Due to rapid urbanisation, people in metropolises spend the majority of their time indoors. Indoor mould contamination, as one of the most pungent biohazards in built environments, can ubiquitously present in humid areas and potentially compromise the health of occupants. Governmental institutions like the World Health Organisation and United States Environmental Protection Agency have put forward guidelines for indoor mould prevention. However, these guidelines normally require occupants to maintain a low indoor humidity (<75% or even 40%), and thus, in tropical and subtropical areas, one of the most widely used approaches to prevent indoor mould contamination is to continuously operate air-conditioners or dehumidifiers (AC/D). The 24 h operation of AC/D, however, conflicts with the requirement of energy sustainability, and hence posits a trade-off between sustainability and indoor mould hygiene. The aim of this study was to facilitate the development of sustainable and effective mould prevention strategies for indoor environments. The literature on currently adopted mould prevention strategies including that target moisture (24 h AC/D), temperature (air-conditioning system and cool wall paint) and nutrient (dust removal) elements as well as new nanoparticles technology (Ag, TiO 2 and MgO nanoparticles), was reviewed and the main limitations of these strategies were discussed. It was found that none of these current mould prevention measures has addressed both sustainability and mould hygiene on balance, urging further investigations. Therefore, the objective of the first phase investigation was to develop sustainable cause-specific mould control measures in built environments. A case study of a mould contaminated site was conducted to illustrate the micro-environments that contribute to mould contamination in buildings. The currently used 24-h AC/D approach was compared with and ranked against other sustainable alternatives. The results of this case study suggest that determining an effective non-24 h AC/D management regime tends to be a sustainable and user-friendly solution. To develop such a regime, understanding the critical mechanisms regulating indoor mould responses to water dynamics is essential. Thus, the objective of the second phase was to characterise the critical mechanism regulating the growth of common indoor moulds under water dynamics. It was hypothesised that oxidative stress is associated with the growth of indoor moulds under water dynamics. Using Cladosporium cladosporioides as a model, both its pre-germination and germinated spores were exposed to daily wet-dry cycles. Afterwards, the growth was assessed and cellular H 2 O 2 concentration and catalase activity were measured. It was found that under water dynamics, the longer growth delay in C. cladosporioides was associated with a higher encountered oxidative stress, with 12-12 wet-dry cycle (12 h wet, 12 h dry) showing the longest delay and highest oxidative stress. Pearson correlation and linear regression analysis suggest a positive correlation between growth delay and oxidative stress under water dynamics (R 2 =0.85, P<0.0001). Moreover, pre- germination spores generally exhibited shorter growth delay, lower cellular H 2 O 2 concentration and higher catalase activity. Collectively, these results suggest that the growth of C. cladosporioides is associated with oxidative stress under water dynamics. After revealing the association between the growth of C. cladosporioides and oxidative stress under water dynamics, at the third phase, this finding was extrapolated to different mould species (C. cladosporioides, Aspergillus niger and Aspergillus penicillioides), water activity (a w ) (0.4 a w , 0.6 a w and 0.8 a w ) and temperature levels (19 °C and 28 °C). In addition, the antioxidant responses of treated moulds, including antioxidant enzymes (superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase) were monitored. The results showed that lower water activity levels imposed higher oxidative stress to moulds, and A. penicillioides exhibited the highest tolerance which displayed the highest antioxidant activities and encountered lowest oxidative damage under water dynamics. Moreover, no significant difference was measured in terms of the survival, oxidative stress and antioxidant responses between these two temperature levels. The third phase of the study, for the first time discovered the reason contributing to the different resistance towards water dynamics among common indoor moulds, and further confirmed the important role of oxidative stress adaptation in withstanding transient water supply. In conclusion, this study reveals the critical role of oxidative stress adaptation in helping moulds to cope with changing water conditions, which may shed light on a new perspective for the future development of indoor mould prevention strategies. It also indicates that longer operation time of AC/D each day may not necessarily lead to better prevention of mould contamination, suggesting that in order to sustainably prevent mould contamination, one should operate reasonable non-24 h AC/D each day (12 h/day according to the examined species in this study) to yield a more stressful wet-dry cycle to moulds. The outcomes of this study foster the development of novel and sustainable indoor mould prevention strategies

Due to rapid urbanisation, people in metropolises spend the majority of their time indoors. Indoor mould contamination, as one of the most pungent biohazards in built environments, can ubiquitously present in humid areas and potentially compromise the health of occupants. Governmental institutions like the World Health Organisation and United States Environmental Protection Agency have put forward guidelines for indoor mould prevention. However, these guidelines normally require occupants to maintain a low indoor humidity (<75% or even 40%), and thus, in tropical and subtropical areas, one of the most widely used approaches to prevent indoor mould contamination is to continuously operate air-conditioners or dehumidifiers (AC/D). The 24 h operation of AC/D, however, conflicts with the requirement of energy sustainability, and hence posits a trade-off between sustainability and indoor mould hygiene. The aim of this study was to facilitate the development of sustainable and effective mould prevention strategies for indoor environments. The literature on currently adopted mould prevention strategies including that target moisture (24 h AC/D), temperature (air-conditioning system and cool wall paint) and nutrient (dust removal) elements as well as new nanoparticles technology (Ag, TiO 2 and MgO nanoparticles), was reviewed and the main limitations of these strategies were discussed. It was found that none of these current mould prevention measures has addressed both sustainability and mould hygiene on balance, urging further investigations. Therefore, the objective of the first phase investigation was to develop sustainable cause-specific mould control measures in built environments. A case study of a mould contaminated site was conducted to illustrate the micro-environments that contribute to mould contamination in buildings. The currently used 24-h AC/D approach was compared with and ranked against other sustainable alternatives. The results of this case study suggest that determining an effective non-24 h AC/D management regime tends to be a sustainable and user-friendly solution. To develop such a regime, understanding the critical mechanisms regulating indoor mould responses to water dynamics is essential. Thus, the objective of the second phase was to characterise the critical mechanism regulating the growth of common indoor moulds under water dynamics. It was hypothesised that oxidative stress is associated with the growth of indoor moulds under water dynamics. Using Cladosporium cladosporioides as a model, both its pre-germination and germinated spores were exposed to daily wet-dry cycles. Afterwards, the growth was assessed and cellular H 2 O 2 concentration and catalase activity were measured. It was found that under water dynamics, the longer growth delay in C. cladosporioides was associated with a higher encountered oxidative stress, with 12-12 wet-dry cycle (12 h wet, 12 h dry) showing the longest delay and highest oxidative stress. Pearson correlation and linear regression analysis suggest a positive correlation between growth delay and oxidative stress under water dynamics (R 2 =0.85, P<0.0001). Moreover, pre- germination spores generally exhibited shorter growth delay, lower cellular H 2 O 2 concentration and higher catalase activity. Collectively, these results suggest that the growth of C. cladosporioides is associated with oxidative stress under water dynamics. After revealing the association between the growth of C. cladosporioides and oxidative stress under water dynamics, at the third phase, this finding was extrapolated to different mould species (C. cladosporioides, Aspergillus niger and Aspergillus penicillioides), water activity (a w ) (0.4 a w , 0.6 a w and 0.8 a w ) and temperature levels (19 °C and 28 °C). In addition, the antioxidant responses of treated moulds, including antioxidant enzymes (superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase) were monitored. The results showed that lower water activity levels imposed higher oxidative stress to moulds, and A. penicillioides exhibited the highest tolerance which displayed the highest antioxidant activities and encountered lowest oxidative damage under water dynamics. Moreover, no significant difference was measured in terms of the survival, oxidative stress and antioxidant responses between these two temperature levels. The third phase of the study, for the first time discovered the reason contributing to the different resistance towards water dynamics among common indoor moulds, and further confirmed the important role of oxidative stress adaptation in withstanding transient water supply. In conclusion, this study reveals the critical role of oxidative stress adaptation in helping moulds to cope with changing water conditions, which may shed light on a new perspective for the future development of indoor mould prevention strategies. It also indicates that longer operation time of AC/D each day may not necessarily lead to better prevention of mould contamination, suggesting that in order to sustainably prevent mould contamination, one should operate reasonable non-24 h AC/D each day (12 h/day according to the examined species in this study) to yield a more stressful wet-dry cycle to moulds. The outcomes of this study foster the development of novel and sustainable indoor mould prevention strategies

Indiana University-Purdue University Indianapolis (IUPUI) / Type 1 and Type 2 diabetes (T1D/T2D) share a common etiology that involves an increase in oxidative stress that leads to dysfunction and subsequent β cell death. Lipoxygenases are enzymes that catalyze the oxygenation of polyunsaturated fatty acids to form lipid metabolites involved in a variety of biological functions including cellular oxidative stress response. On the other hand, Interleukin 6 (IL-6) signaling has been demonstrated to be protective in islets. In this study, we explored the effect of lipoxygenase enzymes 12-Lipoxygenase, 12/15 Lipoxygenase and IL-6 on β cell function and survival in mice using both STZ and high-fat diet (HFD) models of diabetes. Alox12-/- mice showed greater impairment in glucose tolerance following STZ and HFD compared to wild-type mice (WT), whereas Alox15-/- were protected against dysglycemia. These findings were accompanied by evidence of islet oxidative stress in Alox12-/- mice and reduced oxidative stress in Alox15-/- mice, consistent with alterations in the expression of antioxidant response enzymes in islets from these mice. Additionally, islets from Alox12-/- mice showed a compensatory increase in Alox15 gene expression and treatment of these mice with the 12/15-lipoxygenase inhibitor ML-351 rescued the dysglycemic phenotype. IL-6 was able to significantly attenuate the generation of reactive oxygen species by proinflammatory cytokines in human pancreatic islets. Furthermore, we find that IL-6 regulates the master antioxidant response protein NRF2. Collectively these results show that loss of Alox12 activates a compensatory increase in Alox15 that sensitizes β cells to oxidative stress and signaling by IL-6 is required for maximal antioxidant response under conditions of increased ROS formation, such as obesity.

No description available.

Listeria monocytogenes response to oxidative stress by sublethal sodium hypochlorite was investigated in this study. Continuous exposure of sublethal chlorine influenced biofilm formation and stress adaptation (homologous and heterologous) in L. monocytogenes. The biofilm forming ability of oxidative stress adapted and control cells were investigated on polystyrene surface at 22°C and 37°C. The oxidative stress adapted cells were found to form less biofilm in the presence of chlorine (p < 0.10) when compared to non-treated control cells at both the temperatures. In addition, the biofilm forming ability of L. monocytogenes was reduced significantly at higher sublethal chlorine concentrations (p < 0.10). In conclusion, oxidative stress adapted L. monocytogenes has developed tolerance to chlorine and some of the antibiotics. However, oxidative stress those cells did demonstrate an antibiofilm effect. This demonstrates that oxidative stress reduces L. monocytogenes biofilm formation but can also increase antibiotic resistance.

Little research has focused on the involvement of oxidative stress as it relates to the pathophysiology of osteoarthritis (OA); while inflammation has been extensively studied. The present study evaluates the ability to modulate the response of canine chondrocytes to both inflammation and oxidative stress in an in-vitro model. Chondrocytes were incubated and then stimulated to under-go oxidative stress by using hydrogen peroxide or inflammation using interleukin-1 beta and tumor necrosis factor alpha. For inhibition of oxidative stress an antioxidant, N-acetyl-cysteine, was used prior to induction with hydrogen peroxide in a subset of chondrocytes. Measures of oxidative stress were superoxide dismutase and reduced glutathione. Prostaglandin E2 was used as a measurement of inflammation. Chondrocytes responded appropriately to both oxidative stress and inflammation. The antioxidant N-acetyl-cysteine provided adequate protection against oxidative stress. Oxidative stress and inflammation should be considered to play a role in the pathophysiology of canine OA.

The purpose of this study was to induce an oxidative-stress adaptation in Listeria monocytogenes Bug600 (serotype 1/2a) and F1057 (serotype 4b) by pre-exposing to sublethal H2O2 and alkali-stress either singly or sequentially. Our findings show that the sequential pre-exposure of cells to pH 9 for 30 min treatment followed by 50 ppm H2O2 for 30 min at 37°C yielded the highest oxidative-stress resistant phenotypes of L. monocytogenes Bug600 and F1057. The sublethal H2O2 and sublethal alkali-stress induced oxidative-stress adaptations were completely reversible within 60 min at 37°C in the absence of such sublethal stress. However, the oxidative-stress adaptation induced at 37°C was stable at 4°C over a 24 h test period in both L. monocytogenes Bug600 and F1057. Future studies will focus on the potential cross-resistance of oxidative-stress adapted L. monocytogenes serotypes 1/2a and 4b to commonly used disinfectants and GRAS antimicrobials.

Background: The objective of this thesis was to measure oxidative stress (OS) in women with Bipolar Disorder (BD) who used combined oral contraceptives (OCU). Based on our literature review, it was predicted that OCU would increase OS levels relative to non-contraceptive users (NCU) in women. Methods: Thirty-five participants (BD n=25; Control n=10) were recruited from an ongoing study based in British Columbia ‘The Systematic Treatment Optimization Program in Early Mania.’ Participants were administered psychological screening tools (Young Mania Rating Scale (YMRS), Montgomery-Åsberg Depression Rating Scale, Mini International Neuropsychiatric Interview and Hamilton Depression Rating Scale) and provided a blood sample for the assays (Lipid Hydroperoxide (LPH), Protein Carbonylation, 4-Hydroxynonenal, 3-Nitrotyrosine (3-NT) and 17-Beta Estradiol). Results: In our primary analysis we did not find differences in OS between BD and controls relative to OCU. Within our remaining analyses, only BD women (n=17) and who gave smoking status were included. We found 3-NT to be increased in OCU compared to NCU (F (1, 12) = 5.639, p = 0.035). With respect to mood stabilizer use, 3-NT was increased in OCU relative to NCU (F (1, 10) = 6.33, p=0.031). As for atypical antipsychotics, 3-NT was heightened in OCU adjunctive users compared to NCU who did not use atypical antipsychotics (F (3, 10) = 4.822, p = 0.025). As for our correlation analyses, YRMS correlated with 3-NT and LPH in OCU BD women (r(11)= 0.711, p=0.014 and r(11) = 0.676, p=0.022, respectively) and 17-Beta Estradiol correlated with LPH (r(17) = 0.598, p = 0.001). Our results are preliminary and are limited by our small sample size and various other factors (i.e. controls). Conclusion: The association between hormones and oxidative stress still remains controversial. Here we showed, after controlling for smoking, BMI and age the use of a COC significantly increased 3-NT in women with BD. Moreover, hormones may influence the relationship between OS and mood episodes. / Thesis / Master of Science (MSc)

Omi/HtrA2 is a mitochondrial serine protease with a dual and opposite function depending on its subcellular localization. Most of the previous studies focused on Omi/HtrA2’s pro-apoptotic function when the protein is released to the cytoplasm. It is becoming apparent that the main function of Omi/HtrA2 is within the mitochondria, where it has a pro-survival role. However, its mechanism is still poorly understood. To this end, we used the yeast two-hybrid system to dissect the Omi/HtrA2 pathway by identifying novel interactors and substrates. Our studies revealed a novel function of Omi/HtrA2 in the regulation of a deubiquitinating (DUB) complex. In addition we found that Omi/HtrA2 participates in mitophagy by regulating Mulan E3 ubiquitin ligase, which recruits GABARAP (gamma-amino-butyric acid receptor-associated protein) to the mitochondria. Abro1 is the scaffold protein of the DUB complex known as BRISC (BRCC36 isopeptidase complex) that is specific for Lys-63 deubiquitination. This complex is similar to the BRCA-1 complex, a known and important player in DNA damage response. Using the yeast two-hybrid screen and a bait consisting of the unique carboxy-terminus of the Abro1 protein, we identified three transcription factors which are members of the activating protein 1 (AP-1) family, namely ATF4, ATF5 and JunD. The AP-1 family member ATF4 is ubiquitously expressed, like Abro1, and important in cell cycle regulation and survival, thus we further analyzed this interaction. Abro1’s interaction with ATF4 was specific and present only when cells are under cellular stress. When Abro1 protein level is increased it provides protection against stress-induced cell iv death, but interaction between Abro1 and ATF4 is necessary to achieve this protection. The significance of this interaction was the translocation of Abro1 from the cytoplasm to the nucleus. These results establish a new cytoprotective function of cytoplasmic Omi/HtrA2 as a regulator of the BRISC DUB complex. Under normal conditions Omi/HtrA2 is localized in the intermembrane space (IMS) of the mitochondria. We have recently identified that the mitochondrial Mulan E3 ubiquitin ligase is a substrate of Omi/HtrA2 protease. Mulan, along with MARCH5/MITOL and RNF185, are the only three mitochondrial E3 ubiquitin ligases identified thus far. The function of Mulan has been linked to cell growth, cell death and autophagy/mitophagy. In addition, we showed that Omi/HtrA2, through regulation of the Mulan protein level, controls mitophagy, especially during mitochondrial stress. To understand Mulan’s function and its control by Omi/HtrA2, we set out to identify E2 conjugating enzymes that form a complex with Mulan E3 ligase. We isolated four specific interacting E2’s, namely Ube2E2, Ube2E3, Ube2G2 and Ube2L3. To identify substrates for each unique Mulan-E2 complex we used fusion baits in a second yeast two-hybrid screening. One of the interactors isolated against the Mulan-Ube2E3 bait was the GABARAP protein, a member of the Atg8 (autophagy) family. The mammalian Atg8 family is composed of seven members that have been linked to important roles in autophagy/mitophagy. We characterized this interaction both in vitro and in vivo and its role in mitophagy. Our results suggest that Mulan participates in various pathways, depending on the nature of its partner E2 conjugating enzyme. In addition, we identified the pathway by which Mulan participates in mitophagy by recruiting GABARAP to the mitochondria. v I want to dedicate this hard work to the people who mean the most to me, the people who have been more than supportive in these past five years; they are my very small, but very loud and loving, family. My brother Raffaello has always been proud of me and always told his friends how his sister was a "doctor." My father, Alvaro, always told me I had to be better than him, a man who has a master’s degree in chemistry. Finally, but most importantly to me, I want to dedicate this work and degree to my mother, Maria Aparecida Troncon, a woman who is like no other. She has always supported me, and with small gestures like having snacks ready when I came to visit after work or coming with me to lab on the weekends so I would not be alone, she told me every day that she was proud of me. Every time she met someone she told them I was her doctor and she said it with the biggest smile on her face. Unfortunately she passed away on December 17th, 2012, just months shy of the completion of my degree. I remember how proud she was when I received my bachelor’s and I can only imagine the size of her smile when I walk across the stage this time as a Ph.D. I love you mom, you are my rock and my strength; without your constant support and dedication I would have not reached this point. I know you are smiling at me from above. Last but not least I want to dedicate this to the man who is my better-half. Thank you for all your love and support, Guillermo. I love you.

Over 270 million people play soccer worldwide, and its popularity grows every day. In team sport exercise, fatigue may result from numerous factors including limited fuel, depleted energy stores and production of compounds that promote an inflammatory response. While inflammation is an essential mechanism for repairing damaged muscle tissue with exercise, prolonged inflammation leads to increased production of reactive oxygen species that can damage cell membranes, muscle, and signaling proteins. To prevent this response and improve performance, athletes are increasingly looking to nutritional interventions. Carbohydrate and antioxidant supplementation have both shown evidence of producing an ergogenic effect and attenuating inflammation and oxidative stress with prolonged endurance exercise. Less is known about how these interventions may influence intermittent, high-intensity exercise characteristic of soccer. In particular, this exercise presents a unique challenge in that opportunities for nutrient intake are limited to pre-game and half-time. In our first study, we had 10 male collegiate soccer players perform a 90-min. soccer-simulation test, that we developed, which was followed by a progressive shuttle run (PSR) test to exhaustion. They consumed a honey-sweetened beverage (H), a sports drink (S), or a placebo (P) before and half-way through the protocol. Both H and S provided 1.0 g·kg⁻¹ carbohydrate and ~17.6 mL·kg⁻¹ total volume for each trial. Overall, the test resulted in increased fatigue and production of inflammatory markers and antioxidant capacity. There was no significant difference between treatments for any performance measure. Mean times for a high intensity run and rating of perceived exertion increased with time, and there was an overall decrease in PSR time compared to baseline (-22.9%). There was a rise in glucose (15.6%), IL-6 (548%), IL-1ra, IL-10 (514%) and ORAC (15%) post-test but no change in cortisol. Insulin was significantly lower by 1 h-post. IL-1ra levels increased post-test for H (25.8%), S (65.5%), and P (63.9%), but the change for H was less than the other treatments. No treatment effects for the other blood measures were observed. The lack of an ergogenic effect of carbohydrate on soccer performance calls into question the benefit of supplementation at a frequency typical of a regulation soccer match in highly trained athletes with adequate energy stores. Since acute carbohydrate ingestion in the first study did not attenuate some markers of inflammation (e.g. IL-6), we chose to focus on an alternative theory for the rise in inflammatory markers with strenuous exercise in our second study. One aspect of soccer, repeated sprinting, results in increased ROS production partially through the activation of the enzyme xanthine oxidase (XO). Quercetin, a flavonol in plants that has shown some ergogenic effects with endurance exercise, inhibits XO in vitro. The effect of quercetin on team sport exercise had not been studied. We gave recreationally active males a commercial sports drink (S) or S + 500 mg of quercetin (Q) 2x/d for 1 wk prior to a repeated sprint test (RST). Sprint times increased (5.9%) for both treatments as did plasma XO activity (47%), IL-6 (77%), and uric acid (25%) from pre-test to post-test. Q supplementation did not attenuate plasma XO activity or IL-6 and actually increased one calculated index of fatigue, percent fatigue decrement (5.1%- Q and 3.8%- P). These findings add to the growing body of literature that quercetin supplementation does not attenuate exercise-induced inflammation and oxidative stress in vivo. Collectively, this research has practical implications for sports drink companies who are exploring the use of flavonoid compounds in product formulation. Specifically, they should reconsider adding quercetin to their beverages if they are marketing to team sport athletes. Also, soccer players should be made aware that, at ingestion frequencies typical of a soccer match, they may not expect a significant performance benefit from acute carbohydrate supplementation. / Ph. D.