Size of submicrometric and nanometric particles affect cellular uptake and biological activity of macrophages in vitro

Lara Leclerc (1, 2, 3, 4) , Wael Rima (5) , Delphine Boudard (6) , Jérémie Pourchez (2, 3, 4) , Valérie Forest (2, 3, 4) , Valérie Bin (7) , Pierre Mowat (8) , Pascal Perriat (5) , Olivier Tillement (8) , Philippe Grosseau (1, 9, 10) , Didier Bernache-Assollant (1, 2, 3, 4) , Michèle Cottier (6, 11)

1 LPMG-EMSE - Laboratoire des Procédés en Milieux Granulaires
2 IFRESIS - Institut Fédératif de Recherche en Sciences et Ingénierie de la Santé
3 LINA-ENSME - Laboratoire Interdisciplinaire des Nanoparticules Aérosolisées
4 CIS-ENSMSE - Centre Ingénierie et Santé
5 MATEIS - Matériaux, ingénierie et science [Villeurbanne]
6 Service d'Histologie Embryologie
7 BIIO - Biologie, Ingénierie et Imagerie pour l'Ophtalmologie
8 LPCML - Laboratoire de Physico-Chimie des Matériaux Luminescents
9 SPIN-ENSMSE - Centre Sciences des Processus Industriels et Naturels
10 PROPICE-ENSMSE - Département PROcédés Poudres, Interfaces, Cristallisation et Ecoulements
11 LH - Laboratoire d'Histologie

Background: Micrometric and nanometric particles are increasingly used in different fields and may exhibit variable toxicity levels depending on their physicochemical characteristics. The aim of this study was to determine the impact of the size parameter on cellular uptake and biological activity, working with well-characterized fluorescent particles. We focused our attention on macrophages, the main target cells of the respiratory system responsible for the phagocytosis of the particles. Methods: FITC fluorescent silica particles of variable submicronic sizes (850, 500, 250 and 150 nm) but with similar surface coating (COOH) were tailored and physico-chemically characterized. These particles were then incubated with the RAW 264.7 macrophage cell line. After microscopic observations (SEM, TEM, confocal), a quantitative evaluation of the uptake was carried out. Fluorescence detected after a quenching with trypan blue allows us to distinguish and quantify entirely engulfed fluorescent particles from those just adhering to the cell membrane. Finally, these data were compared to the in vitro toxicity assessed in terms of cell damage, inflammation and oxidative stress (evaluated by LDH release, TNF-α and ROS production respectively). Results and conclusion: Particles were well characterized (fluorescence, size distribution, zeta potential, agglomeration and surface groups) and easily visualized after cellular uptake using confocal and electron microscopy. The number of internalized particles was precisely evaluated. Size was found to be an important parameter regarding particles uptake and in vitro toxicity but this latter strongly depends on the particles doses employed.

Mots clés: Uptake quantification; macrophages; fluorescent particles; in vitro toxicity

Domaines: Génie des procédés; Toxicologie

Lara Leclerc, Wael Rima, Delphine Boudard, Jérémie Pourchez, Valérie Forest, et al.. Size of submicrometric and nanometric particles affect cellular uptake and biological activity of macrophages in vitro. Inhalation Toxicology, 2012, 24 (9), pp.580-588. ⟨10.3109/08958378.2012.699984⟩. ⟨hal-00733900⟩