Articular Cartilage Tissue Engineering Research Articles

Impact of interstitial flow on cartilage matrix synthesis and NF-kB transcription factor mRNA expression in a novel perfusion bioreactor.

This work is focused on designing an easy-to-use novel perfusion system for articular cartilage (AC) tissue engineering and using it to elucidate the mechanism by which interstitial shear upregulates matrix synthesis by articular chondrocytes (AChs). Porous chitosan-agarose (CHAG) scaffolds were synthesized and compared to bulk agarose (AG) scaffolds. Both scaffolds were seeded with osteoarthritic human AChs and cultured in a novel perfusion system with a medium flow velocity of 0.33 mm/s corresponding to 0.4 mPa surfice shear and 40 mPa CHAG interstitial shear. While there were no statistical differences in cell viability for perfusion versus static cultures for either scaffold type, CHAG scaffolds exhibited a 3.3-fold higher (p < 0.005) cell viability compared to AG scaffold cultures. Effects of combined superficial and interstitial perfusion for CHAG showed 150- and 45-fold (p < 0.0001) increases in total collagen (COL) and 13- and 2.2-fold (p < 0.001) increases in glycosaminoglycans (GAGs) over AG non-perfusion and perfusion cultures, respectively, and a 1.5-fold and 3.6-fold (p < 0.005) increase over non-perfusion CHAG cultures. Contrasting CHAG perfusion and static cultures, chondrogenic gene comparisons showed a 3.5-fold increase in collagen type II/type I (COL2A1/COL1A1) mRNA ratio (p < 0.05), and a 1.3-fold increase in aggrecan mRNA. Observed effects are linked to NF-κB signal transduction pathway inhibition as confirmed by a 3.2-fold (p < 0.05) reduction of NF-κB mRNA expression upon exposure to perfusion. Our results demonstrate that pores play a critical role in improving cell viability and that interstitial flow caused by medium perfusion through the porous scaffolds enhances the expression of chondrogenic genes and extracellular matrix through downregulating NF-κB1.

Magnetic hydrogel applications in articular cartilage tissue engineering.

Articular cartilage defects afflict millions of individuals worldwide, presenting a significant challenge due to the tissue's limited self-repair capability and anisotropic nature. Hydrogel-based biomaterials have emerged as promising candidates for scaffold production in artificial cartilage construction, owing to their water-rich composition, biocompatibility, and tunable properties. Nevertheless, conventional hydrogels typically lack the anisotropic structure inherent to natural cartilage, impeding their clinical and preclinical applications. Recent advancements in tissue engineering (TE) have introduced magnetically responsive hydrogels, a type of intelligent hydrogel that can be remotely controlled using an external magnetic field. These innovative materials offer a means to create the desired anisotropic architecture required for successful cartilage TE. In this review, we first explore conventional techniques employed for cartilage repair and subsequently delve into recent breakthroughs in the application and utilization of magnetic hydrogels across various aspects of articular cartilage TE.

Alginate-waterborne polyurethane 3D bioprinted scaffolds for articular cartilage tissue engineering

Articular cartilage defects comprise a spectrum of diseases associated with degeneration or damage of the connective tissue present in particular joints, presenting progressive osteoarthritis if left untreated. In vitro tissue regeneration is an innovative treatment for articular cartilage injuries that is attracting not only clinical attention, but also great interest in the development of novel biomaterials, since this procedure involves the formation of a neotissue with the help of material support. In this work, functional alginate and waterborne polyurethane (WBPU) scaffolds have been developed for articular cartilage regeneration using 3D bioprinting technology. The particular properties of alginate-WBPU blends, like mechanical strength, elasticity and moistening, mimic the original cartilage tissue characteristics, being ideal for this application. To fabricate the scaffolds, mature chondrocytes were loaded into different alginate-WBPU inks with rheological properties suitable for 3D bioprinting. Bioinks with high alginate content showed better 3D printing performance, as well as structural integrity and cell viability, being most suitable for scaffolds fabrication. After 28 days of in vitro cartilage formation experiments, scaffolds containing 3.2 and 6.4 % alginate resulted in the maintenance of cell number in the range of 104 chondrocytes/scaffold in differentiated phenotypes, capable of synthesizing specialized extracellular matrix (ECM) up to 6 μg of glycosaminoglycans (GAG) and thus, showing a potential application of these scaffolds for in vitro regeneration of articular cartilage tissue.

Recent progress on 3D-printed gelatin methacrylate-based biomaterials for articular cartilage repair

The structure and composition of articular cartilage is complex, and its self-healing ability is limited, and thus, it is difficult to achieve ideal healing once the articular cartilage is damaged. Recently, three-dimensional (3D) printing technology has provided a new possibility for the repair of articular cartilage. Engineered cartilage tissues can be fabricated by superimposing customized inks, considering different geometric structures and components of tissues. 3D printing can be effectively used to manufacture high-precision structures with complex geometry, solving the shortcomings of traditional scaffold fabrication techniques. Gelatin methacrylate (GelMA) is modified gelatin and is currently a widely used 3D printing ink due to its photocrosslinking properties. With good biocompatibility and tunable physical properties, it can provide a good scaffold platform for cell proliferation and growth factor release. Given that the role of 3D printing technology in cartilage repair has been widely reported, this article reviews the research progress of 3D-printed GelMA-based biomaterials in articular cartilage tissue engineering. We focus primarily on how 3D printing technology addresses the existing challenges inherent to the field of articular cartilage tissue engineering. We accentuate the modifications implemented in GelMA-based 3D printing scaffolds to optimize articular cartilage regeneration. Additionally, we provide a comprehensive summary of the utilization of GelMA-based biomaterials incorporating various cells, growth factors, or other tissue components and highlight how these adaptations, in conjunction with the benefits of 3D printing technology, facilitate improvements the articular cartilage repair.

Biophysical Electrical and Mechanical Stimulations for Promoting Chondrogenesis of Stem Cells on PEDOT:PSS Conductive Polymer Scaffolds.

The investigation of the effects of electrical and mechanical stimulations on chondrogenesis in tissue engineering scaffolds is essential for realizing successful cartilage repair and regeneration. The aim of articular cartilage tissue engineering is to enhance the function of damaged or diseased articular cartilage, which has limited regenerative capacity. Studies have shown that electrical stimulation (ES) promotes mesenchymal stem cell (MSC) chondrogenesis, while mechanical stimulation (MS) enhances the chondrogenic differentiation capacity of MSCs. Therefore, understanding the impact of these stimuli on chondrogenesis is crucial for researchers to develop more effective tissue engineering strategies for cartilage repair and regeneration. This study focuses on the preparation of poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate) (PEDOT:PSS) conductive polymer (CP) scaffolds using the freeze-drying method. The scaffolds were fabricated with varying concentrations (0, 1, 3, and 10 wt %) of (3-glycidyloxypropyl) trimethoxysilane (GOPS) as a crosslinker and an additive to tailor the scaffold properties. To gain a comprehensive understanding of the material characteristics and the phase aggregation phenomenon of PEDOT:PSS scaffolds, the researchers performed theoretical calculations of solubility parameters and surface energies of PSS, PSS-GOPS, and PEDOT polymers, as well as conducted material analyses. Additionally, the study investigated the potential of promoting chondrogenic differentiation of human adipose stem cells by applying external ES or MS on a PEDOT:PSS CP scaffold. Compared to the group without stimulation, the group that underwent stimulation exhibited significantly up-regulated expression levels of chondrogenic characteristic genes, such as SOX9 and COL2A1. Moreover, the immunofluorescence staining images exhibited a more vigorous fluorescence intensity of SOX9 and COL II proteins that was consistent with the trend of the gene expression results. In the MS experiment, the strain excitation exerted on the scaffold was simulated and transformed into stress. The simulated stress response showed that the peak gradually decreased with time and approached a constant value, with the negative value of stress representing the generation of tensile stress. This stress response quantification could aid researchers in determining specific MS conditions for various materials in tissue engineering, and the applied stress conditions could be further optimized. Overall, these findings are significant contributions to future research on cartilage repair and biophysical ES/MS in tissue engineering.

Engineered biomechanical microenvironment of articular chondrocytes based on heterogeneous GelMA hydrogel composites and dynamic mechanical compression

Tissue-engineered articular cartilage constructs are currently not able to equal native tissues in terms of mechanical and biological properties. A major cause lies in the deficiency in engineering the biomechanical microenvironment (BMME) of articular chondrocytes. In this work, to engineer the BMME of articular chondrocytes, heterogeneous hydrogel structures of gelatin methacrylated (GelMA) containing differential-stiffness domains were first fabricated, and then periodic dynamic mechanical stimulations were applied to the hydrogel structures. The chondrocyte phenotype of ATDC5 cells was enhanced as the spatial differentiation in stiffness was increased in the hydrogel structures and was further strengthened by dynamic mechanical stimulation. It was speculated that the mechanical signals generated by the engineered BMME were sensed by the cells through the integrin β1-FAK signaling pathway. This study revealed the key role of the combined effects of differential and dynamic BMME on the chondrocyte phenotype, which could provide theoretical guidance for highly active tissue-engineered articular cartilage.

A modular hydrogel bioink containing microsphere-embedded chondrocytes for 3D-printed multiscale composite scaffolds for cartilage repair

Articular cartilage tissue engineering is being considered an alternative treatment strategy for promoting cartilage damage repair. Herein, we proposed a modular hydrogel-based bioink containing microsphere-embedded chondrocytes for 3D printing multiscale scaffolds integrating the micro and macro environmentof the native articular cartilage. Gelatin methacryloyl (GelMA)/alginate microsphere was prepared by a microfluidic approach, and the chondrocytes embedded in the microspheres remained viable after being frozen and resuscitated. The modular hydrogel bioink could be printed via the gel-in-gel 3D bioprinting strategy for fabricating the multiscale hydrogel-based scaffolds. Meanwhile, the cells cultured in the scaffolds showed good proliferation and differentiation.Furthermore, we also found that the composite hydrogel was biocompatible invivo. These results indicated that the modular hydrogel-based bioinks containing microsphere-embedded chondrocytes for 3D printing multiscale scaffolds could provide a 3D multiscale environment for enhancing cartilage repairing, which would be encouraging considering the numerous alternative applicationsin articular cartilage tissue engineering.

Electrospun Nanocomposite Scaffold Based on Polycaprolactone-decellularized Umbilical Cord Wharton’s Jelly/Multi-walled Carbon Nanotubes: A Biomimetic Substrate for Articular Cartilage Tissue Engineering

Electrospun Nanocomposite Scaffold Based on Polycaprolactone-decellularized Umbilical Cord Wharton’s Jelly/Multi-walled Carbon Nanotubes: A Biomimetic Substrate for Articular Cartilage Tissue Engineering

3D Bioprinting Strategies for Articular Cartilage Tissue Engineering.

Articular cartilage is the avascular and aneural tissue which is the primary connective tissue covering the surface of articulating bone. Traumatic damage or degenerative diseases can cause articular cartilage injuries that are common in the population. As a result, the demand for new therapeutic options is continually increasing for older people and traumatic young patients. Many attempts have been made to address these clinical needs to treat articular cartilage injuries, including osteoarthritis (OA); however, regenerating highly qualified cartilage tissue remains a significant obstacle. 3D bioprinting technology combined with tissue engineering principles has been developed to create biological tissue constructs that recapitulate the anatomical, structural, and functional properties of native tissues. In addition, this cutting-edge technology can precisely place multiple cell types in a 3D tissue architecture. Thus, 3D bioprinting has rapidly become the most innovative tool for manufacturing clinically applicable bioengineered tissue constructs. This has led to increased interest in 3D bioprinting in articular cartilage tissue engineering applications. Here, we reviewed current advances in bioprinting for articular cartilage tissue engineering.

In Vitro and In Vivo Evaluation of a Polycaprolactone (PCL)/Polylactic-Co-Glycolic Acid (PLGA) (80:20) Scaffold for Improved Treatment of Chondral (Cartilage) Injuries.

Articular cartilage is a specialized tissue that provides a smooth surface for joint movement and load transmission. Unfortunately, it has limited regenerative capacity. Tissue engineering, combining different cell types, scaffolds, growth factors, and physical stimulation has become an alternative for repairing and regenerating articular cartilage. Dental Follicle Mesenchymal Stem Cells (DFMSCs) are attractive candidates for cartilage tissue engineering because of their ability to differentiate into chondrocytes, on the other hand, the polymers blend like Polycaprolactone (PCL) and Poly Lactic-co-Glycolic Acid (PLGA) have shown promise given their mechanical properties and biocompatibility. In this work, the physicochemical properties of polymer blends were evaluated by Fourier Transform Infrared Spectroscopy (FTIR) and Scanning Electron Microscope (SEM) and were positive for both techniques. The DFMSCs demonstrated stemness by flow cytometry. The scaffold showed to be a non-toxic effect when we evaluated it with Alamar blue, and the samples were analyzed using SEM and phalloidin staining to evaluate cell adhesion to the scaffold. The synthesis of glycosaminoglycans was positive on the construct in vitro. Finally, the PCL/PLGA scaffold showed a better repair capacity than two commercial compounds, when tested in a chondral defect rat model. These results suggest that the PCL/PLGA (80:20) scaffold may be suitable for applications in the tissue engineering of articular hyaline cartilage.

Application of graphene in articular cartilage tissue engineering and chondrogenic differentiation

Application of graphene in articular cartilage tissue engineering and chondrogenic differentiation

Human fetal cartilage-derived chondrocytes and chondroprogenitors display a greater commitment to chondrogenesis than adult cartilage resident cells.

Obtaining regeneration-competent cells and generating high-quality neocartilage are still challenges in articular cartilage tissue engineering. Although chondroprogenitor cells are a resident subpopulation of native cartilage and possess a high capacity for proliferation and cartilage formation, their potential for regenerative medicine has not been adequately explored. Fetal cartilage, another potential source with greater cellularity and a higher cell-matrix ratio than adult tissue, has been evaluated for sourcing cells to treat articular disorders. This study aimed to compare cartilage resident cells, namely chondrocytes, fibronectin adhesion assay-derived chondroprogenitors (FAA-CPCs) and migratory chondroprogenitors (MCPs) isolated from fetal and adult cartilage, to evaluate differences in their biological properties and their potential for cartilage repair. Following informed consent, three human fetal and three adult osteoarthritic knee joints were used to harvest the cartilage samples, from which the three cell types a) chondrocytes, b) FAA-CPCs, and MCPs were isolated. Assessment parameters consisted of flow cytometry analysis for percentage expression of cell surface markers, population doubling time and cell cycle analyses, qRT-PCR for markers of chondrogenesis and hypertrophy, trilineage differentiation potential and biochemical analysis of differentiated chondrogenic pellets for total GAG/DNA content. Compared to their adult counterparts, fetal cartilage-derived cells displayed significantly lower CD106 and higher levels of CD146 expression, indicative of their superior chondrogenic capacity. Moreover, all fetal groups demonstrated significantly higher levels of GAG/DNA ratio with enhanced uptake of collagen type 2 and GAG stains on histology. It was also noted that fetal FAA CPCs had a greater proliferative ability with significantly higher levels of the primary transcription factor SOX-9. Fetal chondrocytes and chondroprogenitors displayed a superior propensity for chondrogenesis when compared to their adult counterparts. To understand their therapeutic potential and provide an important solution to long-standing challenges in cartilage tissue engineering, focused research into its regenerative properties using in-vivo models is warranted.

Pd@Pt nanoparticle-linked immunosorbent assay for quantification of Collagen type II

The evaluation of specific protein content in engineered tissues provides a gateway for developing regenerative medicine treatments. Since collagen type II, the major component of articular cartilage, is critical for the blossoming field of articular cartilage tissue engineering, the interest in this protein is growing rapidly. Accordingly, the need for quantification of collagen type II is increasing as well. In this study, we provide recent results for a new quantifying nanoparticle sandwich immunoassay technique for collagen type II. Since mesoporous palladium@platinum (Pd@Pt) nanoparticles have peroxidase-like catalytic activities, these nanoparticles were utilized in an enzyme-linked immunosorbent assay (ELISA)-like format to circumvent the need for traditional enzymes. These nanoparticles were easily conjugated with anti-collagen type II antibodies by the natural affinity interaction and used to develop a direct sandwich ELISA-like format for nanoparticle-linked immunosorbent assays. Using this method, we obtained a limit of detection of 1 ng mL−1, a limit of quantification of 9 ng mL-1. and a broad linear range between 1 ng mL−1 and 50 μg mL−1 for collagen type II with an average relative standard deviation of 5.5%, useable over a pH range of 7 – 9 at least. The assay was successfully applied to quantify collagen type II in cartilage tissues and compared with the results of commercial ELISAs and gene expression by reverse transcription-quantitative polymerase chain reaction. This method provides a thermally stable and cost-efficient alternative to traditional ELISAs. It also extends the application of nanoparticle-linked immunosorbent assays, thereby providing the potential to quantify other proteins and apply the technology in the medical, environmental, and biotechnology industry fields.

An assessment of the response of human MSCs to hydrostatic pressure in environments supportive of differential chondrogenesis

Mechanical stimulation can modulate the chondrogenic differentiation of stem/progenitor cells and potentially benefit tissue engineering (TE) of functional articular cartilage (AC). Mechanical cues like hydrostatic pressure (HP) are often applied to cell-laden scaffolds, with little optimization of other key parameters (e.g. cell density, biomaterial properties) known to effect lineage commitment. In this study, we first sought to establish cell seeding densities and fibrin concentrations supportive of robust chondrogenesis of human mesenchymal stem cells (hMSCs). High cell densities (15*106 cells/ml) were more supportive of sGAG deposition on a per cell basis, while collagen deposition was higher at lower seeding densities (5*106 cells/ml). Employment of lower fibrin (2.5 %) concentration hydrogels supported more robust chondrogenesis of hMSCs, with higher collagen type II and lower collagen type X deposition compared to 5 % hydrogels. The application of HP to hMSCs maintained in identified chondro-inductive culture conditions had little effect on overall levels of cartilage-specific matrix production. However, if hMSCs were first temporally primed with TGF-β3 before its withdrawal, they responded to HP by increased sGAG production. The response to HP in higher cell density cultures was also associated with a metabolic shift towards glycolysis, which has been linked with a mature chondrocyte-like phenotype. These results suggest that mechanical stimulation may not be necessary to engineer functional AC grafts using hMSCs if other culture conditions have been optimised. However, such bioreactor systems can potentially be employed to better understand how engineered tissues respond to mechanical loading in vivo once removed from in vitro culture environments.

A core-shell Electrospun Scaffold of polyhydroxybutyrate-starch/halloysite Nanotubes Containing Extracellular Matrix and Chitosan for Articular Cartilage Tissue Engineering Application

A core-shell Electrospun Scaffold of polyhydroxybutyrate-starch/halloysite Nanotubes Containing Extracellular Matrix and Chitosan for Articular Cartilage Tissue Engineering Application

Comparison between articular chondrocytes and mesenchymal stromal cells for the production of articular cartilage implants.

Focal lesions of articular cartilage give rise to pain and reduced joint function and may, if left untreated, lead to osteoarthritis. Implantation of in vitro generated, scaffold-free autologous cartilage discs may represent the best treatment option. Here we compare articular chondrocytes (ACs) and bone marrow-derived mesenchymal stromal cells (MSCs) for their ability to make scaffold-free cartilage discs. Articular chondrocytes produced more extracellular matrix per seeded cell than mesenchymal stromal cells. Quantitative proteomics analysis showed that articular chondrocyte discs contained more articular cartilage proteins, while mesenchymal stromal cell discs had more proteins associated with cartilage hypertrophy and bone formation. Sequencing analysis revealed more microRNAs associated with normal cartilage in articular chondrocyte discs, and large-scale target predictions, performed for the first time for in vitro chondrogenesis, suggested that differential expression of microRNAs in the two disc types were important mechanisms behind differential synthesis of proteins. We conclude that articular chondrocytes should be preferred over mesenchymal stromal cells for tissue engineering of articular cartilage.

Hierarchical porous ECM scaffolds incorporating GDF-5 fabricated by cryogenic 3D printing to promote articular cartilage regeneration

BackgroundIn recent years, there has been significant research progress on in situ articular cartilage (AC) tissue engineering with endogenous stem cells, which uses biological materials or bioactive factors to improve the regeneration microenvironment and recruit more endogenous stem cells from the joint cavity to the defect area to promote cartilage regeneration.MethodIn this study, we used ECM alone as a bioink in low-temperature deposition manufacturing (LDM) 3D printing and then successfully fabricated a hierarchical porous ECM scaffold incorporating GDF-5.ResultsComparative in vitro experiments showed that the 7% ECM scaffolds had the best biocompatibility. After the addition of GDF-5 protein, the ECM scaffolds significantly improved bone marrow mesenchymal stem cell (BMSC) migration and chondrogenic differentiation. Most importantly, the in vivo results showed that the ECM/GDF-5 scaffold significantly enhanced in situ cartilage repair.ConclusionIn conclusion, this study reports the construction of a new scaffold based on the concept of in situ regeneration, and we believe that our findings will provide a new treatment strategy for AC defect repair.

Effects of Electrical Stimulation on Articular Cartilage Regeneration with a Focus on Piezoelectric Biomaterials for Articular Cartilage Tissue Repair and Engineering.

There is increasing evidence that chondrocytes within articular cartilage are affected by endogenous force-related electrical potentials. Furthermore, electrical stimulation (ES) promotes the proliferation of chondrocytes and the synthesis of extracellular matrix (ECM) molecules, which accelerate the healing of cartilage defects. These findings suggest the potential application of ES in cartilage repair. In this review, we summarize the pathogenesis of articular cartilage injuries and the current clinical strategies for the treatment of articular cartilage injuries. We then focus on the application of ES in the repair of articular cartilage in vivo. The ES-induced chondrogenic differentiation of mesenchymal stem cells (MSCs) and its potential regulatory mechanism are discussed in detail. In addition, we discuss the potential of applying piezoelectric materials in the process of constructing engineering articular cartilage, highlighting the important advances in the unique field of tissue engineering.

New refinements aim to optimize articular cartilage tissue engineering.

New refinements aim to optimize articular cartilage tissue engineering.

Mechanical environment for in vitro cartilage tissue engineering assisted by in silico models.

Mechanobiological study of chondrogenic cells and multipotent stem cells for articular cartilage tissue engineering (CTE) has been widely explored. The mechanical stimulation in terms of wall shear stress, hydrostatic pressure and mechanical strain has been applied in CTE in vitro. It has been found that the mechanical stimulation at a certain range can accelerate the chondrogenesis and articular cartilage tissue regeneration. This review explicitly focuses on the study of the influence of the mechanical environment on proliferation and extracellular matrix production of chondrocytes in vitro for CTE. The multidisciplinary approaches used in previous studies and the need for in silico methods to be used in parallel with in vitro methods are also discussed. The information from this review is expected to direct facial CTE research, in which mechanobiology has not been widely explored yet.