Thomas E. Van Dyke

Resolving inflammation: dual anti-inflammatory and pro-resolution lipid mediators

Active resolution of acute inflammation is a previously unrecognized interface between innate and adaptive immunity. Once thought to be a passive process, the resolution of inflammation is now shown to involve active biochemical programmes that enable inflamed tissues to return to homeostasis. This Review presents new cellular and molecular mechanisms for the resolution of inflammation, revealing key roles for eicosanoids, such as lipoxins, and recently discovered families of endogenous chemical mediators, termed resolvins and protectins. These mediators have anti-inflammatory and pro-resolution properties, thereby protecting organs from collateral damage, stimulating the clearance of inflammatory debris and promoting mucosal antimicrobial defence.

Cutting Edge: Different Toll-Like Receptor Agonists Instruct Dendritic Cells to Induce Distinct Th Responses via Differential Modulation of Extracellular Signal-Regulated Kinase-Mitogen-Activated Protein Kinase and c-Fos

Dendritic cells (DCs) are pivotal in determining the class of an adaptive immune response. However, the molecular mechanisms within DCs that determine this decision-making process are unknown. Here, we demonstrate that distinct Toll-like receptor (TLR) ligands instruct human DCs to induce distinct Th cell responses by differentially modulating mitogen-activated protein kinase signaling. Thus, Escherichia coli LPS and flagellin, which trigger TLR4 and TLR5, respectively, instruct DCs to stimulate Th1 responses via IL-12p70 production, which depends on the phosphorylation of p38 and c-Jun N-terminal kinase 1/2. In contrast, the TLR2 agonist, Pam3cys, and the Th2 stimulus, schistosome egg Ags: 1) barely induce IL-12p70; 2) stimulate sustained duration and magnitude of extracellular signal-regulated kinase 1/2 phosphorylation, which results in stabilization of the transcription factor c-Fos, a suppressor of IL-12; and 3) yield a Th2 bias. Thus, distinct TLR agonists differentially modulate extracellular signal-regulated kinase signaling, c-Fos activity, and cytokine responses in DCs to stimulate different Th responses.

Lipopolysaccharides from Distinct Pathogens Induce Different Classes of Immune Responses In Vivo

The adaptive immune system has evolved distinct responses against different pathogens, but the mechanism(s) by which a particular response is initiated is poorly understood. In this study, we investigated the type of Ag-specific CD4+ Th and CD8+ T cell responses elicited in vivo, in response to soluble OVA, coinjected with LPS from two different pathogens. We used Escherichia coli LPS, which signals through Toll-like receptor 4 (TLR4) and LPS from the oral pathogen Porphyromonas gingivalis, which does not appear to require TLR4 for signaling. Coinjections of E. coli LPS + OVA or P. gingivalis LPS + OVA induced similar clonal expansions of OVA-specific CD4+ and CD8+ T cells, but strikingly different cytokine profiles. E. coli LPS induced a Th1-like response with abundant IFN-γ, but little or no IL-4, IL-13, and IL-5. In contrast, P. gingivalis LPS induced Th and T cell responses characterized by significant levels of IL-13, IL-5, and IL-10, but lower levels of IFN-γ. Consistent with these results, E. coli LPS induced IL-12(p70) in the CD8α+ dendritic cell (DC) subset, while P. gingivalis LPS did not. Both LPS, however, activated the two DC subsets to up-regulate costimulatory molecules and produce IL-6 and TNF-α. Interestingly, these LPS appeared to have differences in their ability to signal through TLR4; proliferation of splenocytes and cytokine secretion by splenocytes or DCs from TLR4-deficient C3H/HeJ mice were greatly impaired in response to E. coli LPS, but not P. gingivalis LPS. Therefore, LPS from different bacteria activate DC subsets to produce different cytokines, and induce distinct types of adaptive immunity in vivo.

Yeast zymosan, a stimulus for TLR2 and dectin-1, induces regulatory antigen-presenting cells and immunological tolerance

Emerging evidence suggests critical roles for APCs in suppressing immune responses. Here, we show that zymosan, a stimulus for TLR2 and dectin-1, regulates cytokine secretion in DCs and macrophages to induce immunological tolerance. First, zymosan induces DCs to secrete abundant IL-10 but little IL-6 and IL-12(p70). Induction of IL-10 is dependent on TLR2- and dectin-1-mediated activation of ERK MAPK via a mechanism independent of the activation protein 1 (AP-1) transcription factor c-Fos. Such DCs stimulate antigen-specific CD4+ T cells poorly due to IL-10 and the lack of IL-6. Second, zymosan induces F4-80+ macrophages in the splenic red pulp to secrete TGF-beta. Consistent with these effects on APCs, injection of zymosan plus OVA into mice results in OVA-specific T cells that secrete little or no Th1 or Th2 cytokines, but secrete robust levels of IL-10, and are unresponsive to challenge with OVA plus adjuvant. Finally, coinjection of zymosan with OVA plus LPS suppresses the response to OVA via a mechanism dependent on IL-10, TGF-beta, and lack of IL-6. Together, our data demonstrate that zymosan stimulates IL-10+ IL-12(p70)- IL-6low regulatory DCs and TGF-beta+ macrophages to induce immunological tolerance. These data suggest several targets for pharmacological modulation of immune responses in various clinical settings.

A Toll-Like Receptor 2 Ligand Stimulates Th2 Responses In Vivo, via Induction of Extracellular Signal-Regulated Kinase Mitogen-Activated Protein Kinase and c-Fos in Dendritic Cells

The adaptive immune system can generate distinct classes of responses, but the mechanisms that determine this are poorly understood. In this study, we demonstrate that different Toll-like receptor (TLR) ligands induce distinct dendritic cell (DC) activation and immune responses in vivo. Thus, Escherichia coli LPS (TLR-4 stimulus), activates DCs to produce abundant IL-12(p70), but little IL-10, and stimulates Th1 and Tc1 responses. In contrast, Pam-3-cys (TLR-2 stimulus) elicits less IL-12(p70), but abundant IL-10, and favors Th2 and T cytotoxic 2 (Tc2) responses. These distinct responses likely occur via differences in extracellular signal-regulated kinase signaling in DCs. Thus, Pam-3-cys induces enhanced extracellular signal-regulated kinase signaling, compared with LPS, resulting in suppressed IL-12(p70) and enhanced IL-10 production, as well as enhanced induction of the transcription factor, c-Fos. Interestingly, DCs from c-fos−/− mice produce more IL-12(p70), but less IL-10, compared with control DCs. Therefore, different TLR ligands induce distinct cytokines and signaling in DCs, and differentially bias Th responses in vivo.

Comparisons of Subgingival Microbial Profiles of Refractory Periodontitis, Severe Periodontitis, and Periodontal Health Using the Human Oral Microbe Identification Microarray

BACKGROUND This study compared the subgingival microbiota of subjects with refractory periodontitis (RP) to those in subjects with treatable periodontitis (GRs = good responders) or periodontal health (PH) using the Human Oral Microbe Identification Microarray (HOMIM). METHODS At baseline, subgingival plaque samples were taken from 47 subjects with periodontitis and 20 individuals with PH and analyzed for the presence of 300 species by HOMIM. The subjects with periodontitis were classified as having RP (n = 17) based on mean attachment loss (AL) and/or more than three sites with AL >or=2.5 mm after scaling and root planing, surgery, and systemically administered amoxicillin and metronidazole or as GRs (n = 30) based on mean attachment gain and no sites with AL >or=2.5 mm after treatment. Significant differences in taxa among the groups were sought using the Kruskal-Wallis and chi(2) tests. RESULTS More species were detected in patients with disease (GR or RP) than in those without disease (PH). Subjects with RP were distinguished from GRs or those with PH by a significantly higher frequency of putative periodontal pathogens, such as Parvimonas micra (previously Peptostreptococcus micros or Micromonas micros), Campylobacter gracilis, Eubacterium nodatum, Selenomonas noxia, Tannerella forsythia (previously T. forsythensis), Porphyromonas gingivalis, Prevotella spp., Treponema spp., and Eikenella corrodens, as well as unusual species (Pseudoramibacter alactolyticus, TM7 spp. oral taxon [OT] 346/356, Bacteroidetes sp. OT 272/274, Solobacterium moorei, Desulfobulbus sp. OT 041, Brevundimonas diminuta, Sphaerocytophaga sp. OT 337, Shuttleworthia satelles, Filifactor alocis, Dialister invisus/pneumosintes, Granulicatella adiacens, Mogibacterium timidum, Veillonella atypica, Mycoplasma salivarium, Synergistes sp. cluster II, and Acidaminococcaceae [G-1] sp. OT 132/150/155/148/135) (P <0.05). Species that were more prevalent in subjects with PH than in patients with periodontitis included Actinomyces sp. OT 170, Actinomyces spp. cluster I, Capnocytophaga sputigena, Cardiobacterium hominis, Haemophilus parainfluenzae, Lautropia mirabilis, Propionibacterium propionicum, Rothia dentocariosa/mucilaginosa, and Streptococcus sanguinis (P <0.05). CONCLUSION As determined by HOMIM, patients with RP presented a distinct microbial profile compared to patients in the GR and PH groups.

Reduced Inflammation and Tissue Damage in Transgenic Rabbits Overexpressing 15-Lipoxygenase and Endogenous Anti-inflammatory Lipid Mediators

PGs and leukotrienes (LTs) mediate cardinal signs of inflammation; hence, their enzymes are targets of current anti-inflammatory therapies. Products of arachidonate 15-lipoxygenases (LO) types I and II display both beneficial roles, such as lipoxins (LXs) that stereoselectively signal counterregulation, as well as potential deleterious actions (i.e., nonspecific phospholipid degradation). In this study, we examined transgenic (TG) rabbits overexpressing 15-LO type I and their response to inflammatory challenge. Skin challenges with either LTB4 or IL-8 showed that 15-LO TG rabbits give markedly reduced neutrophil (PMN) recruitment and plasma leakage at dermal sites with LTB4. PMN from TG rabbits also exhibited a dramatic reduction in LTB4-stimulated granular mobilization that was not evident with peptide chemoattractants. Leukocytes from 15-LO TG rabbits gave enhanced LX production, underscoring differences in lipid mediator profiles compared with non-TG rabbits. Microbe-associated inflammation and leukocyte-mediated bone destruction were assessed by initiating acute periodontitis. 15-LO TG rabbits exhibited markedly reduced bone loss and local inflammation. Because enhanced LX production was associated with an increased anti-inflammatory status of 15-LO TG rabbits, a stable analog of 5S,6R,15S-trihydroxyeicosa-7E,9E,11Z,13E-tetraenoic acid (LXA4) was applied to the gingival crevice subject to periodontitis. Topical application with the 15-epi-16-phenoxy-para-fluoro-LXA4 stable analog (ATLa) dramatically reduced leukocyte infiltration, ensuing bone loss as well as inflammation. These results indicate that overexpression of 15-LO type I and LXA4 is associated with dampened PMN-mediated tissue degradation and bone loss, suggesting that enhanced anti-inflammation status is an active process. Moreover, they suggest that LXs can be targets for novel approaches to diseases, e.g., periodontitis and arthritis, where inflammation and bone destruction are features.

Periodontitis and atherosclerotic cardiovascular disease: consensus report of the Joint EFP/AAP Workshop on Periodontitis and Systemic Diseases

BACKGROUND This consensus report is concerned with the association between periodontitis and atherosclerotic cardiovascular disease (ACVD). Periodontitis is a chronic multifactorial inflammatory disease caused by microorganisms and characterized by progressive destruction of the tooth supporting apparatus leading to tooth loss; as such, it is a major public health issue. AIMS This report examined biological plausibility, epidemiology and early results from intervention trials. Plausibility: Periodontitis leads to entry of bacteria in the blood stream. The bacteria activate the host inflammatory response by multiple mechanisms. The host immune response favors atheroma formation, maturation and exacerbation. EPIDEMIOLOGY In longitudinal studies assessing incident cardiovascular events, statistically significant excess risk for ACVD was reported in individuals with periodontitis. This was independent of established cardiovascular risk factors. The amount of the adjusted excess risk varies by type of cardiovascular outcome and across populations by age and gender. Given the high prevalence of periodontitis, even low to moderate excess risk is important from a public health perspective. INTERVENTION There is moderate evidence that periodontal treatment: (i) reduces systemic inflammation as evidenced by reduction in C-reactive protein (CRP) and improvement of both clinical and surrogate measures of endothelial function; but (ii) there is no effect on lipid profiles - supporting specificity. Limited evidence shows improvements in coagulation, biomarkers of endothelial cell activation, arterial blood pressure and subclinical atherosclerosis after periodontal therapy. The available evidence is consistent and speaks for a contributory role of periodontitis to ACVD. There are no periodontal intervention studies on primary ACVD prevention and there is only one feasibility study on secondary ACVD prevention. CONCLUSIONS It was concluded that: (i) there is consistent and strong epidemiologic evidence that periodontitis imparts increased risk for future cardiovascular disease; and (ii) while in vitro, animal and clinical studies do support the interaction and biological mechanism, intervention trials to date are not adequate to draw further conclusions. Well-designed intervention trials on the impact of periodontal treatment on prevention of ACVD hard clinical outcomes are needed.BACKGROUND This consensus report is concerned with the association between periodontitis and atherosclerotic cardiovascular disease (ACVD). Periodontitis is a chronic multifactorial inflammatory disease caused by microorganisms and characterized by progressive destruction of the tooth supporting apparatus leading to tooth loss; as such, it is a major public health issue. AIMS This report examined biological plausibility, epidemiology and early results from intervention trials. PLAUSIBILITY: Periodontitis leads to entry of bacteria in the blood stream. The bacteria activate the host inflammatory response by multiple mechanisms. The host immune response favors atheroma formation, maturation and exacerbation. EPIDEMIOLOGY In longitudinal studies assessing incident cardiovascular events, statistically significant excess risk for ACVD was reported in individuals with periodontitis. This was independent of established cardiovascular risk factors. The amount of the adjusted excess risk varies by type of cardiovascular outcome and across populations by age and gender. Given the high prevalence of periodontitis, even low to moderate excess risk is important from a public health perspective. INTERVENTION There is moderate evidence that periodontal treatment: (i) reduces systemic inflammation as evidenced by reduction in C-reactive protein (CRP) and improvement of both clinical and surrogate measures of endothelial function; but (ii) there is no effect on lipid profiles--supporting specificity. Limited evidence shows improvements in coagulation, biomarkers of endothelial cell activation, arterial blood pressure and subclinical atherosclerosis after periodontal therapy. The available evidence is consistent and speaks for a contributory role of periodontitis to ACVD. There are no periodontal intervention studies on primary ACVD prevention and there is only one feasibility study on secondary ACVD prevention. CONCLUSIONS It was concluded that: (i) there is consistent and strong epidemiologic evidence that periodontitis imparts increased risk for future cardiovascular disease; and (ii) while in vitro, animal and clinical studies do support the interaction and biological mechanism, intervention trials to date are not adequate to draw further conclusions. Well-designed intervention trials on the impact of periodontal treatment on prevention of ACVD hard clinical outcomes are needed.

Resolvin E1 Regulates Inflammation at the Cellular and Tissue Level and Restores Tissue Homeostasis In Vivo

Resolvin E1 (RvE1) is a potent proresolving mediator of inflammation derived from omega-3 eicosapentaenoic acid that acts locally to stop leukocyte recruitment and promote resolution. RvE1 displays potent counter-regulatory and tissue-protective actions in vitro and in vivo. Periodontal disease is a local inflammatory disease initiated by bacteria characterized by neutrophil-mediated tissue injury followed by development of a chronic immune lesion. In this study, we report the treatment of established periodontitis using RvE1 as a monotherapy in rabbits compared with structurally related lipids PGE2 and leukotriene B4. PGE2 and leukotriene B4 each enhanced development of periodontitis and worsened the severity of disease. Promotion of resolution of inflammation as a therapeutic target with RvE1 resulted in complete restoration of the local lesion, and reduction in the systemic inflammatory markers C-reactive protein and IL-1β. This report is the first to show that resolution of inflammation by a naturally occurring endogenous lipid mediator results in complete regeneration of pathologically lost tissues, including bone.

Periodontal Disease Is Associated With Brachial Artery Endothelial Dysfunction and Systemic Inflammation

Objective—The purpose of this study was to determine whether periodontal disease is associated with endothelial dysfunction and systemic inflammation. Epidemiological studies suggest that severe periodontal disease is associated with increased cardiovascular disease risk, but the mechanisms remain unknown. Methods and Results—We assessed flow-mediated dilation and nitroglycerin-mediated dilation of the brachial artery using vascular ultrasound in 26 subjects with advanced periodontal disease and 29 control subjects. The groups were matched for age and sex, and patients with hypercholesterolemia, diabetes mellitus, hypertension, and history of cigarette smoking were excluded. We also examined serum levels of C-reactive protein using an established high-sensitivity method. Subjects with advanced periodontal disease had lower flow-mediated dilation compared with control patients (7.8±4.6% versus 11.7±5.3%, P =0.005). Nitroglycerin-mediated dilation was equivalent in the two groups. Subjects with advanced periodontitis exhibited higher serum levels of high-sensitivity C-reactive protein compared with healthy controls patients (2.3±2.3 versus 1.0±1.0 mg/L, P =0.03). Conclusions—Subjects with advanced periodontal disease exhibit endothelial dysfunction and evidence of systemic inflammation, possibly placing them at increased risk for cardiovascular disease.