The major objective of this proposal is the development and characterization of efficient hematologic, genetic and physiologic assays of hematopoietic failure in the mouse. Although mutagenesis strategies have yielded a number of mouse models of human bone marrow failure syndromes, less attention has been paid to """"""""acquired"""""""" disorders that may result from polygenic traits or gene-environment interactions. This is due in part to the lack of appropriate analytical models and strategies for the detection of these conditions, particularly in their presymptomatic phases. As an outgrowth of the genome project, rapid advances have been made in the development of comprehensive screens for differential gene expression. In parallel, advances in chemical biology have resulted in the development of sensitive methods for the identification of metabolites that may be altered in disease states. The fusion of these two approaches is likely to yield powerful screening tools and facilitate the detection and analysis of complex disorders. Accordingly, they propose to incorporate these assays in a panel that also includes complete blood counts, hematopoietic colony growth assays, and fluorescence-based techniques for the detection of presymptomatic bone marrow failure in mice, Specifically, they will utilize whole-genome microarrays to characterize genes that may have dysregulated expression patterns in bone marrow failure; tandem mass spectrometry to identify metabolites implicated in known syndromes of bone marrow failure; and fluorescence spectrometry for the detection of oxidative stress. These assays will be validated in inbred mouse strains and in mouse models of bone marrow failure at steady-state and after challenge with agents (mitomycin C and chloramphenicol) that can induce bone marrow failure. The data will be correlated with measurements of blood counts and hematopoietic colony growth, and the resulting pre- and post-challenge databases will provide the elements of a diagnostic algorithm for the high- throughput screening of bone marrow failure in mice. Finally, the algorithm will be applied for the detection of early bone marrow failure in a test trial of chemically-mutagenized mice. These studies should result in the development of well-defined assays for pathways that regulate the organismal response to exogenous and endogenous toxins and yield new insights into the pathogenesis of bone marrow failure. (End of Abstract.)
