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Polymer-ceramic composites obtained as the result of a mineralization process hold great promise for the future of tissue engineering. Simulated body fluids (SBFs) are widely used for the mineralization of polymer scaffolds. In this work an exhaustive study with the aim of optimizing the mineralization process on a poly( L-lactic acid) (PLLA) macroporous scaffold has been performed. We observed that when an air plasma treatment is applied to the PLLA scaffold its hydroxyapatite nucleation ability is considerably improved. However, plasma treatment only allows apatite deposition on the surface of the scaffold but not in its interior. When a 5 wt % of synthetic hydroxyapatite (HAp) nanoparticles is mixed with PLLA a more abundant biomimetic hydroxyapatite layer grows inside the scaffold in SBF. The morphology, amount, and composition of the generated biomimetic hydroxyapatite layer on the pores' surface have been analyzed. Large mineralization times are harmful to pure PLLA as it rapidly degrades and its elastic compression modulus significantly decreases. Degradation is retarded in the composite scaffolds because of the faster and extensive biomimetic apatite deposition and the role of HAp to control the pH. Mineralized scaffolds, covered by an apatite layer in SBF, were implanted in osteochondral lesions performed in the medial femoral condyle of healthy sheep. We observed that the presence of biomimetic hydroxyapatite on the pore's surface of the composite scaffold produces a better integration in the subchondral bone, in comparison to bare PLLA scaffolds. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 101B: 173-186, 2013. [ABSTRACT FROM AUTHOR]

Organic scaffolds of poly(ethyl methacrylate‐co‐hydroxyethyl acrylate) [P(EMA‐co‐HEA)] 70/30 wt % ratio, with varying proportions of silica SiO2 from 0 to 20 wt % and aligned tubular pores, were prepared using a fiber‐templating fabrication method, with the aim of mimicking structure and properties of the mineralized tissue of natural dentin. Precursors of the copolymer and silica were simultaneously polymerized in a sol–gel process within the fiber template, which was eventually eliminated to generate homogeneously distributed parallel micrometer‐sized pores in the material. Scaffolds of PEMA and PHEA were obtained by the same approach. The scaffolds were characterized by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy, and thermogravimetric analysis. The specific volume was determined by Archimedes' method and the porosity calculated from the geometry. The mechanical properties were analyzed in tensile and compressive modes. The bioactivity of the scaffolds with 15 wt % SiO2 was tested by immersion in simulated body fluid (SBF) for 7 days followed by immersion in 2× SBF for 7 days. These scaffolds were afterwards characterized by SEM, energy dispersive spectroscopy, and compression assays. Percentages of silica above 10 wt % reinforced mechanically the copolymer, evidenced by the hindrance of the long range motions of the organic chains, altered shrinkage and swelling, and meanwhile conferred bioactivity to its surface. These tubular porous structures, which resemble natural dentin with regard to its structure and properties and induce the precipitation of apatite on their surfaces in vitro, are expected to facilitate the integration in the host mineralized tissue, to stimulate cell growth and to be useful as guiding scaffolds for in vivo dentin regeneration. © 2008 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2009 [ABSTRACT FROM AUTHOR]

Articular cartilage has limited regenerative capacity, so focal lesions generate mechanical stress in the joint that induces an aggravation of the damage, which ultimately leads to osteoarthritis. We recently suggested the use of microgels at the site of the cartilage defect, as a support material, to generate a biomechanical environment where pluripotent cells differentiate towards the hyaline cartilage phenotype. Here we propose a chondral regeneration strategy based on subchondral bone injury, and filling the defect site with an agglomerate of two types of microspheres, some rigid made of a biodegradable polyester (40 μm mean diameter), and others with a gel consistency made of platelet-rich plasma obtained from circulating blood (70-110 μm diameter). A 3-mm diameter defect was made in the articular cartilage of the knee joint in rabbits, exposing the subchondral bone, in which incisions were made to produce bleeding. Microgels were implanted filling the defect, which was covered with a synthetic membrane of the same polyester. Three months later, cartilage regeneration was analyzed according to the International Cartilage Repair Society (ICRS) guidelines. The newly formed tissue showed histological characteristics of hyaline cartilage, being significantly closer to native cartilage than when only the membrane was implanted, mainly in parameters such as tissue (70.0 ± 20.9) and cell morphologies (100.0 ± 0.0), and surface architecture (90.0 ± 22.4) and assessment (70.0 ± 11.2), with native tissue having a value of 100. Polyester microspheres and membrane were not bioreabsorbed during the three months, but rather moved towards the subchondral bone, leaving space for the organization of the newly formed tissue., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Physical cues have been shown to be effective in inducing osteogenic differentiation of mesenchymal stem cells (MSCs). Here, we propose piezoelectric stimulation as a potential osteogenic cue mimicking the electroactive properties of bone's extracellular matrix. When combined with a magnetostrictive component, piezoelectric polymers can be used for MSC stimulation by applying an external magnetic field. The deformation of the magnetostrictive component produces a deformation in the polymer matrix, generating a change in the surface charge that induces an electric field that can be transmitted to the cells. Cell adhesion, cytoskeleton changes, and metabolomics are the first evidence of MSC osteoblastogenesis and can be used to study initial MSC response to this kind of stimulation. In the current study, poly(vinylidene) fluoride (PVDF) piezoelectric films with and without cobalt ferrite oxide (CFO) crystallized from the melt in the presence of the ionic liquid 1-butyl-3-methyl-imidazolium chloride ([Bmim][Cl]) were produced. [Bmim][Cl] allowed the production of the β-phase, the most electroactive phase, even without CFO. After ionic liquid removal, PVDF and PVDF-CFO films presented high percentages of the β-phase and similar crystalline content. Incorporating CFO nanoparticles was effective, allowing the electromechanical stimulation of MSCs by applying a magnetic field with a bioreactor. Before stimulation, the initial response of MSCs was characterized in static conditions, showing that the produced films were biocompatible and noncytotoxic, allowing MSC adhesion and proliferation in the short term. Stimulation experiments revealed that MSCs electromechanically stimulated for 3 days in PVDF-CFO supports showed longer focal adhesions and decreased vimentin cytoskeletal density, both signals of early osteogenic differentiation. Furthermore, they rearranged their energy metabolism toward an osteogenic phenotype after 7 days of culture under the same stimulation. The results prove that MSCs respond to electromechanical stimulation by osteogenic differentiation., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)

Doxorubicin (DOX) is a chemotherapeutic drug used in osteosarcoma treatments, usually administrated in very high dosages. This study proposes novel DOX microcarriers based on chitosan (CHT) physically crosslinked with copper(II) ions that will act synergically to inhibit tumor growth at lower drug dosage without affecting the healthy cells. Spherical CHT-Cu microparticles with a smooth surface and an average size of 30.1 ± 9.1 µm were obtained by emulsion. The release of Cu 2+ ions from the CHT-Cu microparticles showed that 99.4 % of added cupric ions were released in 72 h of incubation in a complete cell culture medium (CCM). DOX entrapment in microparticles was conducted in a phosphate buffer solution (pH 6), utilizing the pH sensitivity of the polymer. The successful drug-loading process was confirmed by DOX emitting red fluorescence from drug-loaded microcarriers (DOX@CHT-Cu). The drug release in CCM showed an initial burst release, followed by sustained release. Biological assays indicated mild toxicity of CHT-Cu microparticles on the MG-63 osteosarcoma cell line, without affecting the viability of human mesenchymal stem cells (hMSCs). The DOX@CHT-Cu microparticles at concentration of 0.5 mg mL ‒ 1 showed selective toxicity toward MG-63 cells., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The aim of this work is to study the effect of platelet factors on the differentiation of mesenchymal stem cells (MSCs) to hyaline cartilage chondrocytes in a three-dimensional environment. MSCs were cultured in a microgel environment with a chondrogenic medium. The microgel consisted of microspheres that combine gelatin and platelet-rich plasma (PRP). The gelatin/PRP microdroplets were produced by emulsion. The gelatin containing the microdroplets was enzymatically gelled, retaining PRP and, just before seeding the cells, platelets were activated by adding calcium chloride so that platelet growth factors were released into the culture media but not before. Platelet activation was analyzed before activation to rule out the possibility that the gelatin cross-linking process itself activated the platelets. The gene expression of characteristic chondrogenic markers and miRNA expression were analyzed in cells cultured in a differentiation medium and significant differences were found between gelation/PRP microgels and those containing only pure gelatin. In summary, the gelatin microspheres effectively encapsulated platelets that secreted and released factors that significantly contributed to cellular chondrogenic differentiation. At the same time, the microgel constituted a 3D medium that provided the cells with adherent surfaces and the possibility of three-dimensional cell-cell contact.

Polymer hydrogels crosslinked by therapeutic metal ions have attracted increased interest in recent years due to their unique and versatile properties. Chitosan hydrogels are widely investigated for various biomedical applications such as tissue engineering and drug delivery. Copper and zinc ions are considered as therapeutic metal ions, that have important roles in bone regeneration. The aim of this study was to investigate the physicochemical and biological properties of bimetallic-chitosan complex hydrogels with different cupric and zinc ions content. Scanning electron microscopy (SEM) revealed changes in the morphology from the microstructure with larger, tubular pores for aerogels with higher Zn content, to the sheets-like structure with long pores for samples with higher Cu content. FTIR analysis indicated the formation of bimetallic-chitosan aerogels. The obtained X-ray diffraction patterns showed a broadening of chitosan's characteristic diffraction maximum, while characterization of physical properties showed decreased swelling ability and increased shear modulus with higher Cu content. ICP-MS results showed a negligible amount of copper and zinc ions released under physiological conditions during 24 h indicating a strong physical crosslink between metal ions and chitosan chains. Furthermore, accelerated in vitro degradation showed that hydrogels maintained good stability during four weeks of lysozyme activity. The MTT assay indicated that the cytotoxicity of Cu 2+ -Zn 2+ /chitosan complexes could be adjusted by the amount of cupric ions. All results imply that Cu 2+ and Zn 2+ ions act as physical crosslinkers of the polymer network. Also, results are in agreement with the prediction of density functional theory (DFT) which indicated stronger chitosan-Cu tetrahedral aqua complex interactions in comparison to the chitosan-[Zn(H 2 O) 4 ] 2+ interactions., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Anamarija Rogina reports financial support was provided by Croatian Science Foundation., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Cell-based liver therapies based on retrieving and steadying failed metabolic function(s) for acute and chronic diseases could be a valuable substitute for liver transplants, even though they are limited by the low engraftment capability and reduced functional quality of primary human hepatocytes (PHH). In this paper we propose the use of gelatin-hyaluronic acid (Gel-HA) scaffolds seeded with PHH for the treatment of liver failure. We first optimized the composition using Gel-HA hydrogels, looking for the mechanical properties closer to the human liver and determining HepG2 cells functionality. Gel-HA scaffolds with interconnected porosity (pore size 102 μm) were prepared and used for PHH culture and evaluation of key hepatic functions. PHH cultured in Gel-HA scaffolds exhibited increased albumin and urea secretion and metabolic capacity (CYP and UGT activity levels) compared to standard monolayer cultures. The transplant of the scaffold containing PHH led to an improvement in liver function (transaminase levels, necrosis) and ameliorated damage in a mouse model of acetaminophen (APAP)-induced liver failure. The study provided a mechanistic understanding of APAP-induced liver injury and the impact of transplantation by analyzing cytokine production and oxidative stress induction to find suitable biomarkers of cell therapy effectiveness., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. The funders had no role in either the design of the study, the collection, analyses, or interpretation of data in writing the manuscript or in the decision to publish the results., (Crown Copyright © 2023. Published by Elsevier B.V. All rights reserved.)

Fibronectin (FN) mediates cell-material interactions during events such as tissue repair, and therefore the biomimetic modeling of this protein in vitro benefits regeneration. The nature of the interface is crucial in determining cell adhesion, morphology, and differentiation. Poly(ethyl acrylate) (PEA) spontaneously organizes FN into biological nanonetworks, resulting in exceptional bone regeneration in animal models. Spontaneous network organization of FN is also observed in poly(buthyl acrylate) (PBA) substrates that have higher surface mobility than PEA. C2C12 myoblasts differentiate efficiently on PEA and PBA substrates. In this study, we investigate if intermediate surface mobilities between PEA and PBA induce cell differentiation more efficiently than PEA. A family of P(EA- co -BA) copolymers were synthesized in the entire range of compositions to finely tune surface mobility between PEA and PBA. Surface characterization demonstrates that FN mobility steadily increased with the PBA content. All compositions allowed the biological organization of FN with similar exposure of cell binding domains. C2C12 myoblasts adhered well in all the materials, with higher focal adhesions in PEA and PBA. The increase of the interfacial mobility had an impact in cell adhesion by increasing the number of FAs per cell. In addition, cell differentiation decreased proportionally with surface mobility, from PEA to PBA.

Hydrogels have been developed for different biomedical applications such as in vitro culture platforms, drug delivery, bioprinting and tissue engineering. Enzymatic cross-linking has many advantages for its ability to form gels in situ while being injected into tissue, which facilitates minimally invasive surgery and adaptation to the shape of the defect. It is a highly biocompatible form of cross-linking, which permits the harmless encapsulation of cytokines and cells in contrast to chemically or photochemically induced cross-linking processes. The enzymatic cross-linking of synthetic and biogenic polymers also opens up their application as bioinks for engineering tissue and tumor models. This review first provides a general overview of the different cross-linking mechanisms, followed by a detailed survey of the enzymatic cross-linking mechanism applied to both natural and synthetic hydrogels. A detailed analysis of their specifications for bioprinting and tissue engineering applications is also included.

Multiple myeloma is a hematologic neoplasm caused by an uncontrolled clonal proliferation of neoplastic plasma cells (nPCs) in the bone marrow. The development and survival of this disease is tightly related to the bone marrow environment. Proliferation and viability of nPCs depend on their interaction with the stromal cells and the extracellular matrix components, which also influences the appearance of drug resistance. Recapitulating these interactions in an in vitro culture requires 3D environments that incorporate the biomolecules of interest. In this work, we studied the proliferation and viability of three multiple myeloma cell lines in a microgel consisting of biostable microspheres with fibronectin (FN) on their surfaces. We also showed that the interaction of the RPMI8226 cell line with FN induced cell arrest in the G0/G1 cell cycle phase. RPMI8226 cells developed a significant resistance to dexamethasone, which was reduced when they were treated with dexamethasone and bortezomib in combination.