Leonard R. Pelgrom

Intestinal dendritic cells (DCs) play a central role in maintaining gut tolerance through priming of peripheral regulatory T cells (pT _reg cells). DC tolerogenicity has been linked to catabolic metabolism, but the role of AMP-activated kinase (AMPK), a key regulator of catabolic metabolism, in regulating intestinal tolerance remains unclear. We found high AMPK activation in intestinal DCs, and loss of AMPKα1 in CD11c-expressing cells (CD11c ^ΔAMPKα1 ) led to reduced frequencies of intestinal RALDH ⁺ CD103 ⁺ cDC2s. This was associated with reduced pT _reg cell induction and consequently increased type 2 immunity in models of intestinal helminth infection. Correspondingly, CD103 ⁺ cDC2s from helminth-infected CD11c ^ΔAMPKα1 mice failed to prime T _reg cells ex vivo. Similarly, T _reg cell induction by AMPK-deficient human retinoic acid (RA)–induced tolerogenic CD103 ⁺ DCs was compromised. Mechanistically, AMPK underpinned RA-driven tolerogenicity by promoting RALDH activity and TGF-β secretion in a FoxO3-dependent manner, independent from metabolic reprogramming. Our findings identify AMPK as a key regulator of intestinal DC-mediated tolerance and as a therapeutic target to counter intestinal inflammatory disease.

Obesity-associated metabolic inflammation drives the development of insulin resistance and type 2 diabetes, notably through modulating innate and adaptive immune cells in metabolic organs. The nutrient sensor liver kinase B1 (LKB1) has recently been shown to control cellular metabolism and T cell priming functions of DCs. Here, we report that hepatic DCs from high-fat diet–fed (HFD-fed) obese mice display increased LKB1 phosphorylation and that LKB1 deficiency in DCs (CD11c ^ΔLKB1 ) worsened HFD-driven hepatic steatosis and impaired glucose homeostasis. Loss of LKB1 in DCs was associated with increased expression of Th17-polarizing cytokines and accumulation of hepatic IL-17A ⁺ Th cells in HFD-fed mice. Importantly, IL-17A neutralization rescued metabolic perturbations in HFD-fed CD11c ^ΔLKB1 mice. Mechanistically, deficiency of the canonical LKB1 target AMPK in HFD-fed CD11c ^ΔAMPKα1 mice recapitulated neither the hepatic Th17 phenotype nor the disrupted metabolic homeostasis, suggesting the involvement of other and/or additional LKB1 downstream effectors. We indeed provide evidence that the control of Th17 responses by DCs via LKB1 is actually dependent on both AMPKα1 and salt-inducible kinase signaling. Altogether, our data reveal a key role for LKB1 signaling in DCs in protection against obesity-induced metabolic dysfunctions by limiting hepatic Th17 responses.

Pro-inflammatory activation of macrophages in metabolic tissues is critically important in induction of obesity-induced metaflammation. Here, we demonstrate that the soluble mannose receptor (sMR) plays a direct, functional role in both macrophage activation and metaflammation. We show that sMR binds CD45 on macrophages and inhibits its phosphatase activity, leading to a Src/Akt/NF-κB-mediated cellular reprogramming towards an inflammatory phenotype both in vitro and in vivo. Remarkably, increased serum sMR levels were observed in obese mice and humans and directly correlated with body weight. Additionally, MR deficiency lowers pro-inflammatory macrophages in metabolic tissues and protects against hepatic steatosis and whole-body metabolic dysfunctions in high-fat diet-induced obese mice. Conversely, administration of sMR in lean mice increases serum pro-inflammatory cytokines, activates tissue macrophages and promotes insulin resistance. Altogether, our results reveal sMR as novel regulator of pro-inflammatory macrophage activation which could constitute a new therapeutic target for metaflammation and other hyperinflammatory diseases.

Type 2 immunity plays an essential role in the maintenance of metabolic homeostasis and its disruption during obesity promotes meta-inflammation and insulin resistance. Infection with the helminth parasite Schistosoma mansoni and treatment with its soluble egg antigens (SEA) can induce a type 2 immune response in metabolic organs and improve insulin sensitivity and glucose tolerance in obese mice, yet a causal relationship remains unproven. Here, we investigated the effects and underlying mechanisms of the T2 ribonuclease omega-1 (ω1), one of the major S. mansoni immunomodulatory glycoproteins, on metabolic homeostasis. Male C57Bl6/J mice were fed a high-fat diet for 12 weeks followed by bi-weekly injection of SEA, ω1 or vehicle for 4 additional weeks. Whole-body metabolic homeostasis and energy expenditure were assessed by glucose/insulin tolerance tests and indirect calorimetry, respectively. Tissue-specific immune cell phenotypes were determined by flow cytometry. We show that treatment of obese mice with plant-produced recombinant ω1, harboring similar glycan motifs as present on the native molecule, decreased body fat mass and improved systemic insulin sensitivity and glucose tolerance in a time-and dose-dependent manner. This effect was associated with an increase in white adipose tissue (WAT) type 2 T helper cells, eosinophils and alternatively-activated macrophages, without affecting type 2 innate lymphoid cells. In contrast to SEA, the metabolic effects of ω1 were still observed in obese STAT6-deficient mice with impaired type 2 immunity, indicating that its metabolic effects are independent of the type 2 immune response. Instead, we found that ω1 inhibited food intake, without affecting locomotor activity, WAT thermogenic capacity or whole-body energy expenditure, an effect also occurring in leptin receptor-deficient obese and hyperphagic db/db mice. Altogether, we demonstrate that while the helminth glycoprotein ω1 can induce type 2 immunity, it improves whole-body metabolic homeostasis in obese mice by inhibiting food intake via a STAT6-independent mechanism. Author summary: The obesity-induced chronic low-grade inflammation, notably in adipose tissue, contributes to insulin resistance and increased risk of type 2 diabetes. We have previously shown that infection with parasitic helminth worms was associated with protection against obesity-related metabolic dysfunctions both in mice and humans. We have also reported that treatment of obese mice with an extract of Schistosoma mansoni eggs (SEA) improves insulin sensitivity and glucose tolerance, a beneficial effect that was associated with a helminth-specific type 2 immune response in metabolic organs. Here, we studied the effects of omega-1 (ω1), a single immunomodulatory molecule from SEA, on metabolic health in obese mice, and investigated the role of the host immune response elicited. We found that ω1 induced a helminth-characteristic type 2 immune response in adipose tissue and improved both insulin sensitivity and glucose tolerance in obese mice. Yet, in contrast to SEA, ω1’s immunomodulatory properties were dispensable for its metabolic effects. Instead, we show that ω1 inhibited food intake, a feature accounting for most of the improvements in metabolic health. Together, our findings indicate that helminth molecules may improve metabolic health through multiple distinct mechanisms, and further characterization of such molecules could lead to new therapeutic strategies to combat obesity.

The molecular mechanisms through which dendritic cells (DCs) prime T helper 2 (Th2) responses, including those elicited by parasitic helminths, remain incompletely understood. Here, we report that soluble egg antigen (SEA) from Schistosoma mansoni, which is well known to drive potent Th2 responses, triggers DCs to produce prostaglandin E2 (PGE₂), which subsequently—in an autocrine manner—induces OX40 ligand (OX40L) expression to license these DCs to drive Th2 responses. Mechanistically, SEA was found to promote PGE₂ synthesis through Dectin-1 and Dectin-2, and via a downstream signaling cascade involving spleen tyrosine kinase (Syk), extracellular signal-regulated kinase (ERK), cytosolic phospholipase A₂ (cPLA₂), and cyclooxygenase 1 and 2 (COX-1 and COX-2). In addition, this pathway was activated independently of the actions of omega-1 (ω-1), a previously described Th2-priming glycoprotein present in SEA. These findings were supported by in vivo murine data showing that ω-1–independent Th2 priming by SEA was mediated by Dectin-2 and Syk signaling in DCs. Finally, we found that Dectin-2^−/−, and to a lesser extent Dectin-1^−/− mice, displayed impaired Th2 responses and reduced egg-driven granuloma formation following S. mansoni infection, highlighting the physiological importance of this pathway in Th2 polarization during a helminth infection. In summary, we identified a novel pathway in DCs involving Dectin-1/2-Syk-PGE₂-OX40L through which Th2 immune responses are induced. T helper 2 (Th2) responses, which are initiated by dendritic cells (DCs), can cause allergic diseases, but they can also provide protection against metabolic disorders and parasitic helminth infections. As such, there is great interest in better understanding how their activity is induced and regulated by DCs. Parasitic helminths can potently induce Th2 responses. However, how helminths condition DCs for priming of Th2 responses remains incompletely understood. Here, we find that egg antigens from the parasitic helminth Schistosoma mansoni bind to pattern-recognition receptors (PRRs) Dectin-1 and Dectin-2 on DCs. This binding triggers a signaling cascade in DCs that results in synthesis of eicosanoid prostaglandin E2 (PGE₂). PGE₂ is sensed by the DCs themselves, resulting in expression of OX40 ligand (OX40L), which subsequently enables the DCs to promote Th2 differentiation. We show that this pathway is activated independently of omega-1 (ω-1), which is a glycoprotein secreted by the eggs and previously shown to condition DCs for priming of Th2 responses. Moreover, we demonstrate that this ω-1–independent pathway is crucial for Th2 induction and egg-driven immunopathology following S. mansoni infection in vivo. In summary, we identified a novel pathway in DCs involving Dectin-1/2–induced autocrine PGE₂ signaling through which Th2 responses are induced.

Recently it has become clear that short chain fatty acids (SCFAs), and in particular butyrate, have anti-inflammatory properties. Murine studies have shown that butyrate can promote regulatory T cells (Tregs) via the induction of tolerogenic dendritic cells (DCs). However, the effects of SCFAs on human DCs and how they affect their capacity to prime and polarize T cell responses has not been addressed. Here we report that butyrate suppresses LPS-induced maturation and metabolic reprogramming of human monocyte-derived DCs (moDCs) and conditions them to polarize naive CD4+ T cells towards IL-10-producing type 1 regulatory T cells (Tr1). This effect was dependent on induction of the retinoic acid-producing enzyme retinaldehyde dehydrogenase 1 (RALDH1) in DCs. The induction of RALDH activity and Tr1 cell differentiation by butyrate was dependent on simultaneous inhibition of histone deacetylases (HDACs) and signaling through G protein-coupled receptor (GPR)109A. Taken together, we reveal that butyrate is a potent inducer of tolerogenic human DCs, thereby shedding new light on the cellular and molecular mechanisms through which SCFAs can exert their immunomodulatory effects in humans.

Type 2 immune responses play key roles in protection against parasitic worm infections, whole‐body metabolic homeostasis, wound healing, and the development of allergies. As a result, there is considerable interest in understanding the pathways that regulate type 2 immunity in order to identify strategies of targeting and controlling these responses. In recent years, it has become increasingly clear that the functional properties of immune cells, including those involved in type 2 immune responses, are dependent on the engagement of specific metabolic pathways such as aerobic glycolysis and fatty acid oxidation (FAO). We here discuss the latest insights in the metabolic regulation of immune cells that initiate type 2 immune responses, such as dendritic cells and innate lymphoid cells, as well as immune cells involved in the effector phase, like T helper 2 (Th2) cells, B cells and alternatively activated macrophages (M2 macrophages). Finally, we consider whether these findings may provide new prospects for the treatment of type 2 immune response‐associated diseases.

Engagement of Toll-like receptors (TLRs) on dendritic cells (DCs) triggers the expression of a large set of genes involved in DC activation and maturation, which allow them to act efficiently as antigen-presenting cells. Recently, it has become clear that TLR signalling in DCs also results in dramatic metabolic changes that are integral to their changed biology. Here, we describe a detailed protocol on how DC metabolism can be studied after TLR stimulation using the 96-well format Extracellular Flux (XF^e96) Analyzer from Seahorse Bioscience, a machine that allows one to simultaneously assess rates of oxidative phosphorylation and glycolysis in real-time, in live cells and in a high-throughput manner.

The endocannabinoid system is an important player in energy metabolism by regulating appetite, lipolysis, and energy expenditure. Chronic blockade of the cannabinoid 1 receptor (CB1R) leads to long‐term maintenance of weight loss and reduction of dyslipidemia in experimental and human obesity. The molecular mechanism by which CB1R blockade reverses dyslipidemia in obesity has not yet been clarified. In this study, we showed that CB1R blockade with the systemic CB1R blocker rimonabant enhanced whole‐body energy expenditure and activated brown adipose tissue (BAT), indicated by increased expression of genes involved in BAT thermogenesis and decreased lipid droplet size in BAT. This was accompanied by selectively increased triglyceride (TG) uptake by BAT and lower plasma TG levels. Interestingly, the effects on BAT activation were still present at thermoneutrality and could be recapitulated by using the strictly peripheral CB1R antagonist AM6545, indicating direct peripheral activation of BAT. Indeed, CB1R blockade directly activated T37i brown adipocytes, resulting in enhanced uncoupled respiration, most likely via enhancing cAMP/PKA signaling via the adrenergic receptor pathway. Our data indicate that selective targeting of the peripheral CB1R in BAT has therapeutic potential in attenuating dyslipidemia and obesity.—Boon, M. R., Kooijman, S., van Dam, A. D., Pelgrom, L. R., Berbée, J. F. P., Visseren, C. A. R., van Aggele, R. C., van den Hoek, A. M., Sips, H. C. M., Lombès, M., Havekes, L. M., Tamsma, J. T., Guigas, B., Meijer, O. C., Jukema, J. W., Rensen, P. C. N., Peripheral cannabinoid 1 receptor blockade activates brown adipose tissue and diminishes dyslipidemia and obesity. FASEB J. 28, 5361–5375 (2014). www.fasebj.org