Evaluation of gene expression profile of keratinocytes in response to JP-8 jet fuel Luis A. Espinoza, a Peijun Li, a Richard Y. Lee, b Yue Wang, c A. Hamid Boulares, a, ** Robert Clarke, b and Mark E. Smulson a, * a Department of Biochemistry and Molecular Biology, Georgetown University School of Medicine, Washington, DC 20057, USA b Department of Oncology, Georgetown University School of Medicine, Washington, DC 20057, USA c Department of Electrical and Computer Engineering, Virginia Polytechnic Institute and State University, Alexandria, VA 22314, USA Received 20 February 2004; accepted 31 March 2004 Available online 15 June 2004 Abstract The skin is the principal barrier against any environmental insult. Therefore, there is a high risk for a large number of military and civilian personnel exposed to jet fuel JP-8 to suffer percutaneous absorption of this fuel. This paper reports the use of cDNA microarray to identify the gene expression profile in normal human epidermal keratinocytes exposed to JP-8 for 24-h and 7-day periods. The effects of JP- 8 exposure on keratinocytes at these two different periods induced a set of genes with altered expression in response to this type of insult. Microarray data were visualized using a novel algorithm based on simple statistical analyses to reduce data dimensionality and identify subsets of discriminant genes. Predictive neural networks were built using a multiplayer perceptron to carry out a proper classification task in microarray data in the untreated versus JP-8-treated samples. The pattern of expressions in response to JP-8 provides evidences that detoxificant-related and cell growth regulator genes with the most variability in the level of expression may be useful genetic markers in adverse health effects of personnel exposed to JP-8. The approaches in our analysis provide a simple, safe, novel, and effective method that is reliable in identifying and analyzing gene expression in samples treated with JP-8 or over potential toxic agents. Gene expression data from these studies can be used to build accurate predictive models that separate different molecular profiles. The data establish the use and effectiveness of these approaches for future prospective studies. D 2004 Elsevier Inc. All rights reserved. Keywords: Keratinocytes; JP-8 jet fuel; cDNA microarrays; Algorithm Introduction JP-8, a kerosene-based fuel used by US and NATO forces, is a complex mixture of both aliphatic and aromatic hydrocarbons, the latter of which is associated with DNA damage and carcinogenesis. JP-8 is toxic if absorbed into the body in sufficient amounts; exposure to JP-8 has been associated with symptoms such as fatigue, headache, and skin irritation (Smith et al., 1997; Zeiger and Smith, 1998). Several studies have shown that exposure of skin to JP- 8 induces immunosuppression in a dose-dependent fashion (Ullrich, 1999) and affects immunological memory function in mice (Ramos et al., 2002). JP-8 treatment also induces skin inflammatory response including the induction of iNOS and IL-1 (Kabbur et al., 2001), development of erythema and edema (Kanikkannan et al., 2001), and histological abnormalities in all the epidermal layers (Rossi et al., 2001). These data from laboratory animals suggest that JP-8 exposure induces several alterations that can lead to tumorigenesis in skin that is continuously exposed to this jet fuel. Accordingly, exposures to petroleum middle distillates have been related with increases in tumor incidence in mice treated for long periods (Nessel et al., 1999). Previous studies in our laboratory have shown that JP- 8 exposure in several human cell lines induces biochem- ical and morphological markers of apoptotic cell death 0041-008X/$ - see front matter D 2004 Elsevier Inc. All rights reserved. doi:10.1016/j.taap.2004.03.022 * Corresponding author. Department of Biochemistry and Molecular Biology, Georgetown University Medical Center, Basic Science Building, Room 351, 3900 Reservoir Road NW, Washington, DC 20057. Fax: +1- 202-687-7186. E-mail address: smulson@georgetown.edu (M.E. Smulson). ** Present address. Louisiana State University Health Science Center, Department of Pharmacology and Experimental Therapeutics and the Stanley Cancer Center, New Orleans, LA, 70112, USA. www.elsevier.com/locate/ytaap Toxicology and Applied Pharmacology 200 (2004) 93– 102