Fasciola hepatica induces weak NETosis and low production of intra- and extracellular ROS in exposed bovine polymorphonuclear neutrophils

Peixoto, Raquel S.; Silva, Liliana; López-Osorio, Sara; Zhou, Ershun; Gärtner, Ulrich; Conejeros, Iván; Taubert, Anja; Hermosilla, Carlos

doi:10.1016/j.dci.2020.103787

Cited by 21 publications

(44 citation statements)

References 57 publications

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“…Cysteine and glycine are necessary for the synthesis of glutathione which plays an important role in detoxification of ROS. Of note, high ROS concentrations are known to impair parasite viability and represent a pivotal mechanism of early host innate immune reactions directed against protozoan and metazoan parasites [99,100].…”

Section: Discussionmentioning

confidence: 99%

Metabolic Signatures of Cryptosporidium parvum-Infected HCT-8 Cells and Impact of Selected Metabolic Inhibitors on C. parvum Infection under Physioxia and Hyperoxia

Vélez

Velásquez

Silva

et al. 2021

Biology

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Cryptosporidium parvum is an apicomplexan zoonotic parasite recognized as the second leading-cause of diarrhoea-induced mortality in children. In contrast to other apicomplexans, C.parvum has minimalistic metabolic capacities which are almost exclusively based on glycolysis. Consequently, C. parvum is highly dependent on its host cell metabolism. In vivo (within the intestine) infected epithelial host cells are typically exposed to low oxygen pressure (1–11% O2, termed physioxia). Here, we comparatively analyzed the metabolic signatures of C. parvum-infected HCT-8 cells cultured under both, hyperoxia (21% O2), representing the standard oxygen condition used in most experimental settings, and physioxia (5% O2), to be closer to the in vivo situation. The most pronounced effect of C. parvum infection on host cell metabolism was, on one side, an increase in glucose and glutamine uptake, and on the other side, an increase in lactate release. When cultured in a glutamine-deficient medium, C. parvum infection led to a massive increase in glucose consumption and lactate production. Together, these results point to the important role of both glycolysis and glutaminolysis during C. parvum intracellular replication. Referring to obtained metabolic signatures, we targeted glycolysis as well as glutaminolysis in C. parvum-infected host cells by using the inhibitors lonidamine [inhibitor of hexokinase, mitochondrial carrier protein (MCP) and monocarboxylate transporters (MCT) 1, 2, 4], galloflavin (lactate dehydrogenase inhibitor), syrosingopine (MCT1- and MCT4 inhibitor) and compound 968 (glutaminase inhibitor) under hyperoxic and physioxic conditions. In line with metabolic signatures, all inhibitors significantly reduced parasite replication under both oxygen conditions, thereby proving both energy-related metabolic pathways, glycolysis and glutaminolysis, but also lactate export mechanisms via MCTs as pivotal for C. parvum under in vivo physioxic conditions of mammals.

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Section: Discussionmentioning

confidence: 99%

Metabolic Signatures of Cryptosporidium parvum-Infected HCT-8 Cells and Impact of Selected Metabolic Inhibitors on C. parvum Infection under Physioxia and Hyperoxia

Vélez

Velásquez

Silva

et al. 2021

Biology

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“…Similarly, macrophages have a low expression of MHC-II and are impaired in their ability to produce pro-inflammatory mediators such as iNOS and TNF, instead adopting a regulatory profile by secreting IL-10 and TGFb (17,19,20). Although not demonstrated in vivo, the exposure of bovine neutrophils to the intra-mammalian life stages of F. hepatica in vitro, failed to induce significant production of reactive oxygen species or NETosis, suggesting that the parasite was impairing the antimicrobial activities of neutrophils (21). Likewise, there is no evidence of eosinophil degranulation in the peritoneal cavity, which indicates that these cells are not activated or have an alternative phenotype that is not contributing to parasite killing (16).…”

Section: Fasciola Hepatica Manipulates the Host Immune Response To Support Successful Invasionmentioning

confidence: 99%

An Evaluation of the Fasciola hepatica miRnome Predicts a Targeted Regulation of Mammalian Innate Immune Responses

et al. 2021

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Understanding mechanisms by which parasitic worms (helminths) control their hosts’ immune responses is critical to the development of effective new disease interventions. Fasciola hepatica, a global scourge of humans and their livestock, suppresses host innate immune responses within hours of infection, ensuring that host protective responses are quickly incapacitated. This allows the parasite to freely migrate from the intestine, through the liver to ultimately reside in the bile duct, where the parasite establishes a chronic infection that is largely tolerated by the host. The recent identification of micro(mi)RNA, small RNAs that regulate gene expression, within the extracellular vesicles secreted by helminths suggest that these non-coding RNAs may have a role in the parasite-host interplay. To date, 77 miRNAs have been identified in F. hepatica comprising primarily of ancient conserved species of miRNAs. We hypothesized that many of these miRNAs are utilized by the parasite to regulate host immune signaling pathways. To test this theory, we first compiled all of the known published F. hepatica miRNAs and critically curated their sequences and annotations. Then with a focus on the miRNAs expressed by the juvenile worms, we predicted gene targets within human innate immune cells. This approach revealed the existence of targets within every immune cell, providing evidence for the universal management of host immunology by this parasite. Notably, there was a high degree of redundancy in the potential for the parasite to regulate the activation of dendritic cells, eosinophils and neutrophils, with multiple miRNAs predicted to act on singular gene targets within these cells. This original exploration of the Fasciola miRnome offers the first molecular insight into mechanisms by which F. hepatica can regulate the host protective immune response.

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“…Key PMN-derived defence mechanisms have been classically defined as a variety of potent intracellular/extracellular microbicidal mechanisms to efficiently kill invasive pathogens, such as bacteria, viruses, fungi [ 18 , 19 ] and large protozoan and helminth parasites [ 20 , 21 , 22 ] and to stimulate adaptive defence mechanisms [ 23 , 24 , 25 , 26 , 27 ]. PMN-derived effector mechanisms include phagocytosis, reactive oxygen species (ROS) production, secretion of granules containing several antimicrobial proteins [ 24 , 28 ], casting of neutrophil extracellular traps (NETs) [ 29 , 30 ] and chemokine/cytokine production, thereby inducing the arrival of other leukocytes to the site of infection or inflammation [ 31 , 32 ].…”

Section: Introductionmentioning

confidence: 99%

The Role of TLR2 and TLR4 in Recognition and Uptake of the Apicomplexan Parasite Eimeria bovis and Their Effects on NET Formation

et al. 2021

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Background: Bovine polymorphonuclear neutrophils (PMN) constitutively express the Toll-like receptors (TLRs) TLR2 and TLR4 and have been shown to generate Neutrophil extracellular traps (NETs) upon exposure to Eimeria bovis. The present work investigated the role of TLR2 and TLR4 in the recognition and uptake of E. bovis sporozoites, IL-8 production and neutrophil extracellular trap (NET) formation. Methods: TLR expression was performed by flow cytometric analysis on PMN exposed to live carboxyfluorescein succinimidyl ester (CFSE)-stained sporozoites. Supernatants of PMN exposed to different E. bovis sporozoite preparations and antigens in the absence or presence of TLR antibodies were assessed for IL-8 secretion. Cells were exposed to sporozoite preparations and assessed for the activation of transcription factor NF-κB using a luciferase reporter assay. Immunofluorescence analysis was done to investigate TLR2 and TLR4 surface expression and NET formation on bovine PMN exposed to vital sporozoites. Results: we observed significantly increased TLR2 and TLR4 expression with a mean increase in expression that was greater for TLR2 than TLR4. This upregulation neither inhibited nor promoted sporozoite phagocytosis by bovine PMN. Live sporozoites together with anti-TLR2 mAb resulted in a significant enhancement of IL-8 production. NF-κB activation was more strongly induced in TLR2-HEK cells than in TLR4/MD2-HEK cells exposed to heat-killed sporozoites and antigens. Immunofluorescence analysis showed TLR-positive signals on the surface of PMN and concomitant NET formation. Conclusions: This is the first report on E. bovis-induced concomitant TLR2 and TLR4 expression during bovine PMN-derived NETosis.

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Fasciola hepatica induces weak NETosis and low production of intra- and extracellular ROS in exposed bovine polymorphonuclear neutrophils

Cited by 21 publications

References 57 publications

Metabolic Signatures of Cryptosporidium parvum-Infected HCT-8 Cells and Impact of Selected Metabolic Inhibitors on C. parvum Infection under Physioxia and Hyperoxia

Metabolic Signatures of Cryptosporidium parvum-Infected HCT-8 Cells and Impact of Selected Metabolic Inhibitors on C. parvum Infection under Physioxia and Hyperoxia

An Evaluation of the Fasciola hepatica miRnome Predicts a Targeted Regulation of Mammalian Innate Immune Responses

The Role of TLR2 and TLR4 in Recognition and Uptake of the Apicomplexan Parasite Eimeria bovis and Their Effects on NET Formation

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