Polyelectrolyte complexes based on a novel and sustainable hemicellulose-rich lignosulphonate for drug delivery applications.

Polyelectrolyte complexes (PECs) are polymeric structures formed by the self-assembly of oppositely charged polymers. Novel biomaterials based on PECs are currently under investigation as drug delivery systems, among other applications. This strategy leverages the ability of PECs to entrap drugs under mild conditions and control their release. In this study, we combined a novel and sustainably produced hemicellulose-rich lignosulphonate polymer (EH, negatively charged) with polyethyleneimine (PEI) or chitosan (CH, positively charged) and agar for the development of drug-releasing PECs. A preliminary screening demonstrated the effect of several parameters (polyelectrolyte ratio, temperature, and type of polycation) on PECs formation. From this, selected formulations were further characterized in terms of thermal properties, surface morphology at the microscale, stability, and ability to load and release methylene blue (MB) as a model drug. EH/PEI complexes had a more pronounced gel-like behaviour compared to the EH/CH complexes. Differential scanning calorimetry (DSC) results supported the establishment of polymeric interactions during complexation. Overall, PECs' stability was positively affected by low pH, ratios close to 1:1, and the addition of agar. PECs with higher EH content showed a higher MB loading, likely promoted by stronger electrostatic interactions. The EH/CH formulation enriched with agar showed the best sustained release profile of MB during the first 30 h in a pH-dependent environment simulating the gastrointestinal tract. Overall, we defined the conditions to formulate novel PECs based on a sustainable hemicellulose-rich lignosulphonate for potential applications in drug delivery, which promotes the valuable synergy between sustainability and the biomedical field.

Integrating bioprinting, cell therapies and drug delivery towards in vivo regeneration of cartilage, bone and osteochondral tissue.

The biological and biomechanical functions of cartilage, bone and osteochondral tissue are naturally orchestrated by a complex crosstalk between zonally dependent cells and extracellular matrix components. In fact, this crosstalk involves biomechanical signals and the release of biochemical cues that direct cell fate and regulate tissue morphogenesis and remodelling in vivo. Three-dimensional bioprinting introduced a paradigm shift in tissue engineering and regenerative medicine, since it allows to mimic native tissue anisotropy introducing compositional and architectural gradients. Moreover, the growing synergy between bioprinting and drug delivery may enable to replicate cell/extracellular matrix reciprocity and dynamics by the careful control of the spatial and temporal patterning of bioactive cues. Although significant advances have been made in this direction, unmet challenges and open research questions persist. These include, among others, the optimization of scaffold zonality and architectural features; the preservation of the bioactivity of loaded active molecules, as well as their spatio-temporal release; the in vitro scaffold maturation prior to implantation; the pros and cons of each animal model and the graft-defect mismatch; and the in vivo non-invasive monitoring of new tissue formation. This work critically reviews these aspects and reveals the state of the art of using three-dimensional bioprinting, and its synergy with drug delivery technologies, to pattern the distribution of cells and/or active molecules in cartilage, bone and osteochondral engineered tissues. Most notably, this work focuses on approaches, technologies and biomaterials that are currently under in vivo investigations, as these give important insights on scaffold performance at the implantation site and its interaction/integration with surrounding tissues.

Biofunctionalization of 3D printed collagen with bevacizumab-loaded microparticles targeting pathological angiogenesis.

Pathological angiogenesis is a crucial attribute of several chronic diseases such as cancer, age-related macular degeneration, and osteoarthritis (OA). In the case of OA, pathological angiogenesis mediated by the vascular endothelial growth factor (VEGF), among other factors, contributes to cartilage degeneration and to implants rejection. In line with this, the use of the anti-VEGF bevacizumab (BVZ) has been shown to prevent OA progression and support cartilage regeneration. The aim of this work was to functionalize a medical grade collagen with poly (lactic-co-glycolic acid) (PLGA) microparticles containing BVZ via three-dimensional (3D) printing to target pathological angiogenesis. First, the effect of several formulation parameters on the encapsulation and release of BVZ from PLGA microparticles was studied. Then, the anti-angiogenic activity of released BVZ was tested in a 3D cell model. The 3D printability of the microparticle-loaded collagen ink was tested by evaluating the shape fidelity of 3D printed structures. Results showed that the release and the encapsulation efficiency of BVZ could be tuned as a function of several formulation parameters. In addition, the released BVZ was observed to reduce vascularization by human umbilical vein endothelial cells. Finally, the collagen ink with embedded BVZ microparticles was successfully printed, leading to shape-stable meniscus-, nose- and auricle-like structures. Taken altogether, we defined the conditions for the successful combination of BVZ-loaded microparticles with the 3D printing of a medical grade collagen to target pathological angiogenesis.

Engineering Anisotropic Meniscus: Zonal Functionality and Spatiotemporal Drug Delivery.

Human meniscus is a fibrocartilaginous structure that is crucial for an adequate performance of the human knee joint. Degeneration of the meniscus is often followed by partial or total meniscectomy, which enhances the risk of developing knee osteoarthritis. The lack of a satisfactory treatment for this condition has triggered a major interest in drug delivery (DD) and tissue engineering (TE) strategies intended to restore a bioactive and fully functional meniscal tissue. The aim of this review is to critically discuss the most relevant studies on spatiotemporal DD and TE, aiming for a multizonal meniscal reconstruction. Indeed, the development of meniscal tissue implants should involve a provision for adequate active molecules and scaffold features that take into account the anisotropic ultrastructure of human meniscus. This zonal differentiation is reflected in the meniscus biochemical composition, collagen fiber arrangement, and cell distribution. In this sense, it is expected that a proper combination of advanced DD and zonal TE strategies will play a key role in the future trends in meniscus regeneration. Impact statement Meniscus degeneration is one of the main causes of knee pain, inflammation, and reduced mobility. Currently used suturing procedures and meniscectomy are far from being ideal solutions to the loss of meniscal function. Therefore, drug delivery (DD) and tissue engineering (TE) strategies are currently under investigation. DD systems aim at an in situ controlled release of growth factors, whereas TE strategies aim at mimicking the anisotropy of native meniscus. The goal of this review is to discuss these two main approaches, as well as synergies between them that are expected to lead to a real breakthrough in the field.

Surface functionalization of polylactic acid fibers with alendronate groups does not improve the mechanical properties of fiber-reinforced calcium phosphate cements.

Calcium phosphate cements (CPCs) are frequently used as synthetic bone substitute, but their intrinsic low fracture toughness impedes their application in highly loaded skeletal sites. However, fibers can be used to reduce the brittleness of these CPCs provided that the affinity between the fibers and cement matrix facilitates the transfer of loads from the matrix to the fibers. The aim of the present work was to improve the interface between hydrophobic polylactic acid (PLA) microfibers and hydrophilic CPC. To this end, calcium-binding alendronate groups were conjugated onto the surface of PLA microfibers via different strategies to immobilize a tunable amount of alendronate onto the fiber surface. CPCs reinforced with PLA fibers revealed toughness values which were up to 50-fold higher than unreinforced CPCs. Nevertheless, surface functionalization of PLA microfibers with alendronate groups did not improve the mechanical properties of fiber-reinforced CPCs.

Complex coacervation-based loading and tunable release of a cationic protein from monodisperse glycosaminoglycan microgels.

Glycosaminoglycans (GAGs) are of interest for biomedical applications because of their ability to retain proteins (e.g. growth factors) involved in cell-to-cell signaling processes. In this study, the potential of GAG-based microgels for protein delivery and their protein release kinetics upon encapsulation in hydrogel scaffolds were investigated. Monodisperse hyaluronic acid methacrylate (HAMA) and chondroitin sulfate methacrylate (CSMA) micro-hydrogel spheres (diameters 500-700 µm), were used to study the absorption of a cationic model protein (lysozyme), microgel (de)swelling, intra-gel lysozyme distribution and its diffusion coefficient in the microgels dispersed in buffers (pH 7.4) of varying ionic strengths. Upon incubation in 20 mM buffer, lysozyme was absorbed up to 3 and 4 mg mg-1 dry microspheres for HAMA and CSMA microgels respectively, with loading efficiencies up to 100%. Binding stoichiometries of disaccharide : lysozyme (10.2 : 1 and 7.5 : 1 for HAMA and CSMA, respectively) were similar to those for GAG-lysozyme complex coacervates based on soluble GAGs found in literature. Complex coacervates inside GAG microgels were also formed in buffers of higher ionic strengths as opposed to GAG-lysozyme systems based on soluble GAGs, likely due to increased local anionic charge density in the GAG networks. Binding of cationic lysozyme to the negatively charged microgel networks resulted in deswelling up to a factor 2 in diameter. Lysozyme release from the microgels was dependent on the ionic strength of the buffer and on the number of anionic groups per disaccharide, (1 for HAMA versus 2 for CSMA). Lysozyme diffusion coefficients of 0.027 in HAMA and <0.006 µm2 s-1 in CSMA microgels were found in 170 mM buffer (duration of release 14 and 28 days respectively). Fluorescence Recovery After Photobleaching (FRAP) measurements yielded similar trends, although lysozyme diffusion was likely altered due to the negative charges introduced to the protein through the FITC-labeling resulting in weaker protein-matrix interactions. Finally, lysozyme-loaded CSMA microgels were embedded into a thermosensitive hydrogel scaffold. These composite systems showed complete lysozyme release in ∼58 days as opposed to only 3 days for GAG-free scaffolds. In conclusion, covalently crosslinked methacrylated GAG hydrogels have potential as controlled release depots for cationic proteins in tissue engineering applications.

3D bioprinting of methacrylated hyaluronic acid (MeHA) hydrogel with intrinsic osteogenicity.

In bone regenerative medicine there is a need for suitable bone substitutes. Hydrogels have excellent biocompatible and biodegradable characteristics, but their visco-elastic properties limit their applicability, especially with respect to 3D bioprinting. In this study, we modified the naturally occurring extracellular matrix glycosaminoglycan hyaluronic acid (HA), in order to yield photo-crosslinkable hydrogels with increased mechanical stiffness and long-term stability, and with minimal decrease in cytocompatibility. Application of these tailor-made methacrylated hyaluronic acid (MeHA) gels for bone tissue engineering and 3D bioprinting was the subject of investigation. Visco-elastic properties of MeHA gels, measured by rheology and dynamic mechanical analysis, showed that irradiation of the hydrogels with UV light led to increased storage moduli and elastic moduli, indicating increasing gel rigidity. Subsequently, human bone marrow derived mesenchymal stromal cells (MSCs) were incorporated into MeHA hydrogels, and cell viability remained 64.4% after 21 days of culture. Osteogenic differentiation of MSCs occurred spontaneously in hydrogels with high concentrations of MeHA polymer, in absence of additional osteogenic stimuli. Addition of bone morphogenetic protein-2 (BMP-2) to the culture medium further increased osteogenic differentiation, as evidenced by increased matrix mineralisation. MeHA hydrogels demonstrated to be suitable for 3D bioprinting, and were printed into porous and anatomically shaped scaffolds. Taken together, photosensitive MeHA-based hydrogels fulfilled our criteria for cellular bioprinted bone constructs within a narrow window of concentration.

A Synthetic Thermosensitive Hydrogel for Cartilage Bioprinting and Its Biofunctionalization with Polysaccharides.

Hydrogels based on triblock copolymers of polyethylene glycol and partially methacrylated poly[N-(2-hydroxypropyl) methacrylamide mono/dilactate] make up an attractive class of biomaterials because of their biodegradability, cytocompatibility, and tunable thermoresponsive and mechanical properties. If these properties are fine-tuned, the hydrogels can be three-dimensionally bioprinted, to generate, for instance, constructs for cartilage repair. This study investigated whether hydrogels based on the polymer mentioned above with a 10% degree of methacrylation (M10P10) support cartilage formation by chondrocytes and whether the incorporation of methacrylated chondroitin sulfate (CSMA) or methacrylated hyaluronic acid (HAMA) can improve the mechanical properties, long-term stability, and printability. Chondrocyte-laden M10P10 hydrogels were cultured for 42 days to evaluate chondrogenesis. M10P10 hydrogels with or without polysaccharides were evaluated for their mechanical properties (before and after UV photo-cross-linking), degradation kinetics, and printability. Extensive cartilage matrix production occurred in M10P10 hydrogels, highlighting their potential for cartilage repair strategies. The incorporation of polysaccharides increased the storage modulus of polymer mixtures and decreased the degradation kinetics in cross-linked hydrogels. Addition of HAMA to M10P10 hydrogels improved printability and resulted in three-dimensional constructs with excellent cell viability. Hence, this novel combination of M10P10 with HAMA forms an interesting class of hydrogels for cartilage bioprinting.

Designing prodrugs for the treatment of Parkinson's disease.

INTRODUCTION: Current Parkinson's disease (PD) therapy is essentially symptomatic, and l-Dopa (LD), is the treatment of choice in more advanced stages of the disease. However, motor complications often develop after long-term treatment, and at this point physicians usually prescribe adjuvant therapy with other classes of antiparkinsonian drugs, including dopamine (DA) agonists, catechol-O-methyl transferase (COMT) or monoamine oxidase (MAO)-B inhibitors. In order to improve bioavailability, the prodrug approach appeared to be the most promising, and some antiparkinsonian prodrugs have been prepared in an effort to solve these problems. AREAS COVERED: This review discusses the evidence of progress in PD therapy, mainly focused on prodrug approach for treatment of this neurological disorder. Several derivatives were studied with the aim of enhancing its chemical stability, water or lipid solubility, as well as diminishing the susceptibility to enzymatic degradation. Chemical structures mainly related to LD, DA and dopaminergic agonists are also reviewed in this paper. EXPERT OPINION: In order to strengthen the pharmacological activity of antiparkinsonian drugs, enhancing their penetration of the blood-brain barrier (BBB), different approaches are possible. Among these, the prodrug approach appeared to be the most promising, and many prodrugs have been prepared in an effort to optimize physicochemical characteristics. In addition, novel therapeutic strategies based on formulations linking dopaminergic drugs with neuroprotective agents, increasing LD striatal levels and offering sustained release of the drug without any fluctuation of brain concentration, offer promising avenues for development of other effective new treatments for PD.