Simultaneous Increases in Intracellular Sodium and Tonicity Boost Antimicrobial Activity of Macrophages.

Inflamed and infected tissues can display increased local sodium (Na+) levels, which can have various effects on immune cells. In macrophages, high salt (HS) leads to a Na+/Ca2+-exchanger 1 (NCX1)-dependent increase in intracellular Na+ levels. This results in augmented osmoprotective signaling and enhanced proinflammatory activation, such as enhanced expression of type 2 nitric oxide synthase and antimicrobial function. In this study, the role of elevated intracellular Na+ levels in macrophages was investigated. Therefore, the Na+/K+-ATPase (NKA) was pharmacologically inhibited with two cardiac glycosides (CGs), ouabain (OUA) and digoxin (DIG), to raise intracellular Na+ without increasing extracellular Na+ levels. Exposure to HS conditions and treatment with both inhibitors resulted in intracellular Na+ accumulation and subsequent phosphorylation of p38/MAPK. The CGs had different effects on intracellular Ca2+ and K+ compared to HS stimulation. Moreover, the osmoprotective transcription factor nuclear factor of activated T cells 5 (NFAT5) was not upregulated on RNA and protein levels upon OUA and DIG treatment. Accordingly, OUA and DIG did not boost nitric oxide (NO) production and showed heterogeneous effects toward eliminating intracellular bacteria. While HS environments cause hypertonic stress and ionic perturbations, cardiac glycosides only induce the latter. Cotreatment of macrophages with OUA and non-ionic osmolyte mannitol (MAN) partially mimicked the HS-boosted antimicrobial macrophage activity. These findings suggest that intracellular Na+ accumulation and hypertonic stress are required but not sufficient to mimic boosted macrophage function induced by increased extracellular sodium availability.

The influence of negative pressure wound therapy on bacterial and fungal growth.

BACKGROUND: The use of negative pressure wound therapy (NPWT) in superinfected wounds is controversial. The mechanism of action is unclear, but recent studies have shown lower atmospheric oxygen levels within the dressing. Therefore, different oxygen-favoring bacteria and fungi might benefit or face impaired thriving conditions. The aim of this in vitro study is to investigate the influence of NPWT on bacterial and fungal growth. METHODS: Salmonella enterica subsp. enterica serovar Typhimurium, Pseudomonas aeruginosa and Candida albicans strains were cultured on concentrated agars and attached to a standard NPWT-device. After 48 hours, colonies were separately harvested from the agar and foam. Optical density (OD) was obtained in order to estimate bacterial loads. RESULTS: For all tested microorganisms, no overall significant differences were found compared to controls. Subanalysis showed lower OD levels from the agar beneath the foam in the NPWT-group. CONCLUSION: NPWT removed bacteria and fungi from the wound surface but accumulation is found within the foam. The use of NPWT showed no influence on bacterial or fungal growth selection. With superinfected wounds, the use of NPWT should thoroughly be evaluated as toxins and virulence factors may not fully be evacuated.

Macrophages inhibit Coxiella burnetii by the ACOD1-itaconate pathway for containment of Q fever.

Infection with the intracellular bacterium Coxiella (C.) burnetii can cause chronic Q fever with severe complications and limited treatment options. Here, we identify the enzyme cis-aconitate decarboxylase 1 (ACOD1 or IRG1) and its product itaconate as protective host immune pathway in Q fever. Infection of mice with C. burnetii induced expression of several anti-microbial candidate genes, including Acod1. In macrophages, Acod1 was essential for restricting C. burnetii replication, while other antimicrobial pathways were dispensable. Intratracheal or intraperitoneal infection of Acod1-/- mice caused increased C. burnetii burden, weight loss and stronger inflammatory gene expression. Exogenously added itaconate restored pathogen control in Acod1-/- mouse macrophages and blocked replication in human macrophages. In axenic cultures, itaconate directly inhibited growth of C. burnetii. Finally, treatment of infected Acod1-/- mice with itaconate efficiently reduced the tissue pathogen load. Thus, ACOD1-derived itaconate is a key factor in the macrophage-mediated defense against C. burnetii and may be exploited for novel therapeutic approaches in chronic Q fever.

Acidic Microenvironments Found in Cutaneous Leishmania Lesions Curtail NO-Dependent Antiparasitic Macrophage Activity.

Local tissue acidosis affects anti-tumor immunity. In contrast, data on tissue pH levels in infected tissues and their impact on antimicrobial activity is sparse. In this study, we assessed the pH levels in cutaneous Leishmania lesions. Leishmania major-infected skin tissue displayed pH levels of 6.7 indicating that lesional pH is acidic. Next, we tested the effect of low extracellular pH on the ability of macrophages to produce leishmanicidal NO and to fight the protozoan parasite Leishmania major. Extracellular acidification led to a marked decrease in both NO production and leishmanicidal activity of lipopolysaccharide (LPS) and interferon γ (IFN-γ)-coactivated macrophages. This was not directly caused by a disruption of NOS2 expression, a shortage of reducing equivalents (NAPDH) or substrate (L-arginine), but by a direct, pH-mediated inhibition of NOS2 enzyme activity. Normalization of intracellular pH significantly increased NO production and antiparasitic activity of macrophages even in an acidic microenvironment. Overall, these findings indicate that low local tissue pH can curtail NO production and leishmanicidal activity of macrophages.

Phagosomal signalling of the C-type lectin receptor Dectin-1 is terminated by intramembrane proteolysis.

Sensing of pathogens by pattern recognition receptors (PRR) is critical to initiate protective host defence reactions. However, activation of the immune system has to be carefully titrated to avoid tissue damage necessitating mechanisms to control and terminate PRR signalling. Dectin-1 is a PRR for fungal ß-glucans on immune cells that is rapidly internalised after ligand-binding. Here, we demonstrate that pathogen recognition by the Dectin-1a isoform results in the formation of a stable receptor fragment devoid of the ligand binding domain. This fragment persists in phagosomal membranes and contributes to signal transduction which is terminated by the intramembrane proteases Signal Peptide Peptidase-like (SPPL) 2a and 2b. Consequently, immune cells lacking SPPL2b demonstrate increased anti-fungal ROS production, killing capacity and cytokine responses. The identified mechanism allows to uncouple the PRR signalling response from delivery of the pathogen to degradative compartments and identifies intramembrane proteases as part of a regulatory circuit to control anti-fungal immune responses.

Differential gene expression response of synovial fibroblasts from temporomandibular joints and knee joints to dynamic tensile stress.

PURPOSE: Apart from other risk factors, mechanical stress on joints can promote the development of osteoarthritis (OA), which can also affect the temporomandibular joint (TMJ), resulting in cartilage degeneration and synovitis. Synovial fibroblasts (SF) play an important role in upkeeping joint homeostasis and OA pathogenesis, but mechanical stress as a risk factor might act differently depending on the type of joint. We thus investigated the relative impact of mechanical stress on the gene expression pattern of SF from TMJs and knee joints to provide new insights into OA pathogenesis. METHODS: Primary SF isolated from TMJs and knee joints of mice were exposed to mechanical strain of varying magnitudes. Thereafter, the expression of marker genes of the extracellular matrix (ECM), inflammation and bone remodelling were analysed by quantitative real-time polymerase chain reaction (RT-qPCR). RESULTS: SF from the knee joints showed increased expression of genes associated with ECM remodelling, inflammation and bone remodelling after mechanical loading, whereas TMJ-derived SF showed reduced expression of genes associated with inflammation and bone remodelling. SF from the TMJ differed from knee-derived SF with regard to expression of ECM, inflammatory and osteoclastogenesis-promoting marker genes during mechanical strain. CONCLUSIONS: Osteoarthritis-related ECM remodelling markers experience almost no changes in strain-induced gene expression, whereas inflammation and bone remodelling processes seem to differ depending on synovial fibroblast origin. Our data indicate that risk factors for the development and progression of osteoarthritis such as mechanical overuse have a different pathological impact in the TMJ compared to the knee joint.

High Na+ Environments Impair Phagocyte Oxidase-Dependent Antibacterial Activity of Neutrophils.

Infection and inflammation can augment local Na+ abundance. These increases in local Na+ levels boost proinflammatory and antimicrobial macrophage activity and can favor polarization of T cells towards a proinflammatory Th17 phenotype. Although neutrophils play an important role in fighting intruding invaders, the impact of increased Na+ on the antimicrobial activity of neutrophils remains elusive. Here we show that, in neutrophils, increases in Na+ (high salt, HS) impair the ability of human and murine neutrophils to eliminate Escherichia coli and Staphylococcus aureus. High salt caused reduced spontaneous movement, degranulation and impaired production of reactive oxygen species (ROS) while leaving neutrophil viability unchanged. High salt enhanced the activity of the p38 mitogen-activated protein kinase (p38/MAPK) and increased the interleukin (IL)-8 release in a p38/MAPK-dependent manner. Whereas inhibition of p38/MAPK did not result in improved neutrophil defense, pharmacological blockade of the phagocyte oxidase (PHOX) or its genetic ablation mimicked the impaired antimicrobial activity detected under high salt conditions. Stimulation of neutrophils with phorbol-12-myristate-13-acetate (PMA) overcame high salt-induced impairment in ROS production and restored antimicrobial activity of neutrophils. Hence, we conclude that high salt-impaired PHOX activity results in diminished antimicrobial activity. Our findings suggest that increases in local Na+ represent an ionic checkpoint that prevents excessive ROS production of neutrophils, which decreases their antimicrobial potential and could potentially curtail ROS-mediated tissue damage.

Effects of mechanical strain on periodontal ligament fibroblasts in presence of Aggregatibacter actinomycetemcomitans lysate.

PURPOSE: Many adult orthodontic patients suffer from periodontitis, which is caused by oral pathogens such as the gram-negative Aggregatibacter actinomycetemcomitans (Agac). Like orthodontic tooth movement, periodontitis is associated with inflammation and alveolar bone remodelling thereby affecting orthodontic treatment. Interactions of both processes, however, are not sufficiently explored, particularly with regard to oxidative stress. METHODS: After preincubation with Agac lysate for 24 h periodontal ligament fibroblasts (PDLF) were either stretched or compressed for further 48 h simulating orthodontic forces in vitro. We analysed the expression of genes and proteins involved in the formation of reactive oxygen species (NOX-4, ROS) and nitric oxide (NOS-2), inflammation (TNF, IL-6, PTGS-2) and bone remodelling (OPG, RANKL). RESULTS: Agac lysate elevated the expression of NOX-4, NOS-2, inflammatory IL-6 and PTGS-2 and the bone-remodelling RANKL/OPG ratio during compressive, but not tensile mechanical strain. Agac lysate stimulated pressure-induced inflammatory signalling, whereas surprisingly ROS formation was reduced. Pressure-induced downregulation of OPG expression was inhibited by Agac lysate. CONCLUSIONS: Agac lysate impact on the expression of genes and proteins involved in inflammation and bone remodelling as well as ROS formation, when PDLF were subjected to mechanical forces occurring during orthodontic tooth movement.

Mechanical Stress Induce PG-E2 in Murine Synovial Fibroblasts Originating from the Temporomandibular Joint.

Genetic predisposition, traumatic events, or excessive mechanical exposure provoke arthritic changes in the temporomandibular joint (TMJ). We analysed the impact of mechanical stress that might be involved in the development and progression of TMJ osteoarthritis (OA) on murine synovial fibroblasts (SFs) of temporomandibular origin. SFs were subjected to different protocols of mechanical stress, either to a high-frequency tensile strain for 4 h or to a tensile strain of varying magnitude for 48 h. The TMJ OA induction was evaluated based on the gene and protein secretion of inflammatory factors (Icam-1, Cxcl-1, Cxcl-2, Il-1ß, Il-1ra, Il-6, Ptgs-2, PG-E2), subchondral bone remodelling (Rankl, Opg), and extracellular matrix components (Col1a2, Has-1, collagen and hyaluronic acid deposition) using RT-qPCR, ELISA, and HPLC. A short high-frequency tensile strain had only minor effects on inflammatory factors and no effects on the subchondral bone remodelling induction or matrix constituent production. A prolonged tensile strain of moderate and advanced magnitude increased the expression of inflammatory factors. An advanced tensile strain enhanced the Ptgs-2 and PG-E2 expression, while the expression of further inflammatory factors were decreased. The tensile strain protocols had no effects on the RANKL/OPG expression, while the advanced tensile strain significantly reduced the deposition of matrix constituent contents of collagen and hyaluronic acid. The data indicates that the application of prolonged advanced mechanical stress on SFs promote PG-E2 protein secretion, while the deposition of extracellular matrix components is decreased.

Mechanisms controlling bacterial infection in myeloid cells under hypoxic conditions.

Various factors of the tissue microenvironment such as the oxygen concentration influence the host-pathogen interaction. During the past decade, hypoxia-driven signaling via hypoxia-inducible factors (HIF) has emerged as an important factor that affects both the pathogen and the host. In this chapter, we will review the current knowledge of this complex interplay, with a particular emphasis given to the impact of hypoxia and HIF on the inflammatory and antimicrobial activity of myeloid cells, the bacterial responses to hypoxia and the containment of bacterial infections under oxygen-limited conditions. We will also summarize how low oxygen concentrations influence the metabolism of neutrophils, macrophages and dendritic cells. Finally, we will discuss the consequences of hypoxia and HIFα activation for the invading pathogen, with a focus on Pseudomonas aeruginosa, Mycobacterium tuberculosis, Coxiella burnetii, Salmonella enterica and Staphylococcus aureus. This includes a description of the mechanisms and microbial factors, which the pathogens use to sense and react to hypoxic conditions.

NCX1 represents an ionic Na+ sensing mechanism in macrophages.

Inflammation and infection can trigger local tissue Na+ accumulation. This Na+-rich environment boosts proinflammatory activation of monocyte/macrophage-like cells (MΦs) and their antimicrobial activity. Enhanced Na+-driven MΦ function requires the osmoprotective transcription factor nuclear factor of activated T cells 5 (NFAT5), which augments nitric oxide (NO) production and contributes to increased autophagy. However, the mechanism of Na+ sensing in MΦs remained unclear. High extracellular Na+ levels (high salt [HS]) trigger a substantial Na+ influx and Ca2+ loss. Here, we show that the Na+/Ca2+ exchanger 1 (NCX1, also known as solute carrier family 8 member A1 [SLC8A1]) plays a critical role in HS-triggered Na+ influx, concomitant Ca2+ efflux, and subsequent augmented NFAT5 accumulation. Moreover, interfering with NCX1 activity impairs HS-boosted inflammatory signaling, infection-triggered autolysosome formation, and subsequent antibacterial activity. Taken together, this demonstrates that NCX1 is able to sense Na+ and is required for amplifying inflammatory and antimicrobial MΦ responses upon HS exposure. Manipulating NCX1 offers a new strategy to regulate MΦ function.

Mononuclear phagocytes orchestrate prolyl hydroxylase inhibition-mediated renoprotection in chronic tubulointerstitial nephritis.

Prolyl hydroxylase domain enzyme inhibitors (PHDIs) stabilize hypoxia-inducible factors (HIFs), and are protective in models of acute ischemic and inflammatory kidney disease. Whether PHDIs also confer protection in chronic inflammatory kidney disease models remains unknown. Here we investigated long-term effects of PHDI treatment in adenine-induced nephropathy as a model for chronic tubulointerstitial nephritis. After three weeks, renal dysfunction and tubulointerstitial damage, including proximal and distal tubular injury, tubular dilation and renal crystal deposition were significantly attenuated in PHDI-treated (the isoquinoline derivative ICA and Roxadustat) compared to vehicle-treated mice with adenine-induced nephropathy. Crystal-induced renal fibrosis was only partially diminished by treatment with ICA. Renoprotective effects of ICA treatment could not be attributed to changes in adenine metabolism or urinary excretion of the metabolite 2,8-dihydroxyadenine. ICA treatment reduced inflammatory infiltrates of F4/80+ mononuclear phagocytes in the kidneys and supported a regulatory, anti-inflammatory immune response. Furthermore, interstitial deposition of complement C1q was decreased in ICA-treated mice fed an adenine-enriched diet. Tubular cell-specific HIF-1α and myeloid cell-specific HIF-1α and HIF-2α expression were not required for the renoprotective effects of ICA. In contrast, depletion of mononuclear phagocytes with clodronate largely abolished the nephroprotective effects of PHD inhibition. Thus, our findings indicate novel and potent systemic anti-inflammatory properties of PHDIs that confer preservation of kidney function and structure in chronic tubulointerstitial inflammation and might counteract kidney disease progression.

HIF1A and NFAT5 coordinate Na+-boosted antibacterial defense via enhanced autophagy and autolysosomal targeting.

Infection and inflammation are able to induce diet-independent Na+-accumulation without commensurate water retention in afflicted tissues, which favors the pro-inflammatory activation of mouse macrophages and augments their antibacterial and antiparasitic activity. While Na+-boosted host defense against the protozoan parasite Leishmania major is mediated by increased expression of the leishmanicidal NOS2 (nitric oxide synthase 2, inducible), the molecular mechanisms underpinning this enhanced antibacterial defense of mouse macrophages with high Na+ (HS) exposure are unknown. Here, we provide evidence that HS-increased antibacterial activity against E. coli was neither dependent on NOS2 nor on the phagocyte oxidase. In contrast, HS-augmented antibacterial defense hinged on HIF1A (hypoxia inducible factor 1, alpha subunit)-dependent increased autophagy, and NFAT5 (nuclear factor of activated T cells 5)-dependent targeting of intracellular E. coli to acidic autolysosomal compartments. Overall, these findings suggest that the autolysosomal compartment is a novel target of Na+-modulated cell autonomous innate immunity. Abbreviations: ACT: actins; AKT: AKT serine/threonine kinase 1; ATG2A: autophagy related 2A; ATG4C: autophagy related 4C, cysteine peptidase; ATG7: autophagy related 7; ATG12: autophagy related 12; BECN1: beclin 1; BMDM: bone marrow-derived macrophages; BNIP3: BCL2/adenovirus E1B interacting protein 3; CFU: colony forming units; CM-H2DCFDA: 5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate, acetyl ester; CTSB: cathepsin B; CYBB: cytochrome b-245 beta chain; DAPI: 4,6-diamidino-2-phenylindole; DMOG: dimethyloxallyl glycine; DPI: diphenyleneiodonium chloride; E. coli: Escherichia coli; FDR: false discovery rate; GFP: green fluorescent protein; GSEA: gene set enrichment analysis; GO: gene ontology; HIF1A: hypoxia inducible factor 1, alpha subunit; HUGO: human genome organization; HS: high salt (+ 40 mM of NaCl to standard cell culture conditions); HSP90: heat shock 90 kDa proteins; LDH: lactate dehydrogenase; LPS: lipopolysaccharide; Lyz2/LysM: lysozyme 2; NFAT5/TonEBP: nuclear factor of activated T cells 5; MΦ: macrophages; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MFI: mean fluorescence intensity; MIC: minimum inhibitory concentration; MOI: multiplicity of infection; MTOR: mechanistic target of rapamycin kinase; NaCl: sodium chloride; NES: normalized enrichment score; n.s.: not significant; NO: nitric oxide; NOS2/iNOS: nitric oxide synthase 2, inducible; NS: normal salt; PCR: polymerase chain reaction; PGK1: phosphoglycerate kinase 1; PHOX: phagocyte oxidase; RFP: red fluorescent protein; RNA: ribonucleic acid; ROS: reactive oxygen species; sCFP3A: super cyan fluorescent protein 3A; SBFI: sodium-binding benzofuran isophthalate; SLC2A1/GLUT1: solute carrier family 2 (facilitated glucose transporter), member 1; SQSTM1/p62: sequestosome 1; ULK1: unc-51 like kinase 1; v-ATPase: vacuolar-type H+-ATPase; WT: wild type.

Limitation of TCA Cycle Intermediates Represents an Oxygen-Independent Nutritional Antibacterial Effector Mechanism of Macrophages.

In hypoxic and inflamed tissues, oxygen (O2)-dependent antimicrobial defenses are impaired due to a shortage of O2. To gain insight into the mechanisms that control bacterial infection under hypoxic conditions, we infected macrophages with the obligate intracellular pathogen Coxiella burnetii, the causative agent of Q fever. Our experiments revealed that hypoxia impeded C. burnetii replication in a hypoxia-inducible factor (HIF) 1α-dependent manner. Mechanistically, under hypoxia, HIF1α impaired the activity of STAT3, which in turn reduced the intracellular level of TCA cycle intermediates, including citrate, and impeded C. burnetii replication in macrophages. However, bacterial viability was maintained, allowing the persistence of C. burnetii, which is a prerequisite for the development of chronic Q fever. This knowledge will open future research avenues on the pathogenesis of chronic Q fever. In addition, the regulation of TCA cycle metabolites by HIF1α represents a previously unappreciated mechanism of host defense against intracellular pathogens.

Hypoxia, Hypoxia-Inducible Factor-1α, and Innate Antileishmanial Immune Responses.

Low oxygen environments and accumulation of hypoxia-inducible factors (HIFs) are features of infected and inflamed tissues. Here, we summarize our current knowledge on oxygen levels found in Leishmania-infected tissues and discuss which mechanisms potentially contribute to local tissue oxygenation in leishmanial lesions. Moreover, we review the role of hypoxia and HIF-1 on innate antileishmanial immune responses.

Dectin-1 Positive Dendritic Cells Expand after Infection with Leishmania major Parasites and Represent Promising Targets for Vaccine Development.

Resistant mouse strains mount a protective T cell-mediated immune response upon infection with Leishmania (L.) parasites. Healing correlates with a T helper (Th) cell-type 1 response characterized by a pronounced IFN-γ production, while susceptibility is associated with an IL-4-dependent Th2-type response. It has been shown that dermal dendritic cells are crucial for inducing protective Th1-mediated immunity. Additionally, there is growing evidence that C-type lectin receptor (CLR)-mediated signaling is involved in directing adaptive immunity against pathogens. However, little is known about the function of the CLR Dectin-1 in modulating Th1- or Th2-type immune responses by DC subsets in leishmaniasis. We characterized the expression of Dectin-1 on CD11c+ DCs in peripheral blood, at the site of infection, and skin-draining lymph nodes of L. major-infected C57BL/6 and BALB/c mice and in peripheral blood of patients suffering from cutaneous leishmaniasis (CL). Both mouse strains responded with an expansion of Dectin-1+ DCs within the analyzed tissues. In accordance with the experimental model, Dectin-1+ DCs expanded as well in the peripheral blood of CL patients. To study the role of Dectin-1+ DCs in adaptive immunity against L. major, we analyzed the T cell stimulating potential of bone marrow-derived dendritic cells (BMDCs) in the presence of the Dectin-1 agonist Curdlan. These experiments revealed that Curdlan induces the maturation of BMDCs and the expansion of Leishmania-specific CD4+ T cells. Based on these findings, we evaluated the impact of Curdlan/Dectin-1 interactions in experimental leishmaniasis and were able to demonstrate that the presence of Curdlan at the site of infection modulates the course of disease in BALB/c mice: wild-type BALB/c mice treated intradermally with Curdlan developed a protective immune response against L. major whereas Dectin-1-/- BALB/c mice still developed the fatal course of disease after Curdlan treatment. Furthermore, the vaccination of BALB/c mice with a combination of soluble L. major antigens and Curdlan was able to provide a partial protection from severe leishmaniasis. These findings indicate that the ligation of Dectin-1 on DCs acts as an important checkpoint in adaptive immunity against L. major and should therefore be considered in future whole-organism vaccination strategies.

Salt-responsive gut commensal modulates TH17 axis and disease.

A Western lifestyle with high salt consumption can lead to hypertension and cardiovascular disease. High salt may additionally drive autoimmunity by inducing T helper 17 (TH17) cells, which can also contribute to hypertension. Induction of TH17 cells depends on gut microbiota; however, the effect of salt on the gut microbiome is unknown. Here we show that high salt intake affects the gut microbiome in mice, particularly by depleting Lactobacillus murinus. Consequently, treatment of mice with L. murinus prevented salt-induced aggravation of actively induced experimental autoimmune encephalomyelitis and salt-sensitive hypertension by modulating TH17 cells. In line with these findings, a moderate high-salt challenge in a pilot study in humans reduced intestinal survival of Lactobacillus spp., increased TH17 cells and increased blood pressure. Our results connect high salt intake to the gut-immune axis and highlight the gut microbiome as a potential therapeutic target to counteract salt-sensitive conditions.

Elementary immunology: Na+ as a regulator of immunity.

The skin can serve as an interstitial Na+ reservoir. Local tissue Na+ accumulation increases with age, inflammation and infection. This increased local Na+ availability favors pro-inflammatory immune cell function and dampens their anti-inflammatory capacity. In this review, we summarize available data on how NaCl affects various immune cells. We particularly focus on how salt promotes pro-inflammatory macrophage and T cell function and simultaneously curtails their regulatory and anti-inflammatory potential. Overall, these findings demonstrate that local Na+ availability is a promising novel regulator of immunity. Hence, the modulation of tissue Na+ levels bears broad therapeutic potential: increasing local Na+ availability may help in treating infections, while lowering tissue Na+ levels may be used to treat, for example, autoimmune and cardiovascular diseases.

Myeloid Cell-Derived HIF-1α Promotes Control of Leishmania major.

Hypoxia-inducible factor-1α (HIF-1α), which accumulates in mammalian host organisms during infection, supports the defense against microbial pathogens. However, whether and to what extent HIF-1α expressed by myeloid cells contributes to the innate immune response against Leishmania major parasites is unknown. We observed that Leishmania-infected humans and L. major-infected C57BL/6 mice exhibited substantial amounts of HIF-1α in acute cutaneous lesions. In vitro, HIF-1α was required for leishmanicidal activity and high-level NO production by IFN-γ/LPS-activated macrophages. Mice deficient for HIF-1α in their myeloid cell compartment had a more severe clinical course of infection and increased parasite burden in the skin lesions compared with wild-type controls. These findings were paralleled by reduced expression of type 2 NO synthase by lesional CD11b+ cells. Together, these data illustrate that HIF-1α is required for optimal innate leishmanicidal immune responses and, thereby, contributes to the cure of cutaneous leishmaniasis.

Ferritin-Mediated Iron Sequestration Stabilizes Hypoxia-Inducible Factor-1α upon LPS Activation in the Presence of Ample Oxygen.

Both hypoxic and inflammatory conditions activate transcription factors such as hypoxia-inducible factor (HIF)-1α and nuclear factor (NF)-κB, which play a crucial role in adaptive responses to these challenges. In dendritic cells (DC), lipopolysaccharide (LPS)-induced HIF1α accumulation requires NF-κB signaling and promotes inflammatory DC function. The mechanisms that drive LPS-induced HIF1α accumulation under normoxia are unclear. Here, we demonstrate that LPS inhibits prolyl hydroxylase domain enzyme (PHD) activity and thereby blocks HIF1α degradation. Of note, LPS-induced PHD inhibition was neither due to cosubstrate depletion (oxygen or α-ketoglutarate) nor due to increased levels of reactive oxygen species, fumarate, and succinate. Instead, LPS inhibited PHD activity through NF-κB-mediated induction of the iron storage protein ferritin and subsequent decrease of intracellular available iron, a critical cofactor of PHD. Thus, hypoxia and LPS both induce HIF1α accumulation via PHD inhibition but deploy distinct molecular mechanisms (lack of cosubstrate oxygen versus deprivation of co-factor iron).