RORA regulates neutrophil migration and activation in zebrafish

Neutrophil migration and activation are essential for defense against pathogens. However, this process may also lead to collateral tissue injury. We used microRNA overexpression as a platform and discovered protein-coding genes that regulate neutrophil migration. Here we show that miR-99 decreased the chemotaxis of zebrafish neutrophils and human neutrophil-like cells. In zebrafish neutrophils, miR-99 directly targets the transcriptional factor RAR-related orphan receptor alpha (roraa). Inhibiting RORα, but not the closely related RORγ, reduced chemotaxis of zebrafish and primary human neutrophils without causing cell death, and increased susceptibility of zebrafish to bacterial infection. Expressing a dominant-negative form of Rorα or disrupting the roraa locus specifically in zebrafish neutrophils reduced cell migration. At the transcriptional level, RORα regulates transmembrane signaling receptor activity and protein phosphorylation pathways. Our results, therefore, reveal previously unknown functions of miR- 99 and RORα in regulating neutrophil migration and anti-microbial defense.

ER-Phagy and Microbial Infection

The endoplasmic reticulum (ER) is an essential organelle in cells that synthesizes, folds and modifies membrane and secretory proteins. It has a crucial role in cell survival and growth, thus requiring strict control of its quality and homeostasis. Autophagy of the ER fragments, termed ER-phagy or reticulophagy, is an essential mechanism responsible for ER quality control. It transports stress-damaged ER fragments as cargo into the lysosome for degradation to eliminate unfolded or misfolded protein aggregates and membrane lipids. ER-phagy can also function as a host defense mechanism when pathogens infect cells, and its deficiency facilitates viral infection. This review briefly describes the process and regulatory mechanisms of ER-phagy, and its function in host anti-microbial defense during infection.

The Role of Toll-like Receptors (TLRs) Mediated Inflammation in Pancreatic Cancer Pathophysiology

Pancreatic cancer (PC) is one of the most lethal forms of cancer, characterized by its aggressiveness and metastatic potential. Despite significant improvements in PC treatment and management, the complexity of the molecular pathways underlying its development has severely limited the available therapeutic opportunities. Toll-like receptors (TLRs) play a pivotal role in inflammation and immune response, as they are involved in pathogen-associated molecular patterns (PAMPs) and danger-associated molecular patterns (DAMPs). Activation of TLRs initiates a signaling cascade, which in turn, leads to the transcription of several genes involved in inflammation and anti-microbial defense. TLRs are also deregulated in several cancers and can be used as prognostic markers and potential targets for cancer-targeted therapy. In this review we discuss the current knowledge about the role of TLRs in PC progression, focusing on the available TLRs-targeting compounds and their possible use in PC therapy.

Immunosuppression Affects Neutrophil Functions: Does Calcineurin-NFAT Signaling Matter?

Neutrophils are innate immune cells with important roles in antimicrobial defense. However, impaired or dysregulated neutrophil function can result in host tissue damage, loss of homeostasis, hyperinflammation or pathological immunosuppression. A central link between neutrophil activation and immune outcomes is emerging to be the calcineurin-nuclear factor of activated T cells (NFAT) signaling pathway, which is activated by neutrophil detection of a microbial threat via pattern recognition receptors and results in inflammatory cytokine production. This potent pro-inflammatory pathway is also the target of several immunosuppressive drugs used for the treatment of autoimmune disorders, during solid organ and hematopoietic cell transplantations, and as a part of anti-cancer therapy: but what effects these drugs have on neutrophil function, and their broader consequences for immune homeostasis and microbial defense are not yet known. Here, we bring together the emerging literature describing pathology- and drug- induced neutrophil impairment, with particular focus on their effects on calcineurin-NFAT signaling in the innate immune compartment.

Disruption of Intestinal Homeostasis Through Altered Responses of the Microbial Community, Energy Metabolites, and Immune System in Zebrafish After Chronic Exposure to DEHP

Di-(2-ethylhexyl) phthalate (DEHP) is ubiquitously reported in global water bodies and exhibits various environmental and human health risks. However, the effects of DEHP chronic exposure on the intestinal microbiota and associated host health concerns in aquatic species are still largely unexplored. In this study, chronic exposure to DEHP at environmental levels significantly increased the body weight, length, and body mass index (BMI), especially in male fish. The microbial community was disrupted with the relative abundance of phylum Firmicutes and genera diversity for Prevotella-7, Deefgea, PeM15, Halomonas, Akkermansia, Chitinibacter, and Roseomonas, which are significantly activated in zebrafish after exposure to DEHP. The height of the gut villus, the thickness of muscularis layer, and the number of goblet cells per villus were significantly decreased, as well as showed differences between female and male zebrafish. Further, the levels of energy-related metabolites in gut tissues were increased, compared to the control group. The expression levels of immune-related genes (interleukin 8, il-8, also referred to as cxcl8a), microbial defense-related genes (lysozyme, lyz, interleukin 10, and il-10), and obesity-related genes (aquaporin 8a, aqp8, mucin 2.1, muc2.1, fibroblast growth factor 2, fgf2, and proopiomelanocortin a, pomca) were significantly up-regulated in zebrafish, except the down-regulated expressions of toll-like receptor-5 (tlr-5) and interleukin 1β (il-1β) in the females and pomca in the males, respectively. Importantly, Spearman’s correlation analyses revealed that the levels of metabolites and gene expressions in the gut were closely related to the dominant microbial genera, such as Aeromonas, Deefgea, Akkermansia, PeM15, Mycobacterium, and Rhodobacter. Taken together, chronic exposure to DEHP at environmental levels disturbed bacterial composition accompanied by the altered expressions of intestinal metabolites and the critical immune and intestinal function-related genes, which provided novel insights into DEHP effects on perturbation of gut microbiota and metabolic homeostasis in zebrafish.

Significance of Mast Cell Formed Extracellular Traps in Microbial Defense

Mast cells (MCs) are critically involved in microbial defense by releasing antimicrobial peptides (such as cathelicidin LL-37 and defensins) and phagocytosis of microbes. In past years, it has become evident that in addition MCs may eliminate invading pathogens by ejection of web-like structures of DNA strands embedded with proteins known together as extracellular traps (ETs). Upon stimulation of resting MCs with various microorganisms, their products (including superantigens and toxins), or synthetic chemicals, MCs become activated and enter into a multistage process that includes disintegration of the nuclear membrane, release of chromatin into the cytoplasm, adhesion of cytoplasmic granules on the emerging DNA web, and ejection of the complex into the extracellular space. This so-called ETosis is often associated with cell death of the producing MC, and the type of stimulus potentially determines the ratio of surviving vs. killed MCs. Comparison of different microorganisms with specific elimination characteristics such as S pyogenes (eliminated by MCs only through extracellular mechanisms), S aureus (removed by phagocytosis), fungi, and parasites has revealed important aspects of MC extracellular trap (MCET) biology. Molecular studies identified that the formation of MCET depends on NADPH oxidase-generated reactive oxygen species (ROS). In this review, we summarize the present state-of-the-art on the biological relevance of MCETosis, and its underlying molecular and cellular mechanisms. We also provide an overview over the techniques used to study the structure and function of MCETs, including electron microscopy and fluorescence microscopy using specific monoclonal antibodies (mAbs) to detect MCET-associated proteins such as tryptase and histones, and cell-impermeant DNA dyes for labeling of extracellular DNA. Comparing the type and biofunction of further MCET decorating proteins with ETs produced by other immune cells may help provide a better insight into MCET biology in the pathogenesis of autoimmune and inflammatory disorders as well as microbial defense.

Human MAIT cells are devoid of alloreactive potential: prompting their use as universal cells for adoptive immune therapy

ABSTRACTBackgroundMucosal associated invariant T (MAIT) cells are semi-invariant T cells that recognize microbial antigens presented by the highly conserved MR1 molecule. MAIT cells are predominantly localized in the liver and barrier tissues and are potent effectors of anti - microbial defense. MAIT cells are very few at birth and accumulate gradually over a period of about 6 years during infancy. The cytotoxic potential of MAIT cells, as well as their newly described regulatory and tissue repair functions, open the possibility of exploiting their properties in adoptive therapy. A prerequisite for their use as “universal” cells would be a lack of alloreactive potential, which remains to be demonstrated.MethodsWe used ex vivo, in vitro and in vivo models to determine if human MAIT cells contribute to allogeneic responses.ResultsWe show that recovery of MAIT cells after allogeneic hematopoietic stem cell transplantation recapitulates their slow physiological expansion in early childhood, independent of recovery of conventional T cells. In vitro, signals provided by allogeneic cells and cytokines do not induce sustained MAIT cell proliferation. In vivo, human MAIT cells do not expand nor accumulate in tissues in a model of T-cell mediated xenogeneic graft-versus-host disease (GVHD) in immunodeficient mice.ConclusionsAltogether, these results provide evidence that MAIT cells are devoid of alloreactive potential and pave the way for harnessing their translational potential in universal adoptive therapy overcoming barriers of HLA disparity.

Role of Kupffer Cells in Systemic Anti-Microbial Defense

The liver has long been recognized as important in digestion. However, the liver’s abundance of innate immune cells strongly suggests that it has specific defense mechanisms. A characteristic anatomical feature of the liver is its large blood flow. The blood flowing out from the whole alimentary tract is transported to the liver via the portal vein and distributed to peripheral structures called sinusoids. Kupffer cells, a typical example of resident macrophages, are located in sinusoids and are in continuous contact with various portal blood components. They have vigorous phagocytic activity and eliminate bacteria coming from the gut before they enter systemic circulation. Based on this framework, Kupffer cells were considered a filter for portal blood pathogens. However, recent evidence reveals that they exert crucial functions in systemic host defense against bacterial infection. To defend against various sources of bacterial pathogens, Kupffer cells construct an efficient surveillance system for systemic circulation, cooperating aggressively with other immune cells. They collaborate with non-immune cells such as hepatocytes and platelets to potentiate defense function. In conclusion, Kupffer cells coordinate immune cell activity to efficiently defend against infections, making them crucial players in systemic antibacterial immunity.

Bioinformatic mapping of a more precise Aspergillus niger degradome

Aspergillus niger has the ability to produce a large variety of proteases, which are of particular importance for protein digestion, intracellular protein turnover, cell signaling, flavour development, extracellular matrix remodeling and microbial defense. However, the A. niger degradome (the full repertoire of peptidases encoded by the A. niger genome) available is not accurate and comprehensive. Herein, we have utilized annotations of A. niger proteases in AspGD, JGI, and version 12.2 MEROPS database to compile an index of at least 232 putative proteases that are distributed into the 71 families/subfamilies and 26 clans of the 6 known catalytic classes, which represents ~ 1.64% of the 14,165 putative A. niger protein content. The composition of the A. niger degradome comprises ~ 7.3% aspartic, ~ 2.2% glutamic, ~ 6.0% threonine, ~ 17.7% cysteine, ~ 31.0% serine, and ~ 35.8% metallopeptidases. One hundred and two proteases have been reassigned into the above six classes, while the active sites and/or metal-binding residues of 110 proteases were recharacterized. The probable physiological functions and active site architectures of these peptidases were also investigated. This work provides a more precise overview of the complete degradome of A. niger, which will no doubt constitute a valuable resource and starting point for further experimental studies on the biochemical characterization and physiological roles of these proteases.