Mapping cisplatin-induced viscosity alterations in cancer cells using molecular rotor and fluorescence lifetime imaging microscopy

Abstract

. Significance: Despite the importance of the cell membrane in regulation of drug activity, the influence of drug treatments on its physical properties is still poorly understood. The combination of fluorescence lifetime imaging microscopy (FLIM) with specific viscosity-sensitive fluorescent molecular rotors allows the quantification of membrane viscosity with high spatiotemporal resolution, down to the individual cell organelles. Aim: The aim of our work was to analyze microviscosity of the plasma membrane of living cancer cells during chemotherapy with cisplatin using FLIM and correlate the observed changes with lipid composition and cell’s response to treatment. Approach: FLIM together with viscosity-sensitive boron dipyrromethene-based fluorescent molecular rotor was used to map the fluidity of the cell’s membrane. Chemical analysis of membrane lipid composition was performed with time-of-flight secondary ion mass spectrometry (ToF-SIMS). Results: We detected a significant steady increase in membrane viscosity in viable cancer cells, both in cell monolayers and tumor spheroids, upon prolonged treatment with cisplatin, as well as in cisplatin-adapted cell line. ToF-SIMS revealed correlative changes in lipid profile of cisplatin-treated cells. Conclusions: These results suggest an involvement of membrane viscosity in the cell adaptation to the drug and in the acquisition of drug resistance.

The Role of Plasma Membrane Viscosity in the Response and Resistance of Cancer Cells to Oxaliplatin

“…After 24 h incubation with the drug, the fluorescence lifetime increased to 3.47 ± 0.41 ns, indicating the viscosity was 593 ± 139 cP. The dynamics of these viscosity changes during chemotherapy treatment agrees with our previous results for the cisplatin treatment of HeLa and CT26 cell lines [ 10 ].…”

“…Todor et al, found that the formation of resistance to cisplatin in MCF-7 human breast cancer cells is accompanied by changes in the composition of lipids, specifically by increases in the content of cholesterol and cholesterol ethers, decreased amounts of monoacylglycerols and triacylglycerols, a higher content of sphingomyelin and phosphatidylserine, and a lower content of phosphatidylcholine and phosphatidylethanolamine [ 21 ], which, together, suggest an increase in viscosity. In our previous study on human cervical carcinoma HeLa cells, we also detected increased membrane viscosity in a cisplatin-adapted subline [ 10 ]. In human colorectal cancer cells, increased levels of all phospholipids, including phosphatidylinositol, phosphatidylserine, phosphatidylethanolamine and phosphatidylcholine, were found, this being associated with enhanced cell membrane synthesis [ 22 , 23 ].…”

“…Further investigations are required to find out how common these effects are for different cancer types. We have studied three cell lines so far: HeLa, CT26 and HCT116 and observed consistent viscosity and lipidome changes in each [ 10 ]. In our present research, lipidomic analysis with ToF-SIMS, a complex, labor- and time-consuming technique, was performed at each time-point of cell adaptation and in all experimental conditions, both in vitro and in vivo, and this confined us to only using one cell line.…”

“…While several studies suggest that cisplatin and its analogues can directly interact with lipids and proteins in the plasma membrane [ 33 , 34 ], there is no consistency about the effects of such interactions on the biophysical properties of membranes. Our previous experiments using FLIM of molecular rotors on model membranes showed no viscosity changes in the presence of cisplatin [ 10 ]. Along with the results of mass-spectroscopy, this indicates that changes in the lipid composition are major causes of viscosity alterations in cells.…”

“…While DNA is the primary target of the platinum complexes, multiple off target effects have been documented, including changes to the plasma membrane [ 9 ]. It is thought that physical properties of the membrane, specifically higher viscosity, are involved in the mechanisms of cell adaptation and resistance to cisplatin [ 10 ]. At the same time, lower viscosity is known to promote apoptotic pathways by facilitating the clustering of death receptors at the plasma membrane, thus sensitizing the cells to cisplatin [ 11 ].…”