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Este estudo visou obter o diagnóstico dos conflitos de uso do solo em áreas de preservação permanentes da microbacia Água da Ema, no município de Rolândia - Paraná, partir da utilização de imagens de satélite. Para a elaboração dos mapas foi utilizado o software AutoCad Map 3D Raster Design. A base cartográfica utilizada foi a carta planialtimétrica em formato digital, Folha de Arapongas, em escala 1:50.000, e a imagem de satélite do Google Earth Pro 2018. Foram identificadas nove classes de uso do solo, sendo culturas anuais a classe de uso predominante, ocupando 892,72 ha (35,32%) da área total da microbacia. A simulação das áreas de preservação permanente mostra que a microbacia deveria ter 94,04 ha de mata ciliar ao redor das nascentes e ao longo dos canais da rede de drenagem. Foram identificadas quatro classes de uso conflitantes em APP, destacando-se culturas anuais com 2,19ha e pasto sujo com 2,13ha. O software AutoCad mostrou-se como uma alternativa de ferramenta para a determinação e mapeamento do uso e conflito de uso do solo em áreas de preservação permanente, servindo de base para estudos futuros visando a conservação e preservação dos recursos hídricos.

Oral immunotherapy (OIT) is an effective approach to controlling food allergy. Although the detailed molecular and cellular mechanisms of OIT are unknown currently, they must be understood to advance the treatment of allergic diseases in general. To elucidate the mechanisms of OIT, especially during the immunological transition from desensitization to allergy regulation, we generated a clinical OIT murine model and used it to examine immunological events of OIT. We found that in mice that completed OIT successfully, desensitized mast cells (MCs) showed functionally beneficial alterations, such as increased induction of regulatory cytokines and enhanced expansion of regulatory T cells. Importantly, these regulatory-T-cell-mediated inhibitions of allergic responses were dramatically decreased in mice lacking OIT-induced desensitized MC. Collectively, these findings show that the desensitization process modulates the activation of MCs, leading directly to enhanced induction of regulatory-T-cell expansion and promotion of clinical allergic unresponsiveness. Our results suggest that efficiently inducing regulatory MCs is a novel strategy for the treatment of allergic disease.

Increases in adhesive and invasive commensal bacteria, such as Escherichia coli, and subsequent disruption of the epithelial barrier is implicated in the pathogenesis of inflammatory bowel disease (IBD). However, the protective systems against such barrier disruption are not fully understood. Here, we show that secretion of luminal glycoprotein 2 (GP2) from pancreatic acinar cells is induced in a TNF–dependent manner in mice with chemically induced colitis. Fecal GP2 concentration is also increased in Crohn’s diease patients. Furthermore, pancreas-specific GP2-deficient colitis mice have more severe intestinal inflammation and a larger mucosal E. coli population than do intact mice, indicating that digestive-tract GP2 binds commensal E. coli, preventing epithelial attachment and penetration. Thus, the pancreas–intestinal barrier axis and pancreatic GP2 are important as a first line of defense against adhesive and invasive commensal bacteria during intestinal inflammation., Glycoprotein-2 (GP-2) can protect the intestinal epithelial barrier from bacteria and is associated with protection against Crohn’s disease. Here, the authors show pancreatic GP-2 is the source of the intestine’s luminal GP-2 that binds bacteria and prevents them from attaching to the epithelium, also limiting pathology in a DSS colitis mouse model.

Mesenchymal cells in the crypt play indispensable roles in the maintenance of intestinal epithelial homeostasis through their contribution to the preservation of stem cells. However, the acquisition properties of the production of stem cell niche factors by the mesenchymal cells have not been well elucidated, due to technical limitations regarding the isolation and subsequent molecular and cellular analyses of cryptal mesenchymal cells. To evaluate the function of mesenchymal cells located at the large intestinal crypt, we established a novel method through which cells are harvested according to the histologic layers of mouse colon, and we compared cellular properties between microenvironmental niches, the luminal mucosa and crypts. The gene expression pattern in the cryptal mesenchymal cells showed that receptors of the hormone/cytokine leptin were highly expressed, and we found a decrease in Wnt2b expression under conditions of leptin receptor deficiency, which also induced a delay in cryptal epithelial proliferation. Our novel stratified layer isolation strategies thus revealed new microenvironmental characteristics of colonic mesenchymal cells, including the intrinsic involvement of leptin in the control of mucosal homeostasis.

This paper presents a remote radioactive seed-loading device for permanent brachytherapy of oral cancer with Au-198 grains to reduce the exposure dose to the doctors. Permanent brachytherapy is a treatment which involve implanting of radioactive Au-198 grains, and requires neither long-term needle insertion nor diet-restriction. This is its advantage over temporary brachytherapy. However, the doctors that perform this therapy are exposed to radiation during the treatment procedures. This hinders permanent brachytherapy from being commonly used. The developed device shoots an Au grain from a distant position using air pressure and loads it onto the applicator. This can reduce the exposure dose of the doctors compared with that of the traditional procedure because the doctors are able to stay as far away from the radioactive seeds as possible. We conducted experiments of motion simulation of the proposed and traditional treatment procedures, and compared the exposed doses. The device can reduce the exposed dose by 62% on average.

Objective The aim of this prospective multicenter study was to identify biomarkers that can be used to predict therapeutic responses to tocilizumab in patients with rheumatoid arthritis (RA). Methods We recruited patients with RA who were treated with tocilizumab for the first time, and determined therapeutic responses at 6 months. In the training cohort (n = 40), gene expression in peripheral blood mononuclear cells (PBMCs) at baseline was analyzed using genome-wide DNA microarray, with 41,000 probes derived from 19,416 genes. In the validation cohort (n = 20), expression levels of the candidate genes in PBMCs at baseline were determined using real-time quantitative polymerase chain reaction (qPCR) analysis. Results We identified 68 DNA microarray probes that showed significant differences in signal intensity between nonresponders and responders in the training cohort. Nineteen putative genes were selected, and a significant correlation between the DNA microarray signal intensity and the qPCR relative expression was confirmed in 15 genes. In the validation cohort, a significant difference in relative expression between nonresponders and responders was reproduced for 3 type I interferon response genes (IFI6, MX2, and OASL) and MT1G. Receiver operating characteristic curve analysis of models incorporating these genes showed that the maximum area under the curve was 0.947 in predicting a moderate or good response to tocilizumab in the validation cohort. Conclusion Using genome-wide DNA microarray analyses, we identified candidate biomarkers that can be used to predict therapeutic responses to tocilizumab in patients with RA. These findings suggest that type I interferon signaling and metallothioneins are involved in the pathophysiology of RA.

Objective The proinflammatory cytokines tumor necrosis factor α and interleukin-6 (IL-6) and the Th17 cell cytokine IL-17A are implicated in the pathogenesis of rheumatoid arthritis (RA), and the blockade of these cytokines by biologic agents provides clinical benefits for RA patients. We undertook this study to clarify the mechanisms underlying the efficacy of IL-6 blockade in RA and to find a novel target for treatment of RA. Methods We examined gene expression profiles of CD4+ T cells by DNA microarray analysis before and after treatment with an anti–IL-6 receptor antibody, tocilizumab (TCZ), in RA patients who exhibited good clinical responses to the treatment. Using murine CD4+ T cells, we then examined the roles of a newly identified molecule whose expression was significantly reduced in CD4+ T cells by TCZ therapy. We also examined the effect of the forced expression of the molecule on retinoic acid receptor–related orphan nuclear receptor γt (RORγt)–induced IL-17A production in CD4+ T cells and on RORγt-induced IL-17A promoter activation. Results We identified AT-rich–interactive domain– containing protein 5A (ARID-5A) as a new molecule down-regulated by IL-6 blockade in the form of TCZ therapy. IL-6 induced the expression of ARID-5A in CD4+ T cells during Th17 cell differentiation by a STAT-3–dependent mechanism, whereas IL-6–induced ARID-5A expression was not affected by the absence of RORγt, a lineage-specifying transcription factor of Th17 cells. Furthermore, ARID-5A physically associated with RORγt through its N-terminal region and inhibited RORγt-induced Th17 cell differentiation. Conclusion ARID-5A is a lineage-specific attenuator of Th17 cell differentiation and may be involved in the pathogenesis of RA.

Although innate immune responses are necessary for the initiation of acquired immune responses and the subsequent successful elimination of pathogens, excessive responses occasionally result in lethal endotoxic shock accompanied by a cytokine storm. B and T lymphocyte attenuator (BTLA), a coinhibitory receptor with similarities to cytotoxic T-lymphocyte antigen (CTLA)-4 and programmed death (PD)-1, is expressed in not only B and T cells but also dendritic cells (DCs) and macrophages (Mϕs). Recently, several studies have reported that BTLA-deficient (BTLA −/− ) mice show enhanced pathogen clearance compared with WT mice in early phase of infections. However, the roles of BTLA expressed on innate cells in overwhelming and uncontrolled immune responses remain unclear. Here, we found that BTLA −/− mice were highly susceptible to LPS-induced endotoxic shock. LPS-induced TNF-α and IL-12 production in DCs and Mϕs was significantly enhanced in BTLA −/− mice. BTLA −/− DCs also produced high levels of TNF-α on stimulation with Pam3CSK4 but not poly(I:C) or CpG, suggesting that BTLA functions as an inhibitory molecule on Toll-like receptor signaling at cell surface but not endosome. Moreover, BTLA −/− DCs showed enhanced MyD88- and toll/IL-1R domain-containing adaptor inducing IFN (TRIF)-dependent signaling on LPS stimulation, which is associated with impaired accumulation of Src homology 2-containing protein tyrosine phosphatase in lipid rafts. Finally, we found that an agonistic anti-BTLA antibody rescued mice from LPS-induced endotoxic shock, even if the antibody was given to mice that had developed a sign of endotoxic shock. These results suggest that BTLA directly inhibits LPS responses in DCs and Mϕs and that agonistic agents for BTLA might have therapeutic potential for LPS-induced endotoxic shock.

The brain capillary endothelial cell (BCEC) is a major functional component of the blood-brain barrier and is an underlying factor in the pathophysiology of various diseases, including brain ischemia, multiple sclerosis, and neurodegenerative disorders. We examined gene silencing in BCECs by using endogenous lipoprotein to introduce short-interfering RNA (siRNA) in vivo. A cholesterol-conjugated 21/23-mer siRNA targeting organic anion transporter 3 (OAT3) mRNA (Chol-siOAT3) was intravenously injected into mice after its incorporation into extracted endogenous lipoproteins. Chol-siOAT3 was not delivered to neurons or glia, but was successfully delivered into BCECs and resulted in a significant reduction of OAT3 mRNA levels when injected after its incorporation into high-density lipoprotein (HDL). Efficient delivery was not achieved, however, when Chol-siOAT3 was injected without any lipoproteins, or after its incorporation into low-density lipoprotein (LDL). Investigations in apolipoprotein E (ApoE)-deficient and LDL receptor (LDLR)-deficient mice revealed that the uptake of HDL-containing Chol-siOAT3 was mainly mediated by ApoE and LDLR in mice. These findings indicate that siRNA can be delivered into BCECs in vivo by using endogenous lipoprotein, which could make this strategy useful as a new gene silencing therapy for diseases involving BCECs.

Species in the wood-feeding genus Salganea within the cockroach subfamily Panesthiinae (Blaberidae) typically live in biparental families; their first instars suffer high mortality when removed from adults, and in at least one species, adults are known to feed neonates on oral liquids. In the closely related gregarious wood-feeding genus Panesthia , no parental interaction with offspring is known. We compared the external morphology of first instars of these two genera and found that eye development and cuticular pigmentation at hatching are correlated with social structure. First instars of Panesthia have a dark cuticle and well-developed eyes. First instars of Salganea have a pale, transparent cuticle, and eyes significantly smaller than those of Panesthia relative to adult eye size. The body size of the first-instar of Salganea esakii is significantly smaller than that of Panesthia angustipennis spadica , relative to adult body size (24.0 and 27.4% of adult head-capsule width, respectively), but a more general survey suggests that, overall, neonate size may be similar in the two genera. We compared the first instars of these two taxa of Panesthiinae to those in the biparental, wood-feeding cockroach Cryptocercus (Cryptocercidae) and discuss how decreased investment in both integumentary and ocular development in subsocial cockroaches parallels that seen in altricial vertebrates.

SUMMARY Atopic disorders are caused by disregulated activation of T helper 2 (Th2) cells that produce IL-4 and IL-5. Because the presence of IL-4 potently augments the differentiation of naive T cells into Th2 cells, it is important to seek the cell population which provides IL-4 for naive T cells. Recently, a unique subpopulation of T cells, natural killer (NK) T cells, has been shown to produce a large amount of IL-4 upon activation, suggesting their regulatory role in initiation of Th2 cell differentiation. To determine whether NK T cells play a regulatory role in human Th2 cell-mediated atopic diseases, we analysed the frequency of invariant Vα24JαQ CD4−CD8− double-negative (DN) T cells, human NK T cells, in patients with atopic asthma and atopic dermatitis. We also studied cytokine production from Vα24+ Vβ11+ DN T cells, which comprise most of Vα24JαQ DN T cells. We found that the invariant Vα24JαQ DN T cells were greatly diminished in patients with asthma and atopic dermatitis. On the other hand, there was no significant difference in Vα24+ CD4+ T cells possessing invariant Vα24JαQ TCR between healthy subjects and atopic patients. We also found that Vα24+ Vβ11+ DN T cells from healthy subjects predominantly produced interferon-gamma (IFN-γ) but not IL-4 upon activation. These results suggest that NK T cells may not be essential for human atopic disease and that the disappearance of NK T cells, most of which produce IFN-γ, may be involved in the pathogenesis of atopic diseases.

The aim of this prospective multicenter study was to identify biomarkers that can be used to predict therapeutic responses to tocilizumab in patients with rheumatoid arthritis (RA).We recruited patients with RA who were treated with tocilizumab for the first time, and determined therapeutic responses at 6 months. In the training cohort (n = 40), gene expression in peripheral blood mononuclear cells (PBMCs) at baseline was analyzed using genome-wide DNA microarray, with 41,000 probes derived from 19,416 genes. In the validation cohort (n = 20), expression levels of the candidate genes in PBMCs at baseline were determined using real-time quantitative polymerase chain reaction (qPCR) analysis.We identified 68 DNA microarray probes that showed significant differences in signal intensity between nonresponders and responders in the training cohort. Nineteen putative genes were selected, and a significant correlation between the DNA microarray signal intensity and the qPCR relative expression was confirmed in 15 genes. In the validation cohort, a significant difference in relative expression between nonresponders and responders was reproduced for 3 type I interferon response genes (IFI6, MX2, and OASL) and MT1G. Receiver operating characteristic curve analysis of models incorporating these genes showed that the maximum area under the curve was 0.947 in predicting a moderate or good response to tocilizumab in the validation cohort.Using genome-wide DNA microarray analyses, we identified candidate biomarkers that can be used to predict therapeutic responses to tocilizumab in patients with RA. These findings suggest that type I interferon signaling and metallothioneins are involved in the pathophysiology of RA.

The proinflammatory cytokines tumor necrosis factor α and interleukin-6 (IL-6) and the Th17 cell cytokine IL-17A are implicated in the pathogenesis of rheumatoid arthritis (RA), and the blockade of these cytokines by biologic agents provides clinical benefits for RA patients. We undertook this study to clarify the mechanisms underlying the efficacy of IL-6 blockade in RA and to find a novel target for treatment of RA.We examined gene expression profiles of CD4+ T cells by DNA microarray analysis before and after treatment with an anti-IL-6 receptor antibody, tocilizumab (TCZ), in RA patients who exhibited good clinical responses to the treatment. Using murine CD4+ T cells, we then examined the roles of a newly identified molecule whose expression was significantly reduced in CD4+ T cells by TCZ therapy. We also examined the effect of the forced expression of the molecule on retinoic acid receptor-related orphan nuclear receptor γt (RORγt)-induced IL-17A production in CD4+ T cells and on RORγt-induced IL-17A promoter activation.We identified AT-rich-interactive domain- containing protein 5A (ARID-5A) as a new molecule down-regulated by IL-6 blockade in the form of TCZ therapy. IL-6 induced the expression of ARID-5A in CD4+ T cells during Th17 cell differentiation by a STAT-3-dependent mechanism, whereas IL-6-induced ARID-5A expression was not affected by the absence of RORγt, a lineage-specifying transcription factor of Th17 cells. Furthermore, ARID-5A physically associated with RORγt through its N-terminal region and inhibited RORγt-induced Th17 cell differentiation.ARID-5A is a lineage-specific attenuator of Th17 cell differentiation and may be involved in the pathogenesis of RA.

This paper presents the development of a remotely operated brachytherapy system for oral cancer. The purpose of the system is to reduce exposure dose to the operator by keeping him or her at a distance of 1m from radioactive 198Au sources. The system is designed to enable the operator to perform the same procedure as for conventional brachytherapy treatment without exposure dose. It consists of an applicator manipulator to implant 198Au seeds, a finger device to firmly grasp organs in the mouth, operational parts for both of them, and a base arm. The applicator manipulator and finger device are fixed on the base arm which is fixed on a radiation shield. The applicator manipulator is a serial manipulator with five degrees of freedom (DOF). The finger device is necessary because organs in the mouth are soft and flexible. Its finger and operational parts are identical three DOF parallel manipulators. This paper details preliminary experiments of grasping and insertion with the system. These experiments verify that an operator can grasp a tongue model with the finger device and can insert the applicator into it with the applicator manipulator. In our developed system, we also confirm that rotation of the applicator needle can reduce insertion force.

Protein phosphorylation can be regulated by changes in kinase activity, phosphatase activity, or both. GABA(B) receptor R2 subunit (GABA(B)R2) is phosphorylated at S783 by 5'-AMP-activated-protein kinase (AMPK), and this phosphorylation modulates GABA(B) receptor desensitization. Since the GABA(B) receptor is an integral membrane protein, solubilizing GABA(B)R2 is difficult. To circumvent this problem and to identify specific phosphatases that dephosphorylate S783, we employed an in vitro assay based on dephosphorylation of proteins on PVDF membranes by purified phosphatases. Our method allowed us to demonstrate that S783 in GABA(B)R2 is directly dephosphorylated by PP2A (but not by PP1, PP2B nor PP2C) in a dose-dependent and okadaic acid-sensitive manner. We also show that the level of phosphorylation of the catalytic subunit of AMPK at T172 is reduced by PP1, PP2A and PP2C. Our data indicate that PP2A dephosphorylates GABA(B)R2(S783) less efficiently than AMPK(T172), and that additional phosphatases might be involved in S783 dephosphorylation.

Background: Lacrimal gland enlargement (LGE) is one of the characteristics of Mikulicz’s disease (MD). Recently, marked serum immunoglobulin (Ig)G4 elevation and infiltration of IgG4-positive plasma cells in the enlarged exocrine glands have been reported in MD patients. Moreover, we have reported that in patients with LGE and elevated serum IgG4 levels (IgG4-related LGE), T helper type 2 (Th2) cell-mediated immune responses are enhanced. Although prostaglandin D2 (PGD2) and its receptor CRTH2 (chemoattractant receptor-homologous molecule expressed on Th2 cells) have been shown to be involved in Th2 cell-related diseases such as bronchial asthma, their roles in IgG4-related diseases remain unknown. Aim: The aim of this study is to address the role of CD4+ T cells expressing CRTH2 (CRTH2+ CD4+ T cells) in IgG4-related LGE. Methods: We examined the expression of CCR4, CXCR3 and CRTH2 on peripheral blood CD4+ T cells in patients with IgG4-related LGE, in patients with bronchial asthma and in healthy controls. Results: The ratio of CCR4+ to CXCR3+ in CD45RO+ CD4+ T cells was increased in patients with IgG4-related LGE when compared to that in healthy controls, confirming that Th2 cells are predominant in patients with IgG4-related LGE. In addition, the frequency of CRTH2+ cells in CD4+ T cells was significantly increased in these patients, compared to healthy controls. Furthermore, although not statistically significant, the frequency of CRTH2+ cells in CD4+ T cells tended to correlate with the levels of serum IgE and the number of blood eosinophils in patients with IgG4-related LGE. Conclusion: CRTH2+ CD4+ T cells may be involved in the pathogenesis of IgG4-related LGE.

「高齢者の食生活は、栄養教育により変容し得る」という視点から、高齢者に短期継続的栄養教育を行い、その効果判定を試みた結果は次のとおりであった。 1.身体的健康度:体脂肪率は増加傾向、血圧値は低下傾向にあり、女性の収縮期血圧は有意に低下した(pく0.01)。 2.精神的健康度:女性の精神生活得点は有意に高まり(pく0.01)、男女とも社会生活得点、休養・余暇生活得点が後期に高まっていたが、女性にのみ有意であった(pく0.01)。家族形態別では一人暮らし高齢者に最も改善がみられた。 3.食品群・栄養素等摂取の変化:食品群摂取は、第5群(穀類、芋類、砂糖類)以外は男女とも所要量充足率が上がり、食生活の改善がみられた。特に女性の第4群(果物類)、第6群(油脂類)摂取増は有意であり(p, Nutrition education for the elderly was conducted. We compared the situation before and after the study, and examined their food intake and their physical and mental health. The results are as follows. 1.Physical health : The rate of body fat rose, though statistically not significant. Blood pressure, both systolic and diastolic lowered. 2.Mental health : Looking over their lifestyle, women's mental conditions improved. Social score and rest-leisure score increased for both men and women, though statistically not significant. Elderly people in elderly couples changed the score remarkably, when compared with the others. 3.Food and nutrition : Eating habits improved significantly(p

Recent studies have suggested that statins, the inhibitors for 3-hydroxy-3-methyglutaryl (HMG)-CoA reductase in the mevalonate pathway, exhibit anti-inflammatory effects. However, the immune modulatory effects of statins on the differentiation of CD4(+) T cells and their underlying mechanisms are still largely unknown. To address these issues, we examined the effect of simvastatin and inhibitors for protein farnesylation and geranylgeranylation on the differentiation of IL-17-producing T cells (T(h)17 cells) and Foxp3(+) CD4(+) T cells. Simvastatin inhibited the differentiation of T(h)17 cells through the inhibition of HMG-CoA reductase activity but enhanced the differentiation of Foxp3(+) CD4(+) T cells. Geranylgeranyltransferase I inhibitor, GGTI-298, but not farnesyltransferase inhibitor, FTI-277, mimicked the effects of simvastatin, indicating that the inhibition of protein geranylgeranylation is responsible for the effects. Moreover, Foxp3(+) CD4(+) T cells developed in the presence of transforming growth factor-beta and GGTI-298 functioned as regulatory T cells (Tregs) in in vitro T cell proliferation assay as well as in an autoimmune colitis model. Finally, GGTI-298 induced SOCS3 expression and inhibited IL-6-induced signal transducers and activators of transcription3 phosphorylation in CD4(+) T cells. Taken together, these results indicate that protein geranylgeranylation enhances the differentiation of T(h)17 cells and inhibits the differentiation of Foxp3(+) Tregs partly via the inhibition of SOCS3 expression.

The mechanism which discriminates the pattern classes at the same $\lambda$, is found. It is closely related to the structure of the rule table and expressed by the numbers of the rules which break the strings of the quiescent states. It is shown that for the N-neighbor and K-state cellular automata, the class I, class II, class III and class IV patterns coexist at least in the range, $\frac{1}{K} \le \lambda \le 1-\frac{1}{K} $. The mechanism is studied quantitatively by introducing a new parameter $F$, which we call quiescent string dominance parameter. It is taken to be orthogonal to $\lambda$. Using the parameter F and $\lambda$, the rule tables of one dimensional 5-neighbor and 4-state cellular automata are classified. The distribution of the four pattern classes in ($\lambda$,F) plane shows that the rule tables of class III pattern class are distributed in larger $F$ region, while those of class II and class I pattern classes are found in the smaller $F$ region and the class IV behaviors are observed in the overlap region between them. These distributions are almost independent of $\lambda$ at least in the range $0.25 \leq \lambda \leq 0.75$, namely the overlapping region in $F$, where the class III and class II patterns coexist, has quite gentle $\lambda$ dependence in this $\lambda$ region. Therefore the relation between the pattern classes and the $\lambda$ parameter is not observed. PACS: 89.75.-k Complex Systems, Comment: 27 pages in latex, 10 Postscript figures

SUMMARYTo study the activation states and cytokine profiles of pulmonary T cells in corticosteroid-resistant and corticosteroid-sensitive interstitial pneumonitis (IP) in dermatomyositis (DM)/polymyositis (PM), we examined the activation markers and cytokine profiles of T cells in bronchoalveolar lavage fluids (BALF) from patients with IP in DM/PM before prednisolone therapy and then compared the activation states of T cells according to the therapeutic response of IP to prednisolone therapy. CD25+ CD4+ T cells in BALF were significantly increased in both corticosteroid-resistant and corticosteroid-sensitive IP in DM/PM as compared with those in controls without IP. Furthermore, CD25+ CD4+ T cells in BALF were significantly more increased in corticosteroid-resistant IP than those in cortico teroid- sensitive IP. Moreover, CD25+ CD8+ T cells in BALF were significantly increased only in corticosteroid-resistant IP, but not in corticosteroid-sensitive IP or controls without IP. IFN-γ mRNA was detected in BALF T cells in corticosteroid-resistant and corticosteroid-sensitive IP but not in controls without IP, whereas IL-4 mRNA was virtually undetected in BALF T cells in both the IP groups. However, there were no significant differences in CD4/CD8 ratio of BALF T cells, HLA-DR+ BALF T cells or CD25+ and HLA-DR+ peripheral blood T cells between the two IP groups. These results indicate that activated Th1-type pulmonary T cells play an important role in the development of corticosteroid- resistant IP in DM/PM and that the increase in CD25+ CD8+ T cells in BALF is a useful indicator for corticosteroid-resistant IP in DM/PM and hence may be an indicator for early use of cyclosporin.

Background Although the overall response rate of rheumatoid arthritis (RA) to tocilizumab is high, improvement of synovitis in patients treated with tocilizumab is frequently slow as compared with that in patients treated with TNF antagonists. Therefore, predicting therapeutic responses of RA to tocilizumab can be particularly beneficial but such methods with sufficient accuracy have not been established yet. Objectives This prospective multicenter study aimed to identify biomarkers to predict therapeutic responses to tocilizumab in patients with RA by analyzing comprehensive gene expression in peripheral blood mononuclear cells (PBMCs). Methods Patients with RA who received treatment with tocilizumab for the first time were recruited and therapeutic responses were determined at 6 months. In the training cohort (n=40), gene expression in PBMCs at baseline was analyzed using genome-wide DNA microarray with 41,000 probes representing 19,416 genes. In the validation cohort (n=20), expression levels of the candidate genes in PBMCs at baseline were determined using real-time quantitative PCR (qPCR) analysis. Results In hierarchical clustering analyses, gene expression patterns of non-responders clustered in the same branch (Figure) and 68 DNA microarray probes which showed a significant difference in signal intensity between non-responders and responders in the training cohort were identified. Nineteen putative genes were selected and a significant correlation between the DNA microarray signal intensity and the qPCR relative expression was confirmed for 15 genes. In the validation cohort, a significant difference in relative expression between non-responders and responders was reproduced for 3 type I interferon response genes ( IFI6 , MX2 , OASL ) and MT1G . Models incorporating these genes provided a maximum area under the receiver operating characteristic curve of 0.947 in predicting a moderate or good response in the validation cohort. Conclusions Our study using genome-wide DNA microarray analyses identified the candidate biomarkers to predict therapeutic responses to tocilizumab in patients with RA and suggests the involvement of type I interferon signaling and metallothioneins in the pathophysiology of RA. Acknowledgements This work was supported by a Health Labour Sciences Research Grant on Allergic Disease and Immunology from the Ministry of Health, Labor and Welfare of Japan, and a Regional Innovation Strategy Support Program and a Grants-in-Aids for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology. Disclosure of Interest Y. Sanayama: None declared, K. Ikeda: None declared, Y. Saito: None declared, S.-I. Kagami: None declared, M. Yamagata: None declared, S. Furuta: None declared, D. Kashiwakuma: None declared, I. Iwamoto: None declared, T. Umibe: None declared, Y. Nawata: None declared, R. Matsumura: None declared, T. Sugiyama: None declared, M. Sueishi: None declared, M. Hiraguri: None declared, K. Nonaka: None declared, O. Ohara: None declared, H. Nakajima Grant/research support: Chugai Pharmaceutical Co., Ltd. DOI 10.1136/annrheumdis-2014-eular.2293

SUMMARYCD4+ T cells are thought to play an important role in airway inflammation in both atopic and non-atopic asthma. However, the mechanism by which T cells are activated in non-atopic asthma, where there is no causative antigen identified, is unknown. To elucidate this issue, we analysed T cell receptor (TCR) Vβ gene clonotypes of T cells in the bronchoalveolar lavage fluids (BALF) of non-atopic asthmatics using polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) analysis and a sequencing method. We found that the numbers of TCR Vβ gene clonotypes of T cells in the BALF of non-atopic asthmatics were significantly increased compared with those of peripheral blood lymphocytes (PBL). We also found that there were several shared amino acid motifs in complementarity-determining region 3 (CDR3) of TCR Vβ genes from those T cell clones in BALF of non-atopic asthmatics, whereas these shared motifs were not found in the same Vβ family genes from PBL in the patients. Moreover, a conserved amino acid sequence was detected in two patients who shared a common HLA-DR allele. These results indicate that the infiltrating T cells in the airways of non-atopic asthmatics recognize relatively limited epitopes of antigens and are clonally expanded by antigen-driven stimulation.