In search of secreted protein biomarkers for the anti-inflammatory effect of β2-adrenergic receptor agonists: Application of DIGE technology in combination with multivariate and univariate data analysis tools
article
Two-dimensional difference gel electrophoresis (DIGE) in combination with univariate (Student's t-test) and multivariate data analysis, principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA) were used to study the anti-inflammatory effects of the β2-adrenergic receptor (β2-AR) agonist zilpaterol. U937 macrophages were exposed to the endotoxin lipopolysaccharide (LPS) to induce an inflammatory reaction, which was inhibited by the addition of zilpaterol (LZ). This inhibition was counteracted by addition of the β2-AR antagonist propranolol (LZP). The extracellular proteome of the U937 cells induced by the three treatments were examined by DIGE. PCA was used as an explorative tool to investigate the clustering of the proteome dataset. Using this tool, the dataset obtained from cells treated with LPS and LZP were separated from those obtained from LZ treated cells. PLS-DA, a multivariate data analysis tool that also takes correlations between protein spots and class assignment into account, correctly classified the different extracellular proteomes and showed that many proteins were differentially expressed between the proteome of inflamed cells (LPS and LZP) and cells in which the inflammatory response was inhibited (LZ). The Student's t-test revealed 8 potential protein biomarkers, each of which was expressed at a similar level in the LPS and LZP treated cells, but differently expressed in the LZ treated cells. Two of the identified proteins, macrophage inflammatory protein-1beta (MIP-1β) and macrophage inflammatory protein-1 alpha (MIP-1α) are known secreted proteins. The inhibition of MIP-1β by zilpaterol and the involvement of the β2-AR and cAMP were confirmed using a specific immunoassay. © 2005 American Chemical Society.
Topics
Analytical researchβ2-adrenergic receptorDifference gel electrophoresisMacrophage inflammatory protein-1βMultivariate data analysisPartial least squares discriminant analysisPrincipal component analysisZilpaterolAdenylate kinaseBeta 2 adrenergic receptorBeta 2 adrenergic receptor blocking agentBeta 2 adrenergic receptor stimulating agentBiological markerButyryl cyclic AMPClenbuterolEndotoxinEscherichia coli lipopolysaccharideFormoterolForskolinHeat shock proteinMacrophage inflammatory protein 1alphaMacrophage inflammatory protein 1betaPhosphoglycerate mutasePropranololProstaglandin E2SalbutamolUnclassified drugZilpaterolAdrenergic systemAnalytic methodAntiinflammatory activityControlled studyData analysisDiscriminant analysisHuman cellLiquid chromatographyMultivariate analysisNucleotide sequencePrincipal component analysisProtein expressionProtein secretionProteomicsStatistical analysisTwo dimensional difference gel electrophoresisTwo dimensional gel electrophoresisAdrenergic AgonistsAdrenergic beta-AntagonistsAnti-Inflammatory AgentsBiological MarkersCluster AnalysisDown-RegulationElectrophoresis, Gel, Two-DimensionalEnzyme-Linked Immunosorbent AssayHumansImmunoassayInflammationLipopolysaccharidesMacrophage Inflammatory Protein-1MacrophagesMass SpectrometryMonocytesMultivariate AnalysisPrincipal Component AnalysisPropranololProteomeProteomicsReceptors, Adrenergic, beta-2StatisticsTrimethylsilyl CompoundsU937 CellsUp-RegulationEscherichia coli
TNO Identifier
238774
ISSN
15353893
Source
Journal of Proteome Research, 4(6), pp. 2015-2023.
Pages
2015-2023
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