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Dietary restriction (DR) and rapamycin extend healthspan and life span across multiple species. We have recently shown that DR in progeroid DNA repair-deficient mice dramatically extended healthspan and trippled life span. Here, we show that rapamycin, while significantly lowering mTOR signaling, failed to improve life span nor healthspan of DNA repair-deficient Ercc1∆/− mice, contrary to DR tested in parallel. Rapamycin interventions focusing on dosage, gender, and timing all were unable to alter life span. Even genetically modifying mTOR signaling failed to increase life span of DNA repair-deficient mice. The absence of effects by rapamycin on P53 in brain and transcription stress in liver is in sharp contrast with results obtained by DR, and appoints reducing DNA damage and transcription stress as an important mode of action of DR, lacking by rapamycin. Together, this indicates that mTOR inhibition does not mediate the beneficial effects of DR in progeroid mice, revealing that DR and rapamycin strongly differ in their modes of action.

RIVM rapport:Veel volwassenen (58 procent) gebruiken in het laatste uur voordat ze gaan slapen een tablet, smartphone of computer. Ze doen dit meerdere keren per week of dagelijks. Het gebruik van deze apparaten kort voor het slapen gaat gepaard met minder en slechter slapen. Of het blauwe licht van deze schermen hier de directe oorzaak van is, is niet bewezen. Mogelijk zijn ook andere factoren van invloed, zoals een hogere alertheid door het gebruik van deze apparaten. Ook is het nog niet bekend of hierdoor op de lange termijn andere gezondheidseffecten ontstaan. Licht regelt in belangrijke mate onze biologische klok. Als die wordt verstoord, kunnen mensen slechter of korter slapen. Slechts vier procent kijkt minder dan één keer in de week voor het slapengaan naar een beeldscherm. Vijf procent van de volwassenen doet dit nooit. Televisiekijken voor het slapengaan, hangt niet samen met veranderingen in slaap. Bij het spelen van spelletjes en het kijken van filmpjes wordt over het algemeen minder blauw licht uitgezonden dan bij activiteiten zoals e-mailen en het volgen van sociale media. Dit blijkt uit onderzoek van het RIVM in samenwerking met het Institute for Risk Assessment Sciences (IRAS), Universiteit Utrecht. Hiervoor is voor het eerst een relatief groot aantal volwassenen geraadpleegd over dit onderwerp (bijna 16.000). Vanwege het hoge aantal dat kort voor het slapengaan schermen gebruikt, wordt verder onderzoek aanbevolen naar de relatie met slaap en mogelijke gezondheidseffecten op de lange termijn., Many adults (58 percent) use a tablet, smartphone or computer within the last hour before going to bed at night. They do this several times a week or daily. The use of these devices shortly before bedtime is associated with less sleep and poor sleep quality. Whether the blue light from these screens is the direct cause of this has not been proven. There may be other factors that influence it, such as greater alertness from the use of these devices. It is also not yet known whether there will be other long-term health effects. Light significantly controls our biological clock. When it is disturbed, people may sleep poorlier or shorter. Only four percent look at a screen less than once a week before bedtime. Five percent of adults never do so. Watching television before going to sleep is not related to changes in sleep. When playing games and watching movies, generally less blue light is transmitted than during activities such as e-mailing and following social media. This is evident from research conducted by RIVM in collaboration with the Institute for Risk Assessment Sciences (IRAS), University of Utrecht. For the first time, a relatively large number of adults were consulted on this subject (almost 16,000). Due to the large group of people who use screens shortly before going to sleep, further research is recommended on the relationship with sleep and possible long-term health effects.

Introduction. Surgery-induced oxidative stress increases the risk of perioperative complications and delay in postoperative recovery. In mice, short-term preoperative dietary and protein restriction protect against oxidative stress. We investigated the feasibility of a calorie-and protein-restricted diet in two patient populations. Methods. In this pilot study, 30 live kidney donors and 38 morbidly obese patients awaiting surgery were randomized into three groups: a restricted diet group, who received a synthetic liquid diet with 30% fewer calories and 80% less protein for five consecutive days; a group who received a synthetic diet containing the daily energy requirements (DER); and a control group. Feasibility was assessed using self-reported discomfort, body weight changes, and metabolic parameters in blood samples. Results. Twenty patients (71%) complied with the restricted and 13 (65%) with the DER-diet. In total, 68% of the patients reported minor discomfort that resolved after normal eating resumed. The mean weight loss on the restricted diet was significantly greater (2.4 kg) than in the control group (0 kg, p = 0.002), but not in the DER-diet (1.5 kg). The restricted diet significantly reduced levels of serum urea and plasma prealbumin (PAB) and retinol binding protein (RBP). Conclusions. A short-term preoperative calorie-and protein-restricted diet is feasible in kidney donors and morbidly obese patients. Compliance is high and can be objectively measured via changes in urea, PAB, and RBP levels. These results demonstrate that this diet can be used to study the effects of dietary restriction on surgery-induced oxidative stress in a clinical setting.

RIVM rapport:Tablets, smartphones, led-tv, led-lampen en dergelijke geven relatief meer blauw licht af dan traditionelere lichtbronnen, zoals gloeilampen en traditionele computer- en televisieschermen. Er zijn aanwijzingen dat de biologische klok van mensen wordt beïnvloedt wanneer zij dergelijke apparaten 's avonds of 's nachts gebruiken. Dit effect lijkt zich voor te doen vanaf een blootstelling van enkele uren. Het is nog niet bekend in welke mate dat schadelijk is. Daarom wordt aanbevolen om te onderzoeken hoe lang en op welke momenten van de dag mensen zulke apparaten gebruiken. Ook dient te worden onderzocht welke effecten optreden, bij welke blootstelling, en in hoeverre ze schadelijk zijn voor de gezondheid, zowel op de korte als de lange termijn. Meer inzicht is nodig omdat het aantal producten dat relatief veel blauw licht uitzendt toeneemt en ze steeds meer worden gebruikt. Het effect van blauw licht op de biologische klok ontstaat doordat het dag-ennachtritme vooral gevoelig is voor licht in het blauwe deel van het spectrum. De biologische klok heeft een ritme van ongeveer één dag, het zogeheten circadiane ritme. Licht is hiervoor de belangrijkste 'tijdgever' en zorgt ervoor dat de biologische klok een ritme van ongeveer 24 uur houdt. Het circadiane systeem stuurt verschillende processen aan, waaronder de slaapwaak- cyclus, de afgifte van hormonen en de energiehuishouding. Het is al bekend dat een langdurige verstoring van het circadiane ritme (jet lag, nachtwerk) negatieve gezondheidseffecten kan veroorzaken, zoals overgewicht, hart- en vaatziekten en borstkanker. Of deze effecten een verband hebben met een blootstelling aan blauw licht is nog niet bewezen. Dit staat in een verkennende studie van het RIVM, in opdracht van de NVWA Hierin is op een rij gezet wat in de wetenschappelijke literatuur bekend is over blauw licht, de effecten daarvan op de gezondheid en welke kennishiaten er zijn., Tablets, smartphones, led-tv, led-lights and similar products emit more blue light compared to more traditional light sources, such as light bulbs and traditional pc and television screens. These devices appear to influence our biological clock when they are used during the evening or night. Effects on the biological clock have been observed after exposure of several hours. Currently, it is unknown if these effects are harmful. For these reasons, it is recommended that future studies investigate the time of day and duration that people use these devices. In addition, effects of these exposures need to be investigated, including the possible harmful health effects after short and long term use. Insight into these effects is required considering the expected increase in the availability and use of products that emit relatively large amounts of blue light. The effect of blue light on the biological clock is related to the sensitivity of our day-and- night rhythm for light of short wave-lengths (blue light) in particular. The biological clock has a rhythm of approximately one day, which is described as the circadian rhythm. Light is an important 'timekeeper' of this rhythm and maintains the rhythm of the biological clock at approximately 24 hours. The circadian system regulates a variety of processes, including the sleep-wake cycle, release of hormones and energy metabolism. It is known that long term disruption of the circadian rhythm (jet lag, night shift work) has negative health effects, including obesity, cardiovascular disease, and breast cancer. If exposure to blue light has related effects is currently unknown. This is described in an exploratory study of the National Institute for Public Health and the Environment (RIVM), in commission of the Netherlands Food and Consumer Product Safety Authority (NVWA). In this study, an overview of scientific literature is presented that describes blue light, its possible health effects, and important knowledge gaps are ident

Frequent shift work causes disruption of the circadian rhythm and might on the long-term result in increased health risk. Current biomarkers evaluating the presence of circadian rhythm disturbance (CRD), including melatonin, cortisol and body temperature, require 24hr ("around the clock") measurements, which is tedious. Therefore, these markers are not eligible to be used in large-scale (human) studies. The aim of the present study was to identify universal biomarkers for CRD independent of time of day using a transcriptomics approach. Female FVB mice were exposed to six shifts in a clockwise (CW) and counterclockwise (CCW) CRD protocol and sacrificed at baseline and after 1 shift, 6 shifts, 5 days recovery and 14 days recovery, respectively. At six time-points during the day, livers were collected for mRNA microarray analysis. Using a classification approach, we identified a set of biomarkers able to classify samples into either CRD or non-disrupted based on the hepatic gene expression. Furthermore, we identified differentially expressed genes 14 days after the last shift compared to baseline for both CRD protocols. Non-circadian genes differentially expressed upon both CW and CCW protocol were considered useful, universal markers for CRD. One candidate marker i. e. CD36 was evaluated in serum samples of the CRD animals versus controls. These biomarkers might be useful to measure CRD and can be used later on for monitoring the effectiveness of intervention strategies aiming to prevent or minimize chronic adverse health effects.

RIVM rapport:Op heel jonge leeftijd zijn proefdieren gevoeliger voor de schadelijke effecten van de stof benzo[a]pyreen dan op volwassen leeftijd. Er ontstaat in de jonge levensfase meer schade aan het DNA. Dit blijkt uit onderzoek van het RIVM. Normaal gesproken worden mogelijke schadelijke effecten van chemische stoffen in kaart gebracht door studies met volwassen proefdieren uit te voeren. Kinderen en volwassenen kunnen echter verschillen in de mate waarin ze gevoelig zijn voor chemische stoffen. Benzo[a]pyreen is een stof die voorkomt in voeding, zoals gebraden vlees, en in tabaksrook en uitlaatgassen. Het is een stof die veranderingen in het erfelijk materiaal kan veroorzaken en bij blootstellingen aan zeer hoge doses tot kanker kan leiden. Langdurige blootstelling op jonge leeftijd leidde niet tot een hoger aantal dieren met tumoren, maar wel tot tumoren in andere organen dan bij volwassen dieren. Momenteel wordt in aanvullende studies onderzocht of deze resultaten specifiek zijn voor benzo[a]pyreen of kenmerkend zijn voor DNAbeschadigende stoffen in het algemeen. Dat onderzoek zal uitwijzen of bij de risicobeoordeling van chemische stoffen meer rekening moeten worden gehouden met kinderen als risicogroep., Experimental animals are more sensitive at a young age to the adverse effects induced upon benzo[a]pyrene exposure compared to adult animals. In research performed at the RIVM, higher amounts of DNA damage were observed when benzo[a]pyrene was given to experimental animals at a young age. Usually, potential adverse human health effects of environmental chemicals are evaluated in toxicity studies using adult laboratory animals. Children and adults, however, may differ in sensitivity to these adverse effects. Benzo[a]pyrene is commonly found in grilled and broiled foods, tobacco smoke and automobile exhaust fumes. Exposure to benzo[a]pyrene may cause DNA damage, and, at very high doses, even induce tumors. Chronic exposure of animals at a very young age did not increase the incidence of tumors, but tumors were primarily found in a different organ compared to animals exposed at an adult age. Further research is ongoing to assess whether these findings are specific for benzo[a]pyrene, or representative for genotoxicants in general. Those studies will provide an insight as to what extent children should be regarded as a high-risk group in human health risk assessment.

RIVM rapport:Jonge dieren lijken niet gevoeliger dan volwassen dieren voor schadelijke effecten van DNA-beschadigende stoffen Op heel jonge leeftijd kunnen proefdieren gevoeliger zijn voor de schadelijke effecten van chemische stoffen dan op volwassen leeftijd. Sommige stoffen veroorzaken in de jonge levensfase meer schade aan het DNA, maar dat is niet altijd het geval. Een hogere gevoeligheid op jonge leeftijd lijkt af te hangen van de manier waarop de stof het erfelijk materiaal beschadigt. Dit blijkt uit onderzoek van het RIVM. Chemische stoffen kunnen op verschillende manieren veranderingen aan het erfelijk materiaal veroorzaken. Normaal gesproken worden mogelijke schadelijke effecten van chemische stoffen in kaart gebracht door studies met volwassen proefdieren uit te voeren. Kinderen en volwassenen kunnen echter verschillen in de mate waarin ze gevoelig zijn voor chemische stoffen. Eerder was een hogere gevoeligheid voor schadelijke effecten bij jonge proefdieren waargenomen in onderzoek van het RIVM naar benzo[a]pyreen. Deze stof, die voorkomt in voeding, zoals gebraden vlees, en in tabaksrook en uitlaatgassen, geeft meer schadelijke effecten op jonge leeftijd dan op volwassen leeftijd. Nadien is dit effect getoetst voor drie andere stoffen. Bij stoffen die op een andere manier DNA beschadigen, heeft leeftijd veel minder invloed. Bij de risicobeoordeling van chemische stoffen zal per stof beoordeeld moeten worden of de standaard veiligheidsfactor voldoende is of aanpassing behoeft., Young animals do not appear to be more susceptible than adult animals to mutagenic effects of environmental chemicals Experimental animals can be more susceptible at a young age to the adverse effects induced upon exposure to environmental chemicals in comparison to adult animals. In research performed at the RIVM, the susceptibility to DNA mutations in young and adult animals was investigated for a selected set of substances. The results from this research suggest that increased susceptibility to mutagenic effects is dependent on the specific mechanism of action. Environmental chemicals may invoke DNA mutations through a variety of mechanisms. Usually, potential adverse human health effects of environmental chemicals are evaluated in toxicity studies using adult laboratory animals. Children and adults, however, may differ in sensitivity to these adverse effects. In previous research we found that exposure to benzo[a]pyrene, a chemical commonly found in grilled and broiled foods, tobacco smoke and automobile exhaust fumes, induced DNA mutations at a higher frequency in animals exposed at a young age in comparison to animals exposed at adult age. For the other three chemicals investigated in the present study we did not find any agerelated differences in genotoxicity. These findings suggest that increased susceptibility to mutagenic effects is dependent on the specific mechanism of action, which then is to be taken into account in chemical risk assessments of children.

RIVM rapport:De laatste jaren groeit de zorg dat langdurig werken in ploegendiensten gezondheidsrisico's kan veroorzaken, waaronder kanker, overgewicht en harten vaatziekten. Het merendeel van de wetenschappelijke literatuur toont aan dat werken in ploegendienst ook tot een verhoogd risico op metabole gezondheidseffecten leidt, zoals een toename van lichaamsgewicht of een verstoorde suiker- en vethuishouding; een substantieel deel laat dit verband echter niet zien. Dit blijkt uit een literatuuroverzicht van het RIVM, dat in opdracht van het ministerie van Sociale Zaken en Werkgelegenheid (SZW) is uitgevoerd. De discrepantie tussen de onderzoeksresultaten wordt waarschijnlijk veroorzaakt door een grote verscheidenheid in de opzet ervan. Zo kan het type van de onderzochte ploegendienst variëren (zoals avond- of, nachtdienst, of wisselende diensten), net als de studiepopulatie (geslacht, beroep en dergelijke). Ook is in sommige onderzoeken niet gekeken naar leefstijlfactoren die de relatie tussen nachtwerk en metabole factoren kunnen beïnvloeden (roken, alcoholgebruik, bewegen en voeding). Het moment waarop wordt gegeten lijkt van invloed op veranderingen in metabole functies als gevolg van werken in ploegendienst. De relatie tussen mogelijke effecten op metabole risicofactoren had tot nu toe minder aandacht in wetenschappelijk onderzoek naar de effecten van werken in ploegendienst. Meer inzicht hierin is belangrijk, aangezien metabole risicofactoren bijdragen aan het ontstaan van diverse chronische aandoeningen, zoals hart- en vaatzieken en diabetes type II. In Nederland werkt 16 procent van de beroepsbevolking soms of regelmatig in de nachtdienst (1,2 miljoen mensen) en 51 procent geeft aan soms of regelmatig tijdens de avonduren te werken (3,8 miljoen mensen). Aanbevolen wordt onder andere om in onderzoek specifieke aandacht te hebben voor de invloed van de leefstijlfactoren bewegen en eten, roosterkenmerken en individuele variaties in dag- en nachtritmes. Deze factoren bieden a, The past few years there has been an increasing concern for the health risk effects of shiftwork (including night work), e.g. risks for cancer, obesity and cardiovascular diseases. A large amount of scientific studies identified an increased risk for metabolic health effects caused by shiftwork, such as increased bodyweight or a disturbed glucose and lipid homeostasis. However, there is also a substantial number of studies that have not observed this relation. These are results from a literature review by the RIVM, in commission of the Ministry of Social Affairs and Employment. An explanation for these contradicting results may be the large heterogeneity in type of shift work investigated (nightshifts, evening shifts or rotating shifts), study population, and factors that can influence this relation (smoking, alcohol intake, activity, and food intake). The moment of food intake appears to have an important role. The relation between shift work and metabolic risk factors has received less attention in contrast to other health effects such as an increased risk for cancer. However, a better understanding of this relation is important, considering the significance of these factors in the development of chronic metabolic disorders, such as cardiovascular disease and diabetes type II. In the Netherlands, 16 percent (1.2 million people) of the working population works regularly or occasionally during the night and 51 percent indicates to sometimes or regularly work during the evening (3.8 million people). Recommendations of this report include research with specific attention for the influence of lifestyle factors, type of shift work en individual differences in circadian rhythms. These aspects are potential targets for future interventions to minimize the health risk effects.

Ku70 and Ku80 form a heterodimer called Ku that forms a holoenzyme with DNA dependent-protein kinase catalytic subunit (DNA-PKCS) to repair DNA double strand breaks (DSBs) through the nonhomologous end joining (NHEJ) pathway. As expected mutating these genes in mice caused a similar DSB repair-defective phenotype. However, ku70-/- cells and ku80 -/- cells also appeared to have a defect in base excision repair (BER). BER corrects base lesions, apurinic/apyrimidinic (AP) sites and single stand breaks (SSBs) utilizing a variety of proteins including glycosylases, AP endonuclease 1 (APE1) and DNA Polymerase β (Pol β). In addition, deleting Ku70 was not equivalent to deleting Ku80 in cells and mice. Therefore, we hypothesized that free Ku70 (not bound to Ku80) and/or free Ku80 (not bound to Ku70) possessed activity that influenced BER. To further test this hypothesis we performed two general sets of experiments. The first set showed that deleting either Ku70 or Ku80 caused an NHEJ-independent defect. We found ku80-/- mice had a shorter life span than dna-pkcs-/- mice demonstrating a phenotype that was greater than deleting the holoenzyme. We also found Ku70-deletion induced a p53 response that reduced the level of small mutations in the brain suggesting defective BER. We further confirmed that Ku80-deletion impaired BER via a mechanism that was not epistatic to Pol β. The second set of experiments showed that free Ku70 and free Ku80 could influence BER. We observed that deletion of either Ku70 or Ku80, but not both, increased sensitivity of cells to CRT0044876 (CRT), an agent that interferes with APE1. In addition, free Ku70 and free Ku80 bound to AP sites and in the case of Ku70 inhibited APE1 activity. These observations support a novel role for free Ku70 and free Ku80 in altering BER. © 2014 Choi et al.

Ku80 and DNA-PKCS are both involved in the repair of double strand DNA breaks via the nonhomologous end joining (NHEJ) pathway. While ku80(-/-) mice exhibit a severely reduced lifespan and size, this phenotype is less pronounced in dna-pk(cs)(-/-) mice. However, these observations are based on independent studies with varying genetic backgrounds. Here, we generated ku80(-/-), dna-pk(cs)(-/-) and double knock out mice in a C57Bl6/J*FVB F1 hybrid background and compared their lifespan, end of life pathology and mutation frequency in liver and spleen using a lacZ reporter. Our data confirm that inactivation of Ku80 and DNA-PKCS causes reduced lifespan and bodyweights, which is most severe in ku80(-/-) mice. All mutant mice exhibited a strong increase in lymphoma incidence as well as other aging-related pathology (skin epidermal and adnexal atrophy, trabacular bone reduction, kidney tubular anisokaryosis, and cortical and medullar atrophy) and severe lymphoid depletion. LacZ mutation frequency analysis did not show strong differences in mutation frequencies between knock out and wild type mice. The ku80(-/-) mice had the most severe phenotype and the Ku80-mutation was dominant over the DNA-PKCS-mutation. Presumably, the more severe degenerative effect of Ku80 inactivation on lifespan compared to DNA-PKCS inactivation is caused by additional functions of Ku80 or activity of free Ku70 since both Ku80 and DNA-PKCS are essential for NHEJ.

Ischemia-reperfusion injury (IRI) is inevitable during kidney transplantation leading to oxidative stress and inflammation. We previously reported that preoperative fasting in young-lean male mice protects against IRI. Since patients are generally of older age with morbidities possibly leading to a different response to fasting, we investigated the effects of preoperative fasting on renal IRI in aged-overweight male and female mice. Male and female F1-FVB/C57BL6-hybrid mice, average age 73 weeks weighing 47.2 grams, were randomized to preoperative ad libitum feeding or 3 days fasting, followed by renal IRI. Body weight, kidney function and survival of the animals were monitored until day 28 postoperatively. Kidney histopathology was scored for all animals and gene expression profiles after fasting were analyzed in kidneys of young and aged male mice. Preoperative fasting significantly improved survival after renal IRI in both sexes compared with normal fed mice. Fasted groups had a better kidney function shown by lower serum urea levels after renal IRI. Histopathology showed less acute tubular necrosis and more regeneration in kidneys from fasted mice. A mRNA analysis indicated the involvement of metabolic processes including fatty acid oxidation and retinol metabolism, and the NRF2-mediated stress response. Similar to young-lean, healthy male mice, preoperative fasting protects against renal IRI in aged-overweight mice of both genders. These findings suggest a general protective response of fasting against renal IRI regardless of age, gender, body weight and genetic background. Therefore, fasting could be a non-invasive intervention inducing increased oxidative stress resistance in older and overweight patients as well.

Aging and age-related pathology is a result of a still incompletely understood intricate web of molecular and cellular processes. We present a C57BL/6J female mice in vivo aging study of five organs (liver, kidney, spleen, lung, and brain), in which we compare genome-wide gene expression profiles during chronological aging with pathological changes throughout the entire murine life span (13, 26, 52, 78, 104, and 130 weeks). Relating gene expression changes to chronological aging revealed many differentially expressed genes (DEGs), and altered gene sets (AGSs) were found in most organs, indicative of intraorgan generic aging processes. However, only ≤ 1% of these DEGs are found in all organs. For each organ, at least one of 18 tested pathological parameters showed a good age-predictive value, albeit with much inter- and intraindividual (organ) variation. Relating gene expression changes to pathology-related aging revealed correlated genes and gene sets, which made it possible to characterize the difference between biological and chronological aging. In liver, kidney, and brain, a limited number of overlapping pathology-related AGSs were found. Immune responses appeared to be common, yet the changes were specific in most organs. Furthermore, changes were observed in energy homeostasis, reactive oxygen species, cell cycle, cell motility, and DNA damage. Comparison of chronological and pathology-related AGSs revealed substantial overlap and interesting differences. For example, the presence of immune processes in liver pathology-related AGSs that were not detected in chronological aging. The many cellular processes that are only found employing aging-related pathology could provide important new insights into the progress of aging.