FLUORESCENT IMAGES OF MITOCHONDRIAL REDOX STATES ININ SITUMOUSE HYPOXIC ISCHEMIC INTESTINES

Abstract

We have imaged mitochondrial oxidation–reduction states by taking a ratio of mitochondrial fluorophores: NADH (reduced nicotinamide adenine dinucleotide) to Fp (oxidized flavoprotein). Although NADH has been investigated for tissue metabolic state in cancer and in oxygen deprived tissues, it alone is not an adequate measure of mitochondrial metabolic state since the NADH signal is altered by dependence on the number of mitochondria and by blood absorption. The redox ratio, NADH/(Fp + NADH), gives a more accurate measure of steady-state tissue metabolism since it is less dependent on mitochondrial number and it compensates effectively for hemodynamic changes. This ratio provides important diagnostic information in living tissues. In this study, the emitted fluorescence of mouse colon in situ is passed through an emission filter wheel and imaged on a CCD camera. Redox ratio images of the healthy and hypoxic mouse intestines clearly showed significant differences. Furthermore, the corrected redox ratio indicated an increase from an average value of 0.51 ± 0.10 in the healthy state to 0.92 ± 0.03 in dead tissue due to severe ischemia (N = 5). We show that the CCD imaging system is capable of displaying the metabolic differences in normal and ischemic tissues as well as quantifying the redox ratio in vivo as a marker of these changes.