Erika H. Noss

Cadherin-11 in Synovial Lining Formation and Pathology in Arthritis

The normal synovium forms a membrane at the edges of joints and provides lubrication and nutrients for the cartilage. In rheumatoid arthritis, the synovium is the site of inflammation, and it participates in an organized tissue response that damages cartilage and bone. We identified cadherin-11 as essential for the development of the synovium. Cadherin-11–deficient mice have a hypoplastic synovial lining, display a disorganized synovial reaction to inflammation, and are resistant to inflammatory arthritis. Cadherin-11 therapeutics prevent and reduce arthritis in mouse models. Thus, synovial cadherin-11 determines the behavior of synovial cells in their proinflammatory and destructive tissue response in inflammatory arthritis.

The role and therapeutic implications of fibroblast-like synoviocytes in inflammation and cartilage erosion in rheumatoid arthritis

Summary: Fibroblast‐like synoviocytes (FLS) are resident mesenchymal cells of synovial joints that have been recognized to play an increasingly important role in the pathogenesis of rheumatoid arthritis (RA). Activation of FLS in the setting of RA leads to the production of a broad array of cell surface and soluble mediators that help to recruit, retain, and activate both cells of the immune system and resident joint cells, leading to the promotion of ongoing inflammation and tissue destruction. The ability of FLS to stimulate both inflammation and tissue damage suggests that this cell type may be a unique target for the treatment of inflammatory arthritis. Greater understanding of how FLS are activated and how they interact with other cells in the RA synovium may provide insights that allow development of novel agents for RA therapy.

Integrins traffic rapidly via circular dorsal ruffles and macropinocytosis during stimulated cell migration

In response to growth factor stimulation, integrins transit through recycling endosomes to reach newly forming focal adhesions at the cell’s leading edge.

Cadherin-11 regulates fibroblast inflammation

Fibroblasts are important participants in inflammation. Although not leukocytes, their capacity to produce cytokines, chemokines, and other inflammatory factors locally in tissues suggests that they can contribute to inflammatory diseases. For example, fibroblasts in a rheumatoid arthritis (RA) joint are a dominant source of IL-6 and RANKL in the synovium, both of which are therapeutic targets for inflammation and bone erosion. Previously, we found that fibroblasts can be targeted by mAb directed against cadherin-11 (cad-11), a mesenchymal cadherin that fibroblasts selectively express. Targeting cad-11 significantly reduced inflammation as assessed by joint swelling and clinical inflammation scores. However, the mechanism by which anti–cad-11 reduced inflammation was not known. Here, we show that cad-11 engagement induces synovial fibroblasts to secret proinflammatory cytokines including IL-6. Cad-11 engagement strongly synergized with TNF-α and IL-1β in the induction of IL-6. Importantly, cad-11 activated MAP kinases and NF-κB for IL-6 induction. IL-6 levels in ankles of inflamed joints were reduced in cad-11 mutant mice compared to wild-type mice with inflammatory arthritis. Thus, we suggest that cad-11 modulates synovial fibroblasts to evoke inflammatory factors that may contribute to the inflammatory process in RA.

Modulation of matrix metalloproteinase production by rheumatoid arthritis synovial fibroblasts after cadherin 11 engagement

OBJECTIVE Cadherin 11 is a homophilic cell-to-cell adhesion molecule expressed on joint synovial fibroblasts. Absence of cadherin 11 in a mouse rheumatoid arthritis (RA) model led to striking reductions in cartilage erosion. Matrix metalloproteinases (MMPs) are enzymes expressed by synovial fibroblasts important for cartilage erosion. The objective of this study was to determine if synovial fibroblast MMP production is regulated by cadherin 11. METHODS To mimic cadherin 11 engagement, human RA synovial fibroblasts were stimulated with a chimeric construct consisting of the cadherin 11 extracellular domain linked to the human IgG1 Fc domain (Cad-11-Fc). Effects on MMP production were measured by enzyme-linked immunosorbent assay, quantitative reverse transcription-polymerase chain reaction analysis, and immunoblotting. RESULTS Human Cad-11-Fc up-regulated MMP-1 and MMP-3 protein production by RA synovial fibroblasts, both alone and in synergy with tumor necrosis factor α. This up-regulation required cell cadherin 11 engagement, since a mutant Cad-11-Fc with reduced binding affinity stimulated significantly less MMP production. Also, short hairpin RNA (shRNA) cadherin 11 silencing almost completely inhibited Cad-11-Fc-induced MMP expression. Cad-11-Fc stimulation increased RA synovial fibroblast MMP messenger RNA levels. It also increased the phosphorylation of the MAPKs JNK, ERK, and p38 kinase, the phosphorylation of NF-κB p65, and the nuclear translocation of activator protein 1 transcription factor. MAPK and NF-κB inhibitors partially blocked RA synovial fibroblast MMP expression. CONCLUSION Cadherin 11 engagement stimulates increased synthesis of several MMPs by RA synovial fibroblasts in a MAPK- and NF-κB-dependent manner. These results underscore the existence of a pathway by which cadherin 11 regulates MMP production and has important implications for joint destruction in RA.

Genetic polymorphism directs IL-6 expression in fibroblasts but not selected other cell types

Significance IL-6 is an important inflammatory cytokine and treatment target in rheumatoid arthritis. Genetic variation in the IL-6 proximal promoter is well-described, but its contribution to IL-6 regulation is uncertain. Numerous studies have shown contradictory associations between disease states and IL-6 promoter polymorphisms. Synovial fibroblasts are a major IL-6 producer in rheumatoid arthritis. We report a striking association between increased fibroblast IL-6 production and the IL-6 proximal promoter single nucleotide polymorphism rs1800795 minor allele genotype. In stark contrast, no association between human monocyte or HeLa cell IL-6 production and rs1800795 association was observed. These results point to fibroblast-specific pathways in IL-6 regulation and highlight that full understanding of the effects of genetic variation requires examination across cell types. Interleukin (IL)-6 blockade is an effective treatment for rheumatoid arthritis (RA), and synovial fibroblasts are a major IL-6 producer in the inflamed joint. We found that human RA and osteoarthritis (OA) synovial fibroblasts derived from independent donors reproducibly segregated into low, medium, and high IL-6 producers, independent of stimulus, cell passage, or disease state. IL-6 expression pattern correlated strongly with total mRNA expression, not mRNA stability, suggesting transcriptional rather than posttranscriptional regulation. High-fibroblast IL-6 expression was significantly associated with the IL-6 proximal promoter single nucleotide polymorphism (SNP) rs1800795 minor allele (CC) genotype. In contrast, no association between this SNP and IL-6 production was detected in CD14+ monocytes, another major producer of synovial IL-6. Luciferase expression assays confirmed that this SNP was associated with differential IL-6 expression in fibroblasts. To date, several association studies examining rs1800795 allele frequency and disease risk have reported seemingly conflicting results ranging from no association to association with either the major or minor allele across a spectrum of conditions, including cancer and autoimmune, cardiovascular, infectious, and metabolic diseases. This study points to a prominent contribution from promoter genetic variation in fibroblast IL-6 regulation, but not in other IL-6–producing cell types. We propose that some of the heterogeneity in these clinical studies likely reflects the cellular source of IL-6 in specific diseases, much of which may be produced by nonhematopoietic cells. These results highlight that functional analysis of disease-associated SNPs on gene expression and pathologic processes must consider variation in diverse cell types.

Functionally distinct disease-associated fibroblast subsets in rheumatoid arthritis

Fibroblasts regulate tissue homeostasis, coordinate inflammatory responses, and mediate tissue damage. In rheumatoid arthritis (RA), synovial fibroblasts maintain chronic inflammation which leads to joint destruction. Little is known about fibroblast heterogeneity or if aberrations in fibroblast subsets relate to pathology. Here, we show functional and transcriptional differences between fibroblast subsets from human synovial tissues using bulk transcriptomics of targeted subpopulations and single-cell transcriptomics. We identify seven fibroblast subsets with distinct surface protein phenotypes, and collapse them into three subsets by integrating transcriptomic data. One fibroblast subset, characterized by the expression of proteins podoplanin, THY1 membrane glycoprotein and cadherin-11, but lacking CD34, is threefold expanded in patients with RA relative to patients with osteoarthritis. These fibroblasts localize to the perivascular zone in inflamed synovium, secrete proinflammatory cytokines, are proliferative, and have an in vitro phenotype characteristic of invasive cells. Our strategy may be used as a template to identify pathogenic stromal cellular subsets in other complex diseases.Synovial fibroblasts are thought to be central mediators of joint destruction in rheumatoid arthritis (RA). Here the authors use single-cell transcriptomics and flow cytometry to identify synovial fibroblast subsets that are expanded and display distinct tissue distribution and function in patients with RA.

Clinical course and management of a consecutive series of patients with "healed temporal arteritis"

Objective. To describe the clinical course and management of patients with a pathologic diagnosis of “healed” giant cell arteritis (GCA), and to determine whether previously published histological descriptions of healed arteritis can identify patients with a greater likelihood of clinically significant arteritis. Methods. All temporal artery biopsy reports between 1994 and 2003 were examined for a diagnosis of “healed arteritis.” Two rheumatologists abstracted the medical record for presenting features, physical findings, comorbid conditions, and data on treatment and outcomes. One pathologist, blinded to the clinical data, reviewed all specimens and reinterpreted the biopsies according to published histological descriptions of healed arteritis. Results. Forty-seven patients with an initial pathologic diagnosis of healed arteritis were identified. In 54% of these patients, corticosteroid therapy did not change after the diagnosis of healed arteritis was documented in the pathology report. Seventy percent were ultimately treated with no corticosteroids or low-moderate corticosteroid regimens. Only 32% of the initial cases were confirmed upon review of the biopsies using standardized histological criteria. Patients with confirmed healed arteritis were more likely to have a documented history of polymyalgia rheumatica/GCA and a longer duration of corticosteroid treatment before biopsy. These patients were not more likely to have adverse outcomes. Conclusion. In this case series, the diagnosis of healed arteritis had little effect on treatment decisions. In most cases, the initial pathologic diagnosis of healed arteritis was not confirmed when biopsies were reviewed by a single pathologist using uniform histological criteria.

Autocrine Loop Involving IL-6 Family Member LIF, LIF Receptor, and STAT4 Drives Sustained Fibroblast Production of Inflammatory Mediators

SUMMARY Fibroblasts are major contributors to and regulators of inflammation and dominant producers of interleukin‐6 (IL‐6) in inflammatory diseases like rheumatoid arthritis. Yet, compared to leukocytes, the regulation of inflammatory pathways in fibroblasts is largely unknown. Here, we report that analyses of genes coordinately upregulated with IL‐6 pointed to STAT4 and leukemia inhibitory factor (LIF) as potentially linked. Gene silencing revealed that STAT4 was required for IL‐6 transcription. STAT4 was recruited to the IL‐6 promoter after fibroblast activation, and LIF receptor (LIFR) and STAT4 formed a molecular complex that, together with JAK1 and TYK2 kinases, controlled STAT4 activation. Importantly, a positive feedback loop involving autocrine LIF, LIFR, and STAT4 drove sustained IL‐6 transcription. Besides IL‐6, this autorine loop also drove the production of other key inflammatory factors including IL‐8, granulocyte‐colony stimulating factor (G‐CSF), IL‐33, IL‐11, IL‐1&agr;, and IL‐1&bgr;. These findings define the transcriptional regulation of fibroblast‐mediated inflammation as distinct from leukocytes. Graphical Abstract Figure. No Caption available. HighlightsDuring inflammation, human fibroblasts upregulate LIF and STAT4LIF acts in an autocrine manner via LIF receptor to promote STAT4 activationActivated STAT4 together with NF‐&kgr;B/p65‐p52 and C/EBP&bgr; enhances IL‐6 transcriptionLIFR/STAT4 circuit also regulates IL‐8, G‐CSF, IL‐33, IL‐11, IL‐1&agr;, and IL‐1&bgr; &NA; Growing evidence implicates fibroblasts as inflammatory cells in sites of peripheral inflammation. Nguyen and colleagues demonstrate that regulation of IL‐6 along with a set of other inflammatory cytokines and chemokines is regulated by a positive feedback loop involving LIF, LIF receptor, and STAT4 that selectively operates in fibroblasts.

Evidence for cadherin-11 cleavage in the synovium and partial characterization of its mechanism

IntroductionEngagement of the homotypic cell-to-cell adhesion molecule cadherin-11 on rheumatoid arthritis (RA) synovial fibroblasts with a chimeric molecule containing the cadherin-11 extracellular binding domain stimulated cytokine, chemokine, and matrix metalloproteinases (MMP) release, implicating cadherin-11 signaling in RA pathogenesis. The objective of this study was to determine if cadherin-11 extracellular domain fragments are found inside the joint and if a physiologic synovial fibroblast cleavage pathway releases those fragments.MethodsCadherin-11 cleavage fragments were detected by western blot in cell media or lysates. Cleavage was interrupted using chemical inhibitors or short-interfering RNA (siRNA) gene silencing. The amount of cadherin-11 fragments in synovial fluid was measured by western blot and ELISA.ResultsSoluble cadherin-11 extracellular fragments were detected in human synovial fluid at significantly higher levels in RA samples compared to osteoarthritis (OA) samples. A cadherin-11 N-terminal extracellular binding domain fragment was shed from synovial fibroblasts after ionomycin stimulation, followed by presenilin 1 (PSN1)-dependent regulated intramembrane proteolysis of the retained membrane-bound C-terminal fragments. In addition to ionomycin-induced calcium flux, tumor necrosis factor (TNF)-α also stimulated cleavage in both two- and three-dimensional fibroblast cultures. Although cadherin-11 extracellular domains were shed by a disintegrin and metalloproteinase (ADAM) 10 in several cell types, a novel ADAM- and metalloproteinase-independent activity mediated shedding in primary human fibroblasts.ConclusionsCadherin-11 undergoes ectodomain shedding followed by regulated intramembrane proteolysis in synovial fibroblasts, triggered by a novel sheddase that generates extracelluar cadherin-11 fragments. Cadherin-11 fragments were enriched in RA synovial fluid, suggesting they may be a marker of synovial burden and may function to modify cadherin-11 interactions between synovial fibroblasts.