O17 Identification and characterisation of phytochemicals activating endogenous cytoprotective mechanisms

Abstract

Improved health span and lifespan extension in a wide phylogenetic range of species is associated with upregulation of the environmental stress response (ESR). The ESR, driven by the transcription factor Nrf2, provides endogenous antioxidant defence to maintain intracellular redox balance. Phytochemicals which stimulate the ESR thus upregulate cytoprotective mechanisms and ultimately contribute towards delaying the onset of age-related degenerative diseases. Two bioluminescent reporter cell lines were constructed to identify and characterise inducers of the ESR, specifically induction of Nrf2 target genes NQO1 and HMOX1 . The first cell line was generated using random integration of an exogenous luciferase gene under control of an NQO1 gene promoter, and allowed rapid and sensitive high throughput screening of natural chemical libraries. The second assay system employed a more targeted strategy using an adeno-associated virus (AAV) vector to mediate the site-specific integration of a luciferase reporter into the locus of the endogenous ESR gene HMOX1 , thus allowing predictive indexing of the gene’s transcription. The two cell lines were validated with potent ESR inducers sulforaphane and curcumin. A compound identified in the screening of a chemical library from the Marine Biodiscovery Centre (University of Aberdeen, UK) was further characterised in a microarray experiment and compared to known phytochemicals. In combination, these in vitro cell-based assay systems will allow a better understanding of the activity-structure relationship of ESR inducers, and assess the bioactivity of phytochemical metabolites from human serum samples following intervention trials.