Sulfur mustard (HD, SM), is a chemical warfare agent that within hours causes extensive blistering at the dermalCepidermal junction of skin. inflammatory response, and chemotaxis; these findings are consistent with other reported results for shorter time periods. Selected genes were chosen for RT-PCR verification and showed correlations in the general trends for the microarrays. Interleukin 1 beta was checked for biological analysis to 490-46-0 IC50 confirm the presence of protein correlated to the corresponding microarray data. The impact of a matrix metalloproteinase inhibitor, MMP-2/MMP-9 inhibitor I, against SM exposure was assessed. These results can help in understanding the molecular mechanism Rabbit Polyclonal to CDC2 of SM-induced blistering, as well as to test the efficacy of different inhibitors. in cell culture (Cowan et al., 2000) and in an mouse model (Powers et al., 2000). It was tested whether or not topical skin treatment with MMP-2/MMP-9 inhibitor I [(2R)-2-[(4-Biphenylylsulfonyl)amino]-3-phenylpropionic acid (Fig. 1B), was effective in reducing the secondary damage caused by MMP-9. Using microarray analysis, the major gene pathways that are activated in response to SM skin exposure were identified. The rationale for using microarray technology was that it may identify potential new target molecules or pathways that could be used for medical intervention against SM-induced injury. It also has the potential to identify biomarkers that could be used as quantitative tools for novel compound evaluation. Fig. 1 Chemical structures of Sulfur Mustard and the MMP inhibitor. To date, there have only been a handful of microarray studies involving analysis of mouse skin treated with sulfur mustard (Rogers et al., 2004; Sabourin et al., 2004; Dillman et al., 2006). These studies only focused on gene changes at early time points within the first 24 h post exposure. The present study expanded the observed time-course to seven days in length. It also analyzed the impact of a specific MMP inhibitor to the SM-induced skin damage by assessing ear tissues from mice exposed to SM for histological damage (with 490-46-0 IC50 and without topical pre-treatment with MMP-2/MMP-9 inhibitor I). Since microarray analysis data vary according to the method employed, analysis was performed using several different techniques in order to compare the gene variations with and without MMP-2/MMP-9 inhibitor I application and generate statistically significant data. A majority of the techniques employed in this study to analyze gene expression microarray data are supported by the USFDA’s ArrayTrack system (Tong et al., 2003; 2004). In the present study, the multiple analysis methods supported by Array-Track were used, both within ArrayTrack and through links to other analysis platforms. Methods Experimental design A schematic depiction of the experiments and subsequent analysis is shown in Fig. 2. The mice were divided into three groups (each group is represented by three post-exposure time-points). The three groups included: 1) untreated, control group; 2) mice treated with sulfur mustard; 3) mice treated with sulfur mustard after pre-treatment with the inhibitor. The microarray gene expression data 490-46-0 IC50 were then analyzed to identify the genes that have been significantly expressed using several different statistical and pathway analysis techniques. Details of the experiments and the analysis methods follow. Fig. 2 Schematic depiction of the microarray experimental 490-46-0 IC50 design and subsequent analyses. *Other controls are described in Methods section (a comparison of all the various control groups showed no significant differences between the groups). SM exposure Animals were exposed to SM as reported in Shakarjian et al. (2006). Briefly, for the mouse ear exposures, male CD1 mice [Charles River Laboratories, Portage, MI; values, novel pathway activations were identified in the sulfur mustard exposure time-points 490-46-0 IC50 when compared to controls (Table 1). The analysis showed that cytokineCcytokine receptor interaction and the Jak-STAT signaling pathways are impacted at all the post-exposure time points. If a comparative analysis was performed in a slightly different way, using the IPA system, it would be possible to identify the top biological functions affected by sulfur mustard exposure. The data is presented in Table 2 and identifies.