CC BY
12B-O-5
Multiphoton microscopy and FLIM metabolic imaging of the hepatocytes during liver regeneration
S.A. Rodimova12, D.S. Kuznetsova12, N.V. Bobrov13, A.A. Gulin4 5, V.V. Elagin1, M.M. Karabut1, V.I. Shcheslavskiy16, V.E. Zagainov13, E.V. Zagaynova12.
1Privolzhsky research medical university, Institute of Experimental Oncology and Biomedical Technologies, 10/1
Minin and Pozharsky sq., Nizhny Novgorod, Russia 2N.I. Lobachevsky Nizhny Novgorod National Research State University, 23 Gagarina ave., Nizhny Novgorod,
Russia
3The Volga District Medical Centre of Federal Medical and Biological Agency, 14 Ilinskaya, Nizhny Novgorod,
Russia
4Russian Academy of Sciences, N.N. Semenov Federal Research Center for Chemical Physics, 4 Kosygina,
Moscow, Russia
5Lomonosov Moscow State University, Department of Chemistry, 1-3 Leninskiye Gory, Moscow, Russia 6Becker&Hickl GmbH, Nunsdorfer Ring 7-9, Berlin, Germany
Surgical liver resection is still the most commonly used method for the treatment of hepatic tumors [1]. However, with a decrease in the regenerative potential of the liver, the risk of developing liver failure during resection increases [2]. Standard clinical methods are not informative enough do not predict the function of the remaining liver [3]. Using modern label-free methods of multiphoton microscopy in combination with FLIM (fluorescence lifetime imaging) and SHG (second harmonic generation) it is possible to assess the metabolic state of hepatocytes during regeneration [4]. In addition, Time-of-Flight Secondary Ion Mass Spectrometry (TOF-SIMS) provides information on the lipid and amino acid composition of liver tissue [8].
The experiments were performed on Wistar rats. Two models were chosen - 70% parietal hepatectomy (PH) and 30% PH. Metabolic imaging was performed on 3rd and 7th days after surgery. We presented a separate analysis of NADH and NADPH to assess the contribution of various metabolic pathways as well as synthetic activity in hepatocytes. TOF-SIMS provides data on the lipid and amino acid composition of liver tissue.
As a result, an increase in the contribution of oxidative phosphorylation and biosynthetic processes in hepatocytes, which compensate for the energy consumption of hepatocytes during regeneration, has been shown. Using TOF-SIMS, we observed simultaneous increase in the signals representing triglycerides and cholesterol in the hepatocytes, combined with a decrease in the signals of amino acids peaks.
Thus, the approach used in this work allowed us also to identify the functional state of the liver tissue before and after resection and to determine the range of cellular metabolic heterogeneity for different stages of regeneration The obtained data will be useful in determining the criteria for intraoperative express assessment of the regenerative potential of the liver after surgery.
The work was supported by the Grant from the Russian Science Foundation №19-15-00263 (metabolic imaging, analysis of FLIM data of liver tissue), and the Grant of the President of the Russian Federation MK-1649.2021.3 (morphological and morphometric study).
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