Photothermal therapy with silver nanoplates in HeLa cells studied by in situ fluorescence microscopy.

María Belén Rivas Aiello,Pedro David Gara,Daniel O. Mártire,Thomas Gensch,Eugenia Zelaya,Julio C. Azcárate,Gabriela N. Bosio

doi:10.1039/d0bm01952f

Abstract

Photothermal therapy (PTT) is a noninvasive treatment for cancer relying on the incorporation of NIR-light absorbing nanomaterials into cells, which upon illumination release heat causing thermally induced cell death. We prove that irradiation of aqueous suspensions of poly(vinylpyrrolidone)-coated silver nanoplates (PVPAgNP) or PVPAgNP in HeLa cells with red or NIR lasers causes a sizeable photothermal effect, which in cells can be visualized with the temperature sensing fluorophore Rhodamine B (RhB) using spinning disk confocal fluorescence microscopy or fluorescence lifetime imaging. Upon red-light irradiation of cells that were incubated with both, RhB and PVPAgNP at concentrations with no adverse effects on cell viability, a substantial heat release is detected. Initiation of cell death by photothermal effect is observed by positive signals of fluorescent markers for early and late apoptosis. Surprisingly, a new nanomaterial-assisted cell killing mode is operating when PVPAgNP-loaded HeLa cells are excited with moderate powers of fs-pulsed NIR light. Small roundish areas are generated with bright and fast (<1 ns) decaying emission, which expand fast and destroy the whole cell in seconds. This characteristic emission is assigned to efficient optical breakdown initiation around the strongly absorbing PVPAgNP leading to plasma formation that spreads fast through the cell.

Highlights

Photothermal therapy (PTT) is defined as the killing of cancer cells, which have taken up light absorbing particles, that release efficiently heat upon irradiation with NIR light.[1]
To prove the ability of the silver nanoplates to act as photothermal agents, we report the use of Spinning Disk Confocal Microscopy (SDCM) and Fluorescence Lifetime Imaging Microscopy (FLIM) techniques to monitor changes in the fluorescence intensity and lifetime of the optical thermometer Rhodamine B (RhB) in HeLa cells incubated with RhB and PVPAgNP
We have demonstrated that there is a photothermal effect upon NIR excitation of both, suspensions of PVPAgNP and HeLa cells incubated with PVPAgNP, but the generation of the short lived emission observed in the FLIM experiments could be a physiological response of the cell upon generation of singlet molecular oxygen, O2(1Δg), photosensitized by PVPAgNP

Summary

Introduction

Photothermal therapy (PTT) is defined as the killing of cancer cells, which have taken up light absorbing particles, that release efficiently heat upon irradiation with NIR (typically laser) light.[1]. Plasmonic nanoparticles based on gold are very popular for PTT applications and have been used as photothermal transducers for the treatment of highly aggressive tumours[6,7,8,9,10,11] in combination with immune therapy.[12] In addition, silverbased nanomaterials have been developed and utilized. Boca et al conducted cytotoxicity assays and found that chitosan-coated silver nanotriangles exhibited good compatibility for healthy human embryonic cells, while showing a dose-dependent phototoxicity towards NCI-H460 cancer cells upon excitation with 800 nm light.[5] These results motivated us to prepare and characterize polyvinylpyrrolidone (PVP)-coated silver nanoplates (PVPAgNP) to be used in PTT assays with HeLa cells. It has been shown that silver nanoplates and in particular silver nanotriangles are best suited among the different types of silver nanomaterials in PTT appli-

Methods

Results

Conclusion