Franny Jongbloed

244 CHAPTER 9 accompanied by a reduction of acute tubular necrosis and more tubular regeneration. These effects were not as robust as in young-lean mice, yet sufficient enough to induce protection. In addition, we showed that female mice were able to withstand longer periods of IRI than male mice as an indication that the female gender has a higher endogenous threshold against IRI. Therefore, these data concluded that the presence of comorbidities as in age and obesity are not able to annul the robust beneficial effects of fasting on the outcome after renal IRI. Although three days of fasting is able to induce its beneficial effects in different mouse models, it does not give insight into which dietary components are precisely responsible for the effects of acute stress. After the first results of calorie restriction and its effects on health span and lifespan, many researchers examined whether the calorie reduction per se, or a specific component, was responsible for these effects. The first studies focused on the macronutrients, being protein, carbohydrates and fat 2-5 . Later the ratio of macronutrients became a subject of interest as well 6,7 . The first evidence of a major role of protein restriction was obtained in fruit flies where long-term protein-restriction resulted in increased lifespan 8-11 . In mice, the translation of long-term protein restriction to short-term regimens found that six days of deprivation was sufficient to induce beneficial effects against multiple forms of IRI 12 . A major issue with protein restriction proved to be the voluntary calorie restriction that mice imposed during the dietary intervention, which made the effects of protein restriction still indistinguishable from calorie restriction 12 . In chapter 4 , we are the first to show the beneficial effects of protein deprivation for only three days, resulting in significant improved mortality and kidney function after renal IRI. In addition, three days of 30% DR did not induce protection and therefore we could distinguish the effects of protein restriction from calorie restriction. Recent literature also implicated the roles of other macronutrients, in particular in the light of aging and health span 13,14 . Solon-Biet et al. showed that the ratio of protein to carbohydrates is responsible for the benefits, in which a low amount of protein and high amount of carbohydrates revealed the largest effect on both health span and lifespan whereas reduction of carbohydrates gave an opposite image 15 . Several studies followed and pointed towards the same direction 6,7 . We could also confirm the detrimental effects of carbohydrate reduction, since our carbohydrate-free diet resulted in an aggravation of mortality and kidney function decline compared to control mice fed AL. In comparison, a fat-free diet showed results similar to the control mice fed AL, and was implicated to have no major role in the effects of calorie restriction on renal IRI. Indeed, Solon-Biet et al. came to the same conclusion in the long-term dietary interventions, revealing that fat intake did not contribute to health span and lifespan changes in mice 6 . The authors also pointed towards the role of amino acids, in particular branched chain amino acids, in the effects of protein restriction on aging 6,15 . In chapter 5 , we also narrowed down the role of protein restriction by providing mice with a 3-day diet lacking one of