Biomarker Discovery by Imaging Mass Spectrometry

Abstract

Adverse drug effects are often associated with pathological changes in tissue. An accurate depiction of the undesired affected area, possibly supported by mechanistic data, is important to classify the effects with regard to relevance for human patients. MALDI imaging MS represents a new analytical tool to directly provide the spatial distribution and the relative abundance of proteins in tissue. Here we evaluate this technique to investigate potential toxicity biomarkers in kidneys of rats that were administered gentamicin, a well known nephrotoxicant. Differential analysis of the mass spectrum profiles revealed a spectral feature at 12,959 Da that strongly correlates with histopathology alterations of the kidney. We unambiguously identified this spectral feature as transthyretin (Ser 28 -Gln 146 ) using an innovative combination of tissue microextraction and fractionation by reverse-phase liquid chromatography followed by a top-down tandem mass spectrometric approach. Our findings clearly demonstrate the emerging role of imaging MS in the discovery of toxicity biomarkers and in obtaining mechanistic insights concerning toxicity mechanisms. Proteomic strategies have been exploited to generate differential protein expression that can be correlated with environmental disease or toxicant exposure. In particular, improvements in two-dimensional gel electrophoresis and liquid chromatography coupled to mass spectrometry and the use of other emerging techniques have demonstrated the usefulness of a proteomic approach in toxicology (1-3). The early detection of disease markers and the rapid screening of experimental compounds either for toxicity or protective efficacy are of particular interest in this field (4). Imaging mass spectrometry (IMS)1 is a technique for the direct analysis of peptides and proteins from thin tissue sections using conventional MALDI-TOF mass spectrometers while preserving the abundance and spatial distribution of each analyte (5). The usefulness of this technique has been demonstrated in experimental models and in clinical settings (6, 7) as in the accurate and sensitive classification of nonsmall cell lung cancer in tumor biopsies (8).In this study we further demonstrated the potential applications of an IMS strategy in the study of a well known nephrotoxicant, the aminoglycoside antibiotic gentamicin (9), in the rat kidney. Aminoglycoside antibiotics are widely used, but nephrotoxicity (10) is a clear risk and occurs in 10 -20% (11) of treated patients. Gentamicin-induced nephrotoxicity is seldom fatal and is usually reversible but often results in long hospital stays. Thus, there is a great interest in finding potential markers for the toxicity event and to further elucidate the toxicity mechanism.We first established our IMS strategy by determining the differential protein expression within the main areas of the rat kidney (cortex, medulla, and papilla) to ensure that we could accurately differentiate the main substructures of this organ. We then investigated whether the kidney lesions sec...

SELDI‐TOF as a method for biomarker discovery in the urine of aristolochic‐acid‐treated mice

“…As shown in Table 1, in the urine of mice treated with AAI, several plasma proteins were identified, among them are serum amyloid and transthyretin. These two proteins are also candidate biomarker for renal injury and cancer, following poisoning with nephrotoxins [24,25]. Inter-alpha-trypsin inhibitor heavy chain 4, which was also identified, is a plasma glycoprotein that acts as an acute-phase protein in several species.…”

Diabetic nephropathy induces alterations in the glomerular and tubule lipid profiles

“…Interestingly, unlike in all other mammals, ganglioside technology enables molecular profi ling of specifi c areas of the tissues while preserving tissue morphology. This technology has been instrumental in understanding molecular changes on the border between normal and cancer tissues ( 42 ), classifying tumor types ( 28,43 ), determining tissue drug distributions ( 44,45 ), and identifying candidate markers of disease ( 20,46 ). As demonstrated in this study, it is now possible to analyze the molecular composition and molecular modifi cations in distinct renal tissue regions, and even within smaller structures such as glomeruli and tubules.…”

Development of operational protocols for the analysis of primary and secondary fingermark lifts by MALDI-MS imaging

“…There are indeed different ways to treat such samples: (a) recovering the fingermark residue remaining after the primary lift with no prior attempt to re-enhance the residue, followed by matrix spray-coating for direct MALDI MS-Imaging/Profiling analysis; (b) subjecting the fingermark residue remaining after the primary lift to a secondary lift following reenhancement using the same FET initially employed, with this lift being subsequently subjected to matrix spray-coating for direct MALDI-MS Imaging/Profiling analysis; (c) prior to MALDI-MS Imaging/Profiling analysis, subjecting the fingermark residue remaining after the primary lift to a secondary lift following re-enhancement by the Dry-Wet method 12,13 with the sample being subsequently either (c-1) spray-coated using a suitable solvent in which both the matrix and analytes can dissolve 12 or (c-2) matrix spray-coated with the matrix re-enhancement using the dry powder as a seeding layer for the co-crystallisation formation 14 . These four methods were trialled, compared and contrasted to provide preliminary recommendations on how to best treat secondary lifts.…”