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Porous titanium addresses the longstanding orthopedic challenges of aseptic loosening and stress shielding. This work expands on the evolution of porous Ti with the manufacturing of hierarchically porous, low stiffness, ductile Ti scaffolds via direct-ink write (DIW) extrusion and sintering of inks containing Ti and NaCl particles. Scaffold macrochannels were filled with a subtherapeutic dose of recombinant bone morphogenetic protein-2 (rhBMP-2) alone or co-delivered within a bioactive supramolecular polymer slurry (SPS) composed of peptide amphiphile nanofibrils and collagen, creating four treatment conditions (Ti struts: microporous vs. fully dense; BMP-2 alone or with SPS). The BMP-2-loaded scaffolds were implanted bilaterally across the L4 and L5 transverse processes in a rat posterolateral lumbar fusion model. In-vivo bone growth in these scaffolds is evaluated with synchrotron X-ray computed microtomography (µCT) to study the effects of strut microporosity and added biological signaling agents on the bone formation response. Optical and scanning electron microscopy confirms the ∼100 µm space-holder micropore size, high-curvature morphology, and pore fenestrations within the struts. Uniaxial compression testing shows that the microporous strut scaffolds have low stiffness and high ductility. A significant promotion in bone formation was observed for groups utilizing the SPS, while no significant differences were found for the scaffolds with the incorporation of micropores. STATEMENT OF SIGNIFICANCE: By 2050, the anticipated number of people aged 60 years and older worldwide is anticipated to double to 2.1 billion. This rapid increase in the geriatric population will require a corresponding increase in orthopedic surgeries and more effective materials for longer indwelling times. Titanium alloys have been the gold standard of bone fusion and fixation, but their use has longstanding limitations in bone-implant stiffness mismatch and insufficient osseointegration. We utilize 3D-printing of titanium with NaCl space holders for large- and small-scale porosity and incorporate bioactive supramolecular polymers into the scaffolds to increase bone growth. This work finds no significant change in bone ingrowth via space-holder-induced microporosity but significant increases in bone ingrowth via the bioactive supramolecular polymers in a rat posterolateral fusion model., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: DCD maintains a financial interest in Metalprinting, Inc. (South Korea) which is active in ink-based materials printing. SIS, NAS, and ELH report a relationship with Amphix Bio that includes equity or stocks. SIS and ELH are inventors on patent #US20170106120A1 issued to Northwestern University., (Copyright © 2024 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

The tumor microenvironment (TME) orchestrates cellular and extracellular matrix (ECM) interactions, playing a key role in tumorigenesis, tumor growth, and metastization. Investigating the interplay between stromal-epithelial cells within the TME is paramount for understanding cancer mechanisms but demands reliable biological models. 3D-models have emerged as powerful in vitro tools, but many fall short in replicating cell-cell/cell-matrix interactions. This study introduces a novel hybrid 3D-model of the breast TME, combining epithelial cells, cancer-associated fibroblasts (CAFs), and their ECM. To build the stromal compartment, porous 3D-printed alginate scaffolds were seeded with CAFs, which proliferated and produced ECM. The pores were infused with oxidized peptide-modified alginate hydrogel laden with MCF10A cells, forming the parenchymal compartment. The hybrid system supported epithelial morphogenesis into acini surrounded by fibroblasts and ECM, and could be readily solubilized to recover cells, their matrix, and sequestered soluble factors. Proteome profiling of the retrieved ECM showed upregulation of proteins associated with matrix assembly/remodeling, epithelial-to-mesenchymal transition (EMT), and cancer. The TME-like microenvironment induced a partial EMT in MCF10A cells, generating a hybrid population with epithelial and mesenchymal features, characteristic of aggressive phenotypes. Our model provided new insights into epithelial-stromal interactions within the TME, offering a valuable tool for cancer research in a physiologically-relevant 3D setting., 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 Ltd.. All rights reserved.)

The complexity in structure and function of the nervous system, as well as its slow rate of regeneration, makes it more difficult to treat it compared to other tissues. Neural tissue engineering aims to create an appropriate environment for nerve cell proliferation and differentiation. Fibrous scaffolds with suitable morphology and topography and better mimicry of the extracellular matrix have been promising for the alignment and migration of neural cells. On this premise, to improve the properties of the scaffold, we combined montmorillonite (MMT) with chitosan (CS) polymer and created microfibers with variable diameters and varied concentrations of MMT using microfluidic technology and tested its suitability for the rat pheochromocytoma cell line (PC12). According to the findings, CS/MMT 0.1 % compared to CS/MMT 0 % microfibers showed a 201 MPa increase in Young's modulus, a 68 mS/m increase in conductivity, and a 1.4-fold increase in output voltage. Analysis of cell mitochondrial activity verified the non-toxicity, resulting in good cell morphology with orientation along the microfiber. Overall, the results of this project showed that with a low concentration of MMT, the properties of microfibers can be significantly improved and a suitable scaffold can be designed for neural tissue engineering., 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. Prof. Mohammad Abdollahi is an Editor-in-Chief for Daru Journal and was not involved in the editorial review or the decision to publish this article., (Copyright © 2024. Published by Elsevier Ltd.)

Mesenchymal stem cell-derived exosomes (MSC-Exos) have emerged as promising candidates for cell-free therapy in tissue regeneration. However, the native osteogenic and angiogenic capacities of MSC-Exos are often insufficient to repair critical-sized bone defects, and the underlying immune mechanisms remain elusive. Furthermore, achieving sustained delivery and stable activity of MSC-Exos at the defect site is essential for optimal therapeutic outcomes. Here, we extracted exosomes from osteogenically pre-differentiated human bone marrow mesenchymal stem cells (hBMSCs) by ultracentrifugation and encapsulated them in gelatin methacryloyl (GelMA) hydrogel to construct a composite scaffold. The resulting exosome-encapsulated hydrogel exhibited excellent mechanical properties and biocompatibility, facilitating sustained delivery of MSC-Exos. Osteogenic pre-differentiation significantly enhanced the osteogenic and angiogenic properties of MSC-Exos, promoting osteogenic differentiation of hBMSCs and angiogenesis of human umbilical vein endothelial cells (HUVECs). Furthermore, MSC-Exos induced polarization of Raw264.7 cells from a pro-inflammatory phenotype to an anti-inflammatory phenotype under simulated inflammatory conditions, thereby creating an immune microenvironment conducive to osteogenesis. RNA sequencing and bioinformatics analysis revealed that MSC-Exos activate the p53 pathway through targeted delivery of internal microRNAs and regulate macrophage polarization by reducing DNA oxidative damage. Our study highlights the potential of osteogenic exosome-encapsulated composite hydrogels for the development of cell-free scaffolds in bone tissue engineering., 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. Published by Elsevier Inc.)

In this study, we report on the fabrication of hybrid nanofibers for labeling and bioimaging applications. Our approach is involved for developing highly fluorescent nanofibers using a blend of polylactic acid, polyethyleneglycol, and perylenediimide dyes, through the solution blow spinning technique. The nanofibers are exhibited diameters ranging from 330 nm to 420 nm. Nanofibers showed excellent red and near-infrared fluorescence emissive properties in fluorescent spectroscopy. Moreover, the strong two-photon absorption phenomenon was observed for nanofibers under confocal microscopy. To assess the applicability of these fluorescent nanofibers in bioimaging settings, we employ two types of mammalian cells B16F1 melanoma cells and J774.A1 macrophages. Both cell types exhibit negligible cytotoxicity after 24 h incubation with the nanofibers, indicating the suitability of nanofibers for cell-based experiments. We also observe strong interactions between the nanofibers and cells, as evidenced by two major events: a) the acquisition of an elongated cellular morphology with the major cellular axis parallel to the nanofibers and b) the accumulation of actin filaments along the points of contact of the cells with the fibers. Our findings demonstrate the suitability of these newly developed fluorescent nanofibers in cell-based applications for guiding cellular behavior. We expect that these fluorescent nanofibers have the potential to serve as scaffold materials for long-time tracking of cell-fiber interactions in fluorescence microscopy., 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.)

Microtumor models, combining cancer and stromal cells within 3D hydrogels, are vital for testing anticancer therapies. Bioprinting hydrogel scaffolds allows tailored in vitro models. We created a 3D microtumor model using a bioprinter, with varying ratios of ovarian stromal cells and leukemia cells (HL-60). PEGylated fibrinogen and alginate hydrogel were used. Cell dynamics and proliferation were assessed via immunofluorescence staining. Microtumors with different HL-60 ratios (1:1, 1:10, 1:100) were cultured for 5 days. Results showed tumor development modulation by cell ratios and culture time. A significant cell density increase occurred in 1:1 ratio microtumors, indicating rapid cancer cell proliferation. No HL-60 cells were found in 1:100 ratio microtumors by day 5. The 1:10 ratio closely mimicked leukemia invasion in ovarian tissue, showing detectable cancer cells by days 3 and 5 without altering total cell density dynamics significantly. This bioprinted leukemia microtumor model offers better physiological relevance than 2D assays, promising applications in cellular analysis and drug screening., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Christiani A. Amorim reports financial support was provided by Louvain Foundation. If there are other authors, they 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 Inc. All rights reserved.)

The integration of functional nanomaterials with tissue engineering scaffolds has emerged as a promising solution for simultaneously treating malignant bone tumors and repairing resected bone defects. However, achieving a uniform bioactive interface on 3D-printing polymer scaffolds with minimized microstructural heterogeneity remains a challenge. In this study, we report a facile metal-coordination self-assembly strategy for the surface engineering of 3D-printed polycaprolactone (PCL) scaffolds with nanostructured two-dimensional conjugated metal-organic frameworks (cMOFs) consisting of Cu ions and 2,3,6,7,10,11-hexahydroxytriphenylene (HHTP). A tunable thickness of Cu-HHTP cMOF on PCL scaffolds was achieved via the alternative deposition of metal ions and HHTP. The resulting composite PCL@Cu-HHTP scaffolds not only demonstrated potent photothermal conversion capability for efficient OS ablation but also promoted the bone repair process by virtue of their cell-friendly hydrophilic interfaces. Therefore, the cMOF-engineered dual-functional 3D-printing scaffolds show promising potential for treating bone tumors by offering sequential anti-tumor effects and bone regeneration capabilities. This work also presents a new avenue for the interface engineering of bioactive scaffolds to meet multifaceted demands in osteosarcoma-related bone defects., 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 Inc. All rights reserved.)

An off-the-shelf scaffold with requisite properties could enable the viable treatment of irregular craniomaxillofacial bone defects. Notably, the scaffold should be conformally fitting, innately bioactive, and bioresorbable. In prior work, we developed a series of shape memory polymer (SMP) scaffolds based on cross-linked poly(ε-caprolactone) (PCL). These were capable of "self-fitting" into complex bone defects when exposed to temperatures above the melt transition of the constituent PCL, either linear -PCL-diacrylate ( linear -PCL-DA, T m ∼55 °C) or star -PCL-tetraacrylate ( star -PCL-TA, T m ∼45 °C) for the potential to improve tissue safety. To achieve favorably increased degradation rates versus PCL-only scaffolds, semi-interpenetrating networks (semi-IPNs) were formed by including linear - or star -poly(l-lactic acid) (PLLA). A potential limitation of these self-fitting scaffolds is the lack of bioactivity, which is essential to osteoinductivity and osseointegration. Herein, analogous composite scaffolds were formed with 45S5 bioglass (BG) to impart bioactivity. The solvent-cast particulate leaching fabrication method was adapted to introduce BG to the fused salt template, resulting in composites with BG concentrated on the pore wall surfaces rather than within pore struts. Composite scaffolds with good pore wall integrity were produced with 2.5, 5, and 10 wt % BG. All composite scaffolds exhibited non-brittle behavior and did not fracture with 85% strain. For semi-IPN composite scaffolds, PLLA crystallinity was lost, and mechanical properties were not appreciably altered versus the non-BG controls. Sufficient retention of PCL crystallinity led to excellent shape memory behavior. The inclusion of 5 and 10 wt % BG led to hydroxyapatite mineralization after 1 day of exposure to simulated body fluid, as well as increased rates of in vitro degradation.

A scaffold is a three-dimensional porous structure that is used as a template to provide structural support for cell adhesion and the formation of new cells. Metallic cellular scaffolds are a good choice as a replacement for human bones in orthopaedic implants, which enhances the quality and longevity of human life. In contrast to conventional methods that produce irregular pore distributions, 3D printing, or additive manufacturing, is characterized by high precision and controlled manufacturing processes. AM processes can precisely control the scaffold's porosity, which makes it possible to produce patient specific implants and achieve regular pore distribution. This review paper explores the potential of Ti-6Al-4V scaffolds produced via the SLM method as a bone substitute. A state-of-the-art review on the effect of design parameters, material, and surface modification on biological and mechanical properties is presented. The desired features of the human tibia and femur bones are compared to bulk and porous Ti6Al4V scaffold. Furthermore, the properties of various porous scaffolds with varying unit cell structures and design parameters are compared to find out the designs that can mimic human bone properties. Porosity up to 65% and pore size of 600 μm was found to give optimum trade-off between mechanical and biological properties. Current manufacturing constraints, biocompatibility of Ti-6Al-4V material, influence of various factors on bio-mechanical properties, and complex interrelation between design parameters are discussed herein. Finally, the most appropriate combination of design parameters that offers a good trade-off between mechanical strength and cell ingrowth are summarized., (© 2024 IOP Publishing Ltd. All rights, including for text and data mining, AI training, and similar technologies, are reserved.)

Biomaterial-based implantable scaffolds capable of promoting physical and functional reconnection of injured spinal cord and nerves represent the latest frontier in neural tissue engineering. Here, we report the fabrication and characterization of self-standing, biocompatible and bioresorbable substrates endowed with both controlled nanotopography and electroactivity, intended for the design of transient implantable scaffolds for neural tissue engineering. In particular, we obtain conductive and nano-modulated poly(D,L-lactic acid) (PLA) and poly(lactic- co -glycolic acid) free-standing films by simply iterating a replica moulding process and coating the polymer with a thin layer of poly(3,4-ethylendioxythiophene) polystyrene sulfonate. The capability of the substrates to retain both surface patterning and electrical properties when exposed to a liquid environment has been evaluated by atomic force microscopy, electrochemical impedance spectroscopy and thermal characterizations. In particular, we show that PLA-based films maintain their surface nano-modulation for up to three weeks of exposure to a liquid environment, a time sufficient for promoting axonal anisotropic sprouting and growth during neuronal cell differentiation. In conclusion, the developed substrates represent a novel and easily-tunable platform to design bioresorbable implantable devices featuring both topographic and electrical cues., (Creative Commons Attribution license.)

In recent years, biological 3D printing has garnered increasing attention for tissue and organ repair. The challenge with 3D-printing inks is to combine mechanical properties as well as biocompatibility. Proteins serve as vital structural components in living systems, and utilizing protein-based inks can ensure that the materials maintain the necessary biological activity. In this study, we incorporated two natural biomaterials, silk fibroin (SF) and collagen (COL), into a low-concentration sodium alginate (SA) solution to create novel composite inks. SF and COL were modified with glycidyl methacrylate (GMA) to impart photo-cross-linking properties. The UV light test and 1 H NMR results demonstrated successful curing of silk fibroin (SF) and collagen (COL) after modification and grafting. Subsequently, the printability of modified silk fibroin (RSFMA)/SA with varying concentration gradients was assessed using a set of three consecutive printing models, and the material's properties were tested. The research results prove that the addition of RSFMA and ColMA enhances the printability of low-concentration SA solutions, with the Pr values increasing from 0.85 ± 0.02 to 0.90 ± 0.03 and 0.92 ± 0.02, respectively, and the mechanical strength increasing from 0.19 ± 0.01 to 0.28 ± 0.01 and 0.38 ± 0.01 MPa; cytocompatibility has also been improved. Furthermore, rheological tests indicated that all of the inks exhibited shear thinning properties. CCK-8 experiments demonstrated that the addition of ColMA increased the cytocompatibility of the ink system. Overall, the utilization of SF and COL-modified SA materials as inks represents a promising advancement in 3D-printed ink technology.

The repair of critical-sized bone defects remains a major challenge for clinical orthopedic surgery. Here, we develop a surface biofunctionalized three-dimensional (3D) porous polyether-ether-ketone (PEEK) scaffold that can simultaneously promote osteogenesis and regulate macrophage polarization. The scaffold is created using polydopamine (PDA)-assisted immobilization of silk fibroin (SF) and the electrostatic self-assembly of nanocrystalline hydroxyapatite (nano-HA) on a 3D-printed porous PEEK scaffold. The SF/nano-HA functionalized surface provides a bone-like microenvironment for osteoblastic cells' adhesion, proliferation, mineralization and osteogenic differentiation. Moreover, the biofunctionalized surface can effectively drive macrophages polarization from the pro-inflammatory M1 phenotype to the anti-inflammatory M2 phenotype. Integrin β1-specific cell-matrix binding and the activation of Ca 2+ receptor-mediated signaling pathway play critical roles in the regulation of macrophage polarization. Compared with the as-printed scaffold, the SF/nano-HA functionalized porous PEEK scaffold induces minimal inflammatory response, enhanced angiogenesis, and substantial new bone formation, resulting in improved osseointegration in vivo . This study not only develops a promising candidate for bone repair but also demonstrates a facile surface biofunctionalization strategy for orthopedic implants to improve osseointegration by stimulating osteogenesis and regulating immunity.

Mesenchymal stromal cells (MSCs) have broad immunomodulatory properties that range from regulation, proliferation, differentiation, and immune cell activation to secreting bioactive molecules that inhibit inflammation and regulate immune response. These properties provide MSCs with high therapeutic potency that has been shown to be relevant to tissue engineering and regenerative medicine. Hence, researchers have explored diverse strategies to control the immunomodulatory potential of stromal cells using polymeric substrates or scaffolds. These substrates alter the immunomodulatory response of MSCs, especially through biophysical cues such as matrix mechanical properties. To leverage these cell-matrix interactions as a strategy for priming MSCs, emerging studies have explored the use of stimuli-responsive substrates to enhance the therapeutic value of stromal cells. This review highlights how stimuli-responsive materials, including chemo-responsive, microenvironment-responsive, magneto-responsive, mechano-responsive, and photo-responsive substrates, have specifically been used to promote the immunomodulatory potential of stromal cells by controlling their secretory activity.

The management of extensive bone loss remains a clinical challenge. Numerous studies are underway to develop a combination of biomaterials, biomolecules, and stem cells to address this challenge. In particular, the systemic administration of antibodies against sclerostin, a regulator of bone formation, was recently shown to enhance the bone repair efficiency of dense collagen hydrogels (DCHs) hosting murine dental pulp stem cells (mDPSCs). The aim of the present study was to assess whether these antibodies, encapsulated and released from DCHs, could promote craniofacial bone repair by the local inhibition of sclerostin. In vitro studies showed that antibody loading modified neither the hydrogel structure nor the viability of seeded mDPSCs. When implanted in a mouse calvaria critical-size bone defect, antibody-loaded DCHs showed repair capabilities similar to those of acellular unloaded DCHs combined with antibody injections. Importantly, the addition of mDPSCs provided no further benefit. Altogether, the local delivery of antisclerostin antibodies from acellular dense collagen scaffolds is highly effective for bone repair. The drastic reduction in the required amount of antibody compared to systemic injection should reduce the cost of the procedure, making the strategy proposed here a promising therapeutic approach for large bone defect repair.

Nanocellulose- and alginate-based hydrogels have been suggested as potential wound-healing materials, but their utilization is limited by the Food and Drug Administration (FDA) requirements regarding endotoxin levels. Cytotoxicity and the presence of endotoxin were assessed after gel sterilization using an autoclave and UV treatment. A new fingerprinting method was developed to characterize the compounds detected in cellulose nanocrystal (CNC)- and cellulose-nanofiber (CNF)-based hydrogels using both positive- and negative-ion mode electrospray ionization Fourier transform ion cyclotron resonance mass spectroscopy (ESI FT-ICR MS). These biobased hydrogels were used as scaffolds for the cultivation and growth of human dermal fibroblasts to test their biocompatibility. A resazurin-based assay was preferred over all other biocompatibility methodologies since it allowed for the evaluation of viability over time in the same sample without causing cell lysis. The CNF dispersion of 6 EU mL -1 was slightly above the limits, and it did not affect the cell viability, whereas CNC hydrogels induced a reduction of metabolic activity by the fibroblasts. The chemical structure of the detected endotoxins did not contain phosphates, but it encompassed hydrophobic sulfonate groups, requiring the development of new high-pressure sterilization methods for the use of cellulose hydrogels in medicine.

Bladder tissue engineering offers significant potential for repairing defects resulting from congenital and acquired conditions. However, the effectiveness of engineered grafts is often constrained by insufficient vascularization and neural regeneration. This study utilized four primary biomaterials─gelatin methacryloyl (GelMA), chitin nanocrystals (ChiNC), titanium carbide (MXene), and adipose-derived stem cells (ADSC)─to formulate two types of bioinks, GCM0.2 and GCM0.2-ADSC, in specified proportions. These bioinks were 3D printed onto bladder acellular matrix (BAM) patches to create BAM-GCM0.2 and BAM-GCM0.2-ADSC patches. The BAM-GCM0.2-ADSC patches underwent electrical stimulation to yield GCM0.2-ADSC-ES bladder patches. Employed for the repair of rat bladder defects, these patches were evaluated against a Control group, which underwent partial cystectomy followed by direct suturing. Our findings indicate that the inclusion of ADSC and electrical stimulation significantly enhances the regeneration of rat bladder smooth muscle (from [24.052 ± 2.782] % to [57.380 ± 4.017] %), blood vessels (from [5.326 ± 0.703] % to [12.723 ± 1.440] %), and nerves (from [0.227 ± 0.017] % to [1.369 ± 0.218] %). This research underscores the superior bladder repair capabilities of the GCM0.2-ADSC-ES patch and opens new pathways for bladder defect repair.

In recent years, there has been increased interest in the use of cellular spheroids, microtissues, and organoids as biological building blocks to engineer functional tissues and organs. Such microtissues are typically formed by the self-assembly of cellular aggregates and the subsequent deposition of a tissue-specific extracellular matrix (ECM). Biofabrication and 3D bioprinting strategies using microtissues may require the development of supporting hydrogels and bioinks to spatially localize such biological building blocks in 3D space and hence enable the engineering of geometrically defined tissues. Therefore, the aim of this work was to engineer scaled-up, geometrically defined cartilage grafts by combining multiple cartilage microtissues within a rapidly degrading oxidized alginate (OA) supporting hydrogel and maintaining these constructs in dynamic culture conditions. To this end, cartilage microtissues were first independently matured for either 2 or 4 days and then combined in the presence or absence of a supporting OA hydrogel. Over 6 weeks in static culture, constructs engineered using microtissues that were matured independently for 2 days generated higher amounts of glycosaminoglycans (GAGs) compared to those matured for 4 days. Histological analysis revealed intense staining for GAGs and negative staining for calcium deposits in constructs generated by using the supporting OA hydrogel. Less physical contraction was also observed in constructs generated in the presence of the supporting gel; however, the remnants of individual microtissues were more observable, suggesting that even the presence of a rapidly degrading hydrogel may delay the fusion and/or the remodeling of the individual microtissues. Dynamic culture conditions were found to modulate ECM synthesis following the OA hydrogel encapsulation. We also assessed the feasibility of 3D bioprinting of cartilage microtissues within OA based bioinks. It was observed that the microtissues remained viable after extrusion-based bioprinting and were able to fuse after 48 h, particularly when high microtissue densities were used, ultimately generating a cartilage tissue that was rich in GAGs and negative for calcium deposits. Therefore, this work supports the use of OA as a supporting hydrogel/bioink when using microtissues as biological building blocks in diverse biofabrication and 3D bioprinting platforms.

Scaffolds play a crucial role in bone tissue engineering to support the defect area through bone regeneration and defect reconstruction. Promising tissue regeneration without negative repercussions and avoidance of the lifelong presence inside the body make bioresorbable metals prosper in the field of regenerative medicine. Recently, Zn and its alloys have emerged as promising biodegradable materials for their moderate degradation rate and satisfactory biocompatibility. Nevertheless, it is very challenging for cells to adhere and grow over the Zn surface alone, which influences the tissue-implant integration. In this study, an attempt has been made to systematically investigate the bioactivity responses in terms of in vitro hemocompatibility, cytotoxicity, antibacterial activity, and in vivo biocompatibility of newly developed Zn-2Cu-0.5Mn/Mg alloy scaffolds with different surface roughness. The rough surface of Zn-2Cu-0.5Mg shows the highest degradation rate of 0.16 mm/yr. The rough surface exhibits a prominent role in the adsorption of protein, further enhancing cell adhesion. Concentration-dependent alloy extract shows the highest cell proliferation for 12.5% of the extract with a maximum cell viability of 101% in Zn-2Cu-0.5Mn and 108% in Zn-2Cu-0.5Mg after 3 d. Acceptable hemolysis percentages (less than 5%) with promising anticoagulation properties are observed for all of the conditions. Enhanced antibacterial ( Staphylococcus aureus and Escherichia coli ) activity due to a significant effect of ions illustrates the maximum killing effect on the bacterial colony for the rough Zn-2Cu-0.5Mg alloy. In addition, it is observed that for rough Zn-2Cu-0.5Mn/Mg alloys, the inflammatory response is minimal after subcutaneous implantation, and neo-bone tissue forms in the defect areas of the rat femur with satisfactory biosafety response. The osseointegration property of the Zn-2Cu-0.5Mg alloy is comparable to that of the Zn-2Cu-0.5Mn alloy. Therefore, the rough surface of the Zn-2Cu-0.5Mg alloy has the potential to enhance biocompatibility and promote better osseointegration activity with host tissues for various biomedical applications.

We fabricated three-dimensional (3D)-printed polycaprolactone (PCL) and PCL/graphene oxide (GO) (PGO) scaffolds for bone tissue engineering. An anti-inflammatory and pro-osteogenesis drug dexamethasone (DEX) was adsorbed onto GO and a 3D-printed PGO/DEX (PGOD) scaffold successfully improved drug delivery with a sustained release of DEX from the scaffold up to 1 month. The physicochemical properties of the PCL, PGO, and PGOD scaffolds were characterized by various analytical techniques. The biological response of these scaffolds was studied for adherence, proliferation, and osteogenic differentiation of seeded rabbit adipose-derived stem cells (ASCs) from DNA assays, alkaline phosphatase (ALP) production, calcium quantification, osteogenic gene expression, and immunofluorescence staining of osteogenic marker proteins. The PGOD scaffold was demonstrated to be the best scaffold for maintaining cell viability, cell proliferation, and osteogenic differentiation of ASCs in vitro. In vivo biocompatibility of PGOD was confirmed from subcutaneous implantation in nude mice where ASC-seeded PGOD can form ectopic bones, demonstrated by microcomputed tomography (micro-CT) analysis and immunofluorescence staining. Furthermore, implantation of PGOD/ASCs constructs into critical-sized cranial bone defects in rabbits form tissue-engineered bones at the defect site, observed using micro-CT and histological analysis.

Bioassembly encompasses various techniques such as bioprinting, microfluidics, organoids, and self-assembly, enabling advances in tissue engineering and regenerative medicine. Advancements in bioassembly technologies have enabled the precise arrangement and integration of various cell types to more closely mimic the complexity functionality of the neurovascular unit (NVU) and that of other biodiverse multicellular tissue structures. In this context, bioprinting offers the ability to deposit cells in a spatially controlled manner, facilitating the construction of interconnected networks. Scaffold-based assembly strategies provide structural support and guidance cues for cell growth, enabling the formation of complex bio-constructs. Self-assembly approaches utilize the inherent properties of cells to drive the spontaneous organization and interaction of neuronal and vascular components. However, recreating the intricate microarchitecture and functional characteristics of a tissue/organ poses additional challenges. Advancements in bioassembly techniques and materials hold great promise for addressing these challenges. The further refinement of bioprinting technologies, such as improved resolution and the incorporation of multiple cell types, can enhance the accuracy and complexity of the biological constructs; however, developing bioinks that support the growth of cells, viability, and functionality while maintaining compatibility with the bioassembly process remains an unmet need in the field, and further advancements in the design of bioactive and biodegradable scaffolds will aid in controlling cell adhesion, differentiation, and vascularization within the engineered tissue. Additionally, integrating advanced imaging and analytical techniques can provide real-time monitoring and characterization of bioassembly, aiding in quality control and optimization. While challenges remain, ongoing research and technological advancements propel the field forward, paving the way for transformative developments in neurovascular research and tissue engineering. This work provides an overview of the advancements, challenges, and future perspectives in bioassembly for fabricating neurovascular constructs with an add-on focus on bioprinting technologies.

Acellular nerve grafts (ANGs) are a promising therapeutic for patients with nerve defects caused by injuries. Conventional decellularization methods utilize a variety of detergents and enzymes. However, these methods have disadvantages, such as long processing times and the presence of detergents that remain on the graft. In this study, we aimed to reduce process time and minimize the risks associated with residual detergents by replacing them with supercritical carbon dioxide (scCO 2 ) and compared the effectiveness to Hudson's decellularization method, which uses several detergents. The dsDNA and the expression of MHC1 and 2 were significantly reduced in both decellularized groups, which confirmed the effective removal of cellular debris. The extracellular matrix proteins and various factors were found to be better preserved in the scCO 2 ANGs compared to the detergent-ANGs. We conducted behavioral tests and histological analyses to assess the impact of scCO 2 ANGs on peripheral nerve regeneration in animal models. Compared with Hudson's method, the scCO 2 method effectively improved the efficacy of peripheral nerve regeneration. Therefore, the decellularization method using scCO 2 is not only beneficial for ANG synthesis, but it may also be helpful for therapeutics by enhancing the efficacy of peripheral nerve regeneration., (© 2024. The Author(s).)

In this work, we report the design and fabrication of a light-addressable, paper-based nanocomposite scaffold for optical pacing and read-out of in vitro grown cardiac tissue. The scaffold consists of paper cellulose microfibers functionalized with gold nanorods (GNRs) and semiconductor quantum dots (QDs), embedded in a cell-permissive collagen matrix. The GNRs enable cardiomyocyte activity modulation through local temperature gradients induced by modulated near-infrared (NIR) laser illumination, with the local temperature changes reported by temperature-dependent QD photoluminescence (PL). The micrometer-sized paper fibers promote the tubular organization of HL-1 cardiac muscle cells, while the NIR plasmonic stimulation modulates reversibly their activity. Given the nanoscale spatial resolution and facile fabrication, paper-based nanocomposite scaffolds with NIR modulation offer excellent alternatives to electrode-based or optogenetic methods for cell activity modulation, at the single cell level, and are compatible with 3D tissue constructs. Such paper-based optical platforms can provide new possibilities for the development of in vitro drug screening assays and heart disease modeling.

Background: Craniofacial bone regeneration represents a dynamic area within tissue engineering and regenerative medicine. Central to this field, is the continual exploration of new methodologies for template fabrication, leveraging established bio ceramic materials, with the objective of restoring bone integrity and facilitating successful implant placements., Methods: Photopolymerized templates were prepared using three distinct bio ceramic materials, specifically a wet chemically synthesized bioactive glass and two commercially sourced hydroxyapatite variants. These templates underwent comprehensive characterization to assess their physicochemical and mechanical attributes, employing techniques including Fourier transform infrared spectroscopy, scanning electron microscopy, and nano-computed tomography. Evaluation of their biocompatibility was conducted through interaction with primary human osteoblasts (hOB) and subsequent examination using scanning electron microscopy., Results: The results demonstrated that composite showed intramolecular hydrogen bonding interactions with the photopolymer, while computerized tomography unveiled the porous morphology and distribution within the templates. A relatively higher porosity percentage (31.55 ± 8.70%) and compressive strength (1.53 ± 0.11 MPa) was noted for bioactive glass templates. Human osteoblast cultured on bioactive glass showed higher viability compared to other specimens. Scanning micrographs of human osteoblast on templated showed cellular adhesion and the presence of filopodia and lamellipodia., Conclusion: In summary these templates have the potential to be used for alveolar bone regeneration in critical size defect. Photopolymerization of bioceramics may be an interesting technique for scaffolds fabrication for bone tissue engineering application but needs more optimization to overcome existing issues like the ideal ratio of the photopolymer to bioceramics., (© 2024. The Author(s).)

The balance of macrophages in immune reactions is crucial for tissue repair. Despite some studies on responsive surfaces for immunomodulation regulation of macrophage phenotypes via external stimuli, 2D and manual interventions are limited. Herein, to address these limitations, we developed an inflammation environment-responsive macrophage-laden hydrogel-filled scaffold for investigating the impact of matrix confinement on macrophage phenotypes adaptively. We fabricated gelatin scaffolds with a controllable pore size and found that macrophages within smaller pores tended to have an anti-inflammation phenotype. We prepared poly(vinyl alcohol) (PVA)-based hydrogels crosslinked with phenylboronic acid (PBA)-based linkers. The hydrogels possessed shear-thinning, cell-loading, and ROS-sensitive-degradation abilities. Subsequently, a macrophage-laden hydrogel-filled scaffold was fabricated by filling the hydrogels into the porous scaffold under vacuum. With the degradation of the hydrogels under the overexpression of ROS in an inflammation environment, the macrophages were transferred from a state with strong matrix confinement to that with a weaker one. Meanwhile, with the change in matrix confinement, the macrophages upregulated the expressions of Arg-1 and IL-10 and downregulated the expressions of IL-1β, TNF-α, and IL-6, indicating polarization toward the anti-inflammatory phenotype. The inflammation environment-adaptive modulation of macrophage phenotypes in 3D provides a smart and biomimetic strategy for immunomodulation and regenerative medicine.

Despite extensive research, current methods for creating three-dimensional (3D) silk fibroin (SF) scaffolds lack control over molecular rearrangement, particularly in the formation of β-sheet nanocrystals that severely embrittle SF, as well as hierarchical fiber organization at both micro- and macroscale. Here, we introduce a fabrication process based on electrowriting of aqueous SF solutions followed by post-processing using an aqueous solution of sodium dihydrogen phosphate (NaH 2 PO 4 ). This approach enables gelation of SF chains via controlled β-sheet formation and partial conservation of compliant random coil structures. Moreover, this process allows for precise architecture control in microfiber scaffolds, enabling the creation of 3D flat and tubular macro-geometries with square-based and crosshatch microarchitectures, featuring inter-fiber distances of 400 μm and ∼97% open porosity. Remarkably, the crosslinked printed structures demonstrated a balanced coexistence of β-sheet and random coil conformations, which is uncommon for organic solvent-based crosslinking methods. This synergy of printing and post-processing yielded stable scaffolds with high compliance (modulus = 0.5-15 MPa) and the ability to support elastic cyclic loading up to 20% deformation. Furthermore, the printed constructs supported in vitro adherence and growth of human renal epithelial and endothelial cells with viability above 95%. These cells formed homogeneous monolayers that aligned with the fiber direction and deposited type-IV collagen as a specific marker of healthy extracellular matrix, indicating that both cell types attach, proliferate, and organize their own microenvironment within the SF scaffolds. These findings represent a significant development in fabricating organized stable SF scaffolds with unique microfiber structures and mechanical and biological properties that make them highly promising for tissue engineering applications.

Treatment of Limbal Stem Cell Deficiency (LSCD), based on autologous transplantation of the patient's stem cells, is one of the few medical stem cell therapies approved by the European Medicines Agency (EMA). It relies on isolating and culturing in vivo Limbal Epithelial Stem Cells (LESC) and then populating them on the fibrin substrate, creating a scaffold for corneal epithelial regeneration. Such a solution is then implanted into the patient's eye. The epithelial cell culture process is specific, and its results strongly depend on the initial cell seeding density. Achieving control of the density and repeatability of the process is a desirable aim and can contribute to the success of the therapy. The study aimed to test bioprinting as a potential technique to increase the control over LESCs seeding on a scaffold and improve process reproducibility. Cells were applied to 0.5 mm thick, flat, transparent fibrin substrates using extrusion bioprinting; the control was the traditional manual application of cells using a pipette. The use of 3D printer enabled uniform coverage of the scaffold surface, and LESCs density in printed lines was close to the targeted value. Moreover, printed cells had higher cell viability than those seeded traditionally (91.1 ± 8.2% vs 82.6 ± 12.8%). The growth rate of the epithelium was higher in bioprinted samples. In both methods, the epithelium had favorable phenotypic features (p63 + and CK14 +). 3D printing constitutes a promising approach in LSCD therapy. It provides favorable conditions for LESCs growth and process reproducibility. Its application may lead to reduced cell requirements, thereby to using fewer cells on lower passages, which will contribute to preserving LESCs proliferative potential., (© 2024. The Author(s).)

Prolonged osteochondral tissue engineering damage can result in osteoarthritis and decreased quality of life. Multiphasic scaffolds, where different layers model different microenvironments, are a promising treatment approach, yet stable joining between layers during fabrication remains challenging. To overcome this problem, in this study, a bilayer scaffold for osteochondral tissue regeneration was fabricated using 3D printing technology which containing a layer of PCL/hydroxyapatite (HA) nanoparticles and another layer of PCL/gelatin with various concentrations of fibrin (10, 20 and 30 wt.%). These printed scaffolds were evaluated with SEM (Scanning Electron Microscopy), FTIR (Fourier Transform Infrared Spectroscopy) and mechanical properties. The results showed that the porous scaffolds fabricated with pore size of 210-255 µm. Following, the ductility increased with the further addition of fibrin in bilayer composites which showed these composites scaffolds are suitable for the cartilage part of osteochondral. Also, the contact angle results demonstrated the incorporation of fibrin in bilayer scaffolds based on PCL matrix, can lead to a decrease in contact angle and result in the improvement of hydrophilicity that confirmed by increasing the degradation rate of scaffolds containing further fibrin percentage. The bioactivity study of bilayer scaffolds indicated that both fibrin and hydroxyapatite can significantly improve the cell attachment on fabricated scaffolds. The MTT assay, DAPI and Alizarin red tests of bilayer composite scaffolds showed that samples containing 30% fibrin have the more biocompatibility than that of samples with 10 and 20% fibrin which indicated the potential of this bilayer scaffold for osteochondral tissue regeneration., (© 2024. The Author(s).)

Hydrogels, composed of hydrophilic polymer networks, have emerged as versatile materials in biomedical applications due to their high water content, biocompatibility, and tunable properties. They mimic natural tissue environments, enhancing cell viability and function. Hydrogels' tunable physical properties allow for tailored antibacterial biomaterial, wound dressings, cancer treatment, and tissue engineering scaffolds. Their ability to respond to physiological stimuli enables the controlled release of therapeutics, while their porous structure supports nutrient diffusion and waste removal, fostering tissue regeneration and repair. In wound healing, hydrogels provide a moist environment, promote cell migration, and deliver bioactive agents and antibiotics, enhancing the healing process. For cancer therapy, they offer localized drug delivery systems that target tumors, minimizing systemic toxicity and improving therapeutic efficacy. Ocular therapy benefits from hydrogels' capacity to form contact lenses and drug delivery systems that maintain prolonged contact with the eye surface, improving treatment outcomes for various eye diseases. In mucosal delivery, hydrogels facilitate the administration of therapeutics across mucosal barriers, ensuring sustained release and the improved bioavailability of drugs. Tissue regeneration sees hydrogels as scaffolds that mimic the extracellular matrix, supporting cell growth and differentiation for repairing damaged tissues. Similarly, in bone regeneration, hydrogels loaded with growth factors and stem cells promote osteogenesis and accelerate bone healing. This article highlights some of the recent advances in the use of hydrogels for various biomedical applications, driven by their ability to be engineered for specific therapeutic needs and their interactive properties with biological tissues.

Gelatin methacryloyl (GelMA), typically derived from mammalian sources, has recently emerged as an ideal bio-ink for three-dimensional (3D) bioprinting. Herein, we developed a fish skin-based GelMA bio-ink for the fabrication of a 3D GelMA skin substitute with a 3D bioprinter. Several concentrations of methacrylic acid anhydride were used to fabricate GelMA, in which their physical-mechanical properties were assessed. This fish skin-based GelMA bio-ink was loaded with human adipose tissue-derived mesenchymal stromal cells (ASCs) and human platelet lysate (HPL) and then printed to obtain 3D ASCs + HPL-loaded GelMA scaffolds. Cell viability test and a preliminary investigation of its effectiveness in promoting wound closure were evaluated in a critical-sized full thickness skin defect in a rat model. The cell viability results showed that the number of ASCs increased significantly within the 3D GelMA hydrogel scaffold, indicating its biocompatibility property. In vivo results demonstrated that ASCs + HPL-loaded GelMA scaffolds could delay wound contraction, markedly enhanced collagen deposition, and promoted the formation of new blood vessels, especially at the wound edge, compared to the untreated group. Therefore, this newly fish skin-based GelMA bio-ink developed in this study has the potential to be utilized for the printing of 3D GelMA skin substitutes., (© 2024. The Author(s).)

Liver disease cases are rapidly expanding worldwide, and transplantation remains the only effective cure for end-stage disease. There is an increasing demand for developing potential drug treatments, and regenerative therapies using in-vitro culture platforms. Human decellularized extracellular matrix (dECM) is an appealing alternative to conventional animal tissues as it contains human-specific proteins and can serve as scaffolding materials. Herein we exploit this with human donor tissue from discarded liver which was not suitable for transplant using a synergistic approach to combining biological and topographical cues in electrospun materials as an in-vitro culture platform. To realise this, we developed a methodology for incorporating human liver dECM into electrospun polycaprolactone (PCL) fibres with surface nanotopographies (230-580 nm). The hybrid scaffolds were fabricated using varying concentrations of dECM; their morphology, mechanical properties, hydrophilicity and stability were analysed. The scaffolds were validated using HepG2 and primary mouse hepatocytes, with subsequent results indicating that the modified scaffolds-maintained cell growth and influenced cell attachment, proliferation and hepatic-related gene expression. This work demonstrates a novel approach to harvesting the potential from decellularized human tissues in the form of innovative in-vitro culture platforms for liver., (© 2024. The Author(s).)

Background: In bone tissue engineering (BTE), cell-laden scaffolds offer a promising strategy for repairing bone defects, particularly when host cell regeneration is insufficient due to age or disease. Exogenous stem cell-based BTE requires bioactive factors to activate these cells. Graphene oxide quantum dots (GOQDs), zero-dimensional derivatives of graphene oxide, have emerged as potential osteogenic nanomedicines. However, constructing biological scaffolds with GOQDs and elucidating their biological mechanisms remain critical challenges., Methods: We utilized GOQDs with a particle size of 10 nm, characterized by a surface rich in C-O-H and C-O-C functional groups. We developed a gelatin methacryloyl (GelMA) hydrogel incorporated with GOQDs-treated dental pulp stem cells (DPSCs). These constructs were transplanted into rat calvarial bone defects to estimate the effectiveness of GOQDs-induced DPSCs in repairing bone defects while also investigating the molecular mechanism underlying GOQDs-induced osteogenesis in DPSCs., Results: GOQDs at 5 μg/mL significantly enhanced the osteogenic differentiation of DPSCs without toxicity. The GOQDs-induced DPSCs showed active osteogenic potential in three-dimensional cell culture system. In vivo, transplantation of GOQDs-preactivated DPSCs/GelMA composite effectively facilitated calvarial bone regeneration. Mechanistically, GOQDs stimulated mitophagy flux through the phosphatase-and-tensin homolog-induced putative kinase 1 (PINK1)/Parkin E3 ubiquitin ligase (PRKN) pathway. Notably, inhibiting mitophagy with cyclosporin A prevented the osteogenic activity of GOQDs., Conclusion: This research presents a well-designed bionic GOQDs/DPSCs/GelMA composite scaffold and demonstrated its ability to promote bone regeneration by enhancing mitophagy. These findings highlight the significant potential of this composite for application in BTE and underscore the crucial role of mitophagy in promoting the osteogenic differentiation of GOQDs-induced stem cells., Competing Interests: The authors report no conflicts of interest in this work., (© 2024 Yan et al.)

Adipose tissue engineering (ATE) has been gaining increasing interest over the past decades, offering promise for new and innovative breast reconstructive strategies. Animal-derived gelatin-methacryloyl (Gel-MA) has already been applied in a plethora of TE strategies. However, due to clinical concerns, related to the potential occurrence of immunoglobulin E-mediated immune responses and pathogen transmission, a shift towards defined, reproducible recombinant proteins has occurred. In the present study, a recombinant protein based on human collagen type I, enriched with arginine-glycine-aspartic acid was functionalized with photo-crosslinkable methacryloyl moieties (RCPhC1-MA), processed into 3D scaffolds and compared with frequently applied Gel-MA from animal origin using an indirect printing method applying poly-lactic acid as sacrificial mould. For both materials, similar gel fractions (>65%) and biodegradation times were obtained. In addition, a significantly lower mass swelling ratio (17.6 ± 1.5 versus 24.3 ± 1.4) and mechanical strength (Young's modulus: 1.1 ± 0.2 kPa versus 1.9 ± 0.3 kPa) were observed for RCPhC1-MA compared to Gel-MA scaffolds. In vitro seeding assays showed similar cell viabilities (>80%) and a higher initial cell attachment for the RCPhC1-MA scaffolds. Moreover, the seeded adipose-derived stem cells could be differentiated into the adipogenic lineage for both Gel-MA and RCPhC1-MA scaffolds, showing a trend towards superior differentiation for the RCPhC1-MA scaffolds based on the triglyceride and Bodipy assay. RCPhC1-MA scaffolds could result in a transition towards the exploitation of non-animal-derived biomaterials for ATE, omitting any regulatory concerns related to the use of animal derived products., (© 2024 IOP Publishing Ltd. All rights, including for text and data mining, AI training, and similar technologies, are reserved.)

A novel avenue for advancing our understanding of kidney disease mechanisms and developing targeted therapeutics lies in overcoming the limitations of the existing in vitro models. Traditional animal models, while useful, do not fully capture the intricacies of human kidney physiology and pathophysiology. Tissue engineering offers a promising solution, yet current models often fall short in replicating the complex microarchitecture and biochemical milieu of the kidney. To address this challenge, we propose the development of a sophisticated in vitro glomerular filtration barrier (GFB) utilizing advanced biomaterials and a kidney decellularized extracellular matrix (kdECM). In our approach, we employ a bacterial cellulose membrane (BC) as a scaffold, providing a robust framework for cell growth and interaction. Coating this scaffold with kdECM hydrogel derived from caprine kidney tissue via a detergent-free decellularization method ensures the preservation of vital extracellular matrix proteins crucial for cellular compatibility and signaling. Our engineered GFB not only supports the growth of endothelial and podocyte cells but also exhibits the presence of key markers such as CD31 and nephrin, indicating successful cellular integration. Furthermore, the expression of collagen IV, an essential extracellular matrix (ECM) protein, validates the fidelity of our model in simulating cellular interactions within a kdECM matrix. Additionally, we assessed the filtration efficiency of the developed GFB model using albumin, a standard protein, to evaluate its performance under conditions that closely mimic the native physiological environment. This innovative approach, which faithfully recapitulates the native microenvironment of the glomerulus, holds immense promise for elucidating kidney disease mechanisms, conducting permeability studies, and advancing personalized therapeutic strategies. By leveraging cutting-edge biomaterials and tissue-specific coculture technology, this study can be further extended to develop GFB for the treatment of renal diseases, ultimately improving patient outcomes and quality of life.

Partial or complete rupture of the tendon can damage the collagen structure, resulting in the disruption of the electrical signal pathway. It is a great challenge to reconstruct the original electrical signal pathway of the tendon and promote the regeneration and functional recovery of defective tendon. In this study, carbon fiber-mediated electrospinning scaffolds were fabricated by wrapping conductive, high-strength, loose single-bundle carbon fibers with nanofiber membranes. Due to the presence of nanofiber membranes, the maximum tensile force of the scaffolds was 2.4 times higher than that of carbon fibers, while providing excellent temporal and spatial prerequisites for tenocytes to adapt to electrical stimulation to accelerate proliferation and expression. The diameter of the carbon fiber monofilaments used in this study was 5.07 ± 1.20 μm, which matched the diameter of tendon collagen, allowing for quickly establishing the connection between the tendon tissue and the scaffold, and better promoting the recovery of the electrical signal pathway. In a rabbit Achilles tendon defect repair model, the carbon fiber-mediated electrospinning scaffold was almost filled with collagen fibers compared to a nonconductive polyethylene glycol terephthalate scaffold. Transcriptome sequencing revealed that fibromodulin and tenomodulin expression were upregulated, and their related proteoglycans and glycosaminoglycan binding proteins pathways were enhanced, which could regulate the TGF-β signaling pathway and optimize the extracellular matrix assembly, thus promoting tendon repair. Therefore, the scaffold in this study makes up for the shortage of conductive scaffolds for repairing tendon defects, revealing the potential impact of conductivity on the signaling pathway of tendon repair and providing a new approach for future clinical studies.

Sea urchin spines are biogenic single crystals of magnesium calcite that are stiff, strong, damage tolerant and light and have a bicontinuous porous structure. Here, we showed that the removal of their intraskeletal organic matrix materials did not affect the compressive mechanical properties and generated an open porosity. This matrix was able to adsorb and release oxytetracycline, a broad-spectrum antibiotic. The drug-loaded sea urchin matrix induced bacterial cell death after 4 and 8 hours of incubation of both Gram-negative E. coli and Gram-positive S. aureus strains and this process induces an inhibition of bacterial cell adhesion. In conclusion, this study shows that thermally treated sea urchin spines are a compressive resistant and lightweight matrix able to load drugs and with potential use in spine fusion, a challenging application that requires withstanding high compressive loading.

In this paper, the mechanical behaviour and failure of porous additively manufactured (AM) polylactide (PLA) scaffolds based on the triply periodic minimal surfaces (TPMS) is investigated using numerical calculations of their unit cells and representative volumes. The strain-amplification factor is chosen as the main parameter, and the most critical locations for failure of different types of scaffold structures are evaluated. The results obtained are presented in comparison with a multiple-crack-growth algorithm using the extended finite element method (XFEM), underpinned by the experimentally obtained fracture properties of PLA. The effect of morphology of TPMS structures on the pre-critical, critical and post-critical behaviours of scaffolds under monotonic loading regimes is assessed. The results provide an understanding of the fracture behaviour and main risk points for crack initiation in structures of AM-PLA scaffolds based on typical commonly used types of TPMS, as well as the influence of structure type and external load on this behaviour., 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 IPEM. Published by Elsevier Ltd. All rights reserved.)

Creating acellular vascularized constructs from animal and plant tissue is one of the well-known strategies for scaffold assembly. Decellularization takes an important position among these strategies. The most common method is chemical decellularization. This approach employs high concentrations of detergents, primarily Triton X-100, sodium dodecyl sulfate (SDS), and sodium hypochlorite (SH). In this work, novel techniques for decellularizing spinach were developed using detergents frequently utilized in laboratories. Spinach leaves were decellularized using Tween-20, SDS, and SH at low concentrations to generate an acellular plant matrix for tissue engineering. We measured the quantities of DNA and protein, as well as the decellularization using hematoxylin and eosin (H&E) staining. The biocompatibility and capacity of the biostructures to stimulate fibroblast wound healing were assessed using MTT and the Scratch assay. The antibacterial activity of the scaffolds was also tested against a gram-positive bacterium, Staphilococcus aureus, which is a common pathogen associated with wound healing. The best shape, evident vascularization, and good biocompatibility were seen in the Tween-20 decellularized samples at 1% concentration at 21°C and 37°C through the enhancement of cell proliferation and wound healing. In terms of antibacterial activity, all scaffold samples had a significant effect on Staphilococcus aureus, where the number of bacterial colonies in all six scaffold groups became zero after 4 h of treatment. The scaffolds also showed a 100% kill rate on Staphilococcus aureus, which could avoid wound infection during the repair process, and that can be suggested as a scaffold for tissue engineering applications and an important constituent for pharmacological activities., (© 2024 The Author(s). Journal of Biomedical Materials Research Part B: Applied Biomaterials published by Wiley Periodicals LLC.)

Bacterial cellulose is a unique biomaterial produced by various species of bacteria that offers a range of potential applications in the biomedical field. To provide a cost-effective alternative to soft-tissue implants used in cavity infills, remodeling, and subdermal wound healing, in vitro cytotoxicity and in vivo biocompatibility of native bacterial cellulose were investigated. Cytotoxicity was assessed using a metabolic assay on Swiss 3T3 fibroblasts and INS-1832/13 rat insulinoma. Results showed no cytotoxicity, whether the cells were seeded over or under the bacterial cellulose scaffolds. Biocompatibility was performed on Sprague-Dawley rats (males and females, 8 weeks old) by implanting bacterial cellulose membranes subcutaneously for 1 or 12 weeks. The explanted scaffolds were then sliced and stained with hematoxylin and eosin for histological characterization. The first series of results revealed acute and chronic inflammation persisting over 12 weeks. Examination of the explants indicated a high number of granulocytes within the periphery of the bacterial cellulose, suggesting the presence of endotoxins within the membrane, confirmed by a Limulus amebocyte lysate test. This discovery motivated the development of non-pyrogenic bacterial cellulose scaffolds. Following this, a second series of animal experiments was done, in which materials were implanted for 1 or 2 weeks. The results revealed mild inflammation 1 week after implantation, which then diminished to minimal inflammation after 2 weeks. Altogether, this study highlights that unmodified, purified native bacterial cellulose membranes may be used as a cost-effective biomedical device provided that proper endotoxin clearance is achieved., (© 2024 The Author(s). Journal of Biomedical Materials Research Part B: Applied Biomaterials published by Wiley Periodicals LLC.)

Stem cell derived small extracellular vesicles (sEVs) have emerged as promising nanomaterials for the repair of bone defects. However, low retention of sEVs affects their therapeutic effects. Clinically used natural substitute inorganic bovine bone mineral (Bio-Oss) bone powder lacks high compactibility and efficient osteo-inductivity that limit its clinical application in repairing large bone defects. In this study, a poly ethylene glycol/hyaluronic acid (PEG/HA) hydrogel was used to stabilize Bio-Oss and incorporate rat bone marrow stem cell-derived sEVs (rBMSCs-sEVs) to engineer a PEG/HA-Bio-Oss (PEG/HA-Bio) composite scaffold. Encapsulation and sustained release of sEVs in hydrogel scaffold can enhance the retention of sEVs in targeted area, achieving long-lasting repair effect. Meanwhile, synergistic administration of sEVs and Bio-Oss in cranial defect can improve therapeutic effects. The PEG/HA-Bio composite scaffold showed good mechanical properties and biocompatibility, supporting the growth of rBMSCs. Furthermore, sEVs enhanced in vitro cell proliferation and osteogenic differentiation of rBMSCs. Implantation of sEVs/PEG/HA-Bio in rat cranial defect model promoted in vivo bone regeneration, suggesting the great potential of sEVs/PEG/HA-Bio composite scaffold for bone repair and regeneration. Overall, this work provides a strategy of combining hydrogel composite scaffold systems and stem cell-derived sEVs for the application of tissue engineering repair., (© 2024 IOP Publishing Ltd. All rights, including for text and data mining, AI training, and similar technologies, are reserved.)

Hydrogels have emerged as potential materials for bone grafting, thanks to their biocompatibility, biodegradation, and flexibility in filling irregular bone defects. In this study, we fabricated a novel NAH hydrogel system, composed of N,O-carboxymethyl chitosan (NOCC), aldehyde hyaluronic acid (AHA), and hydroxyapatite (HAp). To improve the mechanical strength of the fabricated hydrogel, a porous polycaprolactone (PCL) matrix was synthesized and used as a three-dimensional (3D) support template for NAH hydrogel loading, forming a novel PCL/NAH hybrid scaffold. A mixture of monosodium glutamate (M) and sucrose (S) at varied weight ratios (5M:5S, 7M:3S, and 9M:1S) was used for the fabrication of 3D PCL matrices. The morphology, interconnectivity, and water resistance of the porous PCL scaffolds were investigated for optimal hydrogel loading efficiency. The results demonstrated that PCL scaffolds with porogen ratios of 7M:3S and 9M:1S possessed better interconnectivity than 5M:5S ratio. The compressive strength of the PCL/NAH hybrid scaffolds with 9M:1S (561.6 ± 6.1 kPa) and 7M:3S (623.8 ± 6.8 kPa) ratios are similar to cancellous bone and all hybrid scaffolds were biocompatible. Rabbit models with tibial defects were implanted with the PCL/NAH scaffolds to assess the wound healing capability. The results suggest that the PCL/NAH hybrid scaffolds, specifically those with porogen ratio of 7M:3S, exhibit promising bone healing effects., (© 2024 Wiley Periodicals LLC.)

The complete structure-functional repair of volumetric muscle loss (VML) remains a giant challenge and biomedical hydrogels to remodel microenvironment and enhance neurogenesis have appeared to be a promising direction. However, the current hydrogels for VML repair hardly achieve these two goals simultaneously due to their insufficient functionality and the challenge in high-cost of bioactive factors. In this study, a facile strategy using Nb 2 C MXene-functionalized hydrogel (OPTN) as a bioactive scaffold is proposed to promote VML repair with skeletal muscle regeneration and functional restoration. In vitro experiments show that OPTN scaffold can effectively scavenge reactive oxygen species (ROS), guide macrophages polarization toward M2 phenotype, and resist bacterial infection, providing a favorable microenvironment for myoblasts proliferation as well as the endothelial cells proliferation, migration, and tube formation. More importantly, OPTN scaffold with electroactive feature remarkably boosts myoblasts differentiation and mesenchymal stem cells neural differentiation. Animal experiments further confirm that OPTN scaffold can achieve a prominent structure-functional VML repair by attenuating ROS levels, alleviating inflammation, reducing fibrosis, and facilitating angiogenesis, newborn myotube formation, and neurogenesis. Collectively, this study provides a highly promising and effective strategy for the structure-functional VML repair through designing bioactive multifunctional hydrogel with microenvironment remodeling and enhanced neurogenesis., (© 2024 Wiley‐VCH GmbH.)

Mesenchymal stem cell-derived exosomes(MSCs-Exos) offer promising therapeutic potential for a wide range of tissues and organs such as bone/cartilage, nerves, skin, fat, and endocrine organs. In comparison to the application of mesenchymal stem cells (MSCs), MSCs-Exos address critical challenges related to rejection reactions and ethical concerns, positioning themselves as a promising cell-free therapy. As exosomes are extracellular vesicles, their effective delivery necessitates the use of carriers. Consequently, the selection of hydrogel materials as scaffolds for exosome delivery has become a focal point of contemporary research. The diversity of hydrogel scaffolds, which can take various forms such as injectable types, dressings, microneedles, and capsules, leads to differing choices among researchers for treating diseases within the same domain. This variability in hydrogel materials poses challenges for the translation of findings into clinical practice. The review highlights the potential of hydrogel-loaded exosomes in different fields and introduces the advantages and disadvantages of different forms of hydrogel applications. It aims to provide a multifunctional and highly recognized hydrogel scaffold option for tissue regeneration at specific sites, improve clinical translation efficiency, and benefit the majority of patients., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

In this study, a novel bionic periosteum (BP)-bioactive glass fiber membrane (BGFM) is designed. The introduction of magnesium ion (Mg 2+ ) and zinc ion (Zn 2+ ) change the phase separation during the electrospinning (ES) jet stretching process. The fiber's pore structure transitions from connected to closed pores, resulting in a decrease in the rapid release of metal ions while also improving degradation via reducing filling quality. Additionally, the introduction of magnesium (Mg) and zinc (Zn) lead to the formation of negative charged tetrahedral units (MgO 4 2- and ZnO 4 2- ) in the glass network. These units effectively trap positive charged metal ions, further inhibiting ion release. In vitro experiments reveal that the deigned bionic periosteum regulates the polarization of macrophages toward M2 type, thereby establishing a conducive immune environment for osteogenic differentiation. Bioinformatics analysis indicate that BP enhanced bone repair via the JAK-STAT signaling pathway. The slow release of metal ions from the bionic periosteum can directly enhance osteogenic differentiation and vascularization, thereby accelerating bone regeneration. Finally, the bionic periosteum exhibits remarkable capabilities in angiogenesis and osteogenesis, demonstrating its potential for bone repair in a rat calvarial defect model., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

Tissue engineering presents a promising approach for the treatment of meniscal injuries, yet the development of meniscal scaffolds that exhibit both superior biomechanical properties and biocompatibility remains a considerable challenge. In this study, decellularized skin matrix (DSM) scaffolds were first prepared using porcine skin through decellularization and freeze-drying techniques. The DSM scaffold has favorable porosity, hydrophilicity, and biocompatibility. Importantly, the collagen content and tensile modulus of the scaffold are comparable to those of native meniscus (44.13 ± 2.396 mg/g vs. 42.41 ± 2.40 mg/g and 103.30 ± 2.98 MPa vs. 128.80 ± 9.115 MPa). Subsequently, the peptide PFSSTKT (PFS) with mesenchymal stem cells (MSCs) recruitment capability was used to modify DSM to construct DSM-PFS scaffolds. Compared to the DSM scaffold, the optimized DSM-PFS scaffold enhanced in vitro collagen and glycosaminoglycan (GAG) production and upregulated the expression of cartilage-specific genes. Furthermore, the DSM-PFS scaffold was more effective in recruiting MSCs in vitro. In vivo studies in rabbit models showed that the DSM-PFS scaffold successfully promoted meniscus tissue regeneration. Three months post-implantation, meniscus tissue formation can be observable, and after six months, the neo-meniscus exhibited tissue structure and tensile properties similar to the native meniscus. Notably, the DSM-PFS scaffold exhibited significant chondroprotective effects, slowing osteoarthritis (OA) progression. In conclusion, the DSM-PFS scaffold may represent a promising candidate for future applications in meniscus tissue engineering. STATEMENT OF SIGNIFICANCE: We developed a decellularized skin matrix (DSM) meniscus scaffold using whole-layer porcine skin, demonstrating superior biomechanical strength and biocompatibility. Following modification with the stem cell-recruiting peptide PFS, the optimized DSM-PFS scaffold outperformed the DSM scaffold in cell attraction, collagen and glycosaminoglycan production, and cartilage-specific gene expression. Implanted into rabbit knee joints, the cell-free DSM-PFS scaffold induced meniscal tissue formation within three months, achieving the histological structure and tensile strength of the native meniscus by six months. Moreover, it significantly protected the cartilage. Our findings provide new insights into the fabrication of scaffolds for meniscal tissue engineering, with the DSM-PFS scaffold emerging as an ideal candidate for future applications., 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 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Electrospun nanofibrous scaffolds are renowned for their ability to mimic the microstructure of the extracellular matrix (ECM). However, they often fail to replicate the geometry of target tissues, and the biocompatibility of these scaffolds those made from synthetic polymers is always limited due to the lack of cell binding sites. To address these issues, we proposed an innovative approach that combined unidirectional freeze-drying and electrospinning. During this process, electrospun polycaprolactone (PCL) nanofibers were chopped into nanofibrils, which range in size up to several hundred micrometers, and were incorporated into the chitosan scaffolds via unidirectional freeze-drying. In these scaffolds, the chitosan phase was responsible for maintaining the structural integrity at the macroscale, while the embedded nanofibers enhanced the surface topography at the microscale. The resulting scaffolds exhibited a high porosity of 90% and an impressive water uptake capacity of 2500%. Furthermore, 3T3 fibroblast cells showed strong interactions with the scaffolds, characterized by high rates of cell proliferation and viability. The cells also displayed significant orientation along the direction of the pores, suggesting that the scaffolds effectively guided cellular growth., 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 Ltd. All rights reserved.)

Electrical stimulation (ES) of cellular systems can be utilized for biotechnological applications and electroceuticals (bioelectric medicine). Neural cell stimulation especially has a long history in neuroscience research and is increasingly applied for clinical therapies. Application of ES via conventional electrodes requires external connectors and power sources, hindering scientific and therapeutic applications. Here engineering novel 3D scaffold-free human neural stem cell constructs with integrated piezoelectric nanoparticles for enhanced neural tissue induction and function is described. Tetragonal barium titanate (BaTi03) nanoparticles are employed as piezoelectric stimulators prepared as cytocompatible dispersions, incorporated into 3D self-organizing neural spheroids, and activated wirelessly by ultrasound. Ultrasound delivery (low frequency; 40 kHz) is optimized for cell survival, and nanoparticle activation enabled ES throughout the spheroids during differentiation, tissue formation, and maturation. The resultant human neural tissues represent the first example of direct tissue loading with piezoelectric particles for ensuing 3D ultrasound-mediated piezoelectric enhancement of human neuronal induction from stem cells, including augmented neuritogenesis and synaptogenesis. It is anticipated that the platform described will facilitate advanced tissue engineering and in vitro modeling of human neural (and potentially non-neural) tissues, with modeling including tissue development and pathology, and applicable to preclinical testing and prototyping of both electroceuticals and pharmaceuticals., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

Additive manufacturing can develop regenerative scaffolds for wound healing. 3D printing offers meticulous porosity, mechanical integrity, cell adhesion and cost-effectiveness. Herein, we prepared ink composed of carboxymethyl cellulose (CMC), polyvinylpyrrolidone (PVP), collagen, and oregano extract for the fabrication of tissue constructs. The blend was optimized to form a homogeneous ink and rheological characterization demonstrated shear thinning behavior. The scaffolds were printed using Direct Ink Write (DIW) at a flow speed of 4 mm 3 /s and a layer height of 0.18 mm. The fabricated scaffolds demonstrated an ultimate tensile strength (UTS) and toughness of 730 KPa and 2.72 MJ/m 3 , respectively. Scanning Electron Microscopy (SEM) revealed an average pore size of 300 ± 30 μm. Fourier transform infrared spectroscopy (FTIR) analysis confirmed that all materials were present. The contact angle of the composite scaffold was 68° ± 1°. Moreover, the scaffolds presented 82 % mass loss (degradation) in phosphate buffer saline (PBS) over 14 days. The composite scaffold exhibited inhibition zones of 9 mm and 12 mm against Staphylococcus aureus and Escherichia coli, respectively. The PVP/CMC/collagen/oregano 3D printed scaffolds exhibited excellent biocompatibility with the mesenchymal stem cells and humman dermal fibroblast cells, confirmed by water-soluble tetrazolium - 8 (WST-8) assay (test conducted for 7 days). The enhanced angiogenic potential of said scaffold was assesed by release of vascular endothelial growth factor followed by further validation through in-vivo CAM assay. Thus, confirming suitability for the potential wound healing application., 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.)

Salivary glands are the principal organs responsible for secreting saliva in the oral cavity. Tumors, trauma, inflammation, and other factors can cause functional or structural damage to the glands, leading to reduced saliva secretion. In this study, we innovatively prepared a acinar-mimetic silk fibroin-collagen-astragalus polysaccharide (SCA) scaffold using low-temperature three-dimensional (3D) printing and freeze-drying techniques. We evaluated the material properties and cell compatibility of the scaffold in vitro and implanted it into the damaged parotid glands (PG) of rats to assess its efficacy in tissue reconstruction and functional repair. The results demonstrated that the SCA scaffold featured a porous structure resembling natural acini, providing an environment conducive to cell growth and orderly aggregation. It exhibited excellent porosity, water absorption, mechanical properties, and biocompatibility, fulfilling the requirements for tissue engineering scaffolds. In vitro, the scaffold facilitated adhesion, proliferation, orderly polarization, and spherical aggregation of PG cells. In vivo, the SCA scaffold effectively recruited GECs locally, forming gland-like acinar structures that matured gradually, promoting the regeneration of damaged PGs. The SCA scaffold developed in this study supports tissue reconstruction and functional repair of damaged PGs, making it a promising implant material for salivary gland regeneration., Competing Interests: Declaration of competing interest I have nothing to declare., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Hydrogel/fiber composites have received wide attention as tissue engineering scaffolds due to the outstanding properties of fibers and hydrogels. In the current research, a hydrogel/fiber composite scaffold was made based on chitosan-modified polycaprolactone (PCL) microfibers and chitosan hydrogel as a binder. The presence of chitosan as a modifier on the surface of fibers and as a binder between fibers can create scaffolds with excellent structural and mechanical properties. To this end, the three-dimensional microfibers were first functionalized with amine groups. Then, the chitosan chains were attached to the fibers by an aldehyde coupling agent and Schiff base reaction. FTIR and Raman spectroscopies corroborated that chitosan was successfully immobilized on PCL fibers. Chitosan-modified fibers were molded with chitosan solutions of various concentrations and the prepared composite scaffolds were stabilized using ionic crosslinking. The obtained composites represented a porous 3D structure with highly interconnected pores. The compressive modulus increased by 19 and 2.7 folds and the tensile modulus was augmented by 28 and 4 folds, in respective dry and swollen states with increasing hydrogel concentration from 0.1 to 1 %. Hydrogel/fiber composites were able to preserve cell viability, and increasing the hydrogel proportion increased adhesion, proliferation and penetration of cells into the scaffold., 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.)

Infected bone defects are one of the most challenging problems in the treatment of bone defects due to the high antibiotic failure rate and the lack of ideal bone grafts. In this paper, inspired by clinical bone cement filling treatment, α-c phosphate (α-TCP) with self-curing properties is composited with β-tricalcium phosphate (β-TCP) and constructed a bionic cancellous bone scaffolding system α/β-tricalcium phosphate (α/β-TCP) by low-temperature 3D printing, and gelatin is preserved inside the scaffolds as an organic phase, and later loaded with a metal-polyphenol network structure of tea polyphenol-magnesium (TP-Mg) nanoparticles. The scaffolds mimic the structure and components of cancellous bone with high mechanical strength (>100 MPa) based on α-TCP self-curing properties through low-temperature 3D printing. Meanwhile, the scaffolds loaded with TP-Mg exhibit significant inhibition of Staphylococcus aureus (S.aureus) and promote the transition of macrophages from M1 pro-inflammatory to M2 anti-inflammatory phenotype. In addition, the composite scaffold also exhibits excellent bone-enhancing effects based on the synergistic effect of Mg 2+ and Ca 2+ . In this study, a multifunctional ceramic scaffold (α/β-TCP@TP-Mg) that integrates anti-inflammatory, antibacterial, and osteoinduction is constructed, which promotes late bone regenerative healing while modulating the early microenvironment of infected bone defects, has a promising application in the treatment of infected bone defects., (© 2024 The Author(s). Small published by Wiley‐VCH GmbH.)