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Imaging-guided microarray: Identifies molecular markers in the pathogenesis of Parkinson’s disease

ABSTRACT: The full complement of molecular pathways contributing to Parkinson’s disease (PD) pathogenesis remains unknown. Here, to address this issue, we began by using a high-resolution variant of functional magnetic resonance imaging (fMRI) to pinpoint brainstem regions differentially affected by, and resistant to, the disease. Then, relying on the imaging information as a guide, we profiled gene expression levels of postmortem brain samples and used a factorial statistical model to identify a disease related decrease in the expression of the polyamine enzyme spermidine/spermine N1-acetyltransferase 1 (SAT1). Next, a series of studies were performed to confirm the pathogenic relevance of this finding. First, to test for a causal link between polyamines and α-synuclein toxicity, we investigated a yeast model expressing α-synuclein. Polyamines were found to enhance the toxicity of α-synuclein, and an unbiased genome-wide screen for modifiers of α-synuclein toxicity identified Tpo4, a member of a family of proteins responsible for polyamine transport. Second, to test for a causal link between SAT1 activity and PD histopathology we investigated a mouse model expressing α-synuclein. DENSPM (N1, N11-diethylnorspermine), a polyamine analog that increases SAT1 activity, was found to reduce PD histopathology, while Berenil (diminazene aceturate), a pharmacological agent that reduces SAT1 activity, worsened the histopathology. Third, we genotyped PD patients and controls and isolated a rare but novel variant in the SAT1 gene, although the functional significance of this genetic variant was not identified. Taken together, the results suggest that the polyamine pathway contributes to PD pathogenesis.

ORGANISM(S): Homo sapiens

PROVIDER: GSE19587 | GEO | 2010/08/19

SECONDARY ACCESSION(S): PRJNA122363

REPOSITORIES: GEO

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Project description:The full complement of molecular pathways contributing to Parkinson’s disease (PD) pathogenesis remains unknown. Here, to address this issue, we began by using a high-resolution variant of functional magnetic resonance imaging (fMRI) to pinpoint brainstem regions differentially affected by, and resistant to, the disease. Then, relying on the imaging information as a guide, we profiled gene expression levels of postmortem brain samples and used a factorial statistical model to identify a disease related decrease in the expression of the polyamine enzyme spermidine/spermine N1-acetyltransferase 1 (SAT1). Next, a series of studies were performed to confirm the pathogenic relevance of this finding. First, to test for a causal link between polyamines and α-synuclein toxicity, we investigated a yeast model expressing α-synuclein. Polyamines were found to enhance the toxicity of α-synuclein, and an unbiased genome-wide screen for modifiers of α-synuclein toxicity identified Tpo4, a member of a family of proteins responsible for polyamine transport. Second, to test for a causal link between SAT1 activity and PD histopathology we investigated a mouse model expressing α-synuclein. DENSPM (N1, N11-diethylnorspermine), a polyamine analog that increases SAT1 activity, was found to reduce PD histopathology, while Berenil (diminazene aceturate), a pharmacological agent that reduces SAT1 activity, worsened the histopathology. Third, we genotyped PD patients and controls and isolated a rare but novel variant in the SAT1 gene, although the functional significance of this genetic variant was not identified. Taken together, the results suggest that the polyamine pathway contributes to PD pathogenesis. Imaging-guided microarray In principle, gene expression profiling techniques like microarray are well suited to identify molecular pathways contributing to the pathogenesis of complex diseases. In practice, however, microarray applied to diseases of the brain present a number of analytic challenges. By identifying regions within the same brain structure that are differentially targeted by and resistant to a disease, imaging-guided microarray is an approach designed to address these limitations. Specifically, guided by the spatial information generated from high resolution functional imaging, a 2x2 factorial analysis-of-variance can be designed, including both within and between group factors, and this “double subtraction” model is effective in improving signal-to-noise in a microarray experiment. Relying on imaging findings, we harvested the DMNV from 6 postmortem brains with evidence of PD and from 5 control brains. The postmortem PD cases were evaluated for pathological changes (Lewy body-containing neurons and Lewy neurites evidenced with antibodies directed against α-synuclein aggregates) that matched the pattern proposed by Braak. We relied on the imaging results to identify a neighboring medullary region relatively unaffected by the disease to be used as a within-brain control. We decided on the inferior olivary nucleus (ION), because it is histologically identifiable, and harvested the ION from each of the 6 PD cases and 5 controls. Microarray techniques were used to generate gene expression profiles for each of the 22 tissue samples. A repeated-measures 2x2 factorial ANOVA model constructed for the imaging study was applied to the expression dataset, in which expression levels from two regions of the medulla (DMNV vs. ION) were included as the first within group factor, diagnosis (PD vs. controls) was the between group factor, and age and sex were included as covariates. Based on current literature, one of the top hits (SAT1) was investigated further to determine if it played a role in PD pathogenesis.

Project description:Non-small cell lung cancer (NSCLC) is the most lethal and prevalent type of lung cancer. In almost all types of cancer, the levels of polyamines (putrescine, spermidine, and spermine) are increased, playing a pivotal role in tumor proliferation. Indomethacin, a non-steroidal anti-inflammatory drug, increases the abundance of an enzyme termed spermidine/spermine-N1-acetyltransferase (SSAT) encoded by the SAT1 gene. This enzyme is a key player in the export of polyamines from the cell. The aim of this study was to compare the effect of indomethacin on two NSCLC cell lines, and their combinatory potential with polyamine-inhibitor drugs in NSCLC cell lines. A549 and H1299 NSCLC cells were exposed to indomethacin and evaluations included SAT1 expression, SSAT levels, and the metabolic status of cells. Moreover, the difference in polyamine synthesis enzymes among these cell lines as well as the synergistic effect of indomethacin and chemical inhibitors of the polyamine pathway enzymes on cell viability were investigated. Indomethacin increased the expression of SAT1 and levels of SSAT in both cell lines. In A549 cells, it significantly reduced the levels of putrescine and spermidine. However, in H1299 cells, the impact of treatment on the polyamine pathway was insignificant. Also, the metabolic features upstream of the polyamine pathway (i.e., ornithine and methionine) were increased. In A549 cells, the increase of ornithine correlated with the increase of several metabolites involved in the urea cycle. Evaluation of the levels of the polyamine synthesis enzymes showed that ornithine decarboxylase is increased in A549 cells, whereas S-adenosylmethionine-decarboxylase and polyamine oxidase are increased in H1299 cells. This observation correlated with relative resistance to polyamine synthesis inhibitors eflornithine and SAM486 (inhibitors of ornithine decarboxylase and S-adenosyl-L-methionine decarboxylase, respectively), and MDL72527 (inhibitor of polyamine oxidase and spermine oxidase). Finally, indomethacin demonstrated a synergistic effect with MDL72527 in A549 cells and SAM486 in H1299 cells. Collectively, these results indicate that indomethacin alters polyamine metabolism in NSCLC cells and enhances the effect of polyamine synthesis inhibitors, such as MDL72527 or SAM486. However, this effect varies depending on the basal metabolic fingerprint of each type of cancer cell.

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Imaging-guided microarray: Identifies molecular markers in the pathogenesis of Parkinson’s disease

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