Article
Chemistry, Analytical
Alexander S. Goryashchenko, Alexey A. Pakhomov, Anastasia Ryabova, Igor D. Romanishkin, Eugene G. Maksimov, Alexander N. Orsa, Oxana Serova, Andrey A. Mozhaev, Margarita A. Maksimova, Vladimir Martynov, Alexander G. Petrenko, Igor E. Deyev
Summary: A genetically encoded fluorescent sensor, SypHerExtra, was developed to measure extracellular media pH in a mildly alkaline range. This sensor can be used in live eukaryotic cells, along with another sensor, SypHer3s, which measures intracellular pH, for quantitative measurements using the FLIM method.
Review
Chemistry, Analytical
Cong Quang Vu, Satoshi Arai
Summary: Genetically encoded fluorescence lifetime biosensors provide a powerful tool for quantitative imaging, enabling precise measurement of cellular metabolites, molecular interactions, and dynamic cellular processes. This review gives an overview of the principles, applications, and advancements in quantitative imaging with genetically encoded fluorescence lifetime biosensors using fluorescence lifetime imaging microscopy (go-FLIM), highlighting the distinct advantages of fluorescence lifetime-based measurements.
Article
Biochemistry & Molecular Biology
Alena Kashirina, Alena Gavrina, Emil Kryukov, Vadim Elagin, Yuliya Kolesova, Alexander Artyuhov, Ekaterina Momotyuk, Vepa Abdyyev, Natalia Meshcheryakova, Elena Zagaynova, Erdem Dashinimaev, Aleksandra Kashina
Summary: This study used fluorescence microscopy and FLIM to investigate the metabolic status and intracellular pH of spheroids carrying Down syndrome. The results showed a predominance of glycolysis in TS21 spheroids compared to normal karyotype spheroids, as well as a more alkaline intracellular pH in TS21 spheroids. The use of fluorescence imaging allowed for a comprehensive comparative analysis, showing that living cells in 3D models can be studied in real time with minimal invasive effects.
Review
Biochemistry & Molecular Biology
A. V. Mamontova, T. R. Simonyan, A. M. Bogdanov
Summary: This article introduces genetically encoded indicators based on fluorescent proteins that can detect changes in various cellular parameters. Most fluorescent indicators can only qualitatively assess the changes, while quantitative measurements require detection of the indicator signal in the time domain and correlation with fluorescence lifetime.
Article
Biophysics
Nadine Bauer, Ivan Maisuls, Abel Pereira da Graca, Dirk Reinhardt, Raghu Erapaneedi, Nils Kirschnick, Michael Schafers, Carsten Grashoff, Katharina Landfester, Dietmar Vestweber, Cristian A. Strassert, Friedemann Kiefer
Summary: Hypoxia plays a crucial role in regulating cell metabolism, migration, and angiogenesis, and is involved in various pathological conditions. In this study, novel genetically-encoded reporters, dUnORS and dUnOFLS, were developed to accurately assess oxygen gradients in cells using ratiometric measurement and FRET-FLIM analysis. These reporters showed promising results in visualizing oxygen availability in vivo.
BIOSENSORS & BIOELECTRONICS
(2023)
Article
Biochemistry & Molecular Biology
Sviatlana Kalinina, Christian Freymueller, Nilanjon Naskar, Bjoern von Einem, Kirsten Reess, Ronald Sroka, Angelika Rueck
Summary: Metabolic FLIM is used to image bioenergetic status in cells and tissue based on fluorescence lifetime of coenzymes. A new algorithm utilizing NAD(P)H and FAD lifetimes to calculate FLIRR has been developed. The study extended FLIRR approach and introduced FLIRR1, FLIRR2, and FLIRR3 to analyze the redox state of cells with different enzymes. The significance of extended FLIRR was compared to the metabolic index, showing the highest difference for FLIRR1 in tumor and normal cells.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2021)
Article
Chemistry, Multidisciplinary
Tsvetan Kardashliev, Alexandra Weingartner, Elvira Romero, Ulrich Schwaneberg, Marco Fraaije, Sven Panke, Martin Held
Summary: Biocatalysis is increasingly used for synthetic purposes in the chemical and pharmaceutical industry. A genetically encoded sensor was designed to measure the activity of oxidative biocatalysts by the amount of intracellularly-formed H2O2. This sensor system accurately assesses whole-cell oxidations in a high-throughput manner, enabling significant improvements in oxidase and oxygenase variants through mutant library screening.
Review
Biochemistry & Molecular Biology
Dorothy Koveal
Summary: Genetically encoded fluorescent biosensors are used to measure chemical changes in single cells rapidly. These biosensors are mainly used in tracking neural activity and neurotransmitter release, but their use and data interpretation for studying brain metabolism pose challenges. Many biosensors are prone to interferences that can lead to ambiguous results. This review discusses current methods of biosensor quantitation, focusing on cellular interferences, methods to avoid false inferences, and recent advances in sensor optimization.
JOURNAL OF NEUROCHEMISTRY
(2023)
Article
Multidisciplinary Sciences
Fu Zheng, Xinyue Zhou, Peng Zou, Chenxin Yu
Summary: Mapping the subcellular organization of proteins is crucial for understanding their biological functions. In this study, the authors developed a genetically encoded photocatalytic labeling method, RinID, to profile the subcellular proteome in multiple organelles. They used miniSOG, a genetically encoded photocatalyst, to locally generate singlet oxygen that reacts with nearby proteins, and labeled proteins were conjugated with an exogenously supplied nucleophilic probe for subsequent affinity enrichment and mass spectrometry-based identification. The authors demonstrated the spatial specificity and depth of coverage in mammalian cells, capturing 477 mitochondrial proteins with 94% specificity using RinID.
NATURE COMMUNICATIONS
(2023)
Article
Biochemistry & Molecular Biology
Mangyu Choe, Denis V. Titov
Summary: In recent years, there has been a rapid development of novel genetically encoded tools for measuring and manipulating metabolism in live cells and animals. This article reviews the available genetically encoded tools, describes their usage, and outlines future directions for development. The focus is on tools for directly measuring and manipulating metabolites, which are key regulators of metabolism and physiology through their action on enzymes, channels, and factors. The aim of this Perspective is to encourage more individuals to utilize these new tools and potentially engage in the development of novel tools for measuring and manipulating metabolism.
NATURE CHEMICAL BIOLOGY
(2022)
Review
Cell Biology
Panagiotis Chandris, Christina C. Giannouli, George Panayotou
Summary: This article provides an overview of available tools for monitoring the dynamics of biochemical reactions and the kinetics of metabolites, and discusses the potential for developing new methodologies that combine classical biochemistry with advanced imaging.
FRONTIERS IN CELL AND DEVELOPMENTAL BIOLOGY
(2022)
Article
Biology
Veronika Miskolci, Kelsey E. Tweed, Michael R. Lasarev, Emily C. Britt, Alex J. Walsh, Landon J. Zimmerman, Courtney E. McDougal, Mark R. Cronan, Jing Fan, John-Demian Sauer, Melissa C. Skala, Anna Huttenlocher
Summary: This study used two-photon imaging of autofluorescent metabolic coenzymes NAD(P)H and FAD to assess macrophage metabolism in the wound microenvironment. Results showed that inhibiting glycolysis led to a more oxidized intracellular redox state in macrophages, and infection and thermal injury induced a more oxidized redox state in wounded tissues. Kinetic analysis revealed a shift toward a more reduced redox state during tissue repair. Metformin improved tissue repair by reducing TNF alpha+ wound macrophages and making the intracellular redox state more reduced, while depletion of STAT6 impaired regeneration by increasing TNF alpha+ wound macrophages and making the redox state more oxidized. The study suggests that autofluorescence of NAD(P)H and FAD can be used to probe the temporal and spatial regulation of macrophage metabolism during tissue damage and repair.
Article
Biochemistry & Molecular Biology
Xingxiu Pan, Mina L. Heacock, Evana N. Abdulaziz, Sara Violante, Austin L. Zuckerman, Nirajan Shrestha, Canglin Yao, Russell P. Goodman, Justin R. Cross, Valentin Cracan
Summary: Impaired redox metabolism plays a critical role in the development of various diseases. This study introduces a novel genetically encoded tool to promote reductive stress in living cells. By using this tool, metabolic and transcriptomic signatures of NADH reductive stress are found to be cellular background specific.
NATURE CHEMICAL BIOLOGY
(2023)
Article
Biochemistry & Molecular Biology
Escarlata Fernandez-Puente, Jesus Palomero
Summary: The study utilized genetically encoded biosensors and quantitative fluorescence microscopy to investigate the role of reactive oxygen and nitrogen species in skeletal muscle, revealing important insights into the redox biology of skeletal muscle in two different models.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2021)
Review
Biotechnology & Applied Microbiology
Sumeng Wang, Wei Jiang, Xin Jin, Qingsheng Qi, Quanfeng Liang
Summary: This article discusses the importance of cofactor engineering in metabolic engineering and the development of biosensors for detecting, monitoring, and regulating intracellular cofactor levels. It also explores the potential applications of cofactor biosensors in metabolic engineering and identifies current research challenges.
CRITICAL REVIEWS IN BIOTECHNOLOGY
(2023)
Article
Biophysics
Liubov Shimolina, Ekaterina Potekhina, Irina Druzhkova, Maria Lukina, Varvara Dudenkova, Vsevolod Belousov, Vladislav Shcheslavskiy, Elena Zagaynova, Marina Shirmanova
Summary: Changes in intracellular pH reflect the metabolic states of cancer cells during tumor growth and dissemination. Monitoring pH is essential for understanding the metabolic mechanisms supporting cancer progression. Genetically encoded fluorescent pH sensors provide real-time tracking of pH in specific subcellular compartments, but their ratiometric readout is not suitable for thick samples ex vivo or tissues in vivo, including solid tumors. Fluorescence lifetime imaging (FLIM) offers a promising alternative to conventional fluorescence microscopy. This study presents a quantitative approach to map pH in cancer cells and tumors in vivo using the fluorescence lifetime of a genetically encoded pH sensor called SypHerRed. The study demonstrates the utility of SypHerRed in visualizing pH in cancer cell culture and mouse tumor xenografts using FLIM and macroscopy. For the first time, the absolute pH value is obtained for tumors in vivo using an optical technique. Additionally, the study shows the simultaneous detection of pH and the endogenous fluorescence of metabolic cofactor NADH, providing complementary insights into the metabolic aspects of cancer. The fluorescence lifetime-based readout and red-shifted spectra of SypHerRed make it a promising instrument for multiparameter in vivo imaging applications.
BIOPHYSICAL JOURNAL
(2022)
Article
Biochemical Research Methods
Arseniy L. Potapov, Marina A. Sirotkina, Lev A. Matveev, Varvara V. Dudenkova, Vadim V. Elagin, Sergey S. Kuznetsov, Maria M. Karabut, Anastasia D. Komarova, Nailya N. Vagapova, Ivan K. Safonov, Irina A. Kuznetsova, Stefka G. Radenska-Lopovok, Elena V. Zagaynova, Natalia D. Gladkova
Summary: In this article, a novel classification method for progressive changes in connective tissue of vulvar lichen sclerosus (VLS) is proposed based on quantitative assessment of second harmonic generation (SHG) signal. Four degrees of VLS development are distinguished using five quantitative characteristics. Early changes of the dermis are identified and described, providing a way to objectively observe tissue changes during treatment.
JOURNAL OF BIOPHOTONICS
(2022)
Article
Multidisciplinary Sciences
Evgeny A. Shirshin, Marina V. Shirmanova, Alexey V. Gayer, Maria M. Lukina, Elena E. Nikonova, Boris P. Yakimov, Gleb S. Budylin, Varvara V. Dudenkova, Nadezhda I. Ignatova, Dmitry V. Komarov, Vladislav V. Yakovlev, Wolfgang Becker, Elena V. Zagaynova, Vladislav I. Shcheslavskiy, Marlan O. Scully
Summary: The heterogeneity of cells determines their response to external factors, and the metabolic heterogeneity of tumor cells is of clinical relevance. Fluorescence lifetime imaging (FLIM) is a non-invasive technique that can sense and quantify cellular metabolism by measuring the fluorescence decay parameters of endogenous fluorophores. This study demonstrates the unique capability of FLIM in accurately discriminating metabolic heterogeneity using a non-parametric analysis approach, which has been validated in cultured cancer cells treated with chemotherapy.
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
(2022)
Article
Multidisciplinary Sciences
Marina Shirmanova, Alena Gavrina, Tatiana F. Kovaleva, Varvara V. Dudenkova, Ekaterina E. Zelenova, Vladislav Shcheslavskiy, Artem M. Mozherov, Ludmila B. Snopova, Konstantin A. Lukyanov, Elena Zagaynova
Summary: Cellular redox status and ROS level are closely related to the apoptotic potential of cancer cells. This study investigates the correlation between ROS production and redox cofactors using fluorescence imaging. The results suggest new therapeutic strategies based on cellular redox balance regulation to improve anti-tumor activity and overcome apoptotic resistance.
SCIENTIFIC REPORTS
(2022)
Correction
Multidisciplinary Sciences
Marina V. Shirmanova, Alena I. Gavrina, Tatiana F. Kovaleva, Varvara V. Dudenkova, Ekaterina E. Zelenova, Vladislav I. Shcheslavskiy, Artem M. Mozherov, Ludmila B. Snopova, Konstantin A. Lukyanov, Elena V. Zagaynova
SCIENTIFIC REPORTS
(2022)
Article
Chemistry, Analytical
Liubov Shimolina, Alexander Gulin, Aleksandra Khlynova, Nadezhda Ignatova, Irina Druzhkova, Margarita Gubina, Elena Zagaynova, Marina K. Kuimova, Marina Shirmanova
Summary: This study aimed to investigate the mechanisms of acquired resistance of colorectal cancer cells to 5-FU. The results showed that resistant cells exhibited higher membrane viscosity and specific lipid composition, which may contribute to the development of resistance.
METHODS AND APPLICATIONS IN FLUORESCENCE
(2022)
Article
Cell Biology
Anastasia N. N. Kazakova, Ksenia S. S. Anufrieva, Olga M. M. Ivanova, Polina V. V. Shnaider, Irina K. K. Malyants, Olga I. I. Aleshikova, Andrey V. V. Slonov, Lev A. A. Ashrafyan, Nataliya A. A. Babaeva, Artem V. V. Eremeev, Veronika S. S. Boichenko, Maria M. M. Lukina, Maria A. A. Lagarkova, Vadim M. M. Govorun, Victoria O. O. Shender, Georgij P. P. Arapidi
Summary: This study summarized a large number of single-cell RNA-sequencing data from various tumor types and normal tissues to identify marker genes and protein markers for cancer-associated fibroblasts (CAFs). The findings revealed common differences in gene expression patterns between normal and cancer-associated fibroblasts, independent of tumor type.
FRONTIERS IN CELL AND DEVELOPMENTAL BIOLOGY
(2022)
Review
Cell Biology
Olga M. Ivanova, Ksenia S. Anufrieva, Anastasia N. Kazakova, Irina K. Malyants, Polina V. Shnaider, Maria M. Lukina, Victoria O. Shender
Summary: Dysregulation of pre-mRNA splicing is a common characteristic of cancer cells and is linked to changes in expression, localization, and mutations of splicing machinery components. Recent studies have revealed that spliceosome components can also affect cellular processes independently of splicing. This review analyzes open source data and literature relating to the impact of splicing factor knockdown on gene expression and splicing in human cells, particularly regarding cell proliferation, migration, cell cycle regulation, DNA repair, and cell death. The review also explores the involvement of splicing factors in intercellular communication and autoregulatory mechanisms, as well as potential strategies for targeting spliceosome components in anticancer therapy.
CELL DEATH & DISEASE
(2023)
Article
Chemistry, Multidisciplinary
Nataliia A. Petrunina, Alina S. Shtork, Maria M. Lukina, Vladimir B. Tsvetkov, Yuri M. Khodarovich, Alexey V. Feofanov, Anastasia M. Moysenovich, Eugene G. Maksimov, Victoria O. Shipunova, Timofei S. Zatsepin, Alexandra N. Bogomazova, Victoria O. Shender, Andrey V. Aralov, Maria A. Lagarkova, Anna M. Varizhuk
Summary: DNA-intercalated motifs serve as convenient scaffolds for designing pH-responsive nanomachines, including biocompatible pH sensors. By using a simple unimolecular dual-labeled iM scaffold and modifying it with abasic or alkyl linkers, the thermal stability and pH-induced conformational transitions of the iM were improved.
Article
Oncology
Irina Druzhkova, Elena Nikonova, Nadezhda Ignatova, Irina Koryakina, Mikhail Zyuzin, Artem Mozherov, Dmitriy Kozlov, Dmitry Krylov, Daria Kuznetsova, Uliyana Lisitsa, Vladislav Shcheslavskiy, Evgeny A. Shirshin, Elena Zagaynova, Marina Shirmanova
Summary: Understanding the interactions between chemotherapeutic agents and cancer cells in the presence of extracellular matrix (ECM) is crucial for improving drug delivery and treatment efficacy. This study used 3D collagen-based models to show that collagen slows down the diffusion of doxorubicin, reducing cellular drug uptake and leading to changes in cellular metabolic status. The protective role of collagen for cancer cells was observed, as it impedes drug delivery and changes cellular metabolic status, highlighting the potential for modifying ECM structure to improve chemotherapy efficacy.
Article
Medicine, Research & Experimental
D. V. Yuzhakova, M. M. Lukina, D. A. Sachkova, G. M. Yusubalieva, V. P. Baklaushev, A. M. Mozherov, V. V. Dudenkova, A. I. Gavrina, K. S. Yashin, M. V. Shirmanova
Summary: Patient-specific in vitro tumor models are a promising platform for studying the mechanisms of oncogenesis and personalized selection of drugs, especially for glial brain tumors with unsatisfactory treatment effectiveness. This study developed a model of a 3D tumor glioblastoma spheroid based on a patient's surgical material and studied its metabolic characteristics using fluorescence lifetime imaging microscopy of metabolic coenzymes.
SOVREMENNYE TEHNOLOGII V MEDICINE
(2023)
Review
Health Care Sciences & Services
Irina Druzhkova, Nadezhda Ignatova, Marina Shirmanova
Summary: This review provides a summary of cell-in-cell (CIC) structures in gastrointestinal tumors. The formation and consequences of CIC structures, as well as their significance as prognostic factors, are discussed. Studies have shown the presence of CIC structures in all gastrointestinal tumors, with a focus on pancreatic cancer. The role of CIC structures in tumor responses to therapy remains poorly understood.
JOURNAL OF PERSONALIZED MEDICINE
(2023)
Proceedings Paper
Cell Biology
Arseniy L. Potapov, Marina A. Sirotkina, Varvara V. Dudenkova, Vadim V. Elagin, Sergey S. Kuznetsov, Maria M. Karabut, Anastasia D. Komarova, Irina A. Kuznetsova, Stefka G. Radenska-Lopovok, Natalya D. Gladkova
Summary: Lichen sclerosus is a common vulvar dermatosis that damages the connective tissue of the dermis. Nonlinear microscopy can evaluate the changes in tissue structure and aid in clarifying the histological classification system in the future.
MULTIPHOTON MICROSCOPY IN THE BIOMEDICAL SCIENCES XXII
(2022)