Melt electrowritten scaffolds containing fluorescent nanodiamonds for improved mechanical properties and degradation monitoring

Biocompatible fluorescent nanodiamonds (FNDs) were introduced into polycaprolactone (PCL) – the golden standard material in melt electrowriting (MEW). MEW is an advanced additive manufacturing technique capable of depositing high-resolution micrometric fibres. Due to the high printing precision, MEW finds growing interest in tissue engineering applications. Here, we introduced fluorescent nanodiamonds (FNDs) into polycaprolactone prior to printing to fabricate scaffolds for biomedical applications with improved mechanical properties. Further FNDs offer the possibility of their real-time degradation tracking. Compared to pure PCL scaffolds, the functionalized ones containing 0.001 wt% of 70 nm-diameter nanodiamonds (PCL-FNDs) showed increased tensile moduli (1.25 fold) and improved cell proliferation during 7-day cell cultures (2.00 fold increase). Furthermore, the addition of FNDs slowed down the hydrolytic degradation process of the scaffolds, accelerated for the purpose of the study by addition of the enzyme lipase to deionized water. Pure PCL scaffolds showed obvious signs of degradation after 3 h, not observed for PCL-FNDs scaffolds during this time. Additionally, due to the nitrogen-vacancy (NV) centers present on the FNDs, we were able to track their amount and location in real-time in printed fibres using confocal microscopy. This research shows the possibility for high-resolution life-tracking of MEW PCL scaffolds’ degradation.</p

Elastomeric Optical Waveguides by Extrusion Printing

Advances in optogenetics and the increasing use of implantable devices for therapies and health monitoring are driving demand for compliant, biocompatible optical waveguides and scalable methods for their manufacture. Molding, thermal drawing, and dip-coating are the most prevalent approaches in recent literature. Here the authors demonstrate that extrusion printing at room temperature can be used for continuous fabrication of compliant optical waveguides with polydimethylsiloxane (PDMS) core and crosslinked Pluronic F127-diacrylate (Pluronic-DA) cladding. The optical fibers are printed from fluid precursor inks and stabilized by physical interactions and photoinitiated crosslinking in the Pluronic-DA. The printed fibers show optical loss values of 0.13–0.34 dB cm–1 in air and tissue within the wavelength range of 405–520 nm. The fibers have a Young's Modulus (Pluronic cladding) of 150 kPa and can be stretched to more than 5 times their length. The optical loss of the fibers shows little variation with extension. This work demonstrates how printing can simplify the fabrication of compliant and stretchable devices from materials approved for clinical use. These can be of interest for optogenetic or photopharmacology applications in extensible tissues, like muscles or heart

Development of bioactive catechol functionalized nanoparticles applicable for 3D bioprinting

Efficient wound treatments to target specific events in the healing process of chronic wounds constitute a significant aim in regenerative medicine. In this sense, nanomedicine can offer new opportunities to improve the effectiveness of existing wound therapies. The aim of this study was to develop catechol bearing polymeric nanoparticles (NPs) and to evaluate their potential in the field of wound healing. Thus, NPs wound healing promoting activities, potential for drug encapsulation and controlled release, and further incorporation in a hydrogel bioink formulation to fabricate cell-laden 3D scaffolds are studied. NPs with 2 and 29 M % catechol contents (named NP2 and NP29) were obtained by nanoprecipitation and presented hydrodynamic diameters of 100 and 75 nm respectively. These nanocarriers encapsulated the hydrophobic compound coumarin-6 with 70% encapsulation efficiency values. In cell culture studies, the NPs had a protective effect in RAW 264.7 macrophages against oxidative stress damage induced by radical oxygen species (ROS). They also presented a regulatory effect on the inflammatory response of stimulated macrophages and promoted upregulation of the vascular endothelial growth factor (VEGF) in fibroblasts and endothelial cells. In particular, NP29 were used in a hydrogel bioink formulation using carboxymethyl chitosan and hyaluronic acid as polymeric matrices. Using a reactive mixing bioprinting approach, NP-loaded hydrogel scaffolds with good structural integrity, shape fidelity and homogeneous NPs dispersion, were obtained. The in vitro catechol NPs release profile of the printed scaffolds revealed a sustained delivery. The bioprinted scaffolds supported viability and proliferation of encapsulated L929 fibroblasts over 14 days. We envision that the catechol functionalized NPs and resulting bioactive bioink presented in this work offer promising advantages for wound healing applications, as they: 1) support controlled release of bioactive catechol NPs to the wound site; 2) can incorporate additional therapeutic functions by co-encapsulating drugs; 3) can be printed into 3D scaffolds with tailored geometries based on patient requirements

Melt Electrowriting of Graded Porous Scaffolds to Mimic the Matrix Structure of the Human Trabecular Meshwork

The permeability of the human trabecular meshwork (HTM) regulates eye pressure via a porosity gradient across its thickness modulated by stacked layers of matrix fibrils and cells. Changes in HTM porosity are associated with increases in intraocular pressure and the progress of diseases such as glaucoma. Engineered HTMs could help to understand the structure-function relation in natural tissues and lead to new regenerative solutions. Here, melt electrowriting (MEW) is explored as a biofabrication technique to produce fibrillar, porous scaffolds that mimic the multilayer, gradient structure of native HTM. Poly(caprolactone) constructs with a height of 125-500 μm and fiber diameters of 10-12 μm are printed. Scaffolds with a tensile modulus between 5.6 and 13 MPa and a static compression modulus in the range of 6-360 kPa are obtained by varying the scaffold design, that is, the density and orientation of the fibers and number of stacked layers. Primary HTM cells attach to the scaffolds, proliferate, and form a confluent layer within 8-14 days, depending on the scaffold design. High cell viability and cell morphology close to that in the native tissue are observed. The present work demonstrates the utility of MEW for reconstructing complex morphological features of natural tissues

Melt Electrowriting of Scaffolds with a Porosity Gradient to Mimic the Matrix Structure of the Human Trabecular Meshwork

The permeability of the Human Trabecular Meshwork (HTM) regulates eye pressure via a porosity gradient across its thickness modulated by stacked layers of matrix fibrils and cells. Changes in HTM porosity are associated with increases in intraocular pressure and the progress of diseases like glaucoma. Engineered HTMs could help to understand the structure-function relation in natural tissues, and lead to new regenerative solutions. Here, melt electrowriting (MEW) is explored as a biofabrication technique to produce fibrillar, porous scaffolds that mimic the multilayer, gradient structure of native HTM. Poly(caprolactone) constructs with a height of 125-500 μm and fiber diameters of 10-12 μm are printed. Scaffolds with a tensile modulus between 5.6 and 13 MPa, and a static compression modulus in the range of 6-360 kPa are obtained by varying the scaffolds design, i.e., density and orientation of the fibers and number of stacked layers. Primary HTM cells attach to the scaffolds, proliferate, and form a confluent layer within 8-14 days, depending on the scaffold design. High cell viability and cell morphology close to that in the native tissue are observed. The present work demonstrates the utility of MEW to reconstruct complex morphological features of natural tissues

Electrically Conductive and Highly Stretchable Piezoresistive Polymer Nanocomposites via Oxidative Chemical Vapor Deposition

Electrically conductive polymer nanocomposites have been the subject of intense research due to their promising potential as piezoresistive biomedical sensors, leveraging their flexibility and biocompatibility. Although intrinsically conductive polymers such as polypyrrole (PPy) and polyaniline have emerged as lucrative candidates, they are extremely limited in their processability by conventional solution-based approaches. In this work, ultrathin nanostructured coatings of doped PPy are realized on polyurethane films of different architectures via oxidative chemical vapor deposition to develop stretchable and flexible resistance-based strain sensors. Holding the substrates perpendicular to the reactant flows facilitates diffusive transport and ensures excellent conformality of the interfacial integrated PPy coatings throughout the 3D porous electrospun fiber mats in a single step. This allows the mechanically robust (stretchability &gt; 400%, with fatigue resistance up to 1000 cycles) nanocomposites to elicit a reversible change of electrical resistance when subjected to consecutive cycles of stretching and releasing. The repeatable performance of the strain sensor is linear due to dimensional changes of the conductive network in the low-strain regime (ϵ ≤ 50%), while the evolution of nano-cracks leads to an exponential increase, which is observed in the high-strain regime, recording a gauge factor as high as 46 at 202% elongational strain. The stretchable conductive polymer nanocomposites also show biocompatibility toward human dermal fibroblasts, thus providing a promising path for use as piezoresistive strain sensors and finding applications in biomedical applications such as wearable, skin-mountable flexible electronics.</p

Breaking through the barrier: Modelling and exploiting the physical microenvironment to enhance drug transport and efficacy

Pharmaceutical compounds are the main pillar in the treatment of various illnesses. To administer these drugs in the therapeutic setting, multiple routes of administration have been defined, including ingestion, inhalation, and injection. After administration, drugs need to find their way to the intended target for high effectiveness, and this penetration is greatly dependent on obstacles the drugs encounter along their path. Key hurdles include the physical barriers that are present within the body and knowledge of those is indispensable for progress in the development of drugs with increased therapeutic efficacy. In this review, we examine several important physical barriers, such as the blood-brain barrier, the gut-mucosal barrier, and the extracellular matrix barrier, and evaluate their influence on drug transport and efficacy. We explore various in vitro model systems that aid in understanding how parameters within the barrier model affect drug transfer and therapeutic effect. We conclude that physical barriers in the body restrict the quantity of drugs that can pass through, mainly as a consequence of the barrier architecture. In addition, the specific physical properties of the tissue can trigger intracellular changes, altering cell behavior in response to drugs. Though the barriers negatively influence drug distribution, physical stimulation of the surrounding environment may also be exploited as a mechanism to control drug release. This drug delivery approach is explored in this review as a potential alternative to the conventional ways of delivering therapeutics

Thiol-Methylsulfone Based Hydrogels: Enhanced Control on Gelation Kinetics for 3D Cell Encapsulation

Hydrogels are useful temporal matrices for cell culture technologies. The successful mixing and encapsulation of cells within the gel requires the selection of efficient and cytocompatible gelation reactions occurring in the minute timescale under physiological conditions. The thiol-methylsulfonyl (MS) chemical reaction is introduced here as a novel chemistry to encapsulate cells in polymeric matrices. Thiol-MS crosslinking does not require a light activation step and can occur within the seconds-to-minutes timescale by adjusting the pH in the physiological range 8.0-6.6. This reaction is cytocompatible and the reaction product is hydrolytically stable in cell culture media up to 4 weeks. Cell encapsulation protocols enabling comfortable handling and yielding homogenous distribution of the embedded cells are described. All these features are relevant for the application of this crosslinking reaction to biomedical scenarios. Finally, this manuscript also compares the performance of thiol-MS hydrogels with the established thiol-maleimide and thiol-vinylsulfone hydrogels. The benefit of thiol-MS crosslinking in terms of control over hydrogelation kinetics is demonstrated

Chitosan-based inks: 3D printing and bioprinting strategies to improve shape fidelity, mechanical properties, and biocompatibility of 3D scaffolds

El campo de la impresión y la bioimpresión 3D ha alcanzado un gran desarrollo en los últimos años. La llegada de estas técnicas de biofabricación a la ingeniería tisular han supuesto una revolución debido a las estructuras biomiméticas tan complejas que permiten sintetizar. El uso del quitosano como componente de tintas basadas en polímeros naturales es muy atractivo debido a las propiedades beneficiosas que presenta (alta biocompatibilidad, biodegradabilidad, bajo coste). Sin embargo, su aplicación se ve limitada por su baja solubilidad en condiciones fisiológicas y sus pobres propiedades mecánicas. En este artículo, revisamos el estado del arte relacionado con las estrategias actuales de impresión 3D que hacen uso de tintas y biotintas basadas en quitosano para el desarrollo de soportes biomiméticos. También analizamos las estrategias de entrecruzamiento que se aplican actualmente para mejorar su printabilidad, resaltando además los derivados de quitosano disponibles que permiten encapsular células para bioimpresión 3D, incluyendo además nuestra contribución al campo. Prevemos que el uso de quitosano en la impresión 3D aumentará significativamente en los próximos años gracias a los esfuerzos que se están desarrollando en términos de mecanismos de gelificación y derivados de quitosano que permiten la encapsulación celular.El campo de la impresión y la bioimpresión 3D ha alcanzado un gran desarrollo en los últimos años. La llegada de estas técnicas de biofabricación a la ingeniería tisular han supuesto una revolución debido a las estructuras biomiméticas tan complejas que permiten sintetizar. El uso del quitosano como componente de tintas basadas en polímeros naturales es muy atractivo debido a las propiedades beneficiosas que presenta (alta biocompatibilidad, biodegradabilidad, bajo coste). Sin embargo, su aplicación se ve limitada por su baja solubilidad en condiciones fisiológicas y sus pobres propiedades mecánicas. En este artículo, revisamos el estado del arte relacionado con las estrategias actuales de impresión 3D que hacen uso de tintas y biotintas basadas en quitosano para el desarrollo de soportes biomiméticos. También analizamos las estrategias de entrecruzamiento que se aplican actualmente para mejorar su printabilidad, resaltando además los derivados de quitosano disponibles que permiten encapsular células para bioimpresión 3D, incluyendo además nuestra contribución al campo. Prevemos que el uso de quitosano en la impresión 3D aumentará significativamente en los próximos años gracias a los esfuerzos que se están desarrollando en términos de mecanismos de gelificación y derivados de quitosano que permiten la encapsulación celular.Authors thank financial support to Mi- nistry of Science, Innovation and Universi- ties (Spain) (MAT2017-84277-R), “La Caixa” Foundation (ID 100010434, scho- larship of Ana Mora-Boza, code LCF/BQ/ ES16/11570018) and DAAD Research Grants- Short-term grants 201

Development of bioactive catechol functionalized nanoparticles applicable for 3D bioprinting

Efficient wound treatments to target specific events in the healing process of chronic wounds constitute a significant aim in regenerative medicine. In this sense, nanomedicine can offer new opportunities to improve the effectiveness of existing wound therapies. The aim of this study was to develop catechol bearing polymeric nanoparticles (NPs) and to evaluate their potential in the field of wound healing. Thus, NPs wound healing promoting activities, potential for drug encapsulation and controlled release, and further incorporation in a hydrogel bioink formulation to fabricate cell-laden 3D scaffolds are studied. NPs with 2 and 29 M % catechol contents (named NP2 and NP29) were obtained by nanoprecipitation and presented hydrodynamic diameters of 100 and 75 nm respectively. These nanocarriers encapsulated the hydrophobic compound coumarin-6 with 70% encapsulation efficiency values. In cell culture studies, the NPs had a protective effect in RAW 264.7 macrophages against oxidative stress damage induced by radical oxygen species (ROS). They also presented a regulatory effect on the inflammatory response of stimulated macrophages and promoted upregulation of the vascular endothelial growth factor (VEGF) in fibroblasts and endothelial cells. In particular, NP29 were used in a hydrogel bioink formulation using carboxymethyl chitosan and hyaluronic acid as polymeric matrices. Using a reactive mixing bioprinting approach, NP-loaded hydrogel scaffolds with good structural integrity, shape fidelity and homogeneous NPs dispersion, were obtained. The in vitro catechol NPs release profile of the printed scaffolds revealed a sustained delivery. The bioprinted scaffolds supported viability and proliferation of encapsulated L929 fibroblasts over 14 days. We envision that the catechol functionalized NPs and resulting bioactive bioink presented in this work offer promising advantages for wound healing applications, as they: 1) support controlled release of bioactive catechol NPs to the wound site; 2) can incorporate additional therapeutic functions by co-encapsulating drugs; 3) can be printed into 3D scaffolds with tailored geometries based on patient requirements.Authors thank CIBER-BBN (Spain) and the Spanish Ministry of Economy and Competitivity (M. Puertas-Bartolomé scholarship BES-2015-075161) and the Spanish Ministry of Science and Innovation (PID2020-114086RB-100) for supporting this work. The authors acknowledge RegenHu company, and particularly: Sandro Figi, Dominic Ernst, Michael Kuster and Andreas Scheidegger, for the fruitful collaboration, development and providing the mixing tool. The authors thank Dr. Emmanuel Terriac from INM, Germany for assistance in the confocal imaging. B. Vázquez-Lasa is a member of the SusPlast platform from CSI