Osteochondral Tissue Engineering Research Articles

Elderberry (Sambucus nigra) and hawthorn (Crataegus oxyacantha) extract additives in carboxymethyl chitosan scaffolds for osteochondral tissue engineering applications

Elderberry (<i>Sambucus nigra</i>) and hawthorn (<i>Crataegus oxyacantha</i>) extract additives in carboxymethyl chitosan scaffolds for osteochondral tissue engineering applications

A rabbit osteochondral defect (OCD) model for evaluation of tissue engineered implants on their biosafety and efficacy in osteochondral repair.

Osteochondral defect (OCD) is a common but challenging condition in orthopaedics that imposes huge socioeconomic burdens in our aging society. It is imperative to accelerate the R&D of regenerative scaffolds using osteochondral tissue engineering concepts. Yet, all innovative implant-based treatments require animal testing models to verify their feasibility, biosafety, and efficacy before proceeding to human trials. Rabbit models offer a more clinically relevant platform for studying OCD repair than smaller rodents, while being more cost-effective than large animal models. The core-decompression drilling technique to produce full-thickness distal medial femoral condyle defects in rabbits can mimic one of the trauma-relevant OCD models. This model is commonly used to evaluate the implant's biosafety and efficacy of osteochondral dual-lineage regeneration. In this article, we initially indicate the methodology and describe a minimally-invasive surgical protocol in a step-wise manner to generate a standard and reproducible rabbit OCD for scaffold implantation. Besides, we provide a detailed procedure for sample collection, processing, and evaluation by a series of subsequent standardized biochemical, radiological, biomechanical, and histological assessments. In conclusion, the well-established, easy-handling, reproducible, and reliable rabbit OCD model will play a pivotal role in translational research of osteochondral tissue engineering.

Functionally graded hydrogels with opposing biochemical cues for osteochondral tissue engineering.

Osteochondral tissue (OC) repair remains a significant challenge in the field of musculoskeletal tissue engineering. OC tissue displays a gradient structure characterised by variations in both cell types and extracellular matrix components from cartilage to the subchondral bone. These functional gradients in the native tissue have been replicated to engineer osteochondral tissue in vitro. While diverse fabrication methods have been employed to create these microenvironments, emulating the natural gradients and effective regeneration of the tissue continues to present a significant challenge. In this study, we present the design and development of an interpenetrating (IPN) hydrogel with opposing dual biochemical gradients with the aim of regenerating the complete osteochondral unit. The gradients of biochemical cues were generated using an in-house built extrusion system in CMC-Silk IPN hydrogel having improved mechanical strength. Firstly, we fabricated a hydrogel that exhibits a smooth transition of sCMC and TGF-β1 (SCT gradient hydrogel) from the upper to the lower region of the IPN hydrogel, with the intent of regenerating the cartilage layer. Secondly, a hydrogel with a HAp gradient (HAp gradient hydrogel) from the lower to the upper region was fabricated to facilitate the regeneration of the subchondral bone layer. Subsequently, we developed a dual biochemical gradient hydrogel with the goal of regenerating the entire osteochondral unit. The fabricated hydrogel demonstrated a smooth transition of sCMC + TGF-β1 and HAp gradients in opposing directions, along with a blend of both the biochemical cues at the interface. The results showed that the hydrogels having biochemical cues corresponding to the three zones (i.e., cartilage, interface and bone) of dual biochemical gradient hydrogel led to differentiation of BMSCs towards their respective lineages, thereby demonstrating their efficacy in directing the fate of progenitor cells. The developed dual gradient hydrogel provided biochemical signals that have the potential to facilitate tissue growth and regeneration of the entire osteochondral tissue with a smooth transition from cartilage (soft) to bone (hard) tissues. In summary, our study provides a simple and innovative method for incorporating biochemical cues into hydrogels. This promising approach is translatable and has the potential to be extrapolated to other interface tissues.&#xD.

In vitro and in vivo evaluation of periosteum-derived cells and iPSC-derived chondrocytes encapsulated in GelMA for osteochondral tissue engineering.

Osteochondral defects are deep joint surface lesions that affect the articular cartilage and the underlying subchondral bone. In the current study, a tissue engineering approach encompassing individual cells encapsulated in a biocompatible hydrogel is explored in vitro and in vivo. Cell-laden hydrogels containing either human periosteum-derived progenitor cells (PDCs) or human induced pluripotent stem cell (iPSC)-derived chondrocytes encapsulated in gelatin methacryloyl (GelMA) were evaluated for their potential to regenerate the subchondral mineralized bone and the articular cartilage on the joint surface, respectively. PDCs are easily isolated and expanded progenitor cells that are capable of generating mineralized cartilage and bone tissue in vivo via endochondral ossification. iPSC-derived chondrocytes are an unlimited source of stable and highly metabolically active chondrocytes. Cell-laden hydrogel constructs were cultured for up to 28 days in a serum-free chemically defined chondrogenic medium. On day 1 and day 21 of the differentiation period, the cell-laden constructs were implanted subcutaneously in nude mice to evaluate ectopic tissue formation 4 weeks post-implantation. Taken together, the data suggest that iPSC-derived chondrocytes encapsulated in GelMA can generate hyaline cartilage-like tissue constructs with different levels of maturity, while using periosteum-derived cells in the same construct type generates mineralized tissue and cortical bone in vivo. Therefore, the aforementioned cell-laden hydrogels can be an important part of a multi-component strategy for the manufacturing of an osteochondral implant.

Designing Biomimetic 3D-Printed Osteochondral Scaffolds for Enhanced Load-Bearing Capacity.

Osteoarthritis is a debilitating chronic joint disorder that affects millions of people worldwide. Since palliative and surgical treatments cannot completely regenerate hyaline cartilage within the articulating joint, osteochondral (OC) tissue engineering has been explored to heal OC defects. Utilizing computational simulations and three-dimensional (3D) printing, we aimed to build rationale around fabricating OC scaffolds with enhanced biomechanics. First, computational simulations revealed that interfacial fibrils within a bilayer alter OC scaffold deformation patterns by redirecting load-induced stresses toward the top of the cartilage layer. Principal component analysis revealed that scaffolds with 800 μm long fibrils (scaffolds 8A-8H) possessed optimal biomechanical properties to withstand compression and shear forces. While compression testing indicated that OC scaffolds with 800 μm fibrils did not have greater compressive moduli than other scaffolds, interfacial shear tests indicated that scaffold 8H possessed the greatest shear strength. Lastly, failure analysis demonstrated that yielding or buckling models describe interfacial fibril failure depending on fibril slenderness S. Specifically for scaffolds with packing density n = 6 and n = 8, the yielding failure model fits experimental loads with S < 10, while the buckling model fitted scaffolds with S < 10 slenderness. The research presented provides critical insights into designing 3D printed interfacial scaffolds with refined biomechanics toward improving OC tissue engineering outcomes.

Loofah and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) nano-fiber-reinforced chitosan hydrogel composite scaffolds with elderberry (Sambucus nigra) and hawthorn (Crataegus oxyacantha) extracts as additives for osteochondral tissue engineering applications

AbstractIn recent years, people have had more expectations from the developed technology in medicine, especially in the field of orthopedics and traumatology. Tissue engineers are interested in techniques that benefit from patients’ cells and biomaterials, instead of prostheses and implants. On the other hand, researchers have begun to use various medicinal plants for regeneration and anti-cancer studies. In the present study, we aimed to produce cartilage and bone inductive scaffolds for osteochondral tissue engineering applications with the addition of hawthorn or elderberry extracts. Firstly, wet electro-spun poly (3-hydroxybutyric acid-co-3-hydroxyvaleric acid) fibers were integrated with a loofah mat. Then, they were impregnated into chitosan solution with/without hawthorn or elderberry extract. Composite hydrogel scaffolds were obtained by cross-linking with 0.3% (w/v) genipin. Fabricated scaffolds had more than 90% porosity and showed swelling capacity in the range of 1500–2200%. Based on the in vitro biocompatibility analyses using mesenchymal stem cells (MSCs), all the fabricated scaffolds were found to be biocompatible by WST-1, ALP activity, and GAG content analysis. Also, histological/immunohistochemical analyses showed that hawthorn and elderberry extract addition increased MSCs proliferation and collagen type I and II positivity. Consequently, all the scaffolds showed promising features for osteochondral tissue engineering applications. Graphical abstract

Evaluation of rabbit adipose derived stem cells fate in perfused multilayered silk fibroin composite scaffold for Osteochondral repair.

Development of osteochondral tissue engineering approaches using scaffolds seeded with stem cells in association with mechanical stimulations has been recently considered as a promising technique for the repair of this tissue. In this study, an integrated and biomimetic trilayered silk fibroin (SF) scaffold containing SF nanofibers in each layer was fabricated. The osteogenesis and chondrogenesis of stem cells seeded on the fabricated scaffolds were investigated under a perfusion flow. 3-Dimethylthiazol-2,5-diphenyltetrazolium bromide assay showed that the perfusion flow significantly enhanced cell viability and proliferation. Analysis of gene expression by stem cells revealed that perfusion flow had significantly upregulated the expression of osteogenic and chondrogenic genes in the bone and cartilage layers and downregulated the hypertrophic gene expression in the intermediate layer of the scaffold. In conclusion, applying flow perfusion on the prepared integrated trilayered SF-based scaffold can support osteogenic and chondrogenic differentiation for repairing osteochondral defects.

Electromagnetic field-mediated chitosan/gelatin/nano-hydroxyapatite and bone-derived scaffolds regulate the osteoblastic and chondrogenic phenotypes of adipose-derived stem cells to construct osteochondral tissue engineering niche in vitro

It is critical to explore the effects of electromagnetic field (EMF) on the construction of functional osteochondral tissue, which has shown certain clinical significance for the treatment of osteochondral injury. At present, there are few studies on the effect of the direction of EMF on cells. This study aimed to investigate the effects of EMF coupling on different parameters to control adipose-derived stem cells (ADSCs) proliferation and specific chondrogenic and osteogenic differentiation at 2D level and 3D level. The proliferation and differentiation of EMF-induced ADSCs are jointly regulated by EMF and space structure. In this study, Cs7/Gel3/nHAP scaffolds were prepared with good degradation rate (86.75 ± 4.96 %) and absorb water (1100 %), and the pore size was 195.63 ± 54.72 μm. The bone-derived scaffold with a pore size of 267.17 ± 129.18 μm was obtained and its main component was hydroxyapatite. Cs7/Gel3/nHAP scaffolds and bone-derived scaffolds are suitable as 3D level materials. The optimal EMF intensity was 2 mT for chondrogenic differentiation and proliferation and 1 mT for osteogenic differentiation and proliferation. It is noteworthy that EMF has a negative correlation with ADSCs proliferation in the vertical direction at 2D level, while it has a positive correlation with ADSCs proliferation at 3D level. EMF mediated 3D osteochondral scaffold provide good strategy for osteochondral tissue engineering construction.

Microstructurally and mechanically tunable acellular hydrogel scaffold using carboxymethyl cellulose for potential osteochondral tissue engineering

In tissue engineering, scaffold microstructures and mechanical cues play a significant role in regulating stem cell differentiation, proliferation, and infiltration, offering a promising strategy for osteochondral tissue repair. In this present study, we aimed to develop a facile method to fabricate an acellular hydrogel scaffold (AHS) with tunable mechanical stiffness and microstructures using carboxymethyl cellulose (CMC). The impacts of the degree of crosslinking, crosslinker length, and matrix density on the AHS were investigated using different characterization methods, and the in vitro biocompatible of AHS was also examined. Our CMC-based AHS showed tunable mechanical stiffness ranging from 50 kPa to 300 kPa and adjustable microporous size between 50 μm and 200 μm. In addition, the AHS was also proven biocompatible and did not negatively affect rabbit bone marrow stem cells' dual-linage differentiation into osteoblasts and chondrocytes. In conclusion, our approach may present a promising method in osteochondral tissue engineering.

Using loofah reinforced chitosan-collagen hydrogel based scaffolds in-vitro and in-vivo; healing in cartilage tissue defects

The herein article aims to report a new scaffold design as a loofah-reinforced chitosan-collagen hydrogel composite scaffold with three different cross-linker concentrations (0.1, 0.3, and 0.5 wt. /v%). From the analyses, the scaffold crosslinked with 0.5% genipin; collagen-chitosan hydrogel scaffold reinforced with loofah (L-CCol5) was found to be suitable for further in vitro and in vivo studies due to its interconnected porous structure, water content (∼ 97%) and tan delta (0.221 at 1 Hz) values comparable to that of cartilage tissue. In vitro analyses depicted that the L-CCol5 scaffold supported rabbit mesenchymal stem cells (rMSCs) adhesion and proliferation with its non-cytotoxic feature. Moreover, in vivo cartilage healing studies were performed using New Zealand male rabbits in three groups: empty control, cell-free scaffold, and rMSCs-laden scaffold. The elastic moduli of these three groups were 0.69, 0.90, and 1.18 MPa, respectively. Besides, microcomputer tomography (MicroCT) scannings supported the in vivo biomechanical analyses as cell-laden scaffolds showed better osteochondral healing. It can be concluded that the L-CCol5 scaffold could be a promising construct in osteochondral tissue engineering applications. The findings revealed that osteochondral remodeling precedes articular cartilage, providing insight into tailored therapeutic approaches, disease progress, and treatment consequences.

3D-printed gradient scaffolds for osteochondral defects: Current status and perspectives.

Articular osteochondral defects are quite common in clinical practice, and tissue engineering techniques can offer a promising therapeutic option to address this issue.The articular osteochondral unit comprises hyaline cartilage, calcified cartilage zone (CCZ), and subchondral bone.As the interface layer of articular cartilage and bone, the CCZ plays an essentialpart in stress transmission and microenvironmental regulation.Osteochondral scaffolds with the interface structure for defect repair are the future direction of tissue engineering. Three-dimensional (3D) printing has the advantages of speed, precision, and personalized customization, which can satisfy the requirements of irregular geometry, differentiated composition, and multilayered structure of articular osteochondral scaffolds with boundary layer structure. This paper summarizes the anatomy, physiology, pathology, and restoration mechanisms of the articular osteochondral unit, and reviews the necessity for a boundary layer structure in osteochondral tissue engineering scaffolds and the strategy for constructing the scaffolds using 3D printing. In the future, we should not only strengthen the basic research on osteochondral structural units, but also actively explore the application of 3D printing technology in osteochondral tissue engineering. This will enable better functional and structural bionics of the scaffold, which ultimately improve the repair of osteochondral defects caused by various diseases.

Injectable, self-healing poly(amino acid)-hydrogel based on phenylboronate ester bond for osteochondral tissue engineering

A new generation of osteochondral integrated scaffolds is needed for articular osteochondral regeneration, which can not only facilitate the accurate construction of osteochondral scaffolds in a minimally invasive manner but also firmly combine the subchondral bone layer and cartilage layer. Herein, an osteochondral integrated hydrogel scaffold was constructed by the poly(L-glutamic acid) (PLGA) based self-healing hydrogels with phenylboronate ester (PBE) as the dynamic cross-linking. The bone layer self-healing hydrogel (hydrogel O-S) was prepared by physically blending nanohydroxyapatite into the self-healing hydrogel PLGA-PBE-S, which was fabricated by 3-aminophenylboronic acid/glycidyl methacrylate-modified PLGA (PLGA-GMA-PBA) and 3-amino-1,2-propanediol/N-(2-aminoethyl) acrylamide-modified PLGA (PLGA-ADE-AP). The cartilage layer self-healing hydrogel (hydrogel C-S) was prepared by PLGA-GMA-APBA and glucosamine- modified PLGA-ADE-AP (PLGA-ADE-AP-G). Excellent injectability and self-healing profiles of hydrogel O-S and C-S were observed, the self-healing efficiencies were 97.02% ± 1.06% and 99.06% ± 0.57%, respectively. Based on the injectability and spontaneous healing on the interfaces of hydrogel O-S and C-S, the osteochondral hydrogel (hydrogel OC) was conveniently constructed in a minimally invasive manner. In addition, in situ photocrosslinking was used to enhance the mechanical strength and stability of the osteochondral hydrogel. The osteochondral hydrogels exhibited good biodegradability and biocompatibility. The osteogenic differentiation genes BMP-2, ALPL, BGLAP and COL I of adipose-derived stem cells (ASCs) in the bone layer of the osteochondral hydrogel were significantly expressed, and the chondrogenic differentiation genes SOX9, aggrecan and COL II of ASCs in the cartilage layer of the osteochondral hydrogel were obviously upregulated after 14 d of induction. The osteochondral hydrogels could effectively promote repair of osteochondral defects after 3 months post-surgery.

Bioinspired gradient scaffolds for osteochondral tissue engineering.

Repairing articular osteochondral defects present considerable challenges in self-repair due to the complex tissue structure and low proliferation of chondrocytes. Conventional clinical therapies have not shown significant efficacy, including microfracture, autologous/allograft osteochondral transplantation, and cell-based techniques. Therefore, tissue engineering has been widely explored in repairing osteochondral defects by leveraging the natural regenerative potential of biomaterials to control cell functions. However, osteochondral tissue is a gradient structure with a smooth transition from the cartilage to subchondral bone, involving changes in chondrocyte morphologies and phenotypes, extracellular matrix components, collagen type and orientation, and cytokines. Bioinspired scaffolds have been developed by simulating gradient characteristics in heterogeneous tissues, such as the pores, components, and osteochondrogenesis-inducing factors, to satisfy the anisotropic features of osteochondral matrices. Bioinspired gradient scaffolds repair osteochondral defects by altering the microenvironments of cell growth to induce osteochondrogenesis and promote the formation of osteochondral interfaces compared with homogeneous scaffolds. This review outlines the meaningful strategies for repairing osteochondral defects by tissue engineering based on gradient scaffolds and predicts the pros and cons of prospective translation into clinical practice.

A Biphasic Hydrogel with Self-Healing Properties and a Continuous Layer Structure for Potential Application in Osteochondral Defect Repair.

The treatment of osteochondral defects remains challenging due to the limited healing capacity of cartilage and the poor results of traditional methods. Inspired by the structure of natural articular cartilage, we have fabricated a biphasic osteochondral hydrogel scaffold using a Schiff base reaction and a free radical polymerization reaction. Carboxymethyl chitosan (CMCS), oxidized sodium alginate (OSA), and polyacrylamide (PAM) formed a hydrogel (COP) as the cartilage layer, while hydroxyapatite (HAp) was incorporated into the COP hydrogel to obtain a hydrogel (COPH) as an subchondral bone layer. At the same time, hydroxyapatite (HAp) was incorporated into the COP hydrogel to obtain a hydrogel (COPH) as an osteochondral sublayer, combining the two to obtain an integrated scaffold for osteochondral tissue engineering. Interlayer interpenetration through the continuity of the hydrogel substrate and good self-healing properties due to the dynamic imine bonding of the hydrogel resulted in enhanced interlayer bond strength. In addition, in vitro experiments have shown that the hydrogel exhibits good biocompatibility. It shows great potential for osteochondral tissue engineering applications.

Structural and biological investigation of alginate-nano-hydroxyapatite with chitosan-hyaluronic acid for potential osteochondral regeneration

Cartilage joint lesions have created a significant challenge in the field of durable reconstruction and recovery due to their limited long-term repair and reconstruction. In this study, a biphasic scaffold was obtained using chitosan (CS)-hyaluronic acid (HA) and alginate-nanohydroxyapatite (nHAP) for osteoarthritis diseases. The scaffolds revealed porous structures and hydrophilic chemical groups by scanning electron microscopy (SEM) and Fourier-transform infrared spectroscopy (FTIR) methods, respectively. The resulting three-dimensional hydrogels were assembled by using fibrin glue and seeded with human chondrocytes cells (hCHCs) in the chondral layer and human adipose-derived mesenchymal stem cells (hAdMSCs) in the subchondral one. Then, the assessments including 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), SEM, compressive assay, weight loss, alizarin red staining, alkaline phosphatase (ALP) activity, alcian blue staining and Real-Time PCR were performed. The associated cell viability confirmed that there were no interventions between the cell types and SEM illustrated cell attachment and spreading. The subchondral and chondral layers were investigated separately to detect their ability for cellular commitment. Finally, the Real-time PCR was carried out on the bilayer scaffold and the related observations approved the osteogenic/chondrogenic differentiation of both cell types. Taken together, the Alg-nHAP/CS-HA scaffold provided an appropriate environment for cartilage and bone regenerations and could be recommended for all aspects of osteochondral tissue engineering.

Manufacture of Bilayered Composite Hydrogels with Strong, Elastic, and Tough Properties for Osteochondral Repair Applications.

Layered composite hydrogels have been considered attractive materials for use in osteochondral repair and regeneration. These hydrogel materials should be mechanically strong, elastic, and tough besides fulfilling some basic requirements such as biocompatibility and biodegradability. A novel type of bilayered composite hydrogel with multi-network structures and well-defined injectability was thus developed for osteochondral tissue engineering using chitosan (CH), hyaluronic acid (HA), silk fibroin (SF), CH nanoparticles (NPs), and amino-functionalized mesoporous bioglass (ABG) NPs. CH was combined with HA and CH NPs to build the chondral phase of the bilayered hydrogel, and CH, SF, and ABG NPs were used together to construct the subchondral phase of the bilayer hydrogel. Rheological measurements showed that the optimally achieved gels assigned to the chondral and subchondral layers had their elastic moduli of around 6.5 and 9.9 kPa, respectively, with elastic modulus/viscous modulus ratios higher than 36, indicating that they behaved like strong gels. Compressive measurements further demonstrated that the bilayered hydrogel with an optimally formulated composition had strong, elastic, and tough characteristics. Cell culture revealed that the bilayered hydrogel had the capacity to support the in-growth of chondrocytes in the chondral phase and osteoblasts in the subchondral phase. Results suggest that the bilayered composite hydrogel can act as an injective biomaterial for osteochondral repair applications.

Osteochondral Tissue Engineering Dilemma: Scaffolding Trends in Regenerative Medicine.

Orthopedic surgeons face a lot of difficulties in managing and repairing osteochondral defects. Damaged articular cartilage and the subchondral bone underneath are both present in osteochondral defects. The demands of the bone, cartilage, and the contact between the bone and the cartilage must be taken into consideration while repairing an osteochondral defect. Only palliative, not curative, therapeutic interventions are now available for the healing of osteochondral abnormalities. With its ability to successfully rebuild bone, cartilage, and the junction between bone and cartilage, tissue engineering has been recognized as an effective substitute. In correlation, mechanical stress and physical processes are commonly applied to the osteochondral area. Therefore, the ability of chondrocytes and osteoblasts to regenerate is influenced by bioactive molecules and the physicochemical characteristics of the surrounding matrix. The treatment of osteochondral disorders is said to benefit from the use of stem cells as an alternative intervention. In the field of tissue engineering, various approaches have been used such as the direct implantation of scaffolding materials at the site of tissue injury in patients, either alone or loaded with cells and bioactive molecules at the target site to imitate the natural extracellular matrix. Despite the extensive use and advancements of tissue-engineered biomaterials such as natural and synthetic polymer-based scaffolds, their repair capacity is limited due to challenges in combating antigenicity, designed to simulate in vivo microenvironment, and conducting mechanical or metabolic characteristics comparable to native organs/tissues. This study explores numerous osteochondral tissue engineering methodologies focusing on scaffold design, material varieties, manufacturing techniques, and functional features. This review is focused on recent breakthroughs in bioactive scaffolds that aid osteogenic and chondrogenic differentiation for bone and cartilage repair. The topic will cover fundamental anatomy, osteochondral repair methodologies and obstacles, cell selection, biochemical variables, and bioactive materials, as well as the design and manufacture of bioactive scaffolds. Additionally, we focus on the concept and construction of decellularized scaffolds, and the fabrication of dECM scaffolds in tissue engineering from various skin, bone, nerve, heart tissue, lung, liver, and kidney, and their application in osteochondral regeneration.

Bioreactor strategies for tissue-engineered osteochondral constructs: Advantages, present situations and future trends

The aim of osteochondral tissue engineering is to achieve the complex, functional and three-dimensional tissue regeneration under well defined, controlled and reproducible conditions in vitro. To achieve tissue-engineered products in vitro that incorporate rapidly in vivo with healthy tissue, it is essential to develop high-performance cell/scaffold culture systems that mimic the dynamics of the in vivo environment. Bioreactors could provide specific physicochemical culture environment, suitable mechanical stimulation and controlled condition for the development of osteochondral constructs in vitro. This review highlighted the multifunction of bioreactor in tissue engineering, and presented microenvironment and biomechanics of native osteochondral tissue, to illustrate the necessity of establishing osteochondral constructs by bioreactor. Then, we especially emphasized the advantages and limitations of various bioreactors. Furthermore, we systematically summarized and discussed the development of bioreactor-based production systems for bone, cartilage and osteochondral tissue engineering in recent years. Finally, we made a simple conclusion and offered perspectives of bioreactor‐based osteochondral tissue engineering. This review aims to serve as a reference for incorporating bioreactor strategies which could provide mechanical stimulation and physicochemical culture environment into the osteochondral construct culture regimens.

Dual composition Chondroitin Sulfate and gelatin biomimetic hydrogel based on tyramine crosslinking for tissue regenerative medicine

Hydrogels with bone biomimetic microenvironment are of great importance for the development of bone-graft materials. Herein, a biomimetic hydrogel, which was based on biopolymer as an extracellular matrix (ECM) mimicking components with the incorporation of biphasic calcium phosphate (BCP) nanoparticles, was prepared via the enzymatic crosslinking. Chondroitin sulfate and gelatin were first modified to form tyramine (TA) moieties on the natural polymers, which were ultilized as precursors for in-situ forming hydrogel (CDTA-GTA) under physiological conditions. The amount of CDTA to GTA was optimized for gelation time as well as degraded ability with in collagenase medium. Increasing the CDTA amount delayed the time for crosslinking process, but the water holding capacity was increased. The subjection of BCP into CDTA-GTA hydrogel produced an injectable system with suitable biomineralizated property. This hydrogel formed more rapidly in 90–100 s under 3 mM H2O2 and 0.125 mg/mL HRP enzyme. BCP/CDTA-GTA hydrogel exhibited the porous structure (100–300 µm in diameter) with the homogenous distribution of BCP nanoparticles on the wall of the interconnected networks, suggesting the good environment for bone growth. Further, the degradation process of the obtained hydrogel followed pseudo-first-order kinetics in the collagenase condition. After soaking in SBF for 28 days, SEM following EDS technique confirmed the aggregation of the calcium and phosphorus on the surface of CDTA-GTA hydrogel (Ca/P ratio ∼ 1.34) which improved in case of BCP/ CDTA-GTA hydrogel (Ca/P ratios ∼ 1.77). Through X-ray diffraction, the formation of calcite crystals was only detected with BCP hydrogel. In vitro cytotoxicity assays with hMSC cells exhibited non-toxicity of the biomimetic hydrogel. Interestingly, the remarkably enhancement of the level intracellular calcium deposit as well as the activity of ALP-osteogenic related marker were observed in MSC cells treated with biomimetic hydrogel, showing superior scaffold osteogenic potential. These results indicate that enzymatic cross-linked CDTA-GTA gel with BCP nanoparticles would be a favorable option to accelerate bone repair and thus should potentially be useful for osteochondral tissue engineering.

Polysaccharide-Based Composite Scaffolds for Osteochondral and Enthesis Regeneration.

The rotator cuff and Achilles tendons along with the anterior cruciate ligament (ACL) are frequently injured with limited healing capacity. At the soft-hard tissue interface, enthesis is prone to get damaged and its regeneration in osteochondral defects is essential for complete healing. The current clinical techniques used in suturing procedures to reattach tendons to bones need much improvement for the generation of the native interface tissue, that is, enthesis, for patients to regain their full functions. Recently, inspired by the composite native tissue, much effort has been made to fabricate composite scaffolds for enthesis tissue regeneration. This review first focuses on the studies that used composite scaffolds for the regeneration of enthesis. Then, the use of polysaccharides for osteochondral tissue engineering is reviewed and their potential for enthesis regeneration is presented based on their supporting effects on osteogenesis and chondrogenesis. Gellan gum (GG) is selected and reviewed as a promising polysaccharide due to its unique osteogenic and chondrogenic activities that help avoid the inherent weakness of dissimilar materials in composite scaffolds. In addition, original preliminary results showed that GG supports collagen type I production and upregulation of osteogenic marker genes. Impact Statement Enthesis regeneration is essential for complete and functional healing of tendon and ligament tissues. Current suturing techniques to reattach the tendon/ligament to bones have high failure rates. This review highlights the studies on biomimetic scaffolds aimed to regenerate enthesis. In addition, the potential of using polysaccharides to regenerate enthesis is discussed based on their ability to regenerate osteochondral tissues. Gellan gum is presented as a promising biopolymer that can be modified to simultaneously support bone and cartilage regeneration by providing structural continuity for the scaffold.