Despite the wide application of nanomaterials, toxicity studies of nanoparticles (NP) are often limited to in vitro cell models, and the biological impact of NP exposure in mammals has not been thoroughly investigated. Zinc oxide (ZnO) NPs are commonly used in various consumer products. To evaluate the effects of the inhalation of ZnO NP in mice, we studied splice junction expression in the lungs as a proxy to gene expression changes analysis. Female ICR mice were treated with 6.46 × 104 and 1.93 × 106 NP/cm3 for 3 days and 3 months, respectively. An analysis of differential expression and alternative splicing events in 298 targets (splice junctions) of 68 genes involved in the processes relevant to the biological effects of ZnO NP was conducted using next-generation sequencing. Three days of exposure resulted in the upregulation of IL-6 and downregulation of BID, GSR, NF-kB2, PTGS2, SLC11A2, and TXNRD1 splice junction expression; 3 months of exposure increased the expression of splice junctions in ALDH3A1, APAF1, BID, CASP3, DHCR7, GCLC, GCLM, GSR, GSS, EHHADH, FAS, HMOX-1, IFNγ, NF-kB1, NQO-1, PTGS1, PTGS2, RAD51, RIPK2, SRXN1, TRAF6, and TXNRD1. Alternative splicing of TRAF6 and TXNRD1 was induced after 3 days of exposure to 1.93 × 106 NP/cm3. In summary, we observed changes of splice junction expression in genes involved in oxidative stress, apoptosis, immune response, inflammation, and DNA repair, as well as the induction of alternative splicing in genes associated with oxidative stress and inflammation. Our data indicate the potential negative biological effects of ZnO NP inhalation.

*Department of Genetic Toxicology and Nanotoxicology Institute of Experimental Medicine of the Czech Academy of Sciences Prague 14220 Czech Republic

Department for Risk Research and Management Faculty of Safety Engineering VSB Technical University of Ostrava Ostrava 700 30 Czech Republic

Department of Aerosol Chemistry and Physics Institute of Chemical Process Fundamentals of the Czech Academy of Sciences Prague 16502 Czech Republic

Department of Animal Embryology Institute of Animal Physiology and Genetics of the Czech Academy of Sciences Brno 60200 Czech Republic

Department of Chemistry and Toxicology Veterinary Research Institute Brno 62100 Czech Republic

Department of Environmental Analytical Chemistry Institute of Analytical Chemistry of the Czech Academy of Sciences Brno 60200 Czech Republic

Department of Pathological Morphology and Parasitology of Veterinary Medicine University of Veterinary and Pharmaceutical Sciences Brno 612 42 Czech Republic

Department of Pediatric Hematology and Oncology 2nd Faculty of Medicine Charles University Prague and University Hospital Motol Prague 15006 Czech Republic

Department of Transplantation Immunology Institute of Experimental Medicine of the Czech Academy of Sciences Prague 14220 Czech Republic

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