Serum and Hepatic Autofluorescence as a Real-Time Diagnostic Tool for Early Cholestasis Assessment.

While it is well established that various factors can impair the production and flow of bile and lead to cholestatic disease in hepatic and extrahepatic sites, an enhanced assessment of the biomarkers of the underlying pathophysiological mechanisms is still needed to improve early diagnosis and therapeutic strategies. Hence, we investigated fluorescing endogenous biomolecules as possible intrinsic biomarkers of molecular and cellular changes in cholestasis. Spectroscopic autofluorescence (AF) analysis was performed using a fiber optic probe (366 nm excitation), under living conditions and in serum, on the livers of male Wistar rats submitted to bile duct ligation (BDL, 24, 48, and 72 h). Biomarkers of liver injury were assayed biochemically. In the serum, AF analysis distinctly detected increased bilirubin at 24 h BDL. A continuous, significant increase in red-fluorescing porphyrin derivatives indicated the subversion of heme metabolism, consistent with an almost twofold increase in the serum iron at 72 h BDL. In the liver, changes in the AF of NAD(P)H and flavins, as well as lipopigments, indicated the impairment of mitochondrial functionality, oxidative stress, and the accumulation of oxidative products. A serum/hepatic AF profile can be thus proposed as a supportive diagnostic tool for the in situ, real-time study of bio-metabolic alterations in bile duct ligation (BDL) in experimental hepatology, with the potential to eventually translate to clinical diagnosis.

Autofluorescence spectroscopy of rat liver during experimental transplantation procedure. An approach for hepatic metabolism assessment.

Ischemia-reperfusion injury, a major cause of organ metabolic alterations and consequent dysfunction in liver transplantation, could be overcome by optimizing organ preservation procedures. The potential of autofluorescence analysis was investigated with the aim to define parameters suitable for in vivo monitoring tissue functionality. Spectrofluorometric analysis was performed on explanted rat livers during cold storage, under standard (4 degrees C University of Wisconsin medium for 20 h) and purposely damaging (4 degrees C Eurocollins medium for 20, 43 and 72 h) preservation conditions, and reperfusion (rewarming-reoxygenation). For both preservation conditions, cold hypoxia caused a signal amplitude increase, mainly attributable to NAD(P)H, and a spectral shape modification, ascribable to changes in the relative contributions of NAD(P)H and flavins, as a result of the tissue reduced state enhancement. Upon rewarming-reoxygenation the autofluorescence signal decreased with a rate depending on the preservation conditions. The time constant changed according to the extent of the liver functionality impairment, as assessed by conventional biochemical and histochemical analyses, thus providing a parameter exploitable for an in situ, real time monitoring of the efficacy of experimental preservation procedures.