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Title:Hypoxic immunomodulation results in increased disease risk and altered behavior via non-canonical pathways
Author(s):Gainey, Stephen Joseph
Director of Research:Freund, Gregory G
Doctoral Committee Chair(s):Freund, Gregory G
Doctoral Committee Member(s):Beever, Jonathan E; Salak-Johnson, Janeen L; Horn, Gavin P
Department / Program:Animal Sciences
Discipline:Animal Sciences
Degree Granting Institution:University of Illinois at Urbana-Champaign
Degree:Ph.D.
Genre:Dissertation
Subject(s):hypoxia
high-fat
neuroinflammation
caspase-1
memory impairment
novel object
cognition
anxiety
glyburide
adenosine
Abstract:Hypoxia is a potent immunomodulatory condition that underlies a wide array of disease states and associated comorbidities. Inflammation appears to be a critical factor in the adverse psychological outcomes linked with hypoxia/reoxygenation injury. A causative role for brain-based IL-1 has been suggested in hypoxia-induced cognitive impairment and anxiety-like behaviors. However, the exact mechanism responsible has yet to be fully understood. Therefore, in this study we examined the impact of acute hypoxia on hippocampal-independent memory recall and particular cellular localization of inflammasome activity within the amygdala. Mice were exposed to hypoxic (6% oxygen/94% nitrogen) or normoxic (ambient) conditions for a period of 2 hours. Following 2 hours of reoxygenation, mice showed 65% increased caspase-1 activity in the amygdala by ICV-IHC determination with no significant changes observed within the hippocampus. To determine the extent of inflammasome activation in hypoxia/reoxygenation injury, drugs targeting potential activators such as K+ efflux and A2A adenosine receptor were administered. After glyburide (K+ efflux antagonist) and 3,7-dimethyl-1-prop-2-ynyl-purine-2,6-dione (DMPX-A2A adenosine receptor antagonist) administration, both drugs significantly reduced the level of caspase-1 activation in the amygdala correlating with the restoration of performance on the novel object recognition memory (NOR) task and thus mitigating hypoxia-induced retrograde amnesia. To determine the cellular localization of caspase-1 activation within the amygdala, a dual stain protocol was developed and showed that hypoxia induced significantly greater co-localization of caspase-1 and neurons. This result demonstrates the contribution of neuronal cells to the increased activation of caspase-1 following hypoxia/reoxygenation. Interestingly, IGF-I neuron-specific knockout mice showed similarly reduced caspase-1 activation in the amygdala and correction of NOR performance. In addition, hypoxia/reoxygenation significantly increased serine phosphorylation of insulin-receptor substrate 1 (IRS-1) and p70 S6 Kinase (p70SK) in whole brain. However, no change in IGF-I, protein kinase A (PKA), or cyclic AMP (cAMP) was observed in either the amygdala or hippocampus. These results suggest that hypoxia-induced retrograde amnesia is dependent on caspase-1 activation in the amygdala via a neuronal K+ efflux, A2A adenosine receptor, and IGF-I associated mechanism. Next, we examined a real-life scenario in which hypoxic conditions have been observed such as that displayed in obesity or following chronic high-fat diet (HFD) intake. In this study, mice were fed a HFD from weaning for 1, 3 or 6 weeks. Similar to hypoxia, mice displayed impaired NOR performance after 1 and 3 weeks of HFD consumption when compared to low-fat diet (LFD) controls but surprisingly recovered performance after 6 weeks of diet. In a hippocampal-dependent task, object location recognition (OLR), HFD consumption did not impair cognition until 3 and 6 weeks of diet. Additionally, mice displayed anxiety-like behaviors by measure of both the open-field and elevated zero maze tests following 3 weeks of HFD intake but remained comparable to LFD controls at 1 and 6 weeks of diet. These HFD-associated 3 week impairments correlated with the suppression of glutathione (GSH):glutathione disulfide (GSSG) increase in both the amygdala and hippocampus. Glyburide, a second generation sulfonylurea used for type 2 diabetes and K+ efflux antagonism, restored HFD-induced NOR impairment and alleviated anxiety-like behaviors in mice fed HFD for 3 weeks following a single dose. In addition, glyburide administration increased the HFD GSH:GSSG ratio within the amygdala but not the hippocampus. These findings suggest that obesity-associated childhood cognitive and psychopathologies can be corrected by targeting the blocking ATP-sensitive K+ channels with drugs such as glyburide. Finally, we looked to investigate another real-life scenario that places firefighters and first responders at risk for both the acute- and long-term due to hypoxic exposure during various firefighting activities. A rising concern amongst this group is increased cancer incidence associated with carcinogen and asphyxiate contact during firefighting. Therefore, in order to investigate the increased disease risk associated with smoke inhalation, mice were placed in overhaul, the period following suppression where knockdown of fire-materials occurs, after a live-fire scenario for 15 minutes following an interior or transitional tactical choice. Lung tissue was then harvested 2 hours post-overhaul for RNA-seq analysis. The transitional tactical choice reduced environmental significantly by a 10.3% reduction in average temperature when compared to the interior attack. This result correlated with a marked reduction in the number of significantly differentially expressed genes in the transitional tactical choice compared to the interior. Interestingly, interior attack resulted in up-regulation of a number of carcinogenic genes corresponding with down-regulation of numerous immunomodulatory genes. Disease pathways signaling through rap1 signaling, natural killer cell mediated cytotoxicity, and asthma were over-represented within the interior tactic. These results indicate that overhaul exposure after an interior tactical choice show immunosuppression with the advancement of cancer-associated genes whereas the transitional choice mitigates these risks. In conclusion, hypoxia/reoxygenation classically induces retrograde amnesia via increased neuronal-derived caspase-1 through K+ efflux, A2A adenosine receptor, and IGF-I activity. But real-life scenarios of hypoxia, including a high-fat challenge and smoke inhalation, subsequently induce memory impairment, anxiety-like behaviors, and cancer risk via non-canonical caspase-1 independent pathways.
Issue Date:2017-02-28
Type:Thesis
URI:http://hdl.handle.net/2142/97658
Rights Information:Copyright 2017 Stephen Joseph Gainey
Date Available in IDEALS:2017-08-10
Date Deposited:2017-05


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