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In this review, in vitro functional models for human liver diseases and drug testing as an alternative to animal testing are described and discussed, highlighting pros and cons of the current state of the art reported in the scientific literature.

Introduction: For decades, in vitro 2D cell culture techniques have been employed in research, but they fail to recapitulate the complexity of natural tissues. 3D bioprinting could potentially overcome this drawback due to the possibility to control the spatial disposition of living cells and the geometry of the 3D scaffold.Materials and methods: This study reports the design and characterization of a novel bioink for extrusion bioprinting, analyzing different blend formulations composed of alginate, gelatin, and methylcellulose, suitable as cell-laden bioink for lymphoid cells, in particular those isolated from patients with Chronic Lymphocytic Leukemia (CLL). The rheological properties as a function of temperature and the printability of the formulations were investigated to define the optimal printing parameters. In vitro stability of the printed scaffolds was investigated under culture conditions and compression tests were performed on printed and bioprinted scaffolds to compare their mechanical properties with those of fresh lymphoid tissue. Finally, MEC1, a CLL cell line, was bioprinted to investigate cell viability, cell density, and cell capability to be released from the scaffold over time.Results and discussion: Results showed that, for the selected blends, good shape fidelity and printing accuracy were achieved with a limitation on the number of printed layers. Scaffolds withstood culture conditions showing stability for up to 3 weeks and their mechanical properties were similar to those of lymphoid tissues already reported in the literature. High cell viability after 21 days was observed for both MEC1 and primary peripheral mononuclear cells, confirming the possibility to use the selected formulation to successfully bioprint lymphoid cells by possibly mimicking their native lymphoid microenvironment.

Renal tubular cells frequently lose differentiation markers and physiological properties when propagated in conventional cell culture conditions. Embedding cells in 3D microenvironments or controlling their 3D assembly by bioprinting can enhance their physiological properties, which is beneficial for modeling diseases in vitro. A potential cellular source for modeling renal tubular physiology and kidney diseases in vitro are directly reprogrammed induced renal tubular epithelial cells (iRECs). iRECs were cultured in various biomaterials and as bioprinted tubular structures. They showed high compatibility with the embedding substrates and dispensing methods. The morphology of multicellular aggregates was substantially influenced by the 3D microenvironment. Transcriptomic analyses revealed signatures of differentially expressed genes specific to each of the selected biomaterials. Using a new cellular model for autosomal-dominant polycystic kidney disease, Pkd1(−/−) iRECs showed disrupted morphology in bioprinted tubules and a marked upregulation of the Aldehyde dehydrogenase 1a1 (Aldh1a1). In conclusion, 3D microenvironments strongly influence the morphology and expression profiles of iRECs, help to unmask disease phenotypes, and can be adapted to experimental demands. Combining a direct reprogramming approach with appropriate biomaterials will facilitate construction of biomimetic kidney tubules and disease models at the microscale.

In this study, a new edible coating material with enhanced mechanical and gas barrier properties was studied by coupling silk fibroin (SF) with poly(vinyl alcohol) (PVOH). SF and PVOH water suspens...

Author(s): Ghalayani Esfahani, A; Lazazzera, B; Draghi, L; Fare, S; Chiesa, R; De Nardo, L; Billi, F | Abstract: AimsThe aim of this study was to develop a new class of gallium (Ga)-doped chitosan (CS) coatings fabricated by electrophoretic deposition (EPD) in staphylococcal infection therapy.Methods and resultsBiofilm formation on EPD CS/Ga coatings by Staphylococcus epidermidis and Staphylococcus aureus, which are the main strains involved in postarthroplasty infections, was assessed. The codeposition of an antibacterial agent was effective; Ga loaded into CS matrix reduces biofilm viability by up to 86% and 80% for S. epidermidis and S. aureus strains respectively. Lastly, the influence of pulsed electromagnetic field (PEMF) on the bactericidal activity of CS/Ga coatings was investigated in vitro. To this end, the coatings were incubated with S. epidermidis and S. aureus and exposed to the PEMF using two different frequencies and times. Biofilm viability for S. epidermidis was decreased by 35-40% in the presence of low-frequency (LF) and high-frequency (HF) PEMF respectively. Biofilm viability by S. aureus was not further reduced in the presence of LF PEMF, but decreased by 38% at HF PEMF.ConclusionsThis study has established that a combination of PEMFs with the antibacterial agent improves bactericidal activity of Ga against S. epidermidis strain 14990 and S. aureus strain 12600.Significance and impact of the studyThis new integrated approach could reduce the incidence of infection in orthopaedic implant applications. It also clearly demonstrates that the combination of Ga treatment with PEMF could aid biofilm-associated infection therapy due to improved Ga efficiency.

Main problems related to the adoption of magnesium alloys for temporary orthopedic prostheses manufacturing are (i) the need of an efficient production process and (ii) the high corrosion rate compared with the bone healing time. In this work, the single-point incremental forming (SPIF) process, an effective and flexible solution for manufacturing very small batches even composed by one piece, was investigated. Tests were conducted on AZ31B-H24 sheets and were aimed at understanding the effect of temperature on the mechanical characteristics (microstructure, hardness, and roughness) of the sheet after the above-mentioned forming process and their correlation with both the corrosion rate and the cytocompatibility. In addition, after the forming process, samples processed by SPIF were coated by electrospun polycaprolactone (PCL) to reduce the corrosion rate and to further improve the cytocompatibility. Grain refinement was achieved thanks to the combined effect of temperature and strain rate during forming and finer grain size resulted to improve the magnesium corrosion resistance. In simulated body fluids, the electrospun PCL-coated samples exhibited a slower pH increase compared with uncoated samples. No indirect cytotoxic effects were detected in vitro for MC3T3-E1 cells for both coated and uncoated samples. However, cells colonization was observed only on electrospun PCL-coated samples, suggesting the importance of the polymeric coating in promoting the adhesion and survival of seeded MC3T3-E1 cells on the implant surface.

Methylcellulose (MC) hydrogels have been successfully proposed in the field of cell sheet engineering (CSE), allowing cell detachment from their surface by lowering the temperature below their transition temperature (Tt). Among the main limitations of pristine MC hydrogels, low physical stability and mechanical performances limit the breadth of their potential applications. In this study, a crosslinking strategy based on citric acid (CA) was used to prepare thermoresponsive MC hydrogels, with different degrees of crosslinking, to exploit their possible use as substrates in CSE. The investigated amounts of CA did not cause any cytotoxic effect while improving the mechanical performance of the hydrogels (+11-fold increase in E, compared to control MC). The possibility to obtain cell sheets (CSs) was then demonstrated using murine fibroblast cell line (L929 cells). Cells adhered on crosslinked MC hydrogels’ surface in standard culture conditions and then were harvested at selected time points as single CSs. CS detachment was achieved simply by lowering the external temperature below the Tt of MC. The detached CSs displayed adhesive and proliferative activity when transferred to new plastic culture surfaces, indicating a high potential for regenerative purposes.

Aerogels are commonly regarded as soft and versatile materials for multiple applications thanks to their features such as elastic behavior, swelling ability and responsive characteristics. In the last decades these qualities have ensured them multiple uses in biomedical field as controlled drug delivery systems and scaffolds for tissue engineering applications. Recently their employment as sensors has attracted great interest especially thanks to their tunability and the possibility to be used as soft material instead of conventional and many times unreliable sensors in multiple sensing applications. In this work we synthetized graphene oxide/polyaniline aerogels by in situ chemical polymerization of aniline monomer in aqueous dispersion of graphene oxide sheets. We characterized the system through chemical and mechanical analysis, and we its responsivity as gas sensing device was verified. Moreover, we investigated the influence of the encapsulation of graphene nanoplatelets on the responsive ability of the device. The addition of the graphene moieties improves the conductivity of the system, its compressive mechanical resistance and the applicability of these devices as gas sensors.

It is estimated that 10% of the worldwide population suffers from chronic kidney disease (CKD) with a rising tendency. Patients with CKD have limited treatment options and novel therapies that could halt or even reverse the progression of CKD are urgently needed.Bioprinting is considered one of the most promising approaches to generate novel 3D in vitro models and organ-like constructs to investigate underlying pathomechanism of kidney diseases. This study aimed at establishing a method to isolate primary renal cells in an easy and reproducible way. These cells were used in a new bioprinting platform laying the foundation for the development of a 3D renal tubulointerstitium model for in vitro studies. Primary murine tubular (pmTECs), endothelial and fibroblast cells were successfully isolated, but further optimization is required for the culture and expansion of primary endothelial cells. Therefore, an endothelial cell line (HUVECs) and pmTECs were combined with polysaccharide biomaterial ink solutions and processed with a microfluidic 3D bioprinter, leading to high cell viability and metabolic activity. Core-shell bioprinted constructs with HUVECs and pmTECs were manufactured mimicking tubules.In conclusion, microfluidic bioprinting strategy could be used to build a novel 3D kidney in vitro model.

Methylcellulose (MC) is an attractive material used to produce thermo-responsive hydrogels. They undergo sol-gel transition when a critical temperature is reached, thus modifying their properties (e.g., physicochemical and mechanical) in response to temperature changes. This behavior is particularly attractive when the body temperature acts as a trigger to modulate the thermo-responsive behavior of MC hydrogels. In this regard, exciting advances have been achieved in the field of cell and drug delivery, tissue engineering, and regenerative medicine, making MC a very attractive and versatile biomaterial. This review aims to present MC hydrogels, examining their preparation, physical properties, and tunability of thermal response, lastly moving to a comprehensive depiction of both their conventional and innovative applications for tissue regeneration purposes. In particular, three main families of applications are introduced: (1) in situ gelling systems, which undergo sol-gel transition upon delivery into a target site (e.g., tissue or organ), assisting the regeneration of the latter both in the presence or absence of loading components (e.g., cells, biomolecules, and inorganic materials); (2) three-dimensional (3D) (bio)printing, where the sol-gel transition is induced by heating MC-based (bio)inks after printing, obtaining 3D tissue-engineered substitutes with defined geometries and high shape fidelity; (3) smart culture surfaces, where the hydrophilic/hydrophobic transition of MC is exploited to reach a selective attachment/detachment of cells, offering the possibility to obtain cell sheets and cell bodies for tissue reconstruction without the need of any proteolytic enzyme. The main limitations of MC hydrogels will be then examined, together with current solutions to overcome them. Moreover, an overview of the future directions in the field of MC smart hydrogels will be given, with particular focus on the design of multiresponsive systems capable to respond to multiple stimuli (e.g., chemical and biological stimuli), toward the development of more patient-specific treatments. Finally, an overview of the patents and clinical trials describing the use of MC will be given, retracing the abovementioned families of application.

3D scaffolds and 2D matrices fabricated by electrospinnig show morphology similar to that of native ECM, however their mechanical and biological properties are often inadequate, particularly in applications in contact with blood, e.g. in blood vessel substitutes. Biocoatings can improve the performance of these substrates, in particular cross-linked gelatin is among the most used substances. In this work, a gelatin coating was applied to electrospun silk fibroin (ESF) mats and tubes intended for the regeneration of cardiovascular tissues. The crosslinking reaction used is based on a Michael-type addition in water that promotes the formation of covalent bonds between gelatin amino groups and β-carbons of N-N’-methylene bis-acrylamide (MBA)[1]. Interestingly, when the reacting mixture is applied to a substrate containing primary or even secondary amino groups, these groups can participate in the reaction, being incorporated into the gelatin coating, thus increasing the coating stability on the surface. ESF mats and tubes, obtained as described in [2] were coated with gelatin MBA-crosslinked in situ by loading or dipping the ESF samples with the crosslinking solution, by use of static or dynamic home-made systems. SEM analysis on coated samples showed a homogeneous coating with gelatin penetrating the whole thickness of the SF matrix {»120 µm for mats and » 212 µm for tubes), with an increase of thickness of about 40% in wet conditions. Water uptake tests indicated for coated samples a faster and higher swelling (1600% after 14 days) than not coated ones (500%), due to the presence of gelatin. Tensile mechanical tests showed higher values of ultimate stress and elastic modulus for silk fibroin samples (sb=2.4, E=1.82 MPa) compared to gelatin-coated ones (sb=1.2, E=0.58 MPa), with not significant differences in the ultimate deformation (»150%). Indirect cytocompatibility tests, performed by culturing L929 cells in the presence of eluates obtained by immersing coated and uncoated samples up to 7 days in culture medium, demonstrated a cell viability higher than the control. In direct contact tests using L929 cells, a good cytocompatibility was demonstrated by both coated and uncoated ESF samples, with a cell viability increasing with the culture time (up to 7 days) and a flattened and stretched morphology at SEM. Primary human umbilical vein endothelial cell (HUVEC), obtained by enzymatic digestion (Cittadella Hospital, PD,I) were seeded onto ESF and ESF-coated samples and cultured under standard tissue culture conditions. Cell adhesion on the matrices was analysed by OM after fixing with formalin and staining with toluidine blue. After 7 days from seeding, cell proliferation was evaluated by a protein assay (BCA Protein Assay kit) and the results indicated a significantly higher (p

Bioactive glasses (BGs), due to their ability to influence osteogenic cell functions, have become attractive materials to improve loaded and unloaded bone regeneration. BG systems can be easily doped with several metallic ions (e.g., Ag, Sr, Cu, Nb) in order to confer antibacterial properties. In particular, Nb, when compared with other metal ions, has been reported to be less cytotoxic and possess the ability to enhance mineralization process in human osteoblast populations. In this study, we co-deposited, through one-pot electrophoretic deposition (EPD), chitosan (CS), gelatin (GE) and a modified BG containing Nb to obtain substrates with antibacterial activity for unloaded bone regeneration. Self-standing composite scaffolds, with a defined porosity (15–90 μm) and homogeneous dispersion of BGs were obtained. TGA analysis revealed a BG loading of about 10% in the obtained scaffolds. The apatite formation ability of the scaffolds was evaluated in vitro in simulated body fluid (SBF). SEM observations, XRD and FT-IR spectra showed a slow (21–28 days) yet effective nucleation of CaP species on BGs. In particular, FT-IR peak around 603 cm−1 and XRD peak at 2θ = 32°, denoted the formation of a mineral phase after SBF immersion. In vitro biological investigation revealed that the release of Nb from composite scaffolds had no cytotoxic effects. Interestingly, BG-doped Nb scaffolds displayed antibacterial properties, reducing S. lutea and E. coli growth of ≈60% and ≈50%, respectively. Altogether, the obtained results disclose the produced composite scaffolds as promising materials with inherent antibacterial activity for bone tissue engineering applications.

Stable hydrogels with tunable rheological properties were prepared by adding Ca2+ ions to aqueous dispersions of 2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO)-oxidized and ultra-sonicated cellulose nanofibers (TOUS-CNFs). The gelation occurred by interaction among polyvalent cations and the carboxylic units introduced on TOUS-CNFs during the oxidation process. Both dynamic viscosity values and pseudoplastic rheological behaviour increased by increasing the Ca2+ concentration, confirming the cross-linking action of the bivalent cation. The hydrogels were proved to be suitable controlled release systems by measuring the diffusion coefficient of a drug model (ibuprofen, IB) by high-resolution magic angle spinning (HR-MAS) nuclear magnetic resonance (NMR) spectroscopy. IB was used both as free molecule and as a 1:1 pre-formed complex with &beta, cyclodextrin (IB/&beta, CD), showing in this latter case a lower diffusion coefficient. Finally, the cytocompatibility of the TOUS-CNFs/Ca2+ hydrogels was demonstrated in vitro by indirect and direct tests conducted on a L929 murine fibroblast cell line, achieving a percentage number of viable cells after 7 days higher than 70%.

Chondral and osteochondral lesions represent one of the most challenging problems in the orthopedic field, as these types of injuries lead to disability and worsened quality of life for patients and have an economic impact on the healthcare system. The aim of this in vivo study was to develop a new tissue engineering approach through a hybrid scaffold for osteochondral tissue regeneration made of porous polyurethane foam (PU) coated under vacuum with calcium phosphates (PU/VAC). Scaffold characterization showed a highly porous and interconnected structure. Human amniotic mesenchymal stromal cells (hAMSCs) were loaded into scaffolds using pectin (PECT) as a carrier. Osteochondral defects in medial femoral condyles of rabbits were created and randomly allocated in one of the following groups: plain scaffold (PU/VAC), scaffold with hAMSCs injected in the implant site (PU/VAC/hAMSC), scaffold with hAMSCs loaded in pectin (PU/VAC/PECT/hAMSC), and no treated defects (untreated). The therapeutic efficacy was assessed by macroscopic, histological, histomorphometric, microtomographic, and ultrastructural analyses at 3, 6, 12, and 24 weeks. Histological results showed that the scaffold was permissive to tissue growth and penetration, an immature osteocartilaginous tissue was observed at early experimental times, with a more accentuated bone regeneration in comparison with the cartilage layer in the absence of any inflammatory reaction.

In-situ Raman spectroscopy was employed to investigate the thermo-responsive sol-gel transition of methylcellulose (MC) based hydrogels at their lower critical solution temperature (LCST). By comparing the Raman signature of dry and wet MC samples, C-Hx bands in the 2700–3100 cm−1 range (associated with vibrations of CH3 bonds) were found to respond to the hydration level of MC. In particular, the intensity of the C-Hx peaks of wet samples was demonstrated to depend on temperature variations (25–50 °C). Data fitting allowed to identify the LCST (~39 °C) , thereby disclosing the potential of Raman spectroscopy for the precise quantification of the thermo-responsive behavior of MC hydrogels.

Methylcellulose (MC) hydrogels, undergoing sol-gel reversible transition upon temperature changes, lend themselves to smart system applications. However, their reduced stability in aqueous environment and unsatisfactory mechanical properties limit the breadth of their possible applications. Here, a crosslinking strategy based on citric acid (CA) was developed: exploiting three crosslinking parameters (CA concentration, crosslinking time, and crosslinking temperature) by a design of experiment approach, optimized crosslinked MC hydrogels (MC-L, MC-M, MC-H) were obtained and characterized. Swelling tests in water revealed the effectiveness of CA crosslinking in modulating the water uptake of MC hydrogels. Both theoretical and experimental analyses showed an increase in the crosslinking density by the rationale selection of process parameters. The extent of sol-gel transition was assessed by swelling tests, Raman spectroscopy and rheological analyses. MC-M samples demonstrated to preserve their thermo-responsive behavior around their lower critical solution temperature (LCST), while showing increased stability and enhanced mechanical properties when compared to pristine MC hydrogels.

The regeneration of injured or damaged tissues by cell delivery approaches requires the fabrication of cell carriers (e.g., microspheres, MS) that allow for cell delivery to limit cells spreading from the injection site. Ideal MS for cell delivery should allow for cells adhesion and proliferation on the MS before the injection, while they should allow for viable cells release after the injection to promote the damaged tissue regeneration. We optimized a water-in-oil emulsion method to obtain gelatin MS crosslinked by methylenebisacrylamide (MBA). The method we propose allowed obtaining spherical, chemically crosslinked MS characterized by a percentage crosslinking degree of 74.5 ± 2.1%. The chemically crosslinked gelatin MS are characterized by a diameter of 70.9 ± 17.2 μm in the dry state and, at swelling plateau in culture medium at 37 °C, by a diameter of 169.3 ± 41.3 μm. The MS show dimensional stability up to 28 days, after which they undergo complete degradation. Moreover, during their degradation, MS release gelatin that can improve the engraftment of cells in the injured site. The produced MS did not induce any cytotoxic effect in vitro and they supported viable L929 fibroblasts adhesion and proliferation. The MS released viable cells able to colonize and proliferate on the tissue culture plastic, used as release substrate, potentially proving their ability in supporting a simplified in vitro wound healing process, thus representing an optimal tool for cell delivery applications.

Despite their outstanding potential and the success that has already been achieved with three-dimensional (3D) printed hydrogel scaffolds, there has been little investigation into their application in the regeneration of damaged or missing adipose tissue (AT). Due to their macroscopic shape, microarchitecture, extracellular matrix-mimicking structure, degradability and soft tissue biomimetic mechanical properties, 3D printed hydrogel scaffolds have great potential for use in aesthetic, structural and functional restoration of AT. Here, we propose a simple and cost-effective 3D printing strategy using gelatin-based ink to fabricate scaffolds suitable for AT engineering. The ink, successfully printed here for the first time, was prepared by mixing gelatin and methylenebisacrylamide (a crosslinker) to initiate the crosslinking reaction. The solution was then loaded into the cartridge (temperature T = 35 °C) of a pneumatic extrusion-based 3D printer and printed on a cooled surface (T = 4 °C) in the appropriate time window for ink printability as verified by rheological tests. Subsequently, the printed gelatin hydrogels were crosslinked at different temperatures to optimize their stability and fix the printed structure. The gelatin scaffolds crosslinked at 20 °C remained stable for 21 days at physiological temperature, with compressive mechanical properties mimicking those of AT (i.e. elastic modulus = 20 kPa). The 3D printed scaffolds showed no indirect cytotoxic effects on a 3T3-L1 pre-adipocyte cell line. Moreover, the printed scaffolds successfully promoted adhesion and proliferation of primary human pre-adipocytes, as demonstrated by LIVE/DEAD staining and Alamar Blue assay. The differentiation of primary human pre-adipocytes isolated from three different donors according to adipogenic phenotype was demonstrated by an increase in PPARγ gene expression detected by real-time PCR and accumulated lipid droplets stained by Oil Red O, thus proving the potential of the 3D printed gelatin hydrogels as scaffolds for AT engineering.

Among nanocelluloses, bacterial nanocellulose (BNC) has proven to be a promising candidate in a range of biomedical applications, from topical wound dressings to tissue-engineering scaffolds. Chemical modifications and incorporation of bioactive molecules have been obtained, further increasing the potential of BNC. This study describes the incorporation of vancomycin and ciprofloxacin in BNC and in modified BNC to afford bioactive BNCs suitable for topical wound dressings and tissue-engineering scaffolds. BNC was modified by grafting glycidylmethacrylate (GMA) and further cross-linking with ethylene glycol dimethacrylate (EGDMA) with the formation of stable C&ndash, C bonds though a radical Fenton-type process that involves generation of cellulose carbon centred radicals scavenged by methacrylate structures. The average molar substitution degree MS (MS = methacrylate residue per glucose unit, measured by Fourier transform infrared (FT&ndash, IR) analysis) can be modulated in a large range from 0.1 up to 3. BNC-GMA, BNC-EGDMA and BNC-GMA-EGDMA maintain the hydrogel status until MS reaches the value of 1. The mechanical stress resistance increase of BNC-GMA and BNC-GMA-EGDMA of MS around 0.8 with respect to BNC suggests that they can be preferred to BNC for tissue-engineering scaffolds in cases where the resistance plays a crucial role. BNC, BNC-GMA, BNC-EGDMA and BNC-GMA-EGDMA were loaded with vancomycin (VC) and ciprofloxacin (CP) and submitted to release experiments. BNC-GMA-EGDMA of high substitution degree (0.7&ndash, 1) hold up to 50 percentage of the loaded vancomycin and ciprofloxacin amount, suggesting that they can be further investigated for long-term antimicrobial activity. Furthermore, they were not colonized by Staphylococcus aureus (S.A.) and Klebsiella pneumonia (K.P.). Grafting and cross-linking BNC modification emerges from our results as a good choice to improve the BNC potential in biomedical applications like topical wound dressings and tissue-engineering scaffolds.

Methylcellulose (MC) hydrogels change their affinity to water depending on their temperature and can thus be used as substrates for cell sheet engineering. In this work, we characterize the thermo-responsive properties of 8% w/v MC hydrogels, produced in two saline solutions ( i.e. , Na 2 SO 4 and phosphate buffered saline) at different concentrations, by investigating the rheological properties and the UV-absorbance in function of temperature. Both rheological and UV-spectroscopy tests showed that the addition of salts to MC hydrogels allowed lowering the LCST of the MC hydrogel; moreover, hydrogels produced in 0.1 M Na 2 SO 4 or PBS 20 g/L were proved to be particularly promising for cell sheet engineering application, showing a LCST below 37 °C.

Electrospun PET (ePET) is a promising material for small caliber vascular graft applications owing to its tunable mechanical properties, biocompatibility, and nanofibrous structure that mimic the morphology of natural extracellular matrix. However, the inherent inertness of PET impairs the adhesion and proliferation of endothelial cells on the inner surface of ePET tubular grafts, hindering the formation of a functional endothelium. Gelatin coatings, owing to their ability to promote endothelialization, are a valuable approach to overcome the limitations of ePET. Herein, a novel process for the deposition of stable biomimetic coatings of gelatin on ePET tubular grafts is proposed. Electrospun PET was first aminated by plasma treatment and then coated with a gelatin hydrogel cross-linked in situ by a Michael-type addition reaction. Amination provided a superhydrophilic behavior to the ePET surface, allowing easy gelatin interpenetration along the wall thickness of the tubular structure, and the obtainment of thin coatings that maintained the morphology of ePET fibers. Gelatin coating was stable at long term in a physiological-like environment, noncytotoxic and promoted in vitro cell adhesion and proliferation. Noteworthy, the mechanical properties of gelatin-coated ePET tubular grafts were improved in terms of elastic modulus, compliance, and elastic recoil, finally better matching the characteristics of native blood vessels. Altogether, the proposed coating technique successfully combines the advantages of ePET nanofibrous structure with cross-linked gelatin biological cues and mechanical reinforcement, and emerges as a promising strategy for the development of biocompatible small caliber vascular grafts with superior biomimetic and mechanical properties. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2405-2415, 2017.

Background In vitro dynamic culture conditions play a pivotal role in developing engineered tissue grafts, where the supply of oxygen and nutrients, and waste removal must be permitted within construct thickness. For tubular scaffolds, mass transfer is enhanced by introducing a convective flow through rotating bioreactors with positive effects on cell proliferation, scaffold colonization and extracellular matrix deposition. We characterized a novel polyurethane-based tubular scaffold and investigated the impact of 3 different culture configurations over cell behavior: dynamic (i) single-phase (medium) rotation and (ii) double-phase exposure (medium-air) rotation; static (iii) single-phase static culture as control. Methods A new mixture of polyol was tested to create polyurethane foams (PUFs) as 3D scaffold for tissue engineering. The structure obtained was morphologically and mechanically analyzed tested. Murine fibroblasts were externally seeded on the novel porous PUF scaffold, and cultured under different dynamic conditions. Viability assay, DNA quantification, SEM and histological analyses were performed at different time points. Results The PUF scaffold presented interesting mechanical properties and morphology adequate to promote cell adhesion, highlighting its potential for tissue engineering purposes. Results showed that constructs under dynamic conditions contain enhanced viability and cell number, exponentially increased for double-phase rotation; under this last configuration, cells uniformly covered both the external surface and the lumen. Conclusions The developed 3D structure combined with the alternated exposure to air and medium provided the optimal in vitro biochemical conditioning with adequate nutrient supply for cells. The results highlight a valuable combination of material and dynamic culture for tissue engineering applications.

In this study, the structure and properties of polycaprolactone/ibuprofen (PCL/IBP) rods prepared by melt extrusion were investigated by infrared spectroscopy, X-ray diffraction, scanning electron microscopy, flexural tests, dynamic mechanical analysis and drug release analysis. The crystallinity values for the PCL/IBP rods were lower than that for the pure PCL rods. The PCL/IBP rods had higher values for the flexural modulus compared with the pure PCL rods prepared using the same processing temperature, suggesting that ibuprofen has a hardening effect when dispersion in a PCL matrix. Rods prepared at a processing temperature of 130 °C had the highest flexural modulus and glass transition temperature, probably due to better drug dispersion in the PCL matrix at lower temperature. The surfaces of PCL/IBP rods prepared at 150 °C had small particles and molten drug was deposited on the surface. This is probably due to the low melt temperature of ibuprofen and thus at high temperatures the ibuprofen phase migrates from the PLC matrix to the rod surface. The PCL/IBP rods prepared using different processing temperatures provided different drug release behaviors, with fast or slow drug release depending on the ibuprofen distribution. This feature is of great interest in relation to producing implantable drug delivery rods for acute inflammatory crisis or therapeutic treatments via controlled release.

Adequate porosity, appropriate pore size, and 3D-thick shape are crucial parameters in the design of scaffolds, as they should provide the right space for cell adhesion, spreading, migration, and growth. In this work, a novel design for fabricating a 3D nanostructured scaffold by electrospinning was taken into account. Helical spring-shaped collector was purposely designed and used for electrospinning PCL fibers. Improved morphological properties and more uniform diameter distribution of collected nanofibers on the turns of helical spring-shaped collector are confirmed by SEM analysis. SEM images elaboration showed 3D pores with average diameter of 4 and 5.5 micrometer in x-y plane and z-direction, respectively. Prepared 3D scaffold possessed 99.98% porosity which led to the increased water uptake behavior in PBS at 37°C up to 10 days, and higher degradation rate compared to 2D flat structure. Uniaxial compression test on 3D scaffolds revealed an elastic modulus of 7 MPa and a stiffness of 102 MPa, together with very low hysteresis area and residual strain. In vitro cytocompatibility test with MG-63 osteoblast-like cells using AlamarBlue™ colorimetric assay, indicated a continuous increase in cell viability for the 3D structure over the test duration. SEM observation showed enhanced cells spreading and diffusion into the underneath layers for 3D scaffold. Accelerated calcium deposition in 3D substrate was confirmed by EDX analysis. Obtained morphological, physical, and mechanical properties together with in vitro cytocompatibility results, suggest this novel technique as a proper method for the fabrication of 3D nanofibrous scaffolds for the regeneration of critical-size load bearing defects. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1535-1548, 2017.

Background Until now, environmental sustainability issues are almost entirely unsolved for packaging materials. With the final aim of finding materials with a single recycling channel, cellulose fiber/poly(vinyl)alcohol composites were investigated. Methods After extrusion and injection molding, samples of composite with different cellulose fiber content (30%, 50% and 70% w/w) were tested. Results Tensile mechanical tests exhibited an improvement in composite stiffness when the reinforcement content was increased together with a decrease in composite elongation. Solubility tests performed at room temperature and 45°C showed different behavior depending on the water-resistant film applied on the composite (50% cellulose fiber content). In particular, the uncoated composite showed complete solubility after 2 hours, whereas at the same time point, no solubility occurred when a non-water-soluble varnish was used. Conclusions The proposed composites, named Poly-paper, appear to warrant further investigation as highly sustainable packaging.

Electrospinning provides a simple robust method to manufacture scaffolds for tissue engineering applications. Though varieties of materials can be used, optimization and biocompatibility tests are required to provide functional tissue regeneration. Moreover, many studies are limited to 2D electrospun constructs rather than 3D templates due to the production of high density packed fibres, which result in poor cell infiltration. Here, we optimised electrospinning parameters for three different polymers: poly(epsilon-caprolactone) (PCL), polylactic acid (PLA) and poly(ethylene oxide terephthalate)/poly(butylene terephthalate) (PA) copolymers. Human mesenchymal stromal cells (hMSCs) were cultured on scaffolds for 14 days to study the scaffolds' biocompatibility and their multi-lineage differentiation potential or maintenance of stemness in the absence of chemical stimuli. For all scaffolds, a high and stable metabolic activity was measured throughout the culture time with a high proliferation rate compared to day 1 (PCL 5.8-, PLA 4-, PA 4.9-fold). The metabolism of hMSCs was also measured through glucose and lactate concentrations, showing no cytotoxic levels up to 14 days. Total glycosaminoglycan (GAG) production was the highest in PA electrospun scaffolds. When normalized to DNA, GAG production was the highest in PLA and PA scaffolds. All scaffolds were prone to differentiate to an osteogenic lineage, with PCL providing the highest alkaline phosphatase and collagen type Ia gene upregulation. As PA had the most stable fibre formation, it was chosen as a template to further incorporate stromal cell-derived factor-1 (SDF-1) and granulocyte colony-stimulating factor (G-CSF), and stimulate higher hMSC infiltration. These scaffolds provided significantly higher hMSC infiltration than normal PA scaffolds. In conclusion, our optimized biocompatible electrospun scaffolds have shown promising regulation of hMSC fate. When combined with migratory stimulating cytokines, these scaffolds may overcome the known challenges of poor cellular infiltration typical of micro- and nano-fibrillary random meshes.

In this work an innovative method to obtain hybrid bio-functional scaffolds has been developed. Polyether urethane (PU) foam scaffolds were synthetized by one-step gas foaming process. PU foams were coated with crosslinked gelatin hydrogel to promote cell adhesion and proliferation for the regeneration of soft tissues (e.g., adipose tissue). PU foams were coated with inorganic coating (i.e., CaPs) to improve the interaction with osteoblasts for bone tissue regeneration. The functionalized 3D PU porous scaffolds have been characterized investigating morphological properties by SEM and microCT, water uptake and coating stability, and compressive mechanical properties. Adipose tissue derived stem cells (ADSCs), endothelial cells (MS1), amnion mesenchymal cells (AMCs) and chorion mesenchymal cells (CMCs) isolated from human placenta were in vitro cultured on the hybrid functionalized 3D scaffolds. Mechanical properties showed elastic modulus ranging between 15.75 ± 2.14 and 22.9 ± 3.1 kPa; in vitro biological studies showed good cell adhesion, proliferation, and differentiation. In particular, compared to the results with uncoated PU, when cells where differentiated into adipocytes, Oil red O staining confirmed a higher presence of lipid droplets; in case of osteoblasts differentiation, inorganic extracellular matrix deposition was evidenced on CaPs coated PU. The obtained results suggest the important role of an adequate coating on the scaffold to stimulate a better interaction with cells, promoting the differentiation into different cells phenotypes.