Franny Jongbloed

247 9 SUMMARY AND DISCUSSION on both renal and liver array results, in which this protection is induced by activation of nuclear receptor signaling, stress resistance pathways as well as inhibition of cell growth to a more maintenance state, mediated via key transcription factors mandatory for the beneficial effects. The transcriptome profile analyses of all diets as done in chapter 2 , chapter 4 and chapter 5 , were performed in biopsies taken after the dietary intervention but before induction of the acute stress of IRI. In chapter 3 , we focused on the effects shortly after the induction of the stressor, which was in this case the chemotherapeutic agent irinotecan. We applied three days of fasting in C26 colorectal carcinoma (CRC)-bearing BALB/c male mice. Twelve days after tumor inoculation, mice were either fasted for three days or continued to be AL fed after which each group was given one high dose of irinotecan or vehicle. The effects of fasting were investigated on a transcriptional level in liver and tumor tissue collected 12 hours after the administration of irinotecan, when mice were sacrificed. In liver tissue, unbiased transcriptome analysis showed that fasting markedly reduced the number of gene expression changes caused by administration of irinotecan. Together with more homogenous gene expression profiles in liver tissue of mice subjected to fasting, these data point towards a dampening of the changes caused by irinotecan, thereby partially protecting healthy tissue from the response to irinotecan. Further evidence of this hypothesis is provided by the dysregulation of the genotoxic response due to fasting as compared to liver tissue of AL fed mice given irinotecan; the AL fed mice showed activation of DNA damage pathways, including RhoA signaling, as well as cellular stress and cytokine signaling pathways, whereas fasting mice subjected to irinotecan showed indirect inhibition of RhoA via activation of RhoGDI signaling, and vast reduction of chemokine and cytokine pathways 39 . In addition, pathways involved in growth and proliferation, such as the mTOR signaling via suppression of the p7056 pathway, were downregulated which points once more towards the shift from growth to maintenance as described before 38,40 . Interesting, the NRF2-pathway remained activated in the fasting mice after the stressor ( i.e. irinotecan) was given, which may indicate a key role for NRF2 in the protection against acute stress. The fact that these stress-related pathways are already induced before the induction of stress and eventually results in dampening of genotoxicity, implicates that DR is able to induce the right amount of stress to eventually protect against high levels of stress as induced by either IRI or chemotherapy. This hypothesis has been proposed before, and was named hormesis in which low doses of a certain stressor are able to protect against the toxic effects of a stressor in high doses 21 . In this case hormesis is activated via nutrient deprivation, in particular via reduction in proteins or essential amino acids. Even more so, in chapter 3 we showed that fasting has no effect on the antitumor effect of irinotecan as implied by previous findings. For instance, colleagues showed that fasted APC 15lox mutant mice given irinotecan had similar reduction in tumor size, and inhibited tumor cell proliferation as measured