Short-term and long-term outcomes of patients with anti-melanoma differentiation-associated gene 5 antibody-positive dermatomyositis.

OBJECTIVES: Anti-melanoma differentiation-associated gene 5 (MDA5) antibody-positive dermatomyositis (MDA5-DM) is a subtype of dermatomyositis characterized by frequent interstitial lung disease and reduced muscle involvement. This study aimed to determine the short-term and long-term outcomes of patients with MDA5-DM. METHODS: Information on baseline characteristics, treatments, and short-term and long-term outcomes of patients with MDA5-DM including survival, relapse, and the titer of anti-MDA5 antibody, was retrospectively collected. Descriptive statistics regarding clinical outcomes were calculated, and a comparison of clinical parameters between patients with and without relapse was performed. The short-term survival according to the use of Janus kinase inhibitors (JAKi) was also assessed. RESULTS: A total of 154 patients with MDA5-DM were included in the study. Forty patients (25.9%) died during the remission induction phase, with respiratory failure being the most common cause of mortality. Among the 114 patients who survived the remission induction phase, the 5-year cumulative survival and relapse-free survival rates were 96.8% and 77.4%, respectively, and 7.9% of patients achieved complete drug-free remission. Fifty-four patients achieved normalization of anti-MDA5 antibody titers and only two of them relapsed after normalization. In the severe patients, the 6-month survival rate became significantly higher after the emergence of the JAKi treatment compared with before its existence (p= 0.03). CONCLUSIONS: Although relapse often occurs, the long-term survival of MDA5-DM patients who survived the remission induction phase is generally favorable. The status of the anti-MDA5 antibody is associated with relapse. JAKi may improve the survival of refractory patients with severe MDA5-DM.

Prevalence and risk factors of osteonecrosis of the femoral head in patients with ANCA-associated vasculitis: a multicentre cohort study.

OBJECTIVE: We aimed to determine the prevalence and risk factors for osteonecrosis of the femoral head (ONFH) in a multicentre cohort of patients with antineutrophil cytoplasmic antibody-associated vasculitis (AAV). METHODS: One hundred and eighty-six AAV patients who underwent radiographs and MRI screening of bilateral hip joints at more than 6 months after initial remission induction therapy (RIT) were retrospectively assessed for the presence of ONFH. RESULTS: Among 186 AAV patients, 33 (18%) were diagnosed with ONFH. Among the patients with ONFH, 55% were asymptomatic and 64% had bilateral ONFH. Seventy-six per cent of ONFH joints were in precollapse stages (stage ≤2), whereas 24% of ONFH joints were in collapse stages (stage ≥3). Moreover, 56% of the precollapse stage joints were already at risk of future collapse (type ≥C-1). Even in asymptomatic ONFH patients, 39% of the precollapse stage joints were type ≥C-1. Prednisolone dose of ≥20 mg/day on day 90 of RIT was an independent risk factor for ONFH in AAV patients (OR 1.072, 95% CI 1.017 to 1.130, p=0.009). Rituximab use was a significant beneficial factor against ONFH (p=0.019), but the multivariate analysis rejected its significance (p=0.257). CONCLUSION: Eighteen per cent of AAV patients developed ONFH, and two-thirds of the ONFH joints were already in collapse stages or at risk of future collapse. Prednisolone dose of ≥20 mg/day on day 90 of RIT was an independent risk factor for ONFH. A rapid reduction of glucocorticoids in RIT and early detection of precollapse ONFH by MRI may decrease and intervene ONFH development in AAV patients.

A20 (Tnfaip3) expressed in CD4+ T cells suppresses Th2 cell-mediated allergic airway inflammation in mice.

A20 (Tnfaip3), a ubiquitin-editing enzyme, inhibits NF-κB signaling pathways in response to pro-inflammatory cytokines. Previous studies have proved the anti-inflammatory roles of A20 in various cell types, including T cells, B cells, dendritic cells, and intestinal epithelial cells. Moreover, recent studies have shown that A20 expressed in lung epithelial cells is required for LPS-induced protection from asthma. In humans, a single-nucleotide polymorphism in TNFAIP3 is associated with asthma risk. However, the role of A20 expressed in T cells in asthmatic responses has not been elucidated. We addressed this point by generating mice lacking A20 expression in T cells (CD4-CreA20 fl/fl mice). We found that house dust mite (HDM)-induced allergic airway inflammation, mucus production, airway hyperresponsiveness, and Th2 cytokine production were significantly exacerbated in CD4-CreA20 fl/fl mice compared with those in control A20 fl/fl mice. In vitro differentiation of Th2 cells but not of Th1 cells or Th17 cells was enhanced in CD4+ T cells by the absence of A20. Consistently, enforced expression of A20 inhibited the differentiation of Th2 cells but not of Th1 cells or Th17 cells. Notably, the expression of GATA3 was significantly enhanced in A20-deficient CD4+ T cells, and the enhanced GATA3 expression was partly canceled by IL-2 neutralization. These results suggest that A20 functions as a stabilizing factor maintaining GATA3 levels during the induction of Th2 cells to prevent excessive Th2 cell differentiation.

T-bet and STAT6 Coordinately Suppress the Development of IL-9-Mediated Atopic Dermatitis-Like Skin Inflammation in Mice.

T-bet and signal transducer and activator of transcription (STAT) 6 are critical factors for helper T-cell differentiation in humans and mice. Additionally, polymorphisms in TBX21 (T-bet) and STAT6 are associated with the susceptibility of allergic diseases. However, precise mechanisms of the reciprocal regulation between T-bet and STAT6 in allergy remain unclear. To determine the reciprocal regulation in vivo, we investigated the phenotype of T-bet/STAT6 double-deficient (T-bet-/- STAT6-/-) mice. Unexpectedly, T-bet-/- STAT6-/- mice but not T-bet-/- mice or STAT6-/- mice spontaneously developed severe dermatitis. Not only eosinophils and mast cells but also CD4+ T cells infiltrated into the skin of T-bet-/- STAT6-/- mice. Adoptive transfer of CD4+ T cells of T-bet-/- STAT6-/- mice into severe combined immunodeficient mice induced the accumulation of eosinophils and mast cells in the skin, whereas depletion of CD4+ T cells ameliorated the dermatitis in T-bet-/- STAT6-/- mice. Comprehensive transcriptome analyses revealed that IL-9 expression was enhanced in T-bet-/- STAT6-/- CD4+ T cells. Indeed, IL-9 neutralization ameliorated the dermatitis in T-bet-/- STAT6-/- mice. T-bet-/- STAT6-/- CD4+ T cells expressed functional thymic stromal lymphopoietin receptors and produced large amounts of IL-9 on thymic stromal lymphopoietin stimulation. These results indicate that T-bet and STAT6 coordinately suppress atopic dermatitis-like skin inflammation, possibly by inhibiting thymic stromal lymphopoietin-dependent IL-9 production in CD4+ T cells.

Experience of musculoskeletal ultrasound scanning improves physicians' physical examination skills in assessment of synovitis.

OBJECTIVE: Musculoskeletal ultrasound (US) is more sensitive than physical examination in detecting synovitis and helps physicians to understand its pathophysiology. In this study, we aimed to determine if the experience in musculoskeletal US scanning is independently associated with improved physical examination skills to detect synovitis. METHOD: Seventy patients with rheumatoid arthritis and twenty-three physicians were enrolled. Patients were first assessed by multiple physicians with a range of clinical/sonographic experience for the swelling of the wrist, metacarpophalangeal and proximal interphalangeal (PIP) joints and next underwent US assessment performed by another physician experienced in musculoskeletal US. We then calculated the positive/negative predictive values (PPV/NPV) of joint swelling to identify US-detected synovial hypertrophy. Finally, the factors independently associated with the accuracy of clinical assessment were identified by using multivariate analyses. RESULTS: One thousand five hundred forty joints were assessed 6116 times in total for swelling. Overall, PPV and NPV of joint swelling were 51.7% and 88.3%, respectively. Multivariate analyses identified wrist joint, tenderness, male and greater patients' age as the factors significantly associated with higher PPV. In addition, there was a trend that longer experience in rheumatology clinical practice was associated with higher PPV (p = 0.058). On the other hand, longer experience in musculoskeletal US, PIP joint and positive rheumatoid factor were identified as the significant factors for higher NPV, while wrist joint, tenderness, presence of osteophyte and obesity as those for lower NPV. CONCLUSION: Our data suggest that the experience in musculoskeletal US improves physical examination skills particularly to avoid overestimation.Key Points• Physicians with longer US experience are less likely to overestimate synovitis by physical examination.• Musculoskeletal US is a useful tool for rheumatologists to improve their physical examination skill.• Presence of osteophytes, joint tenderness and obesity influence the accuracy of physical examination of joints.

Fucosyltransferase 2 induces lung epithelial fucosylation and exacerbates house dust mite-induced airway inflammation.

BACKGROUND: One of the pathognomonic features of asthma is epithelial hyperproduction of mucus, which is composed of a series of glycoproteins; however, it remains unclear how glycosylation is induced in lung epithelial cells from asthmatic patients and how glycan residues play a role in the pathogenesis of asthma. OBJECTIVE: The objective of this study was to explore comprehensive epithelial glycosylation status induced by allergic inflammation and reveal its possible role in the pathogenesis of asthma. METHODS: We evaluated the glycosylation status of lung epithelium using a lectin microarray. We next searched for molecular mechanisms underlying epithelial glycosylation. We also examined whether epithelial glycosylation is involved in induction of allergic inflammation. RESULTS: On allergen inhalation, lung epithelial cells were heavily α(1,2)fucosylated by fucosyltransferase 2 (Fut2), which was induced by the IL-13-signal transducer and activator of transcription 6 pathway. Importantly, Fut2-deficient (Fut2-/-) mice, which lacked lung epithelial fucosylation, showed significantly attenuated eosinophilic inflammation and airway hyperresponsiveness in house dust mite (HDM)-induced asthma models. Proteome analyses and immunostaining of the HDM-challenged lung identified that complement C3 was accumulated in fucosylated areas. Indeed, Fut2-/- mice showed significantly reduced levels of C3a and impaired accumulation of C3a receptor-expressing monocyte-derived dendritic cells in the lung on HDM challenge. CONCLUSION: Fut2 induces epithelial fucosylation and exacerbates airway inflammation in asthmatic patients in part through C3a production and monocyte-derived dendritic cell accumulation in the lung.

Sox12 promotes T reg differentiation in the periphery during colitis.

Peripherally induced regulatory T (pT reg) cells play indispensable roles in regulating gut inflammation; however, the mechanism underling the differentiation of pT reg cells under inflammatory conditions remains largely unknown. Here, we show that the expression of Sox12, a member of SoxC family, is significantly induced in T reg cells in colitic mice. We also show that TCR-NFAT signaling induces Sox12 expression in CD4+ T cells. Although Sox12 is not required for the development of thymus-derived T reg (tT reg) cells, Sox12 is involved in the development of pT reg cells under inflammatory conditions in an adoptive transfer colitis model. Moreover, we found that enforced expression of Sox12 is sufficient to promote Foxp3 expression in CD4+ T cells even in the absence of TGF-ß or IL-2 and that Sox12 binds to Foxp3 promoter and drives its transcription. These results suggest that TCR-NFAT signaling induces the development of pT reg cells in colitic mice partly through Sox12 induction.

IL-22 induces Reg3γ and inhibits allergic inflammation in house dust mite-induced asthma models.

Previous studies have shown that IL-22, one of the Th17 cell-related cytokines, plays multiple roles in regulating allergic airway inflammation caused by antigen-specific Th2 cells; however, the underlying mechanism remains unclear. Here, we show that allergic airway inflammation and Th2 and Th17 cytokine production upon intratracheal administration of house dust mite (HDM) extract, a representative allergen, were exacerbated in IL-22-deficient mice. We also found that IL-22 induces Reg3γ production from lung epithelial cells through STAT3 activation and that neutralization of Reg3γ significantly exacerbates HDM-induced eosinophilic airway inflammation and Th2 cytokine induction. Moreover, exostatin-like 3 (EXTL3), a functional Reg3γ binding protein, is expressed in lung epithelial cells, and intratracheal administration of recombinant Reg3γ suppresses HDM-induced thymic stromal lymphopoietin and IL-33 expression and accumulation of type 2 innate lymphoid cells in the lung. Collectively, these results suggest that IL-22 induces Reg3γ production from lung epithelial cells and inhibits the development of HDM-induced allergic airway inflammation, possibly by inhibiting cytokine production from lung epithelial cells.

Allergic airway inflammation: key players beyond the Th2 cell pathway.

Allergic asthma is characterized by eosinophilic airway inflammation, mucus hyperproduction, and airway hyperreactivity, causing reversible airway obstruction. Accumulating evidence indicates that antigen-specific Th2 cells and their cytokines such as IL-4, IL-5, and IL-13 orchestrate these pathognomonic features of asthma. However, over the past decade, the understanding of asthma pathogenesis has made a significant shift from a Th2 cell-dependent, IgE-mediated disease to a more complicated heterogeneous disease. Recent studies clearly show that not only Th2 cytokines but also other T cell-related cytokines such as IL-17A and IL-22 as well as epithelial cell cytokines such as IL-25, IL-33, and thymic stromal lymphopoietin (TSLP) are involved in the pathogenesis of asthma. In this review, we focus on the roles of these players beyond Th2 pathways in the pathogenesis of asthma.

T-bet inhibits innate lymphoid cell-mediated eosinophilic airway inflammation by suppressing IL-9 production.

BACKGROUND: Innate lymphoid cells (ILCs) are emerging subsets of immune cells that produce large amounts of cytokines upon cytokine and/or alarmin stimulation. Recent studies have shown that T-bet plays pivotal roles in the development of ILC3s and type 1 ILCs; however, the roles of T-bet in lung type 2 innate lymphoid cells (ILC2s) remain unknown. OBJECTIVE: We sought to determine the role of T-bet in ILC2-mediated airway inflammation. METHODS: The expression of T-bet in lung ILCs (defined as Thy1.2+ Lin- cells) was examined. The roles of T-bet in the development of lung ILC2s and airway inflammation induced by IL-33 administration were examined by using T-bet-deficient (T-bet-/-) mice. Gene expression profiles of T-bet-/- lung ILCs were analyzed by RNA sequencing. RESULTS: T-bet was expressed in lung ILC2s (defined as Thy1.2+ Lin- cells expressing ST2 or CD25) and IFN-γ enhanced its expression. Although the development of lung ILC2s at steady-state conditions was normal in T-bet-/- mice, IL-33-induced accumulation of lung ILC2s and eosinophilic airway inflammation were exacerbated in T-bet-/- mice. The exacerbated accumulation of ILC2s and eosinophilic airway inflammation by the absence of T-bet were evident even in a RAG2-/- background, suggesting that T-bet expressed in non-T/non-B population is involved in the suppression of IL-33-induced eosinophilic airway inflammation. Transcriptome analysis revealed that IL-9 expression in IL-33-stimulated lung ILCs was upregulated in T-bet-/- mice compared with that in wild-type mice. Importantly, neutralization of IL-9 markedly attenuated IL-33-induced accumulation of lung ILC2s and eosinophilic inflammation in T-bet-/- mice. CONCLUSIONS: T-bet suppresses IL-9 production from lung ILC2s and thereby inhibits IL-33-induced eosinophilic airway inflammation.

Dectin-1 Plays an Important Role in House Dust Mite-Induced Allergic Airway Inflammation through the Activation of CD11b+ Dendritic Cells.

It is well known that sensitization against fungi is closely associated with severity of asthma. Dectin-1 (gene symbol Clec7a), a C-type lectin receptor, recognizes the fungal cell wall component ß-glucan, as well as some component(s) in house dust mite (HDM) extract. However, the roles of Dectin-1 in HDM-induced allergic airway inflammation remain unclear. In this study, we used Dectin-1-deficient (Clec7a-/-) mice to examine whether Dectin-1 is involved in HDM-induced allergic airway inflammation. We found that HDM-induced eosinophil and neutrophil recruitment into the airways was significantly attenuated in Clec7a-/- mice compared with that in wild-type mice. In addition, HDM-induced IL-5, IL-13, and IL-17 production from mediastinum lymph node cells was reduced in HDM-sensitized Clec7a-/- mice. Dectin-1 was expressed on CD11b+ dendritic cells (DCs), an essential DC subset for the development of allergic inflammation, but not on CD103+ DCs, plasmacytoid DCs, or lung epithelial cells. Transcriptome analysis revealed that the expression of chemokine/chemokine receptors, including CCR7, which is indispensable for DC migration to draining lymph nodes, was decreased in Clec7a-/- DCs. In accordance with these results, the number of HDM-labeled CD11b+ DCs in mediastinum lymph nodes was significantly reduced in Clec7a-/- mice compared with wild-type mice. Taken together, these results suggest that Dectin-1 expressed on CD11b+ DCs senses some molecule(s) in HDM extract and plays a critical role in the induction of HDM-induced allergic airway inflammation by inducing the expression of chemokine/chemokine receptors in DCs.

Helios Enhances Treg Cell Function in Cooperation With FoxP3.

OBJECTIVE: Helios+FoxP3+CD4+ (Helios+) Treg cells are believed to be involved in the regulation of various autoimmune diseases; however, the regulatory mechanisms underlying the development of Helios+ Treg cells remain uncertain. This study was undertaken to elucidate the regulatory mechanisms of Helios expression in CD4+ T cells and its roles in transforming growth factor ß (TGFß)-induced Treg cell function. METHODS: We examined the expression of Helios in CD4+ T cells in patients with rheumatoid arthritis by DNA microarray analysis before and after treatment with biologic agents. We also examined the effect of interleukin-6 (IL-6) and TGFß on Helios expression in CD4+ T cells in humans and mice. The effect of forced expression of Helios on murine induced Treg cell function was also examined. The role of FoxP3 in the induction and function of Helios was assessed by using CD4+ T cells from FoxP3-deficient scurfy mice. RESULTS: Tocilizumab, but not tumor necrosis factor (TNF) inhibitors or abatacept, increased Helios expression in CD4+ T cells in patients with a good response. IL-6 inhibited the TGFß-induced development of Helios+ induced Treg cells in both humans and mice. Both cell-intrinsic FoxP3 expression and TGFß signaling were required for Helios induction in murine induced Treg cells. The forced expression of Helios enhanced the expression of various Treg cell-related molecules and the suppressive function in murine induced Treg cells. Helios-mediated enhancement of the suppressive function of induced Treg cells was obvious in FoxP3-sufficient CD4+ T cells but not in FoxP3-deficient CD4+ T cells. CONCLUSION: Our findings indicate that Helios enhances induced Treg cell function in cooperation with FoxP3.

Sox5 and c-Maf cooperatively induce Th17 cell differentiation via RORγt induction as downstream targets of Stat3.

Stat3 signaling is essential for the induction of RORγt and subsequent Th17 cell differentiation. However, the downstream targets of Stat3 for RORγt expression remain largely unknown. We show here that a novel isoform of Sox5, named Sox5t, is induced in Th17 cells in a Stat3-dependent manner. In vivo, T cell-specific Sox5-deficient mice exhibit impaired Th17 cell differentiation and are resistant to experimental autoimmune encephalomyelitis and delayed-type hypersensitivity. Retrovirus-mediated induction of Sox5 together with c-Maf induces Th17 cell differentiation even in Stat3-deficient CD4(+) T cells but not in RORγt-deficient CD4(+) T cells, indicating that Sox5 and c-Maf induce Th17 cell differentiation as downstream effectors of Stat3 and as upstream inducers of RORγt. Moreover, Sox5 physically associates with c-Maf via the HMG domain of Sox5 and DNA-binding domain of c-Maf, and Sox5 together with c-Maf directly activates the promoter of RORγt in CD4(+) T cells. Collectively, our results suggest that Sox5 and c-Maf cooperatively induce Th17 cell differentiation via the induction of RORγt as downstream targets of Stat3.

Dectin-2 promotes house dust mite-induced T helper type 2 and type 17 cell differentiation and allergic airway inflammation in mice.

The fact that sensitization against fungi is closely related to the severity of asthma suggests that immune systems recognizing fungi are involved in the pathogenesis of severe asthma. Recently, Dectin-2 (gene symbol, Clec4n), a C-type lectin receptor, has been shown to function as not only a major pattern-recognition receptor for fungi, but also a receptor for some components of house dust mite (HDM) extract, a major allergen for asthma. However, the roles of Dectin-2 in the induction of HDM-induced allergic airway inflammation remain largely unknown. Our objective was to determine the roles of Dectin-2 in HDM-induced allergic airway inflammation. We examined the roles of Dectin-2 in the induction of HDM-induced T helper (Th) 2 and Th17 cell differentiation and subsequent allergic airway inflammation by using Clec4n-deficient (Clec4n(-/-)) mice. We also investigated Dectin-2-expressing cells in the lung and their roles in HDM-induced allergic airway inflammation. Clec4n(-/-) mice showed significantly attenuated HDM-induced allergic airway inflammation and decreased Th2 and Th17 cell differentiation. Dectin-2 mRNA, together with Dectin-3 and Fc receptor-γ mRNAs, was expressed in CD11b(+) dendritic cells (DCs), but not in CD4(+) T cells or epithelial cells in the lung. CD11b(+) DCs isolated from Clec4n(-/-) mice expressed lower amounts of proinflammatory cytokines and costimulatory molecules, which could lead to Th2 and Th17 cell differentiation than those from wild-type mice. HDM-pulsed Clec4n(-/-) DCs were less efficient for the induction of allergic airway inflammation than HDM-pulsed wild-type DCs. In conclusion, Dectin-2 expressed on CD11b(+) DCs promotes HDM-induced Th2 and Th17 cell differentiation and allergic airway inflammation.