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Ina Schuppe-Koistinen

Publications and source records attributed to Ina Schuppe-Koistinen.

12 recordsLinked to original sources
PubMed2006

Second-order peak detection for multicomponent high-resolution LC/MS data.

The first step when analyzing multicomponent LC/MS data from complex samples such as biofluid metabolic profiles is to separate the data into information and noise via, for example, peak detection. Due to the complex nature of this type of data, with problems such as alternating backgrounds and differing peak shapes, this can be a very complex task. This paper presents and evaluates a two-dimensional peak detection algorithm based on raw vector-represented LC/MS data. The algorithm exploits the fact that in high-resolution centroid data chromatographic peaks emerge flanked with data voids in the corresponding mass axis. According to the proposed method, only 4 per thousand of the total amount of data from a urine sample is defined as chromatographic peaks; however, 94% of the raw data variance is captured within these peaks. Compared to bucketed data, results show that essentially the same features that an experienced analyst would define as peaks can automatically be extracted with a minimum of noise and background. The method is simple and requires a priori knowledge of only the minimum chromatographic peak width-a system-dependent parameter that is easily assessed. Additional meta parameters are estimated from the data themselves. The result is well-defined chromatographic peaks that are consistently arranged in a matrix at their corresponding m/z values. In the context of automated analysis, the method thus provides an alternative to the traditional approach of bucketing the data followed by denoising and/or one-dimensional peak detection. The software implementation of the proposed algorithm is available at http://www.anchem.su.se/peakd as compiled code for Matlab.

Algorithms↗
PubMed2006

Predictive metabolite profiling applying hierarchical multivariate curve resolution to GC-MS data--a potential tool for multi-parametric diagnosis.

A method for predictive metabolite profiling based on resolution of GC-MS data followed by multivariate data analysis is presented and applied to three different biofluid data sets (rat urine, aspen leaf extracts, and human blood plasma). Hierarchical multivariate curve resolution (H-MCR) was used to simultaneously resolve the GC-MS data into pure profiles, describing the relative metabolite concentrations between samples, for multivariate analysis. Here, we present an extension of the H-MCR method allowing treatment of independent samples according to processing parameters estimated from a set of training samples. Predictions or inclusion of the new samples, based on their metabolite profiles, into an existing model could then be carried out, which is a requirement for a working application within, e.g., clinical diagnosis. Apart from allowing treatment and prediction of independent samples the proposed method also reduces the time for the curve resolution process since only a subset of representative samples have to be processed while the remaining samples can be treated according to the obtained processing parameters. The time required for resolving the 30 training samples in the rat urine example was approximately 13 h, while the treatment of the 30 test samples according to the training parameters required only approximately 30 s per sample (approximately 15 min in total). In addition, the presented results show that the suggested approach works for describing metabolic changes in different biofluids, indicating that this is a general approach for high-throughput predictive metabolite profiling, which could have important applications in areas such as plant functional genomics, drug toxicity, treatment efficacy and early disease diagnosis.

Animals↗
PubMed2005

Metabolic fingerprinting of rat urine by LC/MS Part 2. Data pretreatment methods for handling of complex data.

Metabolic fingerprinting of biofluids like urine is a useful technique for detecting differences between individuals. With this approach, it might be possible to classify samples according to their biological relevance. In Part 1 of this work a method for the comprehensive screening of metabolites was described, using two different liquid chromatography (LC) column set-ups and detection by electrospray ionization mass spectrometry (ESI-MS). Data pretreatment of the resulting data described in is needed to reduce the complexity of the data and to obtain useful metabolic fingerprints. Three different approaches, i.e., reduced dimensionality (RD), MarkerLynx, and MS Resolver, were compared for the extraction of information. The pretreated data were then subjected to multivariate data analysis by partial least squares discriminant analysis (PLS-DA) for classification. By combining two different chromatographic procedures and data analysis, the detection of metabolites was enhanced as well as the finding of metabolic fingerprints that govern classification. Additional potential biomarkers or xenobiotic metabolites were detected in the fraction containing highly polar compounds that are normally discarded when using reversed-phase liquid chromatography.

Animals↗
PubMed2005

Metabolic fingerprinting of rat urine by LC/MS Part 1. Analysis by hydrophilic interaction liquid chromatography-electrospray ionization mass spectrometry.

Complex biological samples, such as urine, contain a very large number of endogenous metabolites reflecting the metabolic state of an organism. Metabolite patterns can provide a comprehensive signature of the physiological state of an organism as well as insights into specific biochemical processes. Although the metabolites excreted in urine are commonly highly polar, the samples are generally analyzed using reversed-phase liquid chromatography mass spectrometry (RP-LC/MS). In Part 1 of this work, a method for detecting highly polar metabolites by hydrophilic interaction liquid chromatography-electrospray ionization mass spectrometry (HILIC/ESI-MS) is described as a complement to RP-LC/ESI-MS. In addition, in an accompanying paper (Part 2), different multivariate approaches to extracting information from the resulting complex data are described to enable metabolic fingerprints to be obtained. The coverage of the method for the screening of as many metabolites as possible is highly improved by analyzing the urine samples using both a C(18) column and a ZIC-HILIC column. The latter was found to be a good alternative when analyzing highly polar compounds, e.g., hydroxyproline and creatinine, to columns typically used for reversed-phase liquid chromatography.

Animals↗
PubMed2004

Differentiation of human hepatoma cells during confluence as revealed by gene expression profiling.

Certain human hepatocarcinoma cells undergo differentiation when grown at confluence. In order to understand the basis for this differentiation, we investigated the phenotypic changes occurring during confluent growth of the human hepatoma B16A2 cell line. The global gene expression profile of B16A2 cells grown during confluence for 5 weeks was investigated using microarrays containing complementary sequences corresponding to approximately 10,000 genes, and compared with profiles of adult human liver and HepG2 cells. The major part of gene products detected were shared by all three systems and the hepatoma cell lines expressed surprisingly high levels of liver-enriched transcription factors. During confluence of B16A2 cells, the majority of transcriptional changes monitored were directed towards the phenotype of adult human liver in vivo, although the changes accounted for less than 10% of those necessary to acquire a native hepatic phenotype. Several markers of liver differentiation and regeneration were changed in similar manner as observed in developing liver and during liver regeneration. In conclusion, the data indicate that differentiation in vitro of the B16A2 cell line during confluence partially resembles that of hepatic differentiation and regeneration in vivo, implying a partial normalization of a low differentiated phenotype.

Carcinoma, Hepatocellular↗
PubMed2003

Screening of biomarkers in rat urine using LC/electrospray ionization-MS and two-way data analysis.

Biofluids, like urine, form very complex matrixes containing a large number of potential biomarkers, that is, changes of endogenous metabolites in response to xenobiotic exposure. This paper describes a fast and sensitive method of screening biomarkers in rat urine. Biomarkers for phospholipidosis, induced by an antidepressant drug, were studied. Urine samples from rats exposed to citalopram were analyzed using solid-phase extraction (SPE) and liquid chromatography mass spectrometry (LC/MS) analysis detecting negative ions. A fast iterative method, called Gentle, was used for the automatic curve resolution, and metabolic fingerprints were obtained. After peak alignment principal component analysis (PCA) was performed for pattern recognition, PCA loadings were studied as a means of discovering potential biomarkers. In this study a number of potential biomarkers of phospholipidosis in rats are discussed. They are reported by their retention time and base peak, as their identification is not within the scope of the study. In addition to the fact that it was possible to differentiate control samples from dosed samples, the data were very easy to interpret, and signals from xenobiotic-related substances were easily removed without affecting the endogenous compounds. The proposed method is a complement or an alternative to NMR for metabolomic applications.

Animals↗
PubMed2003

Gene and protein expression profiling of human cerebral endothelial cells activated with tumor necrosis factor-alpha.

An increase in permeability of the blood-brain barrier is a critical event in the pathophysiological process of multiple sclerosis and other neurodegenerative diseases. Tumor necrosis factor alpha (TNFalpha) is known to play a crucial role in this process and is a powerful activator of endothelial cell inflammatory responses. Although many reports describe effects of TNFalpha activation in endothelial cells, the molecular mechanisms specific for activation of cerebral endothelial cells remains unclear. The objective of this study was to identify potential pharmaceutical targets for the treatment of multiple sclerosis using molecular profiling techniques. Gene expression measurements (Affymetrix Hu6800 oligonucleotide arrays) and proteomics (two-dimensional gel electrophoresis and mass spectrometry) were applied to analyze early alterations in human cerebral endothelial cells (HCEC) activated by TNFalpha. Human umbilical vein endothelial cells (HUVEC) were used as the reference system. The results presented show that HCEC and HUVEC respond similarly with respect to cell adhesion molecules, chemotaxis, apoptosis and oxidative stress molecules. However, nuclear factors NFkB1 and NFkB2, plasminogen activator inhibitor 1 and cofilin 1 are examples of cerebral specific responses. Our results indicate involvements of the urokinase plasminogen activator system and cytoskeletal rearrangements unique to TNFalpha activation of cerebral endothelial cells.

Antineoplastic Agents↗
PubMed2002

Identification of S-glutathionylated cellular proteins during oxidative stress and constitutive metabolism by affinity purification and proteomic analysis.

Redox modification of proteins is proposed to play a central role in regulating cellular function. However, high-throughput techniques for the analysis of the redox status of individual proteins in complex mixtures are lacking. The aim was thus to develop a suitable technique to rapidly identify proteins undergoing oxidation of critical thiols by S-glutathionylation. The method is based on the specific reduction of mixed disulfides by glutaredoxin, their reaction with N-ethylmaleimide-biotin, affinity purification of tagged proteins, and identification by proteomic analysis. The method unequivocally identified 43 mostly novel cellular protein substrates for S-glutathionylation. These include protein chaperones, cytoskeletal proteins, cell cycle regulators, and enzymes of intermediate metabolism. Comparisons of the patterns of S-glutathionylated proteins extracted from cells undergoing diamide-induced oxidative stress and during constitutive metabolism reveal both common protein substrates and substrates failing to undergo enhanced S-glutathionylation during oxidative stress. The ability to chemically tag, select, and identify S-glutathionylated proteins, particularly during constitutive metabolism, will greatly enhance efforts to establish posttranslational redox modification of cellular proteins as an important biochemical control mechanism in coordinating cellular function.

Cell Line↗
PubMed2002

Molecular profiling of hepatotoxicity induced by a aminoguanidine carboxylate in the rat: gene expression profiling.

The hepatotoxicity of the aminoguanidine carboxylate 2-[1-[hydrazino(imino)methyl]hydrazino]acetic acid was characterized using oligonucleotide micro arrays, with the goal to select compounds from the same class with lower toxicity potential. The approach included a 14-day repeated- and a single-dose study in the rat as well as in vitro studies. Common gene expression changes could be followed from in vivo to in vitro studies. Anyhow, comparing the in vivo and in vitro response of the compound on gene expression, significant discrepancies were detected. Many of the genes whose mRNA levels were increased/decreased in the livers of the animals treated with toxic doses of the compound, were expressed at higher/lower levels in control hepatocytes than in control liver. The expression of the majority of these genes was not affected by in vitro treatment. These data question the use of gene expression analysis as a marker for drug response in vitro and illustrate the need of a careful characterization of in vitro systems. The results presented show that array-based gene expression analysis can lead to a better understanding of the molecular basis of drug-induced liver injury and, potentially, be used in the selection process for compounds and in the design of safer drugs.

Acetates↗
PubMed2002

Identification of end points relevant to detection of potentially adverse drug reactions.

Expectations are high that the use of proteomics, gene arrays and metabonomics will improve risk assessment and enable prediction of toxicity early in drug development. These molecular profiling techniques may be used to classify compounds and to identify predictive markers that can be used to screen large numbers of chemicals. One of the challenges for the scientific community is to discriminate between changes in gene/protein expression and metabolic profiles reflecting physiological/adaptive responses, and changes related to pathology and toxicology. In these proceedings we provide a brief overview of the technologies with focus on proteomics and the possible applications to mechanistic and predictive toxicology. The discussion also includes strengths and limitations of molecular profiling technologies.

Animals↗
PubMed2002

S-glutathionylation of glyceraldehyde-3-phosphate dehydrogenase: role of thiol oxidation and catalysis by glutaredoxin.

The findings in this article illustrate the complexity residing in the regulation of reversible S-glutathionylation of proteins, such as GAPDH. This is clearly reflected in the design of suitable experimental approaches designed to cope with the interaction of several redox-dependent factors. Clear interactions are demonstrated between oxidative modification of GAPDH and its subsequent S-glutathionylation. Similarly, a redox interaction between GSSG and GAPDH with Grx as the catalyst is shown, suggesting that the Grx molecule may participate in catalytic S-glutathionylation in intact cells. Furthermore, Grx itself can readily undergo S-glutathionylation, indicating the potential for regulation of this catalyst of the reversible S-glutathionylation of other proteins. The methodologies detailed in this work may provide a good reference point for other attempts to elucidate the mechanism of reversible S-glutathionylation of purified proteins in a manner that more closely resembles the situation arising in intact cells during the generation of oxidative stress.

Autoradiography↗