Rejection of transplanted hearts in patients evaluated by the component analysis of multi‐wavelength NAD(P)H fluorescence lifetime spectroscopy

Abstract

Rejection of transplanted hearts remains one of the principal reasons for death of paediatric patients, but an appropriate diagnostic tool for the mild rejection in early stages is still missing. Tissue autofluorescence (AF) is one of the most versatile non-invasive tools for mapping the metabolic state in living tissues. Increasing interest in the imaging and diagnosis of living cells and tissues based on their intrinsic fluorescence rather than fluorescence labelling is closely connected to the latest developments in high-performance spectroscopy and microscopy techniques. In this contribution, we investigate individual components in spectrally- and time-resolved NAD(P)H fluorescence, revealed by linear unmixing, responsible for increased fluorescence in patients presenting mild rejection of transplanted hearts. Application of such approach has the potential to improve the diagnostics of the cardiac transplant rejection by helping currently used histological analysis.

Effect of ouabain on metabolic oxidative state in living cardiomyocytes evaluated by time-resolved spectroscopy of endogenous NAD(P)H fluorescence

“…1(a)], as described previously. 7 A linear unmixing approach was used to separate individual NAD(P)H components; component 1 with fluorescence lifetime of around 1.8 ns corresponded to NAD(P)H in a more viscous environment, and this value is in agreement with published values of "bound" NAD(P)H in mitochondria 20,21 [see resolved fluorescence decays in Fig. 1(b)].…”

“…Data were collected from at least three different animals per experimental group, and compared by one-way analysis of variance (ANOVA); p < 0.05 was considered statistically significant. Individual components of NAD(P)H fluorescence were estimated using principal component analysis (PCA), 7 while the linear unmixing approach was applied to separate individual components in the recorded data, as previously reported 13,14 Component amplitude for each resolved fluorescence component was calculated from the fluorescence decay kinetics using a monoexponential fitting procedure at the spectral channel with maximum intensity (450 nm). Fluorescence lifetime for each component was estimated in ns together with the component amplitude.…”

“…7,12 Picosecond laser diode BDL-375 (Becker&Hickl, Boston Electronics, USA) with emission at 375 nm was deployed at the 20 MHz repetition rate as an excitation source. All experiments were performed at room temperature.…”

“…7 This technique allows not only to determine changes in the fluorescence intensity, but also to separate individual NAD(P)H fluorescence components accordingly to their spectral characteristics and fluorescence lifetimes. 8 Consequently, the applied approach allows us to evaluate changes in the cell's metabolic oxidative mitochondrial redox state which correlate with NADH∕NAD þ redox potential.…”

Use of multiphoton tomography and fluorescence lifetime imaging to investigate skin pigmentationin vivo

“…3 in Ref. 66), which is close to the equivalent two-photon excitation wavelength we used (800 nm). From this result we inferred that we could expect differences in melanin concentration in skin in vivo to be measured based on differences in fluorescence intensity.…”