Franny Jongbloed

35 2 FASTING PROTECTS AGED-OVERWEIGHT MICE seen in young animals. Alternatively, or in addition, it could be that fasting of overweight mice triggers less extreme responses because of the presence of larger reserves. Taken together, the survival and kidney function data as well as the gene expression findings suggest that fasting has a more robust effect in young mice, although in aged mice it is still sufficient to induce a protective effect against IRI. Although ECD are getting more and more accepted as an alternative for optimal donors, the outcome after transplantation of an ECD kidney is worse, with higher risks of DGF, acute rejection and graft failure 25 . Since age is a factor that cannot be influenced, reducing the consequences of ischemia is all the more warranted. To elucidate the mechanisms responsible for the protective effect induced by fasting, we performed microarray analysis after fasting. Our analysis revealed several interesting pathways regulated in kidneys from both young-lean and aged-overweight male mice, thereby being independent of age, weight and genetic background. After clustering of the pathways according to their known or predicted biological function, three biological processes stood out, namely: retinol biosynthesis, fatty acid oxidation and the stress response pathway Nrf2 (see Tables 1 and 2). Upregulation of the retinoid X receptors (RXRs) together with retinol biosynthesis was responsible for over a quarter of all overrepresented pathways. RXRs are able to bind with multiple factors like the peroxisome proliferator-activated receptors (PPARs). Depending on the different complexes made, RXRs are involved in many processes. They are able to reduce inflammatory processes and contribute to the reduction of the acute phase response, an inflammatory response occurring after invasive surgery resembling renal IRI 26 . Activation of RXR also stimulates fatty acid oxidation and transport and is thus partially responsible for the enrichment in fatty acid pathways found after short-term fasting, leading to reduced fat synthesis and storage and increased oxidation in order to burn fat for the production of energy as part of a redesign of metabolic processes upon lack of food 27 . Nrf2 and genes involved in the Nrf2 -pathway were found to be significantly upregulated in kidneys of both young and aged mice after three days of fasting, although less pronounced in the aged cohort. Nrf2 , nuclear factor-erythroid 2 p- 45 related factor 2, is known to have both anti-oxidant and anti-inflammatory features and, more specific to IRI, a deficiency of Nrf2 leads to aggravation of ischemic injury in mice 28-30 . Previously, we showed that a downstream product of Nrf2 , heme oxygenase 1 (HO-1), is markedly increased by 3-day fasting in young mice 9 . Together with the pronounced Nrf2 -mediated oxidative stress response pathway found in the current analysis, Nrf2 is likely involved in the stress resistance phenotype after DR in both young-lean and aged-overweight mice. It was recently shown that a Nrf2 activator, CDDO-imidazolide, is able to improve survival after a renal IRI model in mice, inducing similar outcome found with fasting in our mouse model 31 . Whether Nrf2 induction would lead to the same beneficial effects induced by DR and would be safe and feasibly in the clinic, is to be elucidated.