In osteoarthritis, chondrocytes acquire a
This review is aimed at clinicians appraising
preclinical trauma studies and researchers investigating compromised bone
healing or novel treatments for fractures. It categorises the clinical
scenarios of poor healing of fractures and attempts to match them
with the appropriate animal models in the literature. We performed an extensive literature search of animal models
of long bone fracture repair/nonunion and grouped the resulting
studies according to the clinical scenario they were attempting
to reflect; we then scrutinised them for their reliability and accuracy
in reproducing that clinical scenario. Models for normal fracture repair (primary and secondary), delayed
union, nonunion (atrophic and hypertrophic), segmental defects and
fractures at risk of impaired healing were identified. Their accuracy
in reflecting the clinical scenario ranged greatly and the reliability
of reproducing the scenario ranged from 100% to 40%. It is vital to know the limitations and success of each model
when considering its application.
Osteoarthritis (OA) is the most common joint disease, which is characterized by a progressive loss of proteoglycans and the destruction of extracellular matrix (ECM), leading to a loss of cartilage integrity and joint function. During OA development, chondrocytes alter ECM synthesis and change their gene expression profile including upregulation of
Bone healing outcome is highly dependent on the initial mechanical fracture environment [1]. In vivo, direct bone healing requires absolute stability and an interfragmentary strain (IFS) below 2% [2]. In the majority of cases, however, endochondral ossification is engaged where frequency and amplitude of IFS are key factors. Still, at the cellular level, the influence of those parameters remains unknown. Understanding the regulation of naïve hMSC differentiation is essential for developing effective bone healing strategies. Human bone-marrow-derived MSC (KEK-ZH-NR: 2010–0444/0) were embedded in 8% gelatin methacryol. Samples (5mm Ø x 4mm) were subjected to 0, 10 and 30% compressive strain (5sec compression, 2hrs pause sequence for 14 days) using a multi-well uniaxial bioreactor (RISystem) and in presence of chondro-permissive medium (CP, DMEM HG, 1% NEAA, 10 µM ITS, 50 µg/mL ascorbic acid, and 100 mM Dex). Cell differentiation was assessed by qRT-PCR and histo-/immunohistology staining. Experiments were repeated 5 times with cells from 5 donors in duplicate. ANOVA with Tukey post-hoc correction or Kurskal-Wallis test with Dunn's correction was used. Data showed a strong upregulation of
During OA the homeostasis of healthy articular chondrocytes is dysregulated, which leads to a phenotypical transition of the cells, further influenced by external stimuli. Chondrocytes sense those stimuli, integrate them at the intracellular level and respond by modifying their secretory and molecular state. This process is controlled by a complex interplay of intracellular factors. Each factor is influenced by a myriad of feedback mechanisms, making the prediction of what will happen in case of external perturbation challenging. Hampering the
Introduction. Cell-based therapy is needed to overcome the lacking intrinsic ability of cartilage to heal. Generating cartilage tissue from human bone marrow-derived stromal cells (MSC) is limited by up-regulation of COL10, ALP and other hypertrophy markers in vitro and calcifying cartilage at heterotopic sites in vivo. MSC
Integrin α2β1 is one of the major transmembrane receptors for fibrillary collagen. In native bone we could show that the absence of this protein led to a protective effect against age-related osteoporosis. The objective of this study was to elucidate the effects of integrin α2β1 deficiency on fracture repair and its underlying mechanisms. Standardised femoral fractures were stabilised by an intramedullary nail in 12 week old female C57Bl/6J mice (wild type and integrin α2. -/-. ). After 7, 14 and 28 days mice were sacrificed. Dissected femura were subjected to µCT and histological analyses. To evaluate the biomechanical properties, 28-day-healed femura were tested in a torsional testing device. Masson goldner staining, Alizarin blue, IHC and IF staining were performed on paraffin slices. Blood serum of the animals were measured by ELISA for BMP-2. Primary osteoblasts were analysed by in/on-cell western technology and qRT-PCR. Integrin α2β1 deficient animals showed earlier transition from cartilaginous callus to mineralized callus during fracture repair. The shift from chondrocytes over
Virtual mechanical testing is a method for measuring bone healing using finite element models built from computed tomography (CT) scans. Previously, we validated a dual-zone material model for ovine fracture callus that differentiates between mineralized woven bone and soft tissue based on radiodensity. 1. The objective of this study was to translate the dual-zone material model from sheep to two important clinical scenarios: human tibial fractures in early-stage healing and late-stage nonunions. CT scans for N = 19 tibial shaft fractures were obtained prospectively at 12 weeks post-op. A second group of N = 33 tibial nonunions with CT scans were retrospectively identified. The modeling techniques were based on our published method. 2. The dual-zone material model was implemented for humans by performing a cutoff sweep for both the 12-week and nonunion groups. Virtual torsional rigidity (VTR) was calculated as VTR = ML/φ [N-m. 2. /°], where M is the moment reaction, L is the diaphyseal segment length, and φ is the angle of twist. As the soft tissue cutoff was increased, the rigidity of the clinical fractures decreased and soft tissue located within the fracture gaps produced higher strains that are not predicted without the dual zone approach. The structural integrity of the nonunions varied, ranging from very low rigidities in atrophic cases to very high rigidities in highly calcified
The signaling molecule prostaglandin E2 (PGE2), synthesized by cyclooxygenase-2 (COX-2), is immunoregulatory and reported to be essential for skeletal stem cell function. Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used in osteoarthritis (OA) analgesia, but cohort studies suggested that long-term use may accelerate pathology. Interestingly, OA chondrocytes secrete high amounts of PGE2. Mesenchymal stromal cell (MSC) chondrogenesis is an in vitro OA model that phenocopies PGE2 secretion along with a
INTRODUCTION. The generation of cartilage from progenitor cells for the purpose of cartilage repair is often hampered by unwanted
Introduction. Human Mesenchymal stem cells (hMSCs) are a promising source for articular cartilage repair. Unfortunately, under in vitro conditions, chondrogenically differentiated hMSCs have the tendency to undergo hypertrophy similar to growth plate chondrocytes. Retinoic acid (RA) signalling plays a key role in growth plate hypertrophy. Whilst RA agonists block chondrogenesis and foster hypertrophy during later stages, RAR inverse agonists (IA) enhance chondrogenesis when applied early in culture. Therefore, we hypothesized that treatment with RAR IA will attenuate hypertrophy in chondrogenically differentiated hMSCs. To test this hypothesis, we analysed early (initial chondrogenic differentiation) and late treatment (hypertrophy stage) of hMSCs with an RAR IA. Methods. Pellets of passage 2 hMSCs were formed in V-bottom well plates by centrifugation and pre-differentiated in a chemically defined medium containing 10ng/mL TGFß (CM+) for 14 days. Thereafter, pellets were cultured for an additional 14 days under 6 conditions: CM+, CM- (w/out TGFß), and
Design criteria for tissue-engineered materials in regenerative medicine include robust biological effectiveness, off-the-shelf availability, and scalable manufacturing under standardized conditions. For bone repair, existing strategies rely on primary autologous cells, associated with unpredictable performance, limited availability and complex logistic. Here, we report the manufacturing of engineered and devitalized human
Nonunions occur in situations with interrupted fracture healing process and indicate conditions where the fracture has no potential to heal without further intervention. Per definition, no healing is detected nine months post operation and there is no visible progress of healing over the last three months. The classification of nonunions as
INTRODUCTION. The generation of cartilage from progenitor cells for the purpose of cartilage repair is often hampered by unwanted ossification of the generated tissue due to endochondral ossification. Our in vitro data show that celecoxib is able to suppress the
Osteoarthritis (OA) is the most common degenerative joint disease causing joint immobility and chronic pain. Treatment is mainly based on alleviating pain and reducing disease progression. During OA progression the chondrocyte undergoes a
INTRODUCTION. Endochondral ossification in the growth plate is directly responsible for skeletal growth and its de novo bone-generating activity. Growth plates are vulnerable to disturbances that may lead to abnormal skeletal development. Non-steroidal anti-inflammatory drugs (NSAIDs) are widely used analgesics but have been reported to impair endochondral ossification-driven fracture healing. Despite the general awareness that NSAIDs affect endochondral ossification, the consequences of NSAIDs on skeletal development are unknown. We hypothesise that the NSAID celecoxib leads to impaired growth plate development and consequently impairs skeletal development. METHODS. Healthy skeletally immature (5 weeks old) C57BL/6 mice were treated for ten weeks with celecoxib (daily oral administration 10 mg/kg) or placebo (water) (institutional approval 2013–094) (n=12 per group). At 15 weeks postnatally, total growth plate thickness, the thickness of specific growth plate zones, (immuno)histological analysis of extracellular matrix composition in the growth plate, cell number and cell size, longitudinal bone growth and bone micro-architecture by micro-CT were analysed. Inhibition of COX-2 activity was confirmed by determining PGE2 levels in plasma using an ELISA. RESULTS. No significant difference in total growth plate thickness or thickness of the resting zone, proliferative or
Introduction and Objective. Current cartilage repair strategies lack adequate tissue integration capacity and often present mechanical failure at the graft-to-host tissue junction. The design of multilayered osteochondral tissue engineering (TE) constructs is an attractive approach to overcome these problems. However, calcium ion-release from resorbable bone-replacement materials was suggested to compromise chondrogenic differentiation of adjacent cartilage tissue and it is unclear whether articular chondrocytes (AC) or mesenchymal stroma cells (MSC) are more sensitive to such conditions. Aim of the study was to compare how elevated calcium levels affect cartilage matrix production during re-differentiation of AC versus chondrogenic differentiation of MSC. The results of this study will help to identify the ideal cell source for growth of neocartilage adjacent to a calcified bone replacement material for design of multilayered osteochondral TE approaches. Materials and Methods. Expanded human AC and MSC (6–12 donors per group) were seeded in collagen type I/III scaffolds and cultured under standard chondrogenic conditions at control (1.8mM) or elevated (8.0mM) CaCl2 for 35 days. Proteoglycan and collagen production were assessed via radiolabel-incorporation, ELISA, qPCR and Western blotting. Differences between groups or cell types were calculated using the non-parametric Wilcoxon or Mann-Whitney U test, respectively, with p < 0.05 considered significant. Results. Elevated calcium significantly reduced GAG synthesis (63% of control, p=0.04) and chondrogenic marker expression of AC, lowering the GAG/DNA content (47% of control, p=0.004) and collagen type II deposition (24% of control, p=0.05) of neocartilage compared to control conditions. Opposite, at elevated calcium levels MSC-derived chondrocytes significantly increased GAG synthesis (130% of control, p=0.02) and collagen type II content (160% of control, p=0.03) of cartilage compared to control tissue. Chondrogenic and
Heterotopic ossi?cation is the abnormal formation of bone in soft tissues and is a frequent complication of hip replacement surgery. Heterotopic ossi?cations are described to develop via endochondral ossification and standard treatment is administration of indomethacin. It is currently unknown how indomethacin influences heterotopic ossi?cation on a molecular level, therefore we aimed to determine whether indomethacin might influence heterotopic ossi?cation via impairing the chondrogenic phase of endochondral ossification. ATDC5, human bone marrow stem cells (hBMSCs) and rabbit periosteal agarose cultures were employed as progenitor cell models; SW1353, human articular chondrocytes and differentiated ATDC5 cells were used as matured chondrocyte cell models. All cells were cultured in the presence of (increasing) concentrations of indomethacin. The action of indomethacin was confirmed by decreased PGE2 levels in all experiments, and was determined by specific PGE2 ELISA. Gene- and protein expression analyses were employed to determine chondrogenic outcome. Progenitor cell models differentiating in the chondrogenic lineage (ATDC5, primary human bone marrow stem cells and ex vivo periosteal agarose cultures) were treated with increasing concentrations of indomethacin and a dose-dependent decrease in gene- and protein expression of chondrogenic and
Dysplasia Epiphysealis Hemimelica (DEH) also known as Trevor's Disease is a rare developmental disorder resulting in cartilaginous overgrowth of the epiphysis of long bones. DEH is usually diagnosed in children between two and eight years old and it is three times more often diagnosed in boys. The most reported complaints are pain, limitation in range of motion, and deformity or swelling of the affected joint. Treatment of symptomatic lesions consists of surgical resection of the lesion, resulting in good long-term results. Based on histological evaluation, DEH is often described as an osteochondroma or an osteochondroma-like lesion, although there are clinical, radiological and genetic differences between DEH and osteochondromas. To investigate the hypothesis that DEH and osteochondromas are histologically identical, two cases of DEH and two cases of osteochondromas in patients with Hereditary Multiple Osteochondroma (HMO) are compared at histological level. Tissue samples from patients with a histopathologically confirmed diagnosis of DEH were compared with two age and gender matched patients diagnosed with HMO. After tissue sampling and processing, (immuno)histological stainings were performed for Collagen type II, Collagen type X, Sox-9 and Safranin-O. Histologically, clumping of chondrocytes in a fibrillar matrix, a thick disorganized cartilage cap and ossification centres with small amounts of unresorbed cartilage were observed in DEH. In contrast, chondrocyte organisation in cartilage of osteochondromas displays characteristics of the normal growth plate. In addition, differences in expression of collagen type II, collagen type X and Sox9 were observed. Collagen type II was expressed in the extracellular matrix surrounding proliferative and
Growth plates taken from five- to 20-week-old Japanese white rabbits were immunostained for c-Myc protein. This was localised both in the proliferating zone and upper