Cellulose acetate (CA) is an attractive and sustainable biopolymer for biomedical applications due to its biocompatibility, tunable physicochemical properties, and compatibility with microfabrication. Mechanical stimuli such as topographical cues are known to profoundly influence stem cell behavior, yet their employment in cellulose acetate systems remains underexplored. In this study, we present a reproducible soft-lithography approach to engineer CA scaffolds with well-defined microstructured topographies and controlled surface roughness. The interfacial characteristics of the resulting biointerfaces were assessed using atomic force microscopy. Mesenchymal stem cells cultured on these microstructured platforms exhibited enhanced adhesion, cytoskeletal organization, and focal adhesion formation compared with flat CA controls. Notably, surfaces with intermediate roughness modulated the nuclear translocation of the mechanosensitive regulator TAZ, relative to flat and high roughness surfaces, in a topography dependent manner, suggesting activation of adhesive and cytoskeletal signaling pathways. Under osteogenic conditions, these biointerfaces further supported elevated alkaline phosphatase activity and osteopontin expression, indicative of enhanced early osteogenic commitment. Overall, this work demonstrates that microstructured CA biointerfaces act as instructive platforms that modulate mesenchymal stem cell response through topography-mediated mechanical cues, highlighting their potential as sustainable platforms for bone tissue engineering applications.

Two-photon polymerization (2PP) enables the fabrication of complex 3D scaffolds for tissue engineering, yet the lack of biocompatible and bioactive photoinitiators remains a bottleneck for functional bio-manufacturing. Here, a one-step strategy is presented using curcumin (CUR), a naturally derived polyphenol, as a high-efficiency multifunctional photoinitiator for gelatin methacryloyl (GelMA) hydrogels. Nonlinear optical characterization reveals a synergistic interaction between CUR and the GelMA protein backbone, resulting in a significantly enhanced two-photon absorption cross-section (σ ≈ 1500 GM) that outperforms conventional initiators. This synergy facilitates an exceptionally broad processing window and high-speed fabrication (up to 52 mm/s) of high-fidelity, complex triply periodic minimal surface (TPMS) and biomimetic bone-like scaffolds. Beyond its initiation efficiency, CUR imparts intrinsic multifunctionality to the resulting 3D constructs. The scaffolds exhibit exceptional biocompatibility with mesenchymal stem cells while demonstrating a potent dual-action antimicrobial defense: a selective passive antifouling effect against E. coli and active photodynamic eradication (>99.9%) of S. aureus upon blue LED irradiation. This "all-in-one" approach transforms the photoinitiator from an inert processing tool into a source of intrinsic scaffold bioactivity, eliminating the need for complex post-fabrication functionalization.

Laser-induced microfoaming has emerged as an effective strategy for structuring polysaccharide-based films and generating porous architectures with controlled physicochemical properties. However, chitosan foams often suffer from structural instability in aqueous environments. In the present study, chitosan–mastic gum composite films are investigated, where the incorporation of mastic gum, a natural resin with established mechanical robustness and pharmaceutical relevance, enables stabilization of laser-induced porous structures and modulation of surface and mechanical properties. Mastic gum incorporation stabilizes laser-patterned porous architectures while enabling composition-dependent control of mechanical, wettability, and biological properties, resulting in biodegradable matrices with tunable structural and physicochemical characteristics. FTIR spectroscopy indicated the preservation of polymeric structures and intermolecular interactions, while mechanical and wettability analyses demonstrated composition-dependent behavior relevant to biomedical performance. Preliminary in vitro studies demonstrated favorable cytocompatibility of the laser-structured composite surfaces. Overall, these results demonstrate that laser-induced microfoaming enables controlled porous structuring of chitosan-based films, while the incorporation of mastic gum tunes the mechanical and physicochemical properties of the resulting composites, highlighting their potential as tunable polysaccharide-based composite platforms for applications in tissue engineering and drug delivery.

This study presents the development and characterization of cellulose acetate (CA) and CA reinforced with 5 wt % hydroxyapatite (CAHA5) as printable bioinks for extrusion-based 3D printing of scaffolds targeting bone tissue engineering. The printed scaffolds were evaluated for morphology, mechanical performance, surface characteristics, and biological response. Scanning electron microscopy (SEM), Fourier transform infrared (FT-IR) spectroscopy, and energy-dispersive X-ray spectroscopy (EDS) confirmed scaffold integrity and successful HA incorporation, while contact angle, degradation, and swelling measurements revealed tunable surface wettability and fluid uptake. Mechanical testing under compression and tension showed that HA incorporation reduced strength and increased brittleness compared to pure CA, while stiffness values remained within reported ranges for printed scaffolds. Biological assays using mice mesenchymal stem cells (MSCs) showed favorable adhesion and osteogenic differentiation, particularly on CAHA5 scaffolds. These findings suggest that CAHA5 bioinks offer a promising route for fabricating biocompatible and osteoinductive scaffolds, where enhanced bioactivity is achieved despite a moderate reduction in mechanical strength compared to pure CA.

In this study, 3D-printed scaffolds composed of poly(ε-caprolactone) (PCL) and PCL reinforced with 20 wt% cellulose acetate (PCLCA20) were fabricated and evaluated for bone tissue engineering applications. A wall-free woodpile architecture was deliberately introduced to promote multidirectional cell infiltration and nutrient diffusion. Material characterization confirmed the preservation of chemical stability and demonstrated increased hydrophilicity and swelling capacity in the composite scaffolds. Mechanical testing revealed that cellulose acetate incorporation significantly enhanced tensile performance while maintaining sufficient compressive stability. Biological assessment showed improved mesenchymal stem cell adhesion, proliferation, and mineralization on PCLCA20 scaffolds compared to PCL, underscoring the bioactive role of cellulose acetate. Collectively, these findings highlight the synergistic contribution of cellulose acetate reinforcement and wall-free architecture, positioning PCLCA20 scaffolds as promising candidates for load-bearing bone regeneration.

Considering that neurological injuries cannot typically self-recover, there is a need to develop new methods to study neuronal outgrowth in a controllable manner in vitro. In this study, a precise flow-controlled microfluidic system featuring custom-designed chambers that integrate laser-microstructured polyethylene terephthalate (PET) substrates comprising microgrooves (MGs) was developed to investigate the combined effect of shear stress and topography on Neuro-2a (N2a) cells’ behavior. The MGs were positioned parallel to the flow direction and the response of N2a cells was evaluated in terms of growth and differentiation. Our results demonstrate that flow-induced shear stress could inhibit the differentiation of N2a cells. This microfluidic system could potentially be used as a new model system to study the impact of shear stress on cell differentiation.

This study explores the development and characterization of iron oxide nanoclusters (NCs) functionalized with vascular cell adhesion molecule 1 (VCAM-1) for targeted magnetic resonance imaging (MRI) of early atherosclerotic lesions. The NCs were synthesized via a high-temperature polyol method and functionalized using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) chemistry to enable conjugation with VCAM-1 antibodies. Dynamic light scattering and transmission electron microscopy TEM confirmed controlled growth of NCs with a size ranging from 40 nm, in the parent to 110 nm post-functionalization, maintaining though colloidal stability in aqueous media. Cytotoxicity assays using mesenchymal stem cells (MSCs) demonstrated high biocompatibility. Confocal and electron microscopy confirmed specific binding of VCAM-1-NCs to VCAM-1-overexpressing MSCs under inflammatory conditions, with internalization through the endolysosomal pathway. The functionalized NCs remained bound under shear stress in an orbital flow model, mimicking early atherosclerotic conditions. MRI phantom analysis demonstrated preserved contrast capability despite increased T 2 * relaxation times following antibody conjugation. These findings highlight the potential of VCAM-1-NCs as noninvasive imaging agents for early-stage atherosclerosis and vascular inflammation. Although this study is limited by the lack of in vivo validation and therapeutic evaluation, it provides a strong foundation for future translational research.

The non-linear modality of Third Harmonic Generation (THG) can discriminate between senescent and nonsenescent fibroblasts by detecting a distinct lipid content increase of senescent cells, offering a noninvasive approach towards the characterization of cellular senescence. (c) 2025 The Author(s).

Additive manufacturing holds significant potential in the field of tissue engineering, particularly for healing, replacing, and regenerating damaged or diseased tissues. However, the high cost of commercially available bioprinters and the limited availability of suitable biomaterials for bioprinting have hindered its widespread implementation and practical application in clinical settings. The aim of this study was to identify printing parameters tailored to the viscosity of the bioink and the evaporation characteristics of the organic solvent used in its formulation, with the broader goal of developing a cost-effective and accessible bioprinting platform for scaffold fabrication. To this end, we present a novel approach involving the design and fabrication of a cost-effective three-dimensional (3D) bioprinter conversion kit, developed using commercially available 3D printers. Bioprinting high-viscosity bioinks present specific challenges due to their resistance to flow and a high tendency to clog printing nozzles; however, this issue was mitigated through comprehensive rheological characterization. By leveraging the favorable properties of cellulose acetate as the chosen biomaterial, scaffold fabrication via 3D bioprinting was achieved efficiently without the need for curing or post-processing steps. Furthermore, a parametric troubleshooting procedure was developed to optimize printing parameters, elucidate the material behavior, and improve scaffold resolution, as assessed through scanning electron microscopy. Additionally, preliminary cell culture studies were carried out to evaluate the influence of the printed scaffolds’ biophysical cues on cellular responses, including adhesion and proliferation. This innovative and cost-effective solution has great potential to support researchers in tissue engineering and facilitate further exploration of advanced bioprinting techniques.

The interplay between the extracellular matrix and cells significantly impacts cellular survival, proliferation, and differentiation. Cell growth within 3D scaffolds, particularly hydrogels that mimic cellular microenvironments, offers more relevant insights into tissue development compared to traditional 2D systems. This study explores the behavior of neural stem cells and their differentiation within 3D pure adipose tissue derived-ECM (adECM) hydrogels. These hydrogels provide both physical and biochemical cues that closely resemble the 3D microarchitecture of native tissues. Encapsulating neuroectodermal NE-4C cells in adECM hydrogels at different concentrations revealed intriguing divergent cellular responses. While variations in the fiber structure and pore formation between hydrogels did not significantly affect cell survival, they notably influenced the differentiation process. Analysis of neural-lineage-specific markers, such as tubulin beta III and GFAP, demonstrated divergent differentiation outcomes. This biologically derived, tissue-specific 3D platform enables in vitro study of neural differentiation and lays the groundwork for future neural models relevant to regenerative medicine and neurodegenerative research.

Cellular senescence, a state of irreversible growth arrest in response to stress, plays a dual role in physiology and pathology. While essential for processes such as embryogenesis, wound healing, and tumor suppression, senescence also contributes to aging and age-related diseases, including cancer and neurodegeneration. The accumulation of senescent cells is linked to aging and numerous age-associated pathologies, making the detection of these cells crucial for understanding and potentially mitigating age-related diseases. Lipid metabolism is a key feature of senescent cells, which undergo significant alterations in lipid composition that influence membrane remodeling and cellular function. Here, we propose the use of third harmonic generation (THG) microscopy, a label-free imaging modality, to assess lipid profiles in senescent and nonsenescent cells. Our study demonstrated that THG can discriminate between senescent and nonsenescent fibroblasts based on their lipid content, suggesting a noninvasive approach for the detection and characterization of cellular senescence. In addition, these findings reveal that lipid content is increased in senescent cells. This methodology has potential applications in the diagnosis and study of age-related pathologies where lipid dysregulation is a hallmark feature.

Graphene is a nanomaterial used in health and oncology settings. However, several reports have raised the alarm about potential toxicity. This study addressed this concern and determined the in vitro cytotoxicity of few-layer graphene (FLG) flakes produced in bespoke ultrasonic reactors using benign methods. The use of graphene flakes as a potential sensitising agent and a carrier for drug delivery in cancer cells was evaluated. To this end, aqueous based FLG suspensions were systematically characterised using UV-Vis, Raman spectroscopy and High-resolution Transmission electron microscopy (HR-TEM). Cell toxicity characterisation (e.g., cell viability assays using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) and cell membrane integrity) of FLG in water were performed together with charge coupled device (CCD) and second harmonic generation (SHG) imaging of live cells in graphene solutions. Collectively, our findings show that NIH 3T3 mouse fibroblast and human fibroblast cells survival was higher than 80% and 90%, respectively upon treatment with the FLG fraction (~ 16 µg/ml ) recovered after centrifugation at 2000 revolutions per minute (RPM). In contrast, the cervical cancer cell line HeLa exposed to similar concentrations of FLG flakes resulted in approximately 30% cell death arguing in favour of a sensitising effect in cervical cancer cells.

Harnessing the distinctive attributes of three-dimensional auxetic scaffolds in applications related to tissue engineering and regenerative medicine injects fresh momentum into these domains. In this study, we present our findings regarding the creation and characterization of three-dimensional auxetic scaffolds tailored for tissue engineering applications. These scaffolds leverage the well-established re-entrant hexagonal geometry (bowtie) and are manufactured through multiphoton lithography utilizing the organic-inorganic photopolymer SZ2080. Employing in-situ scanning electron microscopy, micro-indentations, and nano-indentation experiments, we meticulously analyze the photocurable resin SZ2080 and the resultant scaffolds. Despite SZ2080 being inherently rigid with a positive Poisson's ratio, our investigation reveals that the scaffolds exhibit a negative Poisson's ratio and remarkable elasticity attributed to their specific architecture. Subsequently, we employ mouse fibroblasts to seed the scaffolds, demonstrating their capacity to efficiently infiltrate and proliferate within, conforming to the scaffold's structure to meet the cells' needs. Furthermore, the scaffold's architecture imparts a directional preference to the cells, a crucial factor in various cell-based applications within regenerative medicine. Our research lays the groundwork for the practical utilization of 3D auxetic metamaterials as cutting-edge, adaptable scaffolds in the realm of tissue engineering.

In the pursuit of advancing neural tissue regeneration, biomaterial scaffolds have emerged as promising candidates, offering potential solutions for nerve disruptions. Among these scaffolds, multichannel hydrogels, characterized by meticulously designed micrometer-scale channels, stand out as instrumental tools for guiding axonal growth and facilitating cellular interactions. This study explores the innovative application of human amniotic membranes modified with methacryloyl domains (AMMA) in neural stem cell (NSC) culture. AMMA hydrogels, possessing a tailored softness resembling the physiological environment, are prepared in the format of multichannel scaffolds to simulate native-like microarchitecture of nerve tracts. Preliminary experiments on AMMA hydrogel films showcase their potential for neural applications, demonstrating robust adhesion, proliferation, and differentiation of NSCs without the need for additional coatings. Transitioning into the 3D realm, the multichannel architecture fosters intricate neuronal networks guiding neurite extension longitudinally. Furthermore, the presence of synaptic vesicles within the cellular arrays suggests the establishment of functional synaptic connections, underscoring the physiological relevance of the developed neuronal networks. This work contributes to the ongoing efforts to find ethical, clinically translatable, and functionally relevant approaches for regenerative neuroscience.

Neural stem cells in vivo receive information from biochemical and biophysical cues of their microenvironment that affect their survival, proliferation and differentiation toward specific lineages. Recapitulation of these conditions in vitro is better achieved in 3D cell cultures. Especially the cells that grow in scaffold-dependent 3D cultures establish more complex cell-cell and cell-material interactions enabling the study of the various signaling pathways. The biochemical signaling from growth factors and hormones has been extensively studied over the years. More recently cumulative evidence demonstrates that cell sensing and response to mechanical stimuli is mediated through mechanotransduction pathways. Although individual signaling pathways activated by biochemical or mechanical cues in cells are well-studied, synergistic or antagonistic effects among them need further research to be fully understood. The understanding of the alteration of the cell behavior due to a microenvironmental cues would be greatly enhanced by the study of key elements that lie in the convergence of biochemical and mechanical pathways. Here we analyzed the effect of the substrate topography on the nerve growth factor (NGF) induced differentiation of PC12 cells. Our results showed that the topography interferes with NGF-induced neuronal differentiation and this is reflected in the reduced activation of the integrin-mediated mechanotransduction.

In this study, we revealed a peculiar morphological feature of 50B11 nociceptive sensory neurons in in vitro culture related to the forskolin-induced differentiation of these cells growing upside-down on cover glass supports. Multi-photon non-linear microscopy was applied to monitor increased neurite arborization and elongation. Under live and unstained conditions, second harmonic generation (SHG) microscopy could monitor microtubule organization inside the cells while also correlating with the detection of cellular multi-photon autofluorescence, probably derived from mitochondria metabolites. Although the differentiated cells of each compartment did not differ significantly in tubulin or multi-photon autofluorescence contents, the upturned neurons were more elongated, presenting a higher length/width cellular ratio and longer neurites, indicative of differentiated cells. SHG originating from the axons’ microtubules represented a proper tool to study neurons’ inverted culture in live conditions without exogenous staining. This work represents the first instance of examining neuronal cell lines growing and differentiated in an upside-down orientation, allowing a possible improvement of 50B11 as a model in physiology studies of sensory neurons in peripheric nervous system disease (e.g., Fabry disease, Friedreich ataxia, Charcot–Marie–Tooth, porphyria, type 1 diabetes, Guillain–Barré syndrome in children) and analgesic drug screening.

Whereas the axons of the peripheral nervous system (PNS) spontaneously regenerate after an injury, the occurring regeneration is rarely successful because axons are usually directed by inappropriate cues. Therefore, finding successful ways to guide neurite outgrowth, in vitro, is essential for neurogenesis. Microfluidic systems reflect more appropriately the in vivo environment of cells in tissues such as the normal fluid flow within the body, consistent nutrient delivery, effective waste removal, and mechanical stimulation due to fluid shear forces. At the same time, it has been well reported that topography affects neuronal outgrowth, orientation, and differentiation. In this review, we demonstrate how topography and microfluidic flow affect neuronal behavior, either separately or in synergy, and highlight the efficacy of microfluidic systems in promoting neuronal outgrowth.

Additive manufacturing is gaining popularity for its limitless possibilities in fields requiring 3D structure fabrication, including tissue engineering. While established methods like Multi-Photon Polymerization and Selective Laser Melting offer high-resolution structures, they exhibit common drawbacks such as high cost, large equipment size and long fabrication times. On the other hand, 3D printers that utilize UV Polymerization are becoming more affordable, allowing for printing 3D scaffolds that could be used as transplantable grafts for tissue regeneration and other biomedical applications. In this study, we used a Light-Emitting-Diode-based 3D printer with a pixel size of 35 μm alongside two commercially available resins to fabricate a variety of high-resolution, large scaffold geometries. Resin and scaffold suitability was examined using 4 common cell lines and various techniques such as MTT cell viability assays, confocal microscopy and Scanning Electron Microscopy. The results highlight the possibility to successfully use widely-available, low-cost 3D printers for high-resolution scaffold fabrication while maintaining cell culture compatibility for biomedical applications.