2D and 3D anticancer properties of C2-functionalised glucosamine-Pt (IV) prodrugs based on cisplatin scaffold.

A series of C2-functionalied Pt (IV) glycoconjugates based on glucosamine have been synthesised, characterised and tested as anticancer agents on a series of different 2D and 3D cancer cell lines. The carbohydrate will act as a targeted delivery system to improve the selectivity, exploiting the Warburg Effect and the GLUTs receptors that are overexpressed in most of the cancer cells. The hydroxyl at C2 of the carbohydrates does not participate in hydrogen bonding with the GLUTs receptors, making C2 an attractive position for drug conjugation as seen in literature. In this study, we use the amino functionality at the C2 position in glucosamine and Copper-catalysed Azide-Alkyne Cycloaddition "click" (CuAAC) reaction to connect the prodrug Pt (IV) scaffold to the carbohydrate. We have investigated complexes with different linker lengths, as well as acetyl protected and free derivatives. To the best of our knowledge, this study represents the first series of Pt (IV) glucosamine-conjugates functionalised at C2.

Designing bioinspired multifunctional nanoplatforms to support wound healing and skin regeneration: Mg-hydroxyapatite meets melanins.

Melanin is a multifunctional biological pigment that recently emerged as endowed with anti-inflammatory, antioxidant, and antimicrobial properties and with high potentialities in skin protection and regenerative medicine. Here, a biomimetic magnesium-doped nano-hydroxyapatite (MgHA) was synthesized and decorated with melanin molecules starting from two different monomeric precursors, i.e. 5,6-dihydroxyindole-2-carboxylic acid (DHICA) and dopamine (DA), demonstrating to be able to polymerize on the surface of MgHA nanostructures, thus leading to a melanin coating. This functionalization was realized by a simple and green preparation method requiring mild conditions in an aqueous medium and room temperature. Complementary spectroscopy and electron imaging analyses were carried out to define the effective formation of a stable coating, the percentage of the organic compounds, and the structural properties of resulting melanin-coated nanostructures, which showed good antioxidant activity. The in vitro interaction with a cell model, i.e. mouse fibroblasts, was investigated. The excellent biocompatibility of all bioinspired nanostructures was confirmed from a suitable cell proliferation. Finally, the enhanced biological performances of the nanostructures coated with melanin from DHICA were confirmed by scratch assays. Jointly our findings indicated that low crystalline MgHA and melanin pigments can be efficiently combined, and the resulting nanostructures are promising candidates as multifunctional platforms for a more efficient approach for skin regeneration and protection.

Biomineralization: A new tool for developing eco-sustainable Ti-doped hydroxyapatite-based hybrid UV filters.

It is well known that the prolonged exposure to UV radiation from sunlight can compromise human health and is particularly damaging to the skin, leading to sunburn, photo-aging and skin cancer. Sunscreen formulations containing UV-filters present a barrier against solar UV and help to mitigate the harmful effects however, concern about their safety for both human and environmental health is still a much-debated topic. EC regulations classify UV-filters depending on their chemical nature, particle size, and mechanism of action. Furthermore, it regulates their use in cosmetic products with specific limitations in terms of concentration (organic UV filters) and particle size and surface modification to reduce their photo-activity (mineral UV filters). The regulations have prompted researchers to identify new materials that show promise for use in sunscreens. In this work, biomimetic hybrid materials composed of titanium-doped hydroxyapatite (TiHA) grown on two different organic templates, derived from animal (gelatin - from pig skin) and vegetable (alginate - from algae) sources. These novel materials were developed and characterized to obtain sustainable UV-filters as a safer alternative for both human and ecosystem health. This 'biomineralization' process yielded TiHA nanoparticles that demonstrated high UV reflectance, low photoactivity, good biocompatibility and an aggregate morphology which prevents dermal penetration. The materials are safe for topical application and for the marine environment; moreover, they can protect organic sunscreen components from photodegradation and yield long-lasting protection.

Nature-Inspired Heat and Moisture Exchanger Filters Composed of Gelatin and Chitosan for the Design of Eco-Sustainable "Artificial Noses".

For long-term mechanical ventilation, during anesthesia or intensive care, it is crucial to preserve a minimum level of humidity to avoid damage to the respiratory epithelium. Heat and moisture exchange filters (HME), also called "artificial noses," are passive systems that contribute to delivering inspired gases at about the same conditions of healthy respiration, i.e., 32 °C and relative humidity higher than 90%. Current HME devices suffer from limitations linked either to performance and filtration efficiency to their inadequate antibacterial efficiency, sterilization methods, and durability. Furthermore, in times of global warming and diminishing petroleum oil reserves, replacing the employing of synthetic materials with biomass biodegradable raw materials has considerable economic and environmental value. In the present study, a generation of eco-sustainable, bioinspired, and biodegradable HME devices are designed and developed through a green-chemistry process based on raw materials deriving from food waste and taking inspiration from the functioning, structure, and chemistry of our respiratory system. In particular, different blends are obtained by mixing aqueous solutions of gelatin and chitosan in various polymer ratios and concentrations and then by cross-linking them with different low amounts of genipin, a natural chemical cross-linker. Finally, the blends, post-gelation, are freeze-dried to obtain three-dimensional (3D) highly porous aerogels reproducing both the highly exposed surface area of the upper respiratory ways and the chemical composition of the mucus secretion covering the nasal mucosae. Results are comparable with accepted standards for HME devices and suitable bacteriostatic potential, thus validating these bioinspired materials as promising candidates to be used as an eco-sustainable generation of HME devices.

Marine-Inspired Approaches as a Smart Tool to Face Osteochondral Regeneration.

The degeneration of osteochondral tissue represents one of the major causes of disability in modern society and it is expected to fuel the demand for new solutions to repair and regenerate the damaged articular joints. In particular, osteoarthritis (OA) is the most common complication in articular diseases and a leading cause of chronic disability affecting a steady increasing number of people. The regeneration of osteochondral (OC) defects is one of the most challenging tasks in orthopedics since this anatomical region is composed of different tissues, characterized by antithetic features and functionalities, in tight connection to work together as a joint. The altered structural and mechanical joint environment impairs the natural tissue metabolism, thus making OC regeneration even more challenging. In this scenario, marine-derived ingredients elicit ever-increased interest for biomedical applications as a result of their outstanding mechanical and multiple biologic properties. The review highlights the possibility to exploit such unique features using a combination of bio-inspired synthesis process and 3D manufacturing technologies, relevant to generate compositionally and structurally graded hybrid constructs reproducing the smart architecture and biomechanical functions of natural OC regions.

Electroconductive scaffolds based on gelatin and PEDOT:PSS for cardiac regeneration.

Electroconductive biomaterials have been emerged to support the recovery of the degenerated electrically conductive tissues, especially the cardiac ones after myocardial infarction. This work describes the development of electroconductive scaffolds for cardiac tissue regeneration by using a biocompatible and conductive polymer - i.e. poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate) (PEDOT:PSS) - combined with a biomimetic polymer network of gelatin. Our approach involves the use of dehydrothermal (DHT) treatment in vacuum conditions to fabricate suitably stable scaffolds without using any additional crosslinking agent. The resulting scaffolds mimic the Young modulus - an essential mechanical performance - of native cardiac tissue and are endowed with a well-interconnected porosity coupled with a good swelling ability and stability in physiological conditions. Additionally, the presence of PEDOT:PSS is able to enhance the electroconductivity of resulting materials. All the scaffolds are non-cytotoxic towards H9C2 cardiomyoblasts and the presence of PEDOT:PSS enhances cell adhesion - especially at early timeframes, an essential condition for a successful outcome after the implantation - proliferation, and spreading on scaffolds. Considering the permissive interaction of scaffolds with cardiomyoblasts, the present biomimetic and electroconductive scaffolds display potential applications as implantable biomaterials for regeneration of electroconductive tissues, especially cardiac tissue, and as a promising 3D tissue model for in vitro biomolecules screening.

Safer and Sustainable-by-Design Hydroxyapatite Nanobiomaterials for Biomedical Applications: Assessment of Environmental Hazards.

Developments in the nanotechnology area occur ensuring compliance with regulatory requirements, not only in terms of safety requirements, but also to meet sustainability goals. Hence, safer and sustainable-by-design (SSbD) materials are also aimed for during developmental process. Similar to with any new materials their safety must be assessed. Nanobiomaterials can offer large advantages in the biomedical field, in areas such as tissue repair and regeneration, cancer therapy, etc. For example, although hydroxyapatite-based nanomaterials (nHA) are among the most studied biomaterials, its ecotoxicological effects are mostly unknown. In the present study we investigated the toxicity of seven nHA-based materials, covering both different biomedical applications, e.g., iron-doped hydroxyapatite designed for theragnostic applications), hybrid collagen/hydroxyapatite composites, designed for bone tissue regeneration, and SSbD alternative materials such as titanium-doped hydroxyapatite/alginate composite, designed as sunscreen. The effects were assessed using the soil model Enchytraeus crypticus (Oligochaeta) in the natural standard LUFA 2.2 soil. The assessed endpoints included the 2, 3 and 4 days avoidance behavior (short-term), 28 days survival, size and reproduction (long term based on the OECD standard reproduction test), and 56 days survival and reproduction (longer-term OECD extension). Although overall results showed little to no toxicity among the tested nHA, there was a significant decrease in animals' size for Ti-containing nHA. Moreover, there was a tendency for higher toxicity at the lowest concentrations (i.e., 100 mg/kg). This requires further investigation to ensure safety.

Environmental Hazards of Nanobiomaterials (Hydroxyapatite-Based NMs)-A Case Study with Folsomia candida-Effects from Long Term Exposure.

Hydroxyapatite (HA) is a calcium phosphate used in many fields, including biomedical applications. In particular, ion-doped HA nanomaterials (nHA) are developed for their increased bioactivity, particularly in the fields of regenerative medicine and nanomedicine. In this study, we assessed the ecotoxicological impact of five nHA materials: a synthesized calcium hydroxyapatite (CaP-HA), superparamagnetic iron-doped hydroxyapatite (Fe-HA), titanium-doped hydroxyapatite (Ti-HA), alginate/titanium-doped hydroxyapatite hybrid composite (Ti-HA-Alg), and a commercial HA. The soil ecotoxicology model species Folsomia candida (Collembola) was used, and besides the standard reproduction test (28 days), an extension to the standard for one more generation was performed (56 days). Assessed endpoints included the standard survival and reproduction, and additionally, growth. Exposure via the standard (28 days) did not cause toxicity, but reproduction increased in commercial HA (significantly at 320 mg HA/kg) whereas via the extension (56 days) it decreased in all tested concentrations. Juveniles' size (56 days) was reduced in all tested nHA materials, except commercial HA. nHA materials seem to trigger a compromise between reproduction and growth. Long-term effects could not be predicted based on the standard shorter exposure; hence, the testing of at least two generations (56 days) is recommended to assess the toxicity of nanomaterials, particularly in F. candida. Further, we found that the inclusion of size as additional endpoint is highly relevant.

Male African Elephant (Loxodonta africana) Behavioral Responses to Estrous Call Playbacks May Inform Conservation Management Tools.

Driven by reproductive motives, male African elephants (Loxodonta africana) in musth often expand their home ranges to locate estrous females. This extended range, coupled with heightened aggression often observed in musth males, can be particularly problematic in regions where human-modified landscapes and elephant territories increasingly overlap. Several mitigation tools have been tested to resolve a wide range of human-elephant conflicts with varying degrees of success due to geographical disparities and habituation. We present findings on the potential application of estrous call playbacks in manipulating the behavior and movement of male elephants non-invasively, particularly mature musth adults and younger post-dispersal males, in Etosha National Park. Estrous vocalizations were presented across 26 experimental trials to mature musth adults (n = 5), mature non-musth adults (n = 6), and non-musth males belonging to younger, post-dispersal age classes (n = 8), with behavioral responses scored on a gradient scale from 0-1. Both mature musth adults and younger non-musth elephants were significantly more likely to respond with the highest intensity by approaching the acoustic source compared to mature non-musth adults that avoided the call. However, younger males tested in the presence of an older, higher-ranking male tended to react with a lower intensity than those tested alone. This result likely demonstrates the influence of social hierarchy and associations on male elephant behavior. We also observed a significant increase in physiological response, measured by defecation rate, across all male groups in response to the estrous call playbacks. Our findings suggest that using estrous calls as acoustic deterrents may effectively and non-invasively aid in reducing tension at the human-elephant interface, depending on the age, social context, and reproductive status of the male elephant.

Additive-Free Gelatine-Based Devices for Chondral Tissue Regeneration: Shaping Process Comparison among Mould Casting and Three-Dimensional Printing.

Gelatine is a well-known and extensively studied biopolymer, widely used in recent decades to create biomaterials in many different ways, exploiting its molecular resemblance with collagen, the main constituent of the extra-cellular matrix, from which it is derived. Many have employed this biopolymer in tissue engineering and chemically modified (e.g., gelatin methacryloyl) or blended it with other polymers (e.g., alginate) to modulate or increase its performances and printability. Nevertheless, little is reported about its use as a stand-alone material. Moreover, despite the fact that multiple works have been reported on the realization of mould-casted and three-dimensional printed scaffolds in tissue engineering, a clear comparison among these two shaping processes, towards a comparable workflow starting from the same material, has never been published. Herein, we report the use of gelatine as stand-alone material, not modified, blended, or admixed to be processed or crosslinked, for the realization of suitable scaffolds for tissue engineering, towards the two previously mentioned shaping processes. To make the comparison reliable, the same pre-process (e.g., the gelatin solution preparation) and post-process (e.g., freeze-drying and crosslinking) steps were applied. In this study, gelatine solution was firstly rheologically characterized to find a formulation suitable for being processed with both the shaping processes selected. The realized scaffolds were then morphologically, phisico-chemically, mechanically, and biologically characterized to determine and compare their performances. Despite the fact that the same starting material was employed, as well as the same pre- and post-process steps, the two groups resulted, for most aspects, in diametrically opposed characteristics. The mould-casted scaffolds that resulted were characterized by small, little-interconnected, and random porosity, high resistance to compression and slow cell colonization, while the three-dimensional printed scaffolds displayed big, well-interconnected, and geometrically defined porosity, high elasticity and recover ability after compression, as well as fast and deep cell colonization.

Electroconductive and injectable hydrogels based on gelatin and PEDOT:PSS for a minimally invasive approach in nervous tissue regeneration.

This work describes the development of electroconductive hydrogels as injectable matrices for neural tissue regeneration by exploiting a biocompatible conductive polymer - poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate) (PEDOT:PSS) - combined with a biomimetic polymer network made of gelatin. Our approach involved also genipin - a natural cross-linking agent - to promote gelation of gelatin networks embedding PEDOT:PSS. The achieved results suggest that physical-chemical properties of the resulting hydrogels, like impedance, gelation time, mechanical properties, swelling and degradation in physiological conditions, can be finely tuned by the amount of PEDOT:PSS and genipin used in the formulation. Furthermore, the presence of PEDOT:PSS (i) enhances the electrical conductivity, (ii) improves the shear modulus of the resulting hydrogels though (iii) partially impairing their resistance to shear deformation, (iv) reduces gelation time and (v) reduces their swelling ability in physiological medium. Additionally, the resulting electroconductive hydrogels demonstrate enhanced adhesion and growth of primary rat cortical astrocytes. Given the permissive interaction of hydrogels with primary astrocytes, the presented biomimetic, electroconductive and injectable hydrogels display potential applications as minimally invasive systems for neurological therapies and damaged brain tissue repair.

Magnetic and radio-labeled bio-hybrid scaffolds to promote and track in vivo the progress of bone regeneration.

This work describes the preparation, characterization and functionalization with magnetic nanoparticles of a bone tissue-mimetic scaffold composed of collagen and hydroxyapatite obtained through a biomineralization process. Bone remodeling takes place over several weeks and the possibility to follow it in vivo in a quick and reliable way is still an outstanding issue. Therefore, this work aims to produce an implantable material that can be followed in vivo during bone regeneration by using the existing non-invasive imaging techniques (MRI). To this aim, suitably designed biocompatible SPIONs were linked to the hybrid scaffold using two different strategies, one involving naked SPIONs (nMNPs) and the other using coated and activated SPIONs (MNPs) exposing carboxylic acid functions allowing a covalent attachment between MNPs and collagen molecules. Physico-chemical characterization was carried out to investigate the morphology, crystallinity and stability of the functionalized materials followed by MRI analyses and evaluation of a radiotracer uptake ([99mTc]Tc-MDP). Cell proliferation assays in vitro were carried out to check the cytotoxicity and demonstrated no side effects due to the SPIONs. The achieved results demonstrated that the naked and coated SPIONs are more homogeneously distributed in the scaffold when incorporated during the synthesis process. This work demonstrated a suitable approach to develop a biomaterial for bone regeneration that allows the monitoring of the healing progress even for long-term follow-up studies.

3D Cocultures of Osteoblasts and Staphylococcus aureus on Biomimetic Bone Scaffolds as a Tool to Investigate the Host-Pathogen Interface in Osteomyelitis.

Osteomyelitis (OM) is an infectious disease of the bone primarily caused by the opportunistic pathogen Staphylococcus aureus (SA). This Gram-positive bacterium has evolved a number of strategies to evade the immune response and subvert bone homeostasis, yet the underlying mechanisms remain poorly understood. OM has been modeled in vitro to challenge pathogenetic hypotheses in controlled conditions, thus providing guidance and support to animal experimentation. In this regard, traditional 2D models of OM inherently lack the spatial complexity of bone architecture. Three-dimensional models of the disease overcome this limitation; however, they poorly reproduce composition and texture of the natural bone. Here, we developed a new 3D model of OM based on cocultures of SA and murine osteoblastic MC3T3-E1 cells on magnesium-doped hydroxyapatite/collagen I (MgHA/Col) scaffolds that closely recapitulate the bone extracellular matrix. In this model, matrix-dependent effects were observed in proliferation, gene transcription, protein expression, and cell-matrix interactions both of the osteoblastic cell line and of bacterium. Additionally, these had distinct metabolic and gene expression profiles, compared to conventional 2D settings, when grown on MgHA/Col scaffolds in separate monocultures. Our study points to MgHA/Col scaffolds as biocompatible and bioactive matrices and provides a novel and close-to-physiology tool to address the pathogenetic mechanisms of OM at the host-pathogen interface.

Medicated Hydroxyapatite/Collagen Hybrid Scaffolds for Bone Regeneration and Local Antimicrobial Therapy to Prevent Bone Infections.

Microbial infections occurring during bone surgical treatment, the cause of osteomyelitis and implant failures, are still an open challenge in orthopedics. Conventional therapies are often ineffective and associated with serious side effects due to the amount of drugs administered by systemic routes. In this study, a medicated osteoinductive and bioresorbable bone graft was designed and investigated for its ability to control antibiotic drug release in situ. This represents an ideal solution for the eradication or prevention of infection, while simultaneously repairing bone defects. Vancomycin hydrochloride and gentamicin sulfate, here considered for testing, were loaded into a previously developed and largely investigated hybrid bone-mimetic scaffold made of collagen fibers biomineralized with magnesium doped-hydroxyapatite (MgHA/Coll), which in the last ten years has widely demonstrated its effective potential in bone tissue regeneration. Here, we have explored whether it can be used as a controlled local delivery system for antibiotic drugs. An easy loading method was selected in order to be reproducible, quickly, in the operating room. The maintenance of the antibacterial efficiency of the released drugs and the biosafety of medicated scaffolds were assessed with microbiological and in vitro tests, which demonstrated that the MgHA/Coll scaffolds were safe and effective as a local delivery system for an extended duration therapy-promising results for the prevention of bone defect-related infections in orthopedic surgeries.

Nature-Inspired Unconventional Approaches to Develop 3D Bioceramic Scaffolds with Enhanced Regenerative Ability.

Material science is a relevant discipline in support of regenerative medicine. Indeed, tissue regeneration requires the use of scaffolds able to guide and sustain the natural cell metabolism towards tissue regrowth. This need is particularly important in musculoskeletal regeneration, such as in the case of diseased bone or osteocartilaginous regions for which calcium phosphate-based scaffolds are considered as the golden solution. However, various technological barriers related to conventional ceramic processing have thus far hampered the achievement of biomimetic and bioactive scaffolds as effective solutions for still unmet clinical needs in orthopaedics. Driven by such highly impacting socioeconomic needs, new nature-inspired approaches promise to make a technological leap forward in the development of advanced biomaterials. The present review illustrates ion-doped apatites as biomimetic materials whose bioactivity resides in their unstable chemical composition and nanocrystallinity, both of which are, however, destroyed by the classical sintering treatment. In the following, recent nature-inspired methods preventing the use of high-temperature treatments, based on (i) chemically hardening bioceramics, (ii) biomineralisation process, and (iii) biomorphic transformations, are illustrated. These methods can generate products with advanced biofunctional properties, particularly biomorphic transformations represent an emerging approach that could pave the way to a technological leap forward in medicine and also in various other application fields.

Targeting the plasticity of mesenchymal stromal cells to reroute the course of acute myeloid leukemia.

Bone marrow (BM) microenvironment contributes to the regulation of normal hematopoiesis through a finely tuned balance of self-renewal and differentiation processes, cell-cell interaction, and secretion of cytokines that during leukemogenesis are altered and favor tumor cell growth. In pediatric acute myeloid leukemia (AML), chemotherapy is the standard of care, but >30% of patients still relapse. The need to accelerate the evaluation of innovative medicines prompted us to investigate the role of mesenchymal stromal cells (MSCs) in the leukemic niche to define its contribution to the mechanism of leukemia drug escape. We generated a humanized 3-dimensional (3D) niche with AML cells and MSCs derived from either patients (AML-MSCs) or healthy donors. We observed that AML cells establish physical connections with MSCs, mediating a reprogrammed transcriptome inducing aberrant cell proliferation and differentiation and severely compromising their immunomodulatory capability. We confirmed that AML cells modulate h-MSCs transcriptional profile promoting functions similar to the AML-MSCs when cocultured in vitro, thus facilitating leukemia progression. Conversely, MSCs derived from BM of patients at time of disease remission showed recovered healthy features at transcriptional and functional levels, including the secretome. We proved that AML blasts alter MSCs activities in the BM niche, favoring disease development and progression. We discovered that a novel AML-MSC selective CaV1.2 channel blocker drug, lercanidipine, is able to impair leukemia progression in 3D both in vitro and when implanted in vivo if used in combination with chemotherapy, supporting the hypothesis that synergistic effects can be obtained by dual targeting approaches.

Biomimetic Mineralization Promotes Viability and Differentiation of Human Mesenchymal Stem Cells in a Perfusion Bioreactor.

In bone tissue engineering, the design of 3D systems capable of recreating composition, architecture and micromechanical environment of the native extracellular matrix (ECM) is still a challenge. While perfusion bioreactors have been proposed as potential tool to apply biomechanical stimuli, its use has been limited to a low number of biomaterials. In this work, we propose the culture of human mesenchymal stem cells (hMSC) in biomimetic mineralized recombinant collagen scaffolds with a perfusion bioreactor to simultaneously provide biochemical and biophysical cues guiding stem cell fate. The scaffolds were fabricated by mineralization of recombinant collagen in the presence of magnesium (RCP.MgAp). The organic matrix was homogeneously mineralized with apatite nanocrystals, similar in composition to those found in bone. X-Ray microtomography images revealed isotropic porous structure with optimum porosity for cell ingrowth. In fact, an optimal cell repopulation through the entire scaffolds was obtained after 1 day of dynamic seeding in the bioreactor. Remarkably, RCP.MgAp scaffolds exhibited higher cell viability and a clear trend of up-regulation of osteogenic genes than control (non-mineralized) scaffolds. Results demonstrate the potential of the combination of biomimetic mineralization of recombinant collagen in presence of magnesium and dynamic culture of hMSC as a promising strategy to closely mimic bone ECM.

Dual-crosslinked 3D printed gelatin scaffolds with potential for temporomandibular joint cartilage regeneration.

A promising alternative to current treatment options for degenerative conditions of the temporomandibular joint (TMJ) is cartilage tissue engineering, using 3D printed scaffolds and mesenchymal stem cells. Gelatin, with its inherent biocompatibility and printability has been proposed as a scaffold biomaterial, but because of its thermoreversible properties, rapid degradation and inadequate strength it must be crosslinked to be stable in physiological conditions. The aim of this study was to identify non-toxic and effective crosslinking methods intended to improve the physical properties of 3D printed gelatin scaffolds for cartilage regeneration. Dehydrothermal (DHT), ribose glycation and dual crosslinking with both DHT and ribose treatments were tested. The crosslinked scaffolds were characterized by chemical, mechanical, and physical analysis. The dual-crosslinked scaffolds had the highest degree of crosslinking and the greatest resistance to hydrolytic and enzymatic degradation. Compared to the dual-crosslinked group, the ribose-crosslinked scaffolds had thinner printed strands, larger pore surface area and higher fluid uptake. The compressive modulus values were 2 kPa for ribose, 37.6 kPa for DHT and 30.9 kPa for dual-crosslinked scaffolds. None of the crosslinking methods had cytotoxic effects on the seeded rat bone marrow-derived mesenchymal stem cells (rBMSC). After 4 and 7 d, the dual-crosslinked scaffolds exhibited better cell proliferation than the other groups. Although all scaffolds supported chondrogenic differentiation of rBMSC, dual-crosslinked scaffolds demonstrated the lowest expression of the hypertrophy-related collagen 10 gene after 21 d. The results show that 3D printed gelatin scaffolds, when dually crosslinked with ribose and DHT methods, are not toxic, promote chondrogenic differentiation of rBMSC and have potential application in tissue engineering of TMJ condylar cartilage.

Bio-inspired polymeric iron-doped hydroxyapatite microspheres as a tunable carrier of rhBMP-2.

Hybrid superparamagnetic microspheres with bone-like composition, previously developed by a bio-inspired assembling/mineralization process, are evaluated for their ability to uptake and deliver recombinant human bone morphogenetic protein-2 (rhBMP-2) in therapeutically-relevant doses along with prolonged release profiles. The comparison with hybrid non-magnetic and with non-mineralized microspheres highlights the role of nanocrystalline, nanosize mineral phases when they exhibit surface charged groups enabling the chemical linking with the growth factor and thus moderating the release kinetics. All the microspheres show excellent osteogenic ability with human mesenchymal stem cells whereas the hybrid mineralized ones show a slow and sustained release of rhBMP-2 along 14 days of soaking into cell culture medium with substantially bioactive effect, as reported by assay with C2C12 BRE-Luc cell line. It is also shown that the release extent can be modulated by the application of pulsed electromagnetic field, thus showing the potential of remote controlling the bioactivity of the new micro-devices which is promising for future application of hybrid biomimetic microspheres in precisely designed and personalized therapies.