RSS 1.0Chemistry and Biochemistry
Recent Submissions
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Ligand substitution versus DNA-binding affinity : ǂb syntheses, structures, and in vitro anticancer activities of Pt(CH₃)₂I₂{2,2'-bipy-n,n'-(CH₃)₂} (n,n' = 4,4'; 5,5'; 6,6')In chemotherapeutic treatment for leukemia, the drug cisplatin is highly effective. However, there is an immense need for cisplatin analogues that are more clinically viable. Cancer cell assay studies have shown that the platinum(IV) complex Pt(CH₃)₂I₂{2,2'-bipyridine} exhibits significant anticancer activity against the human breast cancer cell line ZR-75-1, with an EC50 (effective concentration 50%) concentration of 6.1 μM— substantially lower than that of cisplatin (16.4 μM). To explore the unknown structure-activity relationship between DNA binding and anti-cancer activity, three isomers — Pt(CH₃)₂I₂{2,2'-bipy-n,n'-(CH₃)₂} where n,n’ = 4,4’; 5,5’; 6,6’— were synthesized. The intent was to demonstrate decreased cytotoxicity as complex-DNA steric hindrance increased through the positioning of the methyl groups on the bipyridine ring of the isomers. All isomers were structurally characterized via single-crystal X-ray diffraction, nuclear magnetic resonance (NMR), and infrared spectroscopy (IR). Their cytotoxic profiles were evaluated through the National Cancer Institute’s Sulforhodamine B screening assay. Among them, Pt(CH₃)₂I₂{2,2'-bipy-6,6'-(CH₃)₂} demonstrated the most potent activity with a notable LC50 (lethal concentration 50% ) value of 7.34 μM against ovarian cancer (OVCAR-3). DNA- binding studies using UV-Vis absorption spectroscopy revealed intrinsic binding constants (Kb) of 1.74×10⁴ M⁻¹ and 3.74×10⁴ M⁻¹ for the 6,6'-dimethyl and unsubstituted bipyridine complexes, respectively. A competitive displacement reaction monitored by 'H NMR showed that Pt(CH₃)₂I₂{2,2'-bipy-6,6'-(CH₃)₂} reacts completely with free 2,2’-bipyridine to yield Pt(CH₃)₂I₂{2,2'-bipyridine} and liberated 6,6’-dimethyl bipyridine. These findings suggest that DNA-binding affinity does not directly correlate with cytotoxic potential and that steric factors introduced by ligand substitution play a more critical role in determining anticancer efficacy in these bipyridine-stabilized platinum(IV) compounds.
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Measuring Arctic ground squirrel adult neurogenesis in caudal hippocampus using DAB/DAB-Ni enzyme-based immunohistochemistryUnconscious patients who are resuscitated after cardiac arrest often have a poor prognosis for neurological recovery and are routinely treated with therapeutic hypothermia as a standard of care. This intervention aims to reduce the risk of death and minimize brain damage. However, results from the Targeted Temperature Management 2 (TTM2) trial in 2021 indicate that cooling does not improve neurological outcomes in these patients. This finding raises questions about the neurological benefits, particularly regarding neurogenesis, of the rewarming phase of therapeutic hypothermia, as well as the effects of repeated cooling and rewarming cycles. Neurogenesis has been largely overlooked in therapeutic hypothermia research, leaving significant gaps in our understanding. Notably, hibernation and therapeutic hypothermia share physiological similarities, but hibernation is a more complex process that confers neurological protection against cardiac arrest in the Arctic ground squirrel (AGS), an extreme hibernator. Specifically, during hibernation, AGS repeatedly lower body temperature to as low as 1 to -3°C, along with reduced blood flow and metabolism during torpor, then periodically rewarm to normal levels during brief interbout arousals. These repeated cooling and rewarming cycles happened at least eight times during hibernation. Building on this knowledge, this thesis explores the possibility that cooling and rewarming could activate neurogenesis and thereby improve the benefits of therapeutic hypothermia by replacing dead and damaged neurons. As proof of concept, I developed a method for measuring hippocampal neurogenesis in the AGS across three seasonal states: summer active, hibernation torpor, and interbout arousal.
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Understanding secondary atmospheric chemistry in extremely cold and dark environments through hydroxymethanesulfonate (HMS) measurements in wintertime Fairbanks, AlaskaFairbanks, Alaska is a subarctic city that regularly violates PM2.5 air quality standards in the winter, exacerbated by strong wintertime inversion layers and wood stove heating. While primary sources of PM2.5 in Fairbanks have been studied extensively, secondary chemistry remains not well understood. Here, we focus on the aqueous secondary product hydroxymethanesulfonate (HMS), a major sulfur(IV) particulate species, to help understand what drives secondary pollution in Fairbanks. In 2020 and 2021, we used a particle-into-liquid sampler coupled with ion chromatography (PILS-IC) to measure HMS and sulfate concentrations. We report that HMS concentrations are high compared to other highly polluted regions like Beijing, and HMS makes up a significant portion of PM2.5 mass. In 2022, we repeated these measurements as part of the Alaska Layered Pollution And Chemical Analysis (ALPACA) campaign, and examined the processes that allow accelerated formation of HMS in Fairbanks. We find that HMS formation is likely driven by the extremely cold conditions of Fairbanks. HMS formation occurs in aqueous droplets and is highly pH dependent, forming much faster at higher pH. The extreme cold of Fairbanks lowers the volatility of gas-phase ammonia, which leads to increased solubility of aqueous-phase ammonia and increased partitioning to aqueous-phase ammonium, raising particle pH and causing rapid production of PM2.5 HMS. We show how this mechanism applies to other regions globally, and how it can affect pH-dependent sulfate production. To better understand the HMS precursors formaldehyde and SO2 gas, as well as gas-phase pollutants in general, we made volatile organic compound (VOC) measurements in 2022. Many VOC concentrations are much higher in wintertime Fairbanks compared to wintertime measurements in other US cites. We find that diesel vehicles and air mass transport (possibly from nearby areas with more wood heating) are large sources of formaldehyde. Aromatic pollutants like benzene, toluene, and C8 aromatics largely come from gasoline vehicles and heating oil. Methanol, the single largest VOC, was related to windshield wiper fluid. These measurements and calculations lead to a better understanding of secondary pollutant products and VOC concentrations in cold and dark environments like wintertime Fairbanks, AK, and can hopefully contribute to pollution mitigation strategies in the future.
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Effects of iron nutrients in sub-arctic horticultureWhen managing iron (Fe) nutrients in controlled environment agriculture (CEA) hydroponic systems at sub-arctic regions between latitudes 50 to 70 degrees north, it is important to understand how plants regulate Fe to maintain Fe homeostasis, which is important for the production of Fe proteins and photosynthesis processes. The research objectives of this study are as follows: 1) determine Fe nutrients that promote yearly cycles of high-quality crops, 2) examine the regulation of Fe between plant roots and vegetation, and 3) explore whether seasonal adjustments in Fe nutrients can promote Fe homeostasis and the production of high-quality crops. This study considers the use of ferrous sulfate-heptahydrate (FeSO4·7H2O), ferric citrate (Fe-Cit) and sodium hydrogen ferric diethylenetriamine pentaacetate (Fe-DTPA) in Rosie romaine lettuce. Results demonstrated how Fe-DTPA produced high-quality crops every season compared to FeSO4·7H2O and Fe-Cit. The Fe nutrients had different influences on Fe concentrations in lettuce roots and vegetation. From April to May 2019, high pH and light levels impacted lettuce production. Under Fe-deficient conditions, plants exhibited more Fe in vegetative portions than roots. From December 2020 to January 2021, low pH and the need for supplementary lighting slowed germination and growth. Plant weight decreased and plants exhibited more Fe in roots than vegetative portions. From July to September 2020, low pH and optimal sunlight produced high-quality lettuce and adequate Fe concentrations. Research demonstrated that seasonal adjustments in Fe nutrient management can promote high-quality crops and Fe homeostasis.
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An exploration of panarctic lake formation and methane emission since the Last Glacial MaximumPolar ice cores show that atmospheric methane concentrations nearly doubled in response to rapid climate warming over the last deglacial transition. Since concentrations of this potent greenhouse gas are tightly coupled to the Earth’s climate system, understanding the climate ecosystem interactions that precipitated this event may help predict feedbacks to current and future warming. This work reconstructs panarctic lake areas and methane emissions to assess their contribution to global atmospheric methane budgets since the Last Glacial Maximum. In the first two chapters of my dissertation, I show that climate warming and deglaciation caused widespread lake formation across land surfaces poised toward this trajectory by glaciation. Thermokarst (thaw) lake formation that accelerated in response to climate warming released methane from a mixture of radiocarbon-depleted permafrost soils and contemporary carbon sources, creating a positive climate feedback that helped sustain early Holocene temperature increases. Younger, albeit ultimately larger sources of methane from more extensive glacial lakes, lagged those from thermokarst lakes but were more than twice their magnitude throughout most of the Holocene. These findings are consistent with top-down polar ice core ¹⁴CH₄ constraints. Not included in my initial analysis were exceptionally large proglacial lakes dammed by continental ice sheets. These proglacial lakes, which had never been explored as a methane source in the literature, were the focus of my third dissertation chapter. I found that within a single large proglacial lake, Lake Agassiz, lake lowering and subsequent re-expansion into shallow aquatic and subaerial environments provided the most significant opportunity for methane production, which was otherwise limited by substantial water depth. Since it is unlikely that projected warming will cause extensive lake formation on the order of that observed during last deglacial, much of the 21st century permafrost carbon feedback will ultimately depend on how many new lakes the landscape can support. Over geologic timescales, the function of northern lakes as a carbon source could be unique to early interglacial stages due to the inevitability that lake drainage and terrestrialization will transform these features into climate stabilizing carbon sinks.
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Metabolism and thermogenesis in the hibernating arctic ground squirrelArctic Ground Squirrels (AGS) tolerate up to eight months of fasting, inactivity, and large fluctuations in body mass. AGS utilize their fat stores Fat stores, which account for 25-50% of the total body mass, as a fuel source through the hibernation season to maintain a heterothermic body temperature pattern. Body temperature covers a 42°C range, requiring tight regulation to ensure survival. There are two main groupings of heat production that contribute or may contribute to temperature regulation: shivering and non-shivering thermogenesis (NST). The latter includes uncoupling protein 1 (UCP1) and sarcolipin (SLN) uncoupling of the sarcoendoplasmic reticulum calcium ATPase (SERCA). UCP1, located in the mitochondria of adipose tissue, is a futile proton cycler, while SLN and SERCA, found in muscle tissue, result in futile calcium cycling. However, the regulation and contribution of these individual metabolic pathways to full-body metabolism in AGS is unknown. We utilized three approaches to understand energy balance, metabolism, and thermoregulation in hibernating AGS: protein analysis of squirrels at discrete intervals over the hibernation season, modulation of the temperature conditions of the hibernaculum, and interruption of thermoregulatory pathways via dosing with pharmacological agents. We assessed whole animal metabolism in two ways: by tracking internal body temperature throughout the experiment and by measuring the full body respiratory gas exchange during arousal episodes. During the hibernation season, we observed suppression of muscle NST via SLN uncoupling, while UCP1 uncoupling remained upregulated until after spring terminal arousal. During arousal, lower hibernaculum temperatures increased both UCP1 and muscle NST. Interruption of thermoregulation during arousal, via low doses of the pharmaceutical agents dantrolene and SR59230A, did not change AGS’s ability to arouse but seemed to indicate compensation via intact metabolic pathways. Furthermore, in response to pharmacological intervention AGS were able to recruit white adipose as a thermogenic source under conditions of thermogenic stress. In conclusion, changes in whole animal respirometry, protein expression, and body temperature indicate a tightly regulated metabolism and biological fail-safes to ensure successful hibernation and survival in this novel animal model.
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Synthesis and stereoselective reduction of α-fluoro-β-ketoesters by ketoreductasesUsing commercially available ketoreductase (KRED) enzymes, α-fluoro-β-hydroxy esters were stereoselectively synthesized from racemic α-fluoro-β-keto esters through dynamic reductive kinetic resolution (DYRKR). The α-fluoro-β-keto esters were synthesized via Reformatsky reactions between an aromatic aldehyde and ethyl bromofluoroacetate and subsequent oxidation with Dess Martin Periodinane (DMP). Two enzymes were selected for their ability to yield either syn or anti diastereomers. Three aromatic substrates were reduced in high diastereomeric and enantiomeric excess and good yields. The KRED products were derivatized with (R)- and (S)-α-methoxy-α-trifluoromethylphenylacetic acid (MTPA) and analyzed via ¹⁹F NMR spectroscopy to determine their absolute stereochemistry via Mosher ester analysis. For the three substrates, KRED 110 yielded the anti 2S,3S isomer. KRED 130 predominantly yielded the syn 2S,3R isomer but with less specificity. The use of these commercially available KRED enzymes provides access to enantio- and diastereomerically pure α-fluoro-β-hydroxy esters from readily accessible racemic substrates. Optically pure α-fluoro-β-hydroxy esters may serve as useful intermediates in the synthesis of medicinally relevant compounds such as fluorinated amino acids or fluorinated sphingolipid derivatives.
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Synthesis, characterization and cytotoxicities of novel organoplatinum(IV) complexesBreast cancer is the most common form of cancer in the world. In 2023, nearly 300,000 new cases of breast cancer were projected in the U.S. alone. When treating breast cancer, several therapies are utilized, often including chemotherapy. Organoplatinum(IV) complexes are among the newer chemotherapy drugs being developed as an alternative to cisplatin, the first platinumbased anticancer drug. Two novel organoplatinum(IV) complexes: [Pt(CH₃)₂X₂{bipy-(CO₂H)₂}] (X = Br, I) have been isolated and structurally characterized by single-crystal X-ray diffraction. Nuclear magnetic resonance and infrared spectroscopic data were also collected to confirm these new compounds and provide useful reference values. The anticancer potential of each compound was assessed via in vitro 3-4,5-dimethylthiazol-2,5-diphenyltetrazolium bromide (MTT) assays using human breast cancer cells (cell line ZR-75-1). EC₅₀ values were determined as 3020 μM, for Pt(CH₃)₂Br₂{bipy-(CO₂H)₂} and 86.0 μM, for Pt(CH₃)₂I₂{bipy-(CO₂H)₂}; for comparison, the EC₅₀ value for cisplatin against the ZR 75-1 cells was 16.4 μM. While these compounds were not as effective as cisplatin, the single-crystal X-ray diffraction data showed evidence of intermolecular π-π interactions within the unit cell which suggest that the bipyridyl ligands can potentially interact with tumor DNA nucleobases in the DNA double helix.
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Marine methane: sources and potential adverse effectsRecently, there has been an increase in the occurrence of incidents involving spills of oil and natural gas, such as methane. The world experiences at least one major spill in each decade. An illustrative case is the Deepwater Horizon oil disaster in 2010 in the Gulf of Mexico. This catastrophe resulted in the discharge of about 205.8 million gallons of oil and 225,000 metric tons of methane gas into the Gulf of Mexico. More recently, in 2022, the Nord Stream pipeline leak occurred, which was the largest single methane release ever recorded. It released up to 500K tons of methane underwater, a greenhouse gas with a significantly higher potency than carbon dioxide. Due to the Deepwater Horizon incident, mammals, sea turtles, birds, fish, and invertebrates were adversely affected and caused damage to the corals. More than 90 bird species died, and 1300 miles of shoreline became polluted. The fishing industry suffered a significant reduction. This study systematically reviewed the source and impact of methane in the marine environment, utilizing 271 peer-reviewed academic publications, eight non-peer-reviewed sources, and 44 online resources. In the marine environment, methane can come from various sources such as methane hydrate, methane seeps, pockmarks, mud volcanoes, microbial activities, and anthropogenic sources or human-induced activities. Sediment typically contains methane from methane seeps, methane hydrates, mud volcanoes, and microbial activity. In the water column, methane is produced from diffusion from hydrates, seeps, hydrothermal vents, and thermogenic and anthropogenic sources. On the other hand, air-water interface methane comes from the atmospheric exchange or diffusion from the water column or sediment. In marine water, methane undergoes various reactions. Methane reacts with oxygen, producing carbon dioxide in aerobic conditions. Conversely, in anaerobic conditions, methane is anaerobically oxidized, coupling with sulfate reduction mediating by sulfate-reducing bacteria and methanotrophic archaea. These microorganisms, bacteria, and archaea derive the majority of their carbon and energy from methane, and they can proliferate their number where they find excess methane. However, excess methane can create anoxic conditions by reducing oxygen concentration. Invertebrates utilize methane through a symbiotic relationship with methane-consuming microorganisms. Moreover, the marine ecosystem exhibits complex interdependencies among the organisms and methane.
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Pollution trapping by strong temperature inversions in Fairbanks, AlaskaIn cold climates during winter, surface-based temperature inversions reduce vertical dispersion within the atmospheric boundary layer. Reduced vertical dispersion coupled with stagnant horizontal winds causes pollution emitted near the ground level to accumulate to unhealthy amounts. To study pollution trapping, we measured vertical differences in fine particulate matter and ozone across a shallow 20-meter vertical scale during surface-based inversions in Fairbanks, Alaska and showed that pollution trapping occurs on this extremely shallow scale. In winter 2022, trace gases were measured by long-path differential absorption spectroscopy to probe vertical dispersion on a taller scale, up to ~200 m above the surface. We added horizontal dispersion to a one-dimensional Eulerian chemical transport model and used the model with both vertical and horizontal dispersion, but without chemistry, to simulate the vertically resolved measurements of SO₂. The model achieved excellent results with correlation to the observations having a coefficient of R = 0.88. Steady-state transport residence times calculated from the model were on the order of hours, indicating limited time for chemical processing during winter in Fairbanks. Within the model, only ground-based emissions sources were included and there was no interaction between air above and below the boundary layer height, suggesting that ground-based sources dominate pollution measured at ground level. With the knowledge that ground level sources have a large impact on pollution, we carried out an analysis of trace gases and fine particulate matter measured near ground level that was used to better understand the response to pollution control strategies in Fairbanks. This analysis shows that the total amount of pollution in Fairbanks has been trending down over the past nine years. Following a September 2022 legal change that mandated lower sulfur content in heating oil, the amount of sulfur dioxide gas dropped significantly during winter 2022-2023 as compared to the prior three-year average.
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Effects of calcium magnesium acetate on small ponds in interior AlaskaDeicing winter roads with chlorides has been common practice in northern areas. Corrosion of vehicles, structures, and pavements has resulted in damage, and roadside vegetation and public water supplies have been severely impacted by heavy salt usage. Calcium magnesium acetate (CMA) was identified as a promising alternative to chloride salts. This research focused on the effects on planktonic algae and bacteria in small ponds, to help assess the environmental acceptability of CMA in Alaska. The most serious consequence resulting from CMA was a dissolved oxygen concentration decrease, partly attributable to increased bacteria populations utilizing oxygen during degradation of acetate. Algal biomass recovered toward the end of the summer indicating that the algae may be utilizing some of the carbon dioxide being released by the bacteria. Rapid turnover times of acetate by bacteria in the fall and continued low dissolved oxygen indicated that some of the acetate was being recycled.
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Clearance of beta-amyloid 42 (Aβ-42) and tau proteins in Alzheimers Disease (AD)Alzheimer's disease (AD) is a progressive neurodegenerative disorder and the main cause of dementia among the elderly worldwide. Despite intense efforts to develop drugs for preventing and treating AD, no effective therapies are available yet, posing a growing burden at the personal, medical, and socioeconomic levels. AD is characterized by the production and aggregation of amyloid 0 (A0) peptides derived from amyloid precursor protein (APP), the presence of hyperphosphorylated microtubule-associated protein Tau (MAPT), and chronic inflammation leading to neuronal loss. A0 accumulation and hyperphosphorylated Tau are responsible for the main histopathological features of AD, A0 plaques, and neurofibrillary tangles (NFTs), respectively. However, the full spectrum of molecular factors that contribute to AD pathogenesis is not known. Non-coding (nc)RNAs, including microRNAs (miRNAs), long noncoding RNAs (lncRNAs) regulate gene expression at the transcriptional and posttranscriptional levels in various diseases, serving as biomarkers and potential therapeutic targets. There is rising recognition that ncRNAs have been implicated in both the onset and pathogenesis of AD. Here, the ncRNAs implicated post-transcriptionally in the main AD pathways and discuss the growing interest in targeting regulatory ncRNAs therapeutically to combat AD by using a series of multifunctional molecules that contained APP, and Tau-recognition moieties and E3 ligase-binding moieties to enhance APP, and Tau degradation. The goals involve the exploration into non-invasive biomarker screening for cognitive dementia related to Alzheimer’s Disease (AD) and in canine blood samples. The specific aims are towards the identification and development of non-invasive AD-related biomarkers for detection screening before the onset of pathophysiological symptoms related to cognitive deterioration seen in AD. The purpose of the project is concentration on the clearance of A0 and Tau through the liver.
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Impacts of sex, biometals homeostasis, and dietary fructose on non-alcoholic fatty liver disease pathogenesisNon-alcoholic fatty liver disease (NAFLD) is a broad-spectrum liver disorder that ranges from simple hepatic steatosis to its more severe form, non-alcoholic steatohepatitis, which is marked by hepatic inflammation and cirrhosis. Estimates indicated NAFLD prevalence is currently between 25-30% of adults worldwide. Currently, there are no treatment options for NAFLD other than weight loss, as the etiology of this disease remains elusive. Prior studies have indicated that NAFLD is likely a multifactorial disease arising from interactions between factors like: genetics, nutrition, age, sex, hormone levels, lipid regulation, and biometals homeostasis. While prior studies have largely done an excellent job at defining the NAFLD landscape there are many factors known to impact NAFLD pathogenesis that remain understudied. This study aimed to examine the impacts of some of these factors; namely, biometals homeostasis, dietary fructose intake, and sex. This study first reviewed prior findings to outline the current understanding of the impacts of biological Cu on lipid metabolism and regulation. Next, we used dietary treatments in an animal model (Wistar rat) to examine the interplay between sex, dietary fructose and biometals homeostasis in NAFLD pathogenesis. Finally, an integrated network analysis of human RNA-Seq data was conducted to identify key genes involved in NALFD pathogenesis that were correlated with our factors of interest. In our review we identified some of the potential mechanisms by which copper (Cu) deficiency may promote NAFLD. These included increased immune cell activation, increased lipid biosynthesis, increased oxidative stress, and alterations to mitochondrial metabolism regulation. 1H nuclear magnetic resonance metabolomics results from the animal model identified distinct metabolic phenotypes that were highly correlated with diet and sex. Additionally, ICP-MS and western blot analysis identified previously unknown sex-specific responses to both fructose supplementation and restricted copper intake. Finally, our RNA-Seq integrated network analysis of human samples identified 66 total and 10 novel hub genes that are likely key regulators of NAFLD-related pathways. Enrichment analysis of the hub genes identified overrepresentation of genes related to the CAMKK2 pathway, oxidative phosphorylation, and regulation of mitochondrial transcription. All of the hub genes correlated with both disease status and sex were found to escape X-inactivation, suggesting XIST escapee genes may disproportionately impact sex-specific mechanisms of NAFLD pathogenesis. These results suggest sex, biometals homeostasis, and dietary fructose all impact NAFLD pathogenesis and further investigation into their roles should be pursued.
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Effects of diet and hibernation in skeletal muscle performanceHibernating animals, such as the arctic ground squirrel (AGS), are subjected to a wide range of temperature variations. During hibernation, when they are predominantly physically inactive, body temperature can drop as low as -3°C, while in summer, body temperature can climb as high as 40°C. Torpor is a state of inactivity in an animal induced by a lack of food, which is followed by a fall in body temperature and metabolic rate. Torpor lasts around 21 days in AGS, and the progression of torpor begins with early torpor, followed by mid and late torpor, and culminates with interbout arousal lasting less than 24 hours. AGS exhibit reduced muscular atrophy and protein loss despite lengthy periods of immobility, hypometabolism, and severe hypothermic conditions during hibernation. Skeletal muscle plasticity, unique to mammalian hibernators, may explain why cardiac and respiratory skeletal muscle can function at hypothermic temperatures during and after hibernation. As a result, understanding the effects of ambient temperature on muscle physiology and contractile function is critical. The focus of this research was to investigate skeletal muscle contractile performance and fatigue resistance in ex vivo muscle tissues during hypothermic temperature stress. Ex vivo tissue organ bath functional assays were performed in hibernator and/or non-hibernator rodent models to determine changes in performance and fatigue resistance in the AGS diaphragm induced by polyunsaturated fatty acid dietary modification or ambient hypothermic stress. This study lends support to the idea that diet and hypothermic stress might modify certain functional aspects of skeletal muscle, most likely via membrane lipid composition, ambient temperature, and torpor interaction. Furthermore, summer active AGS has a higher fatigue resistance than mid torpor AGS during the hibernating season. Furthermore, skeletal muscle fatigue resistance was significantly lower in Sprague Dawley rats than in both summer active and hibernating AGS. Preliminary data also suggested that hypothermic stress, to some extent, enhanced fatigue resistance regardless of torpor status or species difference.
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Development of a MUC1 cancer immunotherapy using complement targeted liposomes and organoplatinum (IV) complex as an effective anticancer drugThe development of therapeutic and preventive cancer vaccines has been on the rise in the past decade. Extensive tumor antigen libraries supply researchers with a wide variety of targets for developing cancer vaccines. Cancer vaccines aim to induce tumor regression, eradicate tumors and establish lasting antitumor memory, while avoiding adverse reactions. However, achieving this goal is hampered by the challenges of finding tumor antigens to incite an effective immune response and creating an efficient delivery system to deliver the antigen to target cells. In this thesis, we discuss how to integrate Toll Like Receptor (TLR) agonists with a MUC1 cancer vaccine to improve its effectiveness. The goal of our research was to increase drug delivery targeting efficiency by encapsulating a MUC1 immunogenic peptide within C3 liposomes. The work described herein shows that C3 liposomes were effective at targeting delivery of MUC1 antigen to Antigen Presenting Cells (APCs), which resulted in a T and B cell immune response against MUC1. Furthermore, the addition of TLR agonists to vaccine formulations boosted vaccine effectiveness, creating a more potent treatment against MUC1 expressing cancer. Another goal of this thesis research was to synthesize organoplatinum compounds. Currently, platinum-based chemotherapy drugs are a popular option for treating cancer, however, treatment is still limited by bystander toxicity. Four octahedral organoplatinum (IV) compounds, namely [Pt(CH₃)₂X₂{bipy-R₂}] (X = Br, I; bipy-R₂ = 2,2'-bipyridine, 2,2'-bipyridine-4,4'-dicarboxylic acid), have been isolated and structurally characterized by single-crystal X-ray diffraction. The anticancer potential of each compound was assessed via an in vitro MTT assay.
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An examination of a sleep-related hypermotor epilepsy associated mutation in complex α4β2* nicotinic acetylcholine receptorsEpilepsy is a disease that involves large networks of neurons. As such, studying the pathology of epilepsy has been complicated. Investigating epilepsy through the lens of Sleep-related Hypermotor Epilepsy (SHE) presents itself as a good strategy as SHE is linked to a variety of single point mutations. These mutations affect receptors, the most prominent being mutations in nicotinic acetylcholine receptors (nAChRs). nAChRs are pentameric ligand-gated ion channels. Five individual subunits combine to form a functional receptor. These nAChRs can either be homomeric, where all 5 subunits are the same, or heteromeric where there are different subunits combining. The most common CNS nAChRs are the homomeric [alpha] 7 nAChR and the heteromeric [alpha]4[beta]2 nAChR. Within these heteromeric nAChRs other subunits, such as the [alpha] 3 and [alpha] 5 subunits, can be incorporated into the accessory position. Many of the mutations in nAChRs linked to SHE are present in the transmembrane (TM) regions of the [alpha]4 and [beta]2 subunits of these [alpha]4[beta]2 nAChRs. However, less is known about the SHE mutations present in the intracellular loop regions. The [alpha]4[beta]2 subtype is widely distributed throughout the CNS. Our study looks at both [alpha]4[beta]2 isoforms as well as the more complex [alpha]4[beta]2)₂[alpha]3 and [alpha]4[beta]2)₂[alpha]5. The [alpha]3 and [alpha]5 accessory subunits are more highly expressed within the thalamocortical loop, the network implicated in SHE. Some prior research has been conducted on the [alpha]4[beta]2)₂[alpha]5 subtype and has shown that incorporation of the [alpha] 5 subunit increases the resulting receptor's Ca²⁺ permeability. Minimal work has been done on [alpha]4[beta]2)₂[alpha]3 subtype but research has shown that incorporating the [alpha]3 subunit into the [alpha]4[beta]2 nAChR results in low sensitivity to ACh and can be potentiated by compounds like NS-9283. We investigated the incorporation of the [alpha]4R³³⁶H SHE mutation into the [alpha]4[beta]2 isoforms as well as the ([alpha]4[beta]2)₂[alpha]3 and ([alpha]4[beta]2)₂[alpha]5 subtypes. Our results revealed that the [alpha]4R³³⁶H SHE mutation altered ACh pharmacology and overall receptor function. We also demonstrated that incorporation of the [alpha]3 or [alpha]5 subunits into the [alpha]4[beta]2 subtype caused changes in response to physiologically-relevant ligands. Our study adds to the knowledge of how the [alpha]4R³³⁶H SHE mutation contributes to changes in nAChR pharmacology and function. We also add to the growing knowledge of residues within the intracellular cytoplasmic loop region and their importance. Results from this study may be applicable to future studies involving generalized epilepsy, in which nAChRs are implicated. Study results could also be applicable to other neurological disorders such as Alzheimer's disease, Parkinson's disease, autism, and schizophrenia. These diseases, in many instances, involve alterations to various nAChR subtypes such as [alpha]7, [alpha]4[beta]2, [alpha]5 containing and [alpha]3-containing nAChRs.
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Impairment of DNA repair by oncogenic virus HHV8Cancer is a growing disease burden worldwide. Lifestyle and environmental factors are the largest contributors to cancer development (90-95%) in contrast to genetics (5-10% of cases,); of the environmental factors, infections contribute 15-20% of the "external" risk in developing cancer. In the US, breast cancer is the most common type of cancer with 284,000 new cases expected in 2021, with the next most common being prostate and lung cancer. Viruses have long been suspected to promote cancerous transformations. Kaposi Sarcoma Associated Herpesvirus (KSHV) is implicated in Kaposi Sarcoma (KS) cancer transformation and in vitro is found in BCBL1 cells. KSHV is a herpesvirus, also named Human Herpesvirus 8 (HHV8). The connection between viral infection and increase in cancer has been documented, but the details of cellular mechanisms involved therein are lacking. We hypothesize that during active viral infection, cellular DNA is not repaired properly, leading to the accumulation of DNA damage and mutations; these in turn, could lead to the transformation of normal, albeit virus-infected cells, into cancer-predisposed cells, and later tumor formation. The goal of this thesis was to investigate DNA repair during an active viral infection of HHV8. BCBL1 is a clonal lymphoma cell line isolated in 1995 from an HIV seronegative patient with a body cavity-based lymphoma. Chapter 1 is a mini review on what is known about this oncogenic virus, DNA repair pathways, and cancer and how these three relate to each other is discussed. Chapter 2 compares DNA repair dynamics of two distinct repair pathways in virally active cells versus virally dormant cells. Our research suggests that host processes controlling DNA repair mechanisms are manipulated to favor virus production. Chapter 3 is a general conclusion summarizing our significant findings from Chapter 2.
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3T3-L1 adipocytes as a model of Glut4 translocationType 2 diabetes (T2D), a lethal disease, reveals an alarming increase of epidemic proportions and, by 2050, an estimated 1 in 3 adults in the US will suffer from it. Persistent and systemic inflammatory and oxidative stress in adipose tissue and skeletal muscle are recognized as key players in the progression of T2D. T2D is characterized by insulin resistance, the inability of cells to respond to standard circulating levels of insulin, resulting in hyperglycemia. In adipocytes and skeletal muscle, glucose uptake, a vital step in blood glucose regulation, predominantly occurs via the glucose transporter 4 (Glut4) in an insulindependent recruitment from a cytosolic vesicle. Under inflammatory stress, these cells have demonstrated a lack of Glut4 presence in the plasma membrane, leading to a lack of glucose transport within these cells. 3T3-L1 murine adipocytes are a key model to study insulin signaling in vitro. Mature adipocytes are converted from fibroblasts in a lengthy process requiring chemical induction. Several inducers have been tested in hopes to shorten the time span required for differentiation. However, the use of different inducers and changes in the differentiation programming could have consequences on the phenotype of the mature adipocytes. For this reason, we sought to evaluate the differences in 3T3-L1 fibroblasts induced with either 3-isobutyl-1-methylxanthine (IBMX), or troglitazone. In addition to pharmaceutical recourse to restore insulin signaling and glucose uptake in cells under inflammatory duress, a dietary approach has been considered. Natural products such as blueberries have been of particular interest due to their health benefit including antioxidant benefits and their ability to modulate biochemical pathways. Wild Alaskan blueberries are of particular interest due to their link to a reduced prevalence of T2D in Alaskan Native populations. Here, we report differences in the phenotype of 3T3-L1 adipocytes based on the inducer of adipogenesis and a lack of increase in plasma membrane Glut4. Our studies demonstrate a higher expression of adipocyte biomarkers in cells treated with troglitazone compared to IBMX, but a higher expression of total Glut4 in cells treated with IBMX compared to troglitazone. In both phenotypes, we note a lack of insulin-induced increase of plasma membrane levels of Glut4 in 3T3-L1 mature adipocytes. Finally, we demonstrate an insulin-like benefit of Alaskan blueberry extracts on plasma membrane Glut4.
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Age related dementiasDuring the last few decades, the average life expectancy has dramatically increased in the U.S.A. and globally. As a result, the diseases associated with age, such as dementia, significantly impact families, society, and government. Dementia is a broad term describing memory loss, cognitive dysfunctions, and limited social skills caused by many factors such as aging, stroke, and genetic background. Although the primary etiology remains unclear but inflammatory reactions, neurodegeneration, vascular abnormalities play an important role in the progression of these disorders. Therefore, treatment is mainly supportive to relieve the symptoms of dementia and not to cure dementia or slow down its progression. This review discusses the most common disorders that cause dementia, including Alzheimer's disease, vascular dementia, and Parkinson's disease, as well as their symptoms, diagnosis, and treatments. Also covered are the studies that were performed on these disorders and animal models for each disease.
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Assessment of an Alaskan willow with potential for polychlorinated biphenyl rhizoremediation"Polychlorinated Biphenyls (PCBs) are priority pollutants targeted for remediation efforts in many areas of the world. Rhizoremediation, the use of plants and their associated root microorganisms for the degradation of a pollutant, may be an inexpensive and effective approach to biodegrading PCBs and detoxifying contaminated soils in situ. Two Alaskan native tree species, Salix alaxensis (willow) or Picea glauca (white spruce), were assessed for the ability to stimulate microbial PCB degradation by measuring PCB loss, toxicity and microbial community shifts in soil microcosms following the addition of crushed roots. Incubation of polluted soils with root crushates of S. alaxensis led to significant losses (up to 30% in 180 days) of several PCB congeners, including many of the most toxic congeners, PCB 77, 105 and 169. Soil toxicity, measured using the Microtox assay, also decreased as a result of treatment with willow-roots. Treatment of soils with salicylate, a willow secondary compound hypothesized to promote aromatic pollutant biodegradation, inhibited PCB degradation, suggesting that other willow biodegradation are responsible for biostimulation. The disappearance of PCB congeners, detoxification of soil, and detection of microbes with PCB degrading abilities suggests that S. alaxensis is a promising plant candidate for rhizoremediation of PCBs"--Leaf iii
