Joao H. F. Pedra

Sensing Pathogens and Danger Signals by the Inflammasome

The NLR (nucleotide-binding domain leucine-rich repeat containing) family of intracellular sensors is a crucial component of the innate immune system. A number of NLR family members can form multiprotein complexes, called inflammasomes, and are capable of activating the cysteine protease caspase-1 in response to a wide range of stimuli including both microbial and self-molecules. Caspase-1 activation leads to processing and secretion of the proinflammatory cytokines interleukin-1beta (IL-1beta) and IL-18, which play crucial roles in host defense to infectious insults. Dysregulation of the inflammasome has also been linked to a number of autoinflammatory and autoimmune disorders. Recent advances in the inflammasome field will be discussed in this review.

Sterol Regulatory Element Binding Protein 2 Activation of NLRP3 Inflammasome in Endothelium Mediates Hemodynamic-Induced Atherosclerosis Susceptibility

Background— The molecular basis for the focal nature of atherosclerotic lesions is poorly understood. Here, we explored whether disturbed flow patterns activate an innate immune response to form the NLRP3 inflammasome scaffold in vascular endothelial cells via sterol regulatory element binding protein 2 (SREBP2). Methods and Results— Oscillatory flow activates SREBP2 and induces NLRP3 inflammasome in endothelial cells. The underlying mechanisms involve SREBP2 transactivating NADPH oxidase 2 and NLRP3. Consistently, SREBP2, NADPH oxidase 2, and NLRP3 levels were elevated in atheroprone areas of mouse aortas, suggesting that the SREBP2-activated NLRP3 inflammasome causes functionally disturbed endothelium with increased inflammation. Mimicking the effect of atheroprone flow, endothelial cell–specific overexpression of the activated form of SREBP2 synergized with hyperlipidemia to increase atherosclerosis in the atheroresistant areas of mouse aortas. Conclusions— Atheroprone flow induces NLRP3 inflammasome in endothelium through SREBP2 activation. This increased innate immunity in endothelium synergizes with hyperlipidemia to cause topographical distribution of atherosclerotic lesions.

Tick salivary secretion as a source of antihemostatics.

Ticks are mostly obligatory blood feeding ectoparasites that have an impact on human and animal health. In addition to direct damage due to feeding, some tick species serve as the vectors for the causative agents of several diseases, such as the spirochetes of the genus Borrelia causing Lyme disease, the virus of tick-borne encephalitis, various Rickettsial pathogens or even protozoan parasites like Babesia spp. Hard ticks are unique among bloodfeeders because of their prolonged feeding period that may last up to two weeks. During such a long period of blood uptake, the host develops a wide range of mechanisms to prevent blood loss. The arthropod ectoparasite, in turn, secretes saliva in the sites of bite that assists blood feeding. Indeed, tick saliva represents a rich source of proteins with potent pharmacologic action that target different mechanisms of coagulation, platelet aggregation and vasoconstriction. Tick adaptation to their vertebrate hosts led to the inclusion of a powerful protein armamentarium in their salivary secretion that has been investigated by high-throughput methods. The resulting knowledge can be exploited for the isolation of novel antihemostatic agents. Here we review the tick salivary antihemostatics and their characterized functions at the molecular and cellular levels.

ASC/PYCARD and Caspase-1 Regulate the IL-18/IFN-γ Axis during Anaplasma phagocytophilum Infection

Anaplasma phagocytophilum is an obligate intracellular pathogen that resides within neutrophils and can cause fever, pancytopenia, or death. IFN-γ plays a critical role in the control of A. phagocytophilum; however, the mechanisms that regulate IFN-γ production remain unclear. In this study, we demonstrate that apoptotic specklike protein with a caspase-activating recruiting domain (ASC)/PYCARD, a central adaptor molecule in the Nod-like receptor (NLR) pathway, regulates the IL-18/IFN-γ axis during A. phagocytophilum infection through its effect on caspase-1. Caspase-1- and asc-null mice were more susceptible than control animals to A. phagocytophilum infection due to the absence of IL-18 secretion and reduced IFN-γ levels in the peripheral blood. Moreover, caspase-1 and ASC deficiency reduced CD4+ T cell-mediated IFN-γ after in vitro restimulation with A. phagocytophilum. The NLR family member IPAF/NLRC4, but not NALP3/NLRP3, was partially required for IFN-γ production in response to A. phagocytophilum. Taken together, our data demonstrate that ASC and caspase-1 are critical for IFN-γ-mediated control of A. phagocytophilum infection.

Modulation of host immunity by tick saliva

UNLABELLED Next generation sequencing and proteomics have helped to comprehensively characterize gene expression in tick salivary glands at both the transcriptome and the proteome level. Functional data are, however, lacking. Given that tick salivary secretions are critical to the success of the tick transmission lifecycle and, as a consequence, for host colonization by the pathogens they spread, we thoroughly review here the literature on the known interactions between tick saliva (or tick salivary gland extracts) and the innate and adaptive vertebrate immune system. The information is intended to serve as a reference for functional characterization of the numerous genes and proteins expressed in tick salivary glands with an ultimate goal to develop novel vector and pathogen control strategies. SIGNIFICANCE We overview all the known interactions of tick saliva with the vertebrate immune system. The provided information is important, given the recent developments in high-throughput transcriptomic and proteomic analysis of gene expression in tick salivary glands, since it may serve as a guideline for the functional characterization of the numerous newly-discovered genes expressed in tick salivary glands.

Fucosylation enhances colonization of ticks by Anaplasma phagocytophilum

Fucosylated structures participate in a wide range of pathological processes in eukaryotes and prokaryotes. The impact of fucose on microbial pathogenesis, however, has been less appreciated in arthropods of medical relevance. Thus, we used the tick‐borne bacterium Anaplasma phagocytophilum– the agent of human granulocytic anaplasmosis to understand these processes. Here we show that A. phagocytophilum uses α1,3‐fucose to colonize ticks. We demonstrate that A. phagocytophilum modulates the expression of α1,3‐fucosyltransferases and gene silencing significantly reduces colonization of tick cells. Acquisition but not transmission of A. phagocytophilum was affected when α1,3‐fucosyltransferases were silenced during tick feeding. Our results uncover a novel mechanism of pathogen colonization in arthropods. Decoding mechanisms of pathogen invasion in ticks might expedite the development of new strategies to interfere with the life cycle of A. phagocytophilum.

All For One and One For All on the Tick–Host Battlefield

The saliva of ixodid ticks contains a mixture of bioactive molecules that target a wide spectrum of host defense mechanisms to allow ticks to feed on the vertebrate host for several days. Tick salivary proteins cluster in multigenic protein families, and individual family members display redundancy and pluripotency in their action to ameliorate or evade host immune responses. It is now clear that members of different protein families can target the same cellular or molecular pathway of the host physiological response to tick feeding. We present and discuss our hypothesis that redundancy and pluripotency evolved in tick salivary immunomodulators to evade immune recognition by the host while retaining the immunomodulatory potential of their saliva.

Sialomes and Mialomes: A Systems-Biology View of Tick Tissues and Tick–Host Interactions

Tick saliva facilitates tick feeding and infection of the host. Gene expression analysis of tick salivary glands and other tissues involved in host-pathogen interactions has revealed a wide range of bioactive tick proteins. Transcriptomic analysis has been a milestone in the field and has recently been enhanced by next-generation sequencing (NGS). Furthermore, the application of quantitative proteomics to ticks with unknown genomes has provided deeper insights into the molecular mechanisms underlying tick hematophagy, pathogen transmission, and tick-host-pathogen interactions. We review current knowledge on the transcriptomics and proteomics of tick tissues from a systems-biology perspective and discuss future challenges in the field.

Ixodes scapularis saliva mitigates inflammatory cytokine secretion during Anaplasma phagocytophilum stimulation of immune cells

BackgroundIxodes scapularis saliva enables the transmission of infectious agents to the mammalian host due to its immunomodulatory, anesthetic and anti-coagulant properties. However, how I. scapularis saliva influences host cytokine secretion in the presence of the obligate intracellular rickettsial pathogen Anaplasma phagocytophilum remains elusive.MethodsBone marrow derived macrophages (BMDMs) were stimulated with pathogen associated molecular patterns (PAMPs) and A. phagocytophilum. Cytokine secretion was measured in the presence and absence of I. scapularis saliva. Human peripheral blood mononuclear cells (PBMCs) were also stimulated with Tumor Necrosis Factor (TNF)-α in the presence and absence of I. scapularis saliva and interleukin (IL)-8 was measured.ResultsI. scapularis saliva inhibits inflammatory cytokine secretion by macrophages during stimulation of Toll-like (TLR) and Nod-like receptor (NLR) signaling pathways. The effect of I. scapularis saliva on immune cells is not restricted to murine macrophages because decreasing levels of interleukin (IL)-8 were observed after TNF-α stimulation of human peripheral blood mononuclear cells. I. scapularis saliva also mitigates pro-inflammatory cytokine response by murine macrophages during challenge with A. phagocytophilum.ConclusionsThese findings suggest that I. scapularis may inhibit inflammatory cytokine secretion during rickettsial transmission at the vector-host interface.

The tick salivary protein sialostatin L2 inhibits caspase-1-mediated inflammation during Anaplasma phagocytophilum infection.

ABSTRACT Saliva from arthropod vectors facilitates blood feeding by altering host inflammation. Whether arthropod saliva counters inflammasome signaling, a protein scaffold that regulates the activity of caspase-1 and cleavage of interleukin-1β (IL-1β) and IL-18 into mature molecules, remains elusive. In this study, we provide evidence that a tick salivary protein, sialostatin L2, inhibits inflammasome formation during pathogen infection. We show that sialostatin L2 targets caspase-1 activity during host stimulation with the rickettsial agent Anaplasma phagocytophilum. A. phagocytophilum causes macrophage activation and hemophagocytic syndrome features. The effect of sialostatin L2 in macrophages was not due to direct caspase-1 enzymatic inhibition, and it did not rely on nuclear factor κB or cathepsin L signaling. Reactive oxygen species from NADPH oxidase and the Loop2 domain of sialostatin L2 were important for the regulatory process. Altogether, our data expand the knowledge of immunoregulatory pathways of tick salivary proteins and unveil an important finding in inflammasome biology.