Estdio toxicidad in vitro de nanopartículas

GRADUONDOKO MASTERRA - MASTER DE POSTGRADO - POSTGRADUATE MASTER (2008-2009)

INGURUMEN-KUTSADURA ETA TOXIKOLOGIA CONTAMINACION Y TOXICOLOGIA AMBIENTALES ENVIRONMENTAL CONTAMINATION AND TOXICOLOGY

Mención de Calidad MEC 2005-2012
MASTER TESIAREN PROIEKTUA PROYECTO DE TESIS DE MASTER MASTER THESIS PROJECT

In vitro toxicity assays to evaluate environmental health and risk assessment ofmetals and metallic nanoparticles

ALBERTO KATSUMITI KODO FILHO
Laboratory of Cell Biology and Histology Department of Zoology and Animal Cell Biology University of the Basque Country

Leioa, 2009ko iraila / septiembre 2009 / September 2009

I DEX

1. ABSTRACT……………………………………….………………………………………..1 2. I TRODUCTIO ………………………………………………….………………………5 2.1. MUSSELS AS SE TI ELS SPECIES OF E VIRO ME TALPOLLUTIO ………….............................................................................................................5 2.2. In vitro TOXICITY TESTI G……………………….………………………….……...6 2.3. METALS A D METALLIC A OPARTICLES…………….……………………..10 2.3.1. SOURCES……………………………………………………………………………..10 2.3.2. ZI C……………………………………………………………………………….......11 2.3.3. COPPER……………………………………………….…………….………..............122.3.4. CADMIUM…………………………………………….…………….……….............15 2.3.5. METALLIC A OPARTICLES…………………………………………...............15 3. HYPOTHESIS……………………………………………………………………....…….17 4. OBJECTIVES…………………………………………………………………….............18 5. MATERIAL A D METHODS………………………………………………….............18 5.1. OBTAI I G MUSSEL CELLS.……………………………………...…………........18 5.2. SEX-DEPE DE T DIFFERE CES I SE SITIVITY TO CADMIUM IMUSSEL CELLS.……………………..…………………….……………...……………….20 5.3. EFFECTS OF ACCLIMATIZATIO O MUSSEL CELLS…………..…………..20 5.4. EFFECTS OF METALS A D METALLIC A OPARTICLES O MUSSEL

CELLS………………………………….…………………………...……………………….20 5.5. EFFECTS OF METALLIC A OPARTICLES O HUMA ALVEOLAR LU G

CELLS…………………………………………………………………….............................21 5.6. In vitroASSAYS…………………..……..……………………………………..............21 5.7. A OPARTICLES CHARACTERIZATIO ...…………..………………………...23 5.8. CHEMICAL A ALYSIS…………………….....………...………………….………...28 5.9. STATISTICS…………..………………………………………………………………..29 6. RESULTS A D DISCUSSIO ..……………………………………………...…............29 6.1. SEX-DEPE DE T DIFFERE CES I SE SITIVITY TO CADMIUM I

MUSSEL CELLS…………………………………...………………………...……………..29 6.2. EFFECTS OF ACCLIMATIZATIO O MUSSEL CELLS..……………………..31 6.3. EFFECTS OF ZI C A D COPPER O MUSSELCELLS.....…………………….32

6.4. EFFECTS OF COPPER

A OPARTICLES O

HUMA

ALVEOLAR LU G

CELLS.……………………………………………………………………..….....…………..39 7. CO CLUSIO S...….………………………………………………………...………......43 8. ACK OWLEDGEME TS..………………………………...…………...………...……43 9. REFERE CES......………………..…………………………………………..….............44

1. ABSTRACT

The development of in vitro-based toxicity test methods during thelast four decades has largely occurred as a reaction of the society to reduce and substitute the use of animals in research. Due to the extensive use of bivalve mollusks such as mussels as sentinel species in environmental risk assessment, there is a considerable interest in the development of in vitro models in this species. Mussel gill cells and hemocytes are primary targets for the action ofenvironmental pollutants. An interesting cellular model in human in vitro toxicology is the alveolar lung cell. The alveolar tissue is composed basically of two epithelial cells: type I and type II. Most of the alveolar surface (90%) is covered by Type I (AT I) cells. AT I cells in vivo are easily damaged by environmental challenges from chemical exposures. Metals are important pollutants found inthe aquatic environment whereas metallic nanoparticles (NPs) are emerging pollutants whose effects on living organisms are not fully understood. In the present work, mussel hemocytes and gill cells were exposed to metals: ionic Zn, bulk ZnO and ZnO NPs (0.001 - 10 mg/L) and to ionic Cu, bulk CuO and CuO NPs (0.005 – 0.5 mg/L), and AT I cells were...