MSACL 2019 EU Abstract

Metabotyping Burn Injury Using UPLC-MS Coupled with Microdialysis

Dominic Friston, Helen Laycock, Istvan Nagy, Elizabeth J. Want
Imperial College, London, UK

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Elizabeth Want (Presenter) Imperial College London

Presenter Bio: Dr Elizabeth Want is a Senior Lecturer in Molecular Spectroscopy in the Department of Surgery and Cancer. She joined Imperial College in 2006 after working as a postdoc at the Scripps Research Institute, CA. She is the Director of the Imperial International Phenome Training Centre. Dr Want has >20 yrs of experience in mass spectrometry and chromatographic techniques and has spent the past 18 years working in the field of MS-based metabolite profiling. Her research at Imperial College has involved the development, optimisation and application of UPLC-MS methodologies for the analysis of biological samples, largely in the context of metabolic phenotyping. She is applying these methods to biomedical research areas including toxicology, neonatal disease and development, cardiovascular and neurological diseases, as well as burn injury.

Relevant Financial Disclosures (within past 24 months) Committee/Board/Advisory Board MSACL EU Chair Discussing Products? NO, presenter will NOT present educational content related to products/services of the ineligible company(ies) listed above. Conflict mitigation NOT REQUIRED.

INTRODUCTION: Burn injury can be a devastating trauma, affecting millions of people worldwide, with long-term personal and social implications for patients. Burns can result in life-limiting chronic pain, often refractory to treatment. Mechanisms behind burn injury are poorly understood and there has been little research into the molecular basis of burns and subsequent pain. It is important to explore the local signalling environment, but studies are often destructive in nature and preclude the collection of longitudinal temporal data. Microdialysis is a sampling method allowing the in vivo collection of solutes primarily from the extracellular interstitium and is ideal for the study of burn injury. Metabolomics offers an unbiased approach to the elucidation of metabolites involved in pathological events. Coupled with mass spectrometry, it provides a sensitive platform for the detection of metabolic changes due to burn injury.
OBJECTIVES: Elucidation of altered metabolites due to burn injury - to enhance understanding of the pathological processes occurring and guide future therapeutic strategies.
METHODS: Subcutaneous microdialysis in a burn model was coupled with ultra performance liquid chromatography – mass spectrometry (UPLC-MS) analysis. Microdialysis was conducted for 0.5 hrs pre-burn and 3 hours post-burn, collected in half hour fractions for time series analysis. An Acquity UPLC system with a HSS T3 column was connected to a Synapt G2-S Q-ToF. Data were processed using XCMS software. Principal components analysis (PCA) was applied for unsupervised multivariate comparison of burn and control sites. Partial least squares discriminant-analysis (PLS-DA) was applied to rank metabolite features contributing to these differences. Model robustness was affirmed using CV-ANOVA. MS/MS studies for structural elucidation were performed using the same UPLC-MS system.
RESULTS: Hundreds of polar analytes and lipids were profiled from microdialysate samples over a reversed phase run of 12 minutes, yielding a high number of burn-altered metabolites. PCA scores plots showed tight quality control sample grouping, indicating acquisition stability. Clear metabolic differences were observed between microdialysates collected from burned tissue and the control sites. Two important molecules elevated in burn injury were structurally elucidated; niacinamide and uric acid. These two compounds are potentially involved in the pathology of burn injury.
CONCLUSION: This study demonstrates the application of high throughput metabolomic profiling to samples collected both continuously and specifically from the burn site. Further understanding the metabolic changes induced by burn injury will help to guide therapeutic intervention in the future. This approach is equally applicable to the analysis of other tissues and pathological conditions, so may further improve our understanding of the metabolic changes underlying a wide variety of pathological processes.