Manuel A. de Pablo

Modulatory effects of dietary lipids on immune system functions.

Dietary lipid manipulation may affect a great number of immune parameters, such as lymphocyte proliferation, cytokine synthesis, natural killer (NK) cell activity, phagocytosis and so on. The immunomodulation induced by dietary fatty acids may be applied in the amelioration of inflammatory disorders, such as autoimmune diseases. However, the mechanisms that participate in these processes are still poorly understood. It is probable that modulation of immune system by fatty acids of the diet may occur by alteration of membrane fluidity, lipid peroxide formation, eicosanoid production or regulation of gene expression. However, recent studies have reported the effects of several free fatty acids on apoptosis induction of in vitro cultures. In fact, a possible explanation of the effects that fatty acids promote on the immune system cells could be associated with an apoptotic process performed in an irreversible way. In vivo studies have demonstrated the ability of fatty acids to alter the survival of animals fed diets containing oils and infected with a pathogenic bacterium. Experimental infection in animals fed dietary lipids produces a modification of resistance to micro‐organisms. The present review analyses all of these parameters that dietary fatty acids are capable of altering in order to modify the immune response. Further studies will be needed to establish the mechanisms involved in immune system regulation, reduction of symptoms derived from autoimmune pathologies and so on.

Dietary Antioxidants: Immunity and Host Defense

Natural antioxidants may be defined as molecules that prevent cell damage against free radicals and are critical for maintaining optimum health in both animals and humans. In all living systems, cells require adequate levels of antioxidant defenses in order to avoid the harmful effect of an excessive production of reactive oxygen species (ROS) and to prevent damage to the immune cells. During the inflammatory processes, the activation of phagocytes and/or the action of bacterial products with specific receptors are capable of promoting the assembly of the multicomponent flavoprotein NADPH oxidase, which catalyzes the production of high amounts of the superoxide anion radical (O(2)(-)). Under these particular circumstances, neutrophils and macrophages are recognized to produce superoxide free radicals and H(2)O(2), which are essential for defence against phagocytized or invading microbes. In this state, antioxidants are absolutely necessary to regulate the reactions that release free radicals. Antioxidant nutrients commonly included in the diet such as vitamin E, vitamin C, β-carotene, selenium, copper, iron and zinc improve different immune function exhibiting an important protective role in infections caused by bacteria, viruses or parasites. As a result, dietary antioxidants have been related to modulate the host susceptibility or resistance to infectious pathogens. Overall, numerous studies have suggested that the development of tolerance, and control of inflammation are strongly correlated with specific immune mechanisms that may be altered by an inadequate supply of either macronutrients or micronutrients. Therefore, the present paper will review the effects of dietary antioxidants on immune cell function and the impact on protection against infectious microorganisms.

The antimicrobial peptide cecropin A induces caspase-independent cell death in human promyelocytic leukemia cells

Most antimicrobial peptides have been shown to have antitumoral activity. Cecropin A, a linear 37-residue antimicrobial polypeptide produced by the cecropia moth, has exhibited cytotoxicity in various human cancer cell lines and inhibitory effects on tumor growth. In this study, we investigated the apoptosis induced by cecropin A in the promyelocytic cell line HL-60. Treatment of cells with cecropin A was characterized by loss of viability in a dose-dependent manner, lactate dehydrogenase (LDH) leakage, and modest attenuation of lysosomal integrity measured by neutral red assay. An increase of reactive oxygen species (ROS) generation, DNA fragmentation, and phosphatidylserine externalization were quantified following cecropin A exposure at a concentration of 30 microM, whereas cecropin A-induced apoptosis was independent of caspase family members, because the activity of caspase-8 and -9 were irrelevant. Nevertheless, caspase-3 activity showed a significant increase at concentrations of 20-40 microM, but a considerable reduction at 50 microM. Flow cytometry analysis revealed a dissipation of the mitochondrial transmembrane potential (Deltapsi(m)), and the accumulation of cells at sub-G1 phase measured by FACS analysis of propidium iodide (PI) stained nuclei suggested induction of apoptosis. Morphological changes measured by Hoechst 33342 or acridine orange/ethidium bromide staining showed nuclear condensation, corroborating the apoptotic action of cecropin A. Overall, these data indicate that cecropin A is able to induce apoptosis in HL-60 cells through a signaling mechanism mediated by ROS, but independently of caspase activation.

In vitro biological activities of magainin alone or in combination with nisin.

Antimicrobial peptides have received increasing attention not only as potential candidates to their administration as antimicrobial agents, but also as potential drugs applied in cancer therapy. Here, we have examined the action of both nisin and magainin on human promyelocytic leukemia HL-60 cells. Cells were cultured in presence of either nisin or magainin 1 as well as in combination with both nisin and magainin 1. Results have revealed that magainin, but not nisin, produces a loss of cell viability in HL-60 cells, and a minor increase of hemolysis, whereas it is not responsible for cell membrane disruption and lactate dehydrogenase (LDH) leakage. In addition, magainin is involved in a significant generation of reactive oxygen species (ROS), as well as in an augment of caspase-3 activity. Magainin-induced apoptosis was verified by DNA fragmentation and annexin V-FITC/propidium iodide (PI) staining of the cells. Promotion of cell death by magainin occurs via cytochrome c release accompanied by a substantial increase of proteasome activity. These results underline the importance of magainin as a drug capable of exerting an in vitro antitumoral activity by triggering apoptosis.

Relevance of Dietary Lipids as Modulators of Immune Functions in Cells Infected with Listeria monocytogenes

ABSTRACT Nutritional status may have significant importance for the immune system, and particularly, unsaturated fatty acids may serve as modulators of immune functions. Clinical and epidemiological studies have demonstrated that fatty acids are involved in the reduction of the inflammatory processes that occur in diseases characterized by an overactivation of the immune system. At the same time, an increase in susceptibility to infection has also been reported. The importance of immune system modulation by dietary lipids in the presence of an intracellular bacterial pathogen, such as Listeria monocytogenes, was evaluated in the present study. BALB/c mice were divided into four groups which were each fed a low-fat (2.5% by weight) diet, an olive oil (OO; 20% by weight) diet, a fish oil (FO; 20% by weight) diet, or a hydrogenated coconut oil (HCO; 20% by weight) diet for 4 weeks. In each group, lymphocye proliferation was measured, and a reduction in the stimulation index was observed in the FO and HCO groups. Cytotoxicity exerted by L. monocytogenes was increased in the groups fed diets containing OO and FO after 6 h of incubation with the bacterium. An important increase in the production of reactive oxygen species was found in the groups fed the HCO diet after 12 h of incubation with L. monocytogenes. Finally, invasion and adhesion factors were not modified substantially by the action of dietary lipids, although these factors were reduced in cells from mice fed an FO diet. These results underline the importance of several dietary lipids as biological modulators of immune functions and their crucial role in the alteration of host natural resistance.

Changes in the immune functions and susceptibility to Listeria monocytogenes infection in mice fed dietary lipids

The direct examination of the effects that fish oil diets (composed of long‐chain n‐3 polyunsaturated fatty acids) exert on immune system function indicates a reduction of host natural resistance to infectious diseases mainly because of a suppression of immune function generated by the fatty acids contained in this diet. Here, we evaluated the concentration of IL‐12, IL‐4, prostaglandin E2 and leukotriene B4 in the serum from BALB/c mice receiving four different diets. Each group was fed a diet that differed only in the source of fat: a low‐fat diet (2.5% by weight), an olive oil diet (20% by weight), a fish oil diet (20% by weight) or a hydrogenated coconut oil diet (20% by weight). Mice were fed for 4 weeks and then infected with the intracellular pathogen Listeria monocytogenes. An initial reduction in the Th1‐type response as a result of a decrease in IL‐12p70 secretion, an inefficient action of IL‐4 (Th2‐type response) and no modification of pro‐inflammatory lipid‐mediator production could be, at least in part, the key events responsible for the inadequate elimination of L. monocytogenes from the spleens of mice fed a fish oil diet. Furthermore, our results suggest that the type of dietary lipids may affect the circulating concentration of IL‐12p70 and IL‐4, leading to a modulation in the protective cellular immune response to L. monocytogenes infection.

Significance of olive oil in the host immune resistance to infection

The effects exerted by polyunsaturated fatty acids (PUFA) on immune system functions have been investigated in recent years. These studies have reported the important role that n-3 PUFA play in the diminution of incidence and severity of inflammatory disorders. Nevertheless, less attention has been paid to the action of monounsaturated fatty acids (MUFA) upon the immune system. The administration of a diet containing a high amount of olive oil in experimental animals produces a suppression of lymphocyte proliferation, an inhibition of cytokine production and a reduction in natural killer (NK) cell activity. Despite these alterations in immune functions, it has been reported that olive oil-rich diets are not as immunosuppressive as fish oil diets. An important aspect in immunonutrition is focused on the relationship between fats, the immune system and host resistance to infection, particularly when these nutrients are supplied to patients at risk of sepsis. Different studies have determined that olive oil-rich diets do not impair the host resistance to infection. Therefore, olive oil constitutes a suitable fat that may be applied in clinical nutrition and administered to critically ill patients. In the present review, we summarize the current knowledge on olive oil and immune system functions, the biological consequences derived from the administration of diets containing olive oil and the impact of olive oil on immune defence.

Orally administered Lactobacillus plantarum reduces pro-inflammatory interleukin secretion in sera from Listeria monocytogenes infected mice

Lactic acid bacteria have traditionally been thought to have immunomodulating effects. To verify this property, Lactobacillus plantarum was orally administered to mice (5 x 107 colony forming units (c.f.u.)), prior to infection with Listeria monocytogenes in order to evaluate the host resistance against an infectious micro-organism and to better define the influence of L. plantarum on such responses. Balb/c mice were treated daily with L. plantarum or received PBS (sham-treated mice as controls) for 4 weeks. Subsequently, mice were intravenously infected with a clinical isolate of L. monocytogenes. Our study revealed that the administration of L. plantarum did not significantly increase the survival (P = 0.13) of mice (fifteen in each group) after L. monocytogenes infection (106 c.f.u./ml), whereas a sub-lethal dose of L. monocytogenes (105 c.f.u./ml) was eliminated from liver and spleen 5 d after the challenge in both L. plantarum- and sham-treated mice (n 5). Nevertheless, the levels of IL-1beta and IL-6 from sera of orally administered L. plantarum were drastically reduced at 0, 4 (P < 0.01) and 6 d after L. monocytogenes infection, whereas TNF-alpha production was unaltered. In conclusion, administration of L. plantarum reduced pro-inflammatory IL production after challenge with L. monocytogenes, although it did not significantly impact the survival of mice. We speculate that L. plantarum could exert anti-inflammatory effects, which may represent an important model to reduce inflammatory disorders. Therefore, further studies in human subjects should determine the role of L. plantarum as an immunomodulatory micro-organism and its relationship in the host protection to pathogens.

Assessment of Interleukin-12, Gamma Interferon, and Tumor Necrosis Factor Alpha Secretion in Sera from Mice Fed with Dietary Lipids during Different Stages of Listeria monocytogenes Infection

ABSTRACT Recent experimental observations have determined that long-chain n-3 polyunsaturated fatty acids suppress immune functions and are involved in the reduction of infectious disease resistance. BALB/c mice were fed for 4 weeks with one of four diets containing either olive oil (OO), fish oil (FO), hydrogenated coconut oil, or a low fat level. Interleukin-12p70 (IL-12p70), gamma interferon (IFN-γ), and tumor necrosis factor alpha (TNF-α) production in the sera of mice fed these diets and challenged with Listeria monocytogenes were determined by enzyme-linked immunosorbent assay. In addition, bacterial counts from spleens of mice were carried out at 24, 72, or 96 h of infection. Here, we quantified an initial diminution of production of both IL-12p70 and IFN-γ, which appear to play an important role in the reduction of host resistance to L. monocytogenes infection. In addition, an efficient elimination of L. monocytogenes was observed in spleens of mice fed a diet containing OO at 96 h of infection, despite reductions in IL-12p70 and TNF-α production, suggesting an improvement of immune resistance. Overall, our results indicate that the initial reduction of both IL-12 and IFN-γ production before L. monocytogenes infection represents the most relevant event that corroborates the impairment of immune resistance by n-3 polyunsaturated fatty acids during the different stages of infection. However, we speculate that the modulation of other cytokines must be also involved in this response, because the alteration of cytokine production in mice fed an FO diet in a late phase of L. monocytogenes infection was similar to that in mice fed OO, whereas the ability to eliminate this bacterium from the spleen was improved in the latter group.

Inflammatory and phagocytic response to experimental Campylobacter jejuni infection in mice.

After intraperitoneal inoculation with Campylobacter jejuni BALB/c, Swiss and DBA mice show a peritoneal inflammatory response of different intensity. Only BALB/c mice have a strong peritoneal response. Simultaneous intraperitoneal inoculation of C. jejuni plus FeCl3 increase both inflammatory response and phagocytic activity in Swiss mice, without production of diarrhea. Some thermostable compounds of C. jejuni have a very strong chemotactic activity against peritoneal cells of mice, whereas a diffusible, thermolabile and glutaraldehyde‐resistant factor has an inhibitory effect over murine peritoneal cell phagocytosis. Bactericidal activity of peritoneal cells increased after in vitro re‐challenge with C. jejuni. Bacteremia is present in all the mice strains tested, but the clearance is quick in DBA and slow in BALB/c and Swiss mice. These experiments confirm that in mice, peritoneal non‐specific mechanisms of defense, such as macrophages, play an important role in order to control C. jejuni infection.