Cartilage-like Tissue Research Articles

Bioprinting and regeneration of auricular cartilage using a bioactive bioink based on microporous photocrosslinkable acellular cartilage matrix

Tissue engineering provides a promising strategy for auricular reconstruction. Although the first international clinical breakthrough of tissue-engineered auricular reconstruction has been realized based on polymer scaffolds, this approach has not been recognized as a clinically available treatment because of its unsatisfactory clinical efficacy. This is mainly since reconstruction constructs easily cause inflammation and deformation. In this study, we present a novel strategy for the development of biological auricle equivalents with precise shapes, low immunogenicity, and excellent mechanics using auricular chondrocytes and a bioactive bioink based on biomimetic microporous methacrylate-modified acellular cartilage matrix (ACMMA) with the assistance of gelatin methacrylate (GelMA), poly(ethylene oxide) (PEO), and polycaprolactone (PCL) by integrating multi-nozzle bioprinting technology. Photocrosslinkable ACMMA is used to emulate the intricacy of the cartilage-specific microenvironment for active cellular behavior, while GelMA, PEO, and PCL are used to balance printability and physical properties for precise structural stability, form the microporous structure for unhindered nutrient exchange, and provide mechanical support for higher shape fidelity, respectively. Finally, mature auricular cartilage-like tissues with high morphological fidelity, excellent elasticity, abundant cartilage lacunae, and cartilage-specific ECM deposition are successfully regenerated in vivo, which provides new opportunities and novel strategies for the fabrication and regeneration of patient-specific auricular cartilage.

Jammed Micro-Flake Hydrogel for Four-Dimensional Living Cell Bioprinting.

4D bioprinting is promising to build cell-laden constructs (bioconstructs) with complex geometries and functions for tissue/organ regeneration applications. The development of hydrogel-based 4D bioinks, especially those allowing living cell printing, with easy preparation, defined composition, and controlled physical properties is critically important for 4D bioprinting. Here, a single-component jammed micro-flake hydrogel (MFH) system with heterogeneous size distribution, which differs from the conventional granular microgel, has been developed as a new cell-laden bioink for 4Dbioprinting. This jammed cytocompatible MFH features scalable production and straightforward composition with shear-thinning, shear-yielding, and rapid self-healing properties. As such, it can be smoothly printed into stable 3Dbioconstructs, which can be further cross-linked to form a gradient in cross-linking density when a photoinitiator and a UV absorber are incorporated. After being subject to shape morphing, a variety of complex bioconstructs with well-defined configurations and high cell viability are obtained. Based on this system, 4Dcartilage-like tissue formation is demonstrated as aproof-of-concept. The establishment of this versatile new 4D bioink system may open up anumber of applications in tissue engineering.

Inhibition of BMP signaling with LDN 193189 can influence bone marrow stromal cell fate but does not prevent hypertrophy during chondrogenesis.

SummaryBone morphogenetic protein (BMP) cascades are upregulated during bone marrow-derived stromal cell (BMSC) chondrogenesis, contributing to hypertrophy and preventing effective BMSC-mediated cartilage repair. Previous work demonstrated that a proprietary BMP inhibitor prevented BMSC hypertrophy, yielding stable cartilage tissue. Because of the significant therapeutic potential of a molecule capable of hypertrophy blockade, we evaluated the capacity of a commercially available BMP type I receptor inhibitor with similar properties, LDN 193189, to prevent BMSC hypertrophy. Using 14-day microtissue chondrogenic induction cultures we found that LDN 193189 permitted BMSC chondrogenesis but did not prevent hypertrophy. LDN 193189 was sufficiently potent to counter mineralization and adipogenesis in response to exogenous BMP-2 in osteogenic induction cultures. LDN 193189 did not modify BMSC behavior in adipogenic induction cultures. Although LDN 193189 is effective in countering BMP signaling in a manner that influences BMSC fate, this blockade is insufficient to prevent hypertrophy.

The Evolution of Biomineralization through the Co-Option of Organic Scaffold Forming Networks.

Biomineralization is the process in which organisms use minerals to generate hard structures like teeth, skeletons and shells. Biomineralization is proposed to have evolved independently in different phyla through the co-option of pre-existing developmental programs. Comparing the gene regulatory networks (GRNs) that drive biomineralization in different species could illuminate the molecular evolution of biomineralization. Skeletogenesis in the sea urchin embryo was extensively studied and the underlying GRN shows high conservation within echinoderms, larval and adult skeletogenesis. The organic scaffold in which the calcite skeletal elements form in echinoderms is a tubular compartment generated by the syncytial skeletogenic cells. This is strictly different than the organic cartilaginous scaffold that vertebrates mineralize with hydroxyapatite to make their bones. Here I compare the GRNs that drive biomineralization and tubulogenesis in echinoderms and in vertebrates. The GRN that drives skeletogenesis in the sea urchin embryo shows little similarity to the GRN that drives bone formation and high resemblance to the GRN that drives vertebrates’ vascular tubulogenesis. On the other hand, vertebrates’ bone-GRNs show high similarity to the GRNs that operate in the cells that generate the cartilage-like tissues of basal chordate and invertebrates that do not produce mineralized tissue. These comparisons suggest that biomineralization in deuterostomes evolved through the phylum specific co-option of GRNs that control distinct organic scaffolds to mineralization.

Chondroitin sulfate cross-linked three-dimensional tailored electrospun scaffolds for cartilage regeneration

Degenerated cartilage tissues remain a burgeoning issue to be tackled, while bioactive engineering products available for optimal cartilage regeneration are scarce. In the present study, two-dimensional (2DS) poly(l-lactide-co-ε-caprolactone)/silk fibroin (PLCL/SF)-based scaffolds were fabricated by conjugate electrospinning method, which were then cross-linked with chondroitin sulfate (CS) to further enhance their mechanical and biological performance. Afterwards, three-dimensional (3D) PLCL/SF scaffolds (3DS) and CS-crosslinked 3D scaffolds (3DCSS) with tailored size were successfully fabricated by an in-situ gas foaming in a confined mold followed by freeze-dried. Gas-foamed scaffolds displayed high porosity, rapid water uptake, and stable mechanical properties. While all of the scaffolds exhibited good cytocompatibility in vitro; 3DCSS showed better cell seeding efficiency and chondro-protective effect compared to other scaffolds. Besides, 3DCSS scaffolds supported the formation of more mature cartilage-like tissues along with the best repair outcome in a rabbit articular cartilage defect model in vivo, as well as less expression level of pro-inflammatory cytokines, including interleukin (IL)-1β and tumor necrosis factor (TNF)-α than that of the other groups. Taken together, 3DCSS may provide an alternative therapeutic option for cartilage tissue repair.

Cartilage Repair Using Clematis Triterpenoid Saponin Delivery Microcarrier, Cultured in a Microgravity Bioreactor Prior to Application in Rabbit Model.

Cartilage tissue engineering provides a promising method for the repair of articular cartilage defects, requiring appropriate biological scaffolds and necessary growth factors to enhance the efficiency of cartilage regeneration. Here, a silk fibroin (SF) microcarrier and a clematis triterpenoid saponin delivery SF (CTS-SF) microcarrier were prepared by the high-voltage electrostatic differentiation and lyophilization method, and chondrocytes were carried under the simulated microgravity condition by a rotating cell culture system. SF and CTS-SF microspheres were relatively uniform in size and had a porous structure with good swelling and cytocompatibility. Further, CTS-SF microcarriers could sustainably release CTSs in the monitored 10 days. Compared with the monolayer culture, chondrocytes under the microgravity condition maintained a better chondrogenic phenotype and showed better proliferation ability after culture on microcarriers. Moreover, the sustained release of CTS from CTS-SF microcarriers upregulated transforming growth factor-β, Smad2, and Smad3 signals, contributing to promote chondrogenesis. Hence, the biophysical effects of microgravity and bioactivities of CTS-ST were used for chondrocyte expansion and phenotype maintenance in vitro. With prolonged expansion, SF- and CTS-SF-based microcarrier-cell composites were directly implanted in vivo to repair rabbit articular defects. Gross evaluations, histopathological examinations, and biochemical analysis indicated that SF- and CTS-SF-based composites exhibited cartilage-like tissue repair compared with the nontreated group. Further, CTS-SF-based composites displayed superior hyaline cartilage-like repair that integrated with the surrounding cartilage better and higher cartilage extracellular matrix content. In conclusion, these results provide an alternative preparation method for drug-delivered SF microcarrier and a culture method for maintaining the chondrogenic phenotype of seed cells based on the microgravity environment. CTS showed its bioactive function, and the application of CTS-SF microcarriers can help repair and regenerate cartilage defects.

An antibacterial ε-poly-L-lysine-derived bioink for 3D bioprinting applications.

The limited availability of bioinks has hindered the application of 3D bioprinting to tissue engineering, and bacterial infection is a serious threat to these applications. Aiming to solve this problem, a novel ε-poly-L-lysine (EPL)-derived antibacterial bioink has been developed for 3D bioprinting and tissue-engineering applications. Three glycidyl methacrylate (GMA)-modified EPL products, EPLGMA-1, EPLGMA-2, and EPLGMA-3, were prepared by reacting 3, 4, and 5 mL GMA with 5 g EPL, respectively. EPLGMA-1, EPLGMA-2, and EPLGMA-3 were photocurable and their corresponding photo-crosslinked hydrogels, EPLGMA-1H, EPLGMA-2H, and EPLGMA-3H, all exhibited very high antibacterial rates, good biocompatibility, good degradability, and promising mechanical properties. After EPLGMA-1H, EPLGMA-2H, and EPLGMA-3H with encapsulated chondrocytes were incubated for 4 weeks, EPLGMA-3H was the best one among them for tissue-engineering applications due to its most efficient tissue regeneration. Carrying chondrocytes, the EPLGMA-3 solution was printed into hydrogel products with high-fidelity shapes and high cell viability using a projection-based 3D bioprinter. Following the implantation of chondrocyte-loaded EPLGMA-3H samples into nude mice for 4 weeks, cartilage-like tissue was regenerated, suggesting that EPLGMA-3 is a promising antibacterial bioink for 3D bioprinting and tissue-engineering applications.

Kartogenin regulates hair growth and hair cycling transition.

Background: Kartogenin is a heterocyclic compound able to promote the proliferation, migration, and differentiation of various cell types and induce cartilage-like tissue regeneration. However, the role of kartogenin in hair follicles (HFs), remains unknown. We therefore investigated the effects of kartogenin on the regulation of hair growth and hair growth cycle transition.Methods: The effects of kartogenin on the proliferation, cell cycle status, and migration of primary human outer root sheath cells (ORSCs) were evaluated by MTS assay, flow cytometry, Transwell® and scratch assays, respectively. We exposed ORSCs to kartogenin (1 µM) and determined changes in mRNA and protein levels of transforming growth factor (TGF)-β2/Smad signaling molecules by reverse transcription polymerase chain reaction, western blotting, and immunofluorescence. We also examined the effects of kartogenin (10 µM) on HFs in mice by histology following cutaneous injection.Results: Kartogenin enhanced ORSC proliferation and migration function in a dose-dependent manner, and downregulated the expression of TGF-β2/Smad signaling molecules in vitro. Injection of kartogenin delayed catagen phase and increased regenerated hair length in mice in vivo.Conclusions: Kartogenin modulates HF growth and regulates the hair cycle and the TGF-β2/Smad signaling pathway, providing a potential new approach for the treatment of hair loss.

Intraarticular injection of SHP2 inhibitor SHP099 promotes the repair of rabbit full-thickness cartilage defect.

BackgroundCartilage repair has been a challenge in the field of orthopaedics for decades, highlighting the significance of investigating potential therapeutic drugs. In this study, we explored the effect of the SHP2 inhibitor SHP099, a small-molecule drug, on cartilage repair.MethodsHuman synovial mesenchymal stem cells (SMSCs) were isolated, and their three-way differentiation potential was examined. After treatment with chondrogenic medium, the chondrogenic effect of SHP099 on SMSCs was examined by western blot, qPCR, and immunofluorescence (IF). Micro-mass culture was also used to detect the effect of SHP099. To explore the chondrogenic effects of SHP099 in vivo, full-thickness cartilage defects with microfractures were constructed in the right femoral trochlea of New Zealand White rabbits. Intraarticular injection of SHP099 or normal saline was performed twice a week for 6 weeks. Cartilage repair was evaluated by haematoxylin and eosin (HE) staining and safranin O/fast green staining. Immunohistochemistry (IHC) for collagen II (COL2) was also conducted to verify the abundance of cartilage extracellular matrix after SHP099 treatment. The mechanism involving yes-associated protein (YAP) and WNT signalling was investigated in vitro.ResultsSMSCs isolated from human synovium have optimal multi-differentiation potential. SHP099 increased chondrogenic marker (SOX9, COL2) expression and decreased hypertrophic marker (COL10, RUNX2) expression in SMSCs. In micro-mass culture, the SHP099-induced cartilage tissues had a better result of Safranin O and Toluidine blue staining and are enriched in cartilage-specific collagen II. Inhibition of YAP and WNT signalling was also observed. Moreover, compared to the normal saline group at 6 weeks, intraarticular injection of SHP099 resulted in better defect filling, forming increased hyaline cartilage-like tissue with higher levels of glycosaminoglycan (GAG) and COL2.ConclusionSHP099 promotes the repair of rabbit full-thickness cartilage defects, representing a potential therapeutic drug for cartilage repair.The Translational potential of this articleThis study provides evidence that SHP2 inhibition promotes chondrogenesis and the repair of cartilage in defect area, which could be a novel therapeutic approach for cartilage repair.

Biomimetic Gradient Scaffolds Containing Hyaluronic Acid and Sr/Zn Folates for Osteochondral Tissue Engineering.

Regenerative therapies based on tissue engineering are becoming the most promising alternative for the treatment of osteoarthritis and rheumatoid arthritis. However, regeneration of full-thickness articular osteochondral defects that reproduces the complexity of native cartilage and osteochondral interface still remains challenging. Hence, in this work, we present the fabrication, physic-chemical characterization, and in vitro and in vivo evaluation of biomimetic hierarchical scaffolds that mimic both the spatial organization and composition of cartilage and the osteochondral interface. The scaffold is composed of a composite porous support obtained by cryopolymerization of poly(ethylene glycol) dimethacrylate (PEGDMA) in the presence of biodegradable poly(D,L-lactide-co-glycolide) (PLGA), bioactive tricalcium phosphate β-TCP and the bone promoting strontium folate (SrFO), with a gradient biomimetic photo-polymerized methacrylated hyaluronic acid (HAMA) based hydrogel containing the bioactive zinc folic acid derivative (ZnFO). Microscopical analysis of hierarchical scaffolds showed an open interconnected porous open microstructure and the in vitro behaviour results indicated high swelling capacity with a sustained degradation rate. In vitro release studies during 3 weeks indicated the sustained leaching of bioactive compounds, i.e., Sr2+, Zn2+ and folic acid, within a biologically active range without negative effects on human osteoblast cells (hOBs) and human articular cartilage cells (hACs) cultures. In vitro co-cultures of hOBs and hACs revealed guided cell colonization and proliferation according to the matrix microstructure and composition. In vivo rabbit-condyle experiments in a critical-sized defect model showed the ability of the biomimetic scaffold to promote the regeneration of cartilage-like tissue over the scaffold and neoformation of osteochondral tissue.

Anti-Inflammatory and Prochondrogenic In Situ-Formed Injectable Hydrogel Crosslinked by Strontium-Doped Bioglass for Cartilage Regeneration.

Directed differentiation of bone marrow mesenchymal stem cells (BMSCs) toward chondrogenesis plays a predominant role in cartilage repair. However, the uncontrolled inflammatory response to implants is found to impair the stability of scaffolds and the cartilage regeneration outcome. Herein, we fabricated an injectable hydrogel crosslinked by strontium-doped bioglass (SrBG) to modulate both human BMSC (hBMSC) differentiation and the inflammatory response. The results revealed that the introduction of Sr ions could simultaneously enhance the proliferation of hBMSCs, upregulate cartilage-specific gene expression, and improve the secretion of glycosaminoglycan. Moreover, after cultured with SA/SrBG extracts in vitro, a majority of macrophages were polarized toward the M2 phenotype and subsequently facilitated the chondrogenic differentiation of hBMSCs. Furthermore, after the composite hydrogel was injected into a cartilage defect model, neonatal cartilage-like tissues with a smooth surface and tight integration with original tissues could be found. This study suggests that the synergistic strategy based on an enhanced differentiation ability and a regulated inflammatory response is promising and may lead the way to new anti-inflammatory biomaterials.

A photo-crosslinked proteinogenic hydrogel enabling self-recruitment of endogenous TGF-β1 for cartilage regeneration

Cartilage regeneration is crucial to the functional reconstruction of damaged or degenerated joints, involving a variety of biological factors. Transforming growth factor-beta 1 (TGF-β1) play an important role in chondrogenesis and cartilage regeneration. In this study, we developed a novel photo-crosslinked Gelatin methacryloyl (GelMA) hydrogel modified with TGF-β1-affinity peptide (HSNGLPL), which can self-absorb with TGF-β1, and investigated the effect of GelMA/HSNGLPL (Pep-HGelMA) hydrogels on chondrogenesis and cartilage regeneration. In vitro, Pep-HGelMA hydrogel scaffolds were confirmed to recruit endogenous TGF-β1, and induced mesenchymal stem cells into chondrocytes. In vivo, more cambium and smooth surface was grossly observed at the injured sites of knee joint implanted with Pep-HGelMA hydrogels than those of other groups four and eight weeks post-operation. Micro-CT and histological evaluation further demonstrated more cartilage tissue and higher scores in Pep-HGelMA-implanted knees. In addition, more cartilage-like tissue indicators including collagen II and less matrix metalloproteinase 13 were also observed. All the data suggests that the photo-crosslinked hydrogel scaffold characterized by excellent mechanical properties and biocompatibility, can accelerate cartilage tissue repair by recruiting TGF-β1 automatically, which provides a promising system for cartilage repair in clinical application.

Icariin accelerates cartilage defect repair by promoting chondrogenic differentiation of BMSCs under conditions of oxygen-glucose deprivation.

This study explored the role played by combined ICA and bone mesenchymal stem cells (BMSCs) in repairing rabbit knee cartilage defects. Firstly, rabbit BMSCs were isolated and used to construct an in vitro cellular model of oxygen‐glucose deprivation/reoxygenation (OGD/R). Subsequently, ICA processing, Alcian blue staining, immunofluorescence and Western blot studies were performed to evaluate the ability of BMSCs to display signs of chondrogenic differentiation. Furthermore, a rabbit knee cartilage injury model was established in vivo. International Cartilage Repair Society (ICRS) macroscopic evaluations, H&E, Alcian blue and EdU staining, as well as immunohistochemistry, were analysed cartilage repair and pathological condition of the knee cartilage tissue. Our in vitro results showed that ICA promoted the chondrogenic differentiation of BMSCs, as well as aggrecan (AGR), bone morphogenetic protein 2 (BMP2) and COL2A1 protein expression in BMSCs. In vivo experiments showed that rabbits in the BMSCs or ICA treatment group had higher ICRS scores and displayed a better restoration of cartilage‐like tissue and chondrocyte expression on the surface of their cartilage defects. In conclusion, ICA or BMSCs alone could repair rabbit knee cartilage damage, and combined treatment with ICA and BMSCs showed a better ability to repair rabbit knee cartilage damage.

Converging 3D Printing and Electrospinning: Effect of Poly(l-lactide)/Gelatin Based Short Nanofibers Aerogels on Tracheal Regeneration.

Recently, various tissue engineering based strategies have been pursued for the regeneration of tracheal tissues. However, previously developed tracheal scaffolds do not accurately mimic the microstructure and mechanical behavior of the native trachea, which restrict their clinical translation. Here, tracheal scaffolds are fabricated by using 3D printing and short nanofibers (SF) dispersion of poly(l-lactide)/gelatin (0.5-1.5 wt%) to afford tracheal constructs. The results display that the scaffolds containing 1.0wt % of SF exhibit low density, good water absorption capacity, reasonable degradation rate, and stable mechanical properties, which were comparable to the native trachea. Moreover, the designed scaffolds possess good biocompatibility and promote the growth and infiltration of chondrocytes in vitro. The biocompatibility of tracheal scaffolds is further assessed after subcutaneous implantation in mice for up to 4 and 8 weeks. Histological assessment of tracheal constructs explanted at week 4 shows that scaffolds can maintain their structural integrity and support the formation of neo-vessels. Furthermore, cell-scaffold constructs gradually form cartilage-like tissues, which mature with time. Collectively, these engineered tracheal scaffolds not only possess appropriate mechanical properties to afford a stabilized structure but also a biomimetic extracellular matrix-like structure to accomplish tissue regeneration, which may have broad implications for tracheal regeneration.

Hierarchical porous bacterial cellulose scaffolds with natural biomimetic nanofibrous structure and a cartilage tissue-specific microenvironment for cartilage regeneration and repair

The limited three-dimensional (3D) nano-scale pore structure and lack of biological function hamper the application of bacterial cellulose (BC) in cartilage tissue engineering. To address this challenge, 3D hierarchical porous BC/decellularized cartilage extracellular matrix (DCECM) scaffolds with structurally and biochemically biomimetic cartilage regeneration microenvironment were fabricated by freeze-drying technique after EDC/NHS chemical crosslinking. The BC/DCECM scaffolds exhibited excellent mechanical properties, water superabsorbency and shape-memory properties. Compared with the BC control, the BC/DCECM scaffolds exhibited enhanced cell adhesion and proliferation. Cartilage regeneration in vitro and in vivo indicated that the BC/DCECM scaffolds achieved satisfactory neocartilage tissue regeneration with superior original shape fidelity, exterior natural cartilage-like appearance and histologically cartilage-specific lacuna formation and ECM deposition. Furthermore, the BC/DCECM scaffolds achieved superior repair outcomes, as hyaline cartilage-like tissue formed within the defect sites. The present study constitutes a strong step toward the further application of BC in cartilage tissue engineering.

3D-Printed Reinforcement Scaffolds with Targeted Biodegradation Properties for the Tissue Engineering of Articular Cartilage.

Achieving regeneration of articular cartilage is challenging due to the low healing capacity of the tissue. Appropriate selection of cell source, hydrogel, and scaffold materials are critical to obtain good integration and long-term stability of implants in native tissues. Specifically, biomechanical stability and in vivo integration can be improved if the rate of degradation of the scaffold material matches the stiffening of the sample by extracellular matrix secretion of the encapsulated cells. To this end, a novel 3D-printed lactide copolymer is presented as a reinforcement scaffold for an enzymatically crosslinked hyaluronic acid hydrogel. In this system, the biodegradable properties of the reinforced scaffold are matched to the matrix deposition of articular chondrocytes embedded in the hydrogel. The lactide reinforcement provides stability to the soft hydrogel in the early stages, allowing the composite to be directly implanted in vivo with no need for a preculture period. Compared to pure cellular hydrogels, maturation and matrix secretion remain unaffected by the reinforced scaffold. Furthermore, excellent biocompatibility and production of glycosaminoglycans and collagens are observed at all timepoints. Finally, in vivo subcutaneous implantation in nude mice shows cartilage-like tissue maturation, indicating the possibility for the use of these composite materials in one-step surgical procedures.

Controlled Differentiation of Mesenchymal Stem Cells into Hyaline Cartilage in miR-140-Activated Collagen Hydrogel.

Hypertrophic cartilage formation is a major setback in mesenchymal stem cells (MSCs)-mediated cartilage repair, and overcoming it requires optimization of differentiation. Here, we tested the miR-140 activated collagen hydrogel for the chondrogenic differentiation of MSCs and to produce hyaline cartilage. Bone marrow MSCs isolated from 3 patients were pretreated with miR-140 and then chondrogenic differentiated. The 3-dimensional (3D) transfection potential of 5 different transfection reagents (Polyethylenimine, Lipofectamine, TransIT-X2, Amide:Cholesterol-based liposomes [AmC] and AmC pegylated with Tocofersolan [AmCTOC]) was compared and the reagent that showed higher green fluorescent protein (GFP) expression was selected. Finally, the collagen hydrogel was activated using miR-140-transfection complex and sustained delivered to MSCs during chondrogenic differentiation. After differentiation, the outcome was assessed by reverse transcription-polymerase chain reaction (RT-PCR), histology, immunohistochemistry, and compared with scrambled miRNA treated control. Pretreatment of MSCs with miR-140 significantly increased the expression of cartilage-specific genes (COL2A1, SOX9, and ACAN) with reduced hypertrophic chondrocyte (COL10A1) marker expression and better safranin-O staining than the control. The AmCTOC liposome showed a significant increase in 3D transfection of GFP expressing plasmid than the others. Furthermore, the knockdown of GAPDH using siRNA in HEK cells and expression of GFP mRNA in human bone marrow MSCs confirmed the 3D-transfection efficiency of AmCTOC. The sustained delivery of miR-140 using activated matrix formed a hyaline cartilage-like tissue with minimal COL10A1 expression in RT-PCR and immunohistochemistry. Our results demonstrated the therapeutic potential of miR-140-activated hydrogel for MSCs-based cartilage tissue engineering, which could also be used for endogenous stem cells-mediated cartilage repair.

Ex Vivo and In Vivo Properties of an Injectable Hydrogel Derived From Acellular Ear Cartilage Extracellular Matrix.

Extracellular matrix (ECM) hydrogels provide advantages such as injectability, the ability to fill an irregularly shaped space, and the adequate bioactivity of native matrix. In this study, we developed decellularized cartilage ECM (dcECM) hydrogels from porcine ears innovatively via the main method of enzymatic digestion and verified good biocompatible properties of dcECM hydrogels to deliver chondrocytes and form subcutaneous cartilage in vivo. The scanning electron microscopy and turbidimetric gelation kinetics were used to characterize the material properties and gelation kinetics of the dcECM hydrogels. Then we evaluated the biocompatibility of hydrogels via the culture of chondrocytes in vitro. To further explore the dcECM hydrogels in vivo, grafts made from the mixture of dcECM hydrogels and chondrocytes were injected subcutaneously in nude mice for the gross and histological analysis. The structural and gelation kinetics of the dcECM hydrogels altered according to the variation in the ECM concentrations. The 10 mg/ml dcECM hydrogels could support the adhesion and proliferation of chondrocytes in vitro. In vivo, at 4 weeks after transplantation, cartilage-like tissues were detected in all groups with positive staining of toluidine blue, Safranin O, and collagen II, indicating the good gelation of dcECM hydrogels. While with the increasing concentration, the tissue engineering cartilages formed by 10 mg/ml dcECM hydrogel grafts were superior in weights, volumes, collagen, and glycosaminoglycan (GAG) content compared to the dcECM hydrogels of 1 mg/ml and 5 mg/ml. At 8 weeks after grafting, dcECM hydrogel grafts at 10 mg/ml showed very similar qualities to the control, collagen I grafts. After 12 weeks of in vivo culture, the histological analysis indicated that 10 mg/ml dcECM hydrogel grafts were similar to the normal cartilage from pig ears, which was the source tissue. In conclusion, dcECM hydrogel showed the promising potential as a tissue engineering biomaterial to improve the regeneration and heal injuries of ear cartilage.

RETRACTED: Gas-foaming three-dimensional electrospun nanofiber scaffold improved three-dimensional cartilage regeneration

Repairing cartilage defect is always an intractable problem in joint surgery field. Tissue engineering, in the industry, is universally considered as a decent solution for overcoming this challenge. Especially the three-dimensional (3D) scaffolds play a significant role in cartilage repair. Thereinto, the electrospinning has become a very attractive method for the preparation of scaffolds. In recent years. However, these scaffolds are limited in terms of their three-dimensional (3D) applications due to their two-dimensional (2D) structure and pore size which are smaller than a cartilage cellular diameter and thus limit the cellular migration in these structures. To address this issue, this study will present an promising post electrospinning approach that can transform two-dimensional scaffolds into three-dimensional scaffolds via the way of insitu gas foaming within the pores of the nanofiber membranes as the driving force. Our previous study reported that agelatin/polycaprolactone (GT:PCL) ratio of 7:3 might be suitable for the cartilage regeneration [Zheng R, et al The influence of Gelatin/PCL ratio and 3D construct shape of electrospun membranes on cartilage regeneration. Biomaterials 2014;35:152-164]. Therefore, in the present experiment, we chose the above ratio (GT:PCL = 7:3) to realize two types of scaffolds (2D and 3D scaffolds) transition via the gas-foaming technique and investigated whether the three-dimensional structure was more conducive to cartilage regeneration than 2D.The experiment results have revealed that 3D scaffolds can achieve a larger pore size, higher porosity and higher biocompatibility than 2D scaffolds. In addition, both scaffolds which were implanted with chondrocytes all had formed mature cartilage-like tissues after 8 weeks of culturing in rabbits, and the 3D scaffold formed a three-dimensional structure, whereas the 2D scaffold only formed a thin layer of cartilage. As the macroscopic and histological results showed after 12 weeks postoperation, in the 2D scaffold group, the defect was full of fibrillar connective tissue, and as shown by HE staining, obviously there is no staining with Saf-O/FG and toluidine blue on the surface of repaired site. On the contrary, in the 3D scaffold group, homogeneous and mature cartilaginous tissue were found in the defect area. The defect was filled with numerous new chondrocytes, and the histologicalstaining revealed a large amount of regenerated cartilage tissue which was perfectly integrated with normal cartilage tissue. The results distinctly indicated that the 3D scaffold led to better cartilage repair effects than the 2D scaffold. Generally speaking, the current study demonstrated that a gas-foaming three-dimensional electrospun nanofiber scaffold would be a potential platform for cartilage regeneration and might provide a potential treatment option for repairing articular cartilage defects.

A Comparison of the Capacity of Mesenchymal Stromal Cells for Cartilage Regeneration Depending on Collagen-Based Injectable Biomimetic Scaffold Type.

Mesenchymal stromal cells (MSCs) have shown a high potential for cartilage repair. Collagen-based scaffolds are used to deliver and retain cells at the site of cartilage damage. The aim of the work was a comparative analysis of the capacity of the MSCs from human adipose tissue to differentiate into chondrocytes in vitro and to stimulate the regeneration of articular cartilage in an experimental model of rabbit knee osteoarthrosis when cultured on microheterogenic collagen-based hydrogel (MCH) and the microparticles of decellularized porcine articular cartilage (DPC). The morphology of samples was evaluated using scanning electron microscopy and histological staining methods. On the surface of the DPC, the cells were distributed more uniformly than on the MCH surface. On day 28, the cells cultured on the DPC produced glycosaminoglycans more intensely compared to the MCH with the synthesis of collagen type II. However, in the experimental model of osteoarthrosis, the stimulation of the cartilage regeneration was more effective when the MSCs were administered to the MCH carrier. The present study demonstrates the way to regulate the action of the MSCs in the area of cartilage regeneration: the MCH is more conducive to stimulating cartilage repair by the MSCs, while the DPC is an inducer for a formation of a cartilage-like tissue by the MSCs in vitro.