| 1. |
- Bruce, Stephen J, et al.
(författare)
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Evaluation of a protocol for metabolic profiling studies on human blood plasma by combined ultra-performance liquid chromatography/mass spectrometry : From extraction to data analysis
- 2009
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Ingår i: Analytical Biochemistry. - : Elsevier. - 0003-2697 .- 1096-0309. ; 372:2, s. 237-249
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Tidskriftsartikel (refereegranskat)abstract
- The investigation presented here describes a protocol designed to perform high-throughput metabolic profiling analysis on human blood plasma by ultra-performance liquid chromatography/mass spectrometry (UPLC/MS). To address whether a previous extraction protocol for gas chromatography (GC)/MS-based metabolic profiling of plasma could be used for UPLC/MS-based analysis, the original protocol was compared with similar methods for extraction of low-molecular-weight compounds from plasma via protein precipitation. Differences between extraction methods could be observed, but the previously published extraction method was considered the best. UPLC columns with three different stationary phases (C8, C18, and phenyl) were used in identical experimental runs consisting of a total of 60 injections of extracted male and female plasma samples. The C8 column was determined to be the best for metabolic profiling analysis on plasma. The acquired UPLC/MS data of extracted male and female plasma samples was subjected to principal component analysis (PCA) and orthogonal projections to latent structures discriminant analysis (OPLS–DA). Furthermore, a strategy for compound identification was applied here, demonstrating the strength of high-mass-accuracy time-of-flight (TOF)/MS analysis in metabolic profiling.
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| 2. |
- Jiye, A, et al.
(författare)
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Extraction and GC/MS analysis of the human blood plasma metabolome
- 2005
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Ingår i: ANALYTICAL CHEMISTRY. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 77:24, s. 8086-94
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Tidskriftsartikel (refereegranskat)abstract
- Analysis of the entire set of low molecular weight compounds (LMC), the metabolome, could provide deeper insights into mechanisms of disease and novel markers for diagnosis. In the investigation, we developed an extraction and derivatization protocol, using experimental design theory (design of experiment), for analyzing the human blood plasma metabolome by GC/MS. The protocol was optimized by evaluating the data for more than 500 resolved peaks using multivariate statistical tools including principal component analysis and partial least-squares projections to latent structures (PLS). The performance of five organic solvents (methanol, ethanol, acetonitrile, acetone, chloroform), singly and in combination, was investigated to optimize the LMC extraction. PLS analysis demonstrated that methanol extraction was particularly efficient and highly reproducible. The extraction and derivatization conditions were also optimized. Quantitative data for 32 endogenous compounds showed good precision and linearity. In addition, the determined amounts of eight selected compounds agreed well with analyses by independent methods in accredited laboratories, and most of the compounds could be detected at absolute levels of similar to 0.1 pmol injected, corresponding to plasma concentrations between 0.1 and 1 mu M. The results suggest that the method could be usefully integrated into metabolomic studies for various purposes, e.g., for identifying biological markers related to diseases.
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| 3. |
- Nordin, Angelica, et al.
(författare)
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Extensive size variability of the GGGGCC expansion in C9orf72 in both neuronal and non-neuronal tissues in 18 patients with ALS or FTD
- 2015
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Ingår i: Human Molecular Genetics. - : Oxford University Press (OUP). - 0964-6906 .- 1460-2083. ; 24:11, s. 3133-3142
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Tidskriftsartikel (refereegranskat)abstract
- A GGGGCC-repeat expansion in C9orf72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) among Caucasians. However, little is known about the variability of the GGGGCC expansion in different tissues and whether this correlates with the observed phenotype. Here, we used Southern blotting to estimate the size of hexanucleotide expansions in C9orf72 in neural and non-neural tissues from 18 autopsied ALS and FTD patients with repeat expansion in blood. Digitalization of the Southern blot images allowed comparison of repeat number, smear distribution and expansion band intensity between tissues and between patients. We found marked intra-individual variation of repeat number between tissues, whereas there was less variation within each tissue group. In two patients, the size variation between tissues was extreme, with repeat numbers below 100 in all studied non-neural tissues, whereas expansions in neural tissues were 20-40 times greater and in the same size range observed in neural tissues of the other 16 patients. The expansion pattern in different tissues could not distinguish between diagnostic groups and no correlation was found between expansion size in frontal lobe and occurrence of cognitive impairment. In ALS patients, a less number of repeats in the cerebellum and parietal lobe correlated with earlier age of onset and a larger number of repeats in the parietal lobe correlated with a more rapid progression. In 43 other individuals without repeat expansion in blood, we find that repeat sizes up to 15 are stable, as no size variation between blood, brain and spinal cord was found.
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| 4. |
- Wu, Junfang, et al.
(författare)
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NMR analysis of the CSF and plasma metabolome of rigorously matched amyotrophic lateral sclerosis, Parkinson's disease and control subjects
- 2016
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Ingår i: Metabolomics. - : Springer Science and Business Media LLC. - 1573-3882 .- 1573-3890. ; 12:6
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Tidskriftsartikel (refereegranskat)abstract
- Introduction: Amyotrophic lateral sclerosis (ALS) and Parkinson’s disease (PD) are two severe neurodegenerative disorders for which the disease mechanisms are poorly understood and reliable biomarkers are absent.Objectives: To identify metabolite biomarkers for ALS and PD, and to gain insights into which metabolic pathways are involved in disease.Methods: Nuclear magnetic resonance (NMR) metabolomics was utilized to characterize the metabolite profiles of cerebrospinal fluid (CSF) and plasma from individuals in three age, gender, and sampling-date matched groups, comprising 22 ALS, 22 PD and 28 control subjects.Results: Multivariate analysis of NMR data generated robust discriminatory models for separation of ALS from control subjects. ALS patients showed increased concentrations of several metabolites in both CSF and plasma, these are alanine (CSF fold change = 1.22, p = 0.005), creatine (CSF-fc = 1.17, p = 0.001), glucose (CSF-fc = 1.11, p = 0.036), isoleucine (CSF-fc = 1.24, p = 0.002), and valine (CSF-fc = 1.17, p = 0.014). Additional metabolites in CSF (creatinine, dimethylamine and lactic acid) and plasma (acetic acid, glutamic acid, histidine, leucine, pyruvate and tyrosine) were also important for this discrimination. Similarly, panels of CSF-metabolites that discriminate PD from ALS and control subjects were identified.Conclusions: The results for the ALS patients suggest an affected creatine/creatinine pathway and an altered branched chain amino acid (BCAA) metabolism, and suggest links to glucose and energy metabolism. Putative metabolic markers specific for ALS (e.g. creatinine and lactic acid) and PD (e.g. 3-hydroxyisovaleric acid and mannose) were identified, while several (e.g. creatine and BCAAs) were shared between ALS and PD, suggesting some overlap in metabolic alterations in these disorders.
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| 5. |
- Wuolikainen, Anna, et al.
(författare)
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Multi-platform mass spectrometry analysis of the CSF and plasma metabolomes of rigorously matched amyotrophic lateral sclerosis, Parkinson's disease and control subjects
- 2016
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Ingår i: Molecular Biosystems. - : Royal Society of Chemistry (RSC). - 1742-206X .- 1742-2051. ; 12:4, s. 1287-1298
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Tidskriftsartikel (refereegranskat)abstract
- Amyotrophic lateral sclerosis (ALS) and Parkinson's disease (PD) are protein-aggregation diseases that lack clear molecular etiologies. Biomarkers could aid in diagnosis, prognosis, planning of care, drug target identification and stratification of patients into clinical trials. We sought to characterize shared and unique metabolite perturbations between ALS and PD and matched controls selected from patients with other diagnoses, including differential diagnoses to ALS or PD that visited our clinic for a lumbar puncture. Cerebrospinal fluid (CSF) and plasma from rigorously age-, sex- and sampling-date matched patients were analyzed on multiple platforms using gas chromatography (GC) and liquid chromatography (LC)-mass spectrometry (MS). We applied constrained randomization of run orders and orthogonal partial least squares projection to latent structure-effect projections (OPLS-EP) to capitalize upon the study design. The combined platforms identified 144 CSF and 196 plasma metabolites with diverse molecular properties. Creatine was found to be increased and creatinine decreased in CSF of ALS patients compared to matched controls. Glucose was increased in CSF of ALS patients and alpha-hydroxybutyrate was increased in CSF and plasma of ALS patients compared to matched controls. Leucine, isoleucine and ketoleucine were increased in CSF of both ALS and PD. Together, these studies, in conjunction with earlier studies, suggest alterations in energy utilization pathways and have identified and further validated perturbed metabolites to be used in panels of biomarkers for the diagnosis of ALS and PD.
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