Effect of Clonally Expanded PD-1high CXCR5-CD4+ Peripheral T Helper Cells on B Cell Differentiation in the Joints of Patients With Antinuclear Antibody-Positive Juvenile Idiopathic Arthritis.

OBJECTIVE: Antinuclear antibody (ANA)-positive juvenile idiopathic arthritis (JIA) is characterized by synovial B cell hyperactivity, but the precise role of CD4+ T cells in promoting local B cell activation is unknown. This study was undertaken to determine the phenotype and function of synovial CD4+ T cells that promote aberrant B cell activation in JIA. METHODS: Flow cytometry was performed to compare the phenotype and cytokine patterns of PD-1high CD4+ T cells in the synovial fluid (SF) of patients with JIA and T follicular helper cells in the tonsils of control individuals. TCRVB next-generation sequencing was used to analyze T cell subsets for signs of clonal expansion. The functional impact of these T cell subsets on B cells was examined in cocultures in vitro. RESULTS: Multidimensional flow cytometry revealed the expansion of interleukin-21 (IL-21) and interferon-γ (IFNγ)-coexpressing PD-1high CXCR5-HLA-DR+CD4+ T cells that accumulate in the joints of ANA-positive JIA patients. These T cells exhibited signs of clonal expansion with restricted T cell receptor clonotypes. The phenotype resembled peripheral T helper (Tph) cells with an extrafollicular chemokine receptor pattern and high T-bet and B lymphocyte-induced maturation protein 1 expression, but low B cell lymphoma 6 expression. SF Tph cells, by provision of IL-21 and IFNy, skewed B cell differentiation toward a CD21low/- CD11c+ phenotype in vitro. Additionally, SF Tph cell frequencies correlated with the appearance of SF CD21low/- CD11c+CD27-IgM- double-negative (DN) B cells in situ. CONCLUSION: Clonally expanded CD4+ Tph cells accumulate in the joints of ANA-positive JIA patients and, in particular, promote CD21low/- CD11c+ DN B cell differentiation. The expansion of Tph cells and DN B cells might reflect the autoimmune response in the joints of ANA-positive JIA patients.

Locally renewing resident synovial macrophages provide a protective barrier for the joint.

Macrophages are considered to contribute to chronic inflammatory diseases such as rheumatoid arthritis1. However, both the exact origin and the role of macrophages in inflammatory joint disease remain unclear. Here we use fate-mapping approaches in conjunction with three-dimensional light-sheet fluorescence microscopy and single-cell RNA sequencing to perform a comprehensive spatiotemporal analysis of the composition, origin and differentiation of subsets of macrophages within healthy and inflamed joints, and study the roles of these macrophages during arthritis. We find that dynamic membrane-like structures, consisting of a distinct population of CX3CR1+ tissue-resident macrophages, form an internal immunological barrier at the synovial lining and physically seclude the joint. These barrier-forming macrophages display features that are otherwise typical of epithelial cells, and maintain their numbers through a pool of locally proliferating CX3CR1- mononuclear cells that are embedded into the synovial tissue. Unlike recruited monocyte-derived macrophages, which actively contribute to joint inflammation, these epithelial-like CX3CR1+ lining macrophages restrict the inflammatory reaction by providing a tight-junction-mediated shield for intra-articular structures. Our data reveal an unexpected functional diversification among synovial macrophages and have important implications for the general role of macrophages in health and disease.

Molecular Portraits of Early Rheumatoid Arthritis Identify Clinical and Treatment Response Phenotypes.

There is a current imperative to unravel the hierarchy of molecular pathways that drive the transition of early to established disease in rheumatoid arthritis (RA). Herein, we report a comprehensive RNA sequencing analysis of the molecular pathways that drive early RA progression in the disease tissue (synovium), comparing matched peripheral blood RNA-seq in a large cohort of early treatment-naive patients, namely, the Pathobiology of Early Arthritis Cohort (PEAC). We developed a data exploration website (https://peac.hpc.qmul.ac.uk/) to dissect gene signatures across synovial and blood compartments, integrated with deep phenotypic profiling. We identified transcriptional subgroups in synovium linked to three distinct pathotypes: fibroblastic pauci-immune pathotype, macrophage-rich diffuse-myeloid pathotype, and a lympho-myeloid pathotype characterized by infiltration of lymphocytes and myeloid cells. This is suggestive of divergent pathogenic pathways or activation disease states. Pro-myeloid inflammatory synovial gene signatures correlated with clinical response to initial drug therapy, whereas plasma cell genes identified a poor prognosis subgroup with progressive structural damage.

Lineage tracing of col10a1 cells identifies distinct progenitor populations for osteoblasts and joint cells in the regenerating fin of medaka (Oryzias latipes).

The caudal fin of teleost fish regenerates fully within two weeks of amputation. While various cell lineages have been identified and characterized in the regenerating fin, the origin of bone cells remains debated. Here, we analysed collagen10a1 (col10a1) expressing cells in the regenerating fin of the medaka (Oryzias latipes) and tested whether they represent an alternative progenitor source for regenerating osteoblasts. Under normal conditions, col10a1 cells are positioned along fin ray segments and in intersegmental regions. Lineage tracing in the amputated fin revealed that col10a1 cells from the stump contribute to the regenerating bony fin rays. However, ablation of col10a1 cells did not abolish fin regeneration suggesting that col10a1 expressing osteoblast progenitors are dispensable for regeneration. Intriguingly, however, after ablation of osterix (osx)/sp7-col10a1 double-positive osteoblasts, col10a1 cells exclusively gave rise to joint cells in the intersegmental region thus identifying a pool of lineage-restricted joint progenitor cells. To identify additional sources for regenerating osteoblasts, we performed clonal lineage analysis. Our data provide the first evidence that after ablation of mature osteoblasts in medaka, transdifferentiation does not account for de novo osteoblast generation. Instead, our findings suggest the presence of lineage restricted progenitor pools in medaka, similar to the situation in zebrafish. After osteoblast ablation, these pools become activated and give rise to fin ray osteoblasts and intersegmental joint cells during regeneration. In summary, we conclude that col10a1-positive cells do not represent an exclusive source for osteoblasts but are progenitors of joint cells in the regenerating fin.

Physiologic and pathologic effects of dietary free fatty acids on cells of the joint.

Fatty acids (FAs) are potent organic compounds that not only can be used as an energy source during nutrient deprivation but are also involved in several essential signaling cascades in cells. Therefore, a balanced intake of different dietary FAs is critical for the maintenance of cellular functions and tissue homeostasis. A diet with an imbalanced fat composition creates a risk for developing metabolic syndrome and various musculoskeletal diseases, including osteoarthritis (OA). In this review, we summarize the current state of knowledge and mechanistic insights regarding the role of dietary FAs, such as saturated FAs, omega-6 polyunsaturated FAs (PUFAs), and omega-3 PUFAs on joint inflammation and OA pathogeneses. In particular, we review how different types of dietary FAs and their derivatives distinctly affect a variety of cells within the joint, including chondrocytes, osteoblasts, osteoclasts, and synoviocytes. Understanding the molecular mechanisms underlying the effects of FAs on metabolic behavior, anabolic, and catabolic processes, as well as the inflammatory response of joint cells, may help identify therapeutic targets for the prevention of metabolic joint diseases.

A noncanonical role for the engulfment gene ELMO1 in neutrophils that promotes inflammatory arthritis.

Rheumatoid arthritis is characterized by progressive joint inflammation and affects ~1% of the human population. We noted single-nucleotide polymorphisms (SNPs) in the apoptotic cell-engulfment genes ELMO1, DOCK2, and RAC1 linked to rheumatoid arthritis. As ELMO1 promotes cytoskeletal reorganization during engulfment, we hypothesized that ELMO1 loss would worsen inflammatory arthritis. Surprisingly, Elmo1-deficient mice showed reduced joint inflammation in acute and chronic arthritis models. Genetic and cell-biology studies revealed that ELMO1 associates with receptors linked to neutrophil function in arthritis and regulates activation and early neutrophil recruitment to the joints, without general inhibition of inflammatory responses. Further, neutrophils from the peripheral blood of human donors that carry the SNP in ELMO1 associated with arthritis display increased migratory capacity, whereas ELMO1 knockdown reduces human neutrophil migration to chemokines linked to arthritis. These data identify 'noncanonical' roles for ELMO1 as an important cytoplasmic regulator of specific neutrophil receptors and promoter of arthritis.

Reduction of Articular and Systemic Inflammation by Kava-241 in a Porphyromonas gingivalis-Induced Arthritis Murine Model.

Rheumatoid arthritis (RA) is an inflammatory disease that has been linked to several risk factors, including periodontitis. Identification of new anti-inflammatory compounds to treat arthritis is needed. We had previously demonstrated the beneficial effect of Kava-241, a kavain-derived compound, in the management of Porphyromonas gingivalis-induced periodontitis. The present study evaluated systemic and articular effects of Kava-241 in an infective arthritis murine model triggered by P. gingivalis bacterial inoculation and primed with a collagen antibody cocktail (CIA) to induce joint inflammation and tissular destruction. Clinical inflammation score and radiological analyses of the paws were performed continuously, while histological assessment was obtained at sacrifice. Mice exposed to P. gingivalis and a CIA cocktail and treated concomitantly with Kava-241 exhibited a reduced clinical inflammatory score and a decreased number of inflammatory cells and osteoclasts within joint. Kava-241 treatment also decreased significantly tumor necrosis factor alpha (TNF-α) in serum from mice injected with a Toll-like receptor 2 or 4 (TLR-2/4) ligand, P. gingivalis-lipopolysaccharide (LPS). Finally, bone marrow-derived macrophages infected with P. gingivalis and exposed to Kava-241 displayed reduced TLR-2/4, reduced mitogen-activated protein kinase (MAPK)-related signal elements, and reduced LPS-induced TNF-α factor (LITAF), all explaining the observed reduction of TNF-α secretion. Taken together, these results emphasized the novel properties of Kava-241 in the management of inflammatory conditions, especially TNF-α-related diseases such as infective RA.

Curcumin protects rabbit articular chondrocytes against sodium nitroprusside-induced apoptosis in vitro.

The preventive and therapeutic effects of curcumin on degeneration of articular (joint) cartilage diseases have rarely been investigated. In the present study, the protective effects of curcumin against sodium nitroprusside (SNP)-induced chondrocyte apoptosis were evaluated and the underlying molecular mechanisms were elucidated. Curcumin was used to as a co-treatment with SNP in chondrocytes, and changes occurring in the cells were observed and evaluated. It was shown using a cell counting kit-8 (CCK-8) assay that curcumin protected the viability of chondrocytes against SNP damage. NO (nitric oxide) from SNP could be scavenged by curcumin. Flow cytometry and Hoechst 33342 staining showed that curcumin not only inhibited the cell apoptosis in a concentration-dependent pattern but also ameliorated the SNP-induced nuclear chromatin damage and reduction of the mitochondrial membrane potential in chondrocytes. In SNP-treated chondrocytes, curcumin downregulated the expression of Bax and cleaved caspase-3 but upregulated the expression of Bcl-2, as shown by western blot. Meanwhile, curcumin administration also protected extracellular matrix (ECM) synthesis and prevented its degradation. Taken together, these results support the hypothesis that curcumin exerts its protective effect on chondrocytes against SNP-induced apoptosis, at least partly, via blocking the mitochondrial-dependent apoptotic pathway and maintaining the metabolic balance of ECM. Thus, curcumin may be a potential candidate to be used as a unique biological agent for the prevent and treatment of osteoarthritis (OA).

A computational model for the joint onset and development.

Joints connect the skeletal components and enable movement. The appearance and development of articulations is due to different genetic, biochemical, and mechanical factors. In the embryonic stage, controlled biochemical processes are critical for organized growth. We developed a computational model, which predicts the appearance, location, and development of joints in the embryonic stage. Biochemical events are modeled with reaction diffusion equations with generic molecules representing molecules that 1) determine the site where the articulation will appear, 2) promote proliferation, and matrix synthesis, and 3) define articular cartilage. Our model accounts for cell differentiation from mesenchymal cells to pre-cartilaginous cells, then cartilaginous cells, and lastly articular cartilage. These reaction-diffusion equations were solved using the finite elements method. From a mesenchymal 'bud' of a phalanx, the model predicts growth, joint cleavage, joint morphology, and articular cartilage formation. Our prediction of the gene expression during development agrees with molecular expression profiles of joint development reported in literature. Our computational model suggests that initial rudiment dimensions affect diffusion profiles result in Turing patterns that dictate sites of cleavage thereby determining the number of joints in a rudiment.

Meniscus ECM-functionalised hydrogels containing infrapatellar fat pad-derived stem cells for bioprinting of regionally defined meniscal tissue.

Injuries to the meniscus of the knee commonly lead to osteoarthritis. Current therapies for meniscus regeneration, including meniscectomies and scaffold implantation, fail to achieve complete functional regeneration of the tissue. This has led to increased interest in cell and gene therapies and tissue engineering approaches to meniscus regeneration. The implantation of a biomimetic implant, incorporating cells, growth factors, and extracellular matrix (ECM)-derived proteins, represents a promising approach to functional meniscus regeneration. The objective of this study was to develop a range of ECM-functionalised bioinks suitable for 3D bioprinting of meniscal tissue. To this end, alginate hydrogels were functionalised with ECM derived from the inner and outer regions of the meniscus and loaded with infrapatellar fat pad-derived stem cells. In the absence of exogenously supplied growth factors, inner meniscus ECM promoted chondrogenesis of fat pad-derived stem cells, whereas outer meniscus ECM promoted a more elongated cell morphology and the development of a more fibroblastic phenotype. With exogenous growth factors supplementation, a more fibrogenic phenotype was observed in outer ECM-functionalised hydrogels supplemented with connective tissue growth factor, whereas inner ECM-functionalised hydrogels supplemented with TGFß3 supported the highest levels of Sox-9 and type II collagen gene expression and sulfated glycosaminoglycans (sGAG) deposition. The final phase of the study demonstrated the printability of these ECM-functionalised hydrogels, demonstrating that their codeposition with polycaprolactone microfibres dramatically improved the mechanical properties of the 3D bioprinted constructs with no noticeable loss in cell viability. These bioprinted constructs represent an exciting new approach to tissue engineering of functional meniscal grafts.

[Imaging of bone and joint destruction].

  Osteoclasts are bone-resorbing giant polykaryons that differentiate from mononuclear macrophage/monocyte-lineage hematopoietic precursors. We have originally established an advanced imaging system for visualizing in vivo behavior of osteoclasts and their precursors with intravital two-photon microscopy. By means of the system, we found that sphingosine-1-phosphate, a lipid mediator enriched in blood, controlled the migratory behavior of osteoclast precursors. We also developed pH-sensing chemical fluorescent probes to detect localized acidification by bone-resorbing osteoclasts on the bone surface in vivo, and identified two distinct functional states of differentiated osteoclasts, 'bone-resorptive' and 'non-resorptive'. In this review, we summarize our recent studies on the dynamics and functions of osteoclasts. Our intravital imaging techniques would be beneficial for studying the cellular dynamics in arthritic inflammation and bone destruction in vivo and would thus be useful for evaluating novel therapies targeting aspects of osteoclast dynamics in patients with bone-destructive diseases.

Native joint-resident mesenchymal stem cells for cartilage repair in osteoarthritis.

The role of native (not culture-expanded) joint-resident mesenchymal stem cells (MSCs) in the repair of joint damage in osteoarthritis (OA) is poorly understood. MSCs differ from bone marrow-residing haematopoietic stem cells in that they are present in multiple niches in the joint, including subchondral bone, cartilage, synovial fluid, synovium and adipose tissue. Research in experimental models suggests that the migration of MSCs adjacent to the joint cavity is crucial for chonodrogenesis during embryogenesis, and also shows that synovium-derived MSCs might be the primary drivers of cartilage repair in adulthood. In this Review, the available data is synthesized to produce a proposed model in which joint-resident MSCs with access to superficial cartilage are key cells in adult cartilage repair and represent important targets for manipulation in 'chondrogenic' OA, especially in the context of biomechanical correction of joints in early disease. Growing evidence links the expression of CD271, a nerve growth factor (NGF) receptor by native bone marrow-resident MSCs to a wider role for neurotrophins in OA pathobiology, the implications of which require exploration since anti-NGF therapy might worsen OA. Recognizing that joint-resident MSCs are comparatively abundant in vivo and occupy multiple niches will enable the optimization of single-stage therapeutic interventions for OA.

Hyaluronan supplementation as a mechanical regulator of cartilage tissue development under joint-kinematic-mimicking loading.

Articular cartilage plays an essential role in joint lubrication and impact absorption. Through this, the mechanical signals are coupled to the tissue's physiological response. Healthy synovial fluid has been shown to reduce and homogenize the shear stress acting on the cartilage surfaces due to its unique shear-thinning viscosity. As cartilage tissues are sensitive to mechanical changes in articulation, it was hypothesized that replacing the traditional culture medium with a healthy non-Newtonian lubricant could enhance tissue development in a cartilage engineering model, where joint-kinematic-mimicking mechanical loading is applied. Different amounts of hyaluronic acid were added to the culture medium to replicate the viscosities of synovial fluid at different health states. Hyaluronic acid supplementation, especially at a physiologically healthy concentration (2.0 mg ml-1), promoted a better preservation of chondrocyte phenotype. The ratio of collagen II to collagen I mRNA was 4.5 times that of the control group, implying better tissue development (however, with no significant difference of measured collagen II content), with a good retention of collagen II and proteoglycan in the mechanically active region. Simulating synovial fluid properties by hyaluronic acid supplementation created a favourable mechanical environment for mechanically loaded constructs. These findings may help in understanding the influence of joint articulation on tissue homeostasis, and moreover, improve methods for functional cartilage tissue engineering.

Role of Piezo Channels in Joint Health and Injury.

Cartilage is an intrinsically mechanically sensitive tissue composed of chondrocytes as the only cell type. Chondrocyte mechanotransduction is not well understood, but recently we identified critical components of the mechanotransduction machinery demonstrating how mechanical stimulation of these cells can be converted into cellular calcium signals. Physiologic mechanical cues induce anabolic responses of (post-mitotic) chondrocytes via transient receptor potential vanilloid 4 ion channels, whereas injurious mechanical stress is transduced by Piezo1 jointly with Piezo2 ion channels. This chapter sheds light on the latter discovery and provides a rationale for follow-up questions, such as the nature of interaction between Piezo1 and Piezo2, and their tethering to the cytoskeleton. These recent insights can be leveraged toward translational medical progress to benefit diagnosis and treatment of osteoarthritis, representing a large and growing unmet medical need in the United States and large parts of the world.

Age-related modulation of angiogenesis-regulating factors in the swine meniscus.

An in-depth knowledge of the native meniscus morphology and biomechanics in its different areas is essential to develop an engineered tissue. Meniscus is characterized by a great regional variation in extracellular matrix components and in vascularization. Then, the aim of this work was to characterize the expression of factors involved in angiogenesis in different areas during meniscus maturation in pigs. The menisci were removed from the knee joints of neonatal, young and adult pigs, and they were divided into the inner, intermediate and outer areas. Vascular characterization and meniscal maturation were evaluated by immunohistochemistry and Western blot analysis. In particular, expression of the angiogenic factor Vascular Endothelial Growth Factor (VEGF) and the anti-angiogenic marker Endostatin (ENDO) was analysed, as well as the vascular endothelial cadherin (Ve-CAD). In addition, expression of Collagen II (COLL II) and SOX9 was examined, as markers of the fibro-cartilaginous differentiation. Expression of VEGF and Ve-CAD had a similar pattern in all animals, with a significant increase from the inner to the outer part of the meniscus. Pooling the zones, expression of both proteins was significantly higher in the neonatal meniscus than in young and adult menisci. Conversely, the young meniscus revealed a significantly higher expression of ENDO compared to the neonatal and adult ones. Analysis of tissue maturation markers showed an increase in COLL II and a decrease in SOX9 expression with age. These preliminary data highlight some of the changes that occur in the swine meniscus during growth, in particular the ensemble of regulatory factors involved in angiogenesis.

Analysis of synovial fluid of the Capybara's stifle joints.

BACKGROUND: Although normal synovial fluid has been well characterized in domestic animals such as dogs, cats, horses, and cows, the available information on larger rodents is scarce. OBJECTIVES: The purpose of the study was to analyze the physical, chemical, and cytologic characteristics of the synovial fluid in stifle joints of Capybaras. METHODS: Five free-ranging adult female Capybaras (Hydrochoerus hydrochaeris), weighing from 37 to 56 kg were used. Synovial fluid was obtained by aspiration of 10 stifle joints. Samples were analyzed for physical, chemical, and cytologic properties. RESULTS: Spontaneous clotting was negative in 9 samples. Most synovial fluids had pH 8, and protein concentrations ranged from 1.6 to 3.6 g/dL. The mucin clot test was good in all 6 samples that were tested. Nucleated cell counts ranged from 140 to 508 cells/µL. Relative differential leukocyte counts demonstrated a predominance of mononuclear cells (97.6%), including 76.2% undifferentiated mononuclear cells, 18.1% macrophages, and 3.66% lymphocytes. Polymorphonuclear cells included 1.83% neutrophils and 0.2% eosinophils. CONCLUSION: The synovial stifle joint fluid of healthy free-ranging adult Capybaras is clear, colorless, viscous, and with chemical features and cytologic findings similar to those seen in domestic animals.

Inhibition of cartilage degradation and suppression of PGE2 and MMPs expression by pomegranate fruit extract in a model of posttraumatic osteoarthritis.

OBJECTIVE: Osteoarthritis (OA) is characterized by cartilage degradation in the affected joints. Pomegranate fruit extract (PFE) inhibits cartilage degradation in vitro. The aim of this study was to determine whether oral consumption of PFE inhibits disease progression in rabbits with surgically induced OA. METHODS: OA was surgically induced in the tibiofemoral joints of adult New Zealand White rabbits. In one group, animals were fed PFE in water for 8 wk postsurgery. In the second group, animals were fed PFE for 2 wk before surgery and for 8 wk postsurgery. Histologic assessment and scoring of the cartilage was per Osteoarthritis Research Society International guidelines. Gene expression and matrix metalloproteinases (MMP) activity were determined using quantitative reverse transcriptase polymerase chain reaction and fluorometric assay, respectively. Interleukin (IL)-1 ß, MMP-13, IL-6, prostaglandin (PG)E2, and type II collagen (COL2A1) levels in synovial fluid/plasma/culture media were quantified using enzyme-linked immunosorbent assay. Expression of active caspase-3 and poly (ADP-ribose) polymerase p85 was determined by immunohistochemistry. Effect of PFE and inhibitors of MMP-13, mitogen-activated protein kinase (MAPK) and nuclear factor (NF)-κB was studied in IL-1 ß-stimulated rabbit articular chondrocytes. RESULTS: Safranin-O-staining and chondrocyte cluster formation was significantly reduced in the anterior cruciate ligament transaction plus PFE fed groups. Expression of MMP-3, MMP-9, and MMP-13 mRNA was higher in the cartilage of rabbits given water alone but was significantly lower in the animals fed PFE. PFE-fed rabbits had lower IL-6, MMP-13, and PGE2 levels in the synovial fluid and plasma, respectively, and showed higher expression of aggrecan and COL2A1 mRNA. Significantly higher numbers of chondrocytes were positive for markers of apoptosis in the joints of rabbits with OA given water only compared with those in the PFE-fed groups. PFE pretreatment significantly reduced IL-1 ß induced IL-6 and MMPs expression in rabbit articular chondrocytes. These effects were also mimicked using MMP-13, MAPK, and NF-κB inhibitors in IL-1 ß-stimulated rabbit chondrocytes. In an in vitro activity assay, PFE blocked the activity of MMP-13. Like MAPK and NF-κB inhibitors, PFE was also effective in inhibiting IL-1 ß-induced PGE2 production in rabbit chondrocytes. PFE also reversed the inhibitory effect of IL-1ß on COL2A1 mRNA and protein expression in IL-1 ß-stimulated rabbit chondrocytes. CONCLUSION: The present data highlight the chondroprotective effects of PFE oral consumption in a model of posttraumatic OA and suggest that PFE-derived compounds may have potential value in the management of OA.

Building and maintaining joints by exquisite local control of cell fate.

We owe the flexibility of our bodies to sophisticated articulations between bones. Establishment of these joints requires the integration of multiple tissue types: permanent cartilage that cushions the articulating bones, synovial membranes that enclose a lubricating fluid-filled cavity, and a fibrous capsule and ligaments that provide structural support. Positioning the prospective joint region involves establishment of an "interzone" region of joint progenitor cells within a nascent cartilage condensation, which is achieved through the interplay of activators and inhibitors of multiple developmental signaling pathways. Within the interzone, tight regulation of BMP and TGFß signaling prevents the hypertrophic maturation of joint chondrocytes, in part through downstream transcriptional repressors and epigenetic modulators. Synovial cells then acquire further specializations through expression of genes that promote lubrication, as well as the formation of complex structures such as cavities and entheses. Whereas genetic investigations in mice and humans have uncovered a number of regulators of joint development and homeostasis, recent work in zebrafish offers a complementary reductionist approach toward understanding joint positioning and the regulation of chondrocyte fate at joints. The complexity of building and maintaining joints may help explain why there are still few treatments for osteoarthritis, one of the most common diseases in the human population. A major challenge will be to understand how developmental abnormalities in joint structure, as well as postnatal roles for developmental genes in joint homeostasis, contribute to birth defects and degenerative diseases of joints. WIREs Dev Biol 2017, 6:e245. doi: 10.1002/wdev.245 For further resources related to this article, please visit the WIREs website.

Joint Development Involves a Continuous Influx of Gdf5-Positive Cells.

Synovial joints comprise several tissue types, including articular cartilage, the capsule, and ligaments. All of these compartments are commonly assumed to originate from an early set of Gdf5-expressing progenitors populating the interzone domain. Here, we provide evidence that joints develop through a continuous influx of cells into the interzone, where they contribute differentially to forming joint tissues. Using a knockin Gdf5-CreER(T2) mouse, we show that early labeling of Gdf5-positive interzone cells failed to mark the entire organ. Conversely, multiple Cre activation steps indicated a contribution of these cells to various joint compartments later in development. Spatiotemporal differences between Gdf5 and tdTomato reporter expression support the notion of a continuous recruitment process. Finally, differential contribution of Gdf5-positive cells to various tissues suggests that the spatiotemporal dynamics of Gdf5 expression may instruct lineage divergence. This work supports the influx model of joint development, which may apply to other organogenic processes.