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Presently, there is a need in the developing new approaches to stimulate liver regeneration, which would make its recovery more effective after resection. Application of nanoparticles, loaded with small bioactive molecules, with their targeted delivery into the liver is a promising approach. The aim of the investigation is to study the interaction of nanoparticles with various types of hepatic cells on the models of liver slices and primary hepatic cell cultures using the methods of multiphoton microscopy with fluorescence lifetime imaging., Materials and Methods: Nanoparticles have been synthetized from polylactide (PLA), gold (Au), and silicon (SiO 2 ), and characterized using scanning and transmission electron microscopy. These types of particles were labeled with a fluorescent Cy5 dye for their visualization. Liver slices and a primary hepatocyte culture were used as models for biological testing of nanoparticles. Biodistribution of the nanoparticles in the tissue and cells, their cytotoxicity, and the effect on the cell metabolism were assessed using optical bioimaging methods., Results: The silicon nanoparticles are accumulated mainly by macrophages, which generate reactive oxygen species in a large amount and impair the native metabolic state of hepatocytes. The gold nanoparticles accumulate in all types of the liver cells but possess a marked toxic effect, which is indicated by the appearance of necrotic and apoptotic cells and a sharp change in the hepatocyte metabolic state. The polylactide nanoparticles accumulate most effectively in the liver cells, preferably in hepatocytes, do not change their native metabolic state, making this type of nanoparticles most promising for creating the bioactive molecule delivery systems to stimulate liver regeneration., Competing Interests: Conflicts of interest. The authors have no conflicts of interest to declare.

Liver pathologies remain one of the leading causes of mortality worldwide. Despite a high prevalence of liver diseases, the possibilities of diagnosing, prognosing, and treating non-alcoholic and alcoholic liver diseases still have a number of limitations and require the development of new methods and approaches. In laboratory studies, various models are used to reconstitute the pathological conditions of the liver, including cell cultures, spheroids, organoids, microfluidic systems, tissue slices. We reviewed the most commonly used in vivo , in vitro , and ex vivo models for studying non-alcoholic fatty liver disease and alcoholic liver disease, toxic liver injury, and fibrosis, described their advantages, limitations, and prospects for use. Great emphasis was placed on the mechanisms of development of pathological conditions in each model, as well as the assessment of the possibility of reconstructing various key aspects of pathogenesis for all these pathologies. There is currently no consensus on the choice of the most adequate model for studying liver pathology. The choice of a certain effective research model is determined by the specific purpose and objectives of the experiment., Competing Interests: The authors have no conflicts of interest to declare.

Considering the clinical limitations of individual approaches against metastatic lung cancer, the use of combined therapy can potentially improve the therapeutic effect of treatment. However, determination of the appropriate strategy of combined treatment can be challenging. In this study, combined chemo- and radionuclide therapy has been realized using radionuclide carriers ( 177 Lu-labeled core-shell particles, 177 Lu-MPs) and chemotherapeutic drug (cisplatin, CDDP) for treatment of lung metastatic cancer. The developed core-shell particles can be effectively loaded with 177 Lu therapeutic radionuclide and exhibit good radiochemical stability for a prolonged period of time. In vivo biodistribution experiments have demonstrated the accumulation of the developed carriers predominantly in lungs. Direct radiometry analysis did not reveal an increased absorbance of radiation by healthy organs. It has been shown that the radionuclide therapy with 177 Lu-MPs in mono-regime is able to inhibit the number of metastatic nodules (untreated mice = 120 ± 12 versus 177 Lu-MPs = 50 ± 7). The combination of chemo- and radionuclide therapy when using 177 Lu-MPs and CDDP further enhanced the therapeutic efficiency of tumor treatment compared to the single therapy ( 177 Lu-MPs = 50 ± 7 and CDDP = 65 ± 10 versus 177 Lu-MPs + CDDP = 37 ± 5). Thus, this work is a systematic research on the applicability of the combination of chemo- and radionuclide therapy to treat metastatic lung cancer., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The aim of the investigation was to study the possibility of revealing the heterogeneity of normal liver hepatocytes in terms of metabolic status using the modern methods of multiphoton microscopy and mass spectrometry., Materials and Methods: Heterogeneity of hepatocytes in terms of total metabolic activity was assessed using multiphoton microscopy based on the autofluorescence intensity of intracellular cofactors NAD(P)H and FAD. Hepatocyte heterogeneity in terms of intensity of intracellular metabolic processes was determined using the fluorescence lifetime imaging (FLIM) method based on the data about fluorescence lifetime contributions of various forms of NAD(P)H. The method of time-of-flight secondary ion mass spectrometry (TоF-SIMS) was used to study the lipid and amino acid composition of hepatocytes., Results: It has been revealed using multiphoton microscopy that hepatocytes are heterogeneous in terms of general metabolic activity. Using FLIM, it was established that the heterogeneity degree was high in terms of intensity of oxidative phosphorylation, glycolysis, and synthetic processes (lipogenesis, nucleic acid synthesis, and the pentose phosphate pathway). The TоF-SIMS method revealed the presence of hepatocyte heterogeneity in terms of amino acid and lipid composition, which points to various intensities of synthetic processes in individual hepatocytes. Moreover, differences in the content of PO 3 ions were revealed. The results of ToF-SIMS study correlate with the data obtained by multiphoton microscopy and FLIM, confirming the revealed heterogeneity of hepatocytes in terms of general metabolic activity and intensity of intercellular metabolic processes., Conclusion: The latest methods of fluorescence bioimaging and mass spectrometry proved to be effective in revealing hepatocyte heterogeneity in terms of metabolic status. The presence of heterogeneity should be taken into account in studying the liver tissue under various conditions with the application of fluorescence bioimaging methods., Competing Interests: Conflict of interest. The authors have no conflict of interest to disclose.

Membrane fluidity plays an important role in many cell functions such as cell adhesion, and migration. In stem cell lines membrane fluidity may play a role in differentiation. Here we report the use of viscosity-sensitive fluorophores based on a BODIPY core, termed "molecular rotors", in combination with Fluorescence Lifetime Imaging Microscopy, for monitoring of plasma membrane viscosity changes in mesenchymal stem cells (MSCs) during osteogenic and chondrogenic differentiation. In order to correlate the viscosity values with membrane lipid composition, the detailed analysis of the corresponding membrane lipid composition of differentiated cells was performed by time-of-flight secondary ion mass spectrometry. Our results directly demonstrate for the first time that differentiation of MSCs results in distinct membrane viscosities, that reflect the change in lipidome of the cells following differentiation.

Induced pluripotent stem cells (iPSC) are a promising tool for personalized cell therapy, in particular, in the field of dermatology. Metabolic plasticity of iPSC are not completely understood due to the fact that iPSC have a mixed mitochondrial phenotype, which still resembles that of somatic cells. In this study we investigated the metabolic changes in iPSC undergoing differentiation in two directions, dermal and epidermal, using two-photon fluorescence microscopy combined with FLIM. Directed differentiation of iPSC into dermal fibroblasts and keratinocyte progenitor cells was induced. Cellular metabolism was examined on the basis of the fluorescence of the metabolic cofactors NAD(P)H and FAD. The optical redox ratio (FAD/NAD(P)H) and the fluorescence lifetimes of NAD(P)H and FAD were traced using two-photon fluorescence microscopy combined with FLIM. Evaluation of the intracellular pH was carried out with the fluorescent pH sensor SypHer-2 and fluorescence microscopy. In this study, evaluation of the metabolic status of iPSC during dermal and epidermal differentiation was accomplished for the first time with the use of optical metabolic imaging. Based on the data on the FAD/NAD(P)H redox ratio and on the fluorescence lifetimes of protein-bound form of NAD(P)H and closed form of FAD, we registered a metabolic shift toward a more oxidative status in the process of iPSC differentiation into dermal fibroblasts and keratinocyte progenitor cells. Biosynthetic processes occurring in dermal fibroblasts associated with the synthesis of fibronectin and versican, that stimulate increased energy metabolism and lower the intracellular pH. No intracellular pH shift is observed in the culture of keratinocyte progenitor cells, which reflects the incomplete process of differentiation in this type of cells. Presented results provide the basis for further understanding the metabolic features of iPSC during differentiation process, which is essential for developing new treatment strategies in cell therapy and tissue engineering.