STAT3 and NF-kappaB signal pathway is required for IL-23-mediated IL-17 production in spontaneous arthritis animal model IL-1 receptor antagonist-deficient mice

IL-23 is a heterodimeric cytokine composed of a p19 subunit and the p40 subunit of IL-12. IL-23 has proinflammatory activity, inducing IL-17 secretion from activated CD4(+) T cells and stimulating the proliferation of memory CD4(+) T cells. We investigated the pathogenic role of IL-23 in CD4(+) T cells in mice lacking the IL-1R antagonist (IL-1Ra(-/-)), an animal model of spontaneous arthritis. IL-23 was strongly expressed in the inflamed joints of IL-1Ra(-/-) mice. Recombinant adenovirus expressing mouse IL-23 (rAd/mIL-23) significantly accelerated this joint inflammation and joint destruction. IL-1beta further increased the production of IL-23, which induced IL-17 production and OX40 expression in splenic CD4(+) T cells of IL-1Ra(-/-) mice. Blocking IL-23 with anti-p19 Ab abolished the IL-17 production induced by IL-1 in splenocyte cultures. The process of IL-23-induced IL-17 production in CD4(+) T cells was mediated via the activation of Jak2, PI3K/Akt, STAT3, and NF-kappaB, whereas p38 MAPK and AP-1 did not participate in the process. Our data suggest that IL-23 is a link between IL-1 and IL-17. IL-23 seems to be a central proinflammatory cytokine in the pathogenesis of this IL-1Ra(-/-) model of spontaneous arthritis. Its intracellular signaling pathway could be useful therapeutic targets in the treatment of autoimmune arthritis.     

IL-23 inhibits osteoclastogenesis indirectly through lymphocytes and is required for the maintenance of bone mass in mice

IL-23 stimulates the differentiation and function of the Th17 subset of CD4(+) T cells and plays a critical role in chronic inflammation. The IL-23 receptor-encoding gene is also an inflammatory disease susceptibility gene. IL-23 shares a common subunit with IL-12, a T cell-dependent osteoclast formation inhibitor, and we found that IL-23 also dose-dependently inhibited osteoclastogenesis in a CD4(+) T lymphocyte-dependent manner. When sufficiently enriched, gammadelta T cells also mediated IL-23 inhibition. Like IL-12, IL-23 acted synergistically with IL-18 to block osteoclastogenesis but, unlike IL-12, IL-23 action depended on T cell GM-CSF production. IL-23 did not mediate IL-12 action although IL-12 induced its expression. Male mice lacking IL-23 (IL-23p19(-/-)) had approximately 30% lower bone mineral density and tibial trabecular bone mass (bone volume (BV)/total volume (TV)) than wild-type littermates at 12 wk and 40% lower BV/TV at 26 wk of age; male heterozygotes also had lower bone mass. Female IL-23p19(-/-) mice also had reduced BV/TV. IL-23p19(-/-) mice had no detectable osteoclast defect in trabecular bone but IL-23p19(-/-) had thinner growth plate hypertrophic and primary spongiosa zones (and, in females, less cartilage remnants) compared with wild type. This suggests increased osteoclast action at and below the growth plate, leading to reduced amounts of mature trabecular bone. Thus, IL-23 inhibits osteoclast formation indirectly via T cells in vitro. Under nonpathological conditions (unlike inflammatory conditions), IL-23 favors higher bone mass in long bones by limiting resorption of immature bone forming below the growth plate.     

Tacrolimus potently inhibits human osteoclastogenesis induced by IL-17 from human monocytes alone and suppresses human Th17 differentiation

Tacrolimus (FK506, Prograf?) is an orally available, T cell specific and anti-inflammatory agent that has been proposed as a therapeutic drug in rheumatoid arthritis (RA) patients. It has been known that T cells have a critical role in the pathogenesis of RA. Recent studies suggest that Th17 cells, which mainly produce IL-17, are involved in many autoimmune inflammatory disease including RA. The present study was undertaken to assess the effect of tacrolimus on IL-17-induced human osteoclastogenesis and human Th17 differentiation. Human CD14(+) monocytes were cultured in the presence of macrophage-colony stimulating factor (M-CSF) and IL-17. From day 4, tacrolimus was added to these cultures. Osteoclasts were immunohistologically stained for vitronectin receptor 10days later. IL-17 production from activated T cells stimulated with IL-23 was measured by enzyme-linked immunosorbent assay (ELISA). Th17 differentiation from na?ve T cells was assayed by flow cytometry. Tacrolimus potently inhibited IL-17-induced osteoclastogenesis from human monocytes and osteoclast activation. Addition of tacrolimus also reduced production of IL-17 in human activated T cells stimulated with IL-23. Interestingly, the population of human IL-17(+)IFN-γ(-) CD4 T cells or IL-17(+)TNF-α(+) CD4 T cells were decreased by adding of tacrolimus. The present study demonstrates that the inhibitory effect of tacrolimus on IL-17-induced osteoclastogenesis from human monocytes. Tacrolimus also inhibited expression of IL-17 or TNF-α by reducing the proportion of Th17, suggesting that therapeutic effect on Th17-associated disease such as RA, inflammatory bowel disease, multiple sclerosis, psoriasis, or allograft rejection.     

IL-23 induces human osteoclastogenesis via IL-17 <Emphasis Type="Italic">in vitro</Emphasis>, and anti-IL-23 antibody attenuates collagen-induced arthritis in rats

This study demonstrates that IL-23 stimulates the differentiation of human osteoclasts from peripheral blood mononuclear cells (PBMC). Furthermore, in vivo blockade of endogenous IL-23 activity by treatment with anti-IL-23 antibody attenuates collagen-induced arthritis in rats by preventing both inflammation and bone destruction. IL-23 induced human osteoclastogenesis in cultures of PBMC in the absence of osteoblasts or exogenous soluble-receptor activator of NF-kappaB ligand (RANKL). This IL-23-induced osteoclastogenesis was inhibited by osteoprotegerin, anti-IL-17 antibody, and etanercept, suggesting that RANKL, IL-17, and TNF-alpha are involved. In addition, we found the ratio of production levels of IL-17 to those of IFN-gamma from activated human T cells was elevated at 1 to 10 ng/ml IL-23. The inductive effect of IL-17 and the inhibitory effect of IFN-gamma on osteoclastogenesis indicate that the balance of these two cytokines is particularly important. We also demonstrated that IL-23 administered at a later stage significantly reduced paw volume in rats with collagen-induced arthritis, in a dose-dependent manner. Furthermore, anti-IL-23 antibody reduced synovial tissue inflammation and bone destruction in these rats. These findings suggest that IL-23 is important in human osteoclastogenesis and that neutralizing IL-23 after onset of collagen-induced arthritis has therapeutic potential. Thus, controlling IL-23 production and function could be a strategy for preventing inflammation and bone destruction in patients with rheumatoid arthritis.     

The tachykinins substance P and hemokinin-1 favor the generation of human memory Th17 cells by inducing IL-1β, IL-23, and TNF-like 1A expression by monocytes

The nervous system influences immune responses through the release of neural factors such as neuropeptides. Among them, the tachykinin substance P (SP) signals via the neurokinin 1 receptor (NK-1R), which is expressed by various immune cells. We thereby analyzed in this paper whether tachykinins may participate in human CD4(+) Th cell polarization. We report that SP and hemokinin-1 (HK-1) upregulate IL-17A and IFN-γ production by human memory CD4(+) T cells without affecting IL-4 and IL-10 production. SP and HK-1 switch non-Th17-committed CD4(+) memory T cells into bona fide Th17 cells and Th1/Th17 cells. In contrast, SP and HK-1 do not modulate the polarization of naive CD4(+) T cells. SP- and HK-1-induced Th17 cell generation is mediated through NK-1R and requires the presence of monocytes. SP and HK-1 trigger IL-1β, IL-6, and TNF-α production, upregulate IL-23 production, and enhance TNF-like 1A expression on monocyte surface. Neutralization experiments demonstrated that IL-1β, IL-23, and TNF-like 1A are involved in the SP- and HK-1-induced Th17 cell. The other members of the tachykinin family, neurokinins A and B, have no effect on the differentiation of naive and memory T cells. These results thereby show that SP and HK-1 are novel Th17 cell-inducing factors that may act locally on memory T cells to amplify inflammatory responses.     

IL-23 is critical for induction of arthritis, osteoclast formation, and maintenance of bone mass

The role of IL-23 in the development of arthritis and bone metabolism was studied using systemic IL-23 exposure in adult mice via hydrodynamic delivery of IL-23 minicircle DNA in vivo and in mice genetically deficient in IL-23. Systemic IL-23 exposure induced chronic arthritis, severe bone loss, and myelopoiesis in the bone marrow and spleen, which resulted in increased osteoclast differentiation and systemic bone loss. The effect of IL-23 was partly dependent on CD4(+) T cells, IL-17A, and TNF, but could not be reproduced by overexpression of IL-17A in vivo. A key role in the IL-23-induced arthritis was made by the expansion and activity of myeloid cells. Bone marrow macrophages derived from IL-23p19(-/-) mice showed a slower maturation into osteoclasts with reduced tartrate-resistant acid phosphatase-positive cells and dentine resorption capacity in in vitro osteoclastogenesis assays. This correlated with fewer multinucleated osteoclast-like cells and more trabecular bone volume and number in 26-wk-old male IL-23p19(-/-) mice compared with control animals. Collectively, our data suggest that systemic IL-23 exposure induces the expansion of a myeloid lineage osteoclast precursor, and targeting IL-23 pathway may combat inflammation-driven bone destruction as observed in rheumatoid arthritis and other autoimmune arthritides.     

T cell-mediated increased osteoclast formation from peripheral blood as a mechanism for Crohn's disease-associated bone loss

The pathophysiology of osteoporosis in patients with Crohn's disease (CD) is still not completely elucidated. In this study, we evaluated osteoclastogenesis from peripheral blood cells of CD patients and studied the role of lymphocytes and inflammatory cytokines in this process. Peripheral blood mononuclear cells from seven patients with quiescent CD and matched healthy controls were isolated, and separated into T cells, B cells, and a T- and B-cell depleted fraction. In various culture combinations, osteoclast formation in the absence of the osteoclastogenic factors RANKL and M-CSF was assessed by scoring the number of tartrate-resistant acid phosphatase (TRACP) positive multinucleated cells (MNCs). Cytokine levels in culture supernatants were measured. Formation of heterogeneous cell clusters in culture was noticed; a process that was inhibited by anti-LFA-1. In CD cultures, mean cluster area was up to threefold higher than in control cultures, and shown to be induced by T cells. Over tenfold higher numbers of TRACP(+) MNCs were found in CD cultures, but exclusively in cultures containing T cells. Formation of cell clusters correlated strongly with formation of TRACP(+) MNCs. Both cell cluster formation and osteoclast formation were related to IL-17 levels in vitro. In conclusion, osteoclastogenesis, preceded by cell cluster formation, is T cell-mediated and increased in patients with quiescent CD. Our findings suggest heterotypic interactions between osteoclast precursors and T cells to be a triggering step in osteoclast formation in CD. Furthermore, our results propose a possible role for IL-17 in osteoclastogenesis in CD patients, and as such in CD-associated bone loss.     

Induction of IL-17+ T cell trafficking and development by IFN-gamma: mechanism and pathological relevance in psoriasis

Th1 and Th17 T cells are often colocalized in pathological environments, yet Th1-derived IFN-gamma inhibits Th17 cell development in vitro. We explored the physiologic basis of this paradox in humans. In this study, we demonstrate increased the number of CD4(+) and CD8(+) IL-17(+) T cells in skin lesions of psoriasis. Furthermore, we show that myeloid APCs potently support induction of IL-17(+) T cells, and that this activity is greatly increased in psoriasis. We tested stimuli that might account for this activity. Th1 cells and IFN-gamma are increased in psoriatic blood and lesional skin. We show that IFN-gamma programs myeloid APCs to induce human IL-17(+) T cells via IL-1 and IL-23. IFN-gamma also stimulates APC production of CCL20, supporting migration of IL-17(+) T cells, and synergizes with IL-17 in the production of human beta-defensin 2, an antimicrobial and chemotactic protein highly overexpressed by psoriatic keratinocytes. This study reveals a novel mechanistic interaction between Th1 and IL-17(+) T cells, challenges the view that Th1 cells suppress Th17 development through IFN-gamma, and suggests that Th1 and IL-17(+) T cells may collaboratively contribute to human autoimmune diseases.     

IL-12 alone and in synergy with IL-18 inhibits osteoclast formation in vitro

IL-12, like IL-18, was shown to potently inhibit osteoclast formation in cultures of cocultures of murine osteoblast and spleen cells, as well as in adult spleen cells treated with M-CSF and receptor activator of NF-kappaB ligand (RANKL). Neither IL-12 nor IL-18 was able to inhibit RANKL-induced osteoclast formation in cultured RAW264.7 cells, demonstrating that IL-12, like IL-18, was unable to act directly on osteoclastic precursors. IL-12, like IL-18, was found to act by T cells, since depletion of T cells from the adult spleen cell cultures ablated the inhibitory action of IL-12 and addition of either CD4 or CD8 T cells from C57BL/6 mice to RANKL-stimulated RAW264.7 cultures permitted IL-12 or IL-18 to be inhibitory. Additionally, IL-12 was still able to inhibit osteoclast formation in cocultures with osteoblasts and spleen cells from either GM-CSF R(-/-) mice or IFN-gamma R(-/-) mice, indicating that neither GM-CSF nor IFN-gamma was mediating osteoclast inhibition in these cultures. Combined, IL-18 and IL-12 synergistically inhibited osteoclast formation at concentrations 20- to 1000-fold less, respectively, than when added individually. A candidate inhibitor could not be demonstrated using neutralizing Abs to IL-4, IL-10, or IL-13 or from mRNA expression profiles among known cytokine inhibitors of osteoclastogenesis in response to IL-12 and IL-18 treatment, although the unknown inhibitory molecule was determined to be secreted from T cells.     

Marking and quantifying IL-17A-producing cells in vivo

Interleukin (IL)-17A plays an important role in host defense against a variety of pathogens and may also contribute to the pathogenesis of autoimmune diseases. However, precise identification and quantification of the cells that produce this cytokine in vivo have not been performed. We generated novel IL-17A reporter mice to investigate expression of IL-17A during Klebsiella pneumoniae infection and during experimental autoimmune encephalomyelitis, conditions previously demonstrated to potently induce IL-17A production. In both settings, the majority of IL-17A was produced by non-CD4(+) T cells, particularly γδ T cells, but also invariant NKT cells and other CD4(-)CD3ε(+) cells. As measured in dual-reporter mice, IFN-γ-producing Th1 cells greatly outnumbered IL-17A-producing Th17 cells throughout both challenges. Production of IL-17A by cells from unchallenged mice or by non-T cells under any condition was not evident. Administration of IL-1β and/or IL-23 elicited rapid production of IL-17A by γδ T cells, invariant NKT cells and other CD4(-)CD3ε(+) cells in vivo, demonstrating that these cells are poised for rapid cytokine production and likely comprise the major sources of this cytokine during acute immunologic challenges.     

Regulation of IL-17 in human CCR6+ effector memory T cells

IL-17-secreting T cells represent a distinct CD4(+) effector T cell lineage (Th17) that appears to be essential in the pathogenesis of numerous inflammatory and autoimmune diseases. Although extensively studied in the murine system, human Th17 cells have not been well characterized. In this study, we identify CD4(+)CD45RO(+)CCR7(-)CCR6(+) effector memory T cells as the principal IL-17-secreting T cells. Human Th17 cells have a unique cytokine profile because the majority coexpress TNF-alpha but not IL-6 and a minor subset express IL-17 with IL-22 or IL-17 and IFN-gamma. We demonstrate that the cytokines that promote the differentiation of human naive T cells into IL-17-secreting cells regulate IL-17 production by memory T cells. IL-1beta alone or in association with IL-23 and IL-6 markedly increase IL-17(+) CCR6(+) memory T cells and induce IL-17 production in CCR6(-) memory T cells. We also show that T cell activation induces Foxp3 expression in T cells and that the balance between the percentage of Foxp3(+) and IL-17(+) T cells is inversely influenced by the cytokine environment. These studies suggest that the cytokine environment may play a critical role in the expansion of memory T cells in chronic autoimmune diseases.     

Retinoic acid attenuates rheumatoid inflammation in mice

Retinoic acid is the active vitamin A derivative and is well-known to have diverse immunomodulatory actions. In this study, we investigated the impact of all-trans retinoic acid (ATRA), a biologic key metabolite of vitamin A, on the development of arthritis and the pathophysiologic mechanisms by which ATRA might have antiarthritic effects in animal model of rheumatoid arthritis (RA; collagen-induced arthritis [CIA] in DBA/1J mice). We showed that treatment with ATRA markedly suppressed the clinical and histologic signs of arthritis in the CIA mice. It reduced the expression of IL-17 in the arthritic joints. Interestingly, Foxp3(+) regulatory T cells were markedly increased and IL-17-producing CD4(+) T cells (Th17 cells) were decreased in the spleens of ATRA-treated mice. In vitro treatment with ATRA induced the expression of Foxp3 and repressed the IL-17 expression in the CD4(+) T cells in mice. ATRA suppressed the production of total IgG and IgG2a in splenocytes that were stimulated by LPS. It also reduced serum levels of total IgG and IgG2 anti-collagen Abs and germinal center formation in CIA mice. In addition, the ATRA-treated mice showed decreased osteoclast formation in arthritic joints. Moreover, ATRA downregulated the expression of receptor activator of NF-κB ligand, the leading player of osteoclastogenesis, in the CD4(+) T cells and fibroblast-like synoviocytes from patients with RA. Furthermore, ATRA prevented both human monocytes and mice bone marrow-derived monocytes/macrophage cells from differentiating into osteoclasts. These data suggest ATRA might be an effective treatment modality for RA patients.     

Th17 cells, not IL-17+ γδ T cells, drive arthritic bone destruction in mice and humans

The mechanism whereby IL-17 drives rheumatoid arthritis remains incompletely understood. We demonstrate that anti-IL-17 therapy in collagen-induced arthritis ameliorates bone damage by reducing the number of osteoclasts in joints. We found equal numbers of CD4(+) Th17 and IL-17 producing γδ T cells in the joints of arthritic mice, and in vitro, both populations similarly induced osteoclastogenesis. However, individual depletion and adoptive transfer studies revealed that in vivo, Th17 cells dominated with regard to bone destruction. Unlike γδ T cells, Th17 cells were found in apposition to tartrate-resistant acid phosphatase positive osteoclasts in subchondral areas of inflamed joints, a pattern reproduced in patient biopsies. This localization was caused by Ag-specific retention, because OVA-primed Th17 cells showed a γδ T cell-like diffuse distribution. Because IL-23, as produced by osteoclasts, enhanced T cell-mediated osteoclastogenesis, we propose that Ag-specific juxtaposition is key to foster the molecular cross talk of Th17 cells and osteoclasts, thus driving arthritic bone destruction.     

IL-17 receptor knockout mice have enhanced myelotoxicity and impaired hemopoietic recovery following gamma irradiation

IL-17A is a T cell-derived proinflammatory cytokine required for microbial host defense. In vivo expression profoundly stimulates granulopoiesis. At baseline, the hemopoietic system of IL-17R knockout mice (IL-17Ra(-/-)) is, with the exception of increased splenic progenitor numbers, indistinguishable from normal control mice. However, when challenged with gamma irradiation, hemopoietic toxicity is significantly more pronounced in IL-17Ra(-/-) animals, with the gamma irradiation-associated LD(50) being reduced by 150 rad. In spleen-derived T cells, gamma irradiation induces significant murine IL-17A expression in vivo but not in vitro. After sublethal radiation injury (500 rad), the infusion of purified CD4(+) T cells enhances hemopoietic recovery. This recovery is significantly impaired in IL-17Ra(-/-) animals or after in vivo blockade of IL-17Ra in normal mice, resulting in a reduction of hemopoietic precursors by 50% and of neutrophils by 43%. Following sublethal radiation-induced myelosuppression, in vivo overexpression of murine IL-17A in normal mice substantially enhanced granulopoietic restoration in mice with a 4-fold increase in neutrophils and splenic precursors on day 8 (CFU-granulocyte-macrophage/granulocyte-erythrocyte-megakaryocyte-monocyte, CFU-high proliferative potential), as well as 2- and 3-fold increases of bone marrow precursors, respectively. This establishes IL-17A as a hemopoietic response cytokine to radiation injury in mice and an inducible mechanism that is required for recovery of granulopoiesis after radiation injury.     

The myeloid differentiation factor 88 (MyD88) is required for CD4+ T cell effector function in a murine model of inflammatory bowel disease

Abnormal T cell responses to commensal bacteria are involved in the pathogenesis of inflammatory bowel disease. MyD88 is an essential signal transducer for TLRs in response to the microflora. We hypothesized that TLR signaling via MyD88 was important for effector T cell responses in the intestine. TLR expression on murine T cells was examined by flow cytometry. CD4(+)CD45Rb(high) T cells and/or CD4(+)CD45Rb(low)CD25(+) regulatory T cells were isolated and adoptively transferred to RAG1(-/-) mice. Colitis was assessed by changes in body weight and histology score. Cytokine production was assessed by ELISA. In vitro proliferation of T cells was assessed by [(3)H]thymidine assay. In vivo proliferation of T cells was assessed by BrdU and CFSE labeling. CD4(+)CD45Rb(high) T cells expressed TLR2, TLR4, TLR9, and TLR3, and TLR ligands could act as costimulatory molecules. MyD88(-/-) CD4(+) T cells showed decreased proliferation compared with WT CD4(+) T cells both in vivo and in vitro. CD4(+)CD45Rb(high) T cells from MyD88(-/-) mice did not induce wasting disease when transferred into RAG1(-/-) recipients. Lamina propria CD4(+) T cell expression of IL-2 and IL-17 and colonic expression of IL-6 and IL-23 were significantly lower in mice receiving MyD88(-/-) cells than mice receiving WT cells. In vitro, MyD88(-/-) T cells were blunted in their ability to secrete IL-17 but not IFN-gamma. Absence of MyD88 in CD4(+)CD45Rb(high) cells results in defective T cell function, especially Th17 differentiation. These results suggest a role for TLR signaling by T cells in the development of inflammatory bowel disease.     

Tumour cell generation of inducible regulatory T-cells in multiple myeloma is contact-dependent and antigen-presenting cell-independent

Regulatory T-cells (T(Reg) cells) are increased in patients with multiple myeloma (MM). We investigated whether MM cells could generate and/or expand T(Reg) cells as a method of immuno-surveillance avoidance. In an in vitro model, CD4(+)CD25(-)FoxP3(-) T-cells co-cultured with malignant plasma cells (primary MM cells and cell lines) induced a significant generation of CD4(+)CD25(+)FoxP3(+) inducible T(Reg) cells (tT(Reg) cells; p<0.0001), in a contact-dependent manner. tT(Reg) cells were polyclonal, demonstrated a suppressive phenotype and phenotypically, demonstrated increased FoxP3 (p = 0.0001), increased GITR (p<0.0001), increased PD1 (p = 0.003) and decreased CD62L (p = 0.007) expression compared with naturally occurring T(Reg) cells. FACS-sorted tT(Reg) cells differentiated into FoxP(+)IL-17(+) and FoxP3(-)IL-17(+) CD4(+) cells upon TCR-mediated stimulation. Blocking experiments with anti-ICOS-L MoAb resulted in a significant inhibition of tT(Reg) cell generation whereas both IL-10 & TGFβ blockade did not. MM tumour cells can directly generate functional T(Reg) cells in a contact-dependent manner, mediated by ICOS/ICOS-L. These features suggest that tumour generation of T(Reg) cells may contribute to evasion of immune surveillance by the host.     

Th17 cells expressing KIR3DL2+ and responsive to HLA-B27 homodimers are increased in ankylosing spondylitis

CD4 Th cells producing the proinflammatory cytokine IL-17 (Th17) have been implicated in a number of inflammatory arthritides including the spondyloarthritides. Th17 development is promoted by IL-23. Ankylosing spondylitis, the most common spondyloarthritis (SpA), is genetically associated with both HLA-B27 (B27) and IL-23R polymorphisms; however, the link remains unexplained. We have previously shown that B27 can form H chain dimers (termed B27(2)), which, unlike classical HLA-B27, bind the killer-cell Ig-like receptor KIR3DL2. In this article, we show that B27(2)-expressing APCs stimulate the survival, proliferation, and IL-17 production of KIR3DL2(+) CD4 T cells. KIR3DL2(+) CD4 T cells are expanded and enriched for IL-17 production in the blood and synovial fluid of patients with SpA. Despite KIR3DL2(+) cells comprising a mean of just 15% of CD4 T in the peripheral blood of SpA patients, this subset accounted for 70% of the observed increase in Th17 numbers in SpA patients compared with control subjects. TCR-stimulated peripheral blood KIR3DL2(+) CD4 T cell lines from SpA patients secreted 4-fold more IL-17 than KIR3DL2(+) lines from controls or KIR3DL2(-) CD4 T cells. Strikingly, KIR3DL2(+) CD4 T cells account for the majority of peripheral blood CD4 T cell IL-23R expression and produce more IL-17 in the presence of IL-23. Our findings link HLA-B27 with IL-17 production and suggest new therapeutic strategies in ankylosing spondylitis/SpA.     

Suppression of osteoprotegerin expression by prostaglandin E2 is crucially involved in lipopolysaccharide-induced osteoclast formation

LPS is a potent stimulator of bone resorption in inflammatory diseases. The mechanism by which LPS induces osteoclastogenesis was studied in cocultures of mouse osteoblasts and bone marrow cells. LPS stimulated osteoclast formation and PGE(2) production in cocultures of mouse osteoblasts and bone marrow cells, and the stimulation was completely inhibited by NS398, a cyclooxygenase-2 inhibitor. Osteoblasts, but not bone marrow cells, produced PGE(2) in response to LPS. LPS-induced osteoclast formation was also inhibited by osteoprotegerin (OPG), a decoy receptor of receptor activator of NF-kappaB ligand (RANKL), but not by anti-mouse TNFR1 Ab or IL-1 receptor antagonist. LPS induced both stimulation of RANKL mRNA expression and inhibition of OPG mRNA expression in osteoblasts. NS398 blocked LPS-induced down-regulation of OPG mRNA expression, but not LPS-induced up-regulation of RANKL mRNA expression, suggesting that down-regulation of OPG expression by PGE(2) is involved in LPS-induced osteoclast formation in the cocultures. NS398 failed to inhibit LPS-induced osteoclastogenesis in cocultures containing OPG knockout mouse-derived osteoblasts. IL-1 also stimulated PGE(2) production in osteoblasts and osteoclast formation in the cocultures, and the stimulation was inhibited by NS398. As seen with LPS, NS398 failed to inhibit IL-1-induced osteoclast formation in cocultures with OPG-deficient osteoblasts. These results suggest that IL-1 as well as LPS stimulates osteoclastogenesis through two parallel events: direct enhancement of RANKL expression and suppression of OPG expression, which is mediated by PGE(2) production.