Various genotoxicity endpoints have been used to evaluate the diverse hypotheses on the mechanistic principles of particle-induced tumor development, as reviewed in several recent publications (Gonzalez et al., 2008, Landsiedel et al., 2009, Schins and Knaapen, 2007 and Singh et al., 2009). Nevertheless, knowledge about the in vivo situation is still insufficient. To enlarge the body of knowledge, new experimental approaches are highly needed. In the present study, we therefore investigated whether local DNA damage in particle-exposed lung tissue can be detected and quantified in situ with immunohistochemical methods. One advantage of this approach is the possibility to use paraffin-embedded lung tissue from

previous studies. In the present study, Everolimus mouse we used lung tissue from 3-month satellite groups of an existing carcinogenicity study, where animals had been exposed to particles by intratracheal instillation of high doses of crystalline silica (quartz DQ12), carbon black

(Printex® 90), or amorphous silica (Aerosil® 150). A variety of parallel data on histopathology, inflammation, toxicity, and tumor incidences Pictilisib in vivo enabled assessment of the feasibility and informative value of the approach. A panel of genotoxicity markers with different degrees of informative value was chosen to enable identification of the genotoxic modes of action in alveolar lining cells predominantly consisting of epithelial cells, as target cells of lung tumor development. The well-established genotoxicity markers poly(ADP-ribose) (PAR), phosphorylated H2AX (γ-H2AX), 8-hydroxy-2′-deoxy-guanosine (8-OH-dG), and 8-oxoguanine DNA glycosylase (OGG1) were selected for immunohistochemical detection and quantification in the available lung tissue samples. PAR is a posttranslational protein modification

that has been used as a general, overall marker of genotoxic stress (Bürkle, 2001). Its synthesis reflects an early cellular reaction to DNA Abiraterone single- (SSB) or double-strand breaks (DSB). Additionally, PAR is involved in the regulation of cell division and cell cycle progression (for review, see Hakmé et al., 2008) and plays a role in inflammatory processes in asthma and other lung diseases (Virág, 2005). Gamma-H2AX is a phosphorylated core histone variant phosphorylated after DSB induction (Rogakou et al., 1998) and γ-H2AX-containing foci seem to correlate directly with the number of DSB (Sedelnikova et al., 2002). In addition, γ-H2AX formation also occurs during apoptosis (Sluss and Davis, 2006), but nevertheless can be used as a sensitive genotoxicity marker (Watters et al., 2009). 8-OH-dG, a well-characterized oxidative DNA base lesion, is an important and well-established marker of oxidative stress (Kasai, 1997). It is probably the most mutagenic oxidative DNA base modification (Shibutani et al., 1991) and is commonly found in lung tumors (Husgafvel-Pusiainen et al., 2000).