Development and applications of a novel in vivo dynamic isotope-tracing method to quantitively measure tricarboxylic acid cycle flux in immune cell sub-populations

Abstract

Cellular metabolism plays a key role in the regulation and modulation of cellular function. As it is, methods to quantitatively assess metabolism in vivo are limited, and most research probes into cellular metabolism in vitro. In this study, we identify a dynamic, isotope-tracing method to quantitively assess TCA flux in vivo. Furthermore, we combine applications of this method with immunomagnetic cell isolation in order to characterize immune-cell sub-populations. Thus, a throughput for quantitatively assessing experimental systems in vivo subjected to environmental perturbation was developed. Here, three experimental systems are explored. The first system characterized B cells in the spleen and assessed the impact of vaccination on TCA flux. The second system attempted to quantify metabolism in tumor-infiltrating T cells relative to their latent counterparts in the spleen. The final system was an overall assessment of the immune cell sub-populations present in the bone marrow, with a specific focus on CD11b+ macrophages. Overall, with some minor qualifications, the quantitative assessment of metabolism in the immune cell sub-populations was successful, and our hope is that further studies are warranted in immunology and medicine that employ KFP and quantitative fluxomics to better characterize cellular and organism-level physiology.