Environmental risk assessment of metal bearing nanoparticles using in vitro toxicity tests with mussels (mytilus galloprovincialis) hemocytes and gill cells

Resumen

Since the recent development of nanotechnology industry nanomaterials are increasingly being applied in our daily lives in many consumer products, with domestic, medical, cosmetic, industrial and military uses. At ¿nano¿ size, materials show different physico-chemical properties compared to the same material of larger size (bulk material), particularly with respect to conductivity, density, hardness, surface area and surface layer composition. At the same time, these novel properties of nanoparticles generate special concerns about their potential hazards to humans and other organisms when released into the environment. At the moment, nanomaterials are regulated in the scope of the European Union by REACH, covered by the definition of ¿substance¿ even though there is no explicit reference to nanomaterials. Nevertheless, recommendations for their safe use or waste disposal are still lacking. In this context, studies on the potential toxicity and risk assessment of nanoparticles (NPs) in different systems such as aquatic environments are urgently needed. As suspension-feeders, mussel cells possess highly developed endocytic and phagocytic mechanisms that could be exploited for the internalization of nano- and micro-scale particles and thus mussels have been proposed as key targets for NPs toxicity. The use of in vitro techniques as a tool for environmental risk assessment has been recommended to uncover particular properties that would then trigger more extensive evaluation. In the present PhD thesis primary cell cultures of hemocytes and gill cells of mussels have been used in order to assess the cytotoxicity and mechanisms of action of different metal bearing NPs: Au, ZnO, SiO2, TiO2, CdS, Ag and CuO. In parallel, human alveolar lung cells were also studied in order to compare responses in mussel and human cells in vitro. Selected NPs varied in size, shape, crystal structure, mode of synthesis and presence of additives. Based on the results, cytotoxicity of metal bearing NPs is related to: 1) type of metal (Ag > Cu > Cd > Zn > Ti > Au > Si), the metal form (ionic > NPs > bulk), the size and shape of NPs and finally with the mode of synthesis (milling > wet chemistry > plasma TiO2 NPs). Additives present in the preparation of NPs can influence their toxicity. Main mechanisms of NPs toxicity in mussel cells involved oxidative stress and genotoxicity and damage to different cell compartments. Human and mussel cells showed similar sensitivity to CuO NPs. The present work concludes that physico-chemical properties of metal bearing NPs, especially aggregation and dissolution in exposure media, strongly influence their cytotoxicity and mechanisms of action to mussel cells. The use of in vitro techniques to assess the toxicity of nanoparticles in mussel and human cells provides valuable data to advance in the risk assessment of these novel materials.