Microbiology Commons^™

Il-33 Promotes Increased Replication Of Theiler’S Murine Encephalomyelitis Virus In Raw264.7 Macrophage Cells With An Irf3-Dependent Response, Ahmed Esmael, Thomas M. Petro

Nebraska Center for Virology: Faculty Publications

Interleukin-33 (IL-33), which promotes M2 macrophage development, may influence the control of viruses, such as Theiler’s Murine Encephalomyelitis Virus (TMEV) that infect macrophages. Because Interferon Regulatory Factor-3 (IRF3) is also critical to control of TMEV infection in macrophages, information on the relationship between IL-33 and IRF3 is important. Thus, RAW264.7 Lucia murine macrophage lineage cells with an endogenous IRF3-ISRE promoter driving secreted luciferase and IRF3KO RAW Lucia, a subline deficient in IRF3, were challenged with TMEV. After the challenge, considerable TMEV RNA detected at 18 and 24 h in RAW cells was significantly elevated in IRF3KO RAW cells. TMEV induction …

Il-33 Promotes Increased Replication Of Theiler’S Murine Encephalomyelitis Virus In Raw264.7 Macrophage Cells With An Irf3-Dependent Response, Ahmed Esmael, Thomas M. Petro

Nebraska Center for Virology: Faculty Publications

Interleukin-33 (IL-33), which promotes M2 macrophage development, may influence the control of viruses, such as Theiler’s Murine Encephalomyelitis Virus (TMEV) that infect macrophages. Because Interferon Regulatory Factor-3 (IRF3) is also critical to control of TMEV infection in macrophages, information on the relationship between IL-33 and IRF3 is important. Thus, RAW264.7 Lucia murine macrophage lineage cells with an endogenous IRF3-ISRE promoter driving secreted luciferase and IRF3KO RAW Lucia, a subline deficient in IRF3, were challenged with TMEV. After the challenge, considerable TMEV RNA detected at 18 and 24 h in RAW cells was significantly elevated in IRF3KO RAW cells. TMEV induction …

Effects Of Surface Topography On Macrophages And Bacterial Cells, Joseph Carnicelli

Theses - ALL

An association has been found between the texture of breast implants and anaplastic large cell lymphoma, which led to some textured implants to be withdrawn from the market in 2019. There is evidence that these cancers are associated with the harboring of bacteria on the surfaces of the textured implants. It is possible that specific topographic features hinder the removal of attached bacteria by inhibiting macrophage phagocytosis or promoting biofilm formation. Here we examine how bacteria and macrophages interact with recessive surface topographies as analogs to the surfaces seen on textured breast implants. Changes in bacteria morphology were observed among …

Developing A Microdialysis Sampling-Based Biofilm/Macrophage Co-Culture Model, Alda Diaz Perez

Graduate Theses and Dissertations

The host immune system and bacterial cells are known to interact during the human lifetime. Bacteria secrete a wide variety of signaling molecules, known as quorum sensing (QSC) molecules, that modulate the host immune system. While immune-biofilm interactions involve this chemical signaling network, the mechanisms through which this occurs are not well understood. This work aimed to develop a new method that can be used not only in vitro settings but also in vivo. The microdialysis sampling technique has widely been used in in vitro and in vivo settings in humans, mice, and rats for the collection of neuropeptides, cytokines, …

Examining Early Interactions Between Innate Airway Resident Immune Cells And Mtb-Specific Factors During Pulmonary Infection With Mycobacterium Tuberculosis, Micah D. Dunlap

Arts & Sciences Electronic Theses and Dissertations

Mycobacterium tuberculosis (Mtb) is the leading cause of death by an infectious agent in the world today, infecting roughly one quarter of humans. Despite this, the mechanisms of early pathogenesis and host protective innate immune responses remain poorly understood and uncharacterized.

Lung resident Alveolar Macrophages (AMs) are the first host contact with Mtb bacilli after inhalation and are thus key mediators of the early pulmonary immune response. AMs are generally believed to reside entirely in the airway, but it was recently demonstrated that they have the capacity to egress and enter into granulomas during pulmonary infection with hypervirulent Mtb. Furthermore, …

Evaluation Of Rhodococcus Equi Susceptibility To Silver Nanoparticle Antimicrobials, Elizabeth Boudaher

Theses and Dissertations--Veterinary Science

Rhodococcus equi is a significant cause of pneumonia in foals and immunocompromised humans. Antimicrobial resistance among R. equi isolates has developed as a consequence of inappropriate stewardship and bacterial evolution, leading to an increased rate of treatment failures that typically result in foal fatality. In the current study, we evaluated the efficacy of antimicrobial silver nanoparticle (AgNP) complexes in controlling R. equi growth. Previous studies characterizing AgNP-induced antibacterial effects in other Gram-positive pathogens led us to hypothesize that silver nanoparticle antimicrobials impact R. equi viability and intracellular replication. We therefore investigated the effect of silver nanoparticle complexes on R. equi …

The Histoplasma Capsulatum Ddr48 Gene Is Required For Survival Within Macrophages And Resistance To Oxidative Stress And Antifungal Drugs, Logan Blancett

Dissertations

Histoplasma capsulatum(Hc)is a systemic, dimorphic fungal pathogen that affects upwards of 500,000 individuals in the United States annually. Hc grows as a multicellular mold at environmental temperatures; whereas, upon inhalation into a human or other mammalian host, it transforms into a unicellular, pathogenic yeast. The research presented in this dissertation is focused on characterizing the DNA damage-responsive gene HcDDR48. HcDDR48was originally isolated via a subtractive DNA library enriched for transcripts enriched in the mold-phase of Hcgrowth. Upon further analysis we found that HcDDR48is not just expressed in the mold morphotype, but both growth programs …

Host Mediated Mechanisms Of Fungal Cell Spread In A Transparent Zebrafish Infection Model, Allison Scherer

Electronic Theses and Dissertations

Innate immunity has developed elegant processes for the detection and clearance of invasive fungal pathogens. Disseminated candidiasis is of significant concern for those with suppressed immune systems or indwelling medical equipment, and mortality in these groups approaches 70%. Poor patient outcomes have spurred the need to understand how this non-motile pathogen spreads in the host. Technical limitations have previously hindered our ability to visualize the role of innate immunity and host tissue barriers in the spread of C. albicans in vivo. Using the zebrafish model to overcome these limitations, we have examined three potential host-mediated mechanisms of dissemination: movement …

Sodium Pyruvate Alters The Immune Response To Influenza A Virus Infection In Macrophages, Hazzar Abysalamah

MSU Graduate Theses

ABSTRACT

Pyruvate is the end product of glycolysis. It can either be transported into the mitochondria for use in the TCA cycle or be used to regenerate NAD⁺ during fermentation or aerobic glycolysis (also called the Warburg Effect). I recently discovered that addition of sodium pyruvate to the culture medium during infection of macrophages with influenza A virus affects the production of cytokines involved in immune signaling. While infection of macrophages with influenza A virus resulted in high levels of cytokines (IL-6, IL-1β, and TNF-α) in the absence of sodium pyruvate, the addition of sodium pyruvate significantly impaired cytokine …

Tat Controls Transcriptional Persistence Of Unintegrated Hiv Genome In Primary Human Macrophages., Beatrix Meltzer, Deemah Dabbagh, Jia Guo, Fatah Kashanchi, Mudit Tyagi, Yuntao Wu

Medicine Faculty Publications

In HIV infected macrophages, a large population of viral genomes persists as the unintegrated form (uDNA) that is transcriptionally active. However, how this transcriptional activity is controlled remains unclear. In this report, we investigated whether Tat, the viral transactivator of transcription, is involved in uDNA transcription. We demonstrate that de novo Tat activity is generated from uDNA, and this uDNA-derived Tat (uTat) transactivates the uDNA LTR. In addition, uTat is required for the transcriptional persistence of uDNA that is assembled into repressive episomal minichromatin. In the absence of uTat, uDNA minichromatin is gradually silenced, but remains highly inducible by HDAC …

Nutritional Virulence Of Legionella Pneumophila., Ashley M. Best

Electronic Theses and Dissertations

Legionella pneumophila is an environment organism that parasitizes a wide range of protozoa. Growth within the environmental host primes L. pneumophila for infection of human alveolar macrophages when contaminated aerosols are inhaled. Intracellular replication within either host requires the establishment a replicative niche, known as the Legionella-containing vacuole (LCV). Biogenesis of the LCV depends on the type IVb translocation system, the Dot/Icm, to translocation >320 effectors into the host cytosol. Effectors are responsible for preventing lysosome fusion to the LCV, recruitment of ER-derived vesicles to the LCV, and modulation of a plethora of host processes to promote the intracellular …

The Effect Of Gram-Positive Staphylococcus Aureus Cell Wall Components Lipoteichoic Acid And Peptidoglycan On Cytokine Production, Cytoskeletal Arrangement, And Cell Viability On Raw 264.7 Murine Macrophages, Gabrielle String

Browse all Theses and Dissertations

In this study, gram positive Staphylococcus aureus cell wall components such as lipoteichoic acid (LTA) and peptidoglycan (PGN) were used to study the potential inflammatory and anti-inflammatory cytokine response, cytoskeletal arrangement and cell viability on RAW264.7 murine macrophages over 24 hours. The effect of S.aureus LTA and PGN (5 µg/mL) on RAW 264.7 macrophages was evaluated every six hours for twenty-four hours. Inflammatory cytokine (TNF-a) production peaked at 6 hours before decreasing over time. Anti-inflammatory cytokine (IL-10) production peaked between 12 and 18 hours. During the first twenty-four hours, cytotoxicity of treated macrophages, as defined as the release of lactate …

The Effect Of Gram-Positive Staphylococcus Aureus Cell Wall Components Lipoteichoic Acid And Peptidoglycan On Cytokine Production, Cytoskeletal Arrangement, And Cell Viability On Raw 264.7 Murine Macrophages, Gabrielle String

Browse all Theses and Dissertations

In this study, gram positive Staphylococcus aureus cell wall components such as lipoteichoic acid (LTA) and peptidoglycan (PGN) were used to study the potential inflammatory and anti-inflammatory cytokine response, cytoskeletal arrangement and cell viability on RAW264.7 murine macrophages over 24 hours. The effect of S.aureus LTA and PGN (5 [mu]g/mL) on RAW 264.7 macrophages was evaluated every six hours for twenty-four hours. Inflammatory cytokine (TNF-a) production peaked at 6 hours before decreasing over time. Anti-inflammatory cytokine (IL-10) production peaked between 12 and 18 hours. During the first twenty-four hours, cytotoxicity of treated macrophages, as defined as the release of lactate …

Characterization Of The Ato Gene Family In Alternative Carbon Metabolism, Heather A. Danhof

Dissertations & Theses (Open Access)

As a commensal colonizer and opportunistic pathogen, Candida albicans is the most clinically important human associated fungus. Systemic infection carries an unacceptably high mortality rate of ~40% in the growing population of immunocompromised individuals. Macrophages are important innate immune cells that limit the niches in the human body in which C. albicans can persist through phagocytic removal. However, following phagocytosis C. albicans readily escapes from the immune cell by differentiating into filamentous hyphae, a process that should be inhibited in the normally acidic phagolysosome. We have shown that C. albicans induces germination by neutralizing the phagolysosome. To better understand this …

The Expression Of Major Histocompatibility Class I And Major Histocompatibility Class Ii On Macrophages In The Presence Of Aryl Hydrocarbon Antagonist (Ch-223191), Caitlin Wilson

Browse all Theses and Dissertations

Macrophages are crucial for ridding the body of debris and foreign cells. The aryl hydrocarbon receptor (AhR) also plays a critical role in immunity. This study examined the effect of the AhR on the expression of major histocompatiability complex class I (MHCI) and MHC class II (MHCII) in two murine macrophage cell lines. This study used Raw264.7 and J774A.1 murine macrophage cell lines. The Raw264.7 cells are from male BALB/c mice while the J774A.1 cells are from female BALB/cN mice. The addition of the AhR anatagonist CH-223191 (AhRa) showed that the AhR does not significantly impact MHCI expression. However, MHCII …

Colony Stimulating Factor-1 Receptor (Csf-1r) Expression, Degradation, And Signaling In Lipid-Laden Macrophages, Gregory Thompson

Electronic Theses and Dissertations

Atherosclerosis is a serious disease affecting a large portion of the world’s population. In atherosclerosis, macrophages become filled with lipid droplets and form fatty plaques in artery walls. The formation of these atherosclerotic plaques is dependent on the macrophage growth factor colony stimulating factor-1 (CSF-1); however, understanding the changes in CSF-1 signaling and related pathways in foam cells remain poorly investigated. Lipid-laden macrophages were formed through exposure of murine bone marrow derived macrophages to 100 μg/ml acetylated LDL for 24 h. Non-LDL treated macrophages were grown in parallel for each experiment and used for comparison. Immunofluorescent staining was used to …

Granulocyte Colony-Stimulating Factor: Its Role In Gut-Homing Macrophage Generation And Colitis, And Production By Probiotics, Shahab Meshkibaf

Electronic Thesis and Dissertation Repository

The pleiotropic cytokine granulocyte-colony stimulatory factor (G-CSF) is mainly required for the generation of neutrophils, but its role in macrophage generation has also been reported. In addition, G-CSF is effective for the down-regulation of inflammatory cytokines and ameliorating gut disorders, such as colitis. However, the G-CSF function in macrophage generation and gut immunity remains unclear. The first focus of this thesis was to assess the role of G-CSF in macrophage generation and its contribution to gut immunity. G-CSF was found to promote the generation of Gr-1high/F4/80+ macrophages in macrophage (M)-CSF-treated bone marrow cells, most likely through suppressing cell death. Gr-1high …

Regulation Of The Candida Albicans Arginine Biosynthetic Pathway, Claudia Jimenez Lopez

Dissertations & Theses (Open Access)

Candida albicans is the most importan thuman-associatedfungus.It is a commensal microorganism but also an opportunistic pathogen able to cause superficial infections aswellaslife-threateninginfectionswhich are associated with a highmortalityrateof 50%. The interactions between C. albicans and the cells of the mammalian innate immune system, which confer the most important protecting mechanisms against disseminated infections, are very dynamic and determine the success of C. albicans as a pathogen. Transcriptional profiling has shown that phagocytosis of C. albicans by macrophages results primarily in the activation of alternative carbon metabolism pathways suggesting that the pathogen is exposed to a glucose poor environment. Changes in …

Regulation Of Macrophage Inflammatory Signaling Pathways By Amp-Activated Protein Kinase., Yanfang Zhu

Electronic Theses and Dissertations

AMP-activated protein kinase, AMPK, is a conserved serine/threonine kinase with a critical function in the regulation of metabolic pathways in eukaryotic cells. Recently, AMPK has been shown to play an additional role as a regulator of inflammatory activity in leukocytes. Treatment of macrophages with chemical AMPK activators, or forced expression of a constitutively active form of AMPK, results in polarization to an antiinflammatory phenotype. Additionally, we reported previously that stimulation of macrophages with antiinflammatory cytokines such as IL-10, IL-4 and TGF-ß results in rapid activation of AMPK, suggesting that AMPK contributes to the suppressive function of these cytokines. In the …

Candida Albicans Induces Arginine Biosynthetic Genes In Response To Host-Derived Reactive Oxygen Species, Claudia Jimenez-Lopez, John R. Collette, Kimberly M. Brothers, Kelly M. Shepardson, Robert A. Kramer

Dartmouth Scholarship

The interaction of Candida albicans with phagocytes of the host's innate immune system is highly dynamic, and its outcome directly impacts the progression of infection. While the switch to hyphal growth within the macrophage is the most obvious physiological response, much of the genetic response reflects nutrient starvation: translational repression and induction of alternative carbon metabolism. Changes in amino acid metabolism are not seen, with the striking exception of arginine biosynthesis, which is upregulated in its entirety during coculture with macrophages. Using single-cell reporters, we showed here that arginine biosynthetic genes are induced specifically in phagocytosed cells. This induction is …

Toxoplasma Gondii Rhoptry Kinase Rop16 Activates Stat3 And Stat6 Resulting In Cytokine Inhibition And Arginase-1-Dependent Growth Control, Barbara A. Butcher, Barbara A. Fox, Leah M. Rommereim, Sung Guk Kim, Kirk J. Maurer, Felix Yarovinsky, De'broski R. Herbert, David J. Bzik, Eric Y. Denkers

Dartmouth Scholarship

The ROP16 kinase of Toxoplasma gondii is injected into the host cell cytosol where it activates signal transducer and activator of transcription (STAT)-3 and STAT6. Here, we generated a ROP16 deletion mutant on a Type I parasite strain background, as well as a control complementation mutant with restored ROP16 expression. We investigated the biological role of the ROP16 molecule during T. gondii infection. Infection of mouse bone marrow-derived macrophages with rop16-deleted (ΔROP16) parasites resulted in increased amounts of IL-12p40 production relative to the ROP16-positive RH parental strain. High level IL-12p40 production in ΔROP16 infection was dependent on the host …

Cd1d-Dependent B-Cell Help By Nk-Like T Cells Leads To Enhanced And Sustained Production Of Bacillus Anthracis Lethal Toxin-Neutralizing Antibodies, T. Scott Devera, Lindsay M. Aye, Gillian A. Lang, Sunil K. Joshi, Jimmy D. Ballard, Mark L. Lang

Bioelectrics Publications

The current Bacillus anthracis vaccine consists largely of protective antigen (PA), the protein of anthrax toxin that mediates entry of edema factor (EF) or lethal factor (LF) into cells. PA induces protective antibody (Ab)-mediated immunity against Bacillus anthracis but has limited efficacy and duration. We previously demonstrated that activation of CD1d-restricted natural killer-like T cells (NKT) with a CD1d-binding glycolipid led to enhanced Ab titers specific for foreign antigen (Ag). We therefore tested the hypothesis that activation of NKT cells with the CD1d ligand (alpha-galactosylceramide [alpha-GC]) at the time of immunization improves PA-specific Ab responses. We observed that alpha-GC enhanced …

Candida Albicans Cellwall Components And Farnesol Stimulate The Expression Of Both Inflammatory And Regulatory Cytokines In The Murine Raw264.7 Macrophage Cell Line, Suman Ghosh, Nina Howe, Katie Volk, Swetha Tati, Kenneth W. Nickerson, Thomas M. Petro

Kenneth Nickerson Papers

Candida albicans causes candidiasis, secretes farnesol, and switches from yeast to hyphae to escape from macrophages after phagocytosis. However, before escape, macrophages may respond to C. albicans’ pathogen-associated molecular patterns (PAMPs) through toll-like receptor 2 (TLR2) and dectin-1 receptors by expressing cytokines involved in adaptive immunity, inflammation, and immune regulation. Therefore, macrophages and the RAW264.7 macrophage line were challenged with C. albicans preparations of live wild-type cells, heat-killed cells, a live mutant defective in hyphae formation, a live mutant producing less farnesol, or an isolate producing farnesoic acid instead of farnesol. Interleukin-6 (IL-6), and IL-1b, IL- 10, and tumor …

Interdependent Regulation Of Cytomegalovirus Proteins In Complex, Lisa L. Bolin

Theses and Dissertations in Biomedical Sciences

Human cytomegalovirus (HCMV) is a ubiquitous herpesvirus that causes significant morbidity and mortality in the immune compromised. Using the mutine cytomegalovirus (MCMV) model, we identified two genes, M140 and M141, which are determinants of macrophage tropism. Monocytes/macrophages are a critical cell type for cytomegalovirus (CMV) pathogenesis as sites of viral latency and by supporting viral replication and disseminating virus throughout the body. We previously found that the localization of the M140/M141 gene products (pM140/pM141) is different when the proteins are co-expressed as compared to their individual expression. When the proteins are individually expressed pM141 localizes diffusely throughout the cytoplasm and, …

Protection Of Macrophages J774a.1 By Purine Nucleoside Analogues From Bacillus Anthracis Mediated Necrosis, Zadkiel R. Alvarez

UNLV Theses, Dissertations, Professional Papers, and Capstones

Eight years after the lamentable anthrax attacks, major scientific effort continues to be done, in order to stop imminent acts of bioterrorism. Innovative ways of therapy against the anthrax disease are being investigated. B. anthracis, the etiological agent of the infection, has a dormant stage in its life cycle known as the endospore. When conditions become favorable spores germinate, transforming into vegetative bacteria. In inhalational anthrax, the most fatal manifestation of the disease, spores enter the organism through the respiratory track, and are phagocytosized by alveolar macrophages of the lungs. Spores are able to sense nutrient availability, activating their germination …

Nitric Oxide Production: A Mechanism For Inhibition Of Chlamydia Trachomatis Replication, Bojun Chen

Electronic Theses and Dissertations

Chlamydia trachomatis (CT) replicates in macrophages, but is inhibited by IFN-$\gamma$ or LPS. IFN-$\gamma$ and/or LPS induced nitrite production in mouse peritoneal macrophages, macrophage cell lines (RAW264.7 and J774A.1) and McCoy cells. Kinetic studies indicated that peak production occurred 48 hours post-treatment. CT infection itself was insufficient to induce nitrite production, but resulted in enhancement of nitrite production in IFN-$\gamma$-treated cells. Treatment with IFN-$\gamma$ or LPS resulted in significant inhibition of CT replication in these cells. Strong correlation between nitrite production and inhibition of CT replication was observed in RAW264.7 and J774A.1 cells (correlation coefficients: $-$0.93 and $-$0.94, p $<$ 0.001). N$\sp{\rm g}$- monomethyl-L-arginine (L-NMMA) specifically inhibited nitrite production and partially reversed inhibition of CT replication in macrophage cell lines. NOS mRNA was measured in RAW264.7 cells by Northern blot and Dot blot hybridization. Strong correlation between NOS mRNA expression and inhibition of CT replication (correlation coefficient: $-$0.97, p $<$ 0.05) was observed. Anti-TNF-$\alpha$ antibody completely neutralized the biological activity of TNF-$\alpha$ secreted by LPS-treated RAW264.7 cells, yet the antibody neither reduced nitrite production nor restored CT replication. Combination of the antibody and L-NMMA significantly enhanced restoration of CT replication. In peritoneal macrophages, inhibition of CT replication induced by IFN-$\gamma$ was partially restored by L-NMMA or anti-TNF-$\alpha$ antibody. In McCoy cells, inhibition of CT replication induced by IFN-$\gamma$ and LPS was not significantly restored by L-NMMA. Great restoration of CT replication by 1 mM L-NMMA was observed in LPS-treated J774A.1 cells (31%), but not in IFN-$\gamma$-treated cells (5%). Our data indicate that (1) NO production is one of the mechanisms for inhibition of CT replication in IFN-$\gamma$-activated peritoneal macrophages and RAW264.7 cells; (2) NO plays a significant role in CT inhibition in LPS-treated macrophage cell lines, but not peritoneal macrophages; (3) TNF-$\alpha$ may be associated with inhibition, but the mechanism(s) may not involve NO production; (4) NO production may not be the mechanism for CT inhibition in McCoy cells treated with IFN-$\gamma$ and LPS.