Magnetite nanoparticles (Fe3O4 NPs), due to their unique physicochemical properties, are considered as promising nanomaterials for multiple biomedical applications. However, the development of novel strategies for surface modification and coating of Fe3O4 NPs is needed to fabricate Fe3O4 NPs with improved biocompatibility. In the present study, two polymers, namely, poly(ε-caprolactone) (PCL) and poly(ethylene oxide) (PEO), were applied to produce PCL/PEO microfibers containing Fe3O4 NPs (PCL/PEO/Fe3O4 MFs) using the electrospinning method. Their physicochemical properties, especially magnetically induced hyperthermia effects, were compared to Fe3O4 NPs. The biocompatibility and immunocompatibility of PCL/PEO/Fe3O4 MFs were then tested using four types of human immune cells, namely, CD14+ monocytes, CD4+ helper, CD8+ cytotoxic T cells, and CD56+ NK cells. Monocytes were the most sensitive to PCL/PEO/Fe3O4 MFs as judged by the induction of cell death (apoptosis and necrosis) and micronuclei production, whereas other immune cells were less or not affected by the stimulation with PCL/PEO/Fe3O4 MFs. PCL/PEO/Fe3O4 MFs also did not lower the viability of normal human fibroblasts. Furthermore, a mild immunogenic response was revealed in PCL/PEO/Fe3O4 MF-treated helper T cells based on the analysis of transcriptional activity of 92 genes involved in the NFκB pathway. Observed elevated mRNA levels of NFKB2, TNF, TNFAIP3, TRAF1, and TBK1 may have context-dependent immunomodulatory effects in PCL/PEO/Fe3O4 MF-stimulated helper T cells that should be taken into account while designing novel drug-delivery systems based on PCL/PEO and Fe3O4 NPs.

In vivo, cells are exposed to a dynamic extracellular matrix (ECM) that delivers biochemical, biophysical, and electrical cues essential for regulating cellular behavior. However, most existing in vitro culture systems remain static and fail to reproduce the time-dependent mechanical and electrical signals that characterize native tissues. Developing biomimetic and dynamic platforms that bridge this gap is therefore critical for advancing mechanobiology, tissue engineering, or drug screening studies. Here, we report a 4D cell culture platform based on polyHIPE-PEDOT scaffolds, combining the structural versatility of polyHIPE architectures synthesized from high internal phase emulsion with the electroactive behavior of the conducting polymer poly(3,4-ethylenedioxythiophene) (PEDOT). The resulting electroactive scaffolds exhibit a highly interconnected porous structure with tunable morphology and demonstrate reversible pore deformation under electrical stimulation. To facilitate a broad range of experimental applications, we designed two complementary devices: one optimized for in vitro cell culture under standard incubator conditions, and another tailored for real-time live-cell imaging. To illustrate this integrated workflow, we provide examples with fibroblast cells cultured under electromechanical stimulation using our two devices. The objective of this workflow is to facilitate novel insights into the study of time-resolved cell-matrix interactions and to develop responsive biomimetic microenvironments for advanced biomedical research.

Cellular agriculture offers a transformative approach to protein production by creating biologically derived alternatives to animal-sourced biomaterials. This study focuses on the comprehensive characterization of bovine collagen produced via cellular agriculture. A bovine fibroblast cell line optimized for collagen production was cultured in a custom bioreactor system to produce collagen (≥95% type I), which was extracted, purified, lyophilized, reconstituted at 8 mg/mL, and cast into self-assembling fibrillar hydrogels (Ø8 × 4 mm2) at neutral pH and 37 °C. Functional properties of this cell-based (CB) collagen were benchmarked against traditional animal-derived (AD) bovine collagen. SDS-PAGE confirmed their molecular similarity, revealing characteristic α-chains (∼120 kDa), β-dimers (∼240 kDa), and γ-trimers (∼300 kDa). Amino acid analysis showed nearly identical amino acid profiles, and differential scanning calorimetry analysis highlighted equivalent thermal stability between them. Furthermore, circular dichroism spectroscopy confirmed preservation of the triple-helical structure in the CB collagen. Mechanical testing demonstrated that CB hydrogels possessed significantly higher compressive and dynamic moduli (1260 ± 191 Pa and 6250 ± 1031 Pa, respectively) compared to the AD hydrogels (890 ± 255 Pa and 3134 ± 1857 Pa, respectively) (p < 0.05), indicating better structural integrity and shape retention. Rheological analysis supported these findings, indicated by the higher storage and loss moduli of CB hydrogels (48.40 ± 8.5 Pa and 13.20 ± 1.3 Pa, respectively) than AD hydrogels (25 ± 2.12 Pa and 10 ± 1.58 Pa, respectively) (p < 0.05). Surface wettability testing highlighted comparable hydrophilicity for CB and AD collagens. Finally, the biocompatibility of the CB hydrogels was evident from the notable increase in cultured fibroblast cell density, live cell area, and metabolic activity over 7 days. This comparative assessment underscores the functional equivalence of CB collagen with conventional animal-derived sources and offers critical insights for optimizing cellular agriculture-based production systems and hydrogel formulations for application-specific requirements in the future.

When spheroids are used as building blocks in scaffold-based tissue regeneration strategies, two successive steps are typically involved: the fabrication of the spheroids using dedicated techniques and their incorporation into the scaffold. Here, we studied a scaffold-based approach involving a non-cell-adhesive porous biomaterial, where the cell suspension is seeded directly onto the scaffold, spontaneously forming spheroids within its pores. This "two-step-in-one" route is illustrated with a cross-linked pullulan/dextran hydrogel supplemented with hydroxyapatite (HA) particles (6% w/w in the dry state), entirely dispersed within the hydrogel phase, and textured by freeze-drying. Pores and spheroids were characterized in 3D using state-of-the-art tomographic techniques. The method was applied to the formation and in vitro coculture of MSCs/HUVECs spheroids for vascularized bone tissue engineering. Cells suspended in culture medium were seeded onto the hydrogel scaffold initially in its dry state, and the gel swelled concomitantly with seeding. The scaffold porosity decreased from 80% to 25%, the mean pore size decreased from 350 to 180 μm, and the pores underwent a buckling transition, evolving from spherical to lenticular (sphericity decreased from 0.71 to 0.49). At the same time, the cells condensed into grape-like aggregates, accelerated by two mechanisms: increased local cell density resulting from suction of the culture medium by the dry gel and geometric confinement imposed by shrinking pores. Then, the cell aggregates compacted with a characteristic time of about 3 h, transitioning from a lenticular shape inherited from the pores to a spheroidal shape (mean size of 130 μm, mean sphericity of 0.82). Later, the cellularized HA-supplemented hydrogel scaffolds induced early phase bone differentiation (ALP peaks at day 7), whereas pristine hydrogel scaffolds did not, although cells were cultured in a calcifying medium. Further osteogenic maturation was observed for spheroids cocultured in HA-supplemented scaffolds under dynamic conditions, which exhibited mineralized extracellular matrix. This led us to re-examine the physical mechanisms underlying HA's osteoinductive properties. Together, these findings inform scaffold-based tissue-engineering strategies and open an alternative way for the high-throughput production of mature spheroids.

Spinal fusion is widely performed to treat instability of various spinal pathologies. Currently, the clinical gold standard is still autografts, which unfortunately are limited by donor site morbidity, restricted supply, and inconsistent outcomes. To overcome these challenges, we developed an organic-inorganic nanohybrid (click-ON) cement based on poly(propylene fumarate) (PPF) polymers cross-linked through the strain-promoted azide-alkyne cycloaddition (SPAAC) bioorthogonal click chemistry. This catalyst-free system cures rapidly in situ without external energy or toxic initiators, enabling safe and practical surgical handling. The cement was further reinforced with osteogenic nanohydroxyapatite (nHA) to improve osteoinductivity and microspheres releasing bioactive recombinant human bone morphogenetic protein 2 (rhBMP-2) and recombinant human vascular endothelial growth factor (rhVEGF) to promote coupled osteogenesis and angiogenesis. In a sheep lumbar spinal fusion model, the click-ON cement was implanted with an injectable formulation in the interbody space and a moldable formulation in the posterolateral fusion site with clinically used autograft/rhBMP-2 as the positive control. Longitudinal CT imaging demonstrated fusion with bridging bone formation across both the interbody space and posterolateral region after 6 months. Histological analyses confirmed extensive new bone deposition, integration with host tissue, and vascular ingrowth within the cement, while immunohistochemical staining showed the colocalization of CD31 and alkaline phosphatase (ALP), indicating active angiogenesis and osteogenesis, respectively. The outcomes are comparable to the positive control, which are clinical gold standard bone grafts. Manual palpation further verified the mechanical stability of the fused segments in sheep with a bioactive click-ON cement. These results established click-chemistry-enabled PPF-based cement as a promising alternative to autografts, offering advantages in moldability, biological activity, and functional fusion outcomes.

Effective treatment of infected wounds requires simultaneous eradication of invading pathogens and scavenging of excess reactive oxygen species (ROS). Herein, we fabricated a multifunctional hydrogel dressing (denoted as PVA/CG/Cu-Cys) by hydrogen-bond cross-linking poly(vinyl alcohol) with ι-carrageenan-Ca2+ helices to generate a double network that was subsequently loaded with copper-l-cysteine nanoparticles. The reversible hydrogen-bonded interpenetrating network provided adequate swelling, moldability, mechanical strength, and tissue adhesion. The Cu-Cys nanoparticles endowed the hydrogel with broad-spectrum antioxidant activity, achieving over 60% scavenging efficiency against a range of reactive oxygen (H2O2, •OH, and •O2-) and nitrogen species (DPPH• and ABTS•+). Controlled release of Cu2+ eradicated 50.84 ± 8.21% of S. aureus biofilms and inhibited both E. coli and S. aureus. At the acidic pH of infected wound area (5.6), the hydrogel released only 56.5 ± 3.0% of total Cu2+ in 24 h, significantly less than free Cu-Cys (81.1 ± 2.7%, p = 0.0025), thereby reducing cytotoxicity and maintaining >80% cell viability. The dressing promoted in vitro cell migration and accelerated closure of infected full-thickness wounds in vivo. These results indicate that PVA/CG/Cu-Cys is a readily fabricated, clinically translatable dressing for the management of bacterially infected wounds.

Cardiovascular implants are limited by the lack of materialsthatcombine mechanical strength with bioactivity to support endothelialfunction, prevent thrombosis, and promote long-term integration. Conventionalpolymers often lack these biological features, underscoring the needfor multifunctional biomaterials. This study evaluates poly­[(l-lactide)-co-(ε-caprolactone)]/poly­(d-lactide) (PLLA-co-CL/PDLA) blends processed via3D printing and electrospinning and assesses their suitability ascardiovascular implant coverings through material characterizationand endothelial cell interactions. Blend compositions were fabricatedas films and meshes, and optimized constructs were selected for cytocompatibilityand endothelial biofunctionality testing. Endothelial responses wereevaluated over 7 days through viability assays and quantificationof vasoactive mediators, including Thromboxane B2 (TXB2), Prostacyclin (PGI2), and Nitric Oxide (NO).Among all formulations, the 90:10 PLLA-co-CL/PDLAblend exhibited the most favorable wettability (76.5° ±4.0), indicating enhanced hydrophilicity at intermediate compositions.Electrospun meshes supported high HUVEC viability (>80%) and metabolicactivity, while 90:10 films maintained ∼67% viability after7 days. Functional analysis showed increasing PGI2 releaseover time, reaching ∼966 pg/mL on 90:10 films, whereas TXB2 rose moderately. The PGI2/TXB2 ratioshifted from pro-thrombotic (<1) on day 1 to antithrombotic (>1)by day 7, indicating endothelial stabilization. NO release increasedto ∼0.82 μmol/L on films and ∼0.9 μmol/Lon meshes, reflecting improved endothelial function and vascular homeostasis.Hemolytic potential was assessed via red blood cell lysis assays.All films and meshes exhibited negligible hemolysis (0.027–0.660%),comparable to a silicone elastomer reference (0.153%) and substantiallylower than slightly hemolytic controls (Buna-N, 3.177%), indicatingminimal erythrocyte membrane disruption. Overall, PLLA-co-CL/PDLA blendsparticularly 3D-printed films at a 90:10 ratioprovidehydrophilic, cyto-compatible surfaces that support endothelial viabilityand balanced vasoactive signaling. These findings highlight theirpotential as biodegradable cardiovascular implant coverings.

Osteochondral defects are severe degenerative lesions of the joint that involve the articular cartilage, the cartilage-bone interface, and the underlying subchondral bone. Surgical approaches that have been developed (joint debridement, microfracture, autologous chondrocytes and autologous/allograft cartilage implantation, and whole joint replacement) remain, however, more palliative than curative. Therefore, novel strategies are required, including promising options offered by osteochondral tissue engineering approaches. In this study, a novel chitosan-based scaffold with biomimetic and biodegradable properties was implanted for 16 weeks in a sheep osteochondral defect model in order to evaluate its physical stability, biocompatibility, and regenerative potential. Alongside the chitosan construct, a commercial scaffold made of collagen and ß-TCP was implanted as a positive control, while defects left empty were used as negative control. Postoperative recovery and tissue response were evaluated at different levels including activity monitoring, gross tissue evaluation at sacrifice, imaging, and histological analysis. Despite a transient reaction observed in the first 4 weeks after implantation, the chitosan-based scaffold was well-tolerated, with no clear signs of local or systemic reaction in the long term. The chitosan scaffold was able to support the growth of mature bone tissue with a lamellar conformation at similar levels to the predicate control. It also promoted the formation of a new cartilage layer, although not at a fully mature stage. These proof-of-concept results suggest that the novel chitosan scaffold tested constitutes a possible option for osteochondral regeneration, which should be further analyzed in complementary models to provide information on its long-term performance.

Dental caries, the most common oraldisease with a worldwideburden,causes structural damage to enamel and dentine. Considering the limitationsof conventional restorative approaches and the dynamic nature of caries,biomimetic remineralization strategies offer a promising alternativefor restoring tooth structure. Central to many of these strategiesis the use of polymers to emulate the role of macromolecular templatesin natural mineralization processes. This review summarizes the polymersapplied for biomimetic mineralization of enamel and dentine accordingto their primary functions. Furthermore, perspectives on the trendsand challenges for future research on polymeric biomaterials for enameland dentine repair are also presented.

This review systematically summarizes recent advances in multistimuli-responsive smart hydrogels, with a focus on their response mechanisms, synthesis strategies, and broad applications in the biomedical field. Based on a hydrophilic three-dimensional cross-linked network structure, smart hydrogels can sensitively respond to external stimuli such as temperature, pH, light, electricity, magnetic fields, and enzymes, enabling functions such as structural transformation, controlled drug release, and tissue repair. The article elaborates on various synthesis strategies, including chemical and physical cross-linking, microfluidics and 3D printing, double-network and nanocomposite structures, DNA-based biohydrogels, and self-healing hydrogels. It also analyzes specific response mechanisms─electrical, thermal, photoresponsive, magnetic, pH, and enzymatic─along with their application cases in drug delivery, wound healing, neural repair, and tumor therapy. Although challenges remain in functional integration and clinical translation, such as mechanical performance, biosafety, and scalable production, future developments involving multiresponse synergy, personalized design, and intelligent regulation are expected to promote their applications in precision medicine and flexible electronics.