CC BY
20B-O-6
Visualization of viscous changes of membranes of tumor cell
during chemotherapy
L. Shimolina1' 2, A. Hlynova1,M. Lukina1, N. Ignatova1, M. Kuimova3, E. Zagaynova12, M.
Shirmanova1
1 - Privolzhsky Research Medical University, 603005 Minin and Pozharsky Sq., 10/1, Nizhny Novgorod, Russia
2 - Nizhny Novgorod State University, 603950 Gagarin Av., 23, Nizhny Novgorod, Russia 3 - Imperial College London, Faculty of Natural Sciences, Department of Chemistry, SW7 2AZ, London, United
Kingdom
shimolina. l@mai.ru
The microscopic viscosity is one of the key parameters that controls the diffusion rate of molecular species in living cells at the microscopic level. While methods to measure bulk (macroscopic) viscosity of tissues are well developed, imaging viscosity at the microscopic scale remains a challenge, especially in vivo. The microscopic viscosity of individual domains of live cells can be determined using fluorescence-based methods, including fluorescent molecular rotors. Molecular rotors are small synthetic viscosity-sensitive fluorophores in which fluorescence parameters, in particular fluorescence lifetime, are strongly correlated to the microviscosity of their immediate environment [1]. Although chemotherapy remains one of the main types of treatment for cancer, treatment failure is a frequent occurrence, emphasizing the need for new approaches to the early assessment of tumor response. Recent studies suggest that tumor response to chemotherapy is determined by not only interaction of the drug with the primary target (e.g. nuclear DNA or microtubules) but can include multiple physiological and physicochemical changes [2]. The study of the effects of chemotherapeutic drugs on the viscosity of living cells is important for better understanding the mechanisms of the drug action and evaluating the effectiveness of therapy.
The present work is aimed at the study of microviscosity of plasma membrane in cancer cells using the fluorescent molecular rotor BODIPY 2 and fluorescence lifetime imaging FLIM microscopy during chemotherapy.
The study was performed on cultured cancer cells CT26 (mouse colorectal cancer) and Hela Kyoto (human cervical cancer). Multiphoton tomograph MPTflex (JenLab, Germany) equipped with a TCSPC-based FLIM module (Becker&Hickl Inc., Germany) was used to detect the fluorescence lifetime of a molecular rotor. Viscosity was measured in the plasma membranes of individual cells using the fluorescent molecular rotor BODIPY2 (ex. 800 nm, em. 409-680 nm) First, we developed protocol for imaging plasma membrane viscosity of cancer cells in vitro at the microscopic level. The developed protocols were applied to measure changes in membrane viscosity during chemotherapy in vitro in cell monolayers and 3D spheroids. We showed a significant increase in membrane viscosity in viable cells CT26 and Hela Kyoto in 24 h after cisplatin treatment up to 400 ± 27 cP. At earlier time-points (30-60 min) a tendency to decrease in membrane viscosity was displayed for both cancer cell lines.
The study of the action of drugs on the viscosity of living cells is important for a deep understanding of the mechanisms of their cytotoxicity. This work is supported by the Russian Science Foundation under grant No: 20-14-00111.
[1] Bull J. A. and Kuimova M. K., Chem. Commun., 2013.
[2] Rebillard A. et al. Canc. Res., 2007.
