In this study a combination of autologous chondrocyte implantation (ACI) and the osteochondral autograft transfer system (OATS) was used and evaluated as a treatment option for the repair of large areas of degenerative articular cartilage. We present the results at three years post-operatively. Osteochondral cores were used to restore the contour of articular cartilage in 13 patients with large lesions of the lateral femoral condyle (n = 5), medial femoral condyle (n = 7) and patella (n = 1). Autologous cultured chondrocytes were injected underneath a periosteal patch covering the cores. After one year, the patients had a significant improvement in their symptoms and after three years this level of improvement was maintained in ten of the 13 patients. Arthroscopic examination revealed that the osteochondral cores became well integrated with the surrounding cartilage. We conclude that the hybrid ACI/OATS technique provides a promising surgical approach for the treatment of patients with large degenerative osteochondral defects

An increasing number of patients are treated by autologous chondrocyte implantation (ACI). This study tests the hypothesis that culture within a defined chondrogenic medium containing TGF-β enhances the reexpression of a chondrocytic phenotype and the subsequent production of cartilaginous extracellular matrix by human chondrocytes used in ACI. Chondrocytes surplus to clinical requirements for ACI from 24 patients were pelleted and cultured in either DMEM (Dulbecco’s modified eagles medium)/ITS+Premix/TGF-β1 or DMEM/10%FCS (fetal calf serum) and were subsequently analysed biochemically and morphologically. Pellets cultured in DMEM/ITS+/TGF-β1 stained positively for type-II collagen, while those maintained in DMEM/10%FCS expressed type-I collagen. The pellets cultured in DMEM/ITS+/TGF-β1 were larger and contained significantly greater amounts of DNA and glycosaminoglycans. This study suggests that the use of a defined medium containing TGF-β is necessary to induce the re-expression of a differentiated chondrocytic phenotype and the subsequent stimulation of glycosaminoglycan and type-II collagen production by human monolayer expanded chondrocytes

We compared the quality of debridement of chondral lesions performed by four arthroscopic (SH, shaver; CU, curette; SHCU, shaver and curette; BP, bipolar electrodes) and one open technique (OPEN, scalpel and curette) which are used prior to autologous chondrocyte implantation (ACI). The ex vivo simulation of all five techniques was carried out on six juvenile equine stifle joints. The OPEN, SH and SHCU techniques were tested on knees harvested from six adult human cadavers. The most vertical walls with the least adjacent damage to cartilage were obtained with the OPEN technique. The CU and SHCU methods gave inferior, but still acceptable results whereas the SH technique alone resulted in a crater-like defect and the BP method undermined the cartilage wall. The subchondral bone was severely violated in all the equine samples which might have been peculiar to this model. The predominant depth of the debridement in the adult human samples was at the level of the calcified cartilage. Some minor penetrations of the subchondral end-plate were induced regardless of the instrumentation used. Our study suggests that not all routine arthroscopic instruments are suitable for the preparation of a defect for ACI. We have shown that the preferred debridement technique is either open or arthroscopically-assisted manual curettage. The use of juvenile equine stifles was not appropriate for the study of the cartilage-subchondral bone interface

Osteoarthritis (OA) is a progressive and degenerative joint disease resulting in changes to articular cartilage. In focal early OA defects, autologous chondrocyte implantation (ACI) has a 2-fold failure rate due to poor graft integration and presence of inflammatory factors (e.g. Interleukin-1β). Bone marrow derived mesenchymal stem cells (MSCs) are an alternative cell source for cell-based treatments due to their chondrogenic capacity, though in vivo implantation leads to bone formation. In vivo, chondrocytes reside under an oxygen tension between 2–7% oxygen or physioxia. Physioxia enhances MSC chondrogenesis with reduced hypertrophic marker (collagen X and MMP13) expression compared to hyperoxic conditions (20% oxygen). This study sought to understand whether implantation of physioxic preconditioned MSCs improves cartilage regeneration in an early OA defect model compared to hyperoxic MSCs. Bone marrow extracted from New Zealand white rabbits (male: 5–6 months old; n = 6) was split equally for expansion under 2% (physioxia) or 20% (hyperoxia) oxygen. Chondrogenic pellets (2 × 105 cells/pellet) formed at passage 1 were cultured in the presence of TGF-β1 under their expansion conditions and measured for their wet weight and GAG content after 21 days. During bone marrow extraction, a dental drill (2.5mm diameter) was applied to medial femoral condyle on both the right and left knee and left untreated for 6 weeks. Following this period, physioxia and hyperoxia preconditioned MSCs were seeded into a hyaluronic acid (TETEC) hydrogel. Fibrous tissue was scraped and then MSC-hydrogel was injected into the right (hyperoxic MSCs) and left (physioxia MSCs) knee. Additional control rabbits with drilled defects had fibrous tissue scrapped and then left untreated without MSC-hydrogel treatment for the duration of the experiment. Rabbits were sacrificed at 6 (n = 3) and 12 (n = 3) weeks post-treatment, condyles harvested, decalcified in 10% EDTA and sectioned using a cryostat. Region of interest was identified; sections stained with Safranin-O/Fast green and evaluated for cartilage regeneration using the Sellers scoring system by three blinded observers. Physioxic culture of rabbit MSCs showed significantly shorter doubling time and greater cell numbers compared to hyperoxic culture (∗p < 0.05). Furthermore, physioxia enhanced MSC chondrogenesis via significant increases in pellet wet weight and GAG content (∗p < 0.05). Implantation of physioxic preconditioned MSCs showed significantly improved cartilage regeneration (Mean Sellers score = 7 ± 3; ∗p < 0.05) compared to hyperoxic MSCs (Sellers score = 12 ± 2) and empty defects (Sellers score = 17 ± 3). Physioxia enhances in vitro rabbit MSC chondrogenesis. Subsequent in vivo implantation of physioxia preconditioned MSCs improved cartilage regeneration in an early OA defect model compared to hyperoxic MSCs. Future studies will investigate the mechanisms for enhanced in vivo regeneration using physioxia preconditioned MSCs

Background. Structural and functional outcome of bone graft with first or second generation autologous chondrocyte implantation (ACI) in osteochondral defects has not been reported. Methods. Seventeen patients (mean age of 27±7 years, range 17–40), twelve with osteochondritis dissecans (OD) (ICRS Grade 3 and 4) and five with isolated osteochondral defect (OCD) (ICRS Grade 4) were treated with a combined implantation of a unicortical autologous bone graft with ACI (the Osplug technique). Functional outcome was assessed with Lysholm scores. The repair site was evaluated with the Oswestry Arthroscopy Score (OAS), MOCART score and ICRS II histology score. Formation of subchondral lamina and lateral integration of the bone grafts were evaluated from MRI scans. Results. The mean defect size was 4.5±2.6SD cm. 2. (range 1–9) and depth was 11.3±5SD mm (range 5–18). The pre-operative Lysholm score improved from 45 (IQR 24, range 16–79) to 77 (IQR 28, range 41–100) at 1 year (p-value 0.001) and 70 (IQR 35, range 33–91) at 5 years (p-value 0.009). The mean OAS of the repair site was 6.2 (range 0–9) at a mean of 1.3 years. The mean MOCART score was 61 ± 22SD (range 20–85) at 2.6 ± 1.8SD years. Histology demonstrated generally good integration of the repair cartilage with the underlying bone. Poor lateral integration of the bone graft on the MRI scan and a low OAS were significantly associated with a poor Lysholm score and failure. Conclusion. Osplug technique shows significant improvement of functional outcome for up to 5 years in patients with a high grade OD or OCD. This is the first report describing association of bone graft integration with functional outcome after such a procedure. It also demonstrates histological evidence of integration of the repair cartilage with the underlying bone graft. Level of Evidence. III

Background. Treatment of cartilage defects requires in vitro expansion of human articular chondrocytes (HACs) for autologous chondrocyte implantation (ACI). During standard expansion culture (i.e. plasma osmolarity, 280 mOsm) chondrocytes inevitably lose their specific phenotype (i.e. collagen type II (COL2) expression). This de-differentiation makes them inappropriate for ACI. Physiological osmolarity (i.e. 380 mOsm) improves COL2 expression in vitro, but the underlying reason is unknown. However, an accepted key regulator of chondrocyte differentiation, transforming growth factor beta (TGFβ), is known to stimulate COL2 production. In this study we aimed to elucidate if TGFβ signaling could potentially be driving the COL2 expression under physiological culture conditions. Material and methods. After informed consent was obtained, HACs were isolated from five osteoarthritis (OA) patients and cultured in cytokine-free medium of 280 or 380 mOsm, respectively, under standard 2D in vitro conditions with or without lentiviral TGFβ2 knockdown (RNAi). Expression of TGFβ isoforms, superfamily receptors and chondrocyte marker genes was evaluated by qRT-PCR, TGFβ2 protein secretion by ELISA and TGFβ bioactivity using luciferase reporter assays. Statistical significance was assessed by a student's t-test. Results. TGFβ isoform expression was differentially altered by physiological osmolarity. Specifically, 380 mOsm increased TGFβ2 expression and protein secretion, as well as TGFβ activity. Upon TGFβ2 isoform-specific knockdown COL2 expression was induced. Physiological osmolarity and TGFβ2 RNAi also induced TGFβ1, TGFβ3 and their type I receptor ALK5. Conclusions. We showed that TGFβ2 knockdown increases COL2 expression in human osteoarthritic chondrocytes in vitro, possibly through a regulatory feedback loop involving TGFβ1, TGFβ3 induction and an increased ALK5/ALK1 ratio. This study indicates that TGFβ signalling is involved in osmolarity-induced chondrocyte marker gene expression. Pharmacological targeting of this pathway holds potential to further improve future osmolarity-mediated phenotypic stabilisation in advanced cell-based cartilage repair strategies. Level of Evidence. preclinical. Disclosure. We have nothing to disclose

In order to determine the usefulness of MRI in assessing autologous chondrocyte implantation (ACI) the first 57 patients (81 chondral lesions) with a 12-month review were evaluated clinically and with specialised MRI at three and 12 months. Improvement 12 months after operation was found subjectively (37.6 to 51.9) and in knee function levels (from 85% International Cartilage Repair Society (ICRS) III/IV to 61% I/II). The International Knee Documentation Committee (IKDC) scores showed an initial deterioration at three months (56% IKDC A/B) but marked improvement at 12 months (88% A/B). The MRI at three months showed 82% of patients with at least 50% defect fill, 59% with a normal or nearly normal signal at repair sites, 71% with a mild or no effusion and 80% with a mild or no underlying bone-marrow oedema. These improved at 12 months to 93%, 93%, 94% and 91%, respectively. The overall MR score at 12 months suggested production of normal or nearly normal cartilage in 82%, corresponding to a subjective improvement in 81% of patients and 88% IKDC A/B scores. Second-look surgery and biopsies in 15 patients (22 lesions) showed a moderate correlation of MRI with visual scoring; 70% of biopsies showed hyaline and hyaline-like cartilage. Thus, MRI at 12 months is a reasonable non-invasive means of assessment of ACI

Introduction. Autologous chondrocyte implantation (ACI) is contra-indicated in a joint rendered unstable by a ruptured anterior cruciate ligament (ACL). We present our experience of ACI repair with ACL reconstruction. Methods. Patients underwent arthroscopic examination and cartilage harvesting of the knee. A second operation was undertaken approximately six weeks later to repair the ruptured ACL with hamstring graft or Bone patella-Bone (BPB) and to implant the chondrocytes via formal arthrotomy. Three groups were assessed: Group 1: Simultaneous ACL Reconstruction and ACI; Group 2: Previous ACL Reconstruction with subsequent ACI repair; Group 3: Previously proven partial or complete ACL rupture, deemed stable and not treated with reconstruction with ACI procedure subsequently. Patients then underwent a graduated rehabilitation program and were reviewed using three functional measurements: Bentley functional scale, the modified Cincinnati rating system, and pain measured on a visual analogue scale. All patients also underwent formal clinical examination at review. Results. Those who underwent simultaneous ACL Reconstruction and ACI had a 47% improvement in Bentley functional scale, 36% improvement in visual analogue score and 38% improvement in the modified Cincinnati rating system. This is in contrast to only a 15% improvement in the modified Cincinnati rating system, 30% improvement in Bentley functional scale, and 32% improvement in visual analogue score in patients who had ACI repair after previous ACL reconstruction. 68% of patients who had the procedures simultaneously rated their outcome as excellent/good and 27% felt it was a failure. In contrast 38% of patients rated their outcome as a failure if they had ACI repair without reconstruction of ACL rupture. Conclusion. Symptomatic cartilage defects and ACL deficiency may co-exist in many patients and represent a treatment challenge. Our results suggest that a combined ACL and ACI repair is a viable option in this group of patients and should reduce the anaesthetic and operative risks of a two-stage repair. Patients with complete rupture of ACL despite being deemed stable performed poorly at review and our study suggests all complete ruptures regardless of stability should be treated with a reconstruction when performing an autologous chondrocyte implantation

Autologous chondrocyte implantation (ACI) has been used for many years for the treatment of symptomatic defects in articular joints, predominantly the knee. Traditionally, cells were implanted behind a periosteal membrane, but in more recent times Chondrogide, a membrane consisting of porcine collagens I and III, has been used. There have been trials comparing the clinical outcome of these two groups of patients; in this study we compare the histological outcome using the two different patch types. In a study of 100 patients having received ACI treatment of cartilage defects in the knee, 41 received Chondrogide (ACI-C) and 59 received periosteum (ACI-P). All of these patients had a post-operative biopsy taken at a mean of 16.9±9.2 months and 20.8±23.2 months for ACI-C and ACI-P respectively for histology using the ICRS II scoring system. Lysholm scores, a measure of knee function, were obtained pre- and post-operatively at the time of biopsy and statistical differences tested for via a Mann-Whitney U-test. The mean age of the two groups at treatment was 37±8 and 35±10 years, the size of defect treated was 6.1±5.4 and 4.4±2.7 cm2 and the biopsy follow-up time was 50.6±22.2 and 81.2±34.8 months for ACI-C and ACI-P patients respectively. Both groups exhibited a significant improvement in Lysholm score from pre-operative to the time of biopsy (14.3±25.7; n=100), although there was no significant difference in improvement in Lysholm score between the two patch types. There was no significant difference between the histology score of the two groups, nor was the score found to correlate with the Lysholm score at that time. The individual components of the ICRS II score did not differ significantly with patch type (even for the surface architecture) apart from cellular morphology which was 6.5±3 and 8.2±1.6 for ACI-C and ACI-P respectively. The histological quality of repair tissue formed with ACI-C differed little from that seen with ACI-P, despite the former group being biopsied ∼4 months sooner after treatment and being used to treat defects which were 39% larger. Hence Chondrogide appears just as suitable as periosteum for use as a patch in the procedure of ACI

Objectives

The aim of this study was to investigate the effect of granulocyte-colony stimulating factor (G-CSF) on mesenchymal stem cell (MSC) proliferation in vitro and to determine whether pre-microfracture systemic administration of G-CSF (a bone marrow stimulant) could improve the quality of repaired tissue of a full-thickness cartilage defect in a rabbit model.

Objectives

Mesenchymal stem cells have the ability to differentiate into various cell types, and thus have emerged as promising alternatives to chondrocytes in cell-based cartilage repair methods. The aim of this experimental study was to investigate the effect of bone marrow derived mesenchymal stem cells combined with platelet rich fibrin on osteochondral defect repair and articular cartilage regeneration in a canine model.

The treatment of osteochondral lesions and osteoarthritis remains an ongoing clinical challenge in orthopaedics. This review examines the current research in the fields of cartilage regeneration, osteochondral defect treatment, and biological joint resurfacing, and reports on the results of clinical and pre-clinical studies. We also report on novel treatment strategies and discuss their potential promise or pitfalls. Current focus involves the use of a scaffold providing mechanical support with the addition of chondrocytes or mesenchymal stem cells (MSCs), or the use of cell homing to differentiate the organism’s own endogenous cell sources into cartilage. This method is usually performed with scaffolds that have been coated with a chemotactic agent or with structures that support the sustained release of growth factors or other chondroinductive agents. We also discuss unique methods and designs for cell homing and scaffold production, and improvements in biological joint resurfacing. There have been a number of exciting new studies and techniques developed that aim to repair or restore osteochondral lesions and to treat larger defects or the entire articular surface. The concept of a biological total joint replacement appears to have much potential.

Cite this article: Bone Joint Res 2013;2:193–9.

The aim of this study was to evaluate the cultivation potential of cartilage taken from the debrided edge of a chronic lesion of the articular surface. A total of 14 patients underwent arthroscopy of the knee for a chronic lesion on the femoral condyles or trochlea. In addition to the routine cartilage biopsy, a second biopsy of cartilage was taken from the edge of the lesion. The cells isolated from both sources underwent parallel cultivation as monolayer and three-dimensional (3D) alginate culture. The cell yield, viability, capacity for proliferation, morphology and the expressions of typical cartilage genes (collagen I, COL1; collagen II, COL2; aggrecan, AGR; and versican, VER) were assessed. The cartilage differentiation indices (COL2/COL1, AGR/VER) were calculated. The control biopsies revealed a higher mean cell yield (1346 cells/mg vs 341 cells/mg), but similar cell proliferation, viability and morphology compared with the cells from the edge of the lesion. The cartilage differentiation indices were superior in control cells: COL2/COL1 (threefold in biopsies (non-significant)); sixfold in monolayer cultures (p = 0.012), and 7.5-fold in hydrogels (non-significant), AGR/VER (sevenfold in biopsies (p = 0.04), threefold (p = 0.003) in primary cultures and 3.5-fold in hydrogels (non-significant)).

Our results suggest that the cultivation of chondrocytes solely from the edges of the lesion cannot be recommended for use in autologous chondrocyte implantation.

Implantation of autologous chondrocytes and matrix autologous chondrocytes are techniques of cartilage repair used in the young adult knee which require harvesting of healthy cartilage and which may cause iatrogenic damage to the joint. This study explores alternative sources of autologous cells.

Chondrocytes obtained from autologous bone-marrow-derived cells and those from the damaged cartilage within the lesion itself are shown to be viable alternatives to harvest-derived cells. A sufficient number and quality of cells were obtained by the new techniques and may be suitable for autologous chondrocyte and matrix autologous chondrocyte implantation.

Perilesional changes of chronic focal osteochondral defects were assessed in the knees of 23 sheep. An osteochondral defect was created in the main load-bearing region of the medial condyle of the knees in a controlled, standardised manner. The perilesional cartilage was evaluated macroscopically and biopsies were taken at the time of production of the defect (T0), during a second operation one month later (T1), and after killing animals at three (T3; n = 8), four (T4; n = 8), and seven (T7; n = 8) months. All the samples were histologically assessed by the International Cartilage Repair Society grading system and Mankin histological scores. Biopsies were taken from human patients (n = 10) with chronic articular cartilage lesions and compared with the ovine specimens. The ovine perilesional cartilage presented with macroscopic and histological signs of degeneration. At T1 the International Cartilage Repair Society ‘Subchondral Bone’ score decreased from a mean of 3.0 (sd 0) to a mean of 1.9 (sd 0.3) and the ‘Matrix’ score from a mean of 3.0 (sd 0) to a mean of 2.5 (sd 0.5). This progressed further at T3, with the International Cartilage Repair Society ‘Surface’ grading, the ‘Matrix’ grading, ‘Cell Distribution’ and ‘Cell Viability’ grading further decreasing and the Mankin score rising from a mean of 1.3 (sd 1.4) to a mean of 5.1 (sd 1.6). Human biopsies achieved Mankin grading of a mean of 4.2 (sd 1.6) and were comparable with the ovine histology at T1 and T3.

The perilesional cartilage in the animal model became chronic at one month and its histological appearance may be considered comparable with that seen in human osteochondral defects after trauma.

Ovine articular chondrocytes were isolated from cartilage biopsy and culture expanded in vitro. Approximately 30 million cells per ml of cultured chondrocytes were incorporated with autologous plasma-derived fibrin to form a three-dimensional construct. Full-thickness punch hole defects were created in the lateral and medial femoral condyles. The defects were implanted with either an autologous ‘chondrocyte-fibrin’ construct (ACFC), autologous chondrocytes (ACI) or fibrin blanks (AF) as controls. Animals were killed after 12 weeks. The gross appearance of the treated defects was inspected and photographed. The repaired tissues were studied histologically and by scanning electron microscopy analysis.

All defects were assessed using the International Cartilage Repair Society (ICRS) classification. Those treated with ACFC, ACI and AF exhibited median scores which correspond to a nearly-normal appearance. On the basis of the modified O’Driscoll histological scoring scale, ACFC implantation significantly enhanced cartilage repair compared to ACI and AF. Using scanning electron microscopy, ACFC and ACI showed characteristic organisation of chondrocytes and matrices, which were relatively similar to the surrounding adjacent cartilage.

Implantation of ACFC resulted in superior hyaline-like cartilage regeneration when compared with ACI. If this result is applicable to humans, a better outcome would be obtained than by using conventional ACI.