Bona fide dendritic cells are pivotal precursors for osteoclasts.

OBJECTIVES: Osteoclasts (OCs) are myeloid-derived multinucleated cells uniquely able to degrade bone. However, the exact nature of their myeloid precursors is not yet defined. METHODS: CD11c-diphtheria toxin receptor (CD11cDTR) transgenic mice were treated with diphtheria toxin (DT) or phosphate buffered saline (PBS) during serum transfer arthritis (STA) and human tumour necrosis factor transgenic (hTNFtg) arthritis and scored clinically and histologically. We measured cytokines in synovitis by quantitative polymerase chain reaction (qPCR). We performed ovariectomy in CD11cDTR mice treated with PBS or DT. We analysed CD11cDTR, CD11c-Cre/CX3CR1-STOP-DTR and Zbtb46-DTR-treated mice with DT using histomorphometry and OC of CD11c and Zbtb46 fate reporter mice by fluorescent imaging. We sorted murine and human OC precursors and stimulated them with macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor kappa-B ligand (RANKL) to generate OCs. RESULTS: Targeting CD11c+ cells in vivo in models of inflammatory arthritis (STA and hTNFtg) ameliorates arthritis by reducing inflammatory bone destruction and OC generation. Targeting CD11c-expressing cells in unchallenged mice removes all OCs in their long bones. OCs do not seem to be derived from CD11c+ cells expressing CX3CR1+, but from Zbtb46+conventional dendritic cells (cDCs) as all OCs in Zbtb46-Tomato fate reporter mice are Tomato+. In line, administration of DT in Zbtb46-DTR mice depletes all OCs in long bones. Finally, human CD1c-expressing cDCs readily differentiated into bone resorbing OCs. CONCLUSION: Taken together, we identify DCs as important OC precursors in bone homeostasis and inflammation, which might open new avenues for therapeutic interventions in OC-mediated diseases.

'SMASH' recommendations for standardised microscopic arthritis scoring of histological sections from inflammatory arthritis animal models.

Animal models for inflammatory arthritides such as rheumatoid arthritis (RA) and psoriatic arthritis are widely accepted and frequently used to identify pathological mechanisms and validate novel therapeutic strategies. Unfortunately, many publications reporting on these animal studies lack detailed description and appropriate assessment of the distinct histopathological features of arthritis: joint inflammation, cartilage damage and bone erosion. Therefore, the European consortium BeTheCure, consisting of 38 academic and industrial partners from 15 countries, set as goal to standardise the histological evaluation of joint sections from animal models of inflammatory arthritis. The consensual approach of a task force including 16 academic and industrial scientists as well as laboratory technicians has resulted in the development of the Standardised Microscopic Arthritis Scoring of Histological sections ('SMASH') recommendations for a standardised processing and microscopic scoring of the characteristic histopathological features of arthritis, exemplified by four different rodent models for arthritis: murine collagen-induced arthritis, collagen-antibody-induced arthritis, human tumour necrosis factor transgenic Tg197 mice and rat pristane-induced arthritis, applicable to any other inflammatory arthritis model. Through standardisation, the SMASH recommendations are designed to improve and maximise the information derived from in vivo arthritis experiments and to promote reproducibility and transparent reporting on such studies. In this manuscript, we will discuss and provide recommendations for analysis of histological joint sections: identification of the regions of interest, sample preparation, staining procedures and quantitative scoring methods. In conclusion, awareness of the different features of the arthritis pathology in animal models of inflammatory arthritis is of utmost importance for reliable research outcome, and the standardised histological processing and scoring methods in these SMASH recommendations will help increase uniformity and reproducibility in preclinical research on inflammatory arthritis.

CCR6 controls autoimmune but not innate immunity-driven experimental arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease, characterized by synovial infiltration of various inflammatory cells. Chemokines are involved in controlling the recruitment of different cell types into the synovial membrane. The role of CCR6 in the development of arthritis so far remains unclear. In this study, we investigated the role of CCR6 in the pathogenesis of arthritis using three different murine arthritis models. Compared to WT animals, CCR6-/- mice developed less clinical signs of arthritis in the collagen-induced arthritis model but not in the K/BxN serum transfer arthritis model and in the human tumour necrosis factor transgenic arthritis model, suggesting a defect in adaptive effector functions but intact innate effector functions in the development of arthritis in CCR6-/- animals. In line with this, anti-collagen antibody levels were significantly reduced in CCR6-/- mice compared with WT mice. Moreover, we demonstrate enhanced osteoclastogenesis in vitro in CCR6-/- mice compared with WT mice. However, we did not detect differences in bone mass under steady state conditions in vivo between WT and CCR6-deficient mice. These data suggest that CCR6 is crucially involved in adaptive but not in innate immunity-driven arthritis. CCR6 or its chemokine ligand CCL20 might represent a possible new target for the treatment of RA.

Non-classical monocytes as mediators of tissue destruction in arthritis.

OBJECTIVES: Bone destruction in rheumatoid arthritis is mediated by osteoclasts (OC), which are derived from precursor cells of the myeloid lineage. The role of the two monocyte subsets, classical monocytes (expressing CD115, Ly6C and CCR2) and non-classical monocytes (which are CD115 positive, but low in Ly6C and CCR2), in serving as precursors for OC in arthritis is still elusive. METHODS: We investigated CCR2-/- mice, which lack circulating classical monocytes, crossed into hTNFtg mice for the extent of joint damage. We analysed monocyte subsets in hTNFtg and K/BxN serum transfer arthritis by flow cytometry. We sorted monocyte subsets and analysed their potential to differentiate into OC and their transcriptional response in response to RANKL by RNA sequencing. With these data, we performed a gene ontology enrichment analysis and gene set enrichment analysis. RESULTS: We show that in hTNFtg arthritis local bone erosion and OC generation are even enhanced in the absence of CCR2. We further show the numbers of non-classical monocytes in blood are elevated and are significantly correlated with histological signs of joint destruction. Sorted non-classical monocytes display an increased capacity to differentiate into OCs. This is associated with an increased expression of signal transduction components of RANK, most importantly TRAF6, leading to an increased responsiveness to RANKL. CONCLUSION: Therefore, non-classical monocytes are pivotal cells in arthritis tissue damage and a possible target for therapeutically intervention for the prevention of inflammatory joint damage.

MicroRNA-146a governs fibroblast activation and joint pathology in arthritis.

Synovial fibroblasts are key cells orchestrating the inflammatory response in arthritis. Here we demonstrate that loss of miR-146a, a key epigenetic regulator of the innate immune response, leads to increased joint destruction in a TNF-driven model of arthritis by specifically regulating the behavior of synovial fibroblasts. Absence of miR-146a in synovial fibroblasts display a highly deregulated gene expression pattern and enhanced proliferation in vitro and in vivo. Deficiency of miR-146a induces deregulation of tumor necrosis factor (TNF) receptor associated factor 6 (TRAF6) in synovial fibroblasts, leading to increased proliferation. In addition, loss of miR-146a shifts the metabolic state of fibroblasts towards glycolysis and augments the ability of synovial fibroblasts to support the generation of osteoclasts by controlling the balance of osteoclastogenic regulatory factors receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG). Bone marrow transplantation experiments confirmed the importance of miR-146a in the radioresistant mesenchymal compartment for the control of arthritis severity, in particular for inflammatory joint destruction. This study therefore identifies microRNA-146a as an important local epigenetic regulator of the inflammatory response in arthritis. It is a central element of an anti-inflammatory feedback loop in resident synovial fibroblasts, who are orchestrating the inflammatory response in chronic arthritis. MiR-146a restricts their activation, thereby preventing excessive tissue damage during arthritis.

Tumor necrosis factor-inhibiting therapy preferentially targets bone destruction but not synovial inflammation in a tumor necrosis factor-driven model of rheumatoid arthritis.

OBJECTIVE: To investigate how tumor necrosis factor (TNF)-inhibiting therapy affects bone destruction and inflammation in a TNF-driven mouse model of rheumatoid arthritis. METHODS: In order to evaluate the influence of TNF on osteoclastogenesis in vitro, different concentrations of TNF were added to spleen cell-derived monocytes in the absence or presence of different concentrations of RANKL. In addition, the effects of TNF inhibition on osteoclast precursors as well as local bone destruction in vivo were assessed by treating TNF-transgenic mice with different doses of adalimumab. RESULTS: TNF stimulated osteoclastogenesis mainly by increasing the number of osteoclast precursor cells in vitro. This TNF effect was independent of the presence of RANKL. In the hTNF-transgenic mouse model of destructive arthritis, low-dose TNF-inhibiting therapy with adalimumab had no effect on synovial inflammation but significantly inhibited local bone destruction and the generation of osteoclasts. This inhibition was accompanied by a reduction in the number of c-Fms-positive osteoclast precursor cells in the bone marrow and a reduction of the osteoclast precursor pools in the blood and inflamed synovial membrane of hTNF-transgenic mice. CONCLUSION: Low-dose TNF-inhibiting therapy significantly reduces bone erosions by reducing the number of circulating and joint-invading osteoclast precursors. This effect is uncoupled from its antiinflammatory action.

Early structural changes in cartilage and bone are required for the attachment and invasion of inflamed synovial tissue during destructive inflammatory arthritis.

OBJECTIVE: To elucidate the mechanisms involved in cartilage damage in an experimental model of rheumatoid arthritis (RA) by specifically addressing the time course of extracellular matrix degradation and the contribution of cell-matrix interactions for initiation and perpetuation of this process. METHODS: The human tumour necrosis factor (TNF) transgenic (hTNFtg) mouse model of RA was used to analyse the time course of pannus attachment to the cartilage and cartilage destruction, respectively, and crossed hTNFtg mice with interleukin (IL)-1(-/-) animals were used to investigate the role of IL-1 on these TNF-induced mechanisms in vivo. In addition, an in vitro attachment assay using synovial fibroblasts (SFs) from hTNFtg mice and freshly isolated articular cartilage was used to determine the role of proteoglycan loss in attachment of SFs and the role of the transmembrane heparan sulfate proteoglycan syndecan-4. RESULTS: In vivo analyses of hTNFtg mice showed that proteoglycan loss induced by IL-1 precedes and constitutes an important prerequisite for these processes as, in hTNFtg mice, IL-1 deficiency protected from the loss of cartilage proteoglycans and almost completely prevented the attachment and subsequent invasion of inflamed synovial tissue into cartilage. In vitro studies confirmed that loss of cartilage proteoglycans is required for attachment of SFs and that syndecan-4 is prominently involved in SF attachment and activation. CONCLUSIONS: The results of this study suggest that the loss of cartilage proteoglycans is an early event in the course of destructive arthritis that facilitates the attachment of the inflamed synovial membrane and also initiates matrix degradation and inflammation through cell-matrix interactions.

Essential role of microRNA-155 in the pathogenesis of autoimmune arthritis in mice.

OBJECTIVE: MicroRNAs (miRNA) are a new class of regulatory elements. Altered expression of miRNA has been demonstrated in the inflamed joints of patients with rheumatoid arthritis (RA). The aim of this study was to examine the role of miRNA in the pathogenesis of autoimmune arthritis, using 2 murine models. METHODS: Collagen-induced arthritis (CIA) and K/BxN serum-transfer arthritis were induced in wild-type (WT) and miR-155-deficient (miR-155(-/-) ) mice. The severity of arthritis was determined clinically and histologically. Anticollagen antibodies and cytokines were measured by enzyme-linked immunosorbent assay. The cellular composition of the draining lymph nodes after induction of CIA was measured by flow cytometry. RESULTS: The miR-155(-/-) mice did not develop CIA. Deficiency in miR-155 prevented the generation of pathogenic autoreactive B and T cells, since anticollagen antibodies and the expression levels of antigen-specific T cells were strongly reduced in miR-155(-/-) mice. Moreover, Th17 polarization of miR-155(-/-) mouse T cells was impaired, as shown by a significant decrease in the levels of interleukin-17 (IL-17) and IL-22. In the K/BxN serum-transfer arthritis model, which only depends on innate effector mechanisms, miR-155(-/-) mice showed significantly reduced local bone destruction, attributed to reduced generation of osteoclasts, although the severity of joint inflammation was similar to that in WT mice. CONCLUSION: These results demonstrate that miR-155 is essentially involved in the adaptive and innate immune reactions leading to autoimmune arthritis, and therefore miR-155 might provide a novel target for the treatment of patients with RA.

Analysis of combined deficiency of interleukin-1 and -6 versus single deficiencies in TNF-mediated arthritis and systemic bone loss.

AIMS: Insufficient treatment response in rheumatoid arthritis (RA) patients requires novel treatment strategies to halt disease progression. The potential benefit of combination of cytokine-inhibitors in RA is still unclear and needs further investigation. To explore the impact of combined deficiency of two major cytokines, namely interleukin (IL)-1 and IL-6, in this study double deficient mice for IL-1αß and IL-6 were investigated in different tumour necrosis factor (TNF)-driven inflammatory bone disorders, namely peripheral arthritis and sacroiliitis, as well as systemic bone loss. METHODS: Disease course, histopathological features of arthritis, and micro-CT (µCT) bone analysis of local and systemic bone loss were assessed in 15-week-old IL1-/-IL6-/- hTNFtg in comparison to IL1-/- hTNFtg, IL6-/- hTNFtg, and hTNFtg mice. µCT bone analysis of single deficient and wild-type mice was also performed. RESULTS: Combined deficiency of IL-1/IL-6 markedly ameliorated TNF-mediated arthritis and bilateral sacroiliitis, but without additive benefits compared to single IL-1 deficiency. This finding confirms the important role of IL-1 and the marginal role of IL-6 in TNF-driven pathways of local joint damage, but questions the efficacy of potential combinatorial therapies of IL-1 and IL-6 in treatment of RA. In contrast, combined deficiency of IL-1/IL-6 led to an additive protective effect on TNF-driven systemic bone loss compared to single IL-1 and IL-6 deficiency. This finding clearly indicates a common contribution of both IL-1 and IL-6 in TNF-driven systemic bone loss, and points to a discrepancy of cytokine dependency in local and systemic TNF-driven mechanisms of inflammatory arthritis. CONCLUSION: Combinatorial treatments in RA might provide different benefits to inflammatory local arthritis and systemic comorbidities. Cite this article: Bone Joint Res 2022;11(7):484-493.

CD44 is a determinant of inflammatory bone loss.

Chronic inflammation is a major trigger of local and systemic bone loss. Disintegration of cell-matrix interaction is a prerequisite for the invasion of inflammatory tissue into bone. CD44 is a type I transmembrane glycoprotein that connects a variety of extracellular matrix proteins to the cell surface. Tumor necrosis factor (TNF) is a major inducer of chronic inflammation and its overexpression leads to chronic inflammatory arthritis. By generating CD44(-/-) human TNF-transgenic (hTNFtg) mice, we show that destruction of joints and progressive crippling is far more severe in hTNFtg mice lacking CD44, which also develop severe generalized osteopenia. Mutant mice exhibit an increased bone resorption due to enhanced number, size, and resorptive capacity of osteoclasts, whereas bone formation and osteoblast differentiation are not affected. Responsiveness of CD44-deficient osteoclasts toward TNF is enhanced and associated with increased activation of the p38 mitogen-activated protein kinase. These data identify CD44 as a critical inhibitor of TNF-driven joint destruction and inflammatory bone loss.

Vitamin D receptor regulates TNF-mediated arthritis.

OBJECTIVE: Reduced vitamin D intake has been linked to increased susceptibility to develop rheumatoid arthritis (RA) and vitamin D deficiency is associated with increased disease activity in RA patients. The pathophysiological role of vitamin D in joint inflammation is, however, unclear. METHODS: To determine the influence of absent vitamin D signalling in chronic arthritis, vitamin D receptor (VDR)-deficient mice were crossed with human tumour necrosis factor (TNF) transgenic mice (hTNFtg), which spontaneously develop chronic arthritis. RESULTS: Clinical signs and symptoms of chronic arthritis were aggravated in hTNFtg mice lacking functional VDR signalling. Moreover, synovial inflammation was clearly increased in VDR(-/-)hTNFtg mice as compared to hTNFtg mice and was associated with an increased macrophage influx in inflamed joints. In vitro, VDR-deficient monocytes were proinflammatory and hyper-responsive to TNF stimulation associated with prolonged mitogen-activated protein kinase activation and cytokine secretion. Also, VDR(-/-) monocytes showed enhanced potential to differentiate into bone resorbing osteoclasts in vitro. In line, VDR(-/-)hTNFtg mice had significantly increased cartilage damage and synovial bone erosions. CONCLUSIONS: VDR plays an important role in limiting the inflammatory phenotype in a mouse model of RA. Absent VDR signalling causes a proinflammatory monocyte phenotype associated with increased inflammation, cartilage damage and bone erosion.

Mammalian target of rapamycin signaling is crucial for joint destruction in experimental arthritis and is activated in osteoclasts from patients with rheumatoid arthritis.

OBJECTIVE: Activation of the mammalian target of rapamycin (mTOR) pathway is important for immune cell activation and bone metabolism. To date, the contribution of mTOR signaling to joint inflammation and structural bone and cartilage damage is unknown. The aim of this study was to investigate the potential of inhibiting mTOR as a treatment of inflammatory arthritis. METHODS: Human tumor necrosis factor-transgenic mice in which inflammatory arthritis was developing were treated with 2 different mTOR inhibitors, sirolimus or everolimus. The effects of treatment on clinical disease activity, inflammation, and localized joint and cartilage destruction were studied. In addition, the effects of mTOR inhibition on osteoclast survival and expression of key molecules of osteoclast function were analyzed in vitro. Moreover, synovial tissue from patients with rheumatoid arthritis (RA) was assessed for activation of the mTOR pathway. RESULTS: Inhibition of mTOR by sirolimus or everolimus reduced synovial osteoclast formation and protected against local bone erosions and cartilage loss. Clinical signs of arthritis improved after mTOR inhibition, and histologic evaluation showed a decrease in synovitis. In vitro, mTOR inhibition down-regulated the expression of digestive enzymes and led to osteoclast apoptosis. Moreover, mTOR signaling was shown to be active in the synovial membrane of patients with RA, particularly in synovial osteoclasts. CONCLUSION: Signaling through mTOR is an important link between synovitis and structural damage in inflammatory arthritis. Current pharmacologic inhibitors of mTOR could be effective in protecting joints against structural damage.

Antiinflammatory effects of tumor necrosis factor on hematopoietic cells in a murine model of erosive arthritis.

OBJECTIVE: To investigate the mechanisms leading to the influx of inflammatory hematopoietic cells into the synovial membrane and the role of tumor necrosis factor receptor I (TNFRI) and TNFRII in this process in an animal model of rheumatoid arthritis (RA). METHODS: We performed bone marrow transplantations in human TNF-transgenic mice using hematopoietic cells from wild-type, TNFRI(-/-), TNFRII(-/-), and TNFRI/II(-/-) mice as donors and assessed the severity of arthritis histologically. Generation of osteoclasts from the different genotypes was analyzed in vitro and in vivo. Apoptosis was analyzed using annexin V staining as well as TUNEL assays. RESULTS: Despite lacking responsiveness to TNF in their hematopoietic compartment, mice not only developed full-blown erosive arthritis but even showed increased joint destruction when compared with mice with a TNF-responsive hematopoietic compartment. We demonstrated different roles of the 2 different TNFRs in the regulation of these processes. The absence of TNFRI on hematopoietic cells did not affect joint inflammation but markedly attenuated erosive bone destruction via reduced synovial accumulation of osteoclast precursors. In contrast, the absence of TNFRII on hematopoietic cells increased joint inflammation as well as erosive bone destruction via the regulation of osteoclast precursor apoptosis. CONCLUSION: Our findings indicate that selective blockade of TNFRI, leaving the antiinflammatory effects of TNFRII unaltered instead of unselectively blocking TNF, might be advantageous in patients with RA.

The alpha-isoform of p38 MAPK specifically regulates arthritic bone loss.

Pharmacological inhibitors have provided evidence for the key role of p38 MAPK in osteoclast differentiation and in inflammation-induced bone loss. However, these inhibitors block more than one of the four p38 isoforms, usually p38alpha and p38beta, and sometimes also other kinases such as JNK3. We show in this study that p38alpha is the main p38 isoenzyme expressed in the osteoclast precursors and in the mature osteoclasts. p38alpha as well as its downstream substrates were phosphorylated in osteoclast progenitors stimulated by TNF-alpha. Using Mx-cre-mediated conditional gene inactivation we demonstrated that mice lacking p38alpha were protected against TNF-alpha-induced bone destruction at the site of inflammation as well as against TNF-alpha-mediated systemic bone loss. The bone protection was associated to decreased osteoclast numbers in vivo as well as a decreased IL-1beta expression in the inflamed tissue and in the isolated monocytes. The phenotype was cell autonomous because, similarly to p38alpha-deficient cells, knockdown of p38alpha in monocytes resulted in a decreased osteoclast differentiation in vitro. It was not caused by major changes in RANKL-mediated ERK or JNK activation but rather associated to an increased NF-kappaB activation caused by a decrease in IkappaBalpha recovery. Thus, our data show that developing specific inhibitors of the alpha-isoenzyme of p38 would be beneficial for the treatment of inflammation-induced bone destruction as observed in rheumatoid arthritis.

Interleukin 6 is not a crucial regulator in an animal model of tumour necrosis factor-mediated bilateral sacroiliitis.

OBJECTIVE: To evaluate the role of interleukin 6 (IL-6) in the pathogenesis of bilateral erosive sacroiliitis in human tumour necrosis factor transgenic (hTNFtg) mice, an animal model of ankylosing spondylitis (AS). METHODS: Histological sections of the sacroiliac joints from hTNFtg and IL-6(-/-)hTNFtg mice were evaluated, and wild type and IL-6(-/-)mice served as controls. mRNA levels of inflammation and tissue degradation related genes isolated from sacroiliac joints were also evaluated by quantitative PCR. RESULTS: Severe, erosive bilateral sacroiliitis in 14-week-old hTNFtg animals was accompanied by an upregulation of mRNAs related to tissue inflammation such as matrix metalloproteinase 3 (MMP3), MMP9 and MMP13 or osteoclast activation such as cathepsin K and tartrate-resistant acid phosphatase. In addition, IL-6 was increased in the sera and in the sacroiliac joints of hTNFtg animals. However, high expression of these marker genes in sacroiliac joints from IL-6(-/-)hTNFtg mice was also found. Moreover, absence of IL-6 in these animals did not alter bilateral, erosive sacroiliitis when compared to hTNFtg littermates. CONCLUSIONS: IL-6 is not a crucial regulator in an animal model of TNF-mediated bilateral, erosive sacroiliitis. This finding questions the potential of IL-6 blockade as a new treatment in patients with AS.

PI3Kgamma regulates cartilage damage in chronic inflammatory arthritis.

The gamma isoform of phosphoinositide 3-kinase (PI3Kgamma) has been viewed as restricted to leukocytes mediating the regulation of chemokine-induced migration and recruitment of neutrophils, monocytes, and macrophages. In line with the observation that PI3Kgamma-deficient mice display defects in adaptive immunity, inhibition of PI3Kgamma reduces synovial inflammation in the collagen-induced arthritis mouse model of inflammatory arthritis [rheumatoid arthritis (RA)], which has been attributed to reduced influx of inflammatory cells. Challenging the concept of leukocyte-restricted PI3Kgamma function, we report here a novel, nonredundant function of PI3Kgamma as an important regulator of fibroblast-induced cartilage destruction during chronic destructive arthritis. We show that in human tumor necrosis factor transgenic mice, the loss of PI3Kgamma leads to a milder inflammatory arthritis. Interestingly, PI3Kgamma deficiency does not alter the recruitment of inflammatory cells, but significantly reduces cartilage damage through reduced expression of matrix metalloproteinases in fibroblasts and chondrocytes. In vitro analyses demonstrate that the decreased invasiveness of fibroblasts is mediated by reduced phosphorylation of Akt and extracellular signal-regulated kinase. Using a PI3Kgamma specific inhibitor, these data are confirmed in human synovial fibroblasts from patients with RA who exhibit a disease-specific up-regulation of PI3Kgamma. Our data indicate that in addition to mediating the recruitment of inflammatory cells, PI3Kgamma is an important regulator of fibroblast-mediated joint destruction in RA and suggest that specific inhibitors of PI3Kgamma will interfere with the activation of RA synovial fibroblasts and reduce cartilage destruction in RA.

microRNA-146a controls age-related bone loss.

Bone loss is one of the consequences of aging, leading to diseases such as osteoporosis and increased susceptibility to fragility fractures and therefore considerable morbidity and mortality in humans. Here, we identify microRNA-146a (miR-146a) as an essential epigenetic switch controlling bone loss with age. Mice deficient in miR-146a show regular development of their skeleton. However, while WT mice start to lose bone with age, animals deficient in miR-146a continue to accrue bone throughout their life span. Increased bone mass is due to increased generation and activity of osteoblasts in miR-146a-deficient mice as a result of sustained activation of bone anabolic Wnt signaling during aging. Deregulation of the miR-146a target genes Wnt1 and Wnt5a parallels bone accrual and osteoblast generation, which is accompanied by reduced development of bone marrow adiposity. Furthermore, miR-146a-deficient mice are protected from ovariectomy-induced bone loss. In humans, the levels of miR-146a are increased in patients suffering fragility fractures in comparison with those who do not. These data identify miR-146a as a crucial epigenetic temporal regulator which essentially controls bone homeostasis during aging by regulating bone anabolic Wnt signaling. Therefore, miR-146a might be a powerful therapeutic target to prevent age-related bone dysfunctions such as the development of bone marrow adiposity and osteoporosis.

IRF1 is critical for the TNF-driven interferon response in rheumatoid fibroblast-like synoviocytes : JAKinibs suppress the interferon response in RA-FLSs.

Rheumatoid arthritis (RA) is an autoimmune disease characterized by persistent synovial inflammation. The major drivers of synovial inflammation are cytokines and chemokines. Among these molecules, TNF activates fibroblast-like synoviocytes (FLSs), which leads to the production of inflammatory mediators. Here, we show that TNF regulates the expression of the transcription factor interferon regulatory factor 1 (IRF1) in human FLSs as well as in a TNF transgenic arthritis mouse model. Transcriptomic analyses of IRF1-deficient, TNF-stimulated FLSs define the interferon (IFN) pathway as a major target of IRF1. IRF1 expression is associated with the expression of IFNß, which leads to the activation of the JAK-STAT pathway. Blocking the JAK-STAT pathway with the Janus kinase inhibitor (JAKinib) baricitinib or tofacitinib reduces the expression of IFN-regulated genes (IRGs) in TNF-activated FLSs. Therefore, we conclude that TNF induces a distinct inflammatory cascade, in which IRGs are key elements, in FLSs. The IFN-signature might be a promising biomarker for the efficient and personalized use of new treatment strategies for RA, such as JAKinibs.