Alessandra Folador

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.

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.

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 ω‐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 × 107 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.

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-α (TNF-α), interferon-γ (IFN-γ), 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-α 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-γ 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.

Physical Training Attenuates the Stress-Induced Changes in Rat T-Lymphocyte Function

Backgorund/Aims: Modulations in the immune function by stress are a well-known phenomenon. Acute restraint stress may induce impaired T-lymphocyte responses. Moderate physical training is associated with beneficial effects on immunological functions. We investigated the effects of a moderate physical training on T-lymphocyte function in rats submitted to acute restraint stress. Methods: Thirty male Wistar rats weighing 210–226 g were randomly divided into four groups: non-trained rats (NT, n = 7), and non-trained rats submitted to stress (NT + S, n = 8); trained rats (T, n = 7), and trained rats submitted to stress (T + S, n = 8). Trained rats were submitted to a program of moderate running over a period of 8 weeks. Rats subjected to restraint stress were kept immobilized in glass cylinders (8 cm in diameter and 24 cm long) during 60 min. Plasma corticosterone concentration, peripheral blood leukocyte number, indicators of apoptosis of T lymphocytes in blood and lymphoid organs, and mitogen-induced proliferation of T lymphocytes in lymphoid organs were evaluated. Results: Acute stress exposure raised plasma corticosterone concentration (p < 0.001), but not in previously trained animals. Restraint stress induced an increase in the percentage of lymphocytes in apoptosis, and a decrease in the concanavalin-A-induced proliferation of lymphocytes from the thymus and lymph nodes, and an increase in lymphocytes of the spleen. Neither of these alterations was observed in trained animals submitted to acute restraint stress. Conclusions: Our data confirm that acute restraint stress is associated with changes in T-lymphocyte function. Moreover, moderate physical training attenuates the effects of acute stress by a mechanism that involves the hypothalamic-pituitary-adrenal axis and an increase in tolerance of leukocytes.