Tissue Regeneration Applications Research Articles

Injectable hydrogel combining alginate and Rhodomyrtus tomentosa medicine with antibacterial, anti-inflammatory and cellular proliferation properties as potential wound dressing material

BackgroundAlginate hydrogels have been widely used as wound healing materials because of their high water content, biocompatibility, and easy fabrication. However, it is still challenging to improve their bioactivities due to the inherently bioinert properties of alginate. Herein, we report a simple and effective method to fabricate a multifunctional alginate hydrogel (Alg@Rt) with anti-inflammatory and antibacterial activities, by combining the advantages of alginate and Rhodomyrtus tomentosa leaves extract (Rt). MethodsThe hydrogel is prepared by simply mixing the solutions of alginate and Rt, using CaO2 as the crosslinker. The porous structure of hydrogel is evidenced by the scanning electron microscope. The swelling of the hydrogel was determined by comparing the difference in mass of samples immersed in physiological pH. The polyphenol release was determined by using the Folin-Ciocalteu method. The in vitro anti-inflammatory and antibacterial activities of the Alg@Rt hydrogel were evaluated and compared with the pure Alg hydrogel. The biocompatibility of hydrogels was evaluated towards fibroblasts. Significant findingA new injectable Alg@Rt hydrogel was developed to facilitate the wound healing process. Compared to the pure Alg hydrogel, Alg@Rt hydrogel significantly enhanced the anti-inflammatory, antibacterial and cell proliferative activities, which is beneficial in wound treatment and tissue regenerative applications.

Development of a new biomaterial based on cashew tree gum (Anarcadium occidentale L.) enriched with hydroxyapatite and evaluation of cytotoxicity in adipose-derived stem cell cultures

Cashew tree gum is a polysaccharide material highly available in the Northeast region of Brazil. It has been explored for biocompatibility with human tissues. This research aimed to describe the synthesis and characterization of cashew gum/hydroxyapatite scaffold and evaluate the possible cytotoxicity in murine adipose-derived stem cells (ADSCs) cultures. ADSCs of the subcutaneous fat tissue of Wistar rats were collected, isolated, expanded, differentiated into three strains, and characterized immunophenotypically. The scaffolds were synthesized through chemical precipitation, lyophilized and characterized through scanning electron microscopy (SEM), infrared spectroscopy (FTIR), X-ray diffraction (XRD), thermal analysis (TG and DTG), and mechanical testing. The scaffold presented a crystalline structure and pores with an average diameter of 94.45 ± 50.57 μm. By mechanical tests, the compressive force and modulus of elasticity were like the cancellous bone. The isolated adipose-derived stem cells (ADSCs) presented fibroblast morphology, adhesion capacity to plastic, differentiation in osteogenic, adipogenic and chondrogenic lineages, positive expression for the CD105 and CD90 markers and negative expression for the CD45 and CD14 markers. The MTT test showed increased cell viability, and the biomaterial showed a high level of hemocompatibility (<5 %). This study allowed the development of a new scaffold for future surgical applicability in tissue regeneration.

O103 Digital light processing 3D printed collagen/PEGDA hydrogel for regenerative medicine

Abstract Introduction Digital light processing represents a rapid approach to constructing 3D structures with superior resolution. However, it imposes demanding requirements on the properties of bioink. Methacrylated gelatin has long been the major option but results in limited mechanical properties. The development of collagen-based bioinks provides a wider stiffness tuning range, native bioactivities, and versatility in regenerative medicine applications. Methods Purified collagen solution was obtained via enzymolysis and ultrafiltration, then went through methacrylation. The photo-crosslinkable bioink comprises methacrylated collagen, PEGDA, acetic acid, yellow food dye, and photoinitiator LAP. The 3D structures were fabricated utilising a commercial digital light processing printer with 405 nm visible light. Results The collagen/PEGDA bioink showed translucent and low viscosity suitable for digital light processing printing. The printed scaffolds reached a compressive modulus of ∼100 kPa with 0.6 wt% collagen. Sharp-edged and fine structures (∼250 μm) were obtained via printing. The hydrogels show tuneable mechanical properties and flexibility in post processing. A series of structures were fabricated to test the printability, including ear, nose, osteons, microfluidic pads, and porous scaffolds. Aligned micro structures (∼50 μm) of dehydrated scaffold were also achieved via directional freezing. Moreover, the scaffolds could be further stiffened by immersing into the mineralisation solution, to mimic nanocomposite ECM of native bone. The hydrogels exhibited low cytotoxicity and supported the attachment and proliferation of 3T3 fibroblasts. Conclusion The collagen/PEGDA bioink was fabricated for the digital light processing technique. It is capable of being printed into high-resolution structures with tuneable properties for various tissue regeneration applications.

Influence of Gelatin Source and Bloom Number on Gelatin Methacryloyl Hydrogels Mechanical and Biological Properties for Muscle Regeneration.

Approximately half of an adult human's body weight is made up of muscles. Thus, restoring the functionality and aesthetics of lost muscle tissue is critical. The body is usually able to repair minor muscle injuries. However, when volumetric muscle loss occurs due to tumour extraction, for instance, the body will form fibrous tissue instead. Gelatin methacryloyl (GelMA) hydrogels have been applied for drug delivery, tissue adhesive, and various tissue engineering applications due to their tuneable mechanical properties. Here, we have synthesised GelMA from different gelatin sources (i.e., porcine, bovine, and fish) with varying bloom numbers, which refers to the gel strength, and investigated for the influence of the source of gelatin and the bloom number on biological activities and mechanical properties. The results indicated that the source of the gelatin and variable bloom numbers have an impact on GelMA hydrogel properties. Furthermore, our findings established that the bovine-derived gelatin methacryloyl (B-GelMA) has better mechanical properties than the other varieties composed of porcine and fish with 60 kPa, 40 kPa, and 10 kPa in bovine, porcine, and fish, respectively. Additionally, it showed a noticeably greater swelling ratio (SR) ~1100% and a reduced rate of degradation, improving the stability of hydrogels and giving cells adequate time to divide and proliferate to compensate for muscle loss. Furthermore, the bloom number of gelatin was also proven to influence the mechanical properties of GelMA. Interestingly, although GelMA made of fish had the lowest mechanical strength and gel stability, it demonstrated excellent biological properties. Overall, the results emphasise the importance of gelatin source and bloom number, allowing GelMA hydrogels to have a wide range of mechanical and excellent biological properties and making them suitable for various muscle tissue regeneration applications.

Synthesis of nanostructured silica particles for controlled release of ascorbic acid: Microstructure features and In Vitro scratch wound assay.

To date, the long term stability of ascorbic acid (AA) under physiological conditions represents a major issue for wound healing and tissue regeneration applications. In this study, ascorbylphosphate (AP) was loaded into silica nanoparticles (SiNPs) through a simple one-step procedure, in which sphericalshaped porous SiNPs were obtained via hydrolysis/condensation of tetraethylorthosilicate (TEOS) in the presence of bicarbonate salt and ammonia. The as-prepared SiNPs were characterized by scanning electron microscope (SEM), transmission electron microscope (TEM) and Fourier Transformer Infrared Spectrophotometer (FTIR). Incorporation of bicarbonate salt resulted in the formation of spherical SiNPs with an average diameter of 460±89nm, while further increase of bicarbonate salt led to the formation of silica sheet-like structures.The AP-loaded SiNPs exhibited high loading efficiency from 92.3- 81.5%, according to AP content and sustained release over 3 days. According to cell viability assay, the obtained AP-enriched SiNPS showed no toxicity and supportive effect to the proliferation of human skin fibroblast cells (HSF) at a concentration less than 200μg/mL. Moreover, it was observed that the wound closure percentage (%) after 24h was also shown to increase to 74.1±3.1% for 20AP-loaded SiNPs compared to control samples (50.1±1.8%). The obtained results clearly demonstrated that the developed SiNPs formulation exhibits optimal microstructure features to maintain a sustained release of AA at wound bed for the healing of skin tissue, including acute and chronic wounds. This article is protected by copyright. All rights reserved.

Integrated Osteoinductive Factors─Exosome@MicroRNA-26a Hydrogel Enhances Bone Regeneration.

MicroRNAs (miRNAs) are a new therapeutic tool that can target multiple genes by inducing translation repression and target mRNA degradation. Although miRNAs have gained significant attention in oncology and in work on genetic disorders and autoimmune diseases, their application in tissue regeneration remains hindered by several challenges, such as miRNA degradation. Here, we reported Exosome@MicroRNA-26a (Exo@miR-26a), an osteoinductive factor that can be substituted for routinely used growth factors, which was constructed using bone marrow stem cell (BMSC)-derived exosomes and microRNA-26a (miR-26a). Exo@miR-26a-integrated hydrogels significantly promoted bone regeneration when implanted into defect sites; as the exosome stimulated angiogenesis, miR-26a promoted osteogenesis while the hydrogel enabled a site-directed release. Moreover, BMSC-derived exosomes further facilitated healthy bone regeneration by repressing osteoclast differentiation-related genes rather than damaging osteoclasts. Taken together, our findings demonstrate the promising potential of Exo@miR-26a for bone regeneration and provide a new strategy for the application of miRNA therapy in tissue engineering.

Titanium dioxide doped phosphate glasses modulating pro-inflammatory macrophages reponses for tissue regeneration application

ObjectivesThere is an urgent need to understand how novel biomaterials interact with the immune system to improve patients’ safety. This study aimed to explore the immune-inflammatory responses of titanium dioxide doped phosphate glass (TDPG) using RAW 264.7 macrophage cells. MethodsThe ion release was investigated using Inductively Coupled Plasma Optical Emission Spectroscopy and Ion Chromatography. Cell attachment, viability, percentage apoptosis, cell cycle stages, release of reactive oxygen species (ROS) and pro-inflammatory (IL-6, IL-β & TNF-α) and anti-inflammatory (IL-10) cytokines expression were explored for cells grown directly or indirectly in presence of TDPG using different microscopy techniques, XTT assay, flow cytometry, ROS Assay and RT-PCR respectively. The commercially available ProRoot® MTA was used as a control material and cells grown in the normal tissue culture medium was used as control cells. ResultsTDPG released less calcium but more phosphorus than ProRoot® MTA. Aluminum release was only seen with ProRoot® MTA, but sodium, and titanium were detected with TDPG. Macrophages grown in presence of TDPG or its extract showed rounded morphology with expression of F4/80 surface marker, comparable viability and percentage apoptosis to control cells while with ProRoot® MTA, cells showed an enlarged morphology, lower viability, higher percentage apoptosis than control cells. TDPG even at twice dose as ProRoot® MTA did not affect the cell cycle stages as control cells while ProRoot® MTA induced significant changes in the G1, S, G2/M phases and increased cell granularity. ROS level significantly increased in presence of ProRoot® MTA when compared with TDPG and control cells. A transient induction of pro-inflammatory cytokines (IL-6, IL1β, TNF-α) was observed with TDPG treatments whereas their levels sustained for longer time periods with ProRoot® MTA. ConclusionsTDPG showed better immune response than ProRoot® MTA and can modulate tissue regeneration.

Development of Three-Dimensional (3D) Biodegradable Polyglycolic Acid Fiber (PGA) Preforms for Scaffold Applications: Experimental Patterning and Fiber Volume Fraction-Porosity Modeling Study.

Three-dimensional (3D) biodegradable polyglycolic acid fiber (PGA) preforms were developed as temporary scaffolds for three-dimensional tissue regeneration applications. Three-dimensional biodegradable polyglycolic acid fiber (PGA) preforms including various degrees of interlaced structures called 3D plain, semi-interlaced, and orthogonal woven preforms were designed. Analytical relations and finite element model-based software (TexGen) on fiber volume fraction and porosity fraction were proposed to predict scaffolds' stiffness and strength properties considering micromechanics relations. It was revealed that yarn-to-yarn space, density, and angles of all 3D PGA fiber preforms were heterogeneous and demonstrated direction-dependent features (anisotropy). Total fiber volume fractions (Vfp) and porosity fraction (Vtpr) predicted by analytic and numerical modelling of all 3D scaffolds showed some deviations compared to the measured values. This was because yarn cross-sections in the scaffolds were changed from ideal circular yarn (fiber TOW) geometry to high-order ellipse (lenticular) due to inter-fiber pressure generated under a tensile-based macrostress environment during preform formation. Z-yarn modulus (Ez-yarn) and strength (σz-yarn) were probably critical values due to strong stiffness and strength in the through-the-thickness direction where hydrogel modulus and strengths were negligibly small. Morphology of the scaffold showed that PGA fiber sets in the preform were locally distorted, and they appeared as inconsistent and inhomogeneous continuous fiber forms. Additionally, various porosity shapes in the preform based on the virtual model featured complex shapes from nearly trapezoidal beams to partial or concave rectangular beams and ellipsoid rectangular cylinders. It was concluded that 3D polyglycolic acid fiber preforms could be a temporary supportive substrate for 3D tissue regeneration because cells in the scaffold's thickness can grow via through-the-thickness fiber (z-yarn), including various possible mechanobiology mechanisms.

Microporous/Macroporous Polycaprolactone Scaffolds for Dental Applications.

This study leverages the advantages of two fabrication techniques, namely, melt-extrusion-based 3D printing and porogen leaching, to develop multiphasic scaffolds with controllable properties essential for scaffold-guided dental tissue regeneration. Polycaprolactone-salt composites are 3D-printed and salt microparticles within the scaffold struts are leached out, revealing a network of microporosity. Extensive characterization confirms that multiscale scaffolds are highly tuneable in terms of their mechanical properties, degradation kinetics, and surface morphology. It can be seen that the surface roughness of the polycaprolactone scaffolds (9.41 ± 3.01 µm) increases with porogen leaching and the use of larger porogens lead to higher roughness values, reaching 28.75 ± 7.48 µm. Multiscale scaffolds exhibit improved attachment and proliferation of 3T3 fibroblast cells as well as extracellular matrix production, compared with their single-scale counterparts (an approximate 1.5- to 2-fold increase in cellular viability and metabolic activity), suggesting that these structures could potentially lead to improved tissue regeneration due to their favourable and reproducible surface morphology. Finally, various scaffolds designed as a drug delivery device were explored by loading them with the antibiotic drug cefazolin. These studies show that by using a multiphasic scaffold design, a sustained drug release profile can be achieved. The combined results strongly support the further development of these scaffolds for dental tissue regeneration applications.

Impact of Adipose Tissue Depot Harvesting Site on the Multilineage Induction Capacity of Male Rat Adipose-Derived Mesenchymal Stem Cells: An In Vitro Study.

Recently, substantial attention has been paid toward adipose-derived mesenchymal stem cells (AdMSCs) as a potential therapy in tissue engineering and regenerative medicine applications. Rat AdMSCs (r-AdMSCs) are frequently utilized. However, the influence of the adipose depot site on the multilineage differentiation potential of the r-AdMSCs is still ambiguous. Hence, the main objective of this study was to explore the influence of the adipose tissue harvesting location on the ability of r-AdMSCs to express the stem-cell-related markers and pluripotency genes, as well as their differentiation capacity, for the first time. Herein, we have isolated r-AdMSCs from the inguinal, epididymal, peri-renal, and back subcutaneous fats. Cells were compared in terms of their phenotype, immunophenotype, and expression of pluripotency genes using RT-PCR. Additionally, we investigated their potential for multilineage (adipogenic, osteogenic, and chondrogenic) induction using special stains confirmed by the expression of the related genes using RT-qPCR. All cells could positively express stem cell marker CD 90 and CD 105 with no significant in-between differences. However, they did not express the hematopoietic markers as CD 34 and CD 45. All cells could be induced successfully. However, epididymal and inguinal cells presented the highest capacity for adipogenic and osteogenic differentiation (21.36-fold and 11.63-fold for OPN, 29.69-fold and 26.68-fold for BMP2, and 37.67-fold and 22.35-fold for BSP, respectively, in epididymal and inguinal cells (p < 0.0001)). On the contrary, the subcutaneous cells exhibited a superior potential for chondrogenesis over the other sites (8.9-fold for CHM1 and 5.93-fold for ACAN, (p < 0.0001)). In conclusion, the adipose tissue harvesting site could influence the differentiation capacity of the isolated AdMSCs. To enhance the results of their employment in various regenerative cell-based therapies, it is thus vital to take the collection site selection into consideration.

Application of periodontal tissue regeneration combined with orthodontics in oral prosthodontics and its influence and significance on the expressions of IL-1β, TNF-α and IL-5 in periodontal tissue

ABSTRACT The aim is to investigate the application of periodontal tissue regeneration combined with orthodontics in oral restoration, and explore its effect and significance on the expressions of Interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) and interleukin-5 (IL-5) in periodontal tissue. The patients in observation group were treated with orthodontics combined with periodontal tissue regeneration, and the control group was treated with periodontal tissue regeneration. The total effective rate, adverse reactions, recurrence rate and treatment satisfaction were compared. The masticatory function, language function, aesthetic level, VAS score, quality of life, gingival index (GI), plaque index (PLI), periodontal pocket probing depth (PD), sulcus bleeding index (SBI), IL-1β, TNF-α and IL-5 levels were compared. The recurrence rate of observation group was lower than control group, while the treatment satisfaction was higher after treatment. After treatment, the scores of masticatory, language, aesthetics, physiological, social, emotional, cognitive, and emotional functions and overall health score were higher than before treatment. After treatment, the scores of masticatory and language functions, aesthetics and quality of life of observation group were significantly higher than control group. After treatment, the VAS score, GI, PLI, SBI, PD, IL-1β, TNF-α and IL-5 levels were lower than before. The VAS score, GI, PLI, SBI, PD levels, IL-1β, TNF-α and IL-5 levels of observation group were lower after treatment. Orthodontics combined with periodontal tissue regeneration can help improve the periodontal condition of patients with periodontitis, reduce inflammatory response, improve the level of efficacy and overall safety, and further improve patients’ quality of life and treatment satisfaction.

Gelation in Alginate-Based Magnetic Suspensions Favored by Poor Interaction among Sodium Alginate and Embedded Particles

Alginate gels are extensively tested in biomedical applications for tissue regeneration and engineering. In this regard, the modification of alginate gels and solutions with dispersed magnetic particles gives extra options to control the rheo-elastic properties both for the fluidic and gel forms of alginate. Rheological properties of magnetic suspensions based on Na-alginate water solution with embedded magnetic particles were studied with respect to the interfacial adhesion of alginate polymer to the surface of particles. Particles of magnetite (Fe3O4), metallic iron (Fe), metallic nickel (Ni), and metallic nickel with a deposited carbon layer (Ni@C) were taken into consideration. Storage modulus, loss modulus, and the shift angle between the stress and the strain were characterized by the dynamic mechanical analysis in the oscillatory mode. The intensity of molecular interactions between alginate and the surface of the particles was characterized by the enthalpy of adhesion which was determined from calorimetric measurements using a thermodynamic cycle. Strong interaction at the surface of the particles resulted in the dominance of the “fluidic” rheological properties: the prevalence of the loss modulus over the storage modulus and the high value of the shift angle. Meanwhile, poor interaction of alginate polymer with the surface of the embedded particles favored the “elastic” gel-like properties with the dominance of the storage modulus over the loss modulus and low values of the shift angle.

High-throughput microgel biofabrication via air-assisted co-axial jetting for cell encapsulation, 3D bioprinting, and scaffolding applications

Microgels have recently received widespread attention for their applications in a wide array of domains such as tissue engineering, regenerative medicine, and cell and tissue transplantation because of their properties like injectability, modularity, porosity, and the ability to be customized in terms of size, form, and mechanical properties. However, it is still challenging to mass (high-throughput) produce microgels with diverse sizes and tunable properties. Herein, we utilized an air-assisted co-axial device (ACAD) for continuous production of microgels in a high-throughput manner. To test its robustness, microgels of multiple hydrogels and their combination, including alginate (Alg), gelatin methacrylate (GelMA) and Alg–GelMA, were formed at a maximum production rate of ∼65 000 microgels s−1 while retaining circularity and a size range of 50–500 µm based on varying air pressure levels. The ACAD platform allowed single and multiple cell encapsulation with 74 ± 6% efficiency. These microgels illustrated appealing rheological properties such as yield stress, viscosity, and shear modulus for bioprinting applications. Specifically, Alg microgels have the potential to be used as a sacrificial support bath while GelMA microgels have potential for direct extrusion both on their own or when loaded in a bulk GelMA hydrogel. Generated microgels showed high cell viability (>90%) and proliferation of MDA-MB-231 and human dermal fibroblasts over seven days in both encapsulation and scaffolding applications, particularly for GelMA microgels. The developed strategy provides a facile and rapid approach without any complex or expensive consumables and accessories for scalable high-throughput microgel production for cell therapy, tissue regeneration and 3D bioprinting applications.

Fabrication of Interleukin-4 Encapsulated Bioactive Microdroplets for Regulating Inflammation and Promoting Osteogenesis.

Despite the inherent regenerative ability of bone, large bone defect regeneration remains a major clinical challenge for orthopedic surgery. Therapeutic strategies medicated by M2 phenotypic macrophages or M2 macrophage inducer have been widely used to promote tissue remodeling. In this study, ultrasound-responsive bioactive microdroplets (MDs) encapsulated with bioactive molecule interleukin-4 (IL4, hereafter designated MDs-IL4) were fabricated to regulate macrophage polarization and potentiate the osteogenic differentiation of human mesenchymal stem cells (hBMSCs). The MTT assay, live and dead staining, and phalloidin/DAPI dual staining were used to evaluate biocompatibility in vitro. H&E staining was used to evaluate biocompatibility in vivo. Inflammatory macrophages were further induced via lipopolysaccharide (LPS) stimulation to mimic the pro-inflammatory condition. The immunoregulatory role of the MDs-IL4 was tested via macrophage phenotypic marker gene expression, pro-inflammatory cytokine level, cell morphological analysis, and immunofluorescence staining, etc. The immune-osteogenic response of hBMSCs via macrophages and hBMSCs interactions was further investigated in vitro. The bioactive MDs-IL4 scaffold showed good cytocompatibility in RAW 264.7 macrophages and hBMSCs. The results confirmed that the bioactive MDs-IL4 scaffold could reduce inflammatory phenotypic macrophages, as evidenced by changing in morphological features, reduction in pro-inflammatory marker gene expression, increase of M2 phenotypic marker genes, and inhibition of pro-inflammatory cytokine secretion. Additionally, our results indicate that the bioactive MDs-IL4 could significantly enhance the osteogenic differentiation of hBMSCs via its potential immunomodulatory properties. Our results demonstrate that the bioactive MDs-IL4 scaffold could be used as novel carrier system for other pro-osteogenic molecules, thus having potential applications in bone tissue regeneration.

Front Cover: Digital light processing (DLP)‐based (bio)printing strategies for tissue modeling and regeneration

Digital light processing (DLP)-based bioprinting technology has been exploited for a wide range of applications in tissue repair, tissue regeneration, disease modeling and drug screening. In article number e270, Li and co-workers comprehensively summarize works about DLP-based bioprinting strategy to deliver biomedical materials and/or specific cells to create sophisticated structures for various tissue modeling and regeneration.

Lignin-enriched tricalcium phosphate/sodium alginate 3D scaffolds for application in bone tissue regeneration

The bone is a connective, vascularized, and mineralized tissue that confers protection to organs, and participates in the support and locomotion of the human body, maintenance of homeostasis, as well as in hematopoiesis. However, throughout the lifetime, bone defects may arise due to traumas (mechanical fractures), diseases, and/or aging, which when too extensive compromise the ability of the bone to self-regenerate. To surpass such clinical situation, different therapeutic approaches have been pursued. Rapid prototyping techniques using composite materials (consisting of ceramics and polymers) have been used to produce customized 3D structures with osteoinductive and osteoconductive properties. In order to reinforce the mechanical and osteogenic properties of these 3D structures, herein, a new 3D scaffold was produced through the layer-by-layer deposition of a tricalcium phosphate (TCP), sodium alginate (SA), and lignin (LG) mixture using the Fab@Home 3D-Plotter. Three different TCP/LG/SA formulations, LG/SA ratio 1:3, 1:2, or 1:1, were produced and subsequently evaluated to determine their suitability for bone regeneration. The physicochemical assays demonstrated that the LG inclusion improved the mechanical resistance of the scaffolds, particularly in the 1:2 ratio, since a 15 % increase in the mechanical strength was observed. Moreover, all TCP/LG/SA formulations showed an enhanced wettability and maintained their capacity to promote the osteoblasts' adhesion and proliferation as well as their bioactivity (formation of hydroxyapatite crystals). Such results support the LG inclusion and application in the development of 3D scaffolds aimed for bone regeneration.

Synergetic Effect of Electrical and Topographical Cues in Aniline Trimer-Based Polyurethane Fibrous Scaffolds on Tissue Regeneration.

Processibility and biodegradability of conductive polymers are major concerns when they are applied to tissue regeneration. This study synthesizes dissolvable and conductive aniline trimer-based polyurethane copolymers (DCPU) and processes them into scaffolds by using electrospinning with different patterns (random, oriented, and latticed). The effects of topographic cue changes on electrical signal transmission and further regulation of cell behaviors concerning bone tissue are researched. Results show that DCPU fibrous scaffolds possessed good hydrophilicity, swelling capacity, elasticity, and fast biodegradability in enzymatic liquid. In addition, the conductivity and efficiency of electrical signal transmission can be tuned by changing the surface's topological structure. Among them, oriented DCPU scaffolds (DCPU-O) showed the best conductivity with the lowest ionic resistance value. Furthermore, the viability and proliferation results of bone mesenchymal stem cells (BMSCs) demonstrate a significant increase on three DCPU scaffolds compared to AT-free scaffolds (DPU-R). Especially, DCPU-O scaffolds exhibit superior abilities to promote cell proliferation because of their unique surface topography and excellent electroactivity. Concurrently, the DCPU-O scaffolds can synergistically promote osteogenic differentiation in terms of osteogenic differentiation and gene expression levels when combined with electrical stimulation. Together, these results suggest a promising use of DCPU-O fibrous scaffolds in the application of tissue regeneration.

Immune evaluation of granulocyte-macrophage colony stimulating factor loaded hierarchically 3D nanofiber scaffolds in a humanized mice model.

Background: Immune evaluation of biomaterials for tissue regeneration is a critical preclinical evaluation. The current evaluation criterion (ISO 10993-1 or GB/T 16886) uses rodents to perform the immune evaluation. However, the immune system of rodents is different from humans, the obtained results may not be reliable, which could lead directly to the failure of clinical trials. Granulocyte-macrophage colony-stimulating factor (GM-CSF) shows a great potential application in tissue regeneration by regulating local immune responses. The presented work combines the advantages of GM-CSF (immunoregulation) and hierarchically 3D nanofiber scaffolds (tissue regeneration). Methods: Firstly, we fabricated GM-CSF loaded 3D radially aligned nanofiber scaffolds, and then subcutaneous implantation was performed in humanized mice. The whole scaffold and surrounding tissue were harvested at each indicated time point. Finally, the cell infiltration and local immune responses were detected by histological observations, including H&E and Masson staining and immunochemistry. Results: We found significant cell migration and extracellular matrix deposition within the 3D radially aligned nanofiber scaffold after subcutaneous implantation. The locally released GM-CSF could accelerate the expression of human dendritic cells (CD11c) only 3days after subcutaneous implantation. Moreover, higher expression of human cytotoxic T cells (CD3+/CD8+), M1 macrophages (CD68/CCR7) was detected within GM-CSF loaded radially aligned nanofiber scaffolds and their surrounding tissues. Conclusions: The 3D radially aligned scaffold can accelerate cell migration from surrounding tissues to regenerate the wound area. And the locally released GM-CSF enhances dendritic cell recruitment and activation of cytotoxic T cells and M1 macrophages. Taken together, the GM-CSF loaded 3D radially aligned nanofiber scaffolds have a promising potential for achieving tissue regeneration.

High-resolution lightweight and multifunctional 3D printed scaffolds for cell studies

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.

Designing an Innovative Electrospinning Strategy to Generate PHBV Nanofiber Scaffolds with a Radially Oriented Fibrous Pattern.

Electrospinning has contributed substantially to the construction of nanofibrous scaffolds for potential tissue engineering and regenerative medicine applications. However, conventional electrospinning only has the ability to generate and collect nanofiber scaffolds with a randomly oriented fibrous pattern, which lack the necessary cell alignment guidance function. In this study, a novel electrospinning fiber-collecting device was designed and developed by setting a series of small pin-ring-structured collectors on a large plain plate. Specifically, we demonstrated that the pin-ring-structured collectors, which were constructed by inserting a metal pin into the center of a metal ring, could collect the as-electrospun nanofibers with radially oriented structures in an innovative manner. We first investigated the suitable polymeric concentration for electrospinning poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), and the optimum electrospinning concentration of PHBV was found to be 12% (w/v) PHBV dissolved in hexafluoroisopropyl alcohol (HFIP). Then, 12% (w/v) PHBV solution was electrospun into radially oriented nanofiber scaffolds using our novel electrospinning strategy, and their various performances were further compared with conventionally randomly oriented nanofiber scaffolds that were also produced from 12% (w/v) PHBV solution. The results showed that the radially oriented PHBV nanofiber scaffolds exhibited obviously enhanced mechanical properties and decreased hydrophobicity compared with the randomly oriented PHBV nanofiber scaffold controls. Importantly, the biological properties of radially oriented PHBV nanofiber scaffolds were also demonstrated to be enhanced, compared with randomly oriented PHBV nanofiber scaffolds, by effectively inducing cell alignment and significantly promoting cell proliferation. In sum, the present study indicates that our as-prepared nanofiber scaffolds with a radially oriented pattern are of great interest for advanced applications, such as wound dressings and tissue-engineered scaffolds.