Spectroscopy and Imaging

In this study, thin mouse kidney sections from healthy mice and those infected, leading to acute and chronic sepsis, were examined with two-photon excited fluorescence lifetime imaging (2P-FLIM) using the endogenous fluorescent coenzymes nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD). The results presented show that this approach is a powerful tool for investigating cell metabolism in thin tissue sections. An adapted measurement routine was established for these samples by performing a spectral scan, identifying a combination of two excitation wavelengths and two detection ranges suitable for detailed scan images of NADH and FAD. Selected positions in thin slices of the renal cortex of nine mice (three healthy, three with chronic sepsis, and three with acute sepsis) were studied using 2P-FLIM. In addition, overview images were obtained using two-photon excited fluorescence (2PEF) intensity. This study shows that healthy kidney slices differ considerably from those with acute sepsis with regard to their fluorescence lifetime signatures. The latter shows a difference in metabolism between the inner and outer cortex, indicating that outer cortical tubular cells switch their metabolism from oxidative phosphorylation to glycolysis in kidneys from mice with acute sepsis and back in later stages, as seen for mice with chronic infections. These findings suggest that 2P-FLIM could serve as a powerful tool for early-stage sepsis diagnosis and monitoring metabolic recovery during treatment.