Extracellular Vesicles From Microalgae: Uptake Studies in Human Cells and Caenorhabditis elegans

Abstract

Extracellular vesicles (EVs) are lipid membrane nano-sized vesicles secreted by various cell types for intercellular communication, found in all kingdoms of life. Nanoalgosomes are a subtype of EVs derived from microalgae with a sustainable biotechnological potential. To explore the uptake, distribution and persistence of nanoalgosomes in cells and living organisms, we separated them from a culture of the chlorophyte Tetraselmis chuii cells by tangential flow filtration (TFF), labelled them with different lipophilic dyes and characterized their biophysical attributes. Then we studied the cellular uptake of labelled nanoalgosomes in human cells and in C. elegans, demonstrating that they enter the cells through an energy dependent mechanism and are localized in the cytoplasm of specific cells, where they persist for days. Our data confirm that nanoalgosomes are actively uptaken in vitro by human cells and in vivo by C. elegans cells, supporting their exploitation as potential nanocarriers of bioactive compounds for theranostic applications.

Synthesis and Characterization of a Biocompatible Nanoplatform Based on Silica-Embedded SPIONs Functionalized with Polydopamine

“…Strains used in this work have been provided by the Caenorhabditis Genetics Center (CGC), which is funded by the NIH Office of Research Infrastructure Programs (P40 OD010440): wild-type strain N2, Bristol variety; CL2166 dvIs19 [(pAF15) pgst-4 :GFP:NLS] that expresses an oxidative stress-responsive GFP. Animal treatments with SPIONs@SiO 2 -PDA and Milli-Q water as mock were performed in iquid in 96-multiwell plates for the entire life-cycle of the animals (chronic treatment) . Synchronized eggs, obtained by bleaching, were resuspended in M9 buffer (3 g KH 2 PO 4 ; 6 g Na 2 HPO 4 ; 5 g NaCl; 1 M mgSO 4 ; H 2 O to 1 L) with 2× antibiotic/antimycotic solution (Sigma-Aldrich, A5955), 5 ng/mL cholesterol and OP50.…”

“…Animal treatments with SPIONs@SiO 2 -PDA and Milli-Q water as mock were performed in iquid in 96-multiwell plates for the entire lifecycle of the animals (chronic treatment). 36 Synchronized eggs, obtained by bleaching, were resuspended in M9 buffer (3 g KH 2 PO 4 ; 6 g Na 2 HPO 4 ; 5 g NaCl; 1 M mgSO 4 ; H 2 O to 1 L) with 2× antibiotic/antimycotic solution (Sigma-Aldrich, A5955), 5 ng/mL cholesterol and OP50. 60 μL containing ∼30 eggs were aliquoted in each well.…”

Harnessing Nature’s nanoSecrets: biocompatibility, biodistribution and bioactivity of extracellular vesicles derived from microalgae

“…Our previous study on nanoalgosomes showed that they are highly stable in human blood plasma, non-cytotoxic in vitro and can be taken up by various cellular systems 15,24 . Here, to fully characterize the compatibility, distribution, and bioactivity of nanoalgosomes, we conducted a comprehensive analysis both in vivo, using Caenorhabditis elegans and mice, in vitro with human cell lines, and ex vivo with human basophils from whole blood.…”

“…After quality checks on the nanoalgosome batches (Figure 1), we treated wild-type animals through in liquido culturing for 72 hours with increasing concentrations of nanoalgosomes (1,20,64, 128 µg/mL), first evaluating animal viability and capability of reaching adulthood. These concentrations were chosen because we had previously demonstrated that nanoalgosomes are successfully taken up by C. elegans at 20 µg/mL 24 .…”

“…We focused on sEVs isolated from the marine photosynthetic chlorophyte Tetraselmis chuii, which is rich in vitamin E, carotenoids, chlorophyll, and tocopherols with anti-inflammatory and antioxidant properties 15,16,[21][22][23] . Our analysis showed that nanoalgosomes are non-cytotoxic and can be taken up by different cellular systems in vitro as well as in vivo by the invertebrate model organism Caenorhabditis elegans (Nematoda), an alternative to the mammalian model, that is faster and less expensive to use and whose body transparency is advantageous for studying nanoparticle toxicity, distribution, and uptake 24 . In addition, the current study aims to fully characterize the in vivo toxicity, immuno-safety, biodistribution, and bioactivity of nanoalgosomes through a multi-branched approach starting with studies on living organisms (C. elegans and mice) and delving into molecular mechanisms at the subcellular level.…”