Three-Dimensional Quantitative Morphometric Analysis (QMA) for In Situ Joint and Tissue Assessment of Osteoarthritis in a Preclinical Rabbit Disease Model

This work utilises advances in multi-tissue imaging, and incorporates new metrics which define in situ joint changes and individual tissue changes in osteoarthritis (OA). The aims are to (1) demonstrate a protocol for processing intact animal joints for microCT to visualise relevant joint, bone and cartilage structures for understanding OA in a preclinical rabbit model, and (2) introduce a comprehensive three-dimensional (3D) quantitative morphometric analysis (QMA), including an assessment of reproducibility. Sixteen rabbit joints with and without transection of the anterior cruciate ligament were scanned with microCT and contrast agents, and processed for histology. Semi-quantitative evaluation was performed on matching two-dimensional (2D) histology and microCT images. Subsequently, 3D QMA was performed; including measures of cartilage, subchondral cortical and epiphyseal bone, and novel tibio-femoral joint metrics. Reproducibility of the QMA was tested on seven additional joints. A significant correlation was observed in cartilage thickness from matching histology-microCT pairs. The lateral compartment of operated joints had larger joint space width, thicker femoral cartilage and reduced bone volume, while osteophytes could be detected quantitatively. Measures between the in situ tibia and femur indicated an altered loading scenario. High measurement reproducibility was observed for all new parameters; with ICC ranging from 0.754 to 0.998. In conclusion, this study provides a novel 3D QMA to quantify macro and micro tissue measures in the joint of a rabbit OA model. New metrics were established consisting of: an angle to quantitatively measure osteophytes (σ), an angle to indicate erosion between the lateral and medial femoral condyles (ρ), a vector defining altered angulation (λ, α, β, γ) and a twist angle (τ) measuring instability and tissue degeneration between the femur and tibia, a length measure of joint space width (JSW), and a slope and intercept (m, Χ) of joint contact to demonstrate altered loading with disease progression, as well as traditional bone and cartilage and histo-morphometry measures. We demonstrate correlation of microCT and histology, sensitive discrimination of OA change and robust reproducibility.     

Micro CT and Micro MR imaging of 3D architecture of animal skeleton

Quantitative assessment of three-dimensional (3D) trabecular structural characteristics may improve our ability to understand the pathophysiology of osteoporosis, to test the efficacy of pharmaceutical intervention, and to estimate bone biomechanical properties. Considerable progress has been made in advanced imaging techniques for noninvasive and/or nondestructive assessment of 3D trabecular structure and connectivity. Micro computed tomography (microCT) has been used to measure 3D trabecular bone structure in rats, both in vivo and in vitro. It can directly quantify 3D trabecular bone structure such as trabecular volume, trabecular thickness, number, separation, structure model index, degree of anisotropy, and connectivity, in a model-independent manner. We have used microCT to study ovariectomy (OVX) induced osteopenia in rats and its treatment with agents such as estrogen, and sodium fluoride. We have demonstrated that 3D microCT can quantify mouse trabecular and cortical bone structure with an isotropic resolution of 9 microm(3). It is also useful for studying osteoporosis in mice and in phenotypes of transgenic mice or gene knockout mice. MicroCT can be used to quantify osteogenesis in mouse Ilizarov leg lengthening procedures, to quantify osteoconduction in a rat cranial defect model, and to quantify cortical bone porosity. Recently, microCT using high intensity and tight collimation synchrotron radiation to achieve spatial resolution of 1-2 microm has provided the capability to assess additional features such as resorption cavities. Unlike microCT, micro magnetic resonance imaging (IMRI) is nonionizing. Recently, the ability of microMRI to assess osteoporosis in animal models has been explored. Using a small, high-efficiency coil in a high-field imager, microMRI can give resolutions sufficient to discriminate individual trabeculae. We have shown that, with appropriate settings, it is possible to image trabecular bone in rats in vivo and in vitro. In our study of OVX rats, analysis of microMR images can demonstrate differences in rat trabecular bone that are not detected by DXA measurements. In a rabbit OA model, with the OA induced by meniscectomy or anterior cruciate ligament transection, MRI shows decreased cartilage thickness, subchondral osteosclerosis and osteophytes, while radiographs can only show subchondral osteosclerosis and osteophytes could not be found. Advanced imaging methods are able to measure 3D trabecular structure and connectivity in arbitrary orientations in a highly automated, objective, non-user-specific manner, allowing greater numbers of samples for unbiased comparisons between controls and the disordered or treated. They can be utilized on a large sample leading to fewer sampling errors. They are non-destructive allowing multiple tests such as biomechanical testing and chemical analysis on the same sample; and non-invasive permitting longitudinal studies and reducing the number of animals needed.     

Murine Endoscopy for In Vivo Multimodal Imaging of Carcinogenesis and Assessment of Intestinal Wound Healing and Inflammation

Mouse models are widely used to study pathogenesis of human diseases and to evaluate diagnostic procedures as well as therapeutic interventions preclinically. However, valid assessment of pathological alterations often requires histological analysis, and when performed ex vivo, necessitates death of the animal. Therefore in conventional experimental settings, intra-individual follow-up examinations are rarely possible. Thus, development of murine endoscopy in live mice enables investigators for the first time to both directly visualize the gastrointestinal mucosa and also repeat the procedure to monitor for alterations. Numerous applications for in vivo murine endoscopy exist, including studying intestinal inflammation or wound healing, obtaining mucosal biopsies repeatedly, and to locally administer diagnostic or therapeutic agents using miniature injection catheters. Most recently, molecular imaging has extended diagnostic imaging modalities allowing specific detection of distinct target molecules using specific photoprobes. In conclusion, murine endoscopy has emerged as a novel cutting-edge technology for diagnostic experimental in vivo imaging and may significantly impact on preclinical research in various fields.     

Early and stable upregulation of collagen type II,collagen type I and YKL40 expression levels in cartilage during early experimental osteoarthritis occurs independent of joint location and histological grading

While morphologic and biochemical aspects of degenerative joint disease (osteoarthritis [OA]) have been elucidated by numerous studies, the molecular mechanisms underlying the progressive loss of articular cartilage during OA development remain largely unknown. The main focus of the present study was to gain more insight into molecular changes during the very early stages of mechanically induced cartilage degeneration and to relate molecular alterations to histological changes at distinct localizations of the joint. Studies on human articular cartilage are hampered by the difficulty of obtaining normal tissue and early-stage OA tissue, and they allow no progressive follow-up. An experimental OA model in dogs with a slow natural history of OA (Pond–Nuki model) was therefore chosen. Anterior cruciate ligament transection (ACLT) was performed on 24 skeletally mature dogs to induce joint instability resulting in OA. Samples were taken from different joint areas after 6, 12, 24 and 48 weeks, and gene expression levels of common cartilage molecules were quantified in relation to the histological grading (modified Mankin score) of adjacent tissue. Histological changes reflected early progressive degenerative OA. Soon after ACLT, chondrocytes responded to the altered mechanical conditions by significant and stable elevation of collagen type II, collagen type I and YKL40 expression, which persisted throughout the study. In contrast to the mild to moderate histological alterations, these molecular changes were not progressive and were independent of the joint localization (tibia, femur, lateral, medial) and the extent of matrix degeneration. MMP13 remained unaltered until 24 weeks, and aggrecan and tenascinC remained unaltered until 48 weeks after ACLT. These findings indicate that elevated collagen type II, collagen type I and YKL40 mRNA expression levels are early and sensitive measures of ACLT-induced joint instability independent of a certain grade of morphological cartilage degeneration. A second phase of molecular changes in OA may begin around 48 weeks after ACLT with altered expression of further genes, such as MMP13, aggrecan and tenascin. Molecular changes observed in the present study suggest that dog cartilage responds to degenerative conditions by regulating the same genes in a similar direction as that observed for chondrocytes in late human OA.     

Early and stable upregulation of collagen type II, collagen type I and YKL40 expression levels in cartilage during early experimental osteoarthritis occurs independent of joint location and histological grading

While morphologic and biochemical aspects of degenerative joint disease (osteoarthritis [OA]) have been elucidated by numerous studies, the molecular mechanisms underlying the progressive loss of articular cartilage during OA development remain largely unknown. The main focus of the present study was to gain more insight into molecular changes during the very early stages of mechanically induced cartilage degeneration and to relate molecular alterations to histological changes at distinct localizations of the joint. Studies on human articular cartilage are hampered by the difficulty of obtaining normal tissue and early-stage OA tissue, and they allow no progressive follow-up. An experimental OA model in dogs with a slow natural history of OA (Pond-Nuki model) was therefore chosen. Anterior cruciate ligament transection (ACLT) was performed on 24 skeletally mature dogs to induce joint instability resulting in OA. Samples were taken from different joint areas after 6, 12, 24 and 48 weeks, and gene expression levels of common cartilage molecules were quantified in relation to the histological grading (modified Mankin score) of adjacent tissue. Histological changes reflected early progressive degenerative OA. Soon after ACLT, chondrocytes responded to the altered mechanical conditions by significant and stable elevation of collagen type II, collagen type I and YKL40 expression, which persisted throughout the study. In contrast to the mild to moderate histological alterations, these molecular changes were not progressive and were independent of the joint localization (tibia, femur, lateral, medial) and the extent of matrix degeneration. MMP13 remained unaltered until 24 weeks, and aggrecan and tenascinC remained unaltered until 48 weeks after ACLT. These findings indicate that elevated collagen type II, collagen type I and YKL40 mRNA expression levels are early and sensitive measures of ACLT-induced joint instability independent of a certain grade of morphological cartilage degeneration. A second phase of molecular changes in OA may begin around 48 weeks after ACLT with altered expression of further genes, such as MMP13, aggrecan and tenascin. Molecular changes observed in the present study suggest that dog cartilage responds to degenerative conditions by regulating the same genes in a similar direction as that observed for chondrocytes in late human OA.     

A review of high-resolution X-ray computed tomography and other imaging modalities for small animal research

Dedicated high-resolution small animal systems have recently emerged as important new tools for laboratory animal research. These imaging systems permit researchers to noninvasively screen animal models for mutations or pathologies and to monitor disease progression and response to therapy. The authors survey various small animal imaging modalities, including MRI, PET, SPECT, and microCT, and discuss several representative microCT mouse imaging studies.     

Multimodality Chamber for coregistered anatomical and molecular imaging of small animals

Modern imaging methods are applied extensively in translational animal research, and combined analysis of anatomical and functional imaging results is of increasing importance. Many imaging centers handle multiple independent animal colonies and use several imaging modalities, often in combination. The authors have developed and successfully tested a two-piece acrylic Multimodality Chamber that enables investigators to coregister images from two or more modalities, including microMR, microCT, microPET and optical imaging.     

Lubricin: a novel potential biotherapeutic approaches for the treatment of osteoarthritis

Osteoarthritis (OA) is a multi-factor disorder of sinovial joints, which characterized by escalated degeneration and loss of articular cartilage. Treatment of OA is a critical unmet need in medicine for regeneration of damaged articular cartilage in elderly. On the other hand, lubricin, a glycoprotein specifically synthesized by chondrocytes located at the surface of articular cartilage, has been shown to provide boundary lubrication of congruent articular surfaces under conditions of high contact pressure and near zero sliding speed. Lubrication of these surfaces is critical to normal joint function, while different gene expressions of lubricin had been found in the synovium of rheumatoid arthritis (RA) and OA. Moreover, mutations or lacking of lubricin gene have been shown to link to the joint disease such as camptodactyly-arthropathy-coxa vara-pericarditis syndrome (CACP), synovial hyperplasia and failure of joint function, suggesting an important role of lubricin in the pathogenesis of these joint disease. Recent studies demonstrate that administration with recombinant lubricin in the joint cavity would be effective in the prevention of cartilage degeneration in animal OA models. Therefore, a treatment with lubricin which would protect cartilage in vivo would be desirable. This article reviews recent findings with regard to the possible role of lubricin in the progression of OA, and further discusses lubricin as a novel potential biotherapeutic approaches for the treatment of OA.     

Fourier transform infrared imaging spectroscopy investigations in the pathogenesis and repair of cartilage

Significant complications in the management of osteoarthritis (OA) are the inability to identify early cartilage changes during the development of the disease, and the lack of techniques to evaluate the tissue response to therapeutic and tissue engineering interventions. In recent studies several spectroscopic parameters have been elucidated by Fourier transform infrared imaging spectroscopy (FT-IRIS) that enable evaluation of molecular and compositional changes in human cartilage with progressively severe OA, and in repair cartilage from animal models. FT-IRIS permits evaluation of early-stage matrix changes in the primary components of cartilage, collagen and proteoglycan on histological sections at a spatial resolution of approximately 6.25 microm. In osteoarthritic cartilage, the collagen integrity, monitored by the ratio of peak areas at 1338 cm(-1)/Amide II, was found to correspond to the histological Mankin grade, the gold standard scale utilized to evaluate cartilage degeneration. Apparent matrix degradation was observable in the deep zone of cartilage even in the early stages of OA. FT-IRIS studies also found that within the territorial matrix of the cartilage cells (chondrocytes), proteoglycan content increased with progression of cartilage degeneration while the collagen content remained the same, but the collagen integrity decreased. Regenerative (repair) tissue from microfracture treatment of an equine cartilage defect showed significant changes in collagen distribution and loss in proteoglycan content compared to the adjacent normal cartilage, with collagen fibrils demonstrating a random orientation in most of the repair tissue. These studies demonstrate that FT-IRIS is a powerful technique that can provide detailed ultrastructural information on heterogeneous tissues such as diseased cartilage and thus has great potential as a diagnostic modality for cartilage degradation and repair.     

Animal models of osteoarthritis in an era of molecular biology

Animal models of osteoarthritis (OA) are used to study the pathogenesis of cartilage degeneration and to evaluate potential anti-arthritic drugs for clinical use. In general, these models fall into 2 categories, spontaneous and induced (surgical instability or genetic manipulation). Animal models of naturally occurring OA occur in knee joints of guinea pigs, mice and Syrian hamsters. Commonly utilized surgical instability models include medial meniscal tear in guinea pigs and rats, medial or lateral partial meniscectomy in rabbits, medial partial or total meniscectomy or anterior cruciate transection in dogs. Transgenic models have been developed in mice. These models all have potential use in the study of molecular mechanisms associated with OA development via use of immunohistochemistry, biochemistry and molecular probes to identify altered matrix molecules at different stages in disease progression. Testing of specific types of inhibitors developed through evaluation of matrix changes in the disease process will ultimately help identify key processes which initiate and perpetuate the disease and will lead to discovery of new disease modifying pharmaceutical agents for OA patients. This paper will focus on the discussion of several models which are likely to be useful in the molecular dissection of processes involved in cartilage degeneration.     

Fourier transform infrared imaging spectroscopy investigations in the pathogenesis and repair of cartilage

Significant complications in the management of osteoarthritis (OA) are the inability to identify early cartilage changes during the development of the disease, and the lack of techniques to evaluate the tissue response to therapeutic and tissue engineering interventions. In recent studies several spectroscopic parameters have been elucidated by Fourier transform infrared imaging spectroscopy (FT-IRIS) that enable evaluation of molecular and compositional changes in human cartilage with progressively severe OA, and in repair cartilage from animal models. FT-IRIS permits evaluation of early-stage matrix changes in the primary components of cartilage, collagen and proteoglycan on histological sections at a spatial resolution of ∼6.25 μm. In osteoarthritic cartilage, the collagen integrity, monitored by the ratio of peak areas at 1338 cm⁻¹/Amide II, was found to correspond to the histological Mankin grade, the gold standard scale utilized to evaluate cartilage degeneration. Apparent matrix degradation was observable in the deep zone of cartilage even in the early stages of OA. FT-IRIS studies also found that within the territorial matrix of the cartilage cells (chondrocytes), proteoglycan content increased with progression of cartilage degeneration while the collagen content remained the same, but the collagen integrity decreased. Regenerative (repair) tissue from microfracture treatment of an equine cartilage defect showed significant changes in collagen distribution and loss in proteoglycan content compared to the adjacent normal cartilage, with collagen fibrils demonstrating a random orientation in most of the repair tissue. These studies demonstrate that FT-IRIS is a powerful technique that can provide detailed ultrastructural information on heterogeneous tissues such as diseased cartilage and thus has great potential as a diagnostic modality for cartilage degradation and repair.     

Cell death of chondrocytes is a combination between apoptosis and autophagy during the pathogenesis of Osteoarthritis within an experimental model

The death of chondrocytes and the loss of extracellular matrix are the central features in cartilage degeneration during Osteoarthritis (OA) pathogenesis. The mechanism by which chondrocytes are removed in OA cartilage are still not totally defined, although previous reports support the presence of apoptotic as well as non apoptotic signals. In addition, in 2004 Roach and co-workers suggested the term “Chondroptosis” to design the type of cell death present in articular cartilage, which include the presence of some apoptotic and autophagic processes. To identify the mechanisms, as well as the chronology by which chondrocytes are eliminated during OA pathogenesis, we decided to evaluate apoptosis (by active caspase 3 and TUNEL signal) and autophagy (by LC3II molecule and cytoplasmic vacuolization) using Immunohistochemistry and Western blot techniques in an animal OA model. During OA pathogenesis, chondrocytes exhibit modifications in their death process in each zone of the cartilage. At early stages of OA, the death of chondrocytes starts with apoptosis in the superficial and part of the middle zones of the cartilage, probably as a consequence of a constant mechanical damage in the joint. As the degenerative process progresses, high incidence of active caspase 3 as well as LC3II expression are observed in the same cell, which indicate a combination of both death processes. In contrast, in the deep zone, due the abnormal subchondral bone ossification during the OA pathogenesis, apoptosis is the only mechanism observed.     

Quantitative analysis of micro-CT imaging and histopathological signatures of experimental arthritis in rats

Micro-computed tomographic (micro-CT) imaging provides a unique opportunity to capture 3-D architectural information in bone samples. In this study of pathological joint changes in a rat model of adjuvant-induced arthritis (AA), quantitative analysis of bone volume and roughness were performed by micro-CT imaging and compared with histopathology methods and paw swelling measurement. Micro-CT imaging of excised rat hind paws (n = 10) stored in formalin consisted of approximately 600 30-mum slices acquired on a 512 x 512 image matrix with isotropic resolution. Following imaging, the joints were scored from H&E stained sections for cartilage/bone erosion, pannus development, inflammation, and synovial hyperplasia. From micro-CT images, quantitative analysis of absolute bone volumes and bone roughness was performed. Bone erosion in the rat AA model is substantial, leading to a significant decline in tarsal volume (27%). The result of the custom bone roughness measurement indicated a 55% increase in surface roughness. Histological and paw volume analyses also demonstrated severe arthritic disease as compared to controls. Statistical analyses indicate correlations among bone volume, roughness, histology, and paw volume. These data demonstrate that the destructive progression of disease in a rat AA model can be quantified using 3-D micro-CT image analysis, which allows assessment of arthritic disease status and efficacy of experimental therapeutic agents.     

JUNB‐FBXO21‐ERK axis promotes cartilage degeneration in osteoarthritis by inhibiting autophagy

Osteoarthritis (OA) is a heterogeneous disease that is extremely hard to cure owing to its complex regulation network of pathogenesis, especially cartilage degeneration. FBXO21 is a subunit of ubiquitin E3 ligases that degrades P‐glycoprotein and EID1 by ubiquitination and activates the JNK and p38 pathways; however, its role in OA remains unknown. Here, the main objective of this study was to evaluate the potential effects and mechanism of FBXO21 in OA degeneration, we revealed that FBXO21 is upregulated in the cartilage of patients with OA, aging, and monosodium iodoacetate‐induced OA rats, and chondrocytes treated with interleukin‐1β, tumor necrosis factor‐α, and lipopolysaccharide. Moreover, the in vivo and in vitro knockdown of FBXO21 suppressed OA‐related cartilage degeneration, as evidenced by activated autophagy, upregulated anabolism, alleviated apoptosis, and downregulated catabolism. In contrast, its overexpression promoted OA‐related cartilage degeneration. In addition, using mass spectrometry and co‐immunoprecipitation assay, we demonstrated that the downstream mechanism of FBXO21 inhibits autophagy by interacting with and phosphorylating ERK. Furthermore, FBXO21 alleviated anabolism and enhanced apoptosis and catabolism by inhibiting autophagy in rat chondrocytes. Interestingly, for its upstream mechanism, JUNB promoted FBXO21 expression by directly targeting the FBXO21 promoter, thus further accelerating cartilage degeneration in SW1353 cells and rat chondrocytes. Overall, our findings reveal that the JUNB‐FBXO21‐ERK axis regulates OA apoptosis and cartilage matrix metabolism by inhibiting autophagy. Therefore, FBXO21 is an attractive target for regulating OA pathogenesis, and its knockdown may provide a novel targeted therapy for OA.     

Coenzyme Q10 Ameliorates Pain and Cartilage Degradation in a Rat Model of Osteoarthritis by Regulating Nitric Oxide and Inflammatory Cytokines

Objective

To investigate the effect of CoenzymeQ10 (CoQ10) on pain severity and cartilage degeneration in an experimental model of rat osteoarthritis (OA).

Materials and Methods

OA was induced in rats by intra-articular injection of monosodium iodoacetate (MIA) to the knee. Oral administration of CoQ10 was initiated on day 4 after MIA injection. Pain severity was assessed by measuring secondary tactile allodynia using the von Frey assessment test. The degree of cartilage degradation was determined by measuring cartilage thickness and the amount of proteoglycan. The mankin scoring system was also used. Expressions of matrix metalloproteinase-13 (MMP-13), interleukin-1β (IL-1β), IL-6, IL-15, inducible nitric oxide synthase (iNOS), nitrotyrosine and receptor for advanced glycation end products (RAGE) were analyzed using immunohistochemistry.

Results

Treatment with CoQ10 demonstrated an antinociceptive effect in the OA animal model. The reduction in secondary tactile allodynia was shown by an increased pain withdrawal latency and pain withdrawal threshold. CoQ10 also attenuated cartilage degeneration in the osteoarthritic joints. MMP-13, IL-1β, IL-6, IL-15, iNOS, nitrotyrosine and RAGE expressions were upregulated in OA joints and significantly reduced with CoQ10 treatment.

Conclusion

CoQ10 exerts a therapeutic effect on OA via pain suppression and cartilage degeneration by inhibiting inflammatory mediators, which play a vital role in OA pathogenesis.     

A musculoskeletal finite element model of rat knee joint for evaluating cartilage biomechanics during gait

Abnormal loading of the knee due to injuries or obesity is thought to contribute to the development of osteoarthritis (OA). Small animal models have been used for studying OA progression mechanisms. However, numerical models to study cartilage responses under dynamic loading in preclinical animal models have not been developed. Here we present a musculoskeletal finite element model of a rat knee joint to evaluate cartilage biomechanical responses during a gait cycle. The rat knee joint geometries were obtained from a 3-D MRI dataset and the boundary conditions regarding loading in the joint were extracted from a musculoskeletal model of the rat hindlimb. The fibril-reinforced poroelastic (FRPE) properties of the rat cartilage were derived from data of mechanical indentation tests. Our numerical results showed the relevance of simulating anatomical and locomotion characteristics in the rat knee joint for estimating tissue responses such as contact pressures, stresses, strains, and fluid pressures. We found that the contact pressure and maximum principal strain were virtually constant in the medial compartment whereas they showed the highest values at the beginning of the gait cycle in the lateral compartment. Furthermore, we found that the maximum principal stress increased during the stance phase of gait, with the greatest values at midstance. We anticipate that our approach serves as a first step towards investigating the effects of gait abnormalities on the adaptation and degeneration of rat knee joint tissues and could be used to evaluate biomechanically-driven mechanisms of the progression of OA as a consequence of joint injury or obesity.     

Thermosensitive hybrid hyaluronan/p(HPMAm-lac)-PEG hydrogels enhance cartilage regeneration in a mouse model of osteoarthritis

Osteoarthritis (OA), due to cartilage degeneration, is one of the leading causes of disability worldwide. Currently, there are not efficacious therapies to reverse cartilage degeneration. In this study we evaluated the potential of hybrid hydrogels, composed of a biodegradable and thermosensitive triblock copolymer cross-linked via Michael addition to thiolated hyaluronic acid, in contrasting inflammatory processes underlying OA. Hydrogels composed of different w/w % concentrations of hyaluronan were investigated for their degradation behavior and capacity to release the polysaccharide in a sustained fashion. It was found that hyaluronic acid was controllably released during network degradation with a zero-order release kinetics, and the release rate depended on cross-link density and degradation kinetics of the hydrogels. When locally administered in vivo in an OA mouse model, the hydrogels demonstrated the ability to restore, to some extent, bone remineralization, proteoglycan production, levels of Sox-9 and Runx-2. Furthermore, the downregulation of proinflammatory mediators, such as TNF-α, NFkB, and RANKL and proinflammatory cytokines was observed. In summary, the investigated hydrogel technology represents an ideal candidate for the potential encapsulation and release of drugs relevant in the field of OA. In this context, the hydrogel matrix could act in synergy with the drug, in reversing phenomena of inflammation, cartilage disruption, and bone demineralization associated with OA.     

Small animal models to understand pathogenesis of osteoarthritis and use of stem cell in cartilage regeneration

Osteoarthritis (OA) is one of the most common diseases, which affect the correct functionality of synovial joints and is characterized by articular cartilage degradation. Limitation in the treatment of OA is mostly due to the very limited regenerative characteristic of articular cartilage once is damaged. Small animal models are of particular importance for mechanistic analysis to understand the processes that affect cartilage degradation. Combination of joint injury techniques with the use of stem cells has been shown to be an important tool for understanding the processes of cartilage degradation and regeneration. Implementation of stem cells and small animal models are important tools to help researchers to find a solution that could ameliorate and prevent the symptoms of OA.     

Deletion of Dual Specificity Phosphatase 1 Does Not Predispose Mice to Increased Spontaneous Osteoarthritis

Background

Osteoarthritis (OA) is a degenerative joint disease with poorly understood etiology and pathobiology. Mitogen activated protein kinases (MAPKs) including ERK and p38 play important roles in the mediation of downstream pathways involved in cartilage degenerative processes. Dual specificity phosphatase 1 (DUSP1) dephosphorylates the threonine/serine and tyrosine sites on ERK and p38, causing deactivation of downstream signalling. In this study we examined the role of DUSP1 in spontaneous OA development at 21 months of age using a genetically modified mouse model deficient in Dusp1 (DUSP1 knockout mouse).

Results

Utilizing histochemical stains of paraffin embedded knee joint sections in DUSP1 knockout and wild type female and male mice, we showed similar structural progression of cartilage degeneration associated with OA at 21 months of age. A semi-quantitative cartilage degeneration scoring system also demonstrated similar scores in the various aspects of the knee joint articular cartilage in DUSP1 knockout and control mice. Examination of overall articular cartilage thickness in the knee joint demonstrated similar results between DUSP1 knockout and wild type mice. Immunostaining for cartilage neoepitopes DIPEN, TEGE and C1,2C was similar in the cartilage lesion sites and chondrocyte pericellular matrix of both experimental groups. Likewise, immunostaining for phosphoERK and MMP13 showed similar intensity and localization between groups. SOX9 immunostaining demonstrated a decreased number of positive cells in DUSP1 knockout mice, with correspondingly decreased staining intensity. Analysis of animal walking patterns (gait) did not show a discernable difference between groups.

Conclusion

Loss of DUSP1 does not cause changes in cartilage degeneration and gait in a mouse model of spontaneous OA at 21 months of age. Altered staining was observed in SOX9 immunostaining which may prove promising for future studies examining the role of DUSPs in cartilage and OA, as well as models of post-traumatic OA.     

Kartogenin prevents cartilage degradation and alleviates osteoarthritis progression in mice via the miR-146a/NRF2 axis

Osteoarthritis (OA) is a common articular degenerative disease characterized by loss of cartilage matrix and subchondral bone sclerosis. Kartogenin (KGN) has been reported to improve chondrogenic differentiation of mesenchymal stem cells. However, the therapeutic effect of KGN on OA-induced cartilage degeneration was still unclear. This study aimed to explore the protective effects and underlying mechanisms of KGN on articular cartilage degradation using mice with post-traumatic OA. To mimic the in vivo arthritic environment, in vitro cultured chondrocytes were exposed to interleukin-1β (IL-1β). We found that KGN barely affected the cell proliferation of chondrocytes; however, KGN significantly enhanced the synthesis of cartilage matrix components such as type II collagen and aggrecan in a dose-dependent manner. Meanwhile, KGN markedly suppressed the expression of matrix degradation enzymes such as MMP13 and ADAMTS5. In vivo experiments showed that intra-articular administration of KGN ameliorated cartilage degeneration and inhibited subchondral bone sclerosis in an experimental OA mouse model. Molecular biology experiments revealed that KGN modulated intracellular reactive oxygen species in IL-1β-stimulated chondrocytes by up-regulating nuclear factor erythroid 2-related factor 2 (NRF2), while barely affecting its mRNA expression. Microarray analysis further revealed that IL-1β significantly up-regulated miR-146a that played a critical role in regulating the protein levels of NRF2. KGN treatment showed a strong inhibitory effect on the expression of miR-146a in IL-1β-stimulated chondrocytes. Over-expression of miR-146a abolished the anti-arthritic effects of KGN not only by down-regulating the protein levels of NRF2 but also by up-regulating the expression of matrix degradation enzymes. Our findings demonstrate, for the first time, that KGN exerts anti-arthritic effects via activation of the miR-146a-NRF2 axis and KGN is a promising heterocyclic molecule to prevent OA-induced cartilage degeneration.Subject terms: Osteoarthritis, Drug development