Bone marrow aspirate concentrate (BMAC), together with fibrin glue (Tisseel, Baxter, UK) and Hyaluronic acid (HA) were used as a one-step cell therapy treating patients with ankle cartilage defects in our hospital. This therapy was proven to be safe, with patients demonstrating a significant improvement 12 months post-treatment. Enriched mesenchymal stem cells (MSCs) in BMAC are suggested inducers of cartilage regeneration, however, currently there is no point-of-care assessment for BMAC quality; especially regarding the proportion of MSCs within. This study aims to characterise the cellular component of CCR-generated BMAC using a point-of-care device, and to investigate if the total nucleated cell (TNC) count and patient age are predictive of MSC concentration. During surgery, 35ml of bone marrow aspirate (BMA) was collected from each patients’ iliac crest under anaesthesia, and BMAC was obtained via a commercial kit (Cartilage Regeneration kit, CCR, Innotec®, UK). BMAC was then mixed with thrombin (B+T) for injection with HA and fibrinogen. In our study, donor-matched BMA, BMAC and B+T were obtained from consented patients (n=12, age 41 ± 16years) undergoing surgery with BMAC therapy. TNC, red blood cell (RBC) and platelet (PLT) counts were measured via a haematology analyser (ABX Micros ES 60, Horiba, UK), and the proportion of MSCs in BMA, BMAC and B+T were assessed via colony forming unit-fibroblast (CFU-F) assays. Significant differences data in matched donors were tested using Friedman test. All data were shown as mean ± SD.Abstract
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Methods
Meniscus allograft and synthetic meniscus scaffold (Actifit®) transplantation have shown promising outcomes for symptoms relief in patients with meniscus deficient knees. Untreated chondral defects can place excessive load onto meniscus transplants and cause early graft failure. We hypothesised that combined ACI and allograft or synthetic meniscus replacement might provide a solution for meniscus deficient individuals with co-existing lesions in cartilage and meniscus. We retrospectively collected data from 17 patients (16M, 1F, aged 40±9.26) who had ACI and meniscus allograft transplant (MAT), 8 patients (7M, 1F, aged 42±11) who underwent ACI and Actifit® meniscus scaffold replacement. Other baseline data included BMI, pre-operative procedures and cellular transplant data. Patients were assessed by pre-operative, one-year and last follow-up Lysholm score, one-year repair site biopsy, MRI evaluations.Abstract
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Structural and functional outcome of bone graft with first or second generation autologous chondrocyte implantation (ACI) in osteochondral defects has not been reported. 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.Background
Methods
Extended expansion of cells derived from equine articular cartilage reveal maintenance of chondrogenic potency and no evidence of senescence up to 100 population doublings. The data suggests the reclassification of these cells from progenitor cells to stem cells. One sign of ‘in vitro aging’ is the diminishing capacity for cell division. In contrast to embryonic stem cells that show no loss of proliferative potency, the maximal population doublings (PD) for mesenchymal stem cells (MSCs) in vitro is reported to be between 30 and 40 replications 1,2,3. We have isolated a population of chondroprogenitor cells from articular cartilage of several species, including equine4. These cells have demonstrated functional equivalence in their differentiation capacity when compared with MSCs but have the advantage of retaining the highly desirable stable (permanent) chondrocyte phenotype. In this study, we examined the age-related capacity of these cells for extended division and retention of potency.Summary Statement
Introduction
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.
Osteoarthritis (OA) is the most common form of joint disease leading to disability and dependence. In severe cases of knee OA, the joint is deemed irrecoverable and total knee replacements are indicated. Tissue engineering is a possible solution for this pathology and previous work from our laboratory has demonstrated that it is possible to isolate and expand chondroprogenitor cells in vitro from healthy knee-joint articular cartilage. Work presented here describes the detection and isolation of chondroprogenitor cells derived from osteoarthritic cartilage following total knee replacement in patients with severe OA, suggesting a pool of viable cells from this degenerate region which has been previously deemed non-recoverable. Human articular cartilage was excised from tibial plateaux (TP's) obtained from total knee replacements following the diagnoses of severe OA. Cells were isolated by a sequential pronase and collagenase digestion and subject to a fibronectin adhesion assay. Cells were expanded in monolayer in supplemented growth medium. Clonal 3D pellet cultures were established in chondrogenic and osteogenic differentiation media. Adipogenic cultures were also established in monolayer cultures. Histological procedures, immunohistochemistry and molecular biology were undertaken in order to determine the extent of differentiation. In addition, osteochondral plugs were excised from the TP's and wax embedded for further histological and immunohistochemical analysis. Clonal cell lines obtained from osteoarthritic knee-joint cartilage using the fibronectin adhesion assay were isolated and successfully cultured to a maximum of 60 population doublings whilst still demonstrating a chondrogenic capacity. Three-D pellet cultures after 21 days of chondrogenic induction produced smooth and iridescent pellets which stained positively for toluidine blue and safranin O. Positive labelling for collagen type II and aggrecan were also observed. Following osteogenic induction; evidence of mineralisation was indicated by the von Kossa stain. Adipogenic induction revealed a positive result. Osteochondral plugs demonstrated sporadic positive labelling in the surface region for putative stem cell marker Stro-1. Chondroprogenitor cells isolated from osteoarthritic display a strong chondrogenic phenotype, and have the ability to be induced into different lineages. These findings suggest the presence of a pool of viable chondroprogenitors from osteoarthritic tissue which was otherwise deemed irrecoverable.
A novel scoring system for the grading of osteoarthritis has been developed. Scoring systems for the measurement of Osteoarthritis (OA) are essential for the understanding of the osteoarthritic process. OA is a mutifactorial degenerative joint disease affecting not only hyaline cartilage but also the surrounding tissues and particularly the subchondral bone. It as questionable as to why the articular cartilage remains the sole component used for histopathological assessment. The intimate relationship between the subchondral bone and overlying cartilage provide major difficulty in their independent measurement. A new scoring system has been developed to incorporate the subchondral bone into the assessment process and relating it to the structure of the overlying hyaline cartilage, which together permit a more accurate description of the degree of degenerate change. The new scoring system was developed from the analysis of 26 operative specimens from tibial plateau (TP) from patients who underwent total knee replacement (TKR). Multiple osteochondral plugs were taken from weight-bearing regions of the whole TP. The specimens were fixed and decalcified before being sectioned and stained with Masson's trichrome. Using a standard imaging system (Photoshop) the areas of bone and hyaline cartilage were identified and measured. Further parameters 1) cartilage thickness 2) tidemark integrity, 3) surface integrity 4) cartilage morphology were measured using a numeric measurement scale. The scoring system indicated a relationship between the area of subchondral bone and the hyaline cartilage degeneration. The overall sum of scores was also successful in distinguishing between the milder and more severe samples of OA. More comprehensive inter and intra observer variability needs to be tested in order validate the system. Quantifying changes to the subchondral bone may also serve beneficial to clinicians, as it is possible that monitoring these changes clinically could lead to early identification of OA.
