Toxicological assessment of mesoporous silica particles in the nematode Caenorhabditis elegans
Introduction
In recent years, inorganic nanomaterials have gained appeal as suitable supports for delivery applications (Mo et al., 2014). Among inorganic supports for encapsulation and controlled release, mesoporous silica particles (MSPs) have received great interest (Valtchev and Tosheva, 2013, Stein, 2003, Soler-Illia and Azzaroni, 2011, Angelos et al., 2007). MSPs have tunable and homogeneous pore size distribution (in the 2–10 nm diameter range), and high specific surface area and volume, which provide a large loading capacity (Salonen and Lehto, 2008, Wight and Davis, 2002). Apart from being a porous structure, MSPs stand out for exhibiting a high concentration of structural defects on their surface in the form of silanol (Si-OH) groups, which can easily react with trialkoxysilane derivatives ((R′O)3-Si-R), enabling the generation of organic−inorganic hybrid supports (Vinu et al., 2005, Kickelbick, 2004). This strategy offers a wide range of new perspectives in the design of on-command release particles to control the delivery of a previously entrapped guest (Angelos et al., 2007, Coll et al., 2013, Aznar et al., 2016, Sancenón et al., 2015). In accordance with this concept, the literature reports examples of MSPs functionalized with a number of different molecules and biomolecules able to deliver the cargo upon the application of various stimuli, such as physical (light, temperature, magnetic fields, ultrasounds) (Mal et al., 2003, Agostini et al., 2012, Fu et al., 2003, Aznar et al., 2011, Giri et al., 2005), chemical (anions, cations, neutral molecules, redox-active species and pH) (Fujiwara et al., 2006, Angelos et al., 2009, Casasús et al., 2008) and biochemical (enzymes, DNA and antibodies) (Oroval et al., 2013, Schlossbauer et al., 2009, Bernardos et al., 2010, Park et al., 2009). However, in spite of the promising applicability of MSPs, their toxic effect after oral administration is still poorly understood.
Among the biological models available, the nematode Caenorhabditis elegans has emerged as a well-suited in vivo model for toxicological studies owing to its established biology and readily scorable life traits. C. elegans is a multicellular organism with a short lifespan (21 days). In addition, experiments with C. elegans are less expensive than those carried out with vertebrate models and allow for a wide set of tests under different conditions in a short time span (The C. elegans Sequencing Consortium, 1998). Moreover, research reports that results obtained with C. elegans can be predictive of those in higher eukaryotes because many physiological processes, signal transduction pathways and genes are conserved (Leung et al., 2008). In addition, quantitative parameters of toxic effects on C. elegans can be easily determined through progeny production, mortality (lifespan), sensitivity to oxidative stress and changes in movement capacity (healthy aging evaluation). These features have led to an increase in the use of C. elegans as a suitable model in toxicological studies. Thus, recent toxicological studies with nanomaterials have been carried out in C. elegans (Cha et al., 2012; Wang et al., 2009; Gonzalez-Moragas et al., 2015). Nonetheless, very few studies have been reported with C. elegans and silica-based particles. In particular, amorphous (non-porous) silica nanoparticles have been evaluated (Pluskota et al., 2009, Scharf et al., 2013). Our results suggest that non-porous silica nanoparticles (smaller than 50 nm) induce premature aging, causing progeny reduction and alterations in phenotypes related to aging. However, studies with C. elegans and MSPs are lacking, and there is an absence of correlation studies of lifespan and healthspan of nematodes fed with MSPs.
Taking into account the increasing interest in the design and use of mesoporous silica particles for delivery applications, we report herein the evaluation of toxicity of nano- and micro-sized MSPs based on C. elegans lifespan and healthspan analysis (movement capacity, resistance to acute oxidative stress and offspring production). Moreover, we studied the impact of functionalization of particles. The results show that surface functionalization of MSPs is a suitable procedure to significantly reduce the toxicity of nano-sized particles.
Section snippets
Chemicals
All the chemicals were purchased at the highest possible grade available and were directly used with no further purification. Chemicals tetraethylorthosilicate (TEOS), cetyltrimethylammonium bromide (CTABr), sodium hydroxide, triethanolamine (TEAH), (3-aminopropyl)triethoxysilane (APTES) were provided by Aldrich. Hydrolyzed starch Glucidex® 47 (5% glucose, 50% maltose, 45% oligosaccharides and polysaccharides) was provided by Roquette.
C. elegans strain and maintenance
C. elegans strain Bristol (wild-type) N2 was obtained from
Synthesis and characterization of MSPs
Bare micro- (M0) and nano-sized (N0) MSPs were synthesized using reported procedures (vide supra). Moreover, both M0 and N0 particles were functionalized with hydrolyzed starch to obtain the corresponding micro- (M1) and nano-sized (N1) starch-functionalized particles. Functionalization was carried out by reaction of M0 and N0 with Glu-N. The derivative Glu-N was prepared by reaction of APTES (3-aminopropyl-triethoxysilane) with hydrolyzed starch (Glucidex@ 47) in ethanol (see Fig. 1). The
Conclusions
Here we show the feasibility of using C. elegans as in vivo model to evaluate the toxicity of MSPs. The intake of micro and nano-sized MSPs has been demonstrated, with accumulation occurring mainly in the gastrointestinal tract and the pharynx. Evaluation of lifespan and other age-related parameters in nematodes exposed to micro and nanoparticles has demonstrated the safety of micro-sized MSPs. By contrast, the toxicity of nano-sized MSPs has been shown through the decrease in lifespan,
Acknowledgments
The authors wish to express their gratitude to the Spanish Government (MINECO Projects AGL2012–39597-C02-01, AGL2012–39597-C02-02, AGL2015–70235-C2-1, MAT2012–38429-C04-01 and MAT2015-64139-C4-1), the Generalitat Valenciana (Project PROMETEOII/2014/047) and Colombian Administrative Department of Science, Technology and Research which supported Ms. Acosta Scholarship. We would also like to thank the Institut de Ciència dels Materials (ICMUV), the Microscopy Service of the Universitat Politècnica
Conflict of interest
The authors declare that there is no competing financial interest and nothing to disclose.
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