Fluorescence Lifetime Microscopy (FLIM) for label-free metabolic imaging of living tissues

Please join us on Friday, February 21, at 13:00 CET, for the Virtual Pub. We welcome Chiara Stringari, Laboratory for Optics and Biosciences, CNRS, INSERM, École Polytechnique, France, for a talk entitled, "Fluorescence Lifetime Microscopy (FLIM) for label-free metabolic imaging of living tissues" Everyone is welcome!

Abstract:

Metabolism is increasingly recognized as a hallmark of various physiological and pathological processes, including stem cell differentiation, embryonic development, inflammation, immune response, neurodegenerative diseases and cancer. Understanding metabolic dynamics and mapping real-time changes in redox ratios at the cellular level within the native tissue microenvironment is essential for advancing both basic research and clinical applications. Here, I will discuss the potential of label-free optical techniques for non-invasive monitoring of cellular metabolism in living tissues, specifically by combining two-photon excitation fluorescence lifetime microscopy (2P-FLIM) with the use of endogenous fluorescent biomarkers naturally present in cells and tissues. This technique takes advantage of important and ubiquitous cellular cofactors such as nicotinamide adenine dinucleotide (NAD(P)H) and flavin adenine dinucleotide (FAD), which are crucial for cellular redox reactions and play a central role in energy production and metabolic regulation. The fluorescence lifetimes of NAD(P)H and FAD are highly sensitive to enzymatic binding, allowing 2P-FLIM imaging to probe cellular redox states and the complexity of different metabolic pathways, including glycolysis, oxidative phosphorylation, oxidative stress, and fatty acid metabolism. In this talk I will demonstrate the sensitivity of 2P-FLIM to these pathways through metabolic perturbations, compare the FLIM readout with standard metabolic assays and show how NAD(P)H and FAD are emerging as complementary biomarkers that reveal metabolic heterogeneity and pathway-specific changes. In order to efficiently probe both biomarkers simultaneously and to perform multiparametric metabolic imaging in dynamic biological systems, we have recently developed 2P-FLIM of NAD(P)H and FAD using wavelength mixing. Finally, I will show how 2P-FLIM imaging of NAD(P)H and FAD can be used for non-invasive longitudinal studies in vitro and in vivo with minimal phototoxicity in the context of various biomedical applications, such as stem cell differentiation, embryo development, and immune cell activation (e.g. T cell and microglia activation).