Allogeneic Chondrocytes Research Articles

Tissue-engineered constructs: the effect of scaffold architecture in osteochondral repair

Cartilage has a poor regenerative capacity. Tissue-engineering approaches using porous scaffolds seeded with chondrocytes may improve cartilage repair. The aim of this study was to examine the effect of pore size and pore interconnectivity on cartilage repair in osteochondral defects treated with different scaffolds seeded with allogenic chondrocytes. Scaffolds consisting of 55 wt% poly(ethylene oxide terephthalate) and 45 wt% poly(butylene terephthalate) (PEOT/PBT) with different pore sizes and interconnectivities were made, using a compression moulding (CM) and a three-dimensional fibre (3DF) deposition technique. In these scaffolds, allogenic chondrocytes were seeded, cultured for 3 weeks and implanted in osteochondral defects of skeletally mature rabbits. At 3 weeks no difference in cartilage repair between an empty osteochondral defect, CM or 3DF scaffolds was found. Three months post-implantation, cartilage repair was significantly improved after implantation of a 3DF scaffold compared to a CM scaffold. Although not significant, Mankin scores for osteoarthritis (OA) indicated less OA in the 3DF scaffold group compared to empty defects and CM-treated defects. It is concluded that scaffold pore size and pore interconnectivity influences osteochondral repair and a decreased pore interconnectivity seems to impair osteochondral repair.

Initial phase I safety of retrovirally transduced human chondrocytes expressing transforming growth factor-beta-1 in degenerative arthritis patients

Background aims. TissueGene-C (TG-C) represents a cell-mediated gene therapy for localized delivery of allogeneic chondrocytes expressing transforming growth factor (TGF)-β1 directly to the damaged knee joint. Untransduced human chondrocytes (hChonJ cells) have also been incorporated into the TG-C product at a 3:1 ratio with TGF-β1-expressing chondrocytes (hChonJb#7) in order to help fill in the defect and as target cells for the actions of the expressed TGF-β1.Methods.A phase I dose-escalating clinical trial was performed to evaluate the safety and biologic activity of TG-C in patients with advanced osteoarthritis of the knee joint (full thickness cartilage defect) that was refractory to existing non-operative therapies. Following a single intra-articular injection into the joint space of the damaged knee, patients were monitored for safety, and an evaluation was performed to assess the pharmacokinetics and biologic activity of TG-C.Results.There were no treatment-related serious adverse events. Swelling, effusion and minor localized reactions such as warming sensation or itching were observed in a dose-dependent manner at the injection site. Knee evaluation scores seemed to indicate a dose-dependent trend toward efficacy; however, patient numbers were not sufficient to determine statistical significance.Conclusions.Overall, there were no significant safety issues related to the administration of TG-C, with only some minor injection site reactions observed. Additionally, knee scoring analyzes indicated a possibility that TG-C may contribute to improvement of arthritic symptoms. More study is warranted to evaluate further the safety and determine the potential efficacy of TG-C.

Treatment of subchondral cystic lesions of the medial femoral condyle of mature horses with growth factor enhanced chondrocyte grafts: A retrospective study of 49 cases

To evaluate the long-term clinical outcome after allogeneic chondrocyte and insulin-like growth factor-I (IGF-I) grafting of subchondral cystic lesions (SCLs) of the femoral condyle in horses. To test the hypothesis that chondrocyte and IGF-I grafts will improve the long-term clinical outcome in arthroscopically debrided SCLs. Medical records of 49 horses with SCLs of the femoral condyle treated by debridement and implantation of chondrocytes and IGF-I were reviewed. Preoperative radiographs were obtained, and caudocranial radiographic projections were used to establish a ratio between cyst and femoral condyle size. Arthroscopic cyst debridement followed by filling of the bone void with autologous cancellous bone (45 horses) or tricalcium phosphate granules (4 horses) was performed. A paired syringe containing a fibrinogen and chondrocyte mixture in one syringe and calcium-activated bovine thrombin with IGF-I in the other was used to cover the surface. A successful outcome was defined as a horse that performed to its intended use without lameness. A successful outcome was achieved in 36 of 49 horses (74%). Preoperative radiography was performed in all horses, with 33 horses having unilateral SCLs of the medial femoral condyle, 15 horses having bilateral SCLs of the medial femoral condyle, and one horse having bilateral SCLs of the lateral femoral condyle. Median age of the horses was 3.3 years. Fifteen horses had preoperative radiographic and arthroscopic evidence of osteoarthritis (OA). A successful outcome was not influenced by age of horse, presence of pre-existing osteoarthritis or preoperative size of the subchondral cyst. Grafting resulted in success for 80% of horses >3 years old, and in 80% of horses with OA. Implantation of allogeneic chondrocytes supplemented with IGF-I is an effective treatment for horses with SCLs of the femoral condyle, and particularly for older horses and horses with pre-existing osteoarthritis. Chondrocyte implantation may offer a greater chance of long-term success in older horses and horses with osteoarthritis than has been previously reported with cyst debridement alone.

Midterm Results of the Treatment of Cartilage Defects in the Knee Using Alginate Beads Containing Human Mature Allogenic Chondrocytes

Background: The treatment of chondral lesions is still an important challenge for the orthopaedic surgeon. Attempts have been made to restore cartilage lesions by filling the defects with a temporary biocompatible matrix. Purpose: The authors present their midterm experience with the implantation of alginate beads containing human mature allogenic chondrocytes for the treatment of cartilage lesions in the knee. Study Design: Case series; Level of evidence, 4. Methods: A biodegradable, alginate-based biocompatible scaffold containing human mature allogenic chondrocytes was used for the treatment of cartilage lesions in the knee. Twenty-one patients were clinically prospectively evaluated with use of the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) and a visual analog scale (VAS). The mean follow-up time was 6.3 years (range, 5-8 years). Magnetic resonance imaging (MRI) data were analyzed based on the MOCART (Magnetic Resonance Observation of Cartilage Repair Tissue) system, allowing morphologic assessment of the repair tissue. Magnetic resonance images were taken at 1 year of follow-up and at a mean follow-up of 6.1 years (range, 5-7 years). Results: During the follow-up period, the WOMAC and VAS scores improved significantly. No signs of clinical deterioration or adverse reactions to the alginate beads/allogenic chondrocyte implantation were observed. Four failures occurred during the follow-up period in this study (19.05%). The MOCART scores were moderate and remained stable in time. Conclusion: This investigation provided useful information on the efficacy of the implantation of alginate beads containing human mature allogenic chondrocytes for the treatment of cartilage lesions in the knee. The midterm clinical outcome of the presented technique was satisfactory. However, these results were not confirmed by the MRI findings.

The effect of stress and tissue fluid microenvironment on allogeneic chondrocytes in vivo and the immunological properties of engineered cartilage

Engineered implants derived from neonatal rabbit chondrocytes and collagen type I hydrogel, were loaded in dialyzer pockets and implanted in muscle and articular cavity of rabbits to simulate different stress and tissue fluid micro-environments. After 4 and 12 weeks, the expressions of main histocompatibility complex (MHC) molecules as well as the mixed lymphocyte chondrocytes reactions (MLChR) levels of the seeded cells were detected. The results indicated that with stress and synovial fluid microenvironment, the formation of chondroid tissue was prominently promoted in articular cavity. It gave the seeded chondrocytes lower and gradually decreasing levels of allogeneic lymphocytes activation, however, with the higher cell mortality, the MHC molecules expression, especially MHC-I were up-regulated obviously in early stage. These results are very different to those seen in muscle and prove that stress and tissue fluid micro-environments can greatly impact the differentiation and immunological properties of the engineered cartilage. From the perspective of avoiding severe rejection, to promote the formation of the matrix as fast and select scaffold with higher “isolation” ability may be meaningful. Furthermore, the suitably treated dialyzer pockets model can be used for the study of the differentiation and immunological properties of the tissue engineered cartilage.

In VitroandIn VivoCharacterization of Nonbiomedical- and Biomedical-Grade Alginates for Articular Chondrocyte Transplantation

Alginate is a key hydrogel for cartilage tissue engineering. Here, we systematically evaluated four biomedical- and two nonbiomedical-grade alginates for their capacity to support the in vitro culture and in vivo transplantation of articular chondrocytes. Chondrocytes in all ultrapure alginates maintained high cell viability. Spheres composed of biomedical-grade, low-viscosity, high-mannuronic acid content alginate showed the lowest decrease in size over time. Biomedical-grade, low-viscosity, high-guluronic acid content alginate allowed for optimal cell proliferation. Biomedical-grade, medium-viscosity, high-mannuronic acid content alginate promoted the highest production of proteoglycans. When transplanted into osteochondral defects in the knee joint of sheep in vivo, empty spheres were progressively surrounded by a granulation tissue. In marked contrast with these observations, all alginate spheres carrying allogeneic chondrocytes were gradually invaded by a granulation tissue containing multinucleated giant cells, lymphocytes, and fibroblasts, regardless whether they were based on biomedical- or nonbiomedical-grade alginates. After 21 days in vivo, transplanted chondrocytes were either viable or underwent necrosis, and apoptosis played a minor role in their early fate. The individual characteristics of these alginates may be valuable to tailor specific experimental and clinical strategies for cartilage tissue engineering.

Reattachment of the articular cartilage component of type 1 subchondral cystic lesions of the medial femoral condyle with polydioxanone pins in 3 horses

3 horses were referred for treatment of subchondral cystic lesions of 1 or both medial femoral condyles. All horses had clinically apparent lameness confirmed to be due to a radiographically evident subchondral cystic lesion of the medial femoral condyle with a large articular component (> 15 mm) and shallow subchondral depth (< 10 mm). Arthroscopic assessment of affected cartilage revealed undulating cartilage with a relatively smooth surface and extensive residual perimeter attachment. Resorbable polydioxanone pins were used arthroscopically to reattach the cartilage overlying the subchondral cystic lesions. A biologic graft (bone marrow aspirate concentrate or allogeneic chondrocytes) was injected into the depths of the cystic cavity following cartilage reattachment. Follow-up examination confirmed radiographic resolution of the lesion and elimination of clinical signs within the treated femorotibial joint. Lesions with a large area of affected articular cartilage have been associated with a decreased rate of return to athletic function following arthroscopic debridement, likely secondary to the loss of subchondral architecture and the production of imperfect fibrocartilage repair. Salvage of the affected cartilage in a select population of horses with progressively expanding but shallow subchondral cystic lesions of the medial femoral condyle is possible and may improve radiographic and clinical outcome.

In Vivo Transplantation of Neonatal Ovine Neocartilage Allografts – Determining the Effectiveness of Tissue Transglutaminase

Following transplantation of ovine neocartilage allografts, 26 sheep were divided into groups according to the following weight-bearing schedule: 8-week nonweight bearing (8NWB, n=14), and 8-week nonweight bearing+4-week weight bearing (8NWB+4WB, n=12). In addition, 7 and 6 sheep, respectively, in the 8NWB and 8NWB+4WB groups received tTG treatment after allograft transplantation, whereas the remaining 13 sheep in these groups did not receive tTG. Finally, 8 sheep served as sham-operated controls without allograft transplantation. After euthanasia, stifle joints were harvested for the analysis of gross appearance, chondrocyte viability, histology, and biomechanical testing. No significant differences were noted in macroscopic graft survival and union with host tissue in both 8NWB and 8NWB+4WB groups between the tTG treated and non-tTG treated animals. Analysis of histological scores demonstrated no significant difference between tTG and non-tTG treatments in both 8NWB and 8NWB+4WB groups. Confocal laser microscopic analysis of the explanted defects revealed 70%-100% cell viability in all treatment groups. This study shows that allogeneic chondrocytes harvested from neonatal donors provide sufficient metabolic activity to affect repair. Use of tTG to augment resorbable suture fixation of neocartilage grafts provided no advantage over suture alone in this pilot study.

Treatment of Cartilage Defects in the Knee Using Alginate Beads Containing Human Mature Allogenic Chondrocytes

Background The repair of osteochondral lesions is imperfect and transient; chondral lesions do not heal in mature cartilage. Attempts have been made to restore cartilage lesions by filling the defects with a temporary artificial biocompatible matrix. Purpose To determine whether the implantation of alginate beads containing human mature allogenic chondrocytes is feasible and safe for the treatment of symptomatic cartilage defects in the knee. Study Design Case series; Level of evidence, 4. Methods A biodegradable, alginate-based, biocompatible scaffold containing human mature allogenic chondrocytes was used for the treatment of chondral and osteochondral lesions in the knee. Twenty-one patients were clinically and prospectively evaluated with use of the Western Ontario and McMaster Universities Osteoarthritis Index and a visual analog scale for pain preoperatively and at 3, 6, 9, 12, 18, and 24 months of follow-up. Of the 21 patients, 13 consented to having a biopsy sample taken for investigative purposes from the area of implantation at 12 months of follow-up, allowing histologic assessment of the repair tissue. Results A statistically significant clinical improvement became apparent after 6 months, and patients improved during the 24 months of follow-up. Adverse reactions to the alginate/fibrin matrix seeded with the allogenic cartilage cells were not observed. Histologic analysis of the biopsy specimens rated the repair tissue as hyaline-like in 15.3% of the samples, as mixed tissue in 46.2%, as fibrocartilage in 30.8%, and as fibrous in 7.7%. Conclusion The results of this short-term pilot study show that the alginate-based scaffold containing human mature allogenic chondrocytes is feasible and safe for the treatment of symptomatic cartilage defects of the knee. The described technique provides clinical and histologic outcomes that are equal but not superior to those of other cartilage repair techniques.

Effects of RNA interference-mediated inhibition of aggrecanase-1 and aggrecanase-2 on rat costochondral chondrocytes in vitro

Background and Objectives: Failure of transplanted cartilage or allogenic chondrocytes is attributed mainly to immunologic rejection and cartilage degradation. A major feature is loss of aggrecan from the cartilage matrix, primarily due to the action of the specific proteinases aggrecanase-1 and aggrecanase-2. The aim of this in vitro study was to determine whether specific inhibition of aggrecanase-1 and aggrecanase-2 by RNA interference (RNAi) would mitigate aggrecan loss from cultured chondrocytes.

MRI evaluation of a new scaffold-based allogenic chondrocyte implantation for cartilage repair

Aim The present study was designed to evaluate the implantation of alginate beads containing human mature allogenic chondrocytes for the treatment of symptomatic cartilage defects of the knee. MRI was used for the morphological analysis of cartilage repair. The correlation between MRI findings and clinical outcome was also studied. Methods A biodegradable, alginate-based biocompatible scaffold containing human mature allogenic chondrocytes was used for the treatment of symptomatic chondral and osteochondral lesions in the knee. Twenty-one patients were prospectively evaluated with use of the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) and the Visual Analogue Scale (VAS) for pain preoperatively and at 3, 6, 9 and 12 months of follow-up. Of the 21 patients, 12 had consented to follow the postoperative MRI evaluation protocol. MRI data were analyzed based on the original MOCART (Magnetic Resonance Observation of Cartilage Repair Tissue) and modified MOCART scoring system. The correlation between the clinical outcome and MRI findings was evaluated. Results A statistically significant clinical improvement became apparent after 6 months and patients continued to improve during the 12 months of follow-up. One of the two MRI scoring systems that were used, showed a statistically significant deterioration of the repair tissue at 1 year of follow-up. Twelve months after the operation complete filling or hypertrophy was found in 41.6%. Bone-marrow edema and effusion were seen in 41.7% and 25% of the study patients, respectively. We did not find a consistent correlation between the MRI criteria and the clinical results. Discussion The present study confirmed the primary role of MRI in the evaluation of cartilage repair. Two MOCART-based scoring systems were used in a longitudinal fashion and allowed a practical and morphological evaluation of the repair tissue. However, the correlation between clinical outcome and MRI findings was poor. Further validation of these scoring systems is mandatory. The promising short-term clinical outcome of the allogenic chondrocytes/alginate beads implantation was not confirmed by the short-term MRI findings.

Success Rates and Immunologic Responses of Autogenic, Allogenic, and Xenogenic Treatments to Repair Articular Cartilage Defects

This review examines current approaches available for articular cartilage repair, not only in terms of their regeneration potential, but also as a function of immunologic response. Autogenic repair techniques, including osteochondral plug transplantation, chondrocyte implantation, and microfracture, are the most widely accepted clinical treatment options due to the lack of immunogenic reactions, but only moderate graft success rates have been reported. Although suspended allogenic chondrocytes are shown to evoke an immune response upon implantation, allogenic osteochondral plugs and tissue-engineered grafts using allogenic chondrocytes exhibit a tolerable immunogenic response. Additionally, these repair techniques produce neotissue with success rates approaching those of currently available autogenic repair techniques, while simultaneously obviating their major hindrance of donor tissue scarcity. To date, limited research has been performed with xenogenic tissue, although several studies demonstrate the potential for its long-term success. This article focuses on the various treatment options for cartilage repair and their associated success rates and immunologic responses.

Human Amniotic Mesenchymal Cells Differentiate into Chondrocytes

Recently, cartilage diseases have been treated by auto- or allogenic chondrocyte transplantation. However, such treatments are limited by the necessity of having a large amount of cells for transplantation, the risk of rejection, and donor shortage. Since the human amnion is immune-privileged tissue suitable for allotransplantation, the potential of human amniotic mesenchymal cells (HAMc) to differentiate into chondrocytes was assessed. The expression of gene encoding transcription factors SOXs and bone morphogenetic proteins (BMPs) as well as BMP receptors were assessed. Chondrocyte phenotype was characterized by positive expression of the cartilage marker genes collagen type II and aggrecan by RT-PCR, collagen type II protein were analyzed by immunofluorescence analysis. HAMc expressed chondrocyte-related genes, including SOXs, BMPs, as well as BMP receptors. Collagen type II and aggrecan were detected after the induction of chondrogenesis with BMP-2. HAMc, transplanted into noncartilage tissue of mice with BMP-2, or implanted with collagen-scaffold into the defects generated in a rat's bone, underwent morphological changes with deposition of collagen type II. These results showed that HAMc have the potential to differentiate into chondrocytes in vitro and in vivo, suggesting that they have therapeutic potential for the treatment of damaged or diseased cartilage.

Production of Allogenic Cartilage in a Tube Lined With Epithelium and a Novel Scaffold

To study the feasibility of engineering cartilage tissue in a tube lined with epithelium and implanting allogenic chondrocytes in a novel scaffold that is made of chitosan nonwoven cloth coated with poly(DL-lactide-co-glycolide). Laboratory research. Allogenic chondrocytes were obtained from the auricles of 1-month-old New Zealand white rabbits. After the cells were cultured in vitro for 3 or 4 passages, they were implanted in the scaffolds to form composite grafts and then transplanted into the rabbits. After 6, 12, and 18 weeks, the general histologic characteristics were investigated. The cobweb-like matrix was observed approximately 1 week after the chondrocytes had been implanted in the scaffolds. At 6 weeks, the matrix was secreted, and there were immature chondrocytes in the grafts. At 12 weeks, the allogenic cartilage in the tube lined with epithelium had been created. Chondrocytes were almost mature and the lacunae had formed. At 18 weeks, the neocartilage was similar to native cartilage. It is feasible to produce allogenic cartilage in a tube lined with epithelium by implanting allogenic chondrocytes into a novel scaffold made of chitosan nonwoven cloth coated with poly(DL-lactide-co-glycolide).

Cartilage Repair: Synthetics and Scaffolds

Symptomatic articular cartilage lesions have gained attention and clinical interest in recent years and can be difficult to treat. Historically, various biologic surgical treatment options have yielded inconsistent results because of the inferior biomechanical properties associated with a variable healing response. Improving technology and surgical advances has generated considerable research in cartilage resurfacing and optimizing hyaline tissue restoration. Biologic innovation and tissue engineering in cartilage repair have used matrix scaffolds, autologous and allogenic chondrocytes, cartilage grafts, growth factors, stem cells, and genetic engineering. Numerous evolving technologies and surgical approaches have been introduced into the clinical setting. This review will discuss the basic science, surgical techniques, and clinical outcomes of novel synthetic materials and scaffolds for articular cartilage repair.

Evaluation of articular cartilage repair using biodegradable nanofibrous scaffolds in a swine model: a pilot study

The aim of this study was to evaluate a cell-seeded nanofibrous scaffold for cartilage repair in vivo. We used a biodegradable poly(epsilon-caprolactone) (PCL) nanofibrous scaffold seeded with allogeneic chondrocytes or xenogeneic human mesenchymal stem cells (MSCs), or acellular PCL scaffolds, with no implant as a control to repair iatrogenic, 7 mm full-thickness cartilage defects in a swine model. Six months after implantation, MSC-seeded constructs showed the most complete repair in the defects compared to other groups. Macroscopically, the MSC-seeded constructs regenerated hyaline cartilage-like tissue and restored a smooth cartilage surface, while the chondrocyte-seeded constructs produced mostly fibrocartilage-like tissue with a discontinuous superficial cartilage contour. Incomplete repair containing fibrocartilage or fibrous tissue was found in the acellular constructs and the no-implant control group. Quantitative histological evaluation showed overall higher scores for the chondrocyte- and MSC-seeded constructs than the acellular construct and the no-implant groups. Mechanical testing showed the highest equilibrium compressive stress of 1.5 MPa in the regenerated cartilage produced by the MSC-seeded constructs, compared to 1.2 MPa in the chondrocyte-seeded constructs, 1.0 MPa in the acellular constructs and 0.2 MPa in the no-implant group. No evidence of immune reaction to the allogeneically- and xenogeneically-derived regenerated cartilage was observed, possibly related to the immunosuppressive activities of MSCs, suggesting the feasibility of allogeneic or xenogeneic transplantation of MSCs for cell-based therapy. Taken together, our results showed that biodegradable nanofibrous scaffolds seeded with MSCs effectively repair cartilage defects in vivo, and that the current approach is promising for cartilage repair.

Effects of RNAi-mediated inhibition of aggrecanase-1 and aggrecanase-2 on rat costochondral chondrocytesin vitro1

Failure of transplanted cartilage or allogenic chondrocytes is attributed mainly to immunological rejection and cartilage degradation. A major feature is the loss of aggrecan from the cartilage matrix, primarily due to the action of the specific proteinases aggrecanase-1 and aggrecanase-2. The aim of this in vitro study was to determine whether the specific inhibition of aggrecanase-1 and aggrecanase-2 by RNAi would mitigate aggrecan loss from cultured chondrocytes. Expression plasmid vectors of shRNA targeting aggrecanase-1 and aggrecanase-2 were constructed and transfected into cultured rattus costochondral chondrocytes. The transfected cells were induced with interleukin-1beta (IL-1beta). Gene mRNA levels were analyzed by RT-PCR. Aggrecan and collagen II content were measured by immunohistochemistry and Western blotting. As the chondrocytes underwent dedifferentiation, aggrecanase-1 increased significantly. The specific inhibition of aggrecanase-1 and aggrecanase-2 by RNAi had no negative effect on the morphology and growth velocity of the chondrocytes. The mRNA of aggrecanase-1 and aggrecanase-2 decreased significantly. The alpha-2-macroglobulin expression level was increased by the shRNA specific for aggrecanase-1. Other genes of the chondrocytic extracellular matrix were not affected. RNAi significantly increased the aggrecan and collagen II content of chondrocytes treated with IL-1beta. The results suggest that inhibition of aggrecanase-1 and aggrecanase-2 by RNAi can mitigate aggrecan degradation, without interfering with chondrocytic gene phenotype recovery. RNAi technology can be a useful tool for studying degenerative processes in cartilage.

Evaluation of three-dimensional scaffolds prepared from poly(3-hydroxybutyrate- co-3-hydroxyhexanoate) for growth of allogeneic chondrocytes for cartilage repair in rabbits

Articular cartilage repair using tissue engineering approach generally requires the use of an appropriate scaffold architecture that can support the formation of cartilage tissue. In this investigation, the potential of three-dimensional scaffolds made of poly(3-hydroxybutyrate- co-3-hydroxyhexanoate) (PHBHHx) was evaluated in rabbit articular cartilage defect model. Engineered PHBHHx cartilage constructs inoculated in vitro with rabbit chondrocytes for 30 days were examined. Subsequently the constructs inoculated with chondrocytes for 10 days were selected for transplantation into rabbits. After 16 weeks of in vivo implantation, both the engineered cartilage constructs and the bare scaffolds were found to be filled the defects with white cartilaginous tissue, with the engineered constructs showing histologically good subchondral bone connection and better surrounding cartilage infusion. Owing to pre-seeded chondrocytes in the PHBHHx scaffolds, better surface integrality and more accumulation of extracellular matrix (ECM) including type II collagen and sGAG were achieved in the engineered cartilage constructs. The repaired tissues possessed an average compressive modulus of 1.58 MPa. For comparison, the defects without repair treatments still showed defects with fibrous tissues. These results demonstrated that PHBHHx is a useful material for cartilage tissue engineering.

Growing cartilage for human replacement—where are we?

Articular cartilage injuries predispose to osteoartritis development in the joint and the intrinsic repair capacity of adult hyaline cartilage is limited. Over the years, several surgical techniques have been introduced to improve the repair of injured articular cartilage surfaces, but articular cartilage resurfacing remains a scientific challenge. The use of cultured autologous chondrocytes in patients with cartilage injuries have shown encouraging clinical results in combination with the regeneration of hyaline cartilage [1]. This work started with animal studies in the US in the 1980s [2] and continued in Gothenburg, Sweden, with research on human articular chondrocytes and the potential use in human cell therapy. At that time, other chondrogeneic cells such as auricular chondrocytes, nasal chondrocytes, rib perichondrial cells, allogeneic chondrocytes or mesenchymal bone marrow stromal cells could have been alternative cell sources, but the clinical reality was the driving force behind the decision to use articular cartilage. Since the clinical diagnosis is conducted by arthroscopic inspection and thus the surgeon has sterile access to the joint cavity, there is an opportunity to harvest cartilage conveniently. Other sources of cartilage were less likely to be used, either for practical reasons that other areas of the body had to be cleaned and sterilized (e.g. rib cartilage) or due to the fact that donor site morbidity would give the patient additional suffering. The reason for the in vitro chondrocyte culture is to increase the cell number from an original few 100,000 to several million cells. Chondrocytes are isolated from small slices of cartilage harvested transarthroscopically from a minor weight-bearing area in the injured knee. The amount of cartilage harvested is only a few hundred milligrams. The extracellular matrix is removed from the cartilage by enzymatic digestion and the average cell yield is between 2,000 and 3,000 cells/mg biopsy weight; subsequently, cells are expanded in monolayer culture in a defined cell culture medium with a 10% addition of the patient’s own serum. The knowledge gained over the years has demonstrated that during the expansion of chondrocytes in vitro there is a selection of a limited number of chondrocyte progenitor clones and the culture process allows the cells to revert to a more primitive foetal stage through a process of dedifferentiation. Interestingly, the cellular gene programme activated during the redifferentiation process is similar to programs controlling chondrogenesis in foetal life [3]. Once a suitable number of cells has been reached, the increased number of chondrocytes are suspended in culture medium and then implanted into the area of cartilage defect using a periosteal patch over the defect as a method of cell containment. The closed compartment initiates a process of cell condensation and cartilage neo-formation similar to the mesenchymal condensation seen in the foetal stage of limb formation.

Chondrocyte apoptosis in the regenerated articular cartilage after allogenic chondrocyte transplantation in the rabbit knee

We attempted to repair full-thickness defects in the articular cartilage of the trochlear groove of the femur in 30 rabbit knee joints using allogenic cultured chondrocytes embedded in a collagen gel. The repaired tissues were examined at 2, 4, 8, 12 and 24 weeks after operation using histological and histochemical methods. The articular defect filling index measurement was derived from safranin-O stained sections. Apoptotic cellular fractions were derived from analysis of apoptosis in situ using TUNEL staining, and was confirmed using caspase-3 staining along with quantification of the total cellularity. The mean articular defect filling index decreased with time. After 24 weeks it was 0.7 (SD 0.10), which was significantly lower than the measurements obtained earlier (p < 0.01). The highest mean percentage of apoptotic cells were observed at 12 weeks, although the total cellularity decreased with time. Because apoptotic cell death may play a role in delamination after chondrocyte transplantation, anti-apoptotic gene therapy may protect transplanted chondrocytes from apoptosis.