Browsing by Author "Yipp, Bryan"
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- ItemOpen AccessAn Extract of the Tapeworm, Hymenolepis diminuta, Inhibits Neutrophil Migration in vitro(2017) Graves, Nicholas; McKay, Derek; Gilleard, John; Yipp, Bryan; Finney, ConstanceNeutrophils are not typically the first cell that comes to mind when the immune response to infection with helminths is considered. However, data are emerging showing that these cells may play an overlooked role in response to helminths perhaps early in the infection and particularly those species that cause tissue damage (e.g. Nippostrongylus brasiliensis - a gut nematode (roundworm)). Additionally, different helminth products have been shown to directly activate neutrophils in vitro (e.g. crude extracts of the porcine cestode Cysticercus cellulosae) and directly induce their chemotaxis (e.g. excretory/secretory products of the sheep nematode Haemonchus contortus). Hymenolepis diminuta crude extract (HdE) has been shown to inhibit dinitrobenzene sulfonic acid-induced colitis. Intraperitoneal injection of this crude extract recruited a population of peritoneal exudate cells dominated by neutrophils at 4h post-injection. Remarkably, these neutrophil-predominant PECs evoked increased production of the anti-inflammatory cytokine IL-10 from co-cultures of activated murine splenic CD4+ T cells. HdE did not directly induce murine neutrophil chemotaxis in vitro and direct application of HdE to neutrophils was not preferentially cytotoxic as assessed by trypan blue staining and LDH release. HdE did however induce Ca2+ mobilization and respiratory burst by murine neutrophils in vitro. In contrast to its ability to recruit neutrophils to the peritoneal cavity, HdE inhibited neutrophil chemotaxis in response to WKYMVm and keratinocyte derived chemokine (KC). HdE was unable to modulate neutrophil migration towards leukotriene B4. Biochemical characterizations (boiling, trypsinization, acidification, and sodium metaperiodate treatment) suggested that an acid-stable glycoprotein was responsible for its ability to block migration towards WKYMVm. The ability of HdE to block migration towards WKYMVm appears to be independent of the phosphorylation of the mitogen-activated protein kinase, p38. HdE did not directly induce production of the cytokines, IL-10, TNF-α, and KC. It also did not modulate production of these cytokines induced by other activators. In conclusion: (1) HdE activated neutrophils to mobilize Ca2+ into the cytoplasm and produce ROS; and, (2) an acid-stable glycoprotein component of the HdE inhibited neutrophil migration towards WKYMVm.
- ItemOpen AccessDifferentiating Sepsis-induced ARDS from Ventilated ICU Controls using Serum Metabolomics(2016) Donnelly, Sarah; Winston, Brent; Vogel, Hans; Yipp, Bryan; Stringer, KathleenSepsis-induced ARDS is a significant source of ICU morbidity and mortality with no treatments beyond supportive care. Biomarker identification may help predict development of ARDS, differentiate mild ARDS from moderate/severe ARDS, identify treatment targets, and prognosticate patient outcomes. Metabolomics is a systems biology approach to examine metabolic processes and metabolites in response to physiological stimuli, such as disease. Serum samples from ventilated ICU patients with sepsis or septic shock and ventilated ICU control patients were collected and stored in CCEPTR, an ICU tissue bank. 36 mild ARDS and 36 moderate/severe ARDS patients were age- and sex-matched to 36 ICU controls. Untargeted GC-MS and NMR were performed on ICU day 1 serum samples and last day samples of recovered patients. MVDA was used to identify diagnostic and prognostic biomarker candidates. We identified a candidate biopattern of sepsis-induced ARDS and predicted 28-day/hospital mortality of sepsis-induced ARDS using day 1 serum samples.
- ItemOpen AccessExploring the combinatorial effects of inflammatory stimuli and glucocorticoids on BIRC3 and BIRC2 expression: Differential regulation and functional investigations(2024-01-16) Thorne, Andrew James; Newton, Robert; Hirota, Simon; Yipp, BryanInflammation serves as a crucial innate mechanism in response to injury, infection, or harmful stimuli that activates the immune system and aims to restore homeostasis. Chronic inflammatory disorders like asthma present with ongoing inflammation whereby pulmonary epithelial cells release inflammatory mediators, intensifying airway inflammation, and worsening disease. Synthetic glucocorticoids represent the primary treatment for asthma by utilizing the anti-inflammatory properties of their endogenous counterparts to reduce inflammatory mediators. Their mechanism of action involves binding to the glucocorticoid receptor (GR) in cells and interacting with DNA elements to enhance anti-inflammatory gene expression. However, certain inflammatory genes play protective roles by promoting tissue repair or self-regulatory feedback mechanisms to limit inflammatory signaling. Some of these genes are spared the repressive effects of glucocorticoids and are even augmented, presumably for protection. The BIRC3 gene, while being associated with the inflammatory NF-κB pathway, is a gene upregulated in models of the pulmonary epithelium and in vivo by glucocorticoids. BIRC3, along with its family member BIRC2, may contribute to regulating NF-κB pathways, but also inhibit apoptosis, although their specific roles remain unclear. Consequently, the rationale of glucocorticoid-enhanced BIRC3 remain ambiguous. Given their similarities, BIRC3 and BIRC2 were both investigated, focusing on their expression, regulation, and the interactions between the GR and NF-κB at the gene promoter. These analyses detailed robust expression patterns differentiating BIRC3 and BIRC2 by their response to inflammatory cytokines and glucocorticoids over time. Promoter analysis revealed insights into possible GR and NF-κB interactions to explain the regulation of BIRC3 expression, thus providing an example of GR-NF-κB positive interactions leading to inflammatory gene expression. Further, investigations into the NF-κB pathway as a platform for BIRC2/3 function revealed the established canonical NF-κB pathway may challenge established dogma. This exploratory research highlighted the significance of TAK1 as central kinase within the NF-κB pathway. Moreover, possible redundancy between IKK1 and IKK2, and the presence of additional regulators within canonical NF-κB signaling may exist. While this study provided no greater insights for redundant, or individual roles for BIRC2/3 in NF-κB signaling or apoptosis, it facilitated the development of tools for future investigations.
- ItemOpen AccessMicrobiota-Immune Interactions and Host Defense in Sepsis(2023-04-18) Zucoloto, Amanda; McCoy, Kathy; McDonald, Braedon; Arrieta, Marie Claire; Yipp, BryanGut microbial dysbiosis is associated with increased susceptibility to infectious diseases, including systemic infection and sepsis. While the impact of the microbiota on intestinal homeostasis is well recognized, gut commensals also influence several processes in extraintestinal niches, including host defense against systemic infection. This thesis aimed to study the microbiota-immune interactions that promote host defense against bacterial sepsis at a cellular and molecular level. Using gnotobiotic mouse models and resonant scanning confocal intravital microscopy, I demonstrated that mice devoid of gut commensals show defective intravascular immune responses against staphylococcal sepsis. In germ-free mice, impaired Kupffer cell-mediated bacterial capture and neutrophil trafficking in the liver contribute to increased bacterial dissemination and host mortality. Gut microbiota-dependent regulation of intravascular immunity is dynamic and partly rescued by replenishing gut-derived D-lactate, indicating this bacterial metabolite mediates the crosstalk between the gut microbiota and the liver microenvironment. D-lactate enhances bacterial clearance by Kupffer cells and primes the liver endothelium to respond robustly to inflammatory stimuli, leading to increased neutrophil adherence after infection. Altogether, this thesis uncovers a line of communication between the gut microbiota and the liver microenvironment that is pivotal for intravascular immune defense against sepsis.
- ItemOpen AccessMolecular mechanisms of the CD36-plasmodium falciparum interaction in cytoadherence(2002) Yipp, Bryan; Ho, May; Kubes, Paul
- ItemEmbargoNeutrophil- and B cell-mediated host defense against viral and fungal respiratory infections(2024-03-21) Sarden, Nicole; Yipp, Bryan; Kelly, Margaret; McDonald, BraedonInfections of the lung continue to be a worldwide annual problem affecting millions of people. Fighting acute respiratory infections requires rapid and collaborative responses involving diverse immune cells. An interesting and unexplained clinical problem that sparked this thesis was the clinical observation that influenza A, a common respiratory virus, increases susceptibility to secondary infections with environmental fungi such as Aspergillus fumigatus (Af). By establishing a mouse model of viral-fungal co-infection and using clinical human samples, I discovered an essential collaboration between an underappreciated innate B lymphocyte, called B1a, which produced natural antibodies against Af, which tagged the fungi for elimination by neutrophils. Following influenza A infections, B1a cells died of apoptosis leading to diminished natural anti-Af antibodies and negating the ability of neutrophils to target and eliminate Af. Disruption of this axis explained susceptibility in viral- and steroid-associated infections. Delving deeper into the requirements of immune cell resistance to influenza and Af as individual infections, I discovered unique roles for neutrophils during influenza and innate B cells in aspergillosis. While there is ample research investigating neutrophils and their roles during infections with extracellular pathogens, it remained unclear how the neutrophil pool is affected following viral infections and subsequent functional outcomes. Additionally, it has been recently proposed that an interferon-responsive neutrophil state with unknown functions emerges following viral infections. In isolated viral infection, I found neutrophils adapt to an anti-viral phenotype which is spatially regulated, requires type I interferon licensing and sympathetic drive. On the other hand, using a genetically engineered mouse, we studied the specific roles of innate B1a cells during Aspergillus infection and found that this rare antibody producing cells was essential for host resistance against isolated Af infection. Moreover, I discovered key molecular requirements of the B1a cells that allow them to traffic and recruit properly in the lung, and I discovered that these cells can be amplified to improve host defense against Af. Altogether, this body of work uncovers neutrophils’ effects following isolated viral infection and during co-infection and demonstrates the tight interconnectedness between neutrophils and B lymphocytes which are critical for anti-fungal immunity.
- ItemOpen AccessPseudomonas aeruginosa Exopolysaccharide Biosynthesis and Immunosuppressive Role in Vivo(2016) Rich, Jacquelyn; Harrison, Joe; Morck, Doug; Yipp, BryanPseudomonas aeruginosa is a pathogen that can colonize the lungs of cystic fibrosis patients leading to chronic infections that are thought to have biofilm etiology. This opportunistic bacterium produces three exopolysaccharides found in the biofilm matrix: PEL, PSL and alginate. In this thesis, understanding the synthesis and expression of PEL was conducted by developing complementation vectors for each gene in the pel operon, as well as proposed genes involved in precursor synthesis. These vectors can be manipulated to learn more about how the proteins function to contribute to PEL expression, as shown using the PelA complementation vector to dissect individual residue functions. Results also suggest that the epimerases, GalE and PA4068, contribute to the synthesis of PEL. It was also found that in vivo, PEL and PSL act as immunosuppressive virulence factors, impairing neutrophil activation, potentially leading to immune evasion through alteration in innate immune response.
- ItemOpen AccessThe regulation and function of viperin during rhinovirus infection(2022-12-19) Love, Michelle; Proud, David; Leigh, Richard; Grandvaux, Nathalie; Jenne, Craig; Hirota, Simon; Yipp, BryanThe airway epithelium is the primary site of rhinovirus replication and plays a pivotal role in orchestrating innate immune responses to respiratory viruses, including rhinovirus. Interfacing with inhaled material in the airway lumen, the airway epithelium experiences continuous onslaught from respiratory pathogens. To combat viral infection, cells of the airway epithelium express a range of pattern recognition receptors capable of recognizing diverse viral ligands. Detection of an infectious virus by one of these receptors rapidly upregulates an array of effector molecules, including pro-inflammatory cytokines, interferons and interferon stimulated genes. Viperin is one of the most highly upregulated interferon stimulated genes in rhinovirus infected airway epithelial cells. Viperin expression during viral infection has long been associated with direct antiviral effects across diverse viral infection and cell models, including in rhinovirus infected airway epithelial cells. Despite the long-standing knowledge that viperin expression helps to impedes rhinovirus replication in the airway epithelium, little is known about how it is upregulated in response to rhinoviruses or how viperin proceeds to impact airway epithelial cell biology during infection. In addition to confirming previously published findings demonstrating that viperin is induced in rhinovirus infected primary human airway epithelial cells and is dependent on viral replication, we also demonstrated induction of viperin in airway epithelial cells by interferons. We also highlight the importance of the type III interferon IL-29 as a potent inducer of viperin in the airway epithelium. Finally based on our findings we propose a novel feedback loop, according to this model detection of rhinoviruses by the cytosolic dsRNA receptor melanoma differentiation-associated protein 5 (MDA5), or IL-29 signalling, promotes IRF-1 mediated viperin transcription. Viperin then establishes a protein-protein interaction with MDA5 which results in enhanced MDA5 stability in RV infected cells. This leads to enhanced signalling through MDA5, as measured by increased CXCL10 gene expression and protein secretion. As we have also demonstrated that MDA5 signalling is involved in promoting viperin expression in response to RV we hypothesize that stabilization of MDA5 by viperin may also serves to amplify viperin production during infection.
- ItemOpen AccessThe Role of B-cells in Regulating Pulmonary Neutrophils in vivo(2018-01-17) Kim, Jung Hwan; Yipp, Bryan; Kubes, Paul; Patel, Kamala; Eksteen, BertusNeutrophils, the major innate immune cell, are short-lived and typically considered as pro-inflammatory. As such, its pre-dominant role is to clear infections via the release of cytotoxic granules and phagocytosis. The production of neutrophils is regulated in the bone marrow, which generates about 1011 neutrophils per day in humans. Newly produced neutrophils circulate and transit through all organs receiving blood flow, seeking out infections and inflammatory signals. During homeostasis, neutrophils age overtime and are thought to be cleared from the system by macrophages in lymphoid organs. However, despite t extensive research, a complete life-cycle of neutrophils, including the aging process and regulation of neutrophil clearance, remains poorly understood. Interestingly, the pulmonary vasculature contains a significant amount of retained neutrophils that are higher concentration than their circulating counterpart. These marginated neutrophils were described to be retained within the pulmonary vasculature via CXCR4, a marker that is up-regulated on aged neutrophils. Therefore, we hypothesized that margination of neutrophils in the lung might represent a key intermediate step in the regulation and elimination of neutrophils. In this body of work, we discovered lung neutrophils display an aged phenotype where they express a higher level of CD18 and CXCR4 than the peripheral pool. Interestingly, these aged neutrophils bind pulmonary circulating B-cells via CD18. Moreover, such intercellular interaction between the two cells induces apoptosis in neutrophils that also depends on CD18, leading to the removal of subsequent apoptotic neutrophils by macrophages. B-cell depletion resulted in neutrophil inflammation leading to interstitial lung disease (ILD), and was attenuated by B-cell adoptive transfer or neutrophil depletion. During a model of rheumatoid arthritis (RA), B-cell sufficient animals were protected from neutrophil pulmonary inflammation; however, CD19-deficient mice, which have defective B-cell populations and response regulation, developed lung neutrophilic inflammation in addition to arthritis. Thus, we have defined a new regulatory role of B-cells to control neutrophils in pulmonary capillaries and found that the lung is a key site for neutrophil life-cycle.
- ItemOpen AccessThe Dynamic Role of Pulmonary Neutrophils in the Fungal Clearance and Immunopathology During Systemic Candidiasis in vivo(2017) Lee, Esther Kyung Su; Yipp, Bryan; Mody, Christopher; Proud, DavidNeutrophils are key players in the host immune response against systemic infection with C. albicans, which is the fourth most common bloodstream infection in North America. Using intravital microscopy, we investigated the role of marginated pulmonary neutrophils in the host defense against disseminated C. albicans. We found that this unique population of neutrophils rapidly clears the fungus that sequesters to the pulmonary vasculature in a C5a-dependent fashion. This uptake of C. albicans by the neutrophils then leads to the formation of dense, motile clusters in the vasculature via LTB4 amplification. The formation and persistence of the neutrophil vascular clusters are independent of beta-2 integrins, platelets, and NETs. Although complement-mediated uptake of the fungus is required for the host survival, subsequent neutrophil cluster formation contributes to disease pathology. Therefore, we see that the rapid neutrophilic response in the lung against disseminated candidiasis is a dynamic, double-edged process.
- ItemOpen AccessXenobiotic receptor regulation of Clostridium difficile associated inflammation and tissue damage(2017) Erickson, Sarah Lindsay; Hirota, Simon Andrew; Yipp, Bryan; Giembycz, MarkThe current treatments available for C. difficile infections (CDI) aims to eliminate C. difficile bacteria, however they do not treat the inflammation and tissue damage triggered by the virulence factors, TcdA and TcdB. Indeed, enhanced chemokine/cytokine expression during CDI is associated with poor clinical outcomes. The xenobiotic receptor, pregnane X receptor (PXR) can be activated by antibiotics which are known to target C. difficile. The PXR has also been reported to attenuate intestinal inflammation by inhibiting NFκB signaling. Therefore we hypothesized that targeting the PXR would be an effective strategy to reduce C. difficile toxin-induced inflammation and tissue damage. Activation of the PXR attenuated C. difficile toxin-induced inflammatory mediator release in Caco-2 intestinal epithelial cells (IECs) and in mouse enteroid derived colonic monolayers. In vivo studies demonstrated that PXR activation using the pharmacological agonist pregnenolone-16-α-carbonitrile (PCN) attenuated the expression of pro-inflammatory mediators and significantly reduced neutrophil infiltration to the colonic lamina propria following toxin challenge. Additionally, when compared to wild type mice, PXR-/- mice challenged with toxin displayed a hyper-susceptible phenotype with significant eosinophilic inflammation and increased pro-inflammatory gene expression. Contrary to our hypothesis, this phenotype was not mediated by the loss of the PXR in the epithelium, as determined by the use of a PXRfl/fl x villin cre+ knockout mouse. Taken together, these data will help us expand our knowledge of the PXR’s role in host-pathogen interactions, and its potential to be modulated for the treatment of inflammation and tissue damage in CDI.