It is clear that non-melanoma skin cancers have a significant impact on the health of the general population. With over one million new cases expected this year the extent of the problem is increasing. Extensive research on the molecular mechanisms of tumor promoting agents has yielded a large body of evidence suggesting that activity of the transcription factor activator protein-1 is central to development of benign lesions. Recently, a dominant negative c jun, TAM-67, has been shown to block tumor promotion by 12-O-tetradecanoylphorbol-13-acetate. In order to determine if this is a more general phenomenon, we examined the effects of TAM-67 on okadaic acid induced tumor promotion. We found that TAM-67 could effectively inhibit tumorigenesis induced by okadaic acid. In order to further characterize the activity of TAM-67, we identified its molecular mechanism of action in suppressing okadaic acid induced activator protein-1 activity. These studies led to identification of squelching as the mechanism of action. This work also revealed that TAM-67 could interact with all the proteins of the Jun and Fos families as well as non-activator protein-1 proteins. Thus, TAM-67 may be able to affect multiple pathways including activator protein-1. Other work has identified prostaglandin signaling as a major factor in the development of non-melanoma skin cancer. We identified production of prostaglandin E2 as one of the abnormalities in a model of skin tumor progression. The cell lines examined were able to make prostaglandin E2, and also expressed receptors for it. We therefore examined the importance of this apparent autocrine loop and found that the cells that produce prostaglandin E2 depend on signaling from the prostaglandin E2 receptor EP1 for a normal in vitro growth rate. Both transactivation by activator protein-1 and signaling by prostaglandins are involved in chemically induced skin carcinogenesis. The magnitude of the problem ensures that demand for new treatments will only increase. Obtaining a clear understanding of the molecular events associated with the growth and transformation of cells from normal to benign to malignant is crucial to identifying novel treatment and prevention strategies.

The hypothesis upon which this research is based is that the mitochondrial thioredoxin-2 system, which consists of mitochondrial thioredoxin-2 (Trx-2), mitochondrial thioredoxin reductase (TrxR-2) and mitochondrial thioredoxin peroxidase (Prdx-3), protects cells against apoptosis and regulates cell growth via mitochondrial redox homeostasis. Trx-2 mRNA was found expressed in a panel of cancer cell lines and Trx-2 protein is localized exclusively to the mitochondria. An alternate splice form of Trx-2 lacking exon 2 was identified that does not yield protein. Forced overexpression of Trx-2 in cell lines by using constitutive and inducible promoter systems increased mRNA levels, but did not yield an increase in Trx-2 protein. The role of Trx-2 in mammalian development was examined by generating a mouse deficient in Trx-2. Massive apoptosis, exencephaly and early embryonic lethality was seen in the Trx-2(-/-) homozygous embryos. Trx-2(-/-) embryonic fibroblasts were not viable under normal growth conditions, but could be rescued with maintenance in hypoxia. Trx-2(-/-) cells were also lacking mature cytochrome c, implicating Trx-2 in cytochrome c biogenesis. The Trx-2(+/-) heterozygous mice, which appear normal, had an increased sensitivity to some forms of oxidative toxicity (eg. acute doxorubicin) compared to the wild-type mice. To investigate the role of Prdx-3, WEHI7.2 cells with overexpression of Prdx-3 were generated by stable transfection. Overexpression of Prdx-3 inhibited cell proliferation, reduced cellular hydrogen peroxide levels and altered the mitochondrial membrane potential. Prdx-3 overexpression protected the cell from various drug-induced and hypoxia-induced apoptosis. Prdx-3 protein degradation was decreased during hypoxia, leading to a longer half-life under hypoxic conditions. Additionally, Prdx-3 protein levels were increased in a RCC4 cells expressing wild-type VHL compared to RCC4 cells with mutant VHL.

In humans, livestock and experimental animals, pyrrolizidine alkaloids (PAs) are toxic as a consequence of their hepatic metabolism to reactive pyrrolic esters, or dehydroalkaloids (DHAs). Despite their similarity in structures, PAs often vary markedly in their lethality (LD₅₀s) and in the organs in which toxicity is expressed. We have examined whether there are differences in the physicochemical properties of certain DHAs which are associated with differences in patterns of metabolism and toxicity produced by the parent PA. Using a potentiometric method to measure hydrolysis, it was determined that the half-lives of the corresponding DHAs of retrorsine, seneciphylline, monocrotaline and trichodesmine were 1.06, 1.60, 3.39 and 5.36 sec, respectively. These values were supported by similar results from experiments measuring reactivity of DHAs toward 4-(p-nitrobenzyl)pyridine. Studies from the isolated rat liver perfused with PAs show that DHA stability is related to patterns of metabolism and toxicity. Perfusion of the primarily hepatotoxic retrorsine and seneciphylline is associated with a greater proportion of metabolite released as non-toxic 7-glutathionyl-6,7-dihydro-1-hydroxymethyl-5H-pyrrolizine (7-GSDHP), a greater proportion alkylating liver macromolecules, and a lower proportion released as DHA into the circulation. Perfusion with monocrotaline and trichodesmine, PAs producing extrahepatic toxicity, produced lower proportions of 7-GSDHP release and liver alkylation, and higher proportions of DHA released into the circulation. Other studies characterizing DHAs included the use of an in vitro enzyme assay in which DHAs were shown to inhibit the phosphotransferase activity of yeast and rat brain hexokinase. Parent PAs, and the hydrolysis product of DHAs, (±)-6,7dihydro-1-hydroxymethyl-5H-pyrrolizine (DHP) did not affect enzyme activity. In vivo studies in rats have established that glutathione and cysteine-conjugated pyrrolic metabolites of PAs likely represent detoxication pathways, providing further support for DHAs as the primary toxic metabolite. We have examined the chemical form of sulfur-bound pyrroles to establish the importance of the 7-ester position in PA toxicity. Additionally, we have developed an efficient technique for the rapid separation and purification of large quantities of PAs using high-speed counter-current chromatography.

Peach harvester exposure to azinphos-methyl (AZM) residues estimated by the Transfer Factor (TF) ratio of Dislodgeable Foliar Residue (DFR) to Daily Dermal Exposure (DDE) was compared to estimates based on the relationship between dermal exposure and dialkylphosphate metabolite excretion. DFR was monitored in four orchards from the time of pesticide application through harvest. Eleven male harvesters wore cotton tee-shirt dosimeters and provided area-specific skin washes and wipes for DDE monitoring during work operations. Urinary dialkylphosphate metabolites of AZM were also monitored and compared to DDE results. During a later harvest for which DDE was not determined, the dermal exposure estimated by the TF (1,310 μg) was comparable to the estimates based on dialkylphosphate excretion (1,456-1,534 μg). A repeated measures ANOVA showed that harvesters' cholinesterase levels were significantly lower than 'non-harvesters'. No significant reductions in cholinesterase levels were detected over time.

Three commercially available herbal tea preparations (Weightless, Female Toner, and PMS) and one single ingredient herbal preparation, Chaparral (Larrea tridentata), were sequentially extracted with solvents of decreasing polarity (water, methanol and chloroform) and the crude extracts obtained screened for toxic/mutagenic potentials using the brine shrimp (Artemia sp.), mouse acute toxicity, Salmonella/microsomal mutagenicity, and chicken embryo bioassays. The crude aqueous extract from Weightless Tea was very toxic to brine shrimp larvae and had a cathartic action in mice at the highest concentration tested. While Weightless tea crude water extract was not mutagenic to Salmonella typhimurium TA100 at the concentrations tested, three chromatographic isolates obtained from a silica Gel 60 column were mutagenic to the test organism. Two of these isolates were detoxified after inclusion of a microsomal activation system. The teratogenic potentials of these isolates are unknown as the results obtained from the chicken embryo bioassay were not conclusive.

<p> Toxoplasma gondii is an obligate intracellular parasite of the phylum Apicomplexa. Toxoplasma infection is a serious threat to immunocompromised individuals such as AIDS patients and organ transplant recipients. Side effects associated with current drug treatment calls for identification of new drug targets. DNA repair is essential for cell viability and proliferation. In addition to reactive oxygen species produced as a byproduct of their own metabolism, intracellular parasites also have to manage oxidative stress generated as a defense mechanism by the host immune response. Most of the oxidative DNA damage is repaired through the base excision repair (BER) pathway, of which, the apurinic /apyrimidinic (AP) endonucleases are the rate limiting enzymes. Toxoplasma possesses two different AP endonucleases. The first, TgAPE, is a magnesium-dependent homologue of the human APE1 (hAPE1), but considerably divergent from hAPE1. The second, TgAPN, is a magnesium-independent homologue of yeast (Saccharomyces cerevisiae) APN1 and is not present in mammals. We have expressed and purified recombinant versions of TgAPE and TgAPN in E. coli and shown AP endonuclease activity. Our data shows that TgAPN is the more abundant AP endonuclease and confers protection against a DNA damaging agent when over-expressed in Toxoplasma tachyzoites. We also generated TgAPN knockdown Toxoplasma tachyzoites to establish that TgAPN is important for parasite protection against DNA damage. We have also identified pharmacological inhibitors of TgAPN in a high-throughput screen. The lead compound inhibits Toxoplasma replication at concentrations that do not have overt toxicity to the host cells. The importance of TgAPN in parasite physiology and the fact that humans lack APN1 makes TgAPN a promising candidate for drug development to treat toxoplasmosis.</p>

<p> Growing concerns about the environment and terrorist attacks prompt a search for effective adsorbents for removal of small molecule toxic gases, such as ammonia and hydrogen sulfide, under ambient conditions in the presence of moisture, where physical adsorption is not adequate. We use graphene oxide and CuBTC metal-organic framework as the adsorbents to explore toxic gas removal by reactive adsorption. Using <i>ab initio</i> density functional theory, atomistic reactive molecular dynamics and Monte Carlo simulation strategies, theoretical understanding of the underlying reaction and adsorption mechanisms of ammonia and hydrogen sulfide on graphene oxide and CuBTC metal-organic framework have been gained. </p><p> The <i>ab initio</i> calculation results show that ammonia and hydrogen sulfide decompose on carboxyl and epoxy functional groups and vacancy defects of graphene oxide. The existence of water molecules substantially reduces the adsorption/dissociation of ammonia or hydrogen sulfide on graphene oxide because the water molecules either form hydrogen bonds with the functional groups or adsorb more easily on the vacancy defects. Reactive molecular dynamics calculations by the ReaxFF method have been performed to propose realistic graphene oxide models for theoretical calculations. We also use reactive molecular dynamics simulation to study the thermal and hydrostatic stabilities of the CuBTC metal-organic framework and its application for ammonia removal. We predict the collapse temperature for CuBTC crystal structure and observe the partial collapse of CuBTC at lower temperatures upon ammonia adsorption. The results agree well with experiment data and provide insights on the reaction mechanism involved in such an ammonia removal process. </p><p> The research in this thesis can provide fundamental understanding, at the electronic and atomistic levels, of the roles of surface defects and functionalities for reactive adsorption of toxic gas molecules. In addition to developing experimental and theoretical algorithms to design effective adsorbents, the results are expected to find applications in air cleaning, energy storage, fuel cell technology and other scientific challenges where the separation of reactive molecules is involved.</p>

309 cases of organic dementia, cerebral atrophy or psycho-organic syndrome, admitted for 5 days or more to one of 18 Quebec hospitals, were individually matched to a psychiatric referent, admitted with some other diagnosis, and a general hospital referent. Lifetime occupational history was obtained by telephone. Occupational solvent exposure was assessed by (i) individual ratings blind to case status and (ii) a job-exposure matrix. Subjects working with moderate or high solvent concentrations for at least 10 years were considered exposed. With the psychiatric referent series an odds ratio of 1.44 (90% CI 1.03-2.01) was calculated for individual exposure ratings and 1.41 (90% CI 0.89-2.23) for the job matrix. The increased risk was found largely in those with diagnoses of both organic dementia or cerebral atrophy and an alcohol related condition. A similar pattern of risk was found with the general hospital referents. Adjustment for possible confounders did not appreciably alter the risk estimates.

The hypothesis was that there is an association between cardiovascular, reproductive and oncogenic effects and chemicals (gases, volatile organic compounds (VOC's)) to which firefighters are exposed. The objectives of the study were to assess firefighter exposures to: (1) diesel emissions in firehalls; (2) contaminated self-contained-breathing-apparatus (SCBA's); and, (3) VOC's at fires. Carbon monoxide (CO) was used as a marker for diesel emissions. Air in SCBA cylinders was analyzed for chemical contaminants. The U.S. EPA TO-14 method for ambient air analysis was verified for the analysis of 144 VOC's in air samples collected in SummaTM canisters at experimental and municipal fires. Exposure groups were determined from firehall assignment and vehicle responses to alarms. Estimation of the percentage of time that firefighters used respiratory protection at fires was obtained from records kept for SCBA refills. / The mean maximum concentration of CO measured during the operation of two fire trucks in unventilated garages was 4.1 +/- 2.6 ppm (mean +/- SD, n = 16). A linear relationship existed between levels of CO, CO2 and NO2 in diesel emissions. By implication, using CO as a marker, firefighter exposure to diesel emissions was low. Some SCBA's were found to contain levels of CO (up to 250 ppm) consistent with possible COHb poisoning. Distinctive firefighter exposure groups were identified. A Montreal firefighter attended some 19 to 62 fires per year (depending on firehall assignment), 46% being structural fires. Respiratory protection was used for 6% of the time spent at the firescene. No new or novel toxic VOC's were found in fire samples and a characteristic "fingerprint" of VOC's was identified. Three potential carcinogens (benzene, 1,3-butadiene, styrene) accounted for 25% of the 123 VOC's found in fires, maximum levels of these being 11, 5 and 2 ppm, respectively. Since the results suggest that firefighters are not exposed to high levels of carcinogenic VOC's, why then do epidemiological studies reflect adverse health outcomes?

More than one billion pounds of plasticizers are produced each year to supply the plastics industry. Some of these plasticizers, particularly phthalates, are suspected endocrine disruptors. However, few studies have been conducted to determine if they are susceptible to biodegradation by naturally occurring bacteria once they are released into the environment. / Six organisms were tested for their ability to grow in the presence of six different industrial plasticizers. Two bacteria, Rhodococcus rhodochrous and Arthrobacter paraffineus, grew well in media containing n-hexadecane and one of the plasticizers. / Fermentations in a 2-liter reactor were performed with Rhodococcus rhodochrous and three plasticizers: bis 2-ethylhexyl adipate, dioctyl phthalate and dioctyl terephthalate. The organism degraded all of the adipate, half of the terephthalate was degraded and the phthalate was degraded slightly. / In these growth studies, the toxicity of the media increased as the organism grew. This trend was linked to the accumulation of metabolites from the partial degradation of the plasticizer. The two major metabolites were identified as 2-ethyl hexanol and 2-ethyl hexanoic acid. The alcohol was only observed part way through the growth in the presence of the adipate. Its concentration decreased as it was oxidized to the acid and it was not present at the end of the fermentation. / The acid was observed for all three types of plasticizers and it was present in high concentrations at the end of every experiment. The nature and pattern of production of the metabolites were consistent with a pathway for the degradation of all three plasticizers by hydrolysis of the ester bonds. / The accumulation of toxic metabolites indicates that biodegradation may not be a solution to reducing environmental impacts associated with plasticizers that have leached into the environment.