Lipid droplet accumulation occurs early following Salmonella infection and contributes to intracellular bacterial survival and replication.

Salmonellosis is a public health problem caused by Salmonella sp., a highly adapted facultative intracellular pathogen. After internalization, Salmonella sp. Manipulates several host processes, mainly through the activation of the type III secretion system (T3SS), including modification of host lipid metabolism and lipid droplet (LD) accumulation. LDs are dynamic and complex lipid-rich organelles involved in several cellular processes. The present study investigated the mechanism involved in LD biogenesis in Salmonella-infected macrophages and its role in bacterial pathogenicity. Here, we reported that S. Typhimurium induced a rapid time-dependent increase of LD formation in macrophages. The LD biogenesis was demonstrated to depend on Salmonella's viability and SPI1-related T3SS activity, with the participation of Toll-Like Receptor (TLR) signaling. We also observed that LD accumulation occurs through TLR2-dependent signaling and is counter-regulated by TLR4. Last, the pharmacologic modulation of LD formation by inhibiting diacylglycerol O-acyltransferase 1 (DGAT1) and cytosolic phospholipase A2 (cPLA2) significantly reduced the intracellular bacterial proliferation and impaired the prostaglandin E2 (PGE2 ) synthesis. Collectively, our data suggest the role of LDs on S. typhimurium intracellular survival and replication in macrophages. This data set provides new perspectives for future investigations about LDs in host-pathogen interaction.

Mitochondrial Reactive Oxygen Species Participate in Signaling Triggered by Heme in Macrophages and upon Hemolysis.

Hemolysis causes an increase of intravascular heme, oxidative damage, and inflammation in which macrophages play a critical role. In these cells, heme can act as a prototypical damage-associated molecular pattern, inducing TLR4-dependent cytokine production through the MyD88 pathway, independently of TRIF. Heme promotes reactive oxygen species (ROS) generation independently of TLR4. ROS and TNF production contribute to heme-induced necroptosis and inflammasome activation; however, the role of ROS in proinflammatory signaling and cytokine production remains unknown. In this study, we demonstrate that heme activates at least three signaling pathways that contribute to a robust MAPK phosphorylation and cytokine expression in mouse macrophages. Although heme did not induce a detectable Myddosome formation, the TLR4/MyD88 axis was important for phosphorylation of p38 and secretion of cytokines. ROS generation and spleen tyrosine kinase (Syk) activation induced by heme were critical for most proinflammatory signaling pathways, as the antioxidant N-acetyl-l-cysteine and a Syk inhibitor differentially blocked heme-induced ROS, MAPK phosphorylation, and cytokine production in macrophages. Early generated mitochondrial ROS induced by heme was Syk dependent, selectively promoted the phosphorylation of ERK1/2 without affecting JNK or p38, and contributed to CXCL1 and TNF production. Finally, lethality caused by sterile hemolysis in mice required TLR4, TNFR1, and mitochondrial ROS, supporting the rationale to target these pathways to mitigate tissue damage of hemolytic disorders.

The Role of MIF on Eosinophil Biology and Eosinophilic Inflammation.

Macrophage migration inhibitory factor (MIF) is an inflammatory cytokine that participates in innate and adaptive immune responses. MIF contributes to the resistance against infection agents, but also to the cellular and tissue damage in infectious, autoimmune, and allergic diseases. In the past years, several studies demonstrated a critical role for MIF in the pathogenesis of type-2-mediated inflammation, including allergy and helminth infection. Atopic patients have increased MIF amounts in affected tissues, mainly produced by immune cells such as macrophages, Th2 cells, and eosinophils. Increased MIF mRNA and protein are found in activated Th2 cells, while eosinophils stock pre-formed MIF protein and secrete high amounts of MIF upon stimulation. In mouse models of allergic asthma, the lack of MIF causes an almost complete abrogation of the cardinal signs of the disease including mucus secretion, eosinophilic inflammation, and airway hyper-responsiveness. Additionally, blocking the expression of MIF in animal models leads to significant reduction of pathological signs of eosinophilic inflammation such as rhinitis, atopic dermatitis, eosinophilic esophagitis and helminth infection. A number of studies indicate that MIF is important in the effector phase of type-2 immune responses, while its contribution to Th2 differentiation and IgE production is not consensual. MIF has been found to intervene in different aspects of eosinophil physiology including differentiation, survival, activation, and migration. CD4+ T cells and eosinophils express CD74 and CXCR4, receptors able to signal upon MIF binding. Blockage of these receptors with neutralizing antibodies or small molecule antagonists also succeeds in reducing the signals of inflammation in experimental allergic models. Together, these studies demonstrate an important contribution of MIF on eosinophil biology and in the pathogenesis of allergic diseases and helminth infection.

Co-protoporphyrin IX and Sn-protoporphyrin IX inactivate Zika, Chikungunya and other arboviruses by targeting the viral envelope.

The global situation of diseases transmitted by arthropod-borne viruses such as Dengue (DENV), Yellow Fever (YFV), Chikungunya (CHIKV) and Zika (ZIKV) viruses is alarming and treatment of human infection by these arboviruses faces several challenges. The discovery of broad-spectrum antiviral molecules, able to inactivate different groups of viruses, is an interesting approach. The viral envelope is a common structure among arboviruses, being a potential target for antivirals. Porphyrins are amphipathic molecules able to interact with membranes and absorb light, being widely used in photodynamic therapy. Previously, we showed that heme, Co-protoporphyrin IX (CoPPIX) and Sn-protoporphyrin IX (SnPPIX) directly inactivate DENV and YFV infectious particles. Here we demonstrate that the antiviral activity of these porphyrins can be broadened to CHIKV, ZIKV, Mayaro virus, Sindbis virus and Vesicular Stomatitis virus. Porphyrin treatment causes viral envelope protein loss, affecting viral morphology, adsorption and entry into target cells. Also, light-stimulation enhanced the SnPPIX activity against all tested arboviruses. In summary, CoPPIX and SnPPIX were shown to be efficient broad-spectrum compounds to inactivate medically and veterinary important viruses.

ROS and Trypanosoma cruzi: Fuel to infection, poison to the heart.

The activation of macrophage respiratory burst in response to infection with Trypanosoma cruzi inflicts oxidative damage to the host's tissues. For decades, the role of reactive oxygen species (ROS) in the elimination of T. cruzi was taken for granted, but recent evidence suggests parasite growth is stimulated in oxidative environments. It is still a matter of debate whether indeed oxidative environments provide ideal conditions (e.g., iron availability in macrophages) for T. cruzi growth and whether indeed ROS signals directly to stimulate growth. Nitric oxide (NO) and ROS combine to form peroxynitrite, participating in the killing of phagocytosed parasites by activated macrophages. In response to infection, mitochondrial ROS are produced by cardiomyocytes. They contribute to oxidative damage that persists at the chronic stage of infection and is involved in functional impairment of the heart. In this review, we discuss how oxidative stress helps parasite growth during the acute stage and how it participates in the development of cardiomyopathy at the chronic stage.

Heme and iron induce protein aggregation.

Heme is an essential molecule expressed in many tissues where it plays key roles as the prosthetic group of several proteins involved in vital physiological and metabolic processes such as gas and electron transport. Structurally, heme is a tetrapyrrole ring containing an atom of iron (Fe) in its center. When released into the extracellular milieu, heme exerts several deleterious effects, which make it an important player in infectious and noninfectious hemolytic diseases where large amounts of free heme are observed such as malaria, dengue fever, ß-thalassemia, sickle cell disease and ischemia-reperfusion. Our recent work has uncovered an unappreciated cellular response triggered by heme or Fe, one of its degradation products, on macrophages, which is the formation of protein aggregates known as aggresome-like induced structres (ALIS). This response was shown to be fully dependent on ROS production and the activation of the transcription factor NFE2L2/NRF2. In addition, we have demonstrated that heme degradation by HMOX1/HO-1 (heme oxygenase 1) is required and that Fe is essential for the formation of ALIS, as heme analogs lacking the central atom of Fe are not able to induce these structures. ALIS formation is also observed in vivo, in a model of phenylhydrazine (PHZ)-induced hemolysis, indicating that it is an integral part of the host response to excessive free heme and that it may play a role in cellular homeostasis.

Macrophage-dependent IL-1ß production induces cardiac arrhythmias in diabetic mice.

Diabetes mellitus (DM) encompasses a multitude of secondary disorders, including heart disease. One of the most frequent and potentially life threatening disorders of DM-induced heart disease is ventricular tachycardia (VT). Here we show that toll-like receptor 2 (TLR2) and NLRP3 inflammasome activation in cardiac macrophages mediate the production of IL-1ß in DM mice. IL-1ß causes prolongation of the action potential duration, induces a decrease in potassium current and an increase in calcium sparks in cardiomyocytes, which are changes that underlie arrhythmia propensity. IL-1ß-induced spontaneous contractile events are associated with CaMKII oxidation and phosphorylation. We further show that DM-induced arrhythmias can be successfully treated by inhibiting the IL-1ß axis with either IL-1 receptor antagonist or by inhibiting the NLRP3 inflammasome. Our results establish IL-1ß as an inflammatory connection between metabolic dysfunction and arrhythmias in DM.

Protein aggregation as a cellular response to oxidative stress induced by heme and iron.

Hemolytic diseases include a variety of conditions with diverse etiologies in which red blood cells are destroyed and large amounts of hemeproteins are released. Heme has been described as a potent proinflammatory molecule that is able to induce multiple innate immune responses, such as those triggered by TLR4 and the NLRP3 inflammasome, as well as necroptosis in macrophages. The mechanisms by which eukaryotic cells respond to the toxic effects induced by heme to maintain homeostasis are not fully understood, however. Here we describe a previously uncharacterized cellular response induced by heme: the formation of p62/SQTM1 aggregates containing ubiquitinated proteins in structures known as aggresome-like induced structures (ALIS). This action is part of a response driven by the transcription factor NRF2 to the excessive generation of reactive oxygen species induced by heme that results in the expression of genes involved in antioxidant responses, including p62/SQTM1. Furthermore, we show that heme degradation by HO-1 is required for ALIS formation, and that the free iron released on heme degradation is necessary and sufficient to induce ALIS. Moreover, ferritin, a key protein in iron metabolism, prevents excessive ALIS formation. Finally, in vivo, hemolysis promotes an increase in ALIS formation in target tissues. Our data unravel a poorly understood aspect of the cellular responses induced by heme that can be explored to better understand the effects of free heme and free iron during hemolytic diseases such as sickle cell disease, dengue fever, malaria, and sepsis.

RIPK1 and PGAM5 Control Leishmania Replication through Distinct Mechanisms.

Leishmaniasis is an important parasitic disease found in the tropics and subtropics. Cutaneous and visceral leishmaniasis affect an estimated 1.5 million people worldwide. Despite its human health relevance, relatively little is known about the cell death pathways that control Leishmania replication in the host. Necroptosis is a recently identified form of cell death with potent antiviral effects. Receptor interacting protein kinase 1 (RIPK1) is a critical kinase that mediates necroptosis downstream of death receptors and TLRs. Heme, a product of hemoglobin catabolism during certain intracellular pathogen infections, is also a potent inducer of macrophage necroptosis. We found that human visceral leishmaniasis patients exhibit elevated serum levels of heme. Therefore, we examined the impact of heme and necroptosis on Leishmania replication. Indeed, heme potently inhibited Leishmania replication in bone marrow-derived macrophages. Moreover, we found that inhibition of RIPK1 kinase activity also enhanced parasite replication in the absence of heme. We further found that the mitochondrial phosphatase phosphoglycerate mutase family member 5 (PGAM5), a putative downstream effector of RIPK1, was also required for inhibition of Leishmania replication. In mouse infection, both PGAM5 and RIPK1 kinase activity are required for IL-1ß expression in response to Leishmania However, PGAM5, but not RIPK1 kinase activity, was directly responsible for Leishmania-induced IL-1ß secretion and NO production in bone marrow-derived macrophages. Collectively, these results revealed that RIPK1 and PGAM5 function independently to exert optimal control of Leishmania replication in the host.

Red alert: labile heme is an alarmin.

Alarmins are a heterogeneous group of endogenous molecules that signal cellular damage when sensed extracellularly. Heme is an endogenous molecule that acts as a prosthetic group of hemoproteins, such as hemoglobin and myoglobin. When released from damaged red blood cells or muscle cells, oxidized hemoglobin and myoglobin release their prosthetic heme groups, respectively. This generates labile heme, which is sensed by pattern recognition receptors (PRR) expressed by innate immune cells and possibly regulatory T cells (TREG). The ensuing adaptive response, which alerts for the occurrence of red blood cell or muscle cell damage, regulates the pathologic outcome of hemolysis or rhabdomyolysis, respectively. In conclusion, we propose that labile heme is an alarmin.

CCR4 Controls the Suppressive Effects of Regulatory T Cells on Early and Late Events during Severe Sepsis.

Sepsis is a deadly disease characterized by an overwhelming release of inflammatory mediators and the activation of different types of cells. This altered state of cell activation, termed leukocyte reprogramming, contributes to patient outcome. However, the understanding of the process underlying sepsis and the role of regulatory T cells (Tregs) in sepsis remains to be elucidated. In this study, we investigated the role of CCR4, the CCL17/CCL22 chemokine receptor, in the innate and acquired immune responses during severe sepsis and the role of Tregs in effecting the outcome. In contrast with wild-type (WT) mice subjected to cecal ligation and puncture (CLP) sepsis, CCR4-deficient (CCR4-/-) septic mice presented an increased survival rate, significant neutrophil migration toward the infection site, a low bacterial count in the peritoneum, and reduced lung inflammation and serum cytokine levels. Thus, a better early host response may favor an adequate long-term response. Consequently, the CCR4-/- septic mice were not susceptible to secondary fungal infection, in contrast with the WT septic mice. Furthermore, Tregs cells from the CCR4-/- septic mice showed reduced suppressive effects on neutrophil migration (both in vivo and in vitro), lymphocyte proliferation and ROS production from activated neutrophils, in contrast with what was observed for Tregs from the WT septic mice. These data show that CCR4 is involved in immunosuppression after severe sepsis and suggest that CCR4+ Tregs negatively modulate the short and long-term immune responses.

Hemolysis-induced lethality involves inflammasome activation by heme.

The increase of extracellular heme is a hallmark of hemolysis or extensive cell damage. Heme has prooxidant, cytotoxic, and inflammatory effects, playing a central role in the pathogenesis of malaria, sepsis, and sickle cell disease. However, the mechanisms by which heme is sensed by innate immune cells contributing to these diseases are not fully characterized. We found that heme, but not porphyrins without iron, activated LPS-primed macrophages promoting the processing of IL-1ß dependent on nucleotide-binding domain and leucine rich repeat containing family, pyrin domain containing 3 (NLRP3). The activation of NLRP3 by heme required spleen tyrosine kinase, NADPH oxidase-2, mitochondrial reactive oxygen species, and K(+) efflux, whereas it was independent of heme internalization, lysosomal damage, ATP release, the purinergic receptor P2X7, and cell death. Importantly, our results indicated the participation of macrophages, NLRP3 inflammasome components, and IL-1R in the lethality caused by sterile hemolysis. Thus, understanding the molecular pathways affected by heme in innate immune cells might prove useful to identify new therapeutic targets for diseases that have heme release.

Cryptococcus neoformans glucuronoxylomannan fractions of different molecular masses are functionally distinct.

AIMS: Glucuronoxylomannan (GXM) is the major polysaccharide component of Cryptococcus neoformans. We evaluated in this study whether GXM fractions of different molecular masses were functionally distinct. MATERIALS & METHODS: GXM samples isolated from C. neoformans cultures were fractionated to generate polysaccharide preparations differing in molecular mass. These fractions were used in experiments focused on the association of GXM with cell wall components of C. neoformans, as well as on the interaction of the polysaccharide with host cells. RESULTS & CONCLUSION: GXM fractions of variable molecular masses bound to the surface of a C. neoformans acapsular mutant in a punctate pattern that is in contrast to the usual annular pattern of surface coating observed when GXM samples containing the full molecular mass range were used. The polysaccharide samples were also significantly different in their ability to stimulate cytokine production by host cells. Our findings indicate that GXM fractions are functionally distinct depending on their mass.

Binding of the wheat germ lectin to Cryptococcus neoformans chitooligomers affects multiple mechanisms required for fungal pathogenesis.

The principal capsular component of Cryptococcus neoformans, glucuronoxylomannan (GXM), interacts with surface glycans, including chitin-like oligomers. Although the role of GXM in cryptococcal infection has been well explored, there is no information on how chitooligomers affect fungal pathogenesis. In this study, surface chitooligomers of C. neoformans were blocked through the use of the wheat germ lectin (WGA) and the effects on animal pathogenesis, interaction with host cells, fungal growth and capsule formation were analyzed. Treatment of C. neoformans cells with WGA followed by infection of mice delayed mortality relative to animals infected with untreated fungal cells. This observation was associated with reduced brain colonization by lectin-treated cryptococci. Blocking chitooligomers also rendered yeast cells less efficient in their ability to associate with phagocytes. WGA did not affect fungal viability, but inhibited GXM release to the extracellular space and capsule formation. In WGA-treated yeast cells, genes that are involved in capsule formation and GXM traffic had their transcription levels decreased in comparison with untreated cells. Our results suggest that cellular pathways required for capsule formation and pathogenic mechanisms are affected by blocking chitin-derived structures at the cell surface of C. neoformans. Targeting chitooligomers with specific ligands may reveal new therapeutic alternatives to control cryptococcosis.

Ketoprofen impairs immunosuppression induced by severe sepsis and reveals an important role for prostaglandin E2.

The mechanism of immunosuppression induced by severe sepsis is not fully understood. The production of prostaglandin E2 (PGE2) during sepsis is well known, but its role in long-term consequences of sepsis has not been explored. The current study evaluates the role of PGE2 in the development of immunosuppression secondary to sepsis and its potential as therapeutic target. Cecal ligation and puncture was used as an experimental model for sepsis induction in Balb/c and C57BL/6 mice. Immunosuppression was evaluated by the response to secondary infection with Aspergillus fumigatus in sepsis survivors. The role of prostanoids was evaluated in vivo and in vitro by treatment with the cyclooxygenase inhibitor ketoprofen. Balb/c mice were more susceptible than C57BL/6 to severe sepsis and to secondary infection, with a greater mortality rate. Prostaglandin E2 concentrations found in bronchoalveolar lavage in sham and cecal ligation and puncture group after fungal challenge were much higher in Balb/c than in C57BL/6 mice. Ketoprofen treatment improved survival of septic Balb/c mice subjected to secondary infection, while also enhancing macrophage phagocytosis and neutrophil recruitment to the lungs. We identified a pivotal role for PGE2 acting on EP4 receptors in modulating cytokine production differentially by sham and septic macrophages. Furthermore, sepsis also altered key enzymes in PGE2 synthesis and degradation. Our results indicate the involvement of PGE2 in severe sepsis-induced immunosuppression. Inhibition of PGE2 production represents an attractive target to improve innate immune response against secondary infection in the immunocompromised host.

Heme induces programmed necrosis on macrophages through autocrine TNF and ROS production.

Diseases that cause hemolysis or myonecrosis lead to the leakage of large amounts of heme proteins. Free heme has proinflammatory and cytotoxic effects. Heme induces TLR4-dependent production of tumor necrosis factor (TNF), whereas heme cytotoxicity has been attributed to its ability to intercalate into cell membranes and cause oxidative stress. We show that heme caused early macrophage death characterized by the loss of plasma membrane integrity and morphologic features resembling necrosis. Heme-induced cell death required TNFR1 and TLR4/MyD88-dependent TNF production. Addition of TNF to Tlr4(-/-) or to Myd88(-/-) macrophages restored heme-induced cell death. The use of necrostatin-1, a selective inhibitor of receptor-interacting protein 1 (RIP1, also known as RIPK1), or cells deficient in Rip1 or Rip3 revealed a critical role for RIP proteins in heme-induced cell death. Serum, antioxidants, iron chelation, or inhibition of c-Jun N-terminal kinase (JNK) ameliorated heme-induced oxidative burst and blocked macrophage cell death. Macrophages from heme oxygenase-1 deficient mice (Hmox1(-/-)) had increased oxidative stress and were more sensitive to heme. Taken together, these results revealed that heme induces macrophage necrosis through 2 synergistic mechanisms: TLR4/Myd88-dependent expression of TNF and TLR4-independent generation of ROS.

Maxadilan, the Lutzomyia longipalpis vasodilator, drives plasma leakage via PAC1-CXCR1/2-pathway.

Experiments were designed to determine if the vasodilatory peptides maxadilan and pituitary adenylate cyclase-activating peptide (PACAP-38) may cause plasma leakage through activation of leukocytes and to what extent these effects could be due to PAC1 and CXCR1/2 receptor stimulation. Intravital microscopy of hamster cheek pouches utilizing FITC-dextran and rhodamine, respectively, as plasma and leukocyte markers was used to measure arteriolar diameter, plasma leakage and leukocyte accumulation in a selected area (5mm(2)) representative of the hamster cheek pouch microcirculation. Our studies showed that the sand fly vasodilator maxadilan and PACAP-38 induced arteriolar dilation, leukocyte accumulation and plasma leakage in postcapillary venules. The recombinant mutant of maxadilan M65 and an antagonist of CXCR1/2 receptors, reparixin, and an inhibitor of CD11b/CD18 up-regulation, ropivacaine, inhibited all these effects as induced by maxadilan. Dextran sulfate, a complement inhibitor with heparin-like anti-inflammatory effects, inhibited plasma leakage and leukocyte accumulation but not arteriolar dilation as induced by maxadilan and PACAP-38. In vitro studies with isolated human neutrophils showed that maxadilan is a potent stimulator of neutrophil migration comparable with fMLP and leukotriene B(4) and that M65 and reparixin inhibited such migration. The data suggest that leukocyte accumulation and plasma leakage induced by maxadilan involves a mechanism related to PAC1- and CXCR1/2-receptors on leukocytes and endothelial cells.

Glycoconjugates and polysaccharides from the Scedosporium/Pseudallescheria boydii complex: structural characterisation, involvement in cell differentiation, cell recognition and virulence.

Peptidorhamnomannans (PRMs), rhamnomannans and α-glucans are especially relevant for the architecture of the Scedosporium/Pseudallescheria boydii cell wall, but many of them are immunologically active, with great potential as regulators of pathogenesis and the immune response of the host. In addition, some of them can be specifically recognised by antibodies from the sera of patients, suggesting that they could also be useful in diagnosis of fungal infections. Their primary structures have been determined, based on a combination of techniques including gas chromatography, electrospray ionization - mass spectrometry (ESI-MS), (1)H-COSY and TOCSY, (13)C and (1)H/(13)C NMR spectroscopy. Using monoclonal antibodies to PRM, we showed that it is involved in germination and viability of P. boydii conidia, in the phagocytosis of P. boydii conidia by macrophages and non-phagocytic cells and in the survival of mice with P. boydii infection. Also, components of the fungal cell wall, such as α-glucans, are involved. Rhamnomannans are immunostimulatory and participate in the recognition and uptake of fungal cells by the immune system. These glycosylated polymers, being present in the fungal cell wall, are mostly absent from mammalian cells, and are excellent targets for the design of new agents capable of inhibiting fungal growth and differentiation of pathogens.

Leukotriene B4 mediates neutrophil migration induced by heme.

High concentrations of free heme found during hemolytic events or cell damage leads to inflammation, characterized by neutrophil recruitment and production of reactive oxygen species, through mechanisms not yet elucidated. In this study, we provide evidence that heme-induced neutrophilic inflammation depends on endogenous activity of the macrophage-derived lipid mediator leukotriene B(4) (LTB(4)). In vivo, heme-induced neutrophil recruitment into the peritoneal cavity of mice was attenuated by pretreatment with 5-lipoxygenase (5-LO) inhibitors and leukotriene B(4) receptor 1 (BLT1) receptor antagonists as well as in 5-LO knockout (5-LO(-/-)) mice. Heme administration in vivo increased peritoneal levels of LTB(4) prior to and during neutrophil recruitment. Evidence that LTB(4) was synthesized by resident macrophages, but not mast cells, included the following: 1) immuno-localization of heme-induced LTB(4) was compartmentalized exclusively within lipid bodies of resident macrophages; 2) an increase in the macrophage population enhanced heme-induced neutrophil migration; 3) depletion of resident mast cells did not affect heme-induced LTB(4) production or neutrophil influx; 4) increased levels of LTB(4) were found in heme-stimulated peritoneal cavities displaying increased macrophage numbers; and 5) in vitro, heme was able to activate directly macrophages to synthesize LTB(4). Our findings uncover a crucial role of LTB(4) in neutrophil migration induced by heme and suggest that beneficial therapeutic outcomes could be achieved by targeting the 5-LO pathway in the treatment of inflammation associated with hemolytic processes.

Schistosomal­derived lysophosphatidylcholine are involved in eosinophil activation and recruitment through Toll­like receptor-2-dependent mechanisms.

Parasite­derived lipids may play important roles in host­pathogen interactions and escape mechanisms. Herein, we evaluated the role of schistosomal­derived lipids in Toll­like receptor (TLR)-2 and eosinophil activation in Schistosoma mansoni infection. Mice lacking TLR2 exhibited reduced liver eosinophilic granuloma, compared with that of wild­type animals, following S. mansoni infection. Decreased eosinophil accumulation and eosinophil lipid body (lipid droplet) formation, at least partially due to reduced production of eotaxin, interleukin (IL)­5, and IL­13 in S. mansoni-infected TLR2-/- mice, compared with the corresponding production in wild­type mice, was noted. Although no differences were observed in survival rates during the acute schistosomal infection (up to 50 days), increased survival of TLR2-/- mice, compared with survival of wild­type mice, was observed during the chronic phase of infection. Schistosomal lipid extract­ and schistosomal­derived lysophosphatidylcholine (lyso­PC)-stimulated macrophages in vitro induced TLR2­dependent NF­kB activation and cytokine production. Furthermore, in vivo schistosomal lyso­PC administration induced eosinophil recruitment and cytokine production, in a mechanism largely dependent on TLR2. Taken together, our results suggest that schistosomal­derived lyso­PC may participate in cytokine production and eosinophil activation through a TLR2­dependent pathway in S. mansoni infection. Moreover, our results suggest that TLR2­dependent inflammatory reaction, cytokine production, and eosinophil recruitment and activation may contribute to the pathogenesis and lethality in the chronic phase of infection.