Metabolic Tracer Analysis Core (MTAC) Facility

Metabolic Tracer Analysis Core (MTAC) FacilityMetabolic Tracer Analysis Core (MTAC) focuses on providing investigations to determine metabolic phenotype in vitro and in vivo using high resolution, targeted methodologies.

One particular area of expertise is in the use of stable isotope-enriched nutrients to trace their use by cells, tissues or whole organisms in order to determine activity of specific aspects of the metabolic network. This type of directed analysis of metabolism is complementary to the unbiased metabolomics approaches as performed by the Phenome Centre Birmingham, and alongside them create a complementary suite of technologies available to study cellular metabolism.

Additional capabilities within MTAC include two Oroboros instruments for high resolution respiratory measurements, and the in vivo phenotyping unit to investigate metabolism in a physiological setting in animal models. Through the use of these multiple means of assessing metabolism, we can help interrogate systems in which metabolism changes in order to identify novel approaches to treat disease or improve health-span.

Stable Isotope Analysis

Metabolomics is a powerful approach provide an unbiased view of the cellular metabolic network. However, when a more directed analysis of metabolic pathway activity is required, an alternative method is often more suitable.

Nutrients such as sugars, amino acids and lipid can be followed through a metabolic system when enriched with a stable isotope, such as 13C, 15N or 2H. In this way, the heavy (non-radioactive) isotope can be followed as it is incorporated into different metabolites, allowing us to determine the metabolic pathway used to produce these metabolites as well as their fate.

Our current methods can define the way nutrients are processed through central carbon metabolism, often focusing on mitochondrial phenotype. Indeed, through this approach, specific metabolic enzymes can be identified that are dysfunctional in specific diseases, or that are required for normal tissue function, providing important rationale to further develop them into therapeutic targets.

Stable isotopes can be safely administered to animals and patients, allowing true in vivo analyses of metabolism of complex multi-organ systems.

Mass Spectrometry & NMR spectroscopy

In MTAC, quantitation of relative and absolute pathway activity is facilitated by detection of the enrichment of stable isotopes into cellular metabolites using a combination of mass spectrometry, pseudo-2D and 2D NMR spectroscopic methods. A specific strength of our method is the use of high resolution NMR spectroscopy datasets (derived using narrow-bore cryoprobes and cutting-edge acquisition methodologies) alongside the mass spectrometry data to produce a single read-out of stable isotope incorporation with the positional information often required to define specific enzyme activity. This is achieved by using an in-house software programme – Metabolab, which analyse both datasets simultaneously.

The facility currently relies on an Agilent 7890B GC coupled to 5973A selective MSD with XTR ion source for EI ionisation, as well as a Bruker 600 MHz NMR instrument with 1.7 mm cryoprobe.

In vivo phenotyping centre

Although studying cells and tissues in isolation provides valuable information on the basic metabolic network, whole organism studies are required to understand more complex metabolic interactions both within organs and between them.

The in vivo phenotyping part of MTAC provides the means to investigate the mouse metabolism using PhenoMaster systems capable of monitoring multiple parameters simultaneously. The closed systems allows the assessment of total daily energy expenditure, food and water intake with temporal resolution in order to provide data on day-night variation. The effects of physical activity on energy expenditure can also be measured in a variety of ways using voluntarily exercise wheels, with acute or training protocols possible using treadmills, as well as a calorimetric treadmill.

Meet the Team

Dr. Daniel Tennant is a Reader in Metabolic Biochemistry in the Institute of Metabolism and Systems Research (IMSR), situated within the IBR tower. He uses stable isotope tracers to investigate changes in the use of nutrients in conditions in oxygen limiting conditions or in the presence of genetic mutations (Link to IMSR profile).

Dr Christian Ludwig is lecturer in Metabolic Biophysics in the Institute of Metabolism and Systems Research (IMSR). He expertise in development of novel NMR protocols along with development of software tools to allow mapping and quantitation of system metabolites linked to genetic and proteomic alteration. He led the development of software that combines NMR and MS data that facilitates accurate identification of isotopomer distributions from key metabolites (IMSR profile)

Prof Gareth Lavery is Professor of Molecular Metabolism in the IMSR. He instigated the development of the in vivo phenotyping centre that studies the impact of hormones, nutrients and exercise in mouse models on metabolic pathways in health and disease (IMSR profile)

Other team members

Dr Jennie Roberts runs MTAC, optimising analytical methods and communicating with collaborators in order to facilitate their research through the analysis of stable isotope incorporation into cellular metabolites. 

How can we help?

There are a number of ways in which we can assist with your investigations – please get in touch with us (mtac@contacts.bham.ac.uk) and we can discuss what could best facilitate your research.

We are happy to provide support at the level of experimental design, sample preparation and analysis.

Publications

Katarina Kluckova et al. Succinate dehydrogenase-deficiency in a chromaffin cell model retains metabolic fitness through the maintenance of mitochondrial NADH oxidoreductase function. FASEB J. (2020). 34(1), 303-315

Lucy A. Oakey et al. Metabolic tracing reveals novel adaptations to skeletal muscle cell energy production pathways in response to NAD+ depletion. Wellcome Open Research, 2019. 3:147

TB Smith et al. High-speed tracer analysis of metabolism (HS-TrAM). Wellcome Open Research, 2018. 3

Kate E.R. Hollinshead et al. Oncogenic IDH1 mutations promote enhanced proline synthesis through PYCR1 to support the maintenance of mitochondrial redox homeostasis. Cell Reports, 2018. 22, 3107-3114

Mei Chong et al. Combined NMR and MS analysis for tracer based metabolic flux experiments. Angewandte Chemie, 2017. 15, 4204-4210

Jay Nath et al. Metabolic differences between cold stored and machine perfused porcine kidneys: A 1 H NMR based study. Cryobiology, 2017. (47), 115-120

Jay Nath et al. 13C glucose labelling studies using 2D NMR is a useful tool for determining ex vivo whole organ glycolytic metabolism during hypothermic machine perfusion of kidneys. Transplantation Res, 2016. 5 (1):7

Charlotte Lepoutre-Lussey et al. Loss of succinate dehydrogenase activity results in dependency on pyruvate carboxylation for cellular anabolism. Nature Communications, 2015. 2;6:8784.

C Ludwig et al. Alterations in bone marrow metabolism are an early and consistent feature during the development of MGUS and multiple myeloma. Blood Cancer Journal, 2015. 5(10); e359