Novel microscopy-based screening method reveals regulators of contact-dependent intercellular transfer.

Contact-dependent intercellular transfer (codeIT) of cellular constituents can have functional consequences for recipient cells, such as enhanced survival and drug resistance. Pathogenic viruses, prions and bacteria can also utilize this mechanism to spread to adjacent cells and potentially evade immune detection. However, little is known about the molecular mechanism underlying this intercellular transfer process. Here, we present a novel microscopy-based screening method to identify regulators and cargo of codeIT. Single donor cells, carrying fluorescently labelled endocytic organelles or proteins, are co-cultured with excess acceptor cells. CodeIT is quantified by confocal microscopy and image analysis in 3D, preserving spatial information. An siRNA-based screening using this method revealed the involvement of several myosins and small GTPases as codeIT regulators. Our data indicates that cellular protrusions and tubular recycling endosomes are important for codeIT. We automated image acquisition and analysis to facilitate large-scale chemical and genetic screening efforts to identify key regulators of codeIT.

Vascular toxicity of ultra-small TiO2 nanoparticles and single walled carbon nanotubes in vitro and in vivo.

Ultra-small nanoparticles (USNPs) at 1-3 nm are a subset of nanoparticles (NPs) that exhibit intermediate physicochemical properties between molecular dispersions and larger NPs. Despite interest in their utilization in applications such as theranostics, limited data about their toxicity exist. Here the effect of TiO2-USNPs on endothelial cells in vitro, and zebrafish embryos in vivo, was studied and compared to larger TiO2-NPs (30 nm) and to single walled carbon nanotubes (SWCNTs). In vitro exposure showed that TiO2-USNPs were neither cytotoxic, nor had oxidative ability, nevertheless were genotoxic. In vivo experiment in early developing zebrafish embryos in water at high concentrations of TiO2-USNPs caused mortality possibly by acidifying the water and caused malformations in the form of pericardial edema when injected. Myo1C involved in glomerular development of zebrafish embryos was upregulated in embryos exposed to TiO2-USNPs. They also exhibited anti-angiogenic effects both in vitro and in vivo plus decreased nitric oxide concentration. The larger TiO2-NPs were genotoxic but not cytotoxic. SWCNTs were cytotoxic in vitro and had the highest oxidative ability. Neither of these NPs had significant effects in vivo. To our knowledge this is the first study evaluating the effects of TiO2-USNPs on vascular toxicity in vitro and in vivo and this strategy could unravel USNPs potential applications.