Dietary restriction (DR) extends lifespan and increases resistance to multiple forms of stress, including ischemia reperfusion injury to the brain and heart in rodents. While maximal effects on lifespan require long-term restriction, the kinetics of onset of benefits against acute stress are not known. Here we show that 2–4 weeks of 30% dietary restriction improved survival and kidney function following renal ischemia reperfusion injury in mice. Brief periods of water-only fasting were similarly effective at protecting against ischemic damage. Significant protection occurred within one day, persisted for several days beyond the fasting period and extended to another organ, the liver. Protection by both short-term DR and fasting correlated with improved insulin sensitivity, increased expression of markers of antioxidant defense and reduced expression of markers of inflammation and insulin/insulin-like growth factor-1 signaling. Unbiased transcriptional profiling of kidney from mice subject to short-term DR or fasting revealed a significant enrichment of signature genes of long-term DR. These data demonstrate that brief periods of reduced food intake, including short-term daily restriction and fasting, can increase resistance to ischemia reperfusion injury in rodents and suggest a rapid onset of benefits of DR in mammals.

Our laboratory has reported that glucose is essential for glycolytic enzyme induction and proliferation of mitogen-activated rat thymocytes (41). Here we show that: 1) Resting thymocytes meet their ATP demand mainly by oxidative glucose breakdown (88%), whereas proliferating thymocytes produce 86% by glycolytic degradation of glucose to lactate and only 14% by oxidation to CO2 and water. 2) In contrast to nonstimulated resting thymocytes, production of PMA primed reactive oxygen species (ROS) in the proliferating cells is nearly abolished. 3) Consistent with this finding, no ROS formation is observed in proliferating human promyelocytic HL-60 cells, whereas differentiated, nonproliferating HL-60 cells exert a marked response upon priming with PMA. 4) The observed reduction of ROS formation by resting thymocytes incubated with pyruvate suggests a function of pyruvate as an H(2)O(2) scavenger. 5) The respiratory chain is a potential origin for ROS because inhibitors of the mitochondrial electron transport strongly reduce H(2)O(2) production by resting thymocytes. The results are discussed in the context of aerobic glycolysis by proliferating cells being a means to minimize oxidative stress during the phases of the cell cycle where maximally enhanced biosynthesis and cell division do occur.

Peroxisome proliferator-activated receptor δ (PPARδ) is a critical regulator of energy metabolism in the heart. Here, we propose a mechanism that integrates two deleterious characteristics of heart failure, hypoxia and a metabolic shift toward glycolysis, involving the microRNA cluster miR-199a∼214 and PPARδ. We demonstrate that under hemodynamic stress, cardiac hypoxia activates DNM3os, a noncoding transcript that harbors the microRNA cluster miR-199a∼214, which shares PPARδ as common target. To address the significance of miR-199a∼214 induction and concomitant PPARδ repression, we performed antagomir-based silencing of both microRNAs and subjected mice to biomechanical stress to induce heart failure. Remarkably, antagomir-treated animals displayed improved cardiac function and restored mitochondrial fatty acid oxidation. Taken together, our data suggest a mechanism whereby miR-199a∼214 actively represses cardiac PPARδ expression, facilitating a metabolic shift from predominant reliance on fatty acid utilization in the healthy myocardium toward increased reliance on glucose metabolism at the onset of heart failure.

Energy metabolism in proliferating cultured rat thymocytes was compared with that of freshly prepared non-proliferating resting cells. Cultured rat thymocytes enter a proliferative cycle after stimulation by concanavalin A and Lymphocult T (interleukin-2), with maximal rates of DNA synthesis at 60 h. Compared with incubated resting thymocytes, glucose metabolism by incubated proliferating thymocytes was 53-fold increased; 90% of the amount of glucose utilized was converted into lactate, whereas resting cells metabolized only 56% to lactate. However, the latter oxidized 27% of glucose to CO2, as opposed to 1.1% by the proliferating cells. Activities of hexokinase, 6-phosphofructokinase, pyruvate kinase and aldolase in proliferating thymocytes were increased 12-, 17-, 30- and 24-fold respectively, whereas the rate of pyruvate oxidation was enhanced only 3-fold. The relatively low capacity of pyruvate degradation in proliferating thymocytes might be the reason for almost complete conversion of glucose into lactate by these cells. Glutamine utilization by rat thymocytes was 8-fold increased during proliferation. The major end products of glutamine metabolism are glutamate, aspartate, CO2 and ammonia. A complete recovery of glutamine carbon and nitrogen in the products was obtained. The amount of glutamate formed by phosphate-dependent glutaminase which entered the citric acid cycle was enhanced 5-fold in the proliferating cells: 76% was converted into 2-oxoglutarate by aspartate aminotransferase, present in high activity, and the remaining 24% by glutamate dehydrogenase. With resting cells the same percentages were obtained (75 and 25). Maximal activities of glutaminase, glutamate dehydrogenase and aspartate aminotransferase were increased 3-, 12- and 6-fold respectively in proliferating cells; 32% of the glutamate metabolized in the citric acid cycle was recovered in CO2 and 61% in aspartate. In resting cells this proportion was 41% and 59% and in mitogen-stimulated cells 39% and 65% respectively. Addition of glucose (4 mM) or malate (2 mM) strongly decreased the rates of glutamine utilization and glutamate conversion into 2-oxoglutarate by proliferating thymocytes and also affected the pathways of further glutamate metabolism. Addition of 2 mM-pyruvate did not alter the rate of glutamine utilization by proliferating thymocytes, but decreased the rate of metabolism beyond the stage of glutamate significantly. Formation of acetyl-CoA in the presence of pyruvate might explain the relatively enhanced oxidation of glutamate to CO2 (56%) by proliferating thymocytes.

Photolyase transgenic mice have opened new avenues to improve our understanding of the cytotoxic effects of ultraviolet (UV) light on skin by providing a means to selectively remove either cyclobutane pyrimidine dimers (CPDs) or pyrimidine (6-4) pyrimidone photoproducts. Here, we have taken a genomics approach to delineate pathways through which CPDs might contribute to the harmful effects of UV exposure. We show that CPDs, rather than other DNA lesions or damaged macromolecules, comprise the principal mediator of the cellular transcriptional response to UV. The most prominent pathway induced by CPDs is that associated with DNA double-strand break (DSB) signalling and repair. Moreover, we show that CPDs provoke accumulation of c-H2AX, P53bp1 and Rad51 foci as well as an increase in the amount of DSBs, which coincides with accumulation of cells in S phase. Thus, conversion of unrepaired CPD lesions into DNA breaks during DNA replication may comprise one of the principal instigators of UV-mediated cytotoxicity.