Wireless Electric Cues Mediate Autologous DPSC-Loaded Conductive Hydrogel Microspheres to Engineer the Immuno-Angiogenic Niche for Homologous Maxillofacial Bone Regeneration.

Stem cell therapy serves as an effective treatment for bone regeneration. Nevertheless, stem cells from bone marrow and peripheral blood are still lacking homologous properties. Dental pulp stem cells (DPSCs) are derived from neural crest, in coincidence with maxillofacial tissues, thus attracting great interest in in situ maxillofacial regenerative medicine. However, insufficient number and heterogenous alteration of seed cells retard further exploration of DPSC-based tissue engineering. Electric stimulation has recently attracted great interest in tissue regeneration. In this study, a novel DPSC-loaded conductive hydrogel microspheres integrated with wireless electric generator is fabricated. Application of exogenous electric cues can promote stemness maintaining and heterogeneity suppression for unpredictable differentiation of encapsulated DPSCs. Further investigations observe that electric signal fine-tunes regenerative niche by improvement on DPSC-mediated paracrine pattern, evidenced by enhanced angiogenic behavior and upregulated anti-inflammatory macrophage polarization. By wireless electric stimulation on implanted conductive hydrogel microspheres, loaded DPSCs facilitates the construction of immuno-angiogenic niche at early stage of tissue repair, and further contributes to advanced autologous mandibular bone defect regeneration. This novel strategy of DPSC-based tissue engineering exhibits promising translational and therapeutic potential for autologous maxillofacial tissue regeneration.

In Situ 4D Printing of Polyelectrolyte/Magnetic Composites for Sutureless Gastric Perforation Sealing.

In situ bioprinting has emerged as one of the most promising techniques for the sutureless tissue sealing of internal organs. However, most existing in situ bioprinting methods are limited by the complex and confined printing space inside the organs, harsh curing conditions for printable bioinks, and poor ability to suturelessly seal injured parts. The combination of in situ bioprinting and 4D printing is a promising technique for tissue repair. Herein, the in situ 4D printing of polyelectrolyte/magnetic composites by gastroscopy for sutureless internal tissue sealing is reported. Using gastric perforation as an example, a gelatin/sodium alginate/magnetic bioink is developed, which can be precisely located by a gastroscope with the assistance of an external magnetic field, solidified in gastric fluid, and firmly adhered to tissue surfaces. The solidified bioink along the defect can be attracted by an external magnetic field, resulting in sutureless sealing. A demonstration using a porcine stomach with an artificial perforation confirms the feasibility of sutureless sealing using 4D printing. Moreover, an in vivo investigation on gastric perforation in a rat model identifies the biocompatibility by H&E and CD68+ staining. This study provides a new orientation and concept for functionality-modified in situ 4D bioprinting.

Copper Ion-Modified Germanium Phosphorus Nanosheets Integrated with an Electroactive and Biodegradable Hydrogel for Neuro-Vascularized Bone Regeneration.

Severe bone defects accompanied by vascular and peripheral nerve injuries represent a huge orthopedic challenge and are often accompanied by the risk of infection. Thus, biomaterials with antibacterial and neurovascular regeneration properties are highly desirable. Here, a newly designed biohybrid biodegradable hydrogel (GelMA) containing copper ion-modified germanium-phosphorus (GeP) nanosheets, which act as neuro-vascular regeneration and antibacterial agents, is designed. The copper ion modification process serves to improve the stability of the GeP nanosheets and offers a platform for the sustained release of bioactive ions. Study findings show that GelMA/GeP@Cu has effective antibacterial properties. The integrated hydrogel can significantly boost the osteogenic differentiation of bone marrow mesenchymal stem cells, facilitate angiogenesis in human umbilical vein endothelial cells, and up-regulate neural differentiation-related proteins in neural stem cells in vitro. In vivo, in the rat calvarial bone defect mode, the GelMA/GeP@Cu hydrogel is found to enhance angiogenesis and neurogenesis, eventually contributing to bone regeneration. These findings indicate that in the field of bone tissue engineering, GelMA/GeP@Cu can serve as a valuable biomaterial for neuro-vascularized bone regeneration and infection prevention.

N-cadherin mimetic hydrogel enhances MSC chondrogenesis through cell metabolism.

Mesenchymal stem cells (MSCs) are ideal candidates for tissue engineering and regenerative medicine because of their proliferative capacity and differentiation potential. However, the hypertrophic phenotype occurring in late MSCs chondrogenic differentiation severely limits their clinical translation. While hypertrophy inhibition strategies have been explored, the role of cell metabolism in MSCs chondrogenesis has rarely been studied. In this study, we found that hypertrophy occurred in the late stage of MSCs chondrogenesis with increased fatty acid oxidation (FAO) and decreased glycolysis, as well as cell-cell junctions impairment. Therefore, a N-cadherin mimetic hydrogel was developed to enhance cell-cell junctions via N-cadherin mimetic peptides and high seeding density. The N-cadherin mimetic hydrogel attenuated hypertrophy through regulating glycolysis and FAO. The regulation of cell-cell junctions mechanotransduction on cell metabolism was partly mediated by Hif-1α. In addition, 2D and 3D culture of N-cadherin mimetic hydrogel had similar functions on N-cadherin expression and chondrogenesis in MSCs. Our study is the first to reveal that metabolic remodeling induced hypertrophy during MSCs chondrogenesis, and indicate the effect of N-cadherin mimetic hydrogel on hypertrophy inhibition of MSCs. STATEMENT OF SIGNIFICANCE: The development of hypertrophy during MSCs chondrogenesis severely limits its clinical translation. Various strategies have been explored to inhibit hypertrophy by chemical and/or mechanical stimulation. However, the role of cell metabolism in MSCs chondrogenesis has rarely been studied. In this study, we developed an RNA sequencing at day 0, 7, and 21 of MSCs chondrogenesis to clarify the mechanisms that mediate hypertrophy. We found that hypertrophy occurred in the late stage of MSCs chondrogenesis with increased FAO and decreased glycolysis, as well as impaired cell-cell junctions. We also found that N-cadherin mimetic hydrogel attenuated hypertrophy and enhanced chondrogenesis through regulating glycolysis and FAO. Our finding provides new insights into the application of MSCs in tissue engineering and regenerative medicine.

Spontaneous water-on-water spreading of polyelectrolyte membranes inspired by skin formation.

Stable interfaces between immiscible solvents are crucial for chemical synthesis and assembly, but interfaces between miscible solvents have been less explored. Here the authors report the spontaneous water-on-water spreading and self-assembly of polyelectrolyte membranes. An aqueous mixture solution containing poly(ethyleneimine) and poly(sodium 4-styrenesulfonate) spreads efficiently on acidic water, leading to the formation of hierarchically porous membranes. The reduced surface tension of the polyelectrolyte mixture solution drives the surface spreading, while the interfacial polyelectrolytes complexation triggered by the low pH of water mitigates water-in-water mixing. The synergy of surface tension and pH-dependent complexation represents a generic mechanism governing interfaces between miscible solvents for materials engineering, without the need for surfactants or sophisticated equipment. As a proof-of-concept, porous polyelectrolyte hybrid membranes are prepared by surface spreading, exhibiting exceptional solar thermal evaporation performance (2.8 kg/m2h) under 1-sun irradiation.

Small extracellular vesicles with nanomorphology memory promote osteogenesis.

Nanotopographical cues endow biomaterials the ability to guide cell adhesion, proliferation, and differentiation. Cellular mechanical memory can maintain the cell status by retaining cellular information obtained from past mechanical microenvironments. Here, we propose a new concept "morphology memory of small extracellular vesicles (sEV)" for bone regeneration. We performed nanotopography on titanium plates through alkali and heat (Ti8) treatment to promote human mesenchymal stem cell (hMSC) differentiation. Next, we extracted the sEVs from the hMSC, which were cultured on the nanotopographical Ti plates for 21 days (Ti8-21-sEV). We demonstrated that Ti8-21-sEV had superior pro-osteogenesis ability in vitro and in vivo. RNA sequencing further confirmed that Ti8-21-sEV promote bone regeneration through osteogenic-related pathways, including the PI3K-AKT signaling pathway, MAPK signaling pathway, focal adhesion, and extracellular matrix-receptor interaction. Finally, we decorated the Ti8-21-sEV on a 3D printed porous polyetheretherketone scaffold. The femoral condyle defect model of rabbits was used to demonstrate that Ti8-21-sEV had the best bone ingrowth. In summary, our study demonstrated that the Ti8-21-sEV have memory function by copying the pro-osteogenesis information from the nanotopography. We expect that our study will encourage the discovery of other sEV with morphology memory for tissue regeneration.

m6A hypomethylation of DNMT3B regulated by ALKBH5 promotes intervertebral disc degeneration via E4F1 deficiency.

BACKGROUND: The intervertebral disc (IVD) degeneration is the leading cause of low back pain, which accounts for a main cause of disability. N6-methyladenosine (m6A) is the most abundant internal modification in eukaryotic messenger RNAs and is involved in various diseases and cellular processes by modulating mRNA fate. However, the critical role of m6A regulation in IVD degeneration remains unclear. Nucleus pulposus cell (NPC) senescence is critical for the progression of IVD degeneration. Here, we uncovered the role and explored the regulatory mechanism of m6A in NPC senescence during IVD degeneration. METHODS: Identification of NPC senescence during IVD degeneration was based on the analysis of tissue samples and the cellular model. ALKBH5 upregulation inducing cellular senescence was confirmed by functional experiments in vivo and in vitro. ChIP-qPCR and DNA-Pulldown were used to reveal increased ALKBH5 was regulated by KDM4A-mediated H3K9me3. Furthermore, Me-RIP-seq was performed to identify m6A hypomethylation of DNMT3B transcripts in senescent NPCs. Stability analysis showed that DNMT3B expression was enhanced for less YTHDF2 recognition and increased DNMT3B promoted NPC senescence and IVD degeneration via E4F1 methylation by in vivo and in vitro analyses. RESULTS: Expression of ALKBH5 is enhanced during IVD degeneration and NPC senescence, due to decreased KDM4A-mediated H3K9me3 modification. Functionally, ALKBH5 causes NPC senescence by demethylating DNMT3B transcripts and in turn promoting its expression via less YTHDF2 recognition and following degradation due to transcript hypomethylation in vitro and in vivo. Increased DNMT3B promotes the development of IVD degeneration and NPC senescence, mechanistically by methylating CpG islands of E4F1 at the promoter region and thus restraining its transcription and expression. CONCLUSIONS: Collectively, our findings reveal an epigenetic interplay mechanism in NPC senescence and IVD degeneration, presenting a critical pro-senescence role of ALKBH5 and m6A hypomethylation, highlighting the therapeutic potential of targeting the m6A/DNMT3B/E4F1 axis for treating IVD degeneration.

Nanotopography Sequentially Mediates Human Mesenchymal Stem Cell-Derived Small Extracellular Vesicles for Enhancing Osteogenesis.

Engineered small extracellular vesicles (sEVs) are used as tools to enhance therapeutic efficacy. However, such application of sEVs is associated with several issues, including high costs and a high risk of tumorigenesis. Nanotopography has a greater influence on bone-related cell behaviors. However, whether nanotopography specifically mediate sEV content to perform particular biological functions remains unclear. Here, we demonstrate that selective nanotopography may be used to sequentially mediate human bone mesenchymal stem cell (hBMSC) sEVs to enhance the therapeutic efficacy of hBMSCs-EVs for osteogenesis. We subjected sEVs harvested from hBMSCs cultured on polished titanium plates (Ti) or nanotopographical titanium plates (Ti4) after 7, 14, and 21 d for RNA sequencing, and we found that there was no significant difference in sEV-miRNA expression after 7 d. Differentially expressed osteogenic-related microRNAs were founded after 14 days, and KEGG analysis indicated that the main microRNAs were associated with osteogenesis-related pathways, such as TGF-beta, AMPK, and FoxO. A significant difference was found in sEV-miRNAs expression after 21 d. We loaded sEV secreted from hBMSCs cultured on Ti4 after 21 d on 3D-printed porous PEEK scaffolds with poly dopamine (PDA) and found that such scaffolds showed superior osteogenic ability after 6- and 12-weeks. Here, we demonstrate the alkali- and heat-treated nanotopography with the ability of stimulating osteogenic differentiation of hBMSC can induce the secretion of pro-osteogenesis sEV, and we also found that sEVs meditate osteogenesis through miRNA. Thus, whether nanotopography has the ability to regulate other contents of sEVs such as proteins for enhancing osteogenesis needs further research. These findings may help us use nanotopography to extract sEVs for other biomedical applications, including cancer therapy.

Biomechanical Evaluation of Different Surgical Approaches for the Treatment of Adjacent Segment Diseases After Primary Anterior Cervical Discectomy and Fusion: A Finite Element Analysis.

Symptomatic adjacent segment disease (ASD) is a common challenge after anterior cervical discectomy and fusion (ACDF). The objective of this study was to compare the biomechanical effects of a second ACDF and laminoplasty for the treatment of ASD after primary ACDF. We developed a finite element (FE) model of the C2-T1 based on computed tomography images. The FE models of revision surgeries of ACDF and laminoplasty were simulated to treat one-level and two-level ASD after primary ACDF. The range of motion (ROM) and intradiscal pressure (IDP) of the adjacent segments, and stress in the cord were analyzed to investigate the biomechanical effects of the second ACDF and laminoplasty. The results indicated that revision surgery of one-level ACDF increased the ROM and IDP at the C2-C3 segment, whereas two-level ACDF significantly increased the ROM and IDP at the C2-C3 and C7-T1 segments. Furthermore, no significant changes in the ROM and IDP of the laminoplasty models were observed. The stress in the cord of the re-laminoplasty model decreased to some extent, which was higher than that of the re-ACDF model. In conclusion, both ACDF and laminoplasty can relieve the high level of stress in the spinal cord caused by ASD after primary ACDF, whereas ACDF can achieve better decompression effect. Revision surgery of the superior ACDF or the superior and inferior ACDF after the primary ACDF increased the ROM and IDP at the adjacent segments, which may be the reason for the high incidence of recurrent ASD after second ACDF.

Autophagic Degradation of Gasdermin D Protects against Nucleus Pulposus Cell Pyroptosis and Retards Intervertebral Disc Degeneration In Vivo.

Intervertebral disc degeneration (IDD) is the primary culprit of low back pain and renders heavy social burden worldwide. Pyroptosis is a newly discovered form of programmed cell death, which is also involved in nucleus pulposus (NP) cells during IDD progression. Moderate autophagy activity is critical for NP cell survival, but its relationship with pyroptosis remains unknown. This study is aimed at investigating the relationship between autophagy and pyroptotic cell death. The pyroptosis executor N-terminal domain of gasdermin D (GSDMD-N) and inflammation-related proteins were measured in lipopolysaccharide- (LPS-) treated human NP cells. Inhibition of autophagy by siRNA transfection and chemical drugs aggravated human NP cell pyroptosis. Importantly, we found that the autophagy-lysosome pathway and not the proteasome pathway mediated the degradation of GSDMD-N as lysosome dysfunction promoted the accumulation of cytoplasmic GSDMD-N. Besides, P62/SQSTM1 colocalized with GSDMD-N and mediated its degradation. The administration of the caspase-1 inhibitor VX-765 could reduce cell pyroptosis as confirmed in a rat disc IDD model in vivo, whereas ATG5 knockdown significantly accelerated the progression of IDD. In conclusion, our study indicated that autophagy protects against LPS-induced human NP cell pyroptosis via a P62/SQSTM1-mediated degradation mechanism and the inhibition of pyroptosis retards IDD progression in vivo. These findings deepen the understanding of IDD pathogenesis and hold implications in unraveling therapeutic targets for IDD treatment.

A Superhydrophobic Droplet-Based Magnetoelectric Hybrid System to Generate Electricity and Collect Water Simultaneously.

Traditional electromagnetic generators used in hydraulic power generation are heavy, bulky, and immovable, and are thus unsuitable for low water supply. A portable miniature electromagnetic system that can harvest energy from rainwater is critical for developing a sustainable energy strategy. In this study, a superhydrophobic droplet-based magnetoelectric hybrid system is fabricated, that can generate electricity from tiny water droplets. The magnetoelectric system (MS) comprises three parts: a superhydrophobic surface containing a conductive coil, liquid droplets, and a superhydrophobic magnetic powders/Ecoflex base. The mechanical impact of a falling water droplet onto the assembled system is transformed into electricity. Maxwell numerical simulation is used to analyze the related mechanism; the magnetoelectric performance is further improved by modifying the process parameters such as droplet falling velocity and magnetic powder contents. Furthermore, a model is developed, comprising the MS and a cactus-like superhydrophobic patterned plate that can generate electricity and collect water from fog, simultaneously. The described magnetoelectric strategy is believed to enhance and extend functions in energy harvesting and provide a generalized method to exploit new systems toward sustainable energy development.

Artificial bone scaffolds of coral imitation prepared by selective laser sintering.

Coralline hydroxyapatite (CHA) has been used in clinical for over 20 years. However, coral is an endanger species and has been banned from mining. In addition, coral artificial bone has slow biodegradation of the defects, hindering the growth of new bone. In order to explore the natural coral artificial bone substitute materials, this work proposed using Selective Laser Sintering (SLS) to fabricate natural calcium carbonate/biopolymer composite imitation coral porous structures, and then the surface of the 3D printing product was transformed into a hydroxyapatite thin layer by hydrothermal conversion reaction. The mechanical properties and porosity were optimized by adjusting the SLS processing parameters including laser power, scanning speed and layer thickness. In the composites with the PLLA of 15 wt%, the SLS processing parameters with the laser power of 15 W, laser scanning speed of 1500 mm/s and single layer thickness of 0.08 mm result in the better mechanical properties. After hydrothermal conversion, the products were confirmed to be a mixture of hydroxyapatite (HA) and calcium carbonate by X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR) and energy dispersive X-ray spectroscopy (EDX). The TGA results revealed that increasing the reaction temperature or prolonging the reaction time can increase the degree of hydrothermal reaction and thus promote the transformation of calcium carbonate into hydroxyapatite. The results of cytotoxicity assay and Life/Dead staining showed that the scaffold is not toxic to L929 cells. This work has the materials system innovation and focuses on the study of the effects of the SLS and hydrothermal processes on the mechanical performance and the degree of hydroxylation. Then, the preparation process of imitation coral artificial bone preparation was optimized. it is concluded that the imitation coral artificial bone is a nontoxic biomaterial; however, further study on its osteogenic capacity should be warranted in the future.

Three-dimensional biofabrication of an aragonite-enriched self-hardening bone graft substitute and assessment of its osteogenicity in vitro and in vivo.

A self-hardening three-dimensional (3D)-porous composite bone graft consisting of 65 wt% hydroxyapatite (HA) and 35 wt% aragonite was fabricated using a 3D-Bioplotter®. New tetracalcium phosphate and dicalcium phosphate anhydrous/aragonite/gelatine paste formulae were developed to overcome the phase separation of the liquid and solid components. The mechanical properties, porosity, height and width stability of the end products were optimised through a systematic analysis of the fabrication processing parameters including printing pressure, printing speed and distance between strands. The resulting 3D-printed bone graft was confirmed to be a mixture of HA and aragonite by X-ray diffraction, Fourier transform infrared spectroscopy and energy dispersive X-ray spectroscopy. The compression strength of HA/aragonite was between 0.56 and 2.49 MPa. Cytotoxicity was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in vitro. The osteogenicity of HA/aragonite was evaluated in vitro by alkaline phosphatase assay using human umbilical cord matrix mesenchymal stem cells, and in vivo by juxtapositional implantation between the tibia and the anterior tibialis muscle in rats. The results showed that the scaffold was not toxic and supported osteogenic differentiation in vitro. HA/aragonite stimulated new bone formation that bridged host bone and intramuscular implants in vivo. We conclude that HA/aragonite is a biodegradable and conductive bone formation biomaterial that stimulates bone regeneration. Since this material is formed near 37°C, it will have great potential for incorporating bioactive molecules to suit personalised application; however, further study of its biodegradation and osteogenic capacity is warranted. The study was approved by the Animal Ethical Committee at Tongji Medical School, Huazhong University of Science and Technology (IACUC No. 738) on October 1, 2017.

A novel method based on selective laser sintering for preparing high-performance carbon fibres/polyamide12/epoxy ternary composites.

A novel method based on selective laser sintering (SLS) process is proposed for the first time to prepare complex and high-performance carbon fibres/polyamide12/epoxy (CF/PA12/EP) ternary composites. The procedures are briefly described as follows: prepare polyamide12 (PA12) coated carbon fibre (CF) composite powder; build porous green parts by SLS; infiltrate the green parts with high-performance thermosetting epoxy (EP) resin; and finally cure the resin at high temperature. The obtained composites are a ternary composite system consisting of the matrix of novolac EP resin, the reinforcement of CFs and the transition thin layer of PA12 with a thickness of 595 nm. The SEM images and micro-CT analysis prove that the ternary system is a three-dimensional co-continuous structure and the reinforcement of CFs are well dispersed in the matrix of EP with the volume fraction of 31%. Mechanical tests show that the composites fabricated by this method yield an ultimate tensile strength of 101.03 MPa and a flexural strength of 153.43 MPa, which are higher than those of most of the previously reported SLS materials. Therefore, the process proposed in this paper shows great potential for manufacturing complex, lightweight and high-performance CF reinforced composite components in aerospace, automotive industries and other areas.