Biological Sciences - Theses, Dissertations, and other Required Graduate Degree Essays

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P-Glycoprotein Phosphorylation and Transport Activity in Rainbow Trout (Oncorhynchus mykiss) Hepatocytes.

Date created: 
2014-12-19
Abstract: 

Evidence suggests that P-glycoprotein (P-gp) transport activity can be post-transcriptionally regulated through protein kinase C-mediated phosphorylation. In the present study, P-gp efflux capacity and phosphorylation state were examined in rainbow trout hepatocytes, following protein kinase C (PKC) inhibition with 1-(5-isoquinolinyl-sulfonyl)-3-methyl-piperazine (H-7), or its activation following treatment with Phorbol 12-Myristate 13-Acetate (PMA). Decreased chemical efflux and increased accumulation following H-7 treatment were observed when rhodamine 123 (R123) was used as P-gp substrate, while PMA increased P-gp efflux and decreased the accumulation of daunorubicine (DNR). This substrate-dependent modulation suggests that phosphorylation affects P-gp’s ability to bind to different substrates or, more likely, to its co-substrate ATP. As well, these results raise the possibility that other transporters may be involved, particularly in R123 transport. Immunoblots of solubilized membrane fractions and whole cell extracts following phosphate-affinity SDS-PAGE showed that H-7 and PMA-treated cells produce different P-gp phosphorylation patterns compared to control cells; up to 5 different phosphoisoforms were visualized using the monoclonal antibody C-219. Differences in the distribution, number, and optical densities of bands were visualized, and can be interpreted as PKC activation or inhibition altering the relative abundance of P-gp phosphoisoforms in the cells. Hepatocytes were also treated with H-7 or PMA and radiolabelled in a media containing 32P, were immunoprecipitated, and subjected to conventional SDS-PAGE. The resulting gels confirmed that PMA increased the amount of 32P incorporated into P-gp as compared to controls, while H-7 decreased it, indicating that PKC inhibition prevents or reduces P-gp phosphorylation and the transport of certain substrates, while activation promotes phosphorylation and transport, also in a substrate-dependent manner.

Document type: 
Graduating extended essay / Research project
File(s): 
Senior supervisor: 
Dr. Chris Kennedy
Dr. Margo Moore
Department: 
Science:
Thesis type: 
(Project) M.E.T.

Species richness, abundance and reproductive responses of riparian birds to habitat restoration in the Okanagan Valley

Date created: 
2015-01-26
Abstract: 

In western North America, most riparian habitats have been destroyed or degraded as a result of human settlement and urban development. I examined temporal trends in the abundance, richness and breeding performance of riparian birds in response to restoration of remnant riparian habitat within the south Okanagan Valley, an arid region of Canada. Total abundance and richness increased over the last decade. Restoration increased the abundance of Yellow-breasted Chats (Icteria virens auricollis), the target of management activities, but did not have a detectable effect on the abundance of other songbirds. The habitat characteristics and breeding performance of Yellow-breasted Chats in restored habitat are currently similar to those of Yellow-breasted Chats in reference sites. Habitat characteristics on multiple spatial-scales (shrub cover of the territory and foliage height of the nest patch) influenced the breeding performance of Yellow-breasted Chats. These results provide evidence that limiting grazing is beneficial to some shrub-nesting songbirds.

Document type: 
Thesis
File(s): 
Senior supervisor: 
David Green
Christine Bishop
Department: 
Science:
Thesis type: 
(Thesis) M.Sc.

Sources and effects of persistent organic pollutants and brominated flame retardants in Cooper’s hawks (Accipiter cooperii) of Vancouver, British Columbia

Date created: 
2014-10-14
Abstract: 

Birds of prey are excellent indicators of environmental health. Since they are top predators, they accumulate a high concentration of persistent organic pollutants (POPs). Cooper’s hawks (Accipiter cooperii) are abundant in urban areas but these environments have been associated with POPs known to cause adverse physiological effects. To investigate the exposure and effects of POPs and flame retardants in Vancouver, British Columbia, we assess the influence of diet, and landscape variables, such as land use and population density on plasma concentrations of pollutants in adult and nestling Cooper’s hawks. We then examined how these pollutant influence thyroid hormones and the fledge success. Our results suggest that: 1) concentrations of DDE are relatively high, 2) urbanized areas were most associated with industrial chemicals; ΣPCB and ΣPBDE, which were also negatively influencing thyroid hormone concentrations, and 3) diet most influenced dieldrin concentrations, which were associated with fledge success.

Document type: 
Thesis
File(s): 
Senior supervisor: 
Ronald Ydenberg
Department: 
Science:
Thesis type: 
(Thesis) M.Sc.

Predicting In Vivo Caffeine/ Angelica dahurica and Caffeine/ Salvia miltiorrhiza Pharmacokinetic Interactions in Humans with In Vitro CYP1A2 Inhibition Data

Date created: 
2014-12-15
Abstract: 

Currently, no existing models allow for holistic predictions of drug-herb interactions. To investigate possible interactions between drugs and current use ‘traditional’ medicines, extracts of Angelica dahurica and Salvia miltiorrhiza were examined in vitro and in vivo for their ability to modify human caffeine metabolism. Consumption of either herbal decoction by volunteers significantly elevated caffeine retention (>200%). Human liver microsomes were treated in vitro with candidate precipitants identified in each herb in addition to ethanolic extracts of the whole herb(s). All treatments reversibly inhibited CYP1A2-mediated caffeine metabolism; Ki values of pure precipitants ranged from 0.28 – 2.55 µM. IC50 values for A. dahurica and S. miltiorrhiza extracts were 1.15e-3 and 1.6e-3 mg/L respectively. Human pharmacokinetic values were estimated for each precipitant and used with in vitro PK data to conservatively predict caffeine retention. This study allows for prediction of hepatic threshold precipitant concentrations associated with clinically relevant inhibition of CYP450 function.

Document type: 
Graduating extended essay / Research project
File(s): 
Senior supervisor: 
Francis Law
Department: 
Science:
Thesis type: 
(Project) M.E.T.

The effect of sulphate on selenium bioaccumulation in two freshwater primary producers and a primary consumer

Author: 
Date created: 
2014-12-16
Abstract: 

Site-specific conditions (e.g. presence of sulphate), may be linked to the variability in the uptake of selenium in organisms at the base of a food chain, potentially affecting the risk of adverse effects in higher trophic-level organisms. In this project, the effect of sulphate on selenate bioaccumulation in two primary producers (Lemna minor and Pseudokirchneriella subcapitata) and a primary consumer (Daphnia magna) was explored. When exposed to selenate, all three species exhibited a decrease in selenium tissue concentration with increasing sulphate. When D. magna were exposed to sulphate and dietary selenium, sulphate did not affect selenium tissue concentrations. The results were used to develop equations estimating selenium tissue concentrations when exposed to selenate and sulphate. The strong predictive ability of the equations suggests that selenate, sulphate, and dietary selenium (applicable to D. magna) are important for describing the relationship between selenate and selenium tissue concentrations.

Document type: 
Graduating extended essay / Research project
File(s): 
Senior supervisor: 
Christopher Kennedy
Department: 
Science:
Thesis type: 
(Project) M.E.T.

Palynology and Paleoecology at Kilgii Gwaay Archaeological Site, Southern Haida Gwaii

Date created: 
2014-11-26
Abstract: 

Buried pond sediments at the Kilgii Gwaay archaeological site on Ellen Island in Gwaii Haanas National Park, were analyzed palynologically to produce a high-resolution record of postglacial vegetation during late glacial and early Holocene time. Seven AMS radiocarbon dates provide a sediment core chronology spanning the period 14,470-10,870 cal yr BP. Four biostratigraphic zones are defined, with a transition from early herb-shrub tundra (14,510 cal yr BP) to a Pinus-dominated forest 14,000 cal yr BP. Pinus is largely replaced by Picea 13,290 cal yr BP, followed by establishment of Tsuga heterophylla 12,420 cal yr BP. Changes in vegetation assemblages and stratigraphy may indicate cooling associated with the Younger Dryas event. Charcoal analysis reveal peak abundance during the known occupation of Kilgii Gwaay (10,700 cal yr BP). Charcoal extends below the known occupation horizon, indicating human presence as early as 11,500 cal yr BP, approximately 800 years earlier than previously documented.

Document type: 
Thesis
File(s): 
Senior supervisor: 
Rolf Mathewes
Department: 
Science:
Thesis type: 
(Thesis) M.Sc.

Web reduction behaviour and the chemical communication system of the western black widow, Latrodectus hesperus

Date created: 
2014-12-03
Abstract: 

In spiders, pheromones mediate intersexual attraction, courtship behaviour, and mate recognition. I studied the chemical communication system of the western black widow spider, Latrodectus hesperus. Females attract males to their webs with silk-borne sex pheromones. Contact with the silk elicits courtship behaviour by males. Often, the courting male engages in web reduction behaviour, during which he excises and bundles up sections of the web and wraps them with his own silk. Using analytical chemistry, and behavioural tests in both the laboratory and the field, I investigated (1) the molecular structure and function of the female’s contact sex pheromone, and (2) the function of web reduction behaviour. A silk-borne contact pheromone elicits web reduction behaviour, and a serine derivative similar to the pheromone of congeneric L. hasselti is one constituent of the multi-component pheromone. Web reduction decreases web attractiveness to rival males in the field, thereby limiting male-male competition.

Document type: 
Thesis
File(s): 
Video A1
Video B1
Senior supervisor: 
Gerhard Gries
Department: 
Science:
Thesis type: 
(Thesis) M.Sc.

An evaluative mode for assessing the fate and effects of contaminants in engineered wetlands

Peer reviewed: 
No, item is not peer reviewed.
Date created: 
2014-10-17
Abstract: 

To investigate the potential of engineered wetlands to remove contaminants from wastewaters, a steady-state and temporal multimedia model was developed for a free water surface design. The model estimates contaminant fate throughout wetland media, assesses the potential for bioaccumulation, and evaluates ecological risk using relevant toxicological endpoints (i.e. EC5). Illustrative examples for pyrene, arsenic, a naphthenic acid, and a chemical universe defined by a range of log KOW (-0.5 – 7.5) and log KOA (-4.5 – 11.5) were used to demonstrate the models applications. In these illustrative applications, the steady-state removal efficiencies of pyrene, arsenic, and the naphthenic acid from the wetland were 41%, 83%, and 31%, respectively. The most efficiently removed chemicals had a log KOW ≈ 6 and log KOA ≤ 2.5. Removal efficiency was lowest for substances showing reduced evapotranspiration flux. This study concluded that wetland treatment could be a feasible method of remediation for certain contaminants in wastewater.

Document type: 
Graduating extended essay / Research project
Senior supervisor: 
Frank Gobas
Department: 
Science:
Thesis type: 
(Project) M.E.T.

Downstream processing of a plant-made recombinant human therapeutic enzyme

Author: 
Date created: 
2014-07-25
Abstract: 

Mucopolysaccharidosis I is a lysosomal storage disease that is characterized by a deficiency of the lysosomal hydrolase alpha-L-iduronidase. Enzyme replacement therapy with purified, recombinant IDUA is one treatment option available for individuals that inherit this disease. The goal of my thesis was to engineer a transgenic line of Arabidopsis thaliana that could express, in the seed tissue, human iduronidase with a therapeutically acceptable N-glycan profile. Human iduronidase was expressed at 5.7% of total soluble protein (TSP) in the Arabidopsis cgl mutant line, which is deficient in N-acetylglucosamine transferase I, and at 1.5% of TSP in the Arabidopsis Golgi-Mannosidase I (GM-I) knockout line. The iduronidase purified from both lines was able to hydrolyze the fluorescent iduronide compound, 4-methylumbelliferyliduronide (4-MUI), at a rate comparable to that of iduronidase produced in Chinese Hamster Ovary cells, commercially available as Aldurazyme. Both plant-derived forms of iduronidase possessed primarily high-mannose N-glycans (at 95% for cgl-iduronidase and 100% for GM1-iduronidase); however, the dominant glycoform in the N-glycan profile of cgl-iduronidase was Man5GlcNAc2, whereas most of the N-glycans on GM1-iduronidase contained six to eight mannose sugars. Additionally, cgl-iduronidase contained a small percentage of complex glycans (and thus potentially immunogenic xylose and/or fucose sugars), which were not found on GM1-iduronidase. Surprisingly GM1-iduronidase did not contain an N-glycan at Asn336, which is normally present on the native human iduronidase and on the recombinant iduronidase of cgl Arabidopsis seeds and Chinese hamster ovary cells. Of importance to creating a product that is of therapeutic value, another goal was to determine whether the plant-produced iduronidase was amenable to in vitro phosphorylation to create the mannose-6-phosphate tag necessary for sequestration and lysosomal delivery in human cells. Both plant-derived forms of the enzymes were able to be phosphorylated in vitro by the UDP-GlcNAc:lysosomal enzyme (GlcNAc)-1-phosphotransferase at a rate similar to that of Aldurazyme.

Document type: 
Thesis
File(s): 
Senior supervisor: 
Allison Kermode
Department: 
Science: Biological Sciences Department
Thesis type: 
(Thesis) M.Sc.

The Effects of Legacy and Emerging Persistent Organic Pollutants in Endocrine Signalling Pathways in Mammalian Cancer Cell Lines

Author: 
Date created: 
2014-08-14
Abstract: 

Despite its ban, DDT and DDE are still persistent in the environment. TBECH is a brominated flame retardant that has been detected in the biota. Here, we determined whether p,p’-DDT, p,p’-DDE and TBECH can negatively impact androgen receptor (AR)-regulated expression of prostate-specific antigen (PSA), prostate-specific membrane antigen (PSMA) and AR in human LNCaP cells. Real-time PCR, Western-blotting and COBAS PSA detection system were used to measure mRNA and protein levels. Chromatin-immunoprecipitation assay was used to determine AR-PSA promoter interaction. My results showed that p,p’-DDT and p,p’-DDE repressed R1881-inducible PSA mRNA and protein levels by blocking recruitment of AR to the PSA promoter. p,p’-DDT and p,p’-DDE can also significantly repress AR protein levels and relieve R1881-inducible PSMA repression. TBECH was found to repress R1881-inducible PSA protein levels. I conclude that men who have been exposed to either DDT or DDE may produce a false-negative PSA test when screening for prostate cancer.

Document type: 
Graduating extended essay / Research project
File(s): 
Senior supervisor: 
Timothy Beischlag
Department: 
Science: Biological Sciences Department
Thesis type: 
(Project) M.E.T.