Browsing by Subject "Inflammation"
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Item Characterizing the Role of the E. Coli Cytochrome Oxidase AppBCX During Intestinal Inflammation(August 2021) Chanin, Rachael Beth; Sperandio, Vanessa; Winter, Sebastian E.; Pfeiffer, Julie K.; Moreland, JessicaDuring non-infectious colitis, oral antibiotic treatment, and enteric infection, changes in colonocyte metabolism allow for increased oxygen availability in the gut lumen, supporting the growth of facultative anaerobic bacteria primarily from the family Enterobacteriaceae. Additionally, recruitment of inflammatory cells, especially neutrophils, has also been shown to influence local oxygen levels. The oxidative burst mounted by neutrophils consumes oxygen, which in turn creates a hyper-hypoxic microenvironment. These two seemingly contradictory findings remain to be reconciled. The findings described here delineate an additional mechanism that helps explain these seemingly contradictory observations on oxygen availability and bacterial respiratory processes during non-infectious colitis. The picture emerging from our work is that reactive oxygen species (ROS) generated by the NADPH oxidase 1 (NOX1) at the epithelial interface serve as a local source of oxygen. Specifically, H2O2 deriving from epithelial NOX1, is detoxified via bacterial catalases to molecular oxygen. Facultative anaerobic bacteria can then respire this pool of oxygen. In this work, we have used commensal strains of E. coli as representative members of Enterobacteriaceae, a family of facultative anaerobic bacteria that is observed to outgrow during episodes of inflammation. In particular, we have studied the physiological function of the cytochrome bd oxidase, AppBCX. Using both chemical and genetic models of non-infectious colitis we have shown that it allows E. coli to utilize low levels of oxygen early in inflammation. Additionally, we have characterized the regulation of this enzyme in vivo highlighting the delicate balance between growth and survival during oxidative and nitrosative stress.Item A Chemically Induced Colitis Screen Reveals the Necessity for Membrane Traffic in Intestinal Homeostasis(2019-03-21) McAlpine, William Elliott; Winter, Sebastian E.; Beutler, Bruce; Burstein, Ezra; Schmid, Sandra; Scherer, PhilippInflammatory bowel disease is most commonly a complex disorder caused by the interaction of environmental and genetic aberrations. Under normal conditions, a genetic program actively prevents inflammatory bowel disease, preventing invasion of microbes without permitting severe inflammation of the gut. To identify genes that maintain this balance, we performed a sensitized screen of 49,420 third generation (G3) germline mutant mice derived from N-ethyl-N-nitrosourea-mutagenized grandsires, bearing 104,658 coding/splicing mutations. We induced mild mucosal damage in these mice by orally administering dextran sodium sulfate (DSS) and found mutations that led to diarrhea and weight loss under these conditions. Causative mutations were mapped concurrently with screening using an automated mapping procedure. Among 114 DSS phenotypes identified and mapped, 36 have been validated by CRISPR/Cas9 targeting. Three vesicle trafficking genes, Myo1d, Smcr8, and Tvp23b, were selected for mechanistic evaluation. MYO1D is a class I myosin that binds both actin and lipid. MYO1D localizes to the basolateral membrane of enterocytes and functions in the intestinal epithelium to protect against colitis. SMCR8, along with C9ORF72 and WDR41, is a member of a tripartite complex that functions as a guanine exchange factor. SMCR8 localizes to the lysosome, and its absence results in perturbations to endocytic and phagocytic pathways. Hyperactivation of endosomal Toll-like receptors in Smcr8-/- mice causes spontaneous inflammation, and hyperactivation of multiple pathways contributes to DSS susceptibility. TVP23B is a trans-Golgi protein that binds YIPF6. Both TVP23B and YIPF6 are necessary for the formation of secretory granules in goblet and Paneth cells of the intestinal epithelium. These studies reveal non-redundant molecules required for the return of normal physiologic balance within the intestine after DSS insult.Item Dysbiosis-Associated Changes in Host Metabolism Produce Lactate to Support Enterobacterial Expansion During Inflammation(2019-04-04) Gillis, Caroline Catherine; Hendrixson, David R.; Winter, Sebastian E.; Sperandio, Vanessa; Burstein, EzraThe lumen of the gastrointestinal tract is heavily colonized by microbes, termed the gut microbiota. Under normal conditions, fermentative anaerobes constitute the majority of the gut microbiota. However, during inflammation there is a change in the nutritional environment of the gut that enables the outgrowth of facultative aerobic Enterobacteriaceae through respiratory metabolism. Salmonella enterica serovar Typhimurium (S. Tm) is a pathogenic member of the Enterobacteriaceae family that benefits from inflammation. We found that S. Tm uses lactate as a nutrient during infection, which maximizes colonization of the gut. During S. Tm infection, a profound change in the microbial community of the gut occurs. In particular, butyrate-producing Clostridia species are depleted. Butyrate is the preferred substrate for β-oxidation by intestinal epithelial cells (IEC). In the absence of butyrate, IEC perform a fermentative metabolism that produces lactate as a waste product. Lactate is then used in conjunction with oxygen as a terminal electron acceptor to support growth of S. Tm in the murine gut lumen. We next investigated the regulation of lactate utilization in S. Tm. We found that the lactate utilization genes (lldPRD), were inducible by electron acceptors and L-lactate. The transcriptional response to L-lactate was coordinated by the regulatory protein LldR, which maximized colonization of the murine gut. Under anaerobic conditions, lldPRD expression was repressed by the two-component system ArcAB. Commensal members of the Enterobacteriaceae family also expand during non-infectious colitis. We investigated whether lactate was also produced during non-infectious colitis and if commensal Enterobacteriaceae could use this nutrient. Butyrate was depleted and lactate was abundant in a murine model of colitis. Metagenomic sequencing demonstrated that lactate dehydrogenase genes were more abundant in the microbiome of inflamed mice than control mice. We next began to characterize putative lactate dehydrogenases in E. coli. We identified several putative lactate dehydrogenases, however, their role in E. coli fitness requires further study. In conclusion, we identified an important host-derived nutrient that promotes S. Tm fitness during infection and may serve as a nutrient for commensal Enterobacteriaceae during non-infectious colitis. This illustrates the importance of nutrient acquisition for Enterobacteriaceae during inflammatory colonization of the gut.Item Engineered E. Coli That Detect and Respond to Gut Inflammation Through Nitric Oxide Sensing(2014-07-25) Archer, Eric Jeffry; Tu, Benjamin; Süel, Gürol M.; Mangelsdorf, David J.; Hooper, Lora V.; Gardner, Kevin H.Within the last several years, advances in synthetic biology have allowed for the development of re-programmed microorganisms that perform useful tasks in areas like fuel production, bioremediation, and medicine. Several engineered microorganisms are in pre-clinical development for the treatment of human diseases, but may face critical limitations that decrease their utility in medicine due to adverse events like sepsis, caused by the introduction of bacteria within patients. Here I describe the design, construction, and characterization of a synthetic genetic network that is intended for use by E. coli within lumen of the intestine, which is presumed to be a safer location than other tissues, such as blood, for the introduction of engineered microbes. The synthetic gene regulatory circuit described here regulates gene expression through the activation of a permanent DNA switch in response to nitric oxide produced by inducible nitric oxide synthase. The detection of nitric oxide initiates the expression of a DNA recombinase, causing the permanent genetic rearrangement of a short DNA segment containing a gene promoter, allowing for the regulation of output gene expression upon nitric oxide sensing. Here I demonstrate that E. coli containing this synthetic genetic circuit respond to nitric oxide as designed from both chemical nitric oxide donors and from injured mouse intestinal explants. This synthetic genetic circuit could be optimized for clinical use by allowing E. coli to reliably detect and treat inflammation in patients with inflammatory bowel disease, but the circuit described herein now serves as the proof-of-concept for both bacterial sensing of mammalian inflammation and for the use of DNA recombinases to translate transient environmental signals into permanent responses in engineered bacteria.Item Inflammasome-Independent Interleukin-1 Beta Drives CD4 T Cell Effector Function and Autoimmune Inflammation(2019-03-26) Jain, Aakanksha; Hooper, Lora V.; Pasare, Chandrashekhar; Malter, James; Conrad, Nicholas; Satterthwaite, Anne B.Successful generation of protective immunity is a result highly orchestrated interactions between the innate and the adaptive arms of the immune system. Innate immune cells sense the infectious threat to the host and convert it into meaningful information that is relayed to T and B cells. This relaying of information is the fundamental principle that forms the basis of all protective immune responses and is referred to as the innate control of adaptive immunity. For naïve CD4 T cell activation, innate control is exerted by the three-signal paradigm which dictates that a productive naïve CD4 T cell response requires TCR activation, co-stimulation and innate cytokine cues. The present work uncovers two consequential yet previously unknown aspects of the innate and adaptive immune, specifically dendritic cell (DC)-T cell, crosstalk. First, I found that in addition to naïve CD4 T cell activation, the three-signal paradigm continue to operate during the reactivation phase of memory CD4 T cells. Memory CD4 T cell reactivation was presumed to be independent of innate cytokine cues however, I found that signaling through interleukin (IL)-1 receptor family is required for effector cytokine production by memory CD4 T cells. Mechanistically, IL-1R signaling provides post-transcriptional stability to T cell cytokine transcripts thereby enabling productive secretion of these cytokines. Second, I discovered that DC-T cell interaction is significantly more bidirectional than previously appreciated. Innate cells such as DCs, have evolved to sense "non-self" or "altered-self" ligands via pattern recognition receptors (PRRs) and get activated only in case of pathogen invasion or tissue damage. Surprisingly, I found that autoreactive T cells can also instruct DCs to become activated independent of PRR activation. During their cognate interaction, T cell engage Tumor necrosis factor superfamily (TNFRSF) signaling in DCs to trigger innate cytokine secretion. I particularly, focused on the mechanism of IL-1β production during DC-T cell interaction as IL-1β is mediator of several autoimmune and autoinflammatory diseases. While IL-1β production is commonly attributed to inflammasome activation in autoimmune diseases, I show that that autoreactive T cells elicit IL-1β production by DCs in an inflammasome-independent manner via TNFR-Fas signaling pathway. Furthermore, this novel mechanism of IL-1β secretion drives CD4 T cell effector function, systemic leukocyte infiltration as well as autoimmune inflammation. Altogether, the findings in this dissertation provide a conceptual leap in our understanding of innate and adaptive immune cross-talk, and necessitates revisiting the established paradigm of innate signaling requirements for myeloid cell activation as well as CD4 T cell function. In addition, this work has vast implications on human health as it sheds light on the previously poorly understood mechanism of action of IL-1R blockade therapy as well as offers novel targets for therapeutic intervention of IL-1β mediated autoimmune inflammation.Item Inflammatory Effects of Severe Burn Injury in Rat Intervertebral Disc(2017-01-17) Mitchell, Thomas Wes; Buller, Dustin; Manchanda, Kshitij; Song, Juquan; Hernandez, PaulaIMPORTANCE: Severe burn injury leads to a myriad of clinical effects, some of which are mediated by systemic inflammatory cytokines. Although inflammation has been shown to increase skeletal muscle catabolism, little is known about the effects this inflammation has on the intervertebral disc tissue and whether or not it predisposes to spinal pathology. OBJECTIVE: To investigate the gene expression changes after severe burn injury in rat intervertebral disc tissue and explore the possible role of inflammatory cytokines in tissue remodeling imbalance. METHODS: Rats were given severe burn injury according to protocol, then harvested at multiple time points: 6h, 1d, 3d, 7d, and 14d post-burn. Unburned rats used for control. Rat intervertebral discs were dissected and removed, then used for measurement of gene expression using qPCR. RESULTS: Differences in gene expression of structural matrix proteins between time points were not statistically significant, however, TNF-alpha showed statistically significant (p<.05) decreased expression at 1d, after increased expression at 6h that approached significance (p=.055); this pattern of gene expression is consistent with previous results. In addition, there was a statistically significant (p<.05) increase in expression of the TRPV4 channel, which helps regulates osmolarity of the tissue in the nucleus pulposis. This could explain how osmotic dysregulation induced by pro-inflammatory cytokines leads to loss of water content from nucleus pulposis cells, thus triggering disc degeneration. CONCLUSIONS: These results indicate that in the post-burn state pro-inflammatory cytokines, such as TNF-alpha, likely have an effect on intervertebral disc tissue and the subsequent up-regulation of TRPV4 could have long-term effects that hasten the onset of disc degeneration related to loss of water content.Item Investigating the Effects of Particle Radiation Exposure on Lung Carcinogenesis(2019-04-16) Luitel, Krishna; Shay, Jerry W.; Chen, Benjamin P.; Aroumougame, Asaithamby; Akbay, Esra A.Lung cancer accounts for more cancer-related deaths than any other cancer type among both men and women. The overall increase in radiation risk for human cancer types has been substantiated by the epidemiological data obtained from atomic bomb survivors and uranium mine workers. The lung has a large surface area which makes it a prominent target for radiation exposure making it susceptible to radiation-induced cancer. Recently particle radiation therapy such as the use of protons and carbon has increased in the treatment of cancer. The long-term biological effects of proton radiation remain less well characterized in terms of radiotherapy and well as for astronauts during deep space explorations. We compared the long-term side effects of proton radiation to equivalent doses of X-rays in the initiation and progression of premalignant lesions in a transgenic mouse lung cancer model (K-rasLA1). We show proton irradiation causes more complex DNA damage that is not completely repaired resulting in increased oxidative stress in the lungs both acutely and persistently. Proton irradiated mice had lower median survival and increased carcinoma incidence as compared to un-irradiated controls or X-ray exposed mice. Additionally, the space radiation environment consists of a wide variety of ion species with a various range of energies. To understand the effects of mixed ion beam radiation, we exposed K-rasLA-1 mice with three ion beams: Proton (H), Helium (He), and Silicon (Si) at a low dose rate of 0.5cGy/min. Using the three ion beams, we performed whole body irradiation in two different orders: 3B-1 (H-He-Si) and 3B-2 (Si-He-H) and used only H as a reference. We found that whole-body irradiation with 3B-1 increases the incidence of cancer initiation and systemic oxidative stress in mice 100 days post-irradiation compared to 3B-2 and H irradiation. Additionally, we saw an increase in adenomas with atypia and adenocarcinomas in 3B-1 irradiated mice but not in 3B-2 and H irradiated mice. We also found that a non-toxic anti-inflammatory, anti-oxidative radioprotector (CDDO-EA) reduced 3B-1 induced oxidative stress and cancer initiation almost back to baseline. Thus, exposure to 3B-1 elicits significant changes in lung cancer initiation that can be mitigated using CDDO-EA.Item Mechanisms Governing NF-κB Regulation by the Anti-Inflammatory Protein A20(2013-05-31) Skaug, Brian; Chen, Zhijian J.; Cobb, Melanie H.; Sternweis, Paul C.; Yarovinsky, FelixA20 is a potent anti-inflammatory protein that inhibits NF-ΚB, and A20 dysfunction is associated with autoimmunity and B-cell lymphoma. A20 harbors a deubiquitination enzyme domain and can employ multiple mechanisms to antagonize ubiquitination upstream of NEMO, a regulatory subunit of the IκB kinase complex (IKK). However, direct evidence of IKK inhibition by A20 is lacking, and the inhibitory mechanism remains poorly understood. Here we show that A20 can directly impair IKK activation without deubiquitination or impairment of ubiquitination enzymes. We find that polyubiquitin binding by A20, which is largely dependent on A20’s 7th zinc finger motif (ZnF7), induces specific binding to NEMO. Remarkably, this ubiquitin-induced recruitment of A20 to NEMO is sufficient to block IKK phosphorylation by its upstream kinase TAK1. Our results suggest a novel mechanism of IKK inhibition and a means by which polyubiquitin chains can specify a signaling outcome.Item New methods to treat inflammation in the rheumatic diseases(2001-12-13) Reimold, AndreasItem [News](1982-12-03) Friar, JamieItem The role of inflammation and infection in neurodegeneration(2024-01-12) Beckham, J. DavidItem Severe Remote Burn Injury Results in Early, Elevated Markers of Alzheimer's Disease(2013-01-22) Suleman, Leyya; Gatson, Joshua; Maass, David; Warren, Victoria; Wolf, Steven; Minei, Joseph; Pepe, Paul; Idris, Ahamed; Wigginton, JaneBACKGROUND: Prior studies have found that patients with severe burns may suffer neurocognitive decline. While these observations are frequently attributed to psycho-social causes, our lab recently reported that remote burn injury is associated with significant brain changes, including new data revealing a substantial, rapid and sustained (30 min - 45 day) increase in rat brain inflammation following remote burns. Other acute brain injury processes, such as traumatic brain injury and stroke have been associated with an accelerated accumulation of Aβ40, Aβ42,, and Tau, and ultimately a clinical picture of early-onset Alzheimer's disease (AD). We hypothesized that similar AD-like processes may be triggered in the brain following remote, severe burn injury. METHODS: In this study, 44 male rats received a 3° 40% TBSA back/flank scald burn by immersion (divided into 6 harvest time points), with an additional 8 receiving a sham burn (immersion in room temperature water), totaling 52 rats. Brains of those burned were harvested at 1, 6, 12, 24 hours, 7 days (n=8/each time point) and 45 days (n=4/time point) after injury. Brain tissue IL-6, TNF-α, IL-1β, Aβ40, Aβ42, total Tau and phosphorylated Tau were measured using ELISA methods. RESULTS: Burned animals had significantly increased markers of inflammation and AD at each time point measured compared with those receiving a sham burn injury (see table for data at 1 hour and 45 days). CONCLUSIONS: Severe remote burn injury not only results in early, robust, and sustained neuroinflammation, but also significantly increases brain levels of Aβ40, Aβ42, and Tau. This novel finding may pave the way for future brain-preserving interventional trials in burn patients, as well as provide a more rapid and effective testing-ground for new therapies aimed at slowing and/or preventing AD.Item [Unknown](1998-11-05) Stieglitz, HeatherItem [UT Southwestern Medical Center News](2010-09-14) Shear, Kristen HollandItem [UT Southwestern Medical Center News](2008-12-15) McKenzie, AlineItem [UT Southwestern Medical Center News](2010-12-01) Shear, Kristen Holland