Potency Assay Considerations for Cartilage Repair, Osteoarthritis and Use of Extracellular Vesicles.

Articular cartilage covers the ends of bones in synovial joints acting as a shock absorber that helps movement of bones. Damage of the articular cartilage needs treatment as it does not repair itself and the damage can progress to osteoarthritis. In osteoarthritis all the joint tissues are involved with characteristic progressive cartilage degradation and inflammation. Autologous chondrocyte implantation is a well-proven cell-based treatment for cartilage defects, but a main downside it that it requires two surgeries. Multipotent, aka mesenchymal stromal cell (MSC)-based cartilage repair has gained attention as it can be used as a one-step treatment. It is proposed that a combination of immunomodulatory and regenerative capacities make MSC attractive for the treatment of osteoarthritis. Furthermore, since part of the paracrine effects of MSCs are attributed to extracellular vesicles (EVs), small membrane enclosed particles secreted by cells, EVs are currently being widely investigated for their potential therapeutic effects. Although MSCs have entered clinical cartilage treatments and EVs are used in in vivo efficacy studies, not much attention has been given to determine their potency and to the development of potency assays. This chapter provides considerations and suggestions for the development of potency assays for the use of MSCs and MSC-EVs for the treatment of cartilage defects and osteoarthritis.

3D-printed fish gelatin scaffolds for cartilage tissue engineering.

Knee osteoarthritis is a chronic disease caused by the deterioration of the knee joint due to various factors such as aging, trauma, and obesity, and the nonrenewable nature of the injured cartilage makes the treatment of osteoarthritis challenging. Here, we present a three-dimensional (3D) printed porous multilayer scaffold based on cold-water fish skin gelatin for osteoarticular cartilage regeneration. To make the scaffold, cold-water fish skin gelatin was combined with sodium alginate to increase viscosity, printability, and mechanical strength, and the hybrid hydrogel was printed according to a pre-designed specific structure using 3D printing technology. Then, the printed scaffolds underwent a double-crosslinking process to enhance their mechanical strength even further. These scaffolds mimic the structure of the original cartilage network in a way that allows chondrocytes to adhere, proliferate, and communicate with each other, transport nutrients, and prevent further damage to the joint. More importantly, we found that cold-water fish gelatin scaffolds were nonimmunogenic, nontoxic, and biodegradable. We also implanted the scaffold into defective rat cartilage for 12 weeks and achieved satisfactory repair results in this animal model. Thus, cold-water fish skin gelatin scaffolds may have broad application potential in regenerative medicine.

An Off-the-Shelf Tissue Engineered Cartilage Composed of Optimally Sized Pellets of Cartilage Progenitor/Stem Cells.

Articular cartilage focal lesion remains an intractable challenge in sports medicine, and autologous chondrocytes' implantation (ACI) is one of the most commonly utilized treatment modality for this ailment. However, the current ACI technique requires two surgical steps which increases patients' morbidity and incurs additional medical costs. In the present study, we developed a one-step cryopreserved off-the-shelf ACI tissue-engineered (TE) cartilage by seeding pellets of spheroidal cartilage stem/progenitor cells (CSPCs) on a silk scaffold. The pellets were developed through a hanging-drop method, and the incubation time of 1 day could efficiently produce spheroidal pellets without any adverse influence on the cell activity. The pellet size was also optimized. Under chondrogenic induction, pellets consisting of 40 000 CSPCs were found to exhibit the most abundant cartilage matrix deposition and the highest mRNA expression levels of SOX9, aggrecan, and COL2A1, as compared with pellets consisting of 10 000, 100 000, or 200 000 CSPCs. Scaffolds seeded with CSPCs pellets containing 40 000 cells could be preserved in liquid nitrogen with the viability, migration, and chondrogenic ability remaining unaffected for as long as 3 months. When implanted in a rat trochlear cartilage defect model for 3 months, the ready-to-use, cryopreserved TE cartilage yielded fully cartilage reconstruction, which was comparable with the uncryopreserved control. Hence, our study provided preliminary data that our off-the-shell TE cartilage with optimally sized CSPCs pellets seeded within silk scaffolds exhibited strong cartilage repair capacity, which provided a convenient and promising one-step surgical approach to ACI.

Injectable Ultrasonication-Induced Silk Fibroin Hydrogel for Cartilage Repair and Regeneration.

Articular cartilage lacks both a nutrient supply and progenitor cells. Once damaged, it has limited self-repair capability. Cartilage tissue engineering provides a promising strategy for regeneration, and the use of injectable hydrogels as scaffolds has recently attracted much attention. Silk fibroin (SF) is an advanced natural material used to construct injectable hydrogels that are nontoxic and can be used efficiently in crosslinking applications. The objective of the present work was to develop an injectable hydrogel using SF in a novel one-step ultrasonication crosslinking method. Gelation kinetics and the characteristics of ultrasonication-induced SF (US-SF) hydrogels were systematically evaluated. The cytocompatibility of US-SF hydrogels was evaluated using rabbit chondrocytes, the Cell Counting Kit-8 testing, and immunofluorescence staining. Furthermore, the in vivo cartilage regenerative ability of US-SF hydrogels was confirmed following subcutaneous administration in nude mice and in situ injections in rabbit osteochondral defect models. These results suggest that US-SF hydrogels could be potential candidates for cartilage repair and regeneration. Impact statement Injectable silk fibroin hydrogel is a promising strategy for cartilage tissue engineering. The transition from solution state to gel state can be fabricated by both physical and chemical methods. However, the complexing protocol and toxicity of these methods remain hindrances to further application. In this study, a one-step ultrasonication method was developed. The novel ultrasonication-induced silk fibroin hydrogel showed satisfactory physicochemical and biomechanical properties. In vitro and in vivo experiments proved that it could promote cartilage regeneration, indicating that it may be a potential solution for cartilage repair and regeneration.

Tissue engineering methods for repair of articular cartilage defect under special conditions / 中国组织工程研究

BACKGROUND: Although desired cartilage repair has been realized via tissue engineering technology, these achievements mainly focus on small-size defect under a normal physical condition. However, cartilage defects are always accompanied by the underlying diseases in clinical practice, such as osteoarthritis and rheumatoid arthritis. Meanwhile, the location, scope, and depth of cartilage defects are uncertain, which brings a great challenge in cartilage tissue repair. OBJECTIVE: To summarize the methods of repairing articular cartilage defects at different locations and under inflammatory condition. METHODS: We searched PubMed and CNKI with the search terms “cartilage defect regeneration, osteochondral, growth plate, weight-bearing area, inflammatory” in Chiense and English to retrieve related papers published before March 2019. A total of 209 papers were retrieved and 86 were included in the final analysis according to inclusion and exclusion criteria. RESULTS AND CONCLUSION: For articular cartilage defects under different special conditions, the repair goals and strategies are different. For repair of full-layer cartilage defects and osteochondral structure defects, multi-layered scaffolds are often used to repair the unique stratified cartilage structure and subchondral bone structure, while avoiding the problem of heterotopic ossification in neonatal cartilage. To avoid deformity after long bone maturation, growth factors such as insulin-like growth factor and bone morphogenetic protein 7 should be added to continuously stimulate the repair of the growth plate and promote bone growth. For cartilage repair in the weight-bearing area, the scaffolds should have good mechanical property, which ensure not to undergo severe deformation and structural damage when loaded. In addition, the new cartilage tissue has sufficient mechanical strength to support sustained longitudinal pressure and wear. For cartilage defects in the inflammatory state, both inflammation management and cartilage defect repair should be considered, and introduction of mesenchymal stem cells can regulate immune function and promote tissue regeneration, such that articular cartilage defect can be completely repaired.

Decellularized cartilage matrix scaffolds with laser-machined micropores for cartilage regeneration and articular cartilage repair.

Decellularized allogeneic and xenogeneic articular cartilage matrix scaffolds (CMS) are considered ideal scaffolds for cartilage regeneration owing to their heterogeneous architecture, and biochemical and biomechanical properties of native articular cartilage. However, the dense structure of the articular cartilage extracellular matrix, particularly the arrangement of collagen fibers, limits cellular infiltration, leading to poor cartilage regeneration. In addition, the incomplete removal of xenograft cells is associated with immunogenic reaction in the host. To facilitate the migration of chondrocytes into scaffolds and the rate of decellularization processing, we applied a carbon dioxide laser technique to modify the surface of porcine CMS while retaining major properties of the scaffold. By optimizing the laser parameters, we introduced orderly, lattice-arranged conical micropores of suitable depth and diameter onto the cartilage scaffold surface without affecting the cartilage shape or mechanical properties. We found that laser-modified CMS (LM-CMS) could enhance the degree of decellularization and were conducive to cell adhesion, as compared with the intact CMS. Decellularized scaffolds were seeded with rabbit-derived chondrocytes and cultured for 8 weeks in vitro. We found that cell-scaffold constructs formed cartilage-like tissue within the micropores and on the scaffold surface. In vivo, we found that cell-scaffold constructs subcutaneously implanted into the flanks of nude mice formed ivory-white neocartilage with high contents of DNA and cartilage matrix components, as well as good mechanical strength as compared with native CMS. Furthermore, scaffolds combined with autogenous chondrocytes induced neocartilage and better structural restoration at 8 weeks after transplantation into rabbit knee articular cartilage defects. In conclusion, decellularized xenogeneic CMS with laser-machined micropores offers an ideal scaffold with high fidelity for the functional reconstruction of articular cartilage.

Advances in research of mesenchymal stem cells in the treatment of osteoarthritis / 医学研究生学报

Osteoarthritis is a chronic progressive disease characterized by cartilage degenerative diseases. Due to the lack of vascular supply of articular cartilage and poor regeneration of chondrocytes, it is difficult to repair the cartilage with degenerative wear. A large number of studies have confirmed that bone marrow mesenchymal stem cells, adipose-derived mesenchymal stem cells, umbilical cord blood mesenchymal stem cells, umbilical cord mesenchymal stem cells, synovial mesenchymal stem cells, osteoarthritis joint fluid-derived mesenchymal stem cells, etc. can be effective to relieve osteoarthritis and repair damaged cartilage. Mesenchymal stem cells can directly differentiate into chondrocytes under appropriate microenvironment, and they also have immunosuppressive, anti-inflammatory and paracrine effects. In this paper, the research progress of basic experiments and clinical application of mesenchymal stem cells from different sources in osteoarthritis is reviewed to better promote the research progress of stem cell therapy for osteoarthritis.

Patient-Specific Surgical Guidance System for Intelligent Orthopaedics.

Clinical benefits for image-guided orthopaedic surgical systems are often measured in improved accuracy and precision of tool trajectories, prosthesis component positions and/or reduction of revision rate. However, with an ever-increasing demand for orthopaedic procedures, especially joint replacements, the ability to increase the number of surgeries, as well as lowering the costs per surgery, is generating a similar interest in the evaluation of image-guided orthopaedic systems. Patient-specific instrument guidance has recently gained popularity in various orthopaedic applications. Studies have shown that these guides are comparable to traditional image-guided systems with respect to accuracy and precision of the navigation of tool trajectories and/or prosthesis component positioning. Additionally, reports have shown that these single-use instruments also improve operating room management and reduce surgical time and costs. In this chapter, we discuss how patient-specific instrument guidance provides benefits to patients as well as to the health-care community for various orthopaedic applications.

Silk fibroin-chondroitin sulfate scaffold with immuno-inhibition property for articular cartilage repair.

The demand of favorable scaffolds has increased for the emerging cartilage tissue engineering. Chondroitin sulfate (CS) and silk fibroin have been investigated and reported with safety and excellent biocompatibility as tissue engineering scaffolds. However, the rapid degradation rate of pure CS scaffolds presents a challenge to effectively recreate neo-tissue similar to natural articular cartilage. Meanwhile the silk fibroin is well used as a structural constituent material because its remarkable mechanical properties, long-lasting in vivo stability and hypoimmunity. The application of composite silk fibroin and CS scaffolds for joint cartilage repair has not been well studied. Here we report that the combination of silk fibroin and CS could synergistically promote articular cartilage defect repair. The silk fibroin (silk) and silk fibroin/CS (silk-CS) scaffolds were fabricated with salt-leaching, freeze-drying and crosslinking methodologies. The biocompatibility of the scaffolds was investigated in vitro by cell adhesion, proliferation and migration with human articular chondrocytes. We found that silk-CS scaffold maintained better chondrocyte phenotype than silk scaffold; moreover, the silk-CS scaffolds reduced chondrocyte inflammatory response that was induced by interleukin (IL)-1ß, which is in consistent with the well-documented anti-inflammatory activities of CS. The in vivo cartilage repair was evaluated with a rabbit osteochondral defect model. Silk-CS scaffold induced more neo-tissue formation and better structural restoration than silk scaffold after 6 and 12weeks of implantation in ICRS histological evaluations. In conclusion, we have developed a silk fibroin/ chondroitin sulfate scaffold for cartilage tissue engineering that exhibits immuno-inhibition property and can improve the self-repair capacity of cartilage. STATEMENT OF SIGNIFICANCE: Severe cartilage defect such as osteoarthritis (OA) is difficult to self-repair because of its avascular, aneural and alymphatic nature. Current scaffolds often focus on providing sufficient mechanical support or bio-mimetic structure to promote cartilage repair. Thus, silk has been adopted and investigated broadly. However, inflammation is one of the most important factors in OA. But few scaffolds for cartilage repair reported anti-inflammation property. Meanwhile, chondroitin sulfate (CS) is a glycosaminoglycan present in the natural cartilage ECM, and has exhibited a number of useful biological properties including anti-inflammatory activity. Thus, we designed this silk-CS scaffold and proved that this scaffold exhibited good anti-inflammatory effects both in vitro and in vivo, promoted the repair of articular cartilage defect in animal model.

An in situ photocrosslinkable platelet rich plasma - Complexed hydrogel glue with growth factor controlled release ability to promote cartilage defect repair.

The repair of articular cartilage injury is a great clinical challenge. Platelet-rich plasma (PRP) has attracted much attention for the repair of articular cartilage injury, because it contains various growth factors that are beneficial for wound repair. However, current administration methods of PRP have many shortcomings, such as unstable biological fixation and burst release of growth factors, all of which complicate its application in the repair of articular cartilage and compromise its therapeutic efficacy. In this study, based on our previously reported photoinduced imine crosslinking (PIC) reaction, we developed an in situ photocrosslinkable PRP hydrogel glue (HNPRP) through adding a photoresponsive hyaluronic acid (HA-NB) which could generate aldehyde groups upon light irradiation and subsequently react with amino groups, into autologous PRP. Our study showed that HNPRP hydrogel glue was cytocompatible and could be conveniently and rapidly prepared in situ, forming a robust hydrogel scaffold. In addition, our results demonstrated that HNPRP hydrogel not only achieved controlled release of growth factors, but also showed strong tissue adhesive ability. Therefore, HNPRP hydrogel was quite suitable for cartilage defect regeneration. Our further in vitro experiment showed that HNPRP hydrogel could promote the proliferation and migration of chondrocytes and bone marrow stem cells (BMSCs). In vivo testing using a rabbit full-thickness cartilage defect model demonstrated that HNPRP hydrogel could achieve integrative hyaline cartilage regeneration and its therapeutic efficacy was better than thrombin activated PRP gel. STATEMENT OF SIGNIFICANCE: In this study, we have developed a photocrosslinkable platelet rich plasma (PRP) - complexed hydrogel glue (HNPRP) for cartilage regeneration. The in situ formed HNPRP hydrogel glue showed not only the controlled release ability of growth factors, but also strong tissue adhesiveness, which could resolve the current problems in clinical application of PRP. Furthermore, HNPRP hydrogel glue could promote integrative hyaline cartilage regeneration, and its reparative efficacy for cartilage defect was better than thrombin activated PRP gel. This study provided not only an effective repair material for cartilage regeneration, but also developed an advanced method for PRP application.

Cartilage defect repair in horses: Current strategies and recent developments in regenerative medicine of the equine joint with emphasis on the surgical approach.

Chondral and osteochondral lesions due to injury or other pathology are highly prevalent conditions in horses (and humans) and commonly result in the development of osteoarthritis and progression of joint deterioration. Regenerative medicine of articular cartilage is an emerging clinical treatment option for patients with articular cartilage injury or disease. Functional articular cartilage restoration, however, remains a major challenge, but the field is progressing rapidly and there is an increasing body of supportive clinical and scientific evidence. This review gives an overview of the established and emerging surgical techniques employed for cartilage repair in horses. Through a growing insight in surgical cartilage repair possibilities, surgeons might be more stimulated to explore novel techniques in a clinical setting.

Radially oriented collagen scaffold with SDF-1 promotes osteochondral repair by facilitating cell homing.

The migration of cells from the side and the bottom of the defect is important in osteochondral defect healing. Here, we designed a novel collagen scaffold that possessed channels in both the horizontal and the vertical directions, along with stromal cell-derived factor-1 (SDF-1) to enhance osteochondral regeneration by facilitating cell homing. Firstly we fabricated the radially oriented and random collagen scaffolds, then tested their properties. The radially oriented collagen scaffold had better mechanical properties than the random scaffold, but both supported cell proliferation well. Then we measured the migration of BMSCs in the scaffolds in vitro. The radially oriented collagen scaffold effectively promoted their migration, and this effect was further facilitated by addition of SDF-1. Moreover, we created osteochondral defects in rabbits, and implanted them with random or oriented collagen scaffolds with or without SDF-1, and evaluated cartilage and subchondral bone regeneration at 6 and 12 weeks after surgery. Cartilage regeneration with both the radially oriented scaffold and SDF-1 effectively promoted repair of the cartilage defect. Our results confirmed that the implantation of the radially oriented channel collagen scaffold with SDF-1 could be a promising strategy for osteochondral repair.

Clinical Application of Free Periosteal Autografting in Repairing Articular Cartilage / 中华创伤杂志

We repaired articular cartilage defect in 13 cases using free periosteal autografting. and followed up for 15～29 months with average of 22.4 months. Good results were obtained in 76.9%. The operation method and factors of influencing free periosteal autografting are discussed in the paper.