Synthesis, characterization and bioimaging of fluorescent labeled polyoxometalates

Georg Geisberger,Caroline Maake,Emina Besic Gyenge,Doris Hinger,Peter Bösiger,Greta R Patzke

doi:10.1039/c3dt50414j

Abstract

A fluorescent labeled Wells-Dawson type POM ({P2W17O61Fluo}) was newly synthesized and characterized by a wide range of analytical methods. {P2W17O61Fluo} was functionalized with fluorescein amine through a stable amide bond, and its long time stability was verified by UV/vis spectroscopic techniques at physiologically relevant pH values. No significant impact on the cell viability or morphology of HeLa cells was observed for POM concentrations up to 100 μg mL(-1). Cellular uptake of fluorescent {P2W17O61Fluo} was monitored by confocal laser scanning microscopy. POM uptake occurs mainly after prolonged incubation times of 24 h resulting in different intracellular patterns, i.e. randomly distributed over the entire cytoplasm, or aggregated in larger clusters. This direct monitoring strategy for the interaction of POMs with cells opens up new pathways for elucidating their unknown mode of action on the way to POM-based drug development.

Highlights

Our recent studies were focused on nanocapsules based on bioactive POMs and various chitosan derivatives, preferably carboxymethyl chitosan (CMC) and trimethyl chitosan (TMC)
No cytotoxicity of {P2W17O61Fluo} towards HeLa cells was observed for concentrations up to 100 μg mL−1 regardless of the POM countercation, and cell viability as well as morphology were not significantly changed
Cellular uptake tests provided the first evidence of localization of pristine POMs within cells, and two different intracellular distribution patterns were observed

Summary

Introduction

Polyoxometalates (POMs) have been attracting interdisciplinary research interest for decades due to their virtually unlimited structural flexibility which provides access to a wide range of key material properties.[1,2,3,4,5] The high bioactive potential of POMs has been under intense investigation since the 1980s, and their observed antibacterial, anticancer and antiviral activity opens up new perspectives for the development of lowcost and tuneable inorganic drugs, e.g. against new viral threats and the increasing problem of bacterial resistances.[6,7,8,9,10,11]the multitude of empirical bioactivity data derived from POM-based in vitro and in vivo studies[12] did not afford a proportional degree of insight into their biochemical pathways. We directly observed cellular uptake of bioactive POM composites: complementary CLSM (confocal laser scanning microscopy) and TEM (transmission electron microscopy) techniques on labeled and unlabeled nanocomposites, respectively, revealed that they are taken up within less than 1 h with a strong preference for the perinuclear region of HeLa cells.[19] As the step towards elucidation of biochemical POM routes, we here present new evidence for cellular uptake of pristine POMs. Microscopic tracking of fluorescent POMs containing lanthanoid heteroatoms, such as europium or gadolinium, appears to be the most straightforward approach at first glance. Fluorescein labeled POMs (50 and 100 μg mL−1) were tested for cytotoxicity vs HeLa cells with cell viability (MTT) assays (Fig. 4).

Results

Conclusion