Does Oil Rich in Alpha-Linolenic Fatty Acid Cause the Same Immune Modulation as Fish Oil in Walker 256 Tumor-Bearing Rats?

OBJECTIVE: Polyunsaturated fatty acids n-3 (PUFA n-3) have shown effects in reducing tumor growth, in particular eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) abundantly present in fish oil (FO). When these fatty acids are provided in the diet, they alter the functions of the cells, particularly in tumor and immune cells. However, the effects of α-linolenic fatty acid (ALA), which is the precursor of EPA and DHA, are controversial. Thus, our objective was to test the effect of this parental fatty acid. METHODS: Non-tumor-bearing and tumor-bearing Wistar rats (70 days) were supplemented with 1 g/kg body weight of FO or Oro Inca® (OI) oil (rich in ALA). Immune cells function, proliferation, cytokine production, and subpopulation profile were evaluated. RESULTS: We have shown that innate immune cells enhanced phagocytosis capacity, and increased processing and elimination of antigens. Moreover, there was a decrease in production of pro-inflammatory cytokines (tumor necrosis factor-alpha (TNF-α) and interleukin 6 (IL-6)) by macrophages. Lymphocytes showed decreased proliferation capacity, increased cluster of differentiation 8 (CD8+) subpopulation, and increased TNF-α production. CONCLUSIONS: Oil rich in ALA caused similar immune modulation in cancer when compared with FO.

α-Linolenic Fatty Acid Supplementation Decreases Tumor Growth and Cachexia Parameters in Walker 256 Tumor-Bearing Rats.

Fish oil (FO) has been shown to affect cancer cachexia, tumor mass, and immunity cell. n-3 PUFA, specifically α-linolenic fatty acid (ALA), has controversial effects. We investigated this in nontumor-bearing Wistar rats fed regular chow (C), fed regular chow and supplemented with FO or Oro Inca oil (OI), and Walker 256 tumor-bearing rats fed regular chow (W), fed regular chow and supplemented with FO (WFO) or OI (WOI). Rats were supplemented (1g/kg body weight/day) during 4 wk and then the groups tumor-bearing were inoculated with Walker 256 tumor cells suspension and 14 days later the animals were killed. WFO increased EPA fivefold and DHA 1.5-fold in the tumor tissue compared to W (P < 0.05). OI supplementation increased of threefold of ALA when compared to W (P < 0.05). Tumor mass in WFO and OI was of 2.3-fold lower, as well as tumor cell proliferation of 3.0-fold tumor tissue lipoperoxidation increased of 76.6% and cox-2 expression was 20% lower. Cachexia parameters were attenuate, blood glucose (25% higher), Triacylglycerolemia (50% lower), and plasma TNF-α (65% lower; P < 0.05) and IL-6 (62.5% lower). OI, rich in ALA, caused the same effect on cancer as those seen in FO.

Fish oil supplementation for two generations increases insulin sensitivity in rats.

We investigated the effect of fish oil supplementation for two consecutive generations on insulin sensitivity in rats. After the nursing period (21 days), female rats from the same prole were divided into two groups: (a) control group and (b) fish oil group. Female rats were supplemented with water (control) or fish oil at 1 g/kg body weight as a single bolus for 3 months. After this period, female rats were mated with male Wistar rats fed on a balanced chow diet (not supplemented). Female rats continued to receive supplementation throughout gestation and lactation periods. The same treatment was performed for the next two generations (G1 and G2). At 75 days of age, male offspring from G1 and G2 generations from both groups were used in the experiments. G1 rats did not present any difference with control rats. However, G2 rats presented reduction in glycemia and lipidemia and improvement in in vivo insulin sensitivity (model assessment of insulin resistance, insulin tolerance test) as well as in vitro insulin sensitivity in soleus muscle (glucose uptake and metabolism). This effect was associated with increased insulin-stimulated p38 MAP kinase phosphorylation and lower n-6/n-3 fatty acid ratio, but not with activation of proteins from insulin signaling (IR, IRS-1 and Akt). Global DNA methylation was decreased in liver but not in soleus muscle. These results suggest that long-term fish oil supplementation improves insulin sensitivity in association with increased insulin-stimulated p38 activation and decreased n-6:n-3 ratio in skeletal muscle and decreased global DNA methylation in liver.

Fish oil supplementation reduces cachexia and tumor growth while improving renal function in tumor-bearing rats.

The objective of the present work was to study the renal function of healthy and tumor-bearing rats chronically supplemented with fish oil (FO), a source of n-3 polyunsaturated fatty acids. Weanling male rats were divided in two groups, one control (C) and another orally supplemented for 70 days with FO (1 g/kg body weight). After this time, half the animals of each group were injected in the right flank with a suspension of Walker 256 tumor cells (W and WFO). The W group had less proteinemia reflecting cachectic proteolysis, FO reversed this fact. Tumor weight gain was also reduced in WFO. Glomerular filtration rate (GFR) was not different in FO or W compared to C, but was higher in WFO. Renal plasma flow (RPF) was higher in the FO supplemented groups. The W group had lower plasma osmolality than the C group, but FO supplementation resulted in normalization of this parameter. Fractional sodium excretion (FE(Na+)) of FO rats was similar to C. Proximal Na(+) reabsorption, evaluated by lithium clearance, was similar among the groups. Urinary thromboxane B(2) (TXB(2)) excretion was lower in the supplemented groups. The number of macrophages in renal tissue was higher in W compared to C rats, but was lower in WFO rats compared to W rats. In conclusion, FO supplementation resulted in less tumor growth and cachexia, and appeared to be renoprotective, as suggested by higher RPF and GFR.

Low fish oil intake improves insulin sensitivity, lipid profile and muscle metabolism on insulin resistant MSG-obese rats.

BACKGROUND: Obesity is commonly associated with diabetes, cardiovascular diseases and cancer. The purpose of this study was to determinate the effect of a lower dose of fish oil supplementation on insulin sensitivity, lipid profile, and muscle metabolism in obese rats. METHODS: Monosodium glutamate (MSG) (4 mg/g body weight) was injected in neonatal Wistar male rats. Three-month-old rats were divided in normal-weight control group (C), coconut fat-treated normal weight group (CO), fish oil-treated normal weight group (FO), obese control group (Ob), coconut fat-treated obese group (ObCO) and fish oil-treated obese group (ObFO). Obese insulin-resistant rats were supplemented with fish oil or coconut fat (1 g/kg/day) for 4 weeks. Insulin sensitivity, fasting blood biochemicals parameters, and skeletal muscle glucose metabolism were analyzed. RESULTS: Obese animals (Ob) presented higher Index Lee and 2.5 fold epididymal and retroperitoneal adipose tissue than C. Insulin sensitivity test (Kitt) showed that fish oil supplementation was able to maintain insulin sensitivity of obese rats (ObFO) similar to C. There were no changes in glucose and HDL-cholesterol levels amongst groups. Yet, ObFO revealed lower levels of total cholesterol (TC; 30%) and triacylglycerol (TG; 33%) compared to Ob. Finally, since exposed to insulin, ObFO skeletal muscle revealed an increase of 10% in lactate production, 38% in glycogen synthesis and 39% in oxidation of glucose compared to Ob. CONCLUSIONS: Low dose of fish oil supplementation (1 g/kg/day) was able to reduce TC and TG levels, in addition to improved systemic and muscle insulin sensitivity. These results lend credence to the benefits of n-3 fatty acids upon the deleterious effects of insulin resistance mechanisms.

Effect of fish oil supplementation for two generations on changes of lymphocyte function induced by Walker 256 cancer cachexia in rats.

Fish oil supplementation has been shown to improve the cachectic state of tumor-bearing animals and humans. Our previous study showed that fish oil supplementation (1 g per kg body weight per day) for 2 generations had anticancer and anticachetic effects in Walker 256 tumor-bearing rats as demonstrated by reduced tumor growth and body weight loss and increased food intake and survival. In this study, the effect of fish oil supplementation for 2 generations on membrane integrity, proliferation capacity, and CD4/CD8 ratio of lymphocytes isolated from mesenteric lymph nodes, spleen, and thymus of Walker 256 tumor-bearing animals was investigated. We also determined fish oil effect on plasma concentration and ex vivo production of cytokines [tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), interleukin-4 (IL-4), IL-6, and IL-10]. Lymphocytes from thymus of tumor-bearing rats presented lower viability, but this change was abolished by fish oil supplementation. Tumor growth increased proliferation of lymphocytes from all lymphoid organs, and fish oil supplementation abolished this effect. Ex vivo production of TNF-alpha and IL-6 was reduced in supplemented animals, but IL-4 and IL-10 secretion was stimulated in both nontumor and tumor-bearing rats. IL-10 and IFN-gamma plasma levels was also decreased in supplemented animals. These results suggest that the anticachetic effects of fish oil supplementation for a long period of time (2 generations) in Walker 256 tumor-bearing rats may be associated to a decrease in lymphocyte function as demonstrated by reduced viability, proliferation capacity, and cytokine production.

Glucose metabolism by lymphocytes, macrophages, and tumor cells from Walker 256 tumor-bearing rats supplemented with fish oil for one generation.

Here we investigated the effect of lifelong supplementation of the diet with coconut fat (CO, rich in saturated fatty acids) or fish oil (FO, rich in n-3 polyunsaturated fatty acids) on tumor growth and lactate production from glucose in Walker 256 tumor cells, peritoneal macrophages, spleen, and gut-associated lymphocytes. Female Wistar rats were supplemented with CO or FO prior to mating and then throughout pregnancy and gestation and then the male offspring were supplemented from weaning until 90 days of age. Then they were inoculated subcutaneously with Walker 256 tumor cells. Tumor weight at 14 days in control rats (those fed standard chow) and CO supplemented was approximately 30 g. Supplementation of the diet with FO significantly reduced tumor growth by 76%. Lactate production (nmol h(-1) mg(-1) protein) from glucose by Walker 256 cells in the group fed regular chow (W) was 381.8 +/- 14.9. Supplementation with coconut fat (WCO) caused a significant reduction in lactate production by 1.6-fold and with fish oil (WFO) by 3.8-fold. Spleen lymphocytes obtained from W and WCO groups had markedly increased lactate production (553 +/- 70 and 635 +/- 150) when compared to non-tumor-bearing rats ( approximately 260 +/- 30). FO supplementation reduced significantly the lactate production (297 +/- 50). Gut-associated lymphocytes obtained from W and WCO groups increased lactate production markedly (280 +/- 31 and 276 +/- 25) when compared to non-tumor-bearing rats ( approximately 90 +/- 18). FO supplementation reduced significantly the lactate production (168 +/- 14). Lactate production by peritoneal macrophages was increased by tumor burden but there was no difference between the groups fed the various diets. Lifelong consumption of FO protects against tumor growth and modifies glucose metabolism in Walker tumor cells and lymphocytes but not in macrophages.

Glutamine concentration and immune response of spinal cord-injured rats.

BACKGROUND/OBJECTIVES: Glutamine plays a key role in immune response. Spinal cord injury (SCI) leads to severe loss of muscle mass and to a high incidence of infections. This study investigated the acute effect of SCI (2 and 5 days) on the plasma glutamine and skeletal muscle concentrations and immune responses in rats. METHODS: A total of 29 adult male Wistar rats were divided as follows: control (C; n = 5), sham-operated (S2; n = 5) and spinal cord-transected (T2; n = 7). They were killed on day 2 after surgery/transection (acute phase). Another set was sham-operated (S5; n = 5), spinal cord-transected (T5; n = 7), and killed at day 5 after surgery/transection (secondary phase). Blood was collected; the white portion of the epitrochlearis and gastrocnemius muscles and the red portion of soleus muscles were dissected to measure the glutamine concentration. Gut-associated lymphocytes and peritoneal macrophages were obtained for immune parameters measurements. RESULTS: Glutamine concentration in the plasma, gastrocnemius, and soleus muscles in rats with SCI were significantly reduced but not in the epitrochlearis muscle in the acute (2 days) and secondary (5 days) phases. Phagocytic response was reduced in the acute phase but increased in the secondary phase in rats with SCI. Superoxide production, on the other hand, was significantly increased at days 2 and 5 after SCI, and CD8+ lymphocytes subset decreased significantly on days 2 and 5. CONCLUSIONS: Our results showed reduction in plasma glutamine and skeletal muscle concentrations after spinal cord transection. They also suggest that SCI and glutamine reduction contribute to an alteration in immune competence.

Effect of fish oil supplementation for 2 generations on changes in macrophage function induced by Walker 256 cancer cachexia in rats.

The effect of coconut fat (rich in medium saturated fatty acids) or fish oil (rich in omega-3 polyunsaturated fatty acids) supplementation for 2 generations on tumor growth, cancer cachexia, animal survival and macrophage function was investigated in Walker 256 tumor-bearing rats. Female Wistar rats were supplemented with coconut fat or fish oil prior to mating and then throughout pregnancy and gestation. Both supplementations were daily and orally given at 1 g per kg body weight as a single bolus. Same treatment was performed by the 2 following generations. At 90 days of age, male offspring (50%) from F2 generation were subcutaneously inoculated with 2 x 10(7) Walker 256 tumor cells. At 14 days after tumor implantation, rats not supplemented displayed cancer cachexia characterized by loss of body weight, hypoglycemia, hyperlacticidemia, hypertriglyceridemia, decreased food intake and depletion of glycogen stores in the liver and skeletal muscles. Supplementation with coconut fat did not affect these parameters. However, supplementation with fish oil decreased tumor growth (59%), prevented body weight loss and food intake reduction and attenuated cancer cachexia. In addition, fish oil increased animal survival up to 20 days (from 25% in rats not supplemented to 67% in rats supplemented with fish oil) and improved macrophage function characterized by increased phagocytosis capacity and production of hydrogen peroxide and nitric oxide. These results suggest that fish oil supplementation for 2 generations improves macrophage function in association to reduced tumor growth and attenuated cancer cachexia, maintaining food intake and increasing animal survival.

Ratio of n6 to n-3 fatty acids in the diet affects tumor growth and cachexia in Walker 256 tumor-bearing rats.

In this study we investigate the impact of the dietary ratio of n-6 to n-3 fatty acids (FAs) from postweaning until adult age upon tumor growth, lipid peroxidation in tumor tissue, and metabolic indicators of cancer cachexia in Walker 256 tumor-bearing rats. Weanling male Wistar rats received a normal low-fat (40 g/kg diet) chow diet or high-fat diets (300 g/kg) that included fish oil (FO) or sunflower oil or blends of FO and sunflower oil to yield n-6 to n-3 FA ratios of approximately 6:1, 30:1, and 60:1 ad libitum. After 8 wk, half of each group was inoculated with 1 ml of 2 x 10(7) Walker 256 cells. At the 14th day after tumor inoculation, the animals were killed, and tumors and blood were removed. The different diets did not modify the blood parameters in the absence of tumor bearing, except the high-FO diet, which decreased serum cholesterol and triacylglycerol concentrations. Tumor weight in chow-fed rats was 19 g, and these rats displayed cancer cachexia, characterized by hypoglycemia, hyperlacticidemia, hypertriacylglycerolemia, loss of body weight, and food intake reduction. Tumor weight in FO-fed rats was 7.7 g, and these animals gained body weight (14.6 g) and maintained blood metabolic parameters similar to non-tumor-bearing animals. Tumor weight in rats fed the diet with an n-6 to n-3 FA ratio of 6:1 was similar to tumor-bearing, chow-fed rats, but they gained 2 g in the body weight and blood metabolic parameters were similar to those in non-tumor-bearing rats. However, a further increase in the n-6 FA content of the diet did not change the cachectic state associated with tumor bearing. In this experimental model, a dietary n-6 to n-3 FA ratio of 6:1 was able to increase food intake and body weight, restore the biochemical blood parameters of cachexia, and prevent the development of cancer cachexia.

Fish oil supplementation in F1 generation associated with naproxen, clenbuterol, and insulin administration reduce tumor growth and cachexia in Walker 256 tumor-bearing rats.

Weanling female Wistar rats were supplemented with fish oil (1 g/kg body weight) for one generation. The male offspring received the same supplementation until to adult age. Rats supplemented with coconut fat were used as reference. Some rats were inoculated subcutaneously with a suspension (2 x 10(7) cells/mL) of Walker 256 tumor. At day 3, when the tumor was palpable, rats were treated with naproxen (N) (0.1 mg/mL), clenbuterol (Cb) (0.15 mg/kg body weight), and insulin (I) (10 U/kg body weight). At day 14 after tumor inoculation, the animals were killed. Tumor was removed and weighed. Blood, liver, and skeletal muscles were also collected for measurements of metabolites and insulin. In both tumor-bearing untreated rats and tumor-bearing rats supplemented with coconut fat, tumor growth, triacylglycerol, and blood lactate levels were higher, and glycogen content of the liver, blood glucose, cholesterol and HDL-cholesterol levels were lower as compared with the non-tumor-bearing and fish oil supplemented groups. Fish oil supplementation of tumor-bearing rats led to a partial recovery of the glycogen content in the liver and a full reversion of blood glucose, lactate, cholesterol, and HDL-cholesterol levels. The treatment with N plus Cb plus I attenuated cancer cachexia and decreased tumor growth in both coconut fat and fish oil supplemented rats. In conclusion, chronic fish oil supplementation decreased tumor growth and partially recovered cachexia. This beneficial effect of fish oil supplementation was potentiated by treatment with naproxen plus clenbuterol plus insulin.

Lifelong exposure to dietary fish oil alters macrophage responses in Walker 256 tumor-bearing rats.

Supplementation of the diet with fish oil (FO) decreases growth of the Walker 256 tumor and decreases the cachexia associated with tumor-bearing. The mechanisms by which FO inhibits tumor growth and cachexia are unknown. Macrophages are very important in host defence against tumors since they produce several anti-tumor agents which in turn have been shown to be modified by dietary FO, but rarely in the setting of tumor bearing and never in relation to lifelong exposure. In this study, we compared the effects of supplementation of the diet of pregnant and lactating rats and subsequent supplementation of the offspring with coconut fat or FO on macrophage activities involved in anti-tumor defence. FO supplementation was able to induce an increase in phagocytosis, in O2-, H2O2, nitric oxide, and TNF-alpha production by macrophages and in lysosomal volume in non-tumor-bearing rats. However, phagocytosis, production of O2- and H2O2 and lysosomal volume were not affected by the FO diet when rats were bearing tumors, although nitric oxide production was higher in these animals. It appears that tumor bearing activates the innate immune system and that dietary FO has little effect on innate immunity in the presence of Walker 256 tumors. Thus, it is still unclear how FO decreases the growth of Walker 256 tumors and the associated cachexia.

Cancer cachexia and tumor growth reduction in Walker 256 tumor-bearing rats supplemented with N-3 polyunsaturated fatty acids for one generation.

In this study we investigated the effect of lifelong supplementation of the diet with coconut oil (CO, rich in saturated fatty acids) or fish oil (FO, rich in n-3 polyunsaturated fatty acids, PUFAs) on tumor growth, animal survival, and metabolic indicators of cachexia in adult rats. Female Wistar rats were supplemented with CO or FO prior to mating and then throughout pregnancy and gestation, and then the male offspring were supplemented from weaning until 90 days of age. Then they were inoculated subcutaneously with Walker 256 tumor cells. Tumor weight at 14 days in control rats (those fed standard chow) was approximately 20 g. These animals displayed cancer cachexia, which was characterized by loss of weight, hypoglycemia, hyperlacticidemia, hypertriacylglycerolemia, and depletion of glycogen stores. Supplementation of the diet with CO did not change these parameters, except that there was a smaller decrease in serum triacylglycerol concentration. Supplementation of the diet with FO significantly decreased tumor growth (by approximately 60%), increased survival (50% at 30 days postinoculation vs. 30% in the controls and 13.5% in the CO group), and prevented the fall in body weight. Furthermore, FO supplementation partly abolished the fall in serum glucose, totally prevented the elevation in serum lactate concentrations, partly prevented the hypertriacylgylcerolemia, and preserved tissue glycogen stores. Lifelong consumption of FO, rich in n-3 PUFAs, protects against tumor growth and cancer cachexia and improves survival.