Angelo Turturro

Effect of chronic caloric restriction on physiological variables related to energy metabolism in the male Fischer 344 rat

In the present study, a number of physiological and behavioral measures that are related to metabolism were continuously monitored in 19-month-old male Fischer 344 rats that were fed ad libitum or fed a caloric restricted diet. Caloric restricted rats ate fewer meals but consumed more food during each meal and spent more time eating per meal than did rats fed ad libitum. Therefore, the timing and duration of meals as well as the total number of calories consumed may be associated with life extension. Average body temperature per day was significantly lower in restricted rats but body temperature range per day and motor activity were higher in restricted rats. Dramatic changes in respiratory quotient, indicating rapid changes in metabolic pathway and lower temperature, occurred in caloric restricted rats when carbohydrate reserves were depleted. Lower body temperature and metabolism during this time interval may result in less DNA damage, thereby increasing the survival potential of restricted rats. Nighttime feeding was found to synchronize physiological performance between ad libitum and caloric restricted rats better than daytime feeding, thereby allowing investigators to distinguish the effects of caloric restriction from those related solely to the time-of-day of feeding.

Longevity, Body Weight, and Neoplasia in Ad Libitum-Fed and Diet-Restricted C57BL6 Mice Fed NIH-31 Open Formula Diet

Groups of C57BL6 mice of each sex were assigned to one of 2 dietary regimens, ad libitum (AL) or dietary restriction (DR), to study effects of food restriction on body weight, survival, and neoplasia. The AL and DR groups were subdivided into a scheduled sacrifice group for examination at 6-mo intervals, and a lifetime group to provide longevity data. Necropsies and microscopic examinations were conducted on 911 animals. In the lifetime group food consumption averaged 33.6 and 34.4 g per week by AL males and AL females, respectively; the DR counterparts were given 40% less. The diet contained 4.35 kcal/g. The average lifetime body weights were 34.8, 26.8, 22.6, and 21.6 g for AL males, AL females, DR males, and DR females, respectively, and their age at 50% survival was 27.5, 26.9, 31.7, and 33.5 mo. Maximal lifespan was increased 18% in DR males and females. Lifetime incidence of tumor-bearing mice was 89% and 86% for AL males and females, versus 64% for each sex of DR mice. Dramatic reduction occurred in female DR mice in lymphoma (9% vs 29%), pituitary neoplasms (1% vs 37%), and thyroid neoplasms (0.4% vs 8%). In males, hepatocellular tumors were reduced to 1% from 10% by DR. In contrast, the incidence of histiocytic sarcoma was increased in DR females and unaffected in DR males. Tumor onset was delayed in DR animals; 87% of all neoplasms in males and 95% in females had occurred in the AL mice by 24 mo, whereas the DR animals had only 52% and 39% of their lifetime incidence, respectively, by that age. This study provided comparative AL and DR data from C57BL6 mice examined randomly at 6-mo intervals (cross-sectional group) in parallel with data from animals in similar cohort that was unsampled and allowed to succumb naturally (longevity group). Dietary restriction reduced the lifetime percentage of tumor-bearing animals and the number of tumors per animal, and delayed the age at onset of most neoplasms.

Effect of chronic caloric restriction on hepatic enzymes of intermediary metabolism in the male Fischer 344 rat

It is well established that caloric restriction extends life span and significantly retards the rate of occurrence of most age-associated degenerative disease processes. A paucity of data exists relative to the mechanisms by which caloric restriction accomplishes these events. We have examined the effect of caloric restriction in rats on several hepatic enzymes of intermediary metabolism. The activities of glycolytic and supporting enzymes including lactate dehydrogenase, pyruvate kinase, sorbitol dehydrogenase, and alcohol dehydrogenase were all decreased in response to caloric restriction. Fructose 1-phosphate aldolase and creatine phosphokinase were not altered. Likewise, enzymes associated with lipid metabolism (malic enzyme and glycerokinase) were reduced (fatty acid synthetase was reduced, but not to a statistically significant degree). Activities of enzymes supporting gluconeogenesis (glutamate oxaloacetate transaminase, tyrosine aminotransferase, glutamate pyruvate transaminase, glutamate dehydrogenase, amino acid oxidase, malate dehydrogenase, and glucose 6-phosphatase) were either unchanged or increased significantly by caloric restriction. Glucagon levels were decreased. Comparisons between young ad libitum fed and older calorically restricted rats revealed similar but not identical metabolic activity. These results suggest that caloric restriction produces an effect on intermediary metabolism, favoring the role of glucagon and glucose synthesis; but limiting the role of insulin and glucose catabolism in the liver. The former observation provides for the efficient support of peripheral tissues and the latter a level of energy production necessary only for self maintenance. Limited lipid metabolism suggests decreased potential for fatty acid epoxide formation and free radical damage to cellular macromolecules. Additionally, caloric restriction may delay the progressive age associated changes in the activities of some of the enzymes investigated.

The influence of dietary restriction on DNA repair in rodents: A preliminary study

A range study was undertaken to determine if dietary restriction (DR) affects DNA repair in rodents. Unscheduled DNA synthesis (UDS) was examined in two strains of rat (Brown Norway, BN and Brown Norway X Fischer 344 F1 hybrid, BNF) at 18 months of age. O(6)-Methylguanine-acceptor protein activity (MGAP) was measured across species using rat (Brown Norway X Fischer F-344 F1 hybrid, 18 months) and mouse (B6CB F1 hybrid, 30 months). The rodents were maintained on either an ad libitum (AL) or a restricted diet (60% of the caloric intake of AL rodents). UDS increased approximately 48-65% in freshly isolated skin cells from DR animals opposed to their AL controls after challenge with ultraviolet light (254 nm, 20 J/m2 UV). After treatment with methylmethane sulfonate (0.5 mM MMS), a significant increase in UDS was observed (P less than 0.01, approx, 55% for BN and 52% for BNF rats). Results of measurements for MGAP activity found levels to increase 73% in DR rats and approximately 28% in DR mice when compared to their AL counterparts. In addition MGAP levels in phase shifted mice were examined at three time points during a 24-h period where significant changes were found to occur in the metabolism of DR rodents. The activity of MGAP changed in a circadian fashion with significant increases in MGAP activity in DR mice occurring during the period of highest metabolic activity.

Survival, body weight, and spontaneous neoplasms in ad Libitum-fed and food-restricted Fischer-344 rats.

Ad libitum- fed (AL) and food-restricted (FR) Fischer-344 male and female rats were monitored for survival, body weight, and spontaneous neoplasms. Mean and maximal lifespans for each group were inversely related to mean body weights. AL males were the shortest lived (mean lifespan 101 wk) followed by AL females (118 wk), FR males (125 wk), and FR females (132 wk). Gross and microscopic examinations were performed on 851 rats from cross-sectional and longevity components of the study. In FR groups, the incidence of mammary gland fibroadenomas, testicular interstitial cell tumors, and pituitary neoplasms was decreased while the latency of these neoplasms was increased. In longevity components, most FR groups had a higher incidence of leukemia than AL cohorts, but all FR groups had a higher mean age at death for the rats with leukemia. Higher leukemia rate in the FR groups was thought to be a result of their extended mean lifespan.

Age-Related Neoplasia in a Lifetime Study of Ad Libitum-Fed and Food-Restricted B6C3F1 Mice

Longevity, body weight, and age-specific neoplasia were determined in 1,064 B6C3F1 mice as part of a coordinated study of food restriction (FR). Restricted animals were offered 60% of the diet consumed by the ad libitum (AL) group. Longevity data were derived from a set of 56 animals of each sex from each diet group, which were examined whenever dead or moribund. For cross-sectional data, a parallel set of 210 animals were sacrificed in groups of 12-15 at 6-mo intervals. Lifetime body weight was reduced in the FR mice approximately proportional to restriction (i.e., 40%). Food restriction increased the age at 50% survival (median) by 36% in both sexes and increased the maximal lifespan (mean age of oldest 10%) by 21.5% in males and by 32.5% in females. In 56 males of the longevity groups, there were 89 neoplasms in the AL subgroup versus 53 in FR; 56 AL females had 100, versus 58 in 55 FR females. Increase in lifespan of the restricted animals was achieved primarily by decrease in incidence and delay of onset of fatal tumors, of which lymphoma was the most prominent. This report catalogs all of the neoplasms (1,103) observed in longevity and cross-sectional groups, by diet, sex, and age. These data add to the existing knowledge base needed for future studies of dietary restriction and aging as well to evaluate nutrition of animals used in bioassays.

Mechanisms of Caloric Restriction Affecting Aging and Diseasea

Caloric restriction (CR) appears to affect aging by the inhibition of the specific chronic diseases which occur at increasing frequency with age. A common disease in F-344 rats, granulocytic leukemia, appears to have a window where it is sensitive to the effects of CR. Other diseases, such as pituitary adenomas, appear to have a different relationship to growth in the animal. Additionally, a model for the major disease for a number of long-lived strains of mice, lymphoma, which CR effects by inhibiting the expression of the causative agent, is being developed. Evaluation of the effects of CR on neoplasia, degenerative disease and physiological parameters suggests that the major factors in expression of these diseases is the alteration of growth factors, hormonal status, etc., and that these alterations also affect strain-specific pathologies depending on when they are changed in the life span. Effecting different diseases at different times in the life span, long-term CR, by limiting exposure to endogenous growth factors, altering physiological characteristics, and limiting exposure to food toxicants, inhibits the onset of disease, and its sequela, aging.

Caloric Restriction and Toxicity

The modulatory effects of caloric intake on the rate and extent of both spontaneous and induced disease incidence is well known, but the significance of these effects in the interpretation of testing data has only recently become appreciated. This is especially true relative to the impact of caloric intake on both survival and background incidence for common tumors. In order to enhance the health and survival of animals ongoing chronic toxicity testing it has been suggested that such tests should restrict food consumption. Although this restriction will result in increasing survival of the test animals, it may also effect the expression of toxicity by altering agent metabolism and disease progression. Focus in this symposium is on the necessity to control dietary consumption in toxicity tests (dietary control), and if such a need does exist to what level of consumption should be diet be focused (caloric restriction).

Role of Glucocorticoids and “Caloric Stress” in Modulating the Effects of Caloric Restriction in Rodentsa

Caloric restriction (i.e., the balanced reduction of the protein, carbohydrate and fat content of the diet without reduction of its micronutrient content) has been shown, in rodents, to be an extremely powerful modulator of a broad spectrum of age-associated degenerative diseases as well as life span.’-5 Caloric restriction has been found to delay the occurrence of many age-associated neoplastic diseases or to slow their progression, often to such an extent that the clinical expression of the diseases is eliminated.* Since reducing individual dietary components such as protein or fat without reducing the overall caloric intake is less effective in increasing longevity or suppressing neoplasia than caloric restriction by itself, it is reasonable to assume that the observed effects of caloric restriction are primarily dependent upon a specific reduction in calories and not specific dietary compon e n t ~ . ~ ~ ’ The multiple actions of caloric restriction suggest that it may influence primary aging processes themselves; however, recent examination of pathological data suggests that caloric restriction delays different age-dependent pathological conditions at different rates.8 This observation would argue against the modulation by caloric restriction of a simple fundamental process of cellular aging, but rather, it implies that caloric restriction modifies the organism’s homeostatic set-point to one that is less susceptible to some but not all pathologies. Unfortunately, such theories on the nature of the caloric restriction effect will remain merely speculatory until the precise molecular and biochemical mechanisms by which caloric restriction influences the processes of aging and disease are fully elucidated.

The EFF the effect of dietary restriction of MYC protooncogene expression in mice: A preliminary study

Abstract The effect of chronic dietary restriction on the expression of the c -myc protooncogene was determined in the livers of a hybrid mouse strain (C57B16 × C3H F1 hybrid) at three time points during a 24-h period: 1 h after lights on (1 HALO), 5 h prior to feeding (12 HALO), and 2 h after feeding (19 HALO). In addition, in whole animals studies, changes in core body temperature were monitored. In mice which had been subjected to a chronic diet restriction (60% of the intake of ad libitum controls), c -myc expression was significantly reduced at 1 HALO and 19 HALO compared to corresponding ad libitum animals. In addition, significant differences in c -myc expression were found between time points, in both the ad libitum and restricted groups, suggesting that myc protooncogene expression in the liver may be regulated in a circadian fashion. C -myc expression may correlate with body temperature, suggestng a possible association with metabolic output.