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BACKGROUND: Innate lymphoid cells (ILCs) are the most recently identified leukocytes of the immune system and these cells are increasingly acknowledged to play important roles in host defence and tissue repair. ILCs are also contributors of inflammatory diseases such as asthma and colitis. We analyzed the presence and relative proportions of the different ILC subsets (ILC1, ILC2 and ILC3) in gingivitis and periodontitis. Further, we investigated if ILCs express receptor activator of nuclear factor kappa-B ligand (RANKL), a cytokine crucial for osteoclast differentiation and bone resorption.

METHODS: We collected gingivitis and periodontitis soft tissue and characterized ILC subsets including RANKL expression in single-cell suspensions using flow cytometry.

RESULTS: ILCs were detected both in gingivitis and periodontitis. The majority of ILCs, in both conditions, were ILC1s. Furthermore, RANKL expression was detected on a fraction of the ILC1s.

CONCLUSIONS: Our discovery of the presence of ILCs both in gingivitis and periodontitis and concomitant expression of RANKL on a fraction of the ILC1 population suggest that these cells may be of importance in periodontal disease. In addition, our findings provide a new insight into the field of oral immunology.

Objective: To investigate whether periodontitis, characterized by marginal jawbone loss, precedes the onset of symptoms of rheumatoid arthritis (RA), and to analyze plasma levels of RANKL (a cytokine that is crucial for bone resorption) and anti–citrullinated peptide antibodies (ACPAs) in presymptomatic individuals compared with matched referent controls.

Methods: Marginal jawbone loss was measured on dental radiographs of the premolar/molar regions in the jaws in 176 subjects, 93 of whom subsequently developed RA. Among these participating subjects, 46 had documented radiographs predating symptom onset, and 45 cases could be matched to controls, according to sex, age, and smoking status. Plasma RANKL concentrations were analyzed using enzyme‐linked immunosorbent assay. A receiver operating characteristic curve was used to define the cutoff value for RANKL positivity.

Results: Bone loss was significantly greater in presymptomatic subjects classified as never smokers compared with that in controls, and increasing levels of bone loss were associated with a higher risk of the subsequent development of RA (hazard ratio 1.03, 95% confidence interval 1.01–1.05). No association between jawbone loss and RA was observed in smokers. A significantly greater extent of marginal jawbone loss was detected in RANKL‐positive presymptomatic subjects, and even more pronounced jawbone loss was observed in those who were positive for both RANKL and ACPA.

Conclusion: Marginal jawbone loss preceded the clinical onset of RA symptoms, but this was observed only in nonsmokers. Moreover, marginal jawbone loss was significantly greater in RANKL‐positive presymptomatic subjects compared with RANKL‐negative presymptomatic subjects and was highest in presymptomatic subjects positive for both ACPA and RANKL.

Background/Purpose: Previous studies have shown a higher incidence of alveolar bone loss in patients with rheumatoid arthritis (RA) and that patients with periodontitis are at a greater risk for developing RA. Periodontitis, displayed as marginal jawbone loss was analysed in individuals prior to symptom onset of RA and related to plasma levels of receptor activator of nuclear factor kappa-B (RANKL), a cytokine crucial for bone resorption. Methods: A case-control study performed within the Medical Biobank of Northern Sweden included 232 pre-symptomatic individuals with blood samples donated before symptom onset and 194 controls. A questionnaire on self-assed dental status and smoking status was retrieved. Dental radiographs to evaluate marginal jawbone levels were available from 93 pre-symptomatic individuals (mean age; 56.8 95%CI55.9, 57.7 years and pre-dating time; -5.3 95%CI -12.2, -0.2, 74.2% females) and 83 controls (mean age; 55.5 95%CI54.6, 56.5, 73.5% females) . Of these individuals 45 had radiograph documentations prior to development of RA symptoms and to whom sex, age and smoking status could be matched among the controls. Plasma were analysed for RANKL (BioVendor, Karasek, Czech Republic), and anti-citrullinated peptide antibodies (ACPA) (anti-CCP2 test, Eurodiagnostics, Sweden) from similar time points. Results: Compared to matched controls, total bone loss was significantly higher in never-smokers who developed RA but not in smokers and increasing levels on total jawbone loss was associated with a significantly higher odds to be diagnosed with RA later (OR=1.06, 95%CI 1.01, 1.11). Regardless of smoking status, the number of unaffected teeth did not differ significantly between those who were subsequently diagnosed with RA and their matched controls. In the pre-symptomatic individuals RANKL positive individuals had significantly higher extent of marginal jawbone loss, which was further increased in ACPA positive individuals. Previously documented association between smoking and ageing and marginal jawbone loss was verified. Conclusion: Marginal jawbone loss preceded onset of symptoms of RA but the difference was only manifested in non-smokers. Moreover, marginal jawbone loss and plasma RANKL levels were related in the pre-symptomatic individuals particularly in ACPA positive individuals.

Periodontitis is a chronic inflammatory disease of tooth supporting tissues resulting in periodontal tissue destruction, which may ultimately lead to tooth loss. The disease is characterized by continuous leukocyte infiltration, likely mediated by local chemokine production but the pathogenic mechanisms are not fully elucidated. There are no reliable serologic biomarkers for the diagnosis of periodontitis, which is today based solely on the degree of local tissue destruction, and there is no available biological treatment tool. Prompted by the increasing interest in periodontitis and systemic inflammatory mediators we mapped serum cytokine and chemokine levels from periodontitis subjects and healthy controls. We used multivariate partial least squares (PLS) modeling and identified monocyte chemoattractant protein-1 (MCP-1) and eotaxin as clearly associated with periodontitis along with C-reactive protein (CRP), years of smoking and age, whereas the number of remaining teeth was associated with being healthy. Moreover, body mass index correlated significantly with serum MCP-1 and CRP, but not with eotaxin. We detected higher MCP-1 protein levels in inflamed gingival connective tissue compared to healthy but the eotaxin levels were undetectable. Primary human gingival fibroblasts displayed strongly increased expression of MCP-1 and eotaxin mRNA and protein when challenged with tumor necrosis factor-alpha (TNF-alpha and interleukin-1 beta (IL-1 beta), key mediators of periodontal inflammation. We also demonstrated that the upregulated chemokine expression was dependent on the NF-kappa B pathway. In summary, we identify higher levels of CRP, eotaxin and MCP-1 in serum of periodontitis patients. This, together with our finding that both CRP and MCP-1 correlates with BMI points towards an increased systemic inflammatory load in patients with periodontitis and high BMI. Targeting eotaxin and MCP-1 in periodontitis may result in reduced leukocyte infiltration and inflammation in periodontitis and maybe prevent tooth loss.

Cytokines produced by inflammatory or resident mesenchymal cells play important modulatory roles in the pathogenesis of inflammation induced bone loss. In the present study, the effects of IL-4 and IL-13 on the expression of three osteotropic cytokines in the IL-6 family expressed in human gingival fibroblasts were studied. IL-4R alpha and IL-13R alpha 1 mRNA were constitutively expressed in human gingival fibroblasts. The inflammatory cytokines IL-1 beta and TNF-alpha increased expression of IL-5, LIF, and IL-11 mRNA and protein in the gingival fibroblasts. Addition of IL-4 or IL-13 had no effect on IL-6 expression, but significantly inhibited LIF and IL-11 mRNA and protein stimulated by IL-1 beta and TNF-alpha. No involvement of NF-kappa B or STAT1 was observed in the inhibition. STAT6 was phosphorylated at Y641 by treatment with IL-4 and knockdown of STAT6 with siRNA decreased the inhibition of IL-11 and LIF expression by IL-4 in IL-1 beta and TNF-alpha stimulated cells. This study suggests that activation of STAT6 by IL-4 and IL-13, through type 2 IL-4 receptors, inhibits production of IL-11 and LIF stimulated by IL-1 beta and TNF-alpha in human gingival fibroblasts. A negative modulatory role of IL-4 and IL-13 in osteotropic cytokine production could be a mechanism playing an important inhibitory role in inflammation induced periodontitis. (C) 2011 Elsevier Ltd. All rights reserved.

The osteotropic interleukin-6 (IL-6) types of cytokines IL-6, IL-11, and leukemia inhibitory factor (LIF), but not oncostatin M, are expressed by human gingival fibroblasts, and their expressions are regulated by IL-1 and tumor necrosis factor-alpha (TNF-alpha). In the present study, we investigated whether signaling through Toll-like receptor 2 (TLR2) can affect the expression of these cytokines in human gingival fibroblasts. Lipopolysac-charide (LPS) from P. gingivalis was found to stimulate IL-6 and LIF mRNA and protein, but not IL-11 or OSM mRNA. Using two synthetic ligands acting specifically at TLR2 and siRNA knockdown of TLR2, we demonstrated the important role of TLR2 in the stimulation of IL-6 and LIF in gingival fibroblasts. Analysis of these data suggests that signaling through the innate immune system controls the expression of osteotropic cytokines in human gingival fibroblasts.

Interleukin-6 (IL-6)-type cytokines are pleiotropic molecules capable of stimulating bone resorption and expressed by numerous cell types. In the present study, we tested the hypothesis that gingival fibroblasts may exert local osteotropic effects through production of IL-6 and related cytokines. IL-6-type cytokine expression and regulation by IL-1beta and tumor necrosis factor-alpha (TNF-alpha) were studied in fibroblasts from the non-inflamed gingiva of healthy individuals. Constitutive mRNA expression of IL-6, IL-11, and leukemia inhibitory factor (LIF), but not of oncostatin M (OSM), was demonstrated, as was concentration-dependent stimulation of IL-6 and LIF mRNA and of protein by IL-1beta and TNF-alpha. IL-11 mRNA and protein were concentration-dependently stimulated by IL-1beta. The signaling pathway involved in IL-6 and LIF mRNA stimulation involved MAP kinases, but not NF-kappaB. The findings support the view that resident cells may influence the pathogenesis of periodontal disease through osteotropic IL-6-type cytokine production mediated by activation of MAP kinases. Abbreviations: IL-1alpha (interleukin-1alpha); IL-1beta (interleukin-1beta); IL-6 (interleukin-6); IL-11 (interleukin-11); LIF (leukemia inhibitory factor); OSM (oncostatin M); alpha(1)-coll. I (alpha(1)-collagen I); ALP (alkaline phosphatase); BMP-2 (bone morphogenetic protein-2); OC (osteocalcin); BSP (bone sialoprotein); TNFR I (tumor necrosis factor receptor I); TNFR II (tumor necrosis factor receptor II); IL-1R1 (interleukin-1 receptor 1); GAPDH (glyceraldehyde-3-phosphate dehydrogenase); RPL13A (ribosomal protein L13A); mRNA (messenger ribonucleic acid); cDNA (complementary deoxyribonucleic acid); PCR (polymerase chain-reaction); BCA (bicinchoninic acid); ELISA (enzyme-linked immunosorbent assay); alpha-MEM (alpha modification of Minimum Essential Medium); and FCS (fetal calf serum).

Hur ombyggnaden av benvävnaden i käkar och skelett fungerar och hur processen påverkas vid sjukdomstillstånd som parodontit, benskörhet, tumörer med mera är föremål för omfattande forskning i Umeå.

Interleukin (IL)-4 and IL-13 are cytokines that inhibit bone resorption. Data showing an inhibitory effect of IL-4 and IL-13 on RANK mRNA in mouse calvariae were first reported at the 22nd American Society for Bone and Mineral Research Meeting (Lerner, U.H., and Conaway, H. H. 2000) J. Bone Min. Res. 15, Suppl. 1, Abstr. SU 230). In the present study, release of 45Ca from cultured mouse calvarial bones stimulated by different cytokines, peptides, and steroid hormones was inhibited by IL-4 and IL-13. IL-4 and IL-13 decreased receptor activator of nuclear factor-kappaB ligand (RANKL) and RANK mRNA and increased osteoprotegerin (OPG) mRNA in calvariae. Additionally, the cytokines decreased RANKL protein and increased OPG protein in calvarial bones. In osteoblasts isolated from calvariae, both an increase in RANKL mRNA and a decrease in OPG mRNA and protein elicited by vitamin D3 were reversed by IL-4 and IL-13. IL-4 and IL-13 decreased the number of tartrate-resistant acid phosphatase positive multinucleated cells and the mRNA expression of calcitonin receptor, tartrate-resistant acid phosphatase, and cathepsin K in mouse spleen cells and bone marrow macrophages (BMM) treated with macrophage colony-stimulating factor and RANKL. Inhibition of mRNA for RANK and the transcription factor NFAT2 was also noted in spleen cell and BMM cultures treated with IL-4 and IL-13. In addition, RANK mRNA and RANK protein were decreased by IL-4 and IL-13 in RAW 264.7 cells. Osteoblasts, spleen cells, and BMM expressed mRNA for the four proteins making up the IL-4 and IL-13 receptors. No effects by IL-4 on bone resorption and osteoclast formation or on RANKL and RANK mRNA expression were seen in Stat6-/- mice. The data indicate that IL-4 and IL-13, via a STAT6-dependent pathway, inhibit osteoclast differentiation and bone resorption by activating receptors on osteoblasts and osteoclasts that affect the RANKL/RANK/OPG system.