Association of the FGF4L2 retrogene with fibrocartilaginous embolic myelopathy in dogs.

BACKGROUND: Fibrocartilaginous embolic myelopathy (FCE) is a well-documented condition in dogs although rarely reported in chondrodystrophic breeds. Genetic associations have not been defined. OBJECTIVES: Define the association of the chondrodystrophy-associated FGF4L2 retrogene with histopathologically confirmed cases of FCE. ANIMALS: Ninety-eight dogs with a histopathologic diagnosis of FCE. METHODS: Retrospective multicenter study. Dogs were genotyped for the FGF4L2 and FGF4L1 retrogenes using DNA extracted from formalin-fixed, paraffin-embedded tissue. Associations between breed, FCE and retrogene status were investigated with reference to a hospital population and known breed and general population allele frequencies. RESULTS: FGF4L2 genotype was defined in 89 FCE cases. Fibrocartilaginous embolic myelopathy was present in 22 dogs from FGF4L2-segregating breeds with allele frequencies of ≥5%; however, all dogs were wild type. Two Labrador retrievers with FCE carried FGF4L2 alleles. Frequency of the FGF4L2 allele was significantly (P < .001) and negatively associated with FCE relative to predicted hospital-population dogs. FCE was overrepresented in Boxer, Great Dane, Yorkshire Terrier, Bernese Mountain Dog, Miniature Schnauzer, Rottweiler, and Shetland Sheepdog breeds. CONCLUSIONS AND CLINICAL IMPORTANCE: Study data based on genotypically and histopathologically defined cases support the historical observation that FCE is uncommon in chondrodystrophic dog breeds. FGF4 plays an important role in angiogenesis and vascular integrity; anatomical studies comparing chondrodystrophic and non-chondrodystrophic dogs might provide insight into the pathogenesis of FCE.

The regulatory activities of MALAT1 in the development of bone and cartilage diseases.

Long non-coding RNAs (lncRNAs) have been comprehensively implicated in various cellular functions by mediating transcriptional or post-transcriptional activities. MALAT1 is involved in the differentiation, proliferation, and apoptosis of multiple cell lines, including BMSCs, osteoblasts, osteoclasts, and chondrocytes. Interestingly, MALAT1 may interact with RNAs or proteins, regulating cellular processes. Recently, MALAT1 has been reported to be associated with the development of bone and cartilage diseases by orchestrating the signaling network. The involvement of MALAT1 in the pathological development of bone and cartilage diseases makes it available to be a potential biomarker for clinical diagnosis or prognosis. Although the potential mechanisms of MALAT1 in mediating the cellular processes of bone and cartilage diseases are still needed for further elucidation, MALAT1 shows great promise for drug development.

Synovial mesenchymal stem cell-derived extracellular vesicles containing microRN555A-26a-5p ameliorate cartilage damage of osteoarthritis.

BACKGROUND: Osteoarthritis (OA) is a degenerative disease characterized by cartilage damage. We aimed to improve the understanding of the protective mechanism of synovial mesenchymal stem cell (SMSC)-derived extracellular vesicles (EVs) in cartilage damage of OA. METHODS: SMSCs and SMSC-EVs were isolated from synovial biopsies of patients without OA and then identified. The pathological microenvironment of chondrocytes in OA was simulated by inducing SW1353 cells with interleukin (IL)-1ß, followed by SMSC-EV treatment to assess SW1353 cell proliferation, apoptosis and inflammation. Endocytosis of Dil-labeled EVs by SW1353 cells was observed. microRNA (miR)-26a-5p expression in EVs and EV-treated SW1353 cells was assessed. The effect of miR-26a-5p was evaluated after it was down-regulated in SMSCs, followed by extraction of EVs, which acted on SW1353 cells. The target relationship of miR-26a-5p and phosphatase and tensin homologue (PTEN) was predicted and confirmed. The role of PTEN in OA was evaluated after it was overexpressed. Functional assays were implemented in vivo to certify the role of SMSC-EVs in OA. RESULTS: SMSC-EVs enhanced IL-1ß-induced SW1353 cell proliferation, whereas they inhibited apoptosis and inflammation. EVs were endocytosed by SW1353 cells and delivered miR-26a-5p into SW1353 cells to overexpress miR-26a-5p. Down-regulation of miR-26a-5p in EVs attenuated the protection of EVs against IL-1ß-induced cell damage. miR-26a-5p targeted PTEN, for which overexpression spoiled the protection of EVs against IL-1ß-induced cell damage. SMSC-EVs carrying miR-26a-5p repaired cartilage damage of OA. CONCLUSIONS: SMSC-EVs carried miR-26a-5p into chondrocytes to up-regulate miR-26a-5p and inhibit PTEN, thereby inhibiting apoptosis and inflammation and ameliorating cartilage damage of OA.

Promising diagnostic and therapeutic circRNAs for skeletal and chondral disorders.

Circular RNAs (circRNAs) belong to a highly conserved subtype of non-coding RNAs, produced by the back-splicing of specific regions of pre-mRNA. CircRNAs have wide-ranging effects on eukaryotic physiology and pathology by acting as transcription regulators, miRNA sponges, protein sponges, and templates for translation. Skeletal and chondral disorders are the leading causes of pain and disability, especially for elders, affecting hundreds of millions of people worldwide. Plenty of evidence have shown that circRNAs are dysregulated and play vital roles in the occurrence and progression of skeletal and chondral disorders. Herein, we systematically summarize the emerging roles and underlying molecular mechanisms of hub circRNAs in the pathogenesis of several representative skeletal and chondral disorders. Our findings may provide further insight into the mechanistic details of the role of circRNA in bone or cartilage metabolism, and highlight the promising application of circRNAs in serving as potential diagnostic or therapeutic targets for the prevention and treatment of skeletal and chondral disorders.

An animal model study on the gene expression profile of meniscal degeneration.

Meniscal degeneration is a very common condition in elderly individuals, but the underlying mechanisms of its occurrence are not completely clear. This study examines the molecular mechanisms of meniscal degeneration. The anterior cruciate ligament (ACL) and lateral collateral ligament (LCL) of the right rear limbs of seven Wuzhishan mini-pigs were resected (meniscal degeneration group), and the left rear legs were sham-operated (control group). After 6 months, samples were taken for gene chip analysis, including differentially expressed gene (DEG) analysis, gene ontology (GO) analysis, clustering analysis, and pathway analysis. The selected 12 DEGs were validated by real time reverse transcription-polymerase chain reaction (RT-PCR). The two groups showed specific and highly clustered DEGs. A total of 893 DEGs were found, in which 537 are upregulated, and 356 are downregulated. The GO analysis showed that the significantly affected biological processes include nitric oxide metabolic process, male sex differentiation, and mesenchymal morphogenesis, the significantly affected cellular components include the endoplasmic reticulum membrane, and the significantly affected molecular functions include transition metal ion binding and iron ion binding. The pathway analysis showed that the significantly affected pathways include type II diabetes mellitus, inflammatory mediator regulation of TRP channels, and AMPK signaling pathway. The results of RT-PCR indicate that the microarray data accurately reflects the gene expression patterns. These findings indicate that several molecular mechanisms are involved in the development of meniscal degeneration, thus improving our understanding of meniscal degeneration and provide molecular therapeutic targets in the future.

Effects of Edible Oils with Different n-6/n-3 PUFA Ratios on Articular Cartilage Degeneration via Regulating the NF-κB Signaling Pathway.

Osteoarthritis (OA), a common chronic disease, is characterized by articular cartilage degeneration and inflammation. Recent studies report that n-3 polyunsaturated fatty acids (PUFAs) exhibit protective effects against OA, while n-6 PUFAs are more likely to damage cartilage. However, the effects of edible oils with different n-6/n-3 PUFA ratios on OA are rarely reported. This study investigates the effect of linseed oil (LO), soybean oil (SO), and peanut oil (PO) on cartilage changes in mice joints following destabilization of the medial meniscus. We determined the n-6/n-3 PUFA ratios of LO, SO, and PO used in this experiment to be 1:3.85, 9.15:1, and 372.73:1, respectively. After 12 weeks of LO or SO feeding, OA mice showed increased cartilage thickness and decreased TNF-α in both the serum and cartilage, whereas no improvement was found in the PO group. This may be due to the fact that LO and SO optimized the fatty acid composition of articular cartilage. We further demonstrated that LO or SO activated GPR120 and attenuated EP4, which was followed by inhibition of the NFκB pathway and its downstream matrix-degrading enzymes: MMP13 and ADAMTS5. In conclusion, edible oils with low n-6/n-3 PUFA retard OA progression via inhibiting the NFκB pathway. This study provides a dietary guidance for OA patients.

Overexpressing Runx2 of BMSCs Improves the Repairment of knee Cartilage Defects.

BACKGROUND: Recruitment of gene modifying bone marrow mesenchymal stem cells (BMSCs) has been considered an alternative to single-cell injection in articular cartilage repair. PURPOSE: This study aimed to investigate whether the effect of runt-related transcription factor 2(Runx2) overexpression bone marrow mesenchymal stem cells in vivo could improve the quality of repaired tissue of a knee cartilage defect in a rabbit model. METHODS: Thirty-two New Zealand rabbits were randomly divided into four groups. The blank group (Con) did not receive anything, the model group (Mo) was administered saline, the simple stem cell group (MSCs) received MSCs injection, and the Runx2 transfection group (R-MSCs) received Runx2 overexpression MSCs injection. After adapting to the environment for a week, a 5 mm diameter cylindrical osteochondral defect was created in the center of the medial femoral condyle. Cell and saline injections were performed in the first and third weeks after surgery. The cartilage repair was evaluated by macroscopically and microscopically at 4 and 8 weeks. RESULTS: Macroscopically, defects were filled and surfaces were smoother in the MSCs groups than in the Mo group at 4th week. Microscopically, the R-MSCs group showed coloration similar to surrounding normal articular cartilage tissue at 8 weeks in masson trichrome staining. The COL-II, SOX9, and Aggrecan mRNA expressions of MSCs were enhanced at 4 weeks compared with R-MSCs, then the expression reduced at 8 weeks, but was still higher than Mo group level (P<0.05). The western blot examination revealed that the COL-IIand SOX9 expression of MSCs was higher than R-MSCs at 4 weeks, then the expression reduced at 8 weeks, but was still higher than the Mo level (P<0.05). The IL-1ß content in the joint fluid also revealed that cartilage repair with R-MSCs was better than that with MSCs at 8 weeks (P<0.05). CONCLUSION: The R-MSCs group showed cellular morphology and arrangement similar to surrounding normal articular cartilage tissue, and Runx2 overexpression of MSCs resulted in overall superior cartilage repair as compared with MSCs at 8 weeks.

The long noncoding RNA H19 attenuates force-driven cartilage degeneration via miR-483-5p/Dusp5.

Developmental dysplasia of the hip (DDH) is a common hip disease characterized by abnormal development of the acetabulum and femoral head. In most cases, DDH ultimately leads to osteoarthritis. Anomalous biomechanical force plays an important role in cartilage degeneration in DDH. However, in addition to mechanical wear, the underlying molecular mechanisms in cartilage degeneration in DDH remain unclear. This study analyzed the effect of long noncoding RNA (lncRNA)-H19 on DDH cartilage degradation. To elucidate the specific role of lncRNA H19, we established an intermittent cyclic mechanical stress (ICMS) cell force model to simulate abnormal biomechanical environment in vitro. Then, the roles of lncRNA-H19 were also determined in vivo by establishing a model of swaddling DDH. We observed that patients with DDH possessed low levels of lncRNA-H19, COL2A1, and Aggrecan but high levels of MMP3 and Adamts5. The same results were also obtained in a DDH rat model. Furthermore, the data suggested that ICMS promoted cartilage degeneration and caused reorientation of the cytoskeleton, and lncRNA H19 helped inhibit cartilage degeneration. Bioinformatics analysis and lncRNA sequencing were performed, and luciferase assays showed that lncRNA H19 and Dusp5 are both direct targets of miR-483-5p. Moreover, Dups5 plays a negative role in ICMS-induced cartilage degradation by activating the Erk and p38 pathways. In vivo, lncRNA H19 had protective effects on the swaddling DDH model. These findings indicate that lncRNA-H19 played a positive role in cartilage degradation in DDH through the lncRNA H19/miR-483-5p/Dusp5 axis.

Pathologic Features of the Intervertebral Disc in Young Nova Scotia Duck Tolling Retrievers Confirms Chondrodystrophy Degenerative Phenotype Associated With Genotype.

Chondrodystrophy results in predictable and progressive biochemical and structural changes to the intervertebral disc, resulting in early onset degeneration and dystrophic mineralization of the disc. Accelerated degeneration and mineralization of the intervertebral disc are common in multiple dog breeds and can result in compromised function, herniation, pain, and a variety of neurological sequelae. A mutation responsible for chondrodystrophy in dogs has been identified as an aberrant fibroblast growth factor 4 (FGF4) retrogene insertion on chromosome 12 (CFA12) and is associated with short stature of the Nova Scotia Duck Tolling Retriever. Segregation of the CFA12 FGF4 retrogene in this dog breed provides an opportunity to examine the effect of retrogene presence on radiographic and histologic appearance of chondrodystrophic disc degeneration within a single breed. Here we found that in the intervertebral discs isolated from 2 dogs with the CFA12 FGF4 genotype, the nucleus pulposus was largely replaced by cartilaginous tissue, and physaliferous notochordal cells were rarely if ever identified. These findings are in contrast to the normal histologic findings in 2 breed-matched dogs lacking the mutation. The findings are consistent with premature chondroid degeneration of the intervertebral disc and suggest that the presence of the CFA12 FGF4 retrogene is sufficient to cause the chondrodystrophic phenotype.

Fibrates as drugs with senolytic and autophagic activity for osteoarthritis therapy.

BACKGROUND: Ageing-related failure of homeostasis mechanisms contributes to articular cartilage degeneration and osteoarthritis (OA), for which disease-modifying treatments are not available. Our objective was to identify molecules to prevent OA by regulating chondrocyte senescence and autophagy. METHODS: Human chondrocytes with IL-6 induced senescence and autophagy suppression and SA-ß-gal as a reporter of senescence and LC3 as reporter of autophagic flux were used to screen the Prestwick Chemical Library of approved drugs. Preclinical cellular, tissue and blood from OA and blood from OA and ageing models were used to test the efficacy and relevance of activating PPARα related to cartilage degeneration. FINDINGS: Senotherapeutic molecules with pro-autophagic activity were identified. Fenofibrate (FN), a PPARα agonist used for dyslipidaemias in humans, reduced the number of senescent cells via apoptosis, increased autophagic flux, and protected against cartilage degradation. FN reduced both senescence and inflammation and increased autophagy in both ageing human and OA chondrocytes whereas PPARα knockdown conferred the opposite effect. Moreover, PPARα expression was reduced through both ageing and OA in mice and also in blood and cartilage from knees of OA patients. Remarkably, in a retrospective study, fibrate treatment improved OA clinical conditions in human patients from the Osteoarthritis Initiative (OAI) Cohort. INTERPRETATION: These results demonstrate that FDA-approved fibrate drugs targeting lipid metabolism protect against cartilage degeneration seen with ageing and OA. Thus, these drugs could have immediate clinically utility for age-related cartilage degeneration and OA treatment. FUND: This study was supported by Instituto de Salud Carlos III- Ministerio de Ciencia, Innovación y Universidades, Spain, Plan Estatal 2013-2016 and Fondo Europeo de Desarrollo Regional (FEDER), "Una manera de hacer Europa", PI14/01324 and PI17/02059, by Innopharma Pharmacogenomics platform applied to the validation of targets and discovery of drugs candidates to preclinical phases, Ministerio de Economía y Competitividad, by grants of the National Instiutes of Health to PDR (P01 AG043376 and U19 AG056278). We thank FOREUM Foundation for Research in Rheumatology for their support.

Defective WNT signaling may protect from articular cartilage deterioration - a quantitative MRI study on subjects with a heterozygous WNT1 mutation.

OBJECTIVE: WNT signaling is of key importance in chondrogenesis and defective WNT signaling may contribute to the pathogenesis of osteoarthritis and other cartilage diseases. Biochemical composition of articular cartilage in patients with aberrant WNT signaling has not been studied. Our objective was to assess the knee articular cartilage in WNT1 mutation-positive individuals using a 3.0T MRI unit to measure cartilage thickness, relaxation times, and texture features. DESIGN: Cohort comprised mutation-positive (N = 13; age 17-76 years) and mutation-negative (N = 13; 16-77 years) subjects from two Finnish families with autosomal dominant WNT1 osteoporosis due to a heterozygous missense mutation c.652T>G (p.C218G) in WNT1. All subjects were imaged with a 3.0T MRI unit and assessed for cartilage thickness, T2 and T1ρ relaxation times, and T2 texture features contrast, dissimilarity and homogeneity of T2 relaxation time maps in six regions of interest (ROIs) in the tibiofemoral cartilage. RESULTS: All three texture features showed opposing trends with age between the groups in the medial tibiofemoral cartilage (P = 0.020-0.085 for the difference of the regression coefficients), the mutation-positive individuals showing signs of cartilage preservation. No significant differences were observed in the lateral tibiofemoral cartilage. Cartilage thickness and means of T2 relaxation time did not differ between groups. Means of T1ρ relaxation time were significantly different in one ROI but the regression analysis displayed no differences. CONCLUSIONS: Our results show less age-related cartilage deterioration in the WNT1 mutation-positive than the mutation-negative subjects. This suggests, that the WNT1 mutation may alter cartilage turnover and even have a potential cartilage-preserving effect.

Role of Epigenomics in Bone and Cartilage Disease.

Phenotypic variation in skeletal traits and diseases is the product of genetic and environmental factors. Epigenetic mechanisms include information-containing factors, other than DNA sequence, that cause stable changes in gene expression and are maintained during cell divisions. They represent a link between environmental influences, genome features, and the resulting phenotype. The main epigenetic factors are DNA methylation, posttranslational changes of histones, and higher-order chromatin structure. Sometimes non-coding RNAs, such as microRNAs (miRNAs) and long non-coding RNAs (lncRNAs), are also included in the broad term of epigenetic factors. There is rapidly expanding experimental evidence for a role of epigenetic factors in the differentiation of bone cells and the pathogenesis of skeletal disorders, such as osteoporosis and osteoarthritis. However, different from genetic factors, epigenetic signatures are cell- and tissue-specific and can change with time. Thus, elucidating their role has particular difficulties, especially in human studies. Nevertheless, epigenomewide association studies are beginning to disclose some disease-specific patterns that help to understand skeletal cell biology and may lead to development of new epigenetic-based biomarkers, as well as new drug targets useful for treating diffuse and localized disorders. Here we provide an overview and update of recent advances on the role of epigenomics in bone and cartilage diseases. © 2019 American Society for Bone and Mineral Research.

Idiopathic hip chondrolysis: a case report of a Caucasian HLA-B27 positive adolescent with a history of long walking.

Idiopathic hip chondrolysis is a rare disorder, the pathophysiology of which has not been fully elucidated. Several theories have been proposed regarding the cause of the disease with some of them involving autoimmune-mediated cartilage destruction. There are several similar features between idiopathic hip chondrolysis and rheumatologic diseases such as juvenile idiopathic arthritis, so whether these two disorders are different or not is still debatable. This case report aims to help comprehending this complex disorder by presenting a case of idiopathic hip chondrolysis with apparent risk factors, such as repetitive microtrauma and presence of HLA-B27 antigens. A 15-year-old HLA-B27 positive male presented with idiopathic hip chondrolysis after excessive walking. Initial treatment consisted of medications including corticosteroids, protected weight bearing and surgical soft tissue release. After failure of all these modalities in restoring the decreased range of motion of the hip, a course of a TNF-inhibitor, etanercept was tried. Alleviation of pain achieved early in the treatment period, but range of motion remained mainly unchanged. Although there was a brief improvement of stiffness for a short period after surgery which lasted for about 3 months, stiffness came back afterwards. Administration of a TNF inhibitor in the following period significantly improved his range of motion. The presence of laboratory findings indicating an autoimmune tendency in this patient supports the hypothesis of susceptibility of these patients to autoimmune reactions, while excessive walking was an apparent trigger factor. In future, traditional treatments may be abandoned in favor of novel medications targeting immunologic pathways.

Models of Disease.

Osteochondral (OC) lesions are a major cause of chronic musculoskeletal pain and functional disability, which reduces the quality of life of the patients and entails high costs to the society. Currently, there are no effective treatments, so in vitro and in vivo disease models are critically important to obtain knowledge about the causes and to develop effective treatments for OC injuries. In vitro models are essential to clarify the causes of the disease and the subsequent design of the first barrier to test potential therapeutics. On the other hand, in vivo models are anatomically more similar to humans allowing to reproduce the pattern and progression of the lesion in a controlled scene and offering the opportunity to study the symptoms and responses to new treatments. Moreover, in vivo models are the most suitable preclinical model, being a fundamental and a mandatory step to ensure the successful transfer to clinical trials. Both in vitro and in vitro models have a number of advantages and limitation, and the choice of the most appropriate model for each study depends on many factors, such as the purpose of the study, handling or the ease to obtain, and cost, among others. In this chapter, we present the main in vitro and in vivo OC disease models that have been used over the years in the study of origin, progress, and treatment approaches of OC defects.

Cartilage diseases.

Hyaline cartilages, fibrocartilages and elastic cartilages play multiple roles in the human body including bearing loads in articular joints and intervertebral discs, providing joint lubrication, forming the external ears and nose, supporting the trachea, and forming the long bones during development and growth. The structure and organization of cartilage's extracellular matrix (ECM) are the primary determinants of normal function. Most diseases involving cartilage lead to dramatic changes in the ECM which can govern disease progression (e.g., in osteoarthritis), cause the main symptoms of the disease (e.g., dwarfism caused by genetically inherited mutations) or occur as collateral damage in pathological processes occurring in other nearby tissues (e.g., osteochondritis dissecans and inflammatory arthropathies). Challenges associated with cartilage diseases include poor understanding of the etiology and pathogenesis, delayed diagnoses due to the aneural nature of the tissue and drug delivery challenges due to the avascular nature of adult cartilages. This narrative review provides an overview of the clinical and pathological features as well as current treatment options available for various cartilage diseases. Late breaking advances are also described in the quest for development and delivery of effective disease modifying drugs for cartilage diseases including osteoarthritis, the most common form of arthritis that affects hundreds of millions of people worldwide.

Gene therapy for repair and regeneration of bone and cartilage.

Gene therapy refers to the use of viral and non-viral vectors to deliver nucleic acids to tissues of interest using direct (in vivo) or transduced cell-mediated (ex vivo) approaches. Over the past few decades, strategies have been adopted to express therapeutic transgenes at sites of injury to promote or facilitate repair of bone and cartilage. Targets of interest have typically included secreted proteins such as growth factors and anti-inflammatory mediators; however, work has also begun to focus intracellularly on signaling components, transcription factors and small, regulatory nucleic acids such as microRNAs (miRNAs). In recent years, a number of single therapeutic gene approaches (termed 'monotherapies') have proven effective in preclinical models of disease, and several are being evaluated in clinical trials. In particular, an ex vivo TGF-ß1 gene therapy was approved in Korea in 2017 for treatment of moderate-to-severe osteoarthritis (OA). The ability to utilize viral vectors for context-specific and combinatorial gene therapy is also being investigated, and these strategies are likely to be important in more robustly addressing the complexities of tissue repair and regeneration in skeletal disease. In this review, we provide an overview of viral gene therapies being developed for treatment of bone and cartilage pathologies, with an emphasis on emerging combinatorial strategies as well as those targeting intracellular mediators such as miRNAs.

miR-27b promotes type II collagen expression by targetting peroxisome proliferator-activated receptor-γ2 during rat articular chondrocyte differentiation.

MicroRNAs (miRNAs) play an essential role in articular cartilage development and growth. However, the exact mechanisms involved in this process remain unknown. In the present study, we investigated the biological functions of miR-27b during hypertrophic differentiation of rat articular chondrocytes. Based on in situ hybridization and immunohistochemistry, we report that miR-27b expression is reduced in the hypertrophic zone of articular cartilage, but expression of peroxisome proliferator-activated receptor γ (Pparγ) is increased. Dual-luciferase reporter gene assay and Western blot analysis demonstrated that Pparγ2 is a target of miR-27b Overexpression of miR-27b inhibited expression of Pparγ2, as well as type X collagen (Col10a1) and matrix metalloproteinase 13 (Mmp13), while significantly promoting the expression of Sex-determining Region-box 9 (Sox9) and type II collagen (Col2a1) at both the mRNA and protein levels. Rosiglitazone, a Pparγ agonist, suppressed Col2a1 expression, while promoting expression of runt-related transcription factor 2 (Runx2) and Col10a1 in a concentration-dependent manner. siRNA-mediated knockdown of Pparγ2 caused an increase in protein levels of Col2a1. The present study demonstrates that miR-27b regulates chondrocyte hypertrophy in part by targetting Pparγ2, and that miR-27b may have important therapeutic implications in cartilage diseases.

Different Forms of ER Stress in Chondrocytes Result in Short Stature Disorders and Degenerative Cartilage Diseases: New Insights by Cartilage-Specific ERp57 Knockout Mice.

Cartilage is essential for skeletal development by endochondral ossification. The only cell type within the tissue, the chondrocyte, is responsible for the production of macromolecules for the extracellular matrix (ECM). Before proteins and proteoglycans are secreted, they undergo posttranslational modification and folding in the endoplasmic reticulum (ER). However, the ER folding capacity in the chondrocytes has to be balanced with physiological parameters like energy and oxygen levels. Specific cellular conditions, e.g., a high protein demand, or pathologic situations disrupt ER homeostasis and lead to the accumulation of poorly folded or misfolded proteins. This state is called ER stress and induces a cellular quality control system, the unfolded protein response (UPR), to restore homeostasis. Different mouse models with ER stress in chondrocytes display comparable skeletal phenotypes representing chondrodysplasias. Therefore, ER stress itself seems to be involved in the pathogenesis of these diseases. It is remarkable that chondrodysplasias with a comparable phenotype arise independent from the sources of ER stress, which are as follows: (1) mutations in ECM proteins leading to aggregation, (2) deficiencies in ER chaperones, (3) mutations in UPR signaling factors, or (4) deficiencies in the degradation of aggregated proteins. In any case, the resulting UPR substantially impairs ECM protein synthesis, chondrocyte proliferation, and/or differentiation or regulation of autophagy and apoptosis. Notably, chondrodysplasias arise no matter if single or multiple events are affected. We analyzed cartilage-specific ERp57 knockout mice and demonstrated that the deficiency of this single protein disulfide isomerase, which is responsible for formation of disulfide bridges in ECM glycoproteins, is sufficient to induce ER stress and to cause an ER stress-related bone phenotype. These mice therefore qualify as a novel model for the analysis of ER stress in chondrocytes. They give new insights in ER stress-related short stature disorders and enable the analysis of ER stress in other cartilage diseases, such as osteoarthritis.

Autologous chondrocyte implantation-derived synovial fluids display distinct responder and non-responder proteomic profiles.

BACKGROUND: Autologous chondrocyte implantation (ACI) can be used in the treatment of focal cartilage injuries to prevent the onset of osteoarthritis (OA). However, we are yet to understand fully why some individuals do not respond well to this intervention. Identification of a reliable and accurate biomarker panel that can predict which patients are likely to respond well to ACI is needed in order to assign the patient to the most appropriate therapy. This study aimed to compare the baseline and mid-treatment proteomic profiles of synovial fluids (SFs) obtained from responders and non-responders to ACI. METHODS: SFs were derived from 14 ACI responders (mean Lysholm improvement of 33 (17-54)) and 13 non-responders (mean Lysholm decrease of 14 (4-46)) at the two stages of surgery (cartilage harvest and chondrocyte implantation). Label-free proteome profiling of dynamically compressed SFs was used to identify predictive markers of ACI success or failure and to investigate the biological pathways involved in the clinical response to ACI. RESULTS: Only 1 protein displayed a ≥2.0-fold differential abundance in the preclinical SF of ACI responders versus non-responders. However, there is a marked difference between these two groups with regard to their proteome shift in response to cartilage harvest, with 24 and 92 proteins showing ≥2.0-fold differential abundance between Stages I and II in responders and non-responders, respectively. Proteomic data has been uploaded to ProteomeXchange (identifier: PXD005220). We have validated two biologically relevant protein changes associated with this response, demonstrating that matrix metalloproteinase 1 was prominently elevated and S100 calcium binding protein A13 was reduced in response to cartilage harvest in non-responders. CONCLUSIONS: The differential proteomic response to cartilage harvest noted in responders versus non-responders is completely novel. Our analyses suggest several pathways which appear to be altered in non-responders that are worthy of further investigation to elucidate the mechanisms of ACI failure. These protein changes highlight many putative biomarkers that may have potential for prediction of ACI treatment success.

Inherited Arterial Calcification Syndromes: Etiologies and Treatment Concepts.

PURPOSE OF REVIEW: We give an update on the etiology and potential treatment options of rare inherited monogenic disorders associated with arterial calcification and calcific cardiac valve disease. RECENT FINDINGS: Genetic studies of rare inherited syndromes have identified key regulators of ectopic calcification. Based on the pathogenic principles causing the diseases, these can be classified into three groups: (1) disorders of an increased extracellular inorganic phosphate/inorganic pyrophosphate ratio (generalized arterial calcification of infancy, pseudoxanthoma elasticum, arterial calcification and distal joint calcification, progeria, idiopathic basal ganglia calcification, and hyperphosphatemic familial tumoral calcinosis; (2) interferonopathies (Singleton-Merten syndrome); and (3) others, including Keutel syndrome and Gaucher disease type IIIC. Although some of the identified causative mechanisms are not easy to target for treatment, it has become clear that a disturbed serum phosphate/pyrophosphate ratio is a major force triggering arterial and cardiac valve calcification. Further studies will focus on targeting the phosphate/pyrophosphate ratio to effectively prevent and treat these calcific disease phenotypes.