ToxSci Advance Access published online on September 16, 2008
Toxicological Sciences, doi:10.1093/toxsci/kfn198
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Global Liver Proteomics of Rats Exposed for Five Days to Phenobarbital Identifies Changes Associated with Cancer and with Cyp Metabolism
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* Center for Environmental Health Sciences
Department of Basic Sciences, College of Veterinary Medicine
Mississippi Agriculture and Forestry Experiment Station
Institute for Digital Biology
¶ Life Sciences and Biotechnology Institute, Mississippi State University, Mississippi State, MS, 39762
|| Jointly contributed to this work
111 To whom correspondence should be addressed at the Center for Environmental Health Sciences, College of Veterinary Medicine, Mississippi State University, 240 Wise Center Drive, PO Box 6100, Mississippi State, MS 39762-6100. Phone: (662)325-1255 Fax: (662)325-1031. E-mail: chambers{at}cvm.msstate.edu. Email addresses of additional authors: Dail (dail{at}cvm.msstate.edu), Burgess (burgess{at}cvm.msstate.edu), and Shack (Shack{at}cvm.msstate.edu).
Received June 20, 2008; revision received September 10, 2008; accepted September 10, 2008
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Abstract |
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A global proteomics approach was applied to model the hepatic response elicited by the toxicologically well-characterized xenobiotic phenobarbital (PB), a prototypical inducer of hepatic xenobiotic metabolizing enzymes and a well known nongenotoxic liver carcinogen in rats. Differential detergent fractionation two dimensional liquid chromatography electrospray ionization tandem mass spectrometry (DDF 2D LC ESI MS2) and systems biology modeling were used to identify alterations in toxicologically relevant hepatic molecular functions and biological processes in the livers of rats following a 5 day exposure to PB at 80mg/kg/day or a vehicle control. Of the 3,342 proteins identified, expression of 121 (3.6% of the total proteins) was significantly increased and 127 (3.8%) significantly decreased in the PB group compared to controls. The greatest increase was seen for cytochrome P450 (CYP) 2B2 (167 fold). All proteins with statistically significant differences from control were then analyzed using both Gene Ontology (GO) and Ingenuity Pathways Analysis (5.0 IPA-ToxTM) for cellular location, function, network connectivity, and possible disease processes, especially as they relate to CYP-mediated metabolism and nongenotoxic carcinogenesis mechanisms. The GO results suggested that PB's mechanism of nongenotoxic carcinogenesis involves both increased xenobiotic metabolism, especially induction of the 2B subfamily of CYP enzymes, and increased cell cycle activity. Apoptosis, however, also increased, perhaps as an attempt to counter the rising cancer threat. Of the IPA mapped proteins, 41 have functions which are procarcinogenic and 14 anticarcinogenic according to the hypothesized nongenotoxic mechanism of imbalance between apoptosis and cellular proliferation. Twenty-two additional IPA nodes can be classified as procarcinogenic by the competing theory of increased metabolism resulting in the formation of reactive oxygen species. Since the systems biology modeling corresponded well to PB effects previously elucidated via more traditional methods, the global proteomic approach is proposed as a new screening methodology that can be incorporated into future toxicological studies.
Key Words: cytochrome P450 induction; phenobarbital; global proteome; systems biology; cancer; DDF 2D LC ESI MS2.