffold. The results showed that cells are able to grow within the internal surface of the scaffolds and after 3 weeks they begin to form a ‘‘primordial’’ vessel-like structure. Modeling predictions of the dipcoating process display always an underestimate of experimental data (dependence of wall thickness upon extraction rate)

PLLA biodegradable scaffolds for angiogenesis via Diffusion Induced Phase Separation (DIPS)

A critical obstacle in tissue engineering is the inability to maintain large masses of living cells upon transfer from the in vitro culture conditions into the host in vivo. Capillaries, and the vascular system, are required to supply essential nutrients, including oxygen, remove waste products and provide a biochemical communication “highway”. For this reason it is mandatory to manufacture an implantable structure where the process of vessel formation – the angiogenesis – can take place. In this work PLLA scaffolds for vascular tissue engineering were produced by dip-coating via Diffusion Induced Phase Separation (DIPS) technique. The scaffolds, with a vessel-like shape, were obtained by performing a DIPS process around a nylon fibre whose diameter was 700 μm. The fibre was first immersed into a 4% PLLA dioxane solution and subsequently immersed into a second bath containing distilled water. The covered fibre was then rinsed in order to remove the excess of dioxane and dried; finally the internal nylon fibre was pulled out so as to obtain a hollow biodegradable PLLA fiber. SEM analysis revealed that the scaffolds have a lumen of ca. 700 μm. The internal surface is homogeneous with micropores 1–2 μm large. Moreover, a cross section analysis showed an open structure across the thickness of the scaffold walls. A cell culture of endothelial cells was carried out into the as-prepared scaffolds. The result showed that cells are able to grow within the scaffolds and after 3 weeks they begin to form a “primordial” vessel-like structure

In vivo angiogenic activity induction by collagen- soaked Poly-L-lactic acid scaffolds

Angiogenesis is essential in tissue integration and it is involved in the biological response to biomaterials. Poly-L-lactic acid (PLLA), a synthetic polymer, is utilized as scaffolding to regenerate new tissues. This study investigated the short and long term degradation and the induction of neovascularization of both native PLLA (n-PLLA) and collagen type I soaked PLLA (c-PLLA) porous scaffolds, implanted subcutane- ously in balb/c mice.The comparative analysis by phase contrast, optical, and scanning electron micros- copy (SEM) of scaffolds 7 and 21 days after implantation showed a mild inflam- matory response at the implant site of c-PLLA scaffold. No significant difference in systemic immune response was detected by hematology analyzer, and by histologi- cal evaluation of lymph node and spleen features. On the contrary, immune reaction was moderate in n-PLLA. Pores of both PLLA networks laying on the muscle fibers were partially infiltrated by appositional collagen/elastin tissue, phagocytic cells, and fibroblast, with respect to the inner side. The presence of numerous and large blood vessels into pores of c-PLLA scaffolds showed an enhancing vascularization rate. These characteristics appeared to be less conspicuous in n-PLLA.At longer time points (42 and 84 days), there was low difference in inflammato- ry cell presence into scaffold pores and the number of cells infiltrating each implant was significantly decreased. In fact, we did not observe difference in the migration of inflammatory cells into PLLA scaffolds. Polymer degradation was detected in both PLLA networks, but there are no considerable differences, as confirmed by the SEM analysis.Our results suggest that tissue integration of PLLA is enhanced when it is soaked with collagen, as well as the angiogenic activity on c-PLLA. Furthermore, the colla- gen soaking makes PLLA polymer more suitable for supporting cell attachment, pro- liferation, and function by mimicking the natural extra cellular matrix.

Galvanic Deposition of Hydroxyapatite/Chitosan/Collagen Coatings on 304 Stainless Steel

The galvanic deposition method was used to deposit Hydroxyapatite/Chitosan/Collagen coatings on 304 stainless steel. Galvanic deposition is an alternative and valid way to fabricate bio-coatings with high biocompatibility and good anticorrosion properties. Physical-chemical characterizations were carried out to investigate chemical composition and morphology of the samples. Coatings consist of a mixture of calcium phosphate (Brushite and Hydroxyapatite) with chitosan and collagen. Corrosion tests were performed in the simulated body fluid (SBF) after different aging times. Results show that, in comparison with bare 304 stainless steel, coating shifts corrosion potential to anodic values and reduces corrosion current density. Nevertheless, the aging in SBF led to a completely conversion of brushite into hydroxyapatite. The release of metal ions, measured after 21 days of aging in SBF solution, is very low due to the presence of coating that slow-down the corrosion rate of steel

Modulation of physical and biological properties of a composite PLLA and polyaspartamide derivative obtained via thermally induced phase separation (TIPS) technique

In the present study, blend of poly l-lactic acid (PLLA) with a graft copolymer based on α,β-poly(N-hydroxyethyl)-dl-aspartamide and PLA named PHEA-PLA, has been used to design porous scaffold by using Thermally Induced Phase Separation (TIPS) technique. Starting from a homogeneous ternary solution of polymers, dioxane and deionised water, PLLA/PHEA-PLA porous foams have been produced by varying the polymers concentration and de-mixing temperature in metastable region. Results have shown that scaffolds prepared with a polymer concentration of 4% and de-mixing temperature of 22.5 °C are the best among those assessed, due to their optimal pore size and interconnection. SEM and DSC analysis have been carried out respectively to study scaffold morphology and the influence of PHEA-PLA on PLLA crystallization, while DMF extraction has been carried out in order to quantify PHEA-PLA into the final scaffolds. To evaluate scaffold biodegradability, a hydrolysis study has been performed until 56 days by incubating systems in a media mimicking physiological environment (pH 7.4). Results obtained have highlighted a progressive increase in weight loss with time in PLLA/PHEA-PLA scaffolds, conceivably due to the presence of PHEA-PLA and polymers interpenetration. Viability and adhesion of bovine chondrocytes seeded on the scaffolds have been studied by MTS test and SEM analysis. From results achieved it appears that the presence of PHEA-PLA increases cells affinity, allowing a faster adhesion and proliferation inside the scaffold

Use of Modified 3D Scaffolds to Improve Cell Adhesion and Drive Desired Cell Responses.

In the most common approach of tissue engineering, a polymeric scaffold with a well-defined architecture has emerged as a promising platform for cells adhesion and guide their proliferation and differentiation into the desired tissue or organ. An ideal model for the regeneration should mimic clinical conditions of tissue injury, create a permissive microenvironment for diffusion of nutrients, gases and growth factors and permit angiogenesis. In this work, we used a 3D support made of synthetic resorbable polylactic acid (PLLA), which has considerable potential because of its well-known biocompatibility and biodegradability. One of the factors that influence cell adhesion to the scaffold is its porosity degree, but surface properties represent the main driving forces that influence the composition and orientation of proteins that will be absorbed onto material surfaces. We used scaffolds in which it was possible to control pore size and that had undergone on type-I collagen treatment, to mimic the extra cellular matrix, or plasma enhanced chemical vapor deposition (PE-CVD) combined with plasma treatment, in order to modify surface chemistry of the material. Our results show different cell affinity in non-treated scaffolds compared to type-I collagen or plasma modified ones. These surface changes are of considerable interest for tissue engineering and other areas of biomaterials science, where it can be useful to improve the surface of biomedical polymers to facilitate the colonization of the structure by the cells and obtain a more rapid regeneration or tissue replacement.In the most common approach of tissue engineering, a polymeric scaffold with a well-defined architecture has emerged as a promising platform for cells adhesion and guide their proliferation and differentiation into the desired tissue or organ. An ideal model for the regeneration should mimic clinical conditions of tissue injury, create a permissive microenvironment for diffusion of nutrients, gases and growth factors and permit angiogenesis. In this work, we used a 3D support made of synthetic resorbable polylactic acid (PLLA), which has considerable potential because of its well-known biocompatibility and biodegradability. One of the factors that influence cell adhesion to the scaffold is its porosity degree, but surface properties represent the main driving forces that influence the composition and orientation of proteins that will be absorbed onto material surfaces. We used scaffolds in which it was possible to control pore size and that had undergone on type-I collagen treatment, to mimic the extra cellular matrix, or plasma enhanced chemical vapor deposition (PE-CVD) combined with plasma treatment, in order to modify surface chemistry of the material. Our results show different cell affinity in non-treated scaffolds compared to type-I collagen or plasma modified ones. These surface changes are of considerable interest for tissue engineering and other areas of biomaterials science, where it can be useful to improve the surface of biomedical polymers to facilitate the colonization of the structure by the cells and obtain a more rapid regeneration or tissue replacement. Copyright © 2012, AIDIC Servizi S.r.l

Galvanic deposition of Chitosan-AgNPs as antibacterial coating

Thanks to mechanical properties similar human bones, metallic materials represent the best choice for fabrication of orthopedic implants. Although metals could be widely used in the field of biomedical implants, corrosion phenomena could occur, causing metal ions releasing around periprosthetic tissues leading, in the worst cases, to the development of infections. In these cases, patients need prolonged antibiotic therapies that may cause bacterial resistance. Preventing bacterial colonization of biomedical surfaces is the key to limiting the spread of infections. Antibacterial coatings have become a very active field of research, strongly stimulated by the increasing urgency of identifying alternatives to the traditional administration of antibiotics. Nowadays, the research was focused on coating science to deal with these issues. In particular, the development of the antibacterial composite coatings could be a viable way to provide not only a corrosion resistance but also an antibacterial action and biocompatibility. Chitosan is a great biomaterial used in medicine. It is a natural bioactive polymer and is the second most abundant in nature polysaccharide after cellulose. Chitosan comes from the deacetylation of chitin, a homopolymer of beta-(1-4)-N-acetyl-D-glucosamine, derived from exoskeleton of crustaceans. It is high biocompatible and it is also used in drug delivery. In addition, chitosan has chelating properties due to the amino groups of polysaccharide that are responsible of selective chelation with metal ions. In particular, the attention has been paid to silver nanoparticles for their high stability, low toxicity, biocompatibility and antibacterial properties. These ones are incorporated in polymeric matrix (e.g. chitosan) and they are capable to interact physically with cell walls of bacteria. In this study Chitosan-Silver nanoparticles composite coating on AISI 304L was investigated. These coatings were realized by an alternative method of deposition respect to traditional ones based on galvanic coupling. This process doesn’t request any external power supply and is very easy to carried out. The difference of the electrochemical redox potential between the substrate (cathode) and a sacrificial anode is the pivotal role of the process. Deposition rate is controlled by the ratio of cathodic and anodic area. In practice, electrons generated by anode corrosion flow towards to more noble metal thanks to a short-circuit. As soon electrons arrive to the cathode, the base electrogeneration reactions of nitrate ions and water molecules occur. Production of hydroxyl ions causes an increasing of pH at substrate/solution interface. Hence, deprotonation of amine group leads precipitation of chitosan (pKa=6.4) onto surface. At the same time, silver nanoparticles are incorporated in polymeric matrix of chitosan. Physical-chemical characterizations of the coatings were carried out in order to investigate morphology and chemical composition. In addition, corrosion tests (potentiodynamic polarization and electrochemical impedance spectroscopy) were executed in a simulated body fluid to scrutinize the corrosion resistance. Furthermore, the release of silver nanoparticles from coating in SBF were studied

Galvanic Deposition of Calcium Phosphate/Bioglass Composite Coating on AISI 316L

Calcium phosphate/Bioglass composite coatings on AISI 316L were investigated with regard to their potential role as a beneficial coating for orthopedic implants. These coatings were realized by the galvanic co-deposition of calcium phosphate compounds and Bioglass particles. A different amount of Bioglass 45S5 was used to study its effect on the performance of the composite coatings. The morphology and chemical composition of the coatings were investigated before and after their aging in simulated body fluid. The coatings uniformly covered the AISI 316L substrate and consisted of a brushite and hydroxyapatite mixture. Both phases were detected using X-ray diffraction and Raman spectroscopy. Additionally, both analyses revealed that brushite is the primary phase. The presence of Bioglass was verified through energy-dispersive X-ray spectroscopy, which showed the presence of a silicon peak. During aging in simulated body fluid, the coating was subject to a dynamic equilibrium of dissolution/reprecipitation with total conversion in only the hydroxyapatite phase. Corrosion tests performed in simulated body fluid at different aging times revealed that the coatings made with 1 g/L of Bioglass performed best. These samples have a corrosion potential of −0.068V vs. Ag/AgCl and a corrosion current density of 8.87 × 10−7 A/cm2. These values are better than those measured for bare AISI 316L (−0.187 V vs. Ag/AgCl and 2.52 × 10−6 A/cm2, respectively) and remained superior to pure steel for all 21 days of aging. This behavior indicated the good protection of the coating against corrosion phenomena, which was further confirmed by the very low concentration of Ni ions (0.076 ppm) released in the aging solution after 21 days of immersion. Furthermore, the absence of cytotoxicity, verified through cell viability assays with MC3T3-E1 osteoblastic cells, proves the biocompatibility of the coatings