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Deletion of IFT20 in early stage T lymphocyte differentiation inhibits the development of collagen-induced arthritis.

ABSTRACT: IFT20 is the smallest member of the intraflagellar transport protein (IFT) complex B. It is involved in cilia formation. Studies of IFT20 have been confined to ciliated cells. Recently, IFT20 was found to be also expressed in non-ciliated T cells and have functions in immune synapse formation and signaling in vitro. However, how IFT20 regulates T-cell development and activation in vivo is still unknown. We deleted the IFT20 gene in early and later stages of T-cell development by crossing IFT20(flox/flox) (IFT20(f/f) ) mice with Lck-Cre and CD4-Cre transgenic mice, and investigated the role of IFT20 in T-cell maturation and in the development of T cell-mediated collagen-induced arthritis (CIA). We found that both Lck-Cre/IFT20(f/f) and CD4-Cre/IFT20(f/f) mice were indistinguishable from their wild-type littermates in body size, as well as in the morphology and weight of the spleen and thymus. However, the number of CD4- and CD8-positive cells was significantly lower in thymus and spleen in Lck-Cre/IFT20(f/f) mice. Meanwhile, the incidence and severity of CIA symptoms were significantly decreased, and inflammation in the paw was significantly inhibited in Lck-Cre/IFT20(f/f) mice compared to Lck-Cre/IFT20(+/+) littermates. Deletion IFT20 in more mature T cells of CD4-Cre/IFT20(f/f) mice had only mild effects on the development of T cells and CIA. The expression of IL-1β, IL-6 and TGF-β1 were significantly downregulated in the paw of Lck-Cre/IFT20(f/f) mice, but just slight decreased in CD4-Cre/IFT20(f/f) mice. These results demonstrate that deletion of IFT20 in the early stage of T-cell development inhibited CIA development through regulating T-cell development and the expression of critical cytokines.

SUBMITTER: Yuan X

PROVIDER: S-EPMC4470568 | biostudies-other | 2014

REPOSITORIES: biostudies-other

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Project description:IntroductionIn this article, we present a comparative immunohistochemical evaluation of four clinical-stage antibodies (L19, F16, G11 and F8) directed against splice isoforms of fibronectin and of tenascin-C for their ability to stain synovial tissue alterations in rheumatoid arthritis patients. Furthermore we have evaluated the therapeutic potential of the most promising antibody, F8, fused to the anti-inflammatory cytokine interleukin (IL) 10.MethodsF8-IL10 was produced and purified to homogeneity in CHO cells and shown to comprise biological active antibody and cytokine moieties by binding assays on recombinant antigen and by MC/9 cell proliferation assays. We have also characterized the ability of F8-IL10 to inhibit arthritis progression in the collagen-induced arthritis mouse model.ResultsThe human antibody F8, specific to the extra-domain A of fibronectin, exhibited the strongest and most homogenous staining pattern in synovial biopsies and was thus selected for the development of a fully human fusion protein with IL10 (F8-IL10, also named DEKAVIL). Following radioiodination, F8-IL10 was able to selectively target arthritic lesions and tumor neo-vascular structures in mice, as evidenced by autoradiographic analysis and quantitative biodistribution studies. The subcutaneous administration route led to equivalent targeting results when compared with intravenous administration and was thus selected for the clinical development of the product. F8-IL10 potently inhibited progression of established arthritis in the collagen-induced mouse model when tested alone and in combination with methotrexate. In preparation for clinical trials in patients with rheumatoid arthritis, F8-IL10 was studied in rodents and in cynomolgus monkeys, revealing an excellent safety profile at doses tenfold higher than the planned starting dose for clinical phase I trials.ConclusionsFollowing the encouraging preclinical results presented in this paper, clinical trials with F8-IL10 will now elucidate the therapeutic potential of this product and whether the targeted delivery of IL10 potentiates the anti-arthritic action of the cytokine in rheumatoid arthritis patients.

Project description:Collagen triple helix repeat-containing1 (Cthrc1) has previously been implicated in osteogenic differentiation and positive regulation of bone mass, however, the underlying mechanisms remain unclear. Here we characterized the bone phenotype of a novel Cthrc1 null mouse strain using bone histomorphometry, ?CT analysis and functional readouts for bone strength. In male Cthrc1 null mice both trabecular bone as well as cortical bone formation was impaired, whereas in female Cthrc1 null mice only trabecular bone parameters were altered. Novel and highly specific monoclonal antibodies revealed that CTHRC1 is expressed by osteocytes and osteoblasts, but not osteoclasts. Furthermore, Cthrc1 null mice exhibited increased bone resorption with increased number of osteoclast and increased osteoclast activity together with enhanced expression of osteoclastogenic genes such as c-Fos, Rankl, Trap, and Nfatc1. Differentiation of bone marrow-derived monocytes isolated from Cthrc1 null mice differentiated into osteoclasts as effectively as those from wildtype mice. In the presence of CTHRC1 osteoclastogenic differentiation of bone marrow-derived monocytes was dramatically inhibited as was functional bone resorption by osteoclasts. This process was accompanied by downregulation of osteoclastogenic marker genes, indicating that extrinsically derived CTHRC1 is required for such activity. In vitro, CTHRC1 had no effect on osteogenic differentiation of bone marrow stromal cells, however, calvarial osteoblasts from Cthrc1 null mice exhibited reduced osteogenic differentiation compared to osteoblasts from wildtypes. In a collagen antibody-induced arthritis model Cthrc1 null mice suffered significantly more severe inflammation and joint destruction than wildtypes, suggesting that CTHRC1 expressed by the activated synoviocytes has anti-inflammatory effects. Mechanistically, we found that CTHRC1 inhibited NF?B activation by preventing I?B? degradation while also inhibiting ERK1/2 activation. Collectively our studies demonstrate that CTHRC1 secreted from osteocytes and osteoblasts functions as an inhibitor of osteoclast differentiation via inhibition of NF?B-dependent signaling. Furthermore, our data suggest that CTHRC1 has potent anti-inflammatory properties that limit arthritic joint destruction.

Project description:BackgroundRheumatoid arthritis (RA) is a chronic autoimmune disease. Soufeng sanjie formula (SF), which is composed of scolopendra (dried body of Scolopendra subspinipes mutilans L. Koch), scorpion (dried body of Buthus martensii Karsch), astragali radix (dried root of Astragalus membranaceus (Fisch.) Bge), and black soybean seed coats (seed coats of Glycine max (L.) Merr), is a traditional Chinese prescription for treating RA. However, the mechanism of SF in treating RA remains unclear. This study was aim to investigate the anti-arthritic effects of SF in a collagen-induced arthritis (CIA) mouse model and explore the mechanism by which SF alleviates arthritis in CIA mice.MethodsFor in vivo studies, female DBA/1J mice were used to establish the CIA model, and either SF (183 or 550 mg/kg/day) or methotrexate (MTX, 920 mg/kg, twice/week) was orally administered to the mice from the day of arthritis onset. After administration for 30 days, degree of ankle joint destruction and serum levels of IgG and inflammatory cytokines were determined. The balance of Th17/Treg cells in the spleen and lymph nodes was analyzed using flow cytometry. Moreover, the expression levels of retinoic acid receptor-related orphan nuclear receptor (ROR) γt and phosphorylated STAT3 (pSTAT3, Tyr705) in the spleen were detected by immunohistochemistry. Furthermore, the effect of SF on Th17 cells differentiation in vitro was investigated in CD4+ T cells under Th17 polarization conditions.ResultsSF decreased the arthritis score, ameliorated paw swelling, and reduced cartilage loss in the joint of CIA mice. In addition, SF decreased the levels of bovine collagen-specific IgG in sera of CIA mice. SF decreased the levels of inflammatory cytokines (TNF-α, IL-6, and IL-17A) and increased the level of IL-10 both in the sera and the joint of CIA mice. Moreover, SF treatment rebalanced the Th17/Treg ratio in the spleen and lymph nodes of CIA mice. SF also reduced the expression levels of ROR γt and pSTAT3 (Tyr705) in the spleen of CIA mice. In vitro, SF treatment reduced Th17 cell generation and IL-17A production and inhibited the expression of RORγt, IRF4, IL-17A, and pSTAT3 (Tyr705) under Th17 polarization conditions.ConclusionsOur results suggest that SF exhibits anti-arthritic effects and restores Th17/Treg homeostasis in CIA mice by inhibiting Th17 cell differentiation.

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Deletion of IFT20 in early stage T lymphocyte differentiation inhibits the development of collagen-induced arthritis.

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