FRIVIC · FLUORESCENT ROTORS FOR IMAGING VISCOSITY IN CELLS

FLUORESCENT ROTORS FOR IMAGING VISCOSITY IN CELLS

Measuring viscosity in biological systems is of paramount importance for the understanding of biophysical processes governing both the normal cell function and the cell demise. More specifically, membrane viscosity have been linked with alterations in physiological processes, such as carrier-mediated transport, activities of membrane-bound enzymes and receptor binding, which, in turn, are associated with aging and disease pathogenesis. Three most widespread methods based on fluorescence have been developed to probe the viscosity on a microscopic scale: fluorescence recovery after photobleaching (FRAP), fluorescence anisotropy and the use of molecular rotors. Molecular rotors refer to small synthetic fluorophores in which fluorescence emission is sensitive to the viscosity of the surrounding environment. This sensitivity, based on the non-radiative decay of the excited state, allows a precise calibration of fluorescence parameters with viscosity. Fluorescence lifetime imaging (FLIM) is an excellent technique to probe the viscosity in biological systems as it is concentration independent and a single excitation wavelength/emission interval is required. The aim of this project is to study the sensitivity to viscosity of new BODIPY (Boron DIPYrromethene) derivatives. The design and synthesis of the latest involve two different collaborations. In the first one with Dr Rachel Meallet-Renault and Dr Gilles Clavier, from the Supramolecular and Macromolecular Photophysics and Photochemistry (PPSM) laboratory, the fluorophores were designed to be emitting in the near-infrared of the electromagnetic spectrum, presenting a better match with the tissue therapeutic window. I synthesised two red-emitting BODIPY derivatives and carried out their purification at Imperial College London as well as spectroscopic experiments to explore their properties as molecular rotors. Based on the results obtained so far, we can conclude those compounds don’t show any molecular rotor properties but seemingly prove to be sensitive to the temperature instead, with probably a contribution of the polarity sensitivity as well. These results will be reported in a paper which is currently in preparation and will be submitted to journals such as J Phys Chem B, PCCP or Chem. –Eur. J. In the second collaboration with Dr Ismael Lopez-Duarte and Dr Maria A. Izquierdo-Arcusa, two BODIPY derivatives containing a double positive charge on the aliphatic chain were synthesised, which was aimed to stain specifically the plasma membrane of live cells. Both compounds are shown to be effective molecular rotors and to selectively stain the cell plasma membrane. A very first measurement of the viscosity of the cell plasma membrane has been done and these results have been published in the journal of Chemical Communications where I am a joint first author, along with Dr Lopez Duarte. A first application based on these molecular rotor properties have also been explored, in a collaboration with Dr Ulrike Eggert and Dr Eleonora Muro, from King’s college. In staining experiment on Hela cells in the process of cell division and at different stages, it has been shown that the viscosity in the plasma membrane doesn’t change significantly when the cells undergo the division process. This unexpected result might be the beginning of a new understanding of this biologic process. The differences and complementarity between the properties of these two positively charged BODIPY derivatives as viscosity probes will be reported in a paper which is in preparation and to be submitted to journals such as Chem Commun or Chem- Eur. J.

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