Angela M. Wilson

Expression of the yeast NADH dehydrogenase Ndi1 in Drosophila confers increased lifespan independently of dietary restriction

Mutations in mitochondrial oxidative phosphorylation complex I are associated with multiple pathologies, and complex I has been proposed as a crucial regulator of animal longevity. In yeast, the single-subunit NADH dehydrogenase Ndi1 serves as a non-proton-translocating alternative enzyme that replaces complex I, bringing about the reoxidation of intramitochondrial NADH. We have created transgenic strains of Drosophila that express yeast NDI1 ubiquitously. Mitochondrial extracts from NDI1-expressing flies displayed a rotenone-insensitive NADH dehydrogenase activity, and functionality of the enzyme in vivo was confirmed by the rescue of lethality resulting from RNAi knockdown of complex I. NDI1 expression increased median, mean, and maximum lifespan independently of dietary restriction, and with no change in sirtuin activity. NDI1 expression mitigated the aging associated decline in respiratory capacity and the accompanying increase in mitochondrial reactive oxygen species production, and resulted in decreased accumulation of markers of oxidative damage in aged flies. Our results support a central role of mitochondrial oxidative phosphorylation complex I in influencing longevity via oxidative stress, independently of pathways connected to nutrition and growth signaling.

Phytoremediation of Organic Contaminants: A Review of Phytoremediation Research at the University of Washington

As overwhelmingly positive results have become available regarding the ability of plants to degrade compounds such as trichloroethylene, phytoremediation studies are expanding. Studies to determine the potential for phytoremediation of fully chlorinated compounds, such as carbon tetrachloride and tetrachloroethylene, brominated compounds, such as ethylene dibromide and dibromochloropropane, and nonhalogenated compounds, such as methyl-t-butyl ether (MTBE), are underway. When using phytoremediation, it is important to select not only a plant that is capable of degrading the pollutant in question, but also one that will grow well in that specific environment. In ecologically sensitive areas, such as the Hawaiian Islands, only plants native to the area can be used. One way to supplement the arsenal of plants available for remedial actions is to utilize genetic engineering tools to insert into plants those genes that will enable the plant to metabolize a particular pollutant. Hybrid technologies, such as usin...

Identification of glutathione S-transferase pi as a protein involved in Parkinson disease progression.

Parkinson disease (PD) typically affects the cortical regions during the later stages of disease, with neuronal loss, gliosis, and formation of diffuse cortical Lewy bodies in a significant portion of patients with dementia. To identify novel proteins involved in PD progression, we prepared synaptosomal fractions from the frontal cortices of pathologically verified PD patients at different stages along with age-matched controls. Protein expression profiles were compared using a robust quantitative proteomic technique. Approximately 100 proteins displayed significant differences in their relative abundances between PD patients at various stages and controls; three of these proteins were validated using independent techniques. One of the confirmed proteins, glutathione S-transferase Pi, was further investigated in cellular models of PD, demonstrating that its level was intimately associated with several critical cellular processes that are directly related to neurodegeneration in PD. These results have, for the first time, suggested that the levels of glutathione S-transferase Pi may play an important role in modulating the progression of PD.

Trichloroethylene oxidative metabolism in plants: the trichloroethanol pathway

Trichloroethylene (TCE) is a widespread and persistent environmental contaminant. Recently, plants, poplar trees in particular, have been investigated as a tool to remove TCE from soil and groundwater. The metabolism of TCE in plants is being investigated for two reasons: one, plant uptake and metabolism represent an important aspect of the environmental fate of the contaminant; two, metabolism pattern and metabolite identification will help assess the applicability of phytoremediation. It was previously shown that TCE metabolites in plants are similar to ones that result from cytochrome P450-mediated oxidation in mammals: trichloroethanol, trichloroacetate and dichloroacetate. Our measurements indicate that one of these metabolites, trichloroethanol, is further glycosylated in tobacco and poplar. The glycoside was detected in all tissues (roots, stems and leaves) in comparable levels, and was at least 10 fold more abundant than free trichloroethanol. The glycoside in tobacco was identified as the ss-D-glucoside of trichloroethanol by comparison of the mass spectra and the chromatographic retention time of its acetylation product to that of the synthesized standard. Trichloroethanol and its glucoside did not persist in plant tissue once plants are removed from TCE contaminated water, indicating further metabolism.

Attenuated progression of diet-induced steatohepatitis in glutathione-deficient mice

In nonalcoholic fatty liver disease (NAFLD), depletion of hepatic antioxidants may contribute to the progression of steatosis to nonalcoholic steatohepatitis (NASH) by increasing oxidative stress that produces lipid peroxidation, inflammation, and fibrosis. We investigated whether depletion of glutathione (GSH) increases NASH-associated hepatic pathology in mice fed a diet deficient in methionine and choline (MCD diet). Wild-type (wt) mice and genetically GSH-deficient mice lacking the modifier subunit of glutamate cysteine ligase (Gclm null mice), the rate-limiting enzyme for de novo synthesis of GSH, were fed the MCD diet, a methionine/choline-sufficient diet, or standard chow for 21 days. We assessed NASH-associated hepatic pathology, including steatosis, fibrosis, inflammation, and hepatocyte ballooning, and used the NAFLD Scoring System to evaluate the extent of changes. We measured triglyceride levels, determined the level of lipid peroxidation products, and measured by qPCR the expression of mRNAs for several proteins associated with lipid metabolism, oxidative stress, and fibrosis. MCD-fed GSH-deficient Gclm null mice were to a large extent protected from MCD diet-induced excessive fat accumulation, hepatocyte injury, inflammation, and fibrosis. Compared with wt animals, MCD-fed Gclm null mice had much lower levels of F2-isoprostanes, lower expression of acyl-CoA oxidase, carnitine palmitoyltransferase 1a, uncoupling protein-2, stearoyl-coenzyme A desaturase-1, transforming growth factor-β, and plasminogen activator inhibitor-1 mRNAs, and higher activity of catalase, indicative of low oxidative stress, inhibition of triglyceride synthesis, and lower expression of profibrotic proteins. Global gene analysis of hepatic RNA showed that compared with wt mice, the livers of Gclm null mice have a high capacity to metabolize endogenous and exogenous compounds, have lower levels of lipogenic proteins, and increased antioxidant activity. Thus, metabolic adaptations resulting from severe GSH deficiency seem to protect against the development of steatohepatitis.

Free radical damage to cerebral cortex in Alzheimer's Disease, Microvascular Brain Injury, and Smoking

Evidence supports a pathogenic role for free radical injury to brain in Alzheimers disease; however, clinical trial results are only mildly encouraging. Examining brains from The Adult Changes in Thought study offers a unique perspective. Selectively increased free radical damage to cerebral cortex was associated with Alzheimers disease, microvascular brain injury, and current smoking, but not with antioxidant supplement usage. Our results support suppression of free radical injury to brain as a therapeutic target for Alzheimers disease and microvascular brain injury; however, future clinical trials should consider other antioxidants or doses than those identified in our study. Ann Neurol 2009;65:226–229

Metabolism of the soil and groundwater contaminants, ethylene dibromide and trichloroethylene, by the tropical leguminous tree, Leuceana leucocephala

Ethylene dibromide (EDB; dibromoethane) and trichloroethylene (TCE) are hazardous environmental pollutants. The use of plants to treat polluted sites and groundwater, termed phytoremediation, requires plants that can both effectively remove the pollutant as well as grow in the climatic region of the site. In this paper, we report that the tropical leguminous tree, Leuceana leucocephala var. K636, is able to take up and metabolize EDB and TCE. The plants were grown in sterile hydroponic solution without its symbiont, Rhizobium. EDB and TCE were both metabolized by the plant, as indicated by the formation of bromide ion from EDB and trichloroethanol from TCE. Each plant organ was independently capable of debromination of EDB. L. leucocephala is being used to treat perched groundwater as part of a remedial alternative to address an accidental EDB spill in Hawaii. Bromide levels of plant tissues from the trees grown in the phytoremediation treatment cells at the Hawaii Site were elevated, indicating uptake and degradation of brominated compounds in the trees. This report is the first evidence of a tropical tree effectively metabolizing these common organic pollutants.

Inflammation and Oxidative Stress Markers and Esophageal Adenocarcinoma Incidence in a Barrett's Esophagus Cohort

Background: Persons with Barretts esophagus experience increased risk of esophageal adenocarcinoma. Prediagnostic inflammation markers predict several cancers, but their role in predicting esophageal adenocarcinoma is unknown. Methods: We investigated whether biomarkers of inflammation [C-reactive protein (CRP), interleukin-6 (IL6), soluble tumor necrosis factor (sTNF) receptors I and II], and of oxidative stress (F2-isoprostanes) predicted progression to esophageal adenocarcinoma in a prospective cohort of 397 patients with Barretts esophagus, 45 of whom developed esophageal adenocarcinoma. Biomarkers were measured in stored plasma samples from two time points during follow-up, the mean of which served as the primary predictor. Adjusted hazard ratios (HR) and 95% confidence intervals (CI) were estimated using Cox regression. Results: CRP level above the median was associated with an 80% increased risk of esophageal adenocarcinoma. The HR and 95% CI adjusted for age, gender, and further adjusted for waist–hip ratio and smoking were 1.98 (1.05–3.73) and 1.77 (0.93–3.37), respectively, with Ptrend for continuous CRP = 0.04. Persons with IL6 levels above the median also had almost 2-fold increased risk [HR and 95% CI adjusted for age and gender, and further adjusted for waist–hip ratio and smoking were 1.95 (1.03–3.72) and 1.79 (0.93–3.43), respectively, but no evidence of a trend was observed]. Concentrations of TNF receptors and F2-isoprostanes were not associated with esophageal adenocarcinoma risk. Conclusions: Further research is needed to evaluate the role of inflammation and associated markers in esophageal adenocarcinoma development in persons with Barretts esophagus. Impact: This prospective study suggests that inflammation markers, particularly CRP and IL6, may help identify persons at higher risk of progression to esophageal adenocarcinoma. Cancer Epidemiol Biomarkers Prev; 23(11); 2393–403. ©2014 AACR.

Cerebrospinal Fluid Particles in Alzheimer Disease and Parkinson Disease

Abstract Human cerebrospinal fluid (CSF) contains diverse lipid particles, including lipoproteins that are distinct from their plasma counterparts and contain apolipoprotein (apo) E isoforms, apoJ, and apoAI, and extracellular vesicles, which can be detected by annexin V binding. The aim of this study was to develop a method to quantify CSF particles and evaluate their relationship to aging and neurodegenerative diseases. We used a flow cytometric assay to detect annexin V-, apoE-, apoAI-, apoJ-, and amyloid (A) &bgr;42-positive particles in CSF from 131 research volunteers who were neurologically normal or had mild cognitive impairment (MCI), Alzheimer disease (AD) dementia, or Parkinson disease. APOE &egr;4/&egr;4 participants had CSF apoE-positive particles that were more frequently larger but at an 88% lower level versus those in APOE &egr;3/&egr;3 or APOE &egr;3/&egr;4 patients; this finding was reproduced in conditioned medium from mouse primary glial cell cultures with targeted replacement of apoE. Cerebrospinal fluid apoE-positive and &bgr;-amyloid (A&bgr;42)-positive particle concentrations were persistently reduced one-third to one-half in middle and older age subjects; apoAI-positive particle concentration progressively increased approximately 2-fold with age. Both apoAI-positive and annexin V-positive CSF particle levels were reduced one-third to one-half in CSF of MCI and/or AD dementia patients versus age-matched controls. Our approach provides new methods to investigate CNS lipid biology in relation to neurodegeneration and perhaps develop new biomarkers for diagnosis or treatment monitoring.

Partial depletion of striatal dopamine enhances penetrance of cognitive deficits in a transgenic mouse model of Alzheimer's disease.

Parkinsons disease and Alzheimers disease (AD) are recognized to coexist on a spectrum of neurodegeneration, and it has been proposed that molecular interactions among pathogenic proteins are a basis for the overlap between these two diseases. We instead hypothesized that degeneration of the nigrostriatal dopaminergic system enhances the clinical penetrance of early‐stage AD. To determine the effect of striatal dopamine (DA) on the pathological effects in an experimental model of AD, APPSWE/PS1ΔE9 mice received striatal injections of the neurotoxin 6‐hydroxydopamine (6OHDA). Animals were tested in a Barnes maze protocol and in a water T‐maze protocol at different ages to determine the onset of cognitive impairment. APPSWE/PS1ΔE9 mice that received 6OHDA injections showed significant impairment in Barnes maze performance at an earlier age than controls. Additionally, at 12 months of age, APPswe/PS1ΔE9 + 6OHDA mice demonstrated worse behavioral flexibility than other groups in a task‐switch phase of the water T‐maze. To determine the neuroprotective effects of dopaminergic neurotransmission against amyloid‐β42 (Aβ42) toxicity, neuronal branch order and dendrite length were quantified in primary medium spiny neuron (MSN) cultures pretreated with increasing doses of the D1 and D2 receptor agonists before being exposed to oligomerized Aβ42. Although there were no differences in Aβ peptide levels or plaque burden among the groups, in murine MSN culture dopaminergic agonists prevented a toxic response to Aβ42. Depletion of DA in the striatum exacerbated the cognitive impairment seen in a mouse model of early‐stage AD; this may be due to a protective effect of dopaminergic innervation against Aβ striatal neurotoxicity.