Effects of dietary fat and endurance exercise on plasma cortisol, prostaglandin E2, interferon-gamma and lipid peroxides in runners.

OBJECTIVE: Exercise and the neuroendocrine and oxidative stress it elicits on immune function is modulated by dietary fat intake. The effects of increasing dietary fat on endurance exercise-induced alterations 80% of VO2max for 2 hours) in the plasma levels of cortisol and prostaglandin E2 (PGE2), interferon-gamma (IFN-gamma) and lipid peroxides were investigated. As higher levels of cortisol, PGE2 and lipid peroxides could be immunosuppressive, the effects of different levels of dietary fat on these measures in runners were determined. METHODS: Healthy trained runners (males and females) consumed serially 15% fat diet (of daily energy), 30% fat diet and 40% fat diets for four weeks each. In the last week of each diet period the subjects ran to exhaustion at 80% of their VO2max and blood was drawn pre- and post-run. Cortisol, IFN-gamma, PGE2 and lipid peroxides were determined using standard techniques. RESULTS: Pre-exercise levels of plasma cortisol were elevated, IFN-gamma was unchanged and PGE2 and lipid peroxides decreased on the 40%F diet compared to 30%F and 15%F. Post-exercise levels of plasma cortisol (p < 0.004), PGE2 (p < 0.0057) and lipid peroxide levels increased (p < 0.0001) after endurance exercise on all diets. The rates of increase of plasma cortisol levels during exercise were similar on all three diets. Although absolute cortisol levels were higher in the high fat group, the rate of increase of plasma cortisol level during exercise was similar on each diet. The dietary fat levels did not affect IFN-gamma, however, PGE2 and lipid peroxides decreased with increasing fat at baseline at 40%F level (p < 0.01; 30%F vs. 40% F: p < 0.002; 15%F vs. 40%F: p < 0.007). CONCLUSIONS: Data from the present study suggest that higher levels of fat in the diet, up to 40%, increase endurance running time without adverse effects on plasma cortisol, IFN-gamma, and lipid peroxide levels.

Helical peptide models for protein glycation: proximity effects in catalysis of the Amadori rearrangement.

INTRODUCTION: Non-enzymatic glycation of proteins has been implicated in various diabetic complications and age-related disorders. Proteins undergo glycation at the N-terminus or at the epsilon-amino group of lysine residues. The observation that only a fraction of all lysine residues undergo glycation indicates the role of the immediate chemical environment in the glycation reaction. Here we have constructed helical peptide models, which juxtapose lysine with potentially catalytic residues in order to probe their roles in the individual steps of the glycation reaction. RESULTS: The peptides investigated in this study are constrained to adopt helical conformations allowing residues in the i and i+4 positions to come into spatial proximity, while residues i and i+2 are far apart. The placing of aspartic acid and histidine residues at interacting positions with lysine modulates the steps involved in early peptide glycation (reversible Schiff base formation and its subsequent irreversible conversion to a ketoamine product, the Amadori rearrangement). Proximal positioning of aspartic acid or histidine with respect to the reactive lysine residue retards initial Schiff base formation. On the contrary, aspartic acid promotes catalysis of the Amadori rearrangement. Presence of the strongly basic residue arginine proximate to lysine favorably affects the pK(a) of both the lysine epsilon-amino group and the singly glycated lysine, aiding in the formation of doubly glycated species. The Amadori product also formed carboxymethyl lysine, an advanced glycation endproduct (AGE), in a time-dependent manner. CONCLUSIONS: Stereochemically defined peptide scaffolds are convenient tools for studying near neighbor effects on the reactivity of functional amino acid sidechains. The present study utilizes stereochemically defined peptide helices to effectively demonstrate that aspartic acid is an efficient catalytic residue in the Amadori arrangement. The results emphasize the structural determinants of Schiff base and Amadori product formation in the final accumulation of glycated peptides.

Dietary fats and immune status in athletes: clinical implications.

Athletes are competitive, train at very high levels with inadequate rest, consume too few calories, avoid fats, and may be at increased risk of infections. The immune system is sensitive to both fat intake and intense exercise, suggesting that athletes may have suppressed immune function. It has been reported that many athletes consume about 25% fewer calories than the estimated expenditure, leading to low intakes of some essential micronutrients and fats. Acute exercise has been shown to increase inflammatory and decrease antiinflammatory immune factors and may increase oxidant stress. Chronic exercise appears to improve immune competence. Lipids are powerful mediators of the immune system, and they may modulate the immunosuppressive effects of strenuous exercise. Studies have shown that a low-fat high-carbohydrate diet (15% fat, 65% CHO, 20% protein of total calories), typically eaten by athletes, increases inflammatory and decreases antiinflammatory immune factors, depresses antioxidants, and negatively affects blood lipoprotein ratios. Increasing total caloric intake by 25% to match energy expenditure and the dietary fat intake to 32% in athletes appears to reverse the negative effects on immune function and lipoprotein levels reported on a low-fat diet. Increasing the dietary fat intake of athletes to 42%, while maintaining caloric intake equal to expenditure, does not negatively affect immune competency or blood lipoproteins, whereas it improves endurance exercise performance at 60-80% of VO2max in cyclists, soldiers, and runners. There is no evidence that higher fat intakes (up to 42% of total calories), in calorically balanced diets, increase the risk of cancer, but studies are needed to determine whether the beneficial effects of higher fat diets in athletes reduce their rate of infections.

A perspective on fat intake in athletes.

Performance in endurance events is dependent upon the maximal aerobic power, the percentage of that power that can be sustained and the availability of substrates (carbohydrates [CHO] and fats). The purpose of this paper is to present a perspective of recent studies that demonstrate the role of fat intake and oxidation on endurance performance. Studies have shown that fatigue is associated with reduced muscle glycogen and that increasing muscle glycogen or blood glucose prolongs performance while increasing fat and decreasing CHO decreases performance. This has led to an emphasis on CHO intake in athletes in endurance sports, which quite often leads to low caloric intake. It is well known that trained subjects have higher levels of fat oxidative capacity, which spares glycogen during endurance sports. Data from recent studies in trained athletes, who were fed iso-caloric high-fat diets (42% to 55%) that maintained adequate CHO levels, have shown an increase in endurance in both men and women when compared to diets composed of low fat intake (10% to 15%). The magnitude of the effect on endurance was significant at high percentages of maximal aerobic power and increased as the percentage of maximal aerobic power decreased. Based on this review, a baseline diet comprising 20% protein, 30% CHO and 30% fat, with the remaining 20% of the calories distributed between CHO and fat based on the intensity and duration of the sport, is recommended for discussion and future research.

Effects of dietary omega-3 and omega-6 lipids and vitamin E on serum cytokines, lipid mediators and anti-DNA antibodies in a mouse model for rheumatoid arthritis.

OBJECTIVE: Omega-3 (omega-3) fatty acid rich-fish oil (FO) and vitamin E (vit-E) may delay the progress of certain autoimmune diseases. The present study examined the mechanism of action of omega-3 and omega-6 lipids and vit-E on the serum cytokines and lipid mediators in autoimmune-prone MRL/lpr mice (a model for rheumatoid arthritis, RA). The lpr (lymphoproliferative) gene is overexpressed in these mice causing extensive lymphoproliferation, lupus-like symptoms and accelerated aging. METHODS: Weanling female MRL/lpr and congenic control MRL/++ mice were fed 10% corn oil (CO, omega6) or FO-based semipurified diets containing two levels of vitamin E (vit-E-75, I.U. and vit-E-500 I.U./Kg diet) for four months. At the end of the experiment, serum anti-DNA antibodies, cytokines and lipid mediators levels were determined. RESULTS: The appearance of enlarged lymph nodes was delayed in the mice fed FO, and the FO-500 IU vit-E diet offered further protection against enlargement of lymph nodes. The MRL/lpr mice exhibited significantly higher levels of serum anti-dsDNA antibodies. The FO-fed mice had significantly lower serum IL-6, IL-10, IL-12, TNF-alpha, PGE2, TXB2 and LTB4 levels compared with CO-fed mice. In mice fed 500 IU vit-E diets, the serum IL-6, IL-10, IL-12 and TNF-alpha levels were significantly lower and serum IL-1beta was significantly higher compared to 75 IU-vit-E-fed mice in CO/FO or both. The levels of anti-DNA antibodies, IL-4, IL-6, TNF-alpha, IL-10 and IL-12 were higher in the sera of MRL/lpr mice. The FO diet lowered the levels of these cytokines (except IL-4) and lipid mediators. Adding 500 IU of vit-E to the FO diet further lowered the levels of IL-6, IL-10, IL-12, and TNF-alpha. CONCLUSION: It is clear from our observations that the beneficial effects of FO can be enhanced by the addition of 500 IU of vit-E in the diet. The FO diet containing 500 IU of vit-E may specifically modulate the levels of IL-6, IL-10, IL-12 and TNF-alpha and thereby may delay the onset of autoimmunity in the MRL/lpr mouse model. The observations from this study may form a basis for selective nutrition intervention based on specific fatty acids and antioxidants in delaying the progress of RA.

Fluoroalcohols as structure modifiers in peptides and proteins: hexafluoroacetone hydrate stabilizes a helical conformation of melittin at low pH.

The effect of hexafluoroacetone hydrate (HFA) on the structure of the honey bee venom peptide melittin has been investigated. In aqueous solution at low pH melittin is predominantly unstructured. Addition of HFA at pH approximately 2.0 induces a structural transition from the unstructured state to a predominantly helical conformation as suggested by intense diagnostic far UV CD bands. The structural transition is highly cooperative and complete at 3.6 M (50% v/v) HFA. A similar structural transition is also observed in 2,2,2 trifluoroethanol which is complete only at a cosolvent concentration of approximately 8 M. Temperature dependent CD experiments support a 'cold denaturation' of melittin at low concentrations of HFA, suggesting that selective solvation of peptide by HFA is mediated by hydrophobic interactions. NMR studies in 3.6 M HFA establish a well-defined helical structure of melittin at low pH, as suggested by the presence of strong NH/NHi+1 NOEs throughout the sequence, along with many medium range helical NOEs. Structure calculations using NOE-driven distance constraints reveal a well-ordered helical fold with a relatively flexible segment around residues T10-G11-T12. The helical structure of melittin obtained at 3.6 M HFA at low pH is similar to those determined in methanolic solution and perdeuterated dodecylphosphocholine micelles. HFA as a cosolvent facilitates helix formation even in the highly charged C-terminal segment.

Effects of dietary omega3 and omega6 lipids and vitamin E on proliferative response, lymphoid cell subsets, production of cytokines by spleen cells, and splenic protein levels for cytokines and oncogenes in MRL/MpJ-lpr/lpr mice.

omega3 Fatty acid rich fish oil (FO) and vitamin E may delay the progress of certain autoimmune diseases. The present study examined the mechanisms of action of omega3 lipids and vitamin E in autoimmune-prone MRL/lpr mice suffering from extensive lymphoproliferation, lupus-like symptoms, and accelerated aging. To determine whether the effects of omega3 lipids in autoimmune disease is linked to vitamin E levels, weanling female MRL/lpr and congenic control MRL/++ mice were fed diets containing 10% corn oil (CO) or 10% FO at two levels of vitamin E (75 IU or 500 IU/kg diet) for 4 months. The appearance of lymph nodes was delayed in the mice fed FO, and higher levels of FO offered further protection against the appearance of lymph nodes. Analysis of the spleen cells revealed that the cells positive for Thy.1 and Fas were significantly higher in the MRL/++ mice. The groups fed high levels of vitamin E generally exhibited higher levels of Fas. The proliferative response of splenocytes of MRL/++ mice to mitogens was significantly higher compared with MRL/lpr mice. Interleukin (IL)-10 production by spleen cells was significantly higher in FO-fed MRL/lpr mice than in CO-fed mice. In mice fed a high level of vitamin E, the production of IL-12 and tumor necrosis factor-alpha was significantly lower and IL-2 was significantly higher than in animals fed a low level of vitamin E. Proinflammatory cytokines were higher in the MRL/lpr mice and both FO and vitamin E lowered the levels of proinflammatory cytokines and lipid mediators. Western blots revealed that c-myc and c-ras were significantly lower and IL-2 and transforming growth factor (TGF)-beta1 levels were significantly higher in the spleens of MRL/++ mice. FO lowered c-myc and high levels of vitamin E in the diets normalized the levels of TGF-beta1 in MRL/lpr mice. The observations from this study suggest that both FO and vitamin E modulate the levels of specific cytokines, decrease the levels of proinflammatory cytokines, inflammatory lipid mediators, and c-myc, and increase TGF-beta1 levels in spleens of MRL/lpr mice and thus may delay the progress of autoimmune diseases.

Effects of dietary n-6 and n-3 lipids on antioxidant defense system in livers of exercised rats.

OBJECTIVE: The present study was designed to investigate the effects of dietary n-6 and n-3 lipids and exercise on the activities of hepatic antioxidant enzymes and microsomal lipid composition and peroxidation in Fischer-344 male rats. METHODS: Weanling male Fischer-344 rats were fed ad libitum semipurified diets containing 10% corn oil (CO) or 10% fish oil (FO), with equal levels of antioxidants. After 2 months on the diets, weight-matched animals in each diet group were divided into sedentary (S) and exercised (Ex) groups, and the diets were continued. The animals in the exercise group were run on a treadmill 30 to 40 minutes to exhaustion 6 days/week for 2 months. At the end of 2 months, the rats were sacrificed and livers were collected; antioxidant enzymes were determined in the cytosol, fatty acid composition was analyzed in the microsomes, and vitamin E levels were analyzed in the sera. RESULTS: The rats in the FO-S group exhibited significantly higher liver cytosolic catalase activity, while their superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were significantly lower compared to the CO-S group. The GSH-Px activity was significantly higher in the FO-Ex group compared to FO-S group. The source of dietary lipids significantly influenced the fatty acid composition of the total lipids in the microsomes. Feeding the FO-based diet significantly increased 18:0 and n-3 fatty acids incorporation into the microsomes (18:3, 20:5, 22:5, and 22:6), whereas ingestion of CO resulted in a significant increase in 14:0, 14:1, 18:1, and n-6 fatty acids (18:2 and 20:4). The serum vitamin E levels were significantly higher in the CO groups, and exercise had no effect on vitamin E levels. Exercise significantly decreased the generation of thiobarbituric acid reactive substances (TBARS) by liver microsomes. Consumption of FO, which is highly susceptible to oxidation, did not show any significant changes in membrane lipid peroxidation. CONCLUSIONS: The present study suggests that feeding FO increases the activity of liver cytosolic catalase in FO-S rats and GSH-Px in FO-Ex rats. In addition, exercise significantly decreased the generation of TBARS by the liver microsomal lipids. Serum vitamin E levels were higher in the CO group and exercise did not alter vitamin E levels. This suggests that the amount of vitamin E included in the diets was possibly adequate to cope with the oxidative stress induced during exercise.

Effects of the level of dietary fat intake and endurance exercise on plasma cytokines in runners.

PURPOSE: Chronic exercise and high fat diets have been associated with immune suppression. We have reported the effects of level of dietary fat and exercise on lymphocyte subsets, proliferative response, and in vitro production of cytokines by peripheral blood mononuclear cells of runners. The present study was planned to further investigate whether the mechanisms of action of dietary fats is through their modulation of plasma cytokines in runners. METHODS: This study compared plasma cytokines at rest and after endurance exercise at 80% of V02max in female (N = 8-10) and male (N = 8-10) runners after eating diets comprised of 17% (LF), 32% (MF), and 41% (HF) fats (4 wk each). RESULTS: The level of interleukin-1 beta (IL-1 beta) was independent of gender, exercise, and level of dietary fat. tumor necrosis factor (TNF-alpha) level was higher in the plasma of men compared with that in women runners, and the level of these two cytokines increased with increasing level of dietary fat. Plasma interleukin-2 (IL-2) level (a cytokine involved in enhancing T cell functions for host defense) was significantly higher in men compared with that in women runners and decreased in men with increase dietary fat. Plasma interleukin-6(IL-6) level was significantly lower after the endurance run, and IL-6 levels decreased with increase in dietary fat. CONCLUSIONS: Data from the present study suggest that dietary fat has differential effects on plasma cytokine levels in runners. Increasing the level of dietary fat significantly increased endurance run time and had no adverse effects on the level of plasma IL-2 and pro-inflammatory cytokines (IL-1 beta, IL-6 and TNF-alpha in runners.

Effects of calorie restriction and ω-3 dietary fat on aging in short-and long-lived rodents.

Aging is accompanied by a steady increase in the incidence of spontaneous tumors and a decline in immune function. Calorie restriction (CR) or supplementation with ω-3 fats prolongs life span, suppresses tumorigenesis, and ameliorates immune function in a variety of experimental models. We suggest that decreased oxidant stress and upregulation of apoptosis mediate the effects of calorie restriction on immunity and longevity. CR prolongs life span in several animal models and our studies have examined the effects of CR on the immune system and on tumorigenesis. CR maintains naive T cells, prevents the rise in "double-negative" T cells, maintains lymphocyte responsiveness to mitogens, and preserves Dexamethasone induced apoptosis in spleen cells of MRL/Ipr mice. CR also modulates the expression of inflammatory mediators and cytokines. CR decreases the Sjögren's syndrome-like chronic inflammation of salivary glands of B/W animals while increasing expression of the immunosuppressive cytokine TGFß1 and decreasing expression of the pro-inflammatory cytokines IL-6 and TNFα. The autoimmune disease in the B/W mouse also affects the kidneys, and we find that renal expression of platelet derived growth factor-A, (PDGF-A) and thrombin receptor are decreased in CR animals. Similarly, CR decreases the expression and localization of plasminogen activator inhibitor type 1 in glomeruli of B/W animals. CR also modulates expression and function of androgen receptors and the binding of insulin to liver nuclei. Finally, CR suppresses the development of breast tumors in the Ras oncomouse. These effects of calorie restriction are paralleled in short-lived B/W animals fed diets supplemented with ω-3 fatty acids. Omega-3 fatty acids induce the expression of hepatic antioxidant enzymes, and enhance apoptosis in lymphocytes of B/W animals.

X-ray studies on crystalline complexes involving amino acids and peptides. XXXII. Effect of chirality on ionisation state, stoichiometry and aggregation in the complexes of oxalic acid with DL- and L-lysine.

Crystals of the oxalic acid complex of DL-lysine (triclinic P1; a = 5.540(1), b = 10.764(2), c = 12.056(2) A, alpha = 77.8(1), beta = 80.6(1), gamma = 75.6(1).; R = 4.7% for 2023 observed reflections) contain lysine and semioxalate ions in the 1:1 ratio, whereas the ratio of lysine and semioxalate/oxalate ions is 2:3 in the crystals of the L-lysine complex (monoclinic P2(1); alpha = 4.906(1), b = 20.145(4), c = 12.455(1) A, beta = 92.5(1).; R = 4.4% for 1494 observed reflections). The amino acid molecule in the L-lysine complex has an unusual ionisation state with positively charged alpha- and side-chain amino groups and a neutral carboxyl group. The unlike molecules aggregate into separate alternating layers in the DL-lysine complex in a manner similar to that observed in several of the amino acid complexes. The L-lysine complex exhibits a new aggregation pattern which cannot be easily explained in terms of planar features, thus emphasizing the fundamental dependence of aggregation on molecular characteristics. Despite the differences in stoichiometry, ionisation state and long-range aggregation patterns, the basic element of aggregation in the two complexes exhibits considerable similarity.

Influence of the level of dietary lipid intake and maximal exercise on the immune status in runners.

Chronic exercise and high fat diets are associated with immune suppression. This study compares cellular immune responses at rest and after maximal exercise in runners after eating diets comprised of 17% low fat (LF), 32% medium fat (MF), and 41% high fat (HF) (4 wk each). VO2max increased significantly from the 17% to 41% fat diet. The leukocyte cell counts were significantly increased after exercise. In men, significantly higher proliferative response to phytohemagglutinin (PHA) (P < 0.004) was observed with MF diet, while response to pokeweed mitogen (PWM) was significantly decreased by MF and HF diets. The number of CD8+ (suppressor) T cells was significantly higher in men and exercise increased it significantly, while CD4+ (helper) T cells were not affected. Natural killer cells number was significantly increased 2.5 fold by exercise and with increase in dietary fat. The production of IL-2 by peripheral blood mononuclear cells was significantly higher in men (P < 0.0001) and increasing dietary fat significantly increased IL-2 production (P < 0.001). In men, exercise decreased the level of the proinflammatory cytokines (IL-1, IL-6 and TNF-alpha), whereas in women, with the exception of MF diet for IL-6, exercise had no effect. This study indicates that short, intense bouts of exercise in runners training 40 miles.wk-1 have mixed effects on the immune system. A high percentage of fat intake (41%) did not have any deleterious effects on the immune system of the well-trained runners.

Effect of food restriction on life span and immune functions in long-lived Fischer-344 x Brown Norway F1 rats.

Life-long food restriction is known to slow aging and reduce the rate of occurrence of age-associated disease processes, but the mechanism by which this is accomplished is unknown. In this study we have examined the effect of food restriction on the proliferative response of spleen cells to mitogens and lymphokine production in 6-, 18-, and 30-month-old AL and FR Fischer-344 x Brown Norway (F-344 x BNF1) female rats whose average life span is 137 weeks on an ad libitum (AL) diet and 177 weeks on a food-restricted (FR) diet. In addition, the ability of food restriction to recall antigens was tested in 10-month-old rats by immunizing them with keyhole limpet and hen's egg albumin and measuring proliferative response of draining lymph node cells to these antigens. Our results indicated that the spleen-cell proliferative response to phytohemagglutinin and concanavalin A (Con A) was equal in 6- and 18-month-old rats but declined significantly in 30-month-old AL rats compared to FR rats. Although flow cytometric analyses did not reveal differences for CD4, CD8, and Ig+ cells with age, a significant rise in memory T cells (Ox-22low) in both CD4+ and CD8+ T-cell subset lineage was noted in AL-fed rats at 30 months of age. In FR rats, however, only a minimal shift of naive T cells (Ox-22high) to memory cells was observed. In FR rats, the observed changes in the naive and memory T-cell subsets correlate well with the observed higher levels of the antiinflammatory interleukin-2 (IL-2) and lower levels of the proinflammatory cytokines such as IL-6 and tumor necrosis factor-alpha. The ability of food-restricted animals to recall antigens was lower compared to their age-matched controls, though the proliferative response to T-cell mitogen Con A and superantigen staphylococcal enterotoxin B was higher. These findings indicate that food restriction may selectively act to maintain a lower number of antigen-induced memory T cells with age, thereby maintaining the organism's ability to produce higher levels of IL-2 with age. In summary, the increased cell-mediated immune function noted in aged FR rats appears to be due to the presence of a higher number of naive T cells, which are known to produce elevated levels of the antiinflammatory cytokines, which may in part be responsible for reducing the observed age-related rise in disease.

The role of dietary fat on performance, metabolism, and health.

This paper presents a model to evaluate the nutritional status of trained athletes based on work in our laboratory as well as others. The model proposes that substrate use is set by the muscle fibers recruited, based on the exercise intensity. Second, the substrate available is primarily determined by the intramuscular stores. In trained athletes, intramuscular fat plays an important role in metabolism at exercise intensities as high as 80% of maximal aerobic power. Based on these factors, increasing the fat in the diet (while maintaining adequate intramuscular glycogen) increases VO2max and intramuscular stores of fat (presumably due to increased mitochondrial volume). These two factors result in a significant increase in the time to exhaustion at set levels of exercise (endurance). It also appears that fatigue is associated with depletion of either glycogen or fat. These conclusions hold true for athletes on diets where sufficient calories are taken in to meet demands and for exercise levels below 80% of VO2max, where primarily slow-twitch oxidative fibers are used. These data may not apply in exercise where predominantly fast-twitch fibers are used. Also, these data do not apply to runners eating a hypocaloric diet, where reducing the percentage of carbohydrates may compromise their glycogen stores. It would appear that the fat in the diet can be increased to a very high level without compromising the cardiovascular or immune systems of athletes. Moreover, it can be proposed that these data could be applied to sedentary persons, as long as they are isocaloric. This would imply that the fat consumed in the diet would be used in the muscle, as in the runners, although at a lower level. Thus, the dietary intake should be matched in both total calories and percentage of fats and carbohydrates to calories consumed by daily activity. It should be cautioned that if glycogen and fat stores are compromised, protein resynthesis is inhibited and loss of muscle mass may result. This has a negative effect on the athlete's ability to perform at high levels.

Dietary lipids and calorie restriction affect mammary tumor incidence and gene expression in mouse mammary tumor virus/v-Ha-ras transgenic mice.

We have studied the effects of food restriction (FR) and substitution of fish oil (FO; omega 3) for corn oil (CO; omega 6) on breast tumor incidence and survival in mouse mammary tumor virus/v-Ha-ras transgenic (Onco) mice. The diets were as follows: group 1, 5% (wt/wt) CO fed ad libitum (AL); group 2, 5% CO, restricted calories (40% fewer calories than AL; FR); group 3, 20% CO fed AL; and group 4, 20% FO fed AL. After 3 years, 40% of FR Onco (group 2) mice were alive, whereas there were no survivors in the other three groups. Similarly, tumor incidence was reduced to 27% (5 out of 18) in FR animals (group 2), whereas it was 83% (11 out of 13) in group 1 mice, 89% (16 out of 18) in group 3 mice, and 71% (10 out of 14) in group 4 mice. These protective effects of FR on survival and tumor incidence were paralleled by higher expression of the tumor suppressor gene p53 (wild type) and free-radical scavenging enzymes (catalase and superoxide dismutase) in breast tumors. Immunoblotting showed less ras gene product, p21, and increased p53 levels in the tumors of FR mice. In addition, FR decreased RNA levels of c-erbB-2, interleukin 6, and the transgene v-Ha-ras in tumors. In contrast, analysis of hepatic mRNA from tumor-bearing FR mice revealed higher expression of catalase, glutathione peroxidase, and superoxide dismutase. Survival and tumor incidence were not influenced significantly by dietary supplementation with FO in place of CO. Taken together, our studies suggest that moderate restriction of energy intake significantly inhibited the development of mammary tumors and altered expression of cytokines, oncogenes, and free-radical scavenging enzymes.

Dietary omega-3 lipids delay the onset and progression of autoimmune lupus nephritis by inhibiting transforming growth factor beta mRNA and protein expression.

The present study was carried out to test whether transforming growth factor beta (TGF beta) plays a pathological role in the induction or progression of glomerulonephritis in a murine model of systemic lupus erythematosus (SLE), and whether dietary supplementation with fish oil (FO) can modulate the expression of TGF beta. Weanling female (NZB x NZW) F1 (B/W) mice were divided into three groups. One group was fed an unmanipulated diet (lab. chow; LC) and the other two groups were fed a nutritionally adequate semipurified diet supplemented with 10% CO or FO. Both water and food were provided ad libitum. Proteinuria and serum anti-dsDNA antibody levels were measured to assess disease progression. Mice were killed at 3.5 and 6.5 months of age and renal mRNA levels for TGF beta isoforms, fibronectin-1 (FN-1) and intercellular adhesion molecule-1 (ICAM-1) were studied by Northern blot analysis. TGF beta 1 protein levels were also examined in kidneys by Western blot analysis. Our results indicate that at 3.5 months of age, when urinary protein levels were undetectable and very low levels of anti-dsDNA were detected, no mRNA signal could be detected for TGF beta isoforms, ICAM-1 and FN-1 in either dietary group. However, at 6.5 months, the FO-fed mice, compared to LC and CO, had [1] greatly reduced proteinuria (LC: 2-3+, CO: 2-3+; FO: trace -1+) and serum anti-dsDNA antibodies; [2] improved survival (CO: 100% death (15/15) occurred by 8 months; FO: 50% were alive at 12 months (8/15) and [3] reduced renal TGF beta 1 mRNA and protein levels. TGF beta 2 and beta 3 were not significantly affected by FO diet. Similarly, lower levels of renal FN-1 and ICAM-1 mRNA were observed in FO fed mice. These data indicate that in B/W mice on a FO diet, prolonged survival and amelioration of renal disease may be attributed at least in part to lower levels of TGF beta 1 mRNA and protein in the kidneys.

Modulation of antioxidant activities and immune response by food restriction in aging Fisher-344 rats.

Food restriction delays the loss of several cellular immune functions, retards the onset of many diseases during aging and, consequently, extends life span significantly in laboratory rodents. The present study was undertaken to determine whether the age-associated loss in immune function is linked to changes in microsomal and mitochondrial membranes of spleens in Fischer-344 (F-344) male rats. In this study, we determined cytosolic superoxide dismutase activity (SOD), fluidity and cholesterol content in the splenic microsomal and mitochondrial membranes, and DNA synthesis and IL-2 production in spleen cells from young and old ad libitum-fed (AL) and food restricted (FR) rats. The results show that proliferative response to phytohemagglutinin (PHA) and concanavalin A (Con-A) was significantly higher in the spleen cells of 18-month- and 24-month-old FR rats, as compared to their age-matched AL controls. Cytosolic SOD activity in the 24-month-old AL rats decreased by 28% as compared to 6-month-old AL rats, whereas in FR old rats, the loss was only 12%, suggesting that food restriction prevents loss in cytosolic SOD activity in spleens. Our data are consistent with the notion that food restriction modulates loss in immune response of splenocytes by maintaining both cytosolic SOD activity and membrane fluidity during aging.

Effects of n-3 and n-6 fatty acids on the activities and expression of hepatic antioxidant enzymes in autoimmune-prone NZBxNZW F1 mice.

Menhaden fish oil (FO) containing n-3 fatty acids dramatically extends the life span and delays the onset and progression of autoimmune disease in (NZBxNZW)F1 (B/W) female mice as compared to those fed corn oil (CO) rich in n-6 lipids. As an inefficient antioxidant defense system has been linked to autoimmune diseases, the present study was undertaken to determine whether the protective action of n-3 lipids is mediated through their antioxidant defense system. Weanling B/W mice were fed a nutritionally adequate, semipurified diet containing CO or krill oil (KO) or FO at 10% level (w/w) ad libitum until the mice were 6.5 months old. All diets contained the same level of vitamin E (21.5 mg/100 g diet). We compared the effects of feeding n-6 and n-3 lipids on survival, kidney disease, hepatic microsomal lipid composition, peroxidation, and on the activity and mRNA expression of the antioxidant enzymes catalase, glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) in 6.5-month-old B/W mice. The results showed that when compared to livers from CO-fed mice, livers from KO- and FO-fed mice showed: (i) significantly higher (P < 0.001) activities and expression of CAT, GSH-Px and SOD; (ii) significantly lower (P < 0.001) arachidonic acid (20:4n-6) and linoleic acid (18:2n-6) and higher (P < 0.001) eicosapentaenoic acid (20:5n-3) and docosahexaenoic acid (22:6n-3) levels in hepatic microsomes; and (iii) significantly lower (P < 0.001) estimated peroxidation indices and thiobarbituric acid reactive substances generation.(ABSTRACT TRUNCATED AT 250 WORDS)