Scaffold For Cartilage Repair Research Articles

Small Ruminant Models for Articular Cartilage Regeneration by Scaffold-Based Tissue Engineering.

Animal models play an important role in preclinical studies, especially in tissue engineering scaffolds for cartilage repair, which require large animal models to verify the safety and effectiveness for clinical use. The small ruminant models are most widely used in this field than other large animals because they are cost-effective, easy to raise, not to mention the fact that the aforementioned animal presents similar anatomical features to that of humans. This review discusses the experimental study of tissue engineering scaffolds for knee articular cartilage regeneration in small ruminant models. Firstly, the selection of these scaffold materials and the preparation process in vitro that have been already used in vivo are briefly reviewed. Moreover, the major factors influencing the rational design and the implementation as well as advantages and limitations of small ruminants are also demonstrated. As regards methodology, this paper applies principles and methods followed by most researchers in the process of experimental design and operation of this kind. By summarizing and comparing different therapeutic concepts, this paper offers suggestions aiming to increase the effectiveness of preclinical research using small ruminant models and improve the process of developing corresponding therapies.

Graphene oxide–modified 3D acellular cartilage extracellular matrix scaffold for cartilage regeneration

Articular cartilage regeneration is a challenge in orthopedics and tissue engineering. This study prepared a graphene oxide (GO)-modified 3D acellular cartilage extracellular matrix (ACM) scaffold for cartilage repair. Cartilage slices were decellularized using a combination of physical and chemical methods of fabricating ACM particles. GO was crosslinked with the ACM by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride and N-hydroxy succinimide to prepare a composite scaffold. GO modification improved the internal structure and mechanical properties of the scaffold. The GO-modified (2 mg/mL) composite scaffold promoted cell adhesion, cell proliferation, and chondrogenic differentiation in vitro. Experiments on subcutaneous implantation in rats demonstrated that the composite scaffold had good biocompatibility and mild inflammatory response. After 12 weeks of implantation, the composite scaffold loaded with bone marrow mesenchymal stem cells completely bridged the cartilage defects in the rabbit knee with hyaline cartilage. Results indicated that the GO-modified 3D ACM composite scaffold can provide a powerful platform for cartilage tissue engineering and articular cartilage injury treatment.

Design and evaluation of nano-hydroxyapatite/poly(vinyl alcohol) hydrogels coated with poly(lactic-co-glycolic acid)/nano-hydroxyapatite/poly(vinyl alcohol) scaffolds for cartilage repair

BackgroundPoly(vinyl alcohol) (PVA) hydrogels have been widely used in synthetic cartilage materials. However, limitations of PVA hydrogels such as poor biomechanics and limited cell ingrowth remain challenges in this field.MethodsThis work aimed to design novel nano-hydroxyapatite (nano-HA)/poly(vinyl alcohol) (PVA) hydrogels coated with a poly(lactic-co-glycolic acid) (PLGA)/nano-HA/PVA scaffold to counter the limitations of PVA hydrogels. The core, comprising nano-HA/PVA hydrogel, had the primary role of bearing the mechanical load. The peripheral structure, composed of PLGA/nano-HA/PVA, was designed to favor interaction with surrounding cartilage.ResultsThe double-layer HA/PVA hydrogel coated with PLGA/HA/PVA scaffold was successfully prepared using a two-step molding method, and the mechanical properties and biocompatibility were characterized. The mechanical properties of the novel PLGA/HA/PVA scaffold modified HA/PVA hydrogel were similar to those of native cartilage and showed greater sensitivity to compressive stress than to tensile stress. Rabbit chondrocytes were seeded in the composites to assess the biocompatibility and practicability in vitro. The results showed that the peripheral component comprising 30 wt% PLGA/5 wt% HA/15 wt% PVA was most conducive to rabbit chondrocyte adhesion and proliferation.ConclusionsThe study indicated that the double-layer HA/PVA hydrogel coated with PLGA/HA/PVA scaffold has the potential for cartilage repair.

Cell-free cartilage repair in large defects of the knee: increased failure rate 5years after implantation of a collagen type I scaffold.

Cartilage defects of the knee remain a challenging problem in orthopedic surgery despite the ongoing improvements in regenerative procedures such as the autologous chondrocyte transplantation. Due to the lack of donor-site morbidity and the single-stage procedure cell-free scaffolds are an interesting alternative to cell-based procedures. But as currently mid- and long-term data are lacking, the aim of the present study was to present mid-term clinical, radiological and histological results of a cell-free collagen type I scaffolds for cartilage repair. Twenty-eight patients were followed prospectively. Clinical evaluation using patient-reported outcome measures (KOOS, IKDC; VAS for pain, Tegner score for activity) as well as radiologic evaluation of the repair tissue (MOCART) was performed at 1year, 2years and 5years. Histologic evaluation of the repair tissue was done in case of revision surgery using the ICRS II score for human cartilage repair. In these large cartilage defects with a mean defect size of 3.7 ± 1.9cm2, clinical failure necessitating revision surgery was seen in 5 of 28 patients (18%). While the remaining patients showed good-to-excellent clinical results (KOOS, IKDC, VAS, Tegner), the radiologic appearance of the repair tissue showed a reduction of the MOCART score between the 2- and 5-year follow-up. Histologic evaluation of the repair tissue showed a cartilage-like appearance with no signs of inflammation or cell death but an overall medium tissue quality according to the ICRS II Score. The use of this cell-free collagen type I scaffold for large defects showed increased wear of the repair tissue and clinical failure in 18% of cases at 5-year follow-up.

Comparison of Fixation Techniques of 3D-Woven Poly(ϵ-Caprolactone) Scaffolds for Cartilage Repair in a Weightbearing Porcine Large Animal Model.

Objective To test different fixation methods of a 3-dimensionally woven poly(ϵ-caprolactone) (PCL) scaffold within chondral defects of a weightbearing large animal model. Methods Full thickness chondral defects were made in the femoral condyles of 15 adult male Yucatan mini-pigs. Two surgical approaches were compared including total arthrotomy (traditional) and a retinaculum-sparing, minimally invasive surgery (MIS) approach. Following microfracture (MFX), scaffolds were placed without fixation or were fixed with fibrin glue, suture, or subchondral anchor. Experimental endpoints were between 1 and 6 weeks. Micro-computed tomography and histology were used to assess samples. Results The MIS approach was superior as the traditional approach caused medial condyle cartilage wear. One of 13 (7.7%) of scaffolds without fixation, 4 of 11 (36.3%) fibrin scaffolds, 1 of 4 (25%) of sutured scaffolds, and 9 of 9 (100%) of anchor-fixed scaffolds remained in place. Histology demonstrated tissue filling with some overgrowth of PCL scaffolds. Conclusions Of the methods tested, the MIS approach coupled with subchondral anchor fixation provided the best scaffold retention in a mini-pig chondral defect model. This finding has implications for fixation strategies in future animal studies and potential future human use.

A scaffold-filter model for studying the chondrogenic differentiation of stem cells in vitro

This study was undertaken to explore the synergistic effect of scaffold materials and a cartilage-like environment on the chondrogenic differentiation of stem cells. Because stem cells encapsulated in a cartilage scaffold will be induced by scaffold molecules as well as permeable molecules from the surroundings, it is impossible to optimize a chondro-inducible scaffold without considering environmental sensitivity. How do we know if a designed scaffold will be sufficient prior to implantation? In this study, bone marrow mesenchymal stem cells (bMSCs) were seeded in various scaffolds, including collagen hydrogel, collage/sodium alginate hydrogel, collagen sponge and silk fibroin sponge. The cell-scaffold complex was encapsulated in a filter pocket to avoid direct contact with co-cultured chondrocytes. Scaffolds differed in the ability to adsorb inducible molecules expressed by chondrocytes, as evidenced by various expressions of cartilage specific proteins and genes. Collagen hydrogel unexpectedly supported chondrogenic differentiation in an environment filled with chondrocytes secretion better than other reinforced scaffolds, which is consistent with the previous experiment in vivo. This result indicated that the environmental sensitivity of a scaffold is important for in vivo chondro-induction. This in vitro scaffold-filter model may be useful as a precursor to investigate the chondro-inducing potential of various scaffolds for cartilage repair.

Material-induced chondrogenic differentiation of mesenchymal stem cells is material-dependent.

Certain materials may mimic natural cartilage to provide an amenable cellular microenvironment for the chondrogenic differentiation of mesenchymal stem cells. The chondrogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) has been demonstrated to be induced by collagen-based hydrogels in vivo, but whether the induction is material-driven or self-differentiation has not been elucidated. In the present study, BMSCs were encapsulated in porous materials, namely, a biphasic calcium phosphate ceramic (BCP), silk fibroin protein matrix (SFP) and collagen sponge (CS), to further study the chondrogenic effects of various materials. Diffusion chambers that allow the body fluid to permeate and deter the host cells from invasion were also loaded with the cell-scaffold constructs. Chambers containing the scaffold-BMSC composites were implanted subcutaneously in the dorsa of rabbits. The specimens in the chamber were harvested for histological and immunohistochemical analyses eight weeks after implantation. The results showed that no chondrogenic differentiation of the BMSCs occurred when the BMSCs were encapsulated in BCP, SFP and CS, indicating that chondrogenesis induced by materials is material-dependent and that these particular porous materials are not suitable for inducing chondrogenesis. However, the diffusion chamber was effective in preventing host immune rejection, host cell invasion and vascular invasion. The results are likely to serve as a valuable clinical reference when selecting an appropriate scaffold for cartilage repair.

An International Randomized Clinical Trial Evaluating Bst-Cargel®, A Novel Scaffold For Cartilage Repair, Demonstrates Superiority of Repair When Compared with Microfracture Alone Treatment

An international RCT with 80 patients evaluating BST-CarGel®, a novel chitosan-based scaffold applied to a cartilage lesion, showed superiority to microfracture alone for both the degree of lesion filling and the quality of cartilage repair tissue at 12 months by quantitative MRI and microscopic analyses of biopsies. Equivalent safety and clinical improvement at 12 months was also demonstrated. An international, multicenter randomized clinical trial was conducted to evaluate the efficacy and safety of BST-CarGel® (Piramal Healthcare, Laval, Canada), for the repair of articular cartilage lesions, compared to microfracture alone. BST-CarGel® is a device composed of the biomaterial chitosan (polyglucosamine) which is applied over a microfractured lesion. 80 patients at international sites (Canada, Spain, South Korea) were randomized (1:1) to either BST-CarGel®+microfracture or microfracture alone. Enrolled patients were aged 18 to 55, with BMIs=30 and had a single, symptomatic, grade III or IV lesion on their femoral condyles. They followed standardized 12-week post-operative rehabilitation. Quantity and quality of cartilage repair tissue was assessed at 12 months using quantitative MRI techniques. Supplemental structural data (ICRS I and II Histology Scoring and Polarized Light Microscopy (PLM) scoring) was also obtained from elective osteochondral biopsies taken at an average of 13 months from 38 consenting patients. Clinical improvement was measured at 12 months using WOMAC and SF-36 questionnaires, and safety was evaluated by monitoring adverse events. Blinded MRI analysis demonstrated that BST-CarGel® met both primary endpoints by achieving statistical superiority for BST-CarGel® in greater % Lesion Fill (p=0.0105) and more cartilage-like repair tissue T2 values (p=0.0330) when compared with microfracture alone. WOMAC assessments for pain, stiffness and function yielded equivalent improvement for both groups at 12 months, which were statistically significant (p<0.0001) from baseline. Safety was comparable for both groups. When compared to microfracture, BST-CarGel® repair tissue showed improved ICRS macroscopic grading (p=0.0002) at second-look arthroscopy, superior collagen organization by PLM (p=0.0003), and improvements in most histological parameters (4/6 ICRS-I and 10/14 ICRS-II, with 8 at (p<0.05) or near significance (p<0.11)). When compared to microfracture alone, the treatment of focal cartilage lesions with BST-CarGel® resulted in greater quantity and superior quality of cartilage repair tissue at 12 months as demonstrated by multiple, independent indicators.

Repair of osteochondral defects with recombinant human type II collagen gel and autologous chondrocytes in rabbit

Recombinant human type II collagen (rhCII) gels combined with autologous chondrocytes were tested as a scaffold for cartilage repair in rabbits invivo. Autologous chondrocytes were harvested, expanded and combined with rhCII-gel and further pre-cultivated for 2weeks prior to transplantation into a 4mm diameter lesion created into the rabbit's femoral trochlea (n=8). Rabbits with similar untreated lesions (n=7) served as a control group. Six months after the transplantation the repair tissue in both groups filled the lesion site, but in the rhCII-repair the filling was more complete. Both repair groups also had high proteoglycan and type II collagen contents, except in the fibrous superficial layer. However, the integration to the adjacent cartilage was incomplete. The O'Driscoll grading showed no significant differences between the rhCII-repair and spontaneous repair, both representing lower quality than intact cartilage. In the repair tissues the collagen fibers were abnormally organized and oriented. No dramatic changes were detected in the subchondral bone structure. The repair cartilage was mechanically softer than the intact tissue. Spontaneously repaired tissue showed lower values of equilibrium and dynamic modulus than the rhCII-repair. However, the differences in the mechanical properties between all three groups were insignificant. When rhCII was used to repair cartilage defects, the repair quality was histologically incomplete, but still the rhCII-repairs showed moderate mechanical characteristics and a slight improvement over those in spontaneous repair. Therefore, further studies using rhCII for cartilage repair with emphasis on improving integration and surface protection are required.

Dermatan sulphate in methoxy polyethylene glycol-polylactide-co-glycolic acid scaffolds upregulates fibronectin gene expression but has no effect on in vivo osteochondral repair

The purpose of the study was to investigate the effect of dermatan sulphate (DS) addition to biodegradable methoxy polyethylene glycol (MPEG) substituted polylactide-co-glycolic acid (PLGA) scaffolds for cartilage repair in vitro and in vivo. Human chondrocytes from eight patients undergoing anterior cruciate ligament reconstruction were isolated and cultured in 5% oxygen on MPEG-PLGA scaffolds±DS for one, three, seven and 14 days. Analyses were performed using quantitative gene expression analysis for chondrogenic and cell attachment markers. An osteochondral drill hole defect was created in the intertrochlear groove of the distal femur in 20 New Zealand white rabbits (defects n=20). When bleeding was observed, the defects were treated with MPEG-PLGA scaffolds±DS. Twelve weeks after surgery the rabbits were sacrificed and the defects were analysed using histological grading with O'Driscoll scoring. DS addition to MPEG-PLGA scaffolds resulted in a significant upregulation of fibronectin gene expression on day 1. No differences were observed in chondrogenic gene expression. There were no differences between the two groups in histological grading (+DS 10.3 and -DS 9.6). Upregulation of fibronectin in vitro indicating early cell-scaffold interaction and attachment did not result in improved cartilage repair in an osteochondral defect model in rabbits.

Tissue engineering applications: cartilage lesions repair by the use of autologous chondrocytes

Promising new therapies based on tissue engineering have been recently developed for cartilage repair. The association of biomaterials with autologous chondrocytes expanded in vitro can represent a useful tool to regenerate this tissue. The scaffolds utilised in such therapeutical applications should provide a pre-formed three-dimensional shape, prevent cells from floating out of the defect, have sufficient mechanical strength, facilitate uniform spread of cells and stimulate the phenotype of transplanted cells. Hyaff-11 is a hyaluronic-acid based biodegradable polymer, that has been shown to provide successful cell carrier for tissue-engineered repair. From our findings we can state that human chondrocytes seeded on Hyaff-11 are able to maintain in vitro the characteristic of differentiated cells, expressing and producing collagen type II and aggrecan which are the main markers of cartilage phenotype, down-regulating collagen type I. Moreover, it seems to be a useful scaffold for cartilage repair both in animal models and clinical trials in humans, favouring the formation of a hyaline-like tissue. In the light of these data, we can hypothesise, for the future, the use of autologous chondrocyte transplantation together with gene therapy as a treatment for rheumatic diseases such as osteoarthritis.

The use of de-differentiated chondrocytes delivered by a heparin-based hydrogel to regenerate cartilage in partial-thickness defects

Partial-thickness cartilage defects, with no subchondral bone injury, do not repair spontaneously, thus there is no clinically effective treatment for these lesions. Although the autologous chondrocyte transplantation (ACT) is one of the promising approaches for cartilage repair, it requires in vitro cell expansion to get sufficient cells, but chondrocytes lose their chondrogenic phenotype during expansion by monolayer culture, leading to de-differentiation. In this study, a heparin-based hydrogel was evaluated and optimized to induce cartilage regeneration with de-differentiated chondrocytes. First, re-differentiation of de-differentiated chondrocytes encapsulated in heparin-based hydrogels was characterized in vitro with various polymer concentrations (from 3 to 20 wt.%). Even under a normal cell culture condition (no growth factors or chondrogenic components), efficient re-differentiation of cells was observed with the optimum at 10 wt.% hydrogel, showing the complete re-differentiation within a week. Efficient re-differentiation and cartilage formation of de-differentiated cell/hydrogel construct were also confirmed in vivo by subcutaneous implantation on the back of nude mice. Finally, excellent cartilage regeneration and good integration with surrounding, similar to natural cartilage, was also observed by delivering de-differentiated chondrocytes using the heparin-based hydrogel in partial-thickness defects of rabbit knees whereas no healing was observed for the control defects. These results demonstrate that the heparin-based hydrogel is very efficient for re-differentiation of expanded chondrocytes and cartilage regeneration without using any exogenous inducing factors, thus it could serve as an injectable cell-carrier and scaffold for cartilage repair. Excellent chondrogenic nature of the heparin-based hydrogel might be associated with the hydrogel characteristic that can secure endogenous growth factors secreted from chondrocytes, which then can promote the chondrogenesis, as suggested by the detection of TGF- β1 in both in vitro and in vivo cell/hydrogel constructs.

A Collagen–Poly(Vinyl Alcohol) Nanofiber Scaffold for Cartilage Repair

Articular cartilage has a limited capacity for self-repair. Untreated injuries of cartilage may lead to osteoarthritis. This problem demands new effective methods to reconstruct articular cartilage. Mesenchymal stem cells (MSCs) have the proclivity to differentiate along multiple lineages giving rise to new bone, cartilage, muscle, or fat. This study was an animal model for autologous effects of transplantation of MSCs with a collagen–poly(vinyl alcohol) (PVA) scaffold into full-thickness osteochondral defects of the stifle joint in the rabbit as an animal model. A group of 10 rabbits had a defect created experimentally in the full thickness of articular cartilage penetrated into the subchondral space in the both stifle joints. The defect in the right stifle was filled with MSCs/collagen–PVA scaffold (group I), and in the left stifle, the defect was left without any treatment as the control group (group II). Specimens were harvested at 12 weeks after implantation, examined histologically for morphologic features, and stained immunohistochemically for type-II collagen. Histology observation showed that the MSCs/collagen–PVA repair group had better chondrocyte morphology, continuous subchondral bone, and much thicker newly formed cartilage compared with the control group at 12 weeks post operation. There was a significant difference in histological grading score between these two groups. The present study suggested that the hybrid collagen–PVA scaffold might serve as a new way to keep the differentiation of MSCs for enhancing cartilage repair.

Chondrogenic differentiation of mesenchymal stem cells induced by collagen‐based hydrogel: An in vivo study

Chondrogenic differentiation of mesenchymal stem cells (MSCs) relies on inductive media of chondrogenic environment. With proper design, a cellular microenvironment mimicking chondrogenic environment might be created to induce chondrogenesis of MSCs. In this study, bone marrow mesenchymal cells (BMSCs) were encapsulated in collagen-based hydrogel, and then enclosed in diffusion-chambers which allow the body fluid to permeate and preclude the host cells to invade. Then, the chamber with the hydrogel-BMSCs composite was implanted in the back of rabbits subcutaneously. The specimens in the chamber were harvested for histological, immunohistochemical, and RT-PCR analyses after 8 weeks. The results showed that cells with the characteristic of chondrocytes were homogenously distributed and the extracellular matrix (ECM) of cartilage has been secreted, indicating the chondrogenic differentiation of BMSCs. As control, nothing was obtained with only BMSCs. Moreover, the expression of collagen type II, indicator of cartilage ECM, was less in tissues with collagen-alginate-hydrogel (CAH) than that with collagen-hydrogel (CH). The results showed that both CH and CAH may induce the chondrogenesis and the induction is materials dependent. From in vitro experiments, TGF-beta is a necessary signal molecule for chondrogenesis, and it was suggested that the material may take in vivo growth factors to trigger chondrogenesis. From the studies, the chondrogenic induction of the hydrogel may be ascribed to that the hydrogel may provide a suitable environment and aggregate the signal molecule for chondrogenesis in vivo. The results would lend valuable reference in clinical for selection of appropriate scaffold for cartilage repair.

P116 Novel ECM-based Scaffolds for Cartilage Repair

P116 Novel ECM-based Scaffolds for Cartilage Repair

A Peek Into the Possible Future of Management of Articular Cartilage Injuries: Gene Therapy and Scaffolds for Cartilage Repair

Two rapidly progressing areas of research will likely contribute to cartilage repair procedures in the foreseeable future: gene therapy and synthetic scaffolds. Gene therapy refers to the transfer of new genetic information to cells that contribute to the cartilage repair process. This approach allows for manipulation of cartilage repair at the cellular and molecular level. Scaffolds are the core technology for the next generation of autologous cartilage implantation procedures in which synthetic matrices are used in conjunction with chondrocytes. This approach can be improved further using bioreactor technologies to enhance the production of extracellular matrix proteins by chondrocytes seeded onto a scaffold. The resulting "neo-cartilage implant" matures within the bioreactor, and can then be used to fill cartilage defects.

Bovine primary chondrocyte culture in synthetic matrix metalloproteinase-sensitive poly(ethylene glycol)-based hydrogels as a scaffold for cartilage repair.

A poly(ethylene glycol) (PEG)-based hydrogel was used as a scaffold for chondrocyte culture. Branched PEG-vinylsulfone macromers were end-linked with thiol-bearing matrix metalloproteinase (MMP)-sensitive peptides (GCRDGPQGIWGQDRCG) to form a three-dimensional network in situ under physiologic conditions. Both four- and eight-armed PEG macromer building blocks were examined. Increasing the number of PEG arms increased the elastic modulus of the hydrogels from 4.5 to 13.5 kPa. PEG-dithiol was used to prepare hydrogels that were not sensitive to degradation by cell-derived MMPs. Primary bovine calf chondrocytes were cultured in both MMP-sensitive and MMP-insensitive hydrogels, formed from either four- or eight-armed PEG. Most (>90%) of the cells inside the gels were viable after 1 month of culture and formed cell clusters. Gel matrices with lower elastic modulus and sensitivity to MMP-based matrix remodeling demonstrated larger clusters and more diffuse, less cell surface-constrained cell-derived matrix in the chondron, as determined by light and electron microscopy. Gene expression experiments by real-time RT-PCR showed that the expression of type II collagen and aggrecan was increased in the MMP-sensitive hydrogels, whereas the expression level of MMP-13 was increased in the MMP-insensitive hydrogels. These results indicate that cellular activity can be modulated by the composition of the hydrogel. This study represents one of the first examples of chondrocyte culture in a bioactive synthetic material that can be remodeled by cellular protease activity.