Conventional Scaffolds Research Articles

Low temperature hybrid 3D printing of hierarchically porous bone tissue engineering scaffolds with in situ delivery of osteogenic peptide and mesenchymal stem cells

Compared to other conventional scaffold fabrication techniques, three-dimensional (3D) printing is advantageous in producing bone tissue engineering scaffolds with customized shape, tailored pore size/porosity, required mechanical properties and even desirable biomolecule delivery capability. However, for scaffolds with a large volume, it is highly difficult to get seeded cells to migrate to the central region of the scaffolds, resulting in an inhomogeneous cell distribution and therefore lowering the bone forming ability. To overcome this major obstacle, in this study, cell-laden bone tissue engineering scaffolds consisting of osteogenic peptide (OP) loaded β-tricalcium phosphate (TCP)/poly(lactic-co-glycolic acid) (PLGA) (OP/TCP/PLGA, designated as OTP) nanocomposite struts and rat bone marrow derived mesenchymal stem cell (rBMSC)-laden gelatin/GelMA hydrogel rods were produced through ‘dual-nozzle’ low temperature hybrid 3D printing. The cell-laden scaffolds exhibited a bi-phasic structure and had a mechanical modulus of about 19.6 MPa, which was similar to that of human cancellous bone. OP can be released from the hybrid scaffolds in a sustained manner and achieved a cumulative release level of about 78% after 24 d. rBMSCs encapsulated in the hydrogel rods exhibited a cell viability of about 87.4% right after low temperature hybrid 3D printing and could be released from the hydrogel rods to achieve cell anchorage on the surface of adjacent OTP struts. The OP released from OTP struts enhanced rBMSCs proliferation. Compared to rBMSC-laden hybrid scaffolds without OP incorporation, the rBMSC-laden hybrid scaffolds incorporated with OP significantly up-regulated osteogenic differentiation of rBMSCs by showing a higher level of alkaline phosphatase expression and calcium deposition. This ‘proof-of-concept’ study has provided a facile method to form cell-laden bone tissue engineering scaffolds with not only required mechanical strength, biomimetic structure and sustained biomolecule release profile but also excellent cell delivery capability with uniform cell distribution, which can improve the bone forming ability in the body.

Ultrasound induced, easy-to-store porous poly(amino acid) based electrospun scaffolds

• Ultrasonication affected the mechanical properties of the electrospun scaffolds. • Enhanced cellular infiltration to the electrospun scaffold. • Presented an alternative storage condition and increasing shelf life for hydrogels. • Decreasing the probability of microbiological damage of the fibrous scaffolds. • Huge step towards an off-the-shelf and effective product for tissue regeneration. The aim of tissue engineering is to develop methods to restore, maintain or improve tissue functions. To imitate the fibrous structure of the native extracellular matrix, the electrospinning technique is widely used. However, the dense packing of fibers results in small pores and hereby the inhibition of cellular penetration. In this study, we used biocompatible and biodegradable poly(aspartic acid) based fibrous hydrogel scaffolds to enhance the cell infiltration using ultrasonication (US). The US can enlarge the space between the fibers in the scaffold and create a 3D structure based on the thickness increase of the samples. To prevent the scaffolds from degradation and create an easy-to-store sample beyond the US treatment, a freeze-drying process was also introduced in this work. After all these treatments, the scaffold’s specific load capacity was 0.11 ± 0.01 Nm 2 /g which did not change after a rehydration cycle and the elongation break became almost two times higher than before the US treatment. The cytotoxicity results demonstrated that the cellular viability did not show any significant decrease compared to the control groups for none of the samples. The cellular penetration was visualized by multiphoton microscopy. In summary, we were able to overcome the major limitation of conventional electrospun scaffolds regarding their application in tissue engineering. We also improved the storing conditions of fibrous hydrogel scaffolds and extend their shelf life without degradation.

Three-Dimensional Culture of Cartilage Tissue on Nanogel-Cross-Linked Porous Freeze-Dried Gel Scaffold for Regenerative Cartilage Therapy: A Vibrational Spectroscopy Evaluation.

This study presents a set of vibrational characterizations on a nanogel-cross-linked porous freeze-dried gel (NanoCliP-FD gel) scaffold for tissue engineering and regenerative therapy. This scaffold is designed for the in vitro culture of high-quality cartilage tissue to be then transplanted in vivo to enable recovery from congenital malformations in the maxillofacial area or crippling jaw disease. The three-dimensional scaffold for in-plate culture is designed with interface chemistry capable of stimulating cartilage formation and maintaining its structure through counteracting the dedifferentiation of mesenchymal stem cells (MSCs) during the formation of cartilage tissue. The developed interface chemistry enabled high efficiency in both growth rate and tissue quality, thus satisfying the requirements of large volumes, high matrix quality, and superior mechanical properties needed in cartilage transplants. We characterized the cartilage tissue in vitro grown on a NanoCliP-FD gel scaffold by human periodontal ligament-derived stem cells (a type of MSC) with cartilage grown by the same cells and under the same conditions on a conventional (porous) atelocollagen scaffold. The cartilage tissues produced by the MSCs on different scaffolds were comparatively evaluated by immunohistochemical and spectroscopic analyses. Cartilage differentiation occurred at a higher rate when MSCs were cultured on the NanoCliP-FD gel scaffold compared to the atelocollagen scaffold, and produced a tissue richer in cartilage matrix. In situ spectroscopic analyses revealed the cell/scaffold interactive mechanisms by which the NanoCliP-FD gel scaffold stimulated such increased efficiency in cartilage matrix formation. In addition to demonstrating the high potential of human periodontal ligament-derived stem cell cultures on NanoCliP-FD gel scaffolds in regenerative cartilage therapy, the present study also highlights the novelty of Raman spectroscopy as a non-destructive method for the concurrent evaluation of matrix quality and cell metabolic response. In situ Raman analyses on living cells unveiled for the first time the underlying physiological mechanisms behind such improved chondrocyte performance.

Two-Dimensional Metal-Organic Framework-Based Cellular Scaffolds with High Protein Adsorption, Retention, and Replenishment Capabilities.

Metal-organic frameworks (MOFs) are porous materials with adsorption, storage, and separation capabilities due to their high specific surface areas and large pore volumes. MOFs are thus used in biomedical applications, and MOF nanoparticles have been widely studied as nanocarriers for drug delivery systems. Several research groups recently reported that specific MOF nanoparticles can adsorb and retain proteins, suggesting to us that MOF nanoparticles may have advantages as novel cell culture scaffolds. However, MOF nanoparticles cannot be used as two-dimensional scaffolds for cells. We therefore established a bottom-up technique to construct two-dimensional MOFs [MIL-53 (Al)] on polymer films. The developed two-dimensional MIL-53 (Al) film [fMIL-53 (Al)] exhibited high serum protein adsorption, retention, and replenishment capabilities as compared to conventional cell culture scaffolds. β-Galactosidase, used as a model protein, adsorbed on fMIL-53 (Al) exhibited original enzymatic activity, indicating that proteins are not denatured during the adsorption process. The viability of mouse myoblast cells (C2C12) cultured on fMIL-53 (Al) was 100%, indicating the cell compatibility of fMIL-53 (Al). Importantly, C2C12 cells cultured on serum protein-preadsorbed fMIL-53 (Al) exhibited excellent long-term adhesion, morphology, and proliferation even in a medium lacking serum proteins, demonstrating an important advantage of fMIL-53 (Al) as a cell culture scaffold, given that conventional cell culture scaffolds typically require a serum-containing medium to support stable cell adhesion and proliferation. To our knowledge, this is the first report regarding the application of MOFs as cell culture scaffolds and will serve as a starting point for studying two- and three-dimensional MOF-based cellular scaffolds for cell culture systems and for in vitro and in vivo tissue engineering.

Fabrication and Characterization of Layer-by-Layer Janus Base Nano-Matrix to Promote Cartilage Regeneration.

Various biomaterial scaffolds have been developed to guide cell adhesion and proliferation in hopes to promote specific functions for in vitro and in vivo uses. The addition of growth factors into these biomaterial scaffolds is generally done to provide an optimal cell culture environment, mediating cell differentiation and its subsequent functions. However, the growth factors in a conventional biomaterial scaffold are typically designed to be released upon implantation, which could result in unintended side effects on surrounding tissue or cells. Here, the DNA-inspired Janus base nano-matrix (JBNm) has successfully achieved a highly localized microenvironment with a layer-by-layer structure for self-sustainable cartilage tissue constructs. JBNms are self-assembled from Janus base nanotubes (JBNts), matrilin-3, and transforming growth factor beta-1 (TGF-β1) via bioaffinity. The JBNm was assembled at a TGF-β1:matrilin-3:JBNt ratio of 1:4:10, as this has been the determined ratio at which proper assembly into the layer-by-layer structure could occur. First, the TGF-β1 solution was added to the matrilin-3 solution. Then, this mixture was pipetted several times to ensure sufficient homogeneity before the addition of the JBNt solution. This formed the layer-by-layer JBNm, after pipetting several times again. A variety of experiments were performed to characterize the layer-by-layer JBNm structure, JBNts alone, matrilin-3 alone, and TGF-β1 alone. The formation of JBNm was studied with UV-Vis absorption spectra, and the structure of the JBNm was observed with transmission electron microscopy (TEM). As the innovative layer-by-layer JBNm scaffold is formed on a molecular scale, the fluorescent dye-labeled JBNm could be observed. The TGF-β1 is confined within the inner layer of the injectable JBNm, which can prevent the release of growth factors to surrounding areas, promote localized chondrogenesis, and promote an anti-hypertrophic microenvironment.

Methacrylate-Modified Gold Nanoparticles Enable Non-Invasive Monitoring of Photocrosslinked Hydrogel Scaffolds.

Photocrosslinked hydrogels, such as methacrylate-modified gelatin (gelMA) and hyaluronic acid (HAMA), are widely utilized as tissue engineering scaffolds and/or drug delivery vehicles, but lack a suitable means for non-invasive, longitudinal monitoring of surgical placement, biodegradation, and drug release. Therefore, we developed a novel photopolymerizable X-ray contrast agent, methacrylate-modified gold nanoparticles (AuMA NPs), to enable covalent-linking to methacrylate-modified hydrogels (gelMA and HAMA) in one-step during photocrosslinking and non-invasive monitoring by X-ray micro-computed tomography (micro-CT). Hydrogels exhibited a linear increase in X-ray attenuation with increased Au NP concentration to enable quantitative imaging by contrast-enhanced micro-CT. The enzymatic and hydrolytic degradation kinetics of gelMA-Au NP hydrogels were longitudinally monitored by micro-CT for up to one month in vitro, yielding results that were consistent with concurrent measurements by optical spectroscopy and gravimetric analysis. Importantly, AuMA NPs did not disrupt the hydrogel network, rheology, mechanical properties, and hydrolytic stability compared with gelMA alone. GelMA-Au NP hydrogels were thus able to be bioprinted into well-defined three-dimensional architectures supporting endothelial cell viability and growth. Overall, AuMA NPs enabled the preparation of both conventional photopolymerized hydrogels and bioprinted scaffolds with tunable X-ray contrast for noninvasive, longitudinal monitoring of placement, degradation, and NP release by micro-CT.

The assessment of average cell number inside in-flight 3D printed droplets in microvalve-based bioprinting

3D cell bioprinting is an innovative and time-saving additive manufacturing technology; it precisely generates complex cell-laden constructs to overcome the limitations of 2D cell culture and conventional tissue engineering scaffold technology. Many efforts have been made to evaluate the bioprinter performance by considering printed cell number and the consistency of printed cell number. In this paper, a modified droplet imaging system is used to study the printing performance for a micro-valve-based 3D bioprinter using fluorescence MCF-7 cells. The effects of droplet dispensing physics (dosing energy Ed), ink properties (Z number—the inverse of the Ohnesorge number and particle sedimentation velocity), and input cell concentration are considered. The droplet imaging system demonstrates a strong capability and accuracy in analyzing bioprinting performance for printed cell density less than 300 cells/droplet. The average printed cell number is positively correlated with the increasing input cell concentration, dosing energy, and printing time. Printing ink, with Z number ranging from 4 to 7.41 and cell sedimentation velocity at 9.45×10−8 m/s, can provide the estimated printed cell number and consistent cell printing results within 2 min printing time. Printing inks with higher Z number or cell sedimentation velocity should be ejected under dosing energy below 2.1 La and printed right after filling the reservoir to achieve reliable and stable printing results.

Improvement of the mechanical properties and osteogenic activity of 3D-printed polylactic acid porous scaffolds by nano-hydroxyapatite and nano-magnesium oxide

Porous bone scaffolds based on high-precision 3D printing technology gave recently been developed for use in bone defect repair. However, conventional scaffold materials have poor mechanical properties and low osteogenic activity, limiting their clinical use. In this study, a porous composite tissue-engineered bone scaffold was prepared using polylactic acid, nano-hydroxyapatite, and nano-magnesium oxide as raw materials for high-precision 3D printing. The composite scaffold takes full advantage of the personalized manufacturing features of 3D printers and can be used to repair complex bone defects in clinical settings. The composite scaffold combines the advantages of nano-hydroxyapatite, which improves the formability of scaffold printing, and of nano-magnesium oxide, which regulates pH during degradation and provide a good environment for cell growth. Additionally, nano-magnesium oxide and nano-hydroxyapatite have a bidirectional effect on promoting the compressive strength and osteogenic activity of the scaffolds. The prepared composite porous scaffolds based on 3D printing technology show promise for bone defect repair.

Three-dimensional-printed polycaprolactone scaffolds with interconnected hollow-pipe structures for enhanced bone regeneration.

Three-dimensional (3D)-printed scaffolds are widely used in tissue engineering to help regenerate critical-sized bone defects. However, conventional scaffolds possess relatively simple porous structures that limit the delivery of oxygen and nutrients to cells, leading to insufficient bone regeneration. Accordingly, in the present study, perfusable and permeable polycaprolactone scaffolds with highly interconnected hollow-pipe structures that mimic natural micro-vascular networks are prepared by an indirect one-pot 3D-printing method. In vitro experiments demonstrate that hollow-pipe-structured (HPS) scaffolds promote cell attachment, proliferation, osteogenesis and angiogenesis compared to the normal non-hollow-pipe-structured scaffolds. Furthermore, in vivo studies reveal that HPS scaffolds enhance bone regeneration and vascularization in rabbit bone defects, as observed at 8 and 12 weeks, respectively. Thus, the fabricated HPS scaffolds are promising candidates for the repair of critical-sized bone defects.

Ultralight electrospun fiber foam with tunable lamellar macropores for efficient interfacial evaporation

Solar-driven interfacial evaporation is regarded as one of the most promising technologies to address the critical issue of global water scarcity. However, a great deal of energy is lost via conduction during the interfacial evaporation process. Herein, a self-floating Janus evaporator containing micron-sized isolated bubbles that can enhance heat localization is designed. The evaporator is synthesized by gas foaming polyacrylonitrile (PAN) nanofibers and subsequently depositing polypyrrole (PPy). The hydrophilic PAN nanofiber layer on the bottom of the evaporator with abundant macropores can continuously pump water upward through the capillary force. The hydrophobic PAN@PPy composite nanofiber layer on the top possesses a broadband solar absorption property, converting solar irradiation to heat efficiently. The low thermal conductivity of the isolated micron-sized bubbles enhances the thermal insulation of the obtained PAN foam@PPy Janus evaporator, while the stretching and expanding of the nanofibers induced by the bubbles contribute to its increased mechanical stability. This work offers a simple route to fabricate evaporators that can continuously produce drinking water from brine under sunlight, and provides a fresh idea to enhance the thermal insulation and mechanical properties of the conventional electrospinning-based fiber scaffold.

Bio-inspired zonal-structured matrices for bone-cartilage interface engineering

Design and development of scaffold structures for osteochondral (OC) interface regeneration is a significant engineering challenge. Recent efforts are aimed at recapitulating the unique compositional and hierarchical structure of an OC interface. Conventional scaffold fabrication techniques often have limited design control and reproducibility, and the development of OC scaffolds with zonal hierarchy and structural integrity between zones is especially challenging. In this study, a series of multi-zonal and gradient structures were designed and fabricated using three-dimensional bioprinting. We developed OC scaffolds with bi-phasic and tri-phasic configurations to support the zonal structure of OC tissue, and gradient scaffold configurations to enable smooth transitions between the zones to more closely mimic a bone-cartilage interface. A biodegradable polymer, polylactic acid, was used for the fabrication of zonal/gradient scaffolds to provide mechanical strength and support OC function. The formation of the multi-zonal and gradient scaffolds was confirmed through scanning electron microscopy imaging and micro-computed tomography scanning. Precisely controlled hierarchy with tunable porosity along the scaffold length established the formation of the bio-inspired scaffolds with different zones/gradient structure. In addition, we also developed a novel bioprinting method to selectively introduce cells into desired scaffold zones of the zonal/gradient scaffolds via concurrent printing of a cell-laden hydrogel within the porous template. Live/dead staining of the cell-laden hydrogel introduced in the cartilage zone showed uniform cell distribution with high cell viability. Overall, our study developed bio-inspired scaffold structures with structural hierarchy and mechanical integrity for bone-cartilage interface engineering.

Multiscale porous scaffolds constructed of carbonate apatite honeycomb granules for bone regeneration

The use of scaffolds with pores ranging from macroscale to nanoscale (i.e., multiscale pores), is an effective strategy to achieve favorable bone regeneration. Here, we report the fabrication of multiscale porous scaffolds (MPSs) and evaluate the effects of macropores (>100 μm) and micropores (<10 μm) on bone regeneration in the early to medium term. MPSs were constructed from interconnecting carbonate apatite honeycomb granules. The uniaxial macropores penetrating the honeycomb granules linked the gaps among the granules, creating an interconnected macroporous architecture. MPSs with different proportions of macropores and micropores were implanted into rabbit femurs. In an early stage, macropores played a crucial role in promoting new bone formation. In the medium term, micropores, rather than macropores, were the dominant factor affecting the replacement of MPSs with new bone. Notably, micropores of 100 nm to 10 μm primarily promoted MPS resorption by osteoclasts, stimulating secondary osteogenesis. Our findings demonstrated that pore size distribution, rather than porosity, was crucial for osteogenesis in the early-to-medium term. Although conventional scaffold development sacrifices mechanical strength to improve bone formation, our findings indicate that optimizing the pore size distribution is a promising strategy to develop scaffolds that satisfy the requirements of both osteogenesis and mechanical strength.

Bio-inspired zonal-structured matrices for bone-cartilage interface engineering.

Design and development of scaffold structures for osteochondral (OC) interface regeneration is a significant engineering challenge. Recent efforts are aimed at recapitulating the unique compositional and hierarchical structure of an OC interface. Conventional scaffold fabrication techniques often have limited design control and reproducibility, and the development of OC scaffolds with zonal hierarchy and structural integrity between zones is especially challenging. In this study, a series of multi-zonal and gradient structures were designed and fabricated using three-dimensional (3D) bioprinting. We developed OC scaffolds with bi-phasic and tri-phasic configurations to support the zonal structure of OC tissue, and gradient scaffold configurations to enable smooth transitions between the zones to more closely mimic a bone-cartilage interface. A biodegradable polymer, polylactic acid (PLA), was used for the fabrication of zonal/gradient scaffolds to provide mechanical strength and support OC function. The formation of the multi-zonal and gradient scaffolds was confirmed through SEM imaging and micro-CT scanning. Precisely controlled hierarchy with tunable porosity along the scaffold length established the formation of the bio-inspired scaffolds with different zones/gradient structure. In addition, we also developed a novel bioprinting method to selectively introduce cells into desired scaffold zones of the zonal/gradient scaffolds via concurrent printing of a cell-laden hydrogel within the porous template. Live/dead staining of the cell-laden hydrogel introduced in the cartilage zone showed uniform cell distribution with high cell viability. Overall, our study developed bio-inspired scaffold structures with structural hierarchy and mechanical integrity for bone-cartilage interface engineering.

A Minireview of Microfluidic Scaffold Materials in Tissue Engineering.

In 2020, nearly 107,000 people in the U.S needed a lifesaving organ transplant, but due to a limited number of donors, only ∼35% of them have actually received it. Thus, successful bio-manufacturing of artificial tissues and organs is central to satisfying the ever-growing demand for transplants. However, despite decades of tremendous investments in regenerative medicine research and development conventional scaffold technologies have failed to yield viable tissues and organs. Luckily, microfluidic scaffolds hold the promise of overcoming the major challenges associated with generating complex 3D cultures: 1) cell death due to poor metabolite distribution/clearing of waste in thick cultures; 2) sacrificial analysis due to inability to sample the culture non-invasively; 3) product variability due to lack of control over the cell action post-seeding, and 4) adoption barriers associated with having to learn a different culturing protocol for each new product. Namely, their active pore networks provide the ability to perform automated fluid and cell manipulations (e.g., seeding, feeding, probing, clearing waste, delivering drugs, etc.) at targeted locations in-situ. However, challenges remain in developing a biomaterial that would have the appropriate characteristics for such scaffolds. Specifically, it should ideally be: 1) biocompatible—to support cell attachment and growth, 2) biodegradable—to give way to newly formed tissue, 3) flexible—to create microfluidic valves, 4) photo-crosslinkable—to manufacture using light-based 3D printing and 5) transparent—for optical microscopy validation. To that end, this minireview summarizes the latest progress of the biomaterial design, and of the corresponding fabrication method development, for making the microfluidic scaffolds.

A porous calcium-zirconia scaffolds composed of magnetic nanoparticles for bone cancer treatment: Fabrication, characterization and FEM analysis

One of the most critical challenges in tissue engineering is the fabrication of porous scaffolds with high porosity and proper mechanical properties. Conventional synthetic scaffolds are typically made of ceramic composed with other ceramics; however, a better combination of nanocomposite can be obtained using magnetite nanoparticles (MNPs), which benefits both ceramic materials. The current study used the space holder method and mechanical activation to create a porous magnetic-zirconia calcium bio-nanocomposite scaffold for bone tissue applications. The samples were made with varying amounts of reinforcement (0, 5, 10, and 15 wt%). The addition of sodium chloride has the main effect on the porosity size of the architecture. The prepared magnetic-zirconia calcium bio-nanocomposite scaffold was coated with a 5% solution of chitosan polymer, and its mechanical and biological properties were investigated. Powders and scaffolds were examined by X-ray diffraction (XRD), scanning electron microscopy (SEM), and Fourier transform infrared (FTIR) spectroscopy. The cell viability and non-toxicity of porous scaffolds were evaluated by MTT assay. The results showed that the addition of 15 wt% of MNPs changed the mechanical and biological properties of the composite scaffold. According to the SEM results, addition of 15% MNPs did not significantly change the porosity. However, it improves the porosity percentages. After being placed in the simulated body fluid (SBF), SEM images of the scaffolds showed the formation of a bone-like apatite layer on the surfaces of the sample with a higher amount of MNPs. Biocompatibility assessment by MTT test showed that composite scaffolds did not show any toxicity in contact with bone marrow stem cells and increased cell growth and proliferation. The mechanical simulation shows that the specimen's mechanical strength can be predicted by low error and can be a suitable option for bone tissue engineering applications.

Material Challenges and Opportunities in 3D Printing for Hip Implant Applications

There is a current need for tissue and organ repairs, replacement, and regeneration for patients who suffer from diseased or damaged tissues or organs. This situation is continuously on the rise and the supply of this form of therapy does not meet the patients demand mostly due to lack of donors and biocompatibility issues which causes immune system rejection of the implants. To succeed through these limitations, researchers are currently investigating the use of scaffolds as another approach for implants. The conventional scaffold fabrication technique is limited due to the precision of pore design. The 3D printing technology on the other side can produce an extracellular matrix with a higher degree of complexity and matching details such as pore size and geometry suitably based on certain factors including tissue engineering, hip biomechanism, material suitability, ethical standards, future, and challenges. This paper in particular focuses on materials challenges and opportunities addressing various issues at various levels to the materials-process-property relationship. It is comprehensive as it starts with hip biomechanism in gait and stress distribution to give the reader a clear perspective of the magnitude of challenges for hip implants and details to consider when designing the materials. This is followed by 3D printing for orthopaedic applications and 3D hip tissue regeneration. The hip replacement materials including polymers, composites, and metals are explored and correlated to conventional hip replacement materials. The work is concluded with some concluding remarks on opportunities, challenges, and future trends. The goal is to have scaffolds that have the capability of having a biomimicking design similar to the extracellular matrix with the advantage being the provision of structural supports for cell attachment, growth, and differentiation with the main goal of producing an operational organ or tissue. The knowledge derived from this review offers huge potential for providing a pathway for sustainable healing.

Two-Dimensional Nanomaterials for Peripheral Nerve Engineering: Recent Advances and Potential Mechanisms.

Peripheral nerve tissues possess the ability to regenerate within artificial nerve scaffolds, however, despite the advance of biomaterials that support nerve regeneration, the functional nerve recovery remains unsatisfactory. Importantly, the incorporation of two-dimensional nanomaterials has shown to significantly improve the therapeutic effect of conventional nerve scaffolds. In this review, we examine whether two-dimensional nanomaterials facilitate angiogenesis and thereby promote peripheral nerve regeneration. First, we summarize the major events occurring after peripheral nerve injury. Second, we discuss that the application of two-dimensional nanomaterials for peripheral nerve regeneration strategies by facilitating the formation of new vessels. Then, we analyze the mechanism that the newly-formed capillaries directionally and metabolically support neuronal regeneration. Finally, we prospect that the two-dimensional nanomaterials should be a potential solution to long range peripheral nerve defect. To further enhance the therapeutic effects of two-dimensional nanomaterial, strategies which help remedy the energy deficiency after peripheral nerve injury could be a viable solution.

Cell-Incorporated Bioactive Tissue Engineering Scaffolds made by Concurrent Cell Electrospinning and Emulsion Electrospinning

Electrospun fibrous scaffolds attract great attention in tissue engineering owing to their high similarity in architecture to the extracellular matrix (ECM) that support cell attachment and growth in human bodies. Although they have shown superiority in promoting cell attachment and proliferation on their surfaces and hence, hold great promise for the regeneration of body tissues, the research still faces a great challenge of three-dimensional (3D) cell incorporation in electrospun scaffolds to form thick and cell-dense constructs because deep cell infiltration is hard to achieve in conventional electrospun scaffolds that normally have very small diameters of interconnected pores. Such hindrance has severely limited the clinical application of electrospun fibrous scaffolds to repair/regenerate various body tissues, particularly those with complex anatomies. To address this challenge, we have developed a concurrent cell electrospinning and emulsion electrospinning technique for fabricating bioactive bio-hybrid scaffolds with 3D and high-density cell incorporation. Through concurrent electrospinning, cell-encapsulated hydrogel fibers (“cell fibers”) and growth factor-containing ultrafine fibers are simultaneously deposited to form two-component scaffolds (i.e., scaffolds composed of two types of fibers) according to the design. With the breakup of cell fibers, live cells with well-preserved cell viability are released in situ inside the scaffolds, resulting in the creation of cell-incorporated bioactive scaffolds with ECM-mimicking fibrous architectures and 3D and high-density incorporation of cells. The growth and functions of incorporated cells in the scaffolds can be enhanced by the released growth factor from the emulsion electrospun fibrous component. The bioactive bio-hybrid scaffolds fabricated via concurrent electrospinning mimic the cell-matrix organization of body tissues and therefore have great potential for regenerating body tissues such as tendon and ligament.

Nanofiber configuration affects biological performance of decellularized meniscus extracellular matrix incorporated electrospun scaffolds

Electrospinning represents the simplest approach to fabricate nanofiber scaffolds that approximate the heterogeneous fibrous structure of the meniscus. More effort is needed to understand the relationship between scaffold properties and cell responses to determine the appropriate scaffolds supporting meniscus tissue repair and regeneration. In this study, we investigate the influence of nanofiber configuration of electrospun scaffolds on phenotype and matrix production of meniscus cells, as well as on scaffold degradation behaviors and biocompatibility. Twisting electrospun nanofibers into yarns not only recapitulates the major collagen bundles of the meniscus but also increases the pore size and porosity of resultant scaffolds. The yarn scaffold significantly regulated expression levels of meniscus-associated genes and promoted extracellular matrix production compared with conventional electrospun scaffolds with random or aligned nanofiber orientation. Additionally, the yarn scaffold allowed considerable cell infiltration and experienced faster degradation and tissue remodeling upon subcutaneous implantation in a rat model. These results suggest that nanofiber configuration dictates cell interactions, scaffold degradation and integration with host tissue, providing design parameters of porosity and pore size of electrospun scaffolds toward meniscus repair.

A Three-in-One Strategy: Injectable Biomimetic Porous Hydrogels for Accelerating Bone Regeneration via Shape-Adaptable Scaffolds, Controllable Magnesium Ion Release, and Enhanced Osteogenic Differentiation.

The repair of bone defects with irregular shapes, particularly in a minimally invasive manner, remains a major challenge. For synthetic bone grafts, injectable hydrogels are superior to conventional scaffolds because they can adapt satisfactorily to the defect margins and can be injected into deeper areas of injury via a minimally invasive procedure. Based on the poly(lactide-co-glycolide)(PLGA)/1-methyl-2-pyrrolidinone solution reported in our previous study, we successfully synthesized injectable MgO/MgCO3@PLGA (PMM) hydrogels, namely, injectable biomimetic porous hydrogels (IBPHs), to accelerate bone regeneration. In addition to exhibiting excellent injectability, PMM hydrogels could transform into porous scaffolds in situ through a liquid-to-solid phase transition and completely fill irregular bone defects via their superb shape adaptability. Moreover, sustainable and steady release of Mg2+ was achieved by regulating the weight ratio of the incorporated MgO and MgCO3 particles. Via controlled release of Mg2+, PMM hydrogels significantly promoted proliferation, osteogenic differentiation, migration, and biomineral deposition of immortalized mouse embryonic fibroblasts. More importantly, micro-CT imaging and histological analysis indicated that concomitant with their gradual degradation, PMM hydrogels effectively stimulated in situ bone regeneration in rat calvarial defects with an increase in the bone volume fraction of almost 2-fold compared with that in the control group. These findings suggest that injectable PMM hydrogels can satisfactorily match bone defects and form porous scaffolds in situ and can significantly promote bone regeneration via controllable Mg2+ release. The remarkable features of IPBHs may open a new avenue for the exploration of in situ repair systems for irregular bone defects to accelerate bone regeneration and have great potential for clinical translation.