Elizabeth A. Miles

Is deficiency of interferon gamma production by allergen triggered cord blood cells a predictor of atopic eczema

Abstract. Peripheral blood mononuclear cell proliferative responses and interferon‐γ product ion to anti‐CD3, cat fur extract, betalactoglobulin and ovalbumin were determined al birth in a group of 34 babies born to families where at least one parent was; atopic. The development of atopic eczema with positive allergy skin‐prick tests to cows’ milk and egg at 1 year of age, where the symptoms improved on an egg and milk‐free diet, was significantly associated with raised proliferative responses and defective IFN‐γ production to stimulation with betalactoglobulin with a trend for similar responses to ovalbumin. This was not observed in those who did not develop atopic eczema or those with atopic eczema not associated with foods. Responses to cat fur extract were not significantly different between those with and without atopic eczema. This has important implications for the prediction at birth, not only of the probability of being allergic, but also of the specific allergens which will cause problems. The implementation of specific targeted allergen avoidance during the critical period in infancy, therefore, should be more easily applied and should facilitate attempts to prevent disease.

Fatty acids and lymphocyte functions

The immune system acts to protect the host against pathogenic invaders. However, components of the immune system can become dysregulated such that their activities are directed against host tissues, so causing damage. Lymphocytes are involved in both the beneficial and detrimental effects of the immune system. Both the level of fat and the types of fatty acid present in the diet can affect lymphocyte functions. The fatty acid composition of lymphocytes, and other immune cells, is altered according to the fatty acid composition of the diet and this alters the capacity of those cells to produce eicosanoids, such as prostaglandin E2, which are involved in immunoregulation. A high fat diet can impair lymphocyte function. Cell culture and animal feeding studies indicate that oleic, linoleic, conjugated linoleic, gamma-linolenic, dihomo-gamma-linolenic, arachidonic, alpha-linolenic, eicosapentaenoic and docosahexaenoic acids can all influence lymphocyte proliferation, the production of cytokines by lymphocytes, and natural killer cell activity. High intakes of some of these fatty acids are necessary to induce these effects. Among these fatty acids the long chain n-3 fatty acids, especially eicosapentaenoic acid, appear to be the most potent when included in the human diet. Although not all studies agree, it appears that fish oil, which contains eicosapentaenoic acid, down regulates the T-helper 1-type response which is associated with chronic inflammatory disease. There is evidence for beneficial effects of fish oil in such diseases; this evidence is strongest for rheumatoid arthritis. Since n-3 fatty acids also antagonise the production of inflammatory eicosanoid mediators from arachidonic acid, there is potential for benefit in asthma and related diseases. Recent evidence indicates that fish oil may be of benefit in some asthmatics but not others.

Fetal peripheral blood mononuclear cell proliferative responses to mitogenic and allergenic stimuli during gestation

Blood samples were obtained from fetuses and premature babies (n=51) (15‐34 weeks gestation) to determine at what stage the fetal immune system was able to produce a positive proliferative response to common allergens. Peripheral blood mononuclear cells (PB MC) were stimulated with the mitogen, phytohaemagglutinin (PHA), and the allergens, house dust mite, cat fur. birch tree pollen, β‐lactoglobulin, ovalbumin and bee venom (mellitin). Results were expressed as ratios of stimulated to unstimulated 3H thymidine incorporation, and as percent positive responders. There was an increase in proliferation ratio which correlated with increasing gestational age for PHA (p < 0.0001), cat fur (p=0.042), birch pollen (p=0.022) and β‐lactoglobulin (p=0, 006). The point in gestation when cells from some individuals began responding to the allergens with a ratio of 2. 0 was at approximately 22 weeks. PBMC proliferative response ratios were higher from samples from babies > 22 weeks gestation compared to < 22 weeks for the mitogen and all allergens, except mellitin. There was also a greater proportion of positive responders from samples > 22 weeks compared to < 22 weeks for the mitogen and all allergens, except mellitin. Maternal exposure to birch pollen, which has a discrete season, was assessed to determine whether exposure had occurred at 22 weeks gestation or beyond. Results showed a higher prolifera tive response in infant cells stimulated with birch pollen (p=0.005) and higher proportion of positive responders (p=0.01) in the group of babies whose mothers had been exposed to hirch pollen beyond 22 weeks, compared to those whose mothers had not been so exposed. These results suggest that in utero fetal exposure to an allergen from around 22 weeks gestation may result in primary sensitisation to that allergen, leading to positive proliferative responses, at birth.

Influence of dietary supplementation with long-chain n−3 or n−6 polyunsaturated fatty acids on blood inflammatory cell populations and functions and on plasma soluble adhesion molecules in healthy adults

Greatly increasing the amounts of flaxseed oil [rich in α-linolenic acid (ALNA)] or fish oil (FO); [rich in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] in the diet can decrease inflammatory cell functions and so might impair host defense. The objective of this study was to determine the effect of dietary supplementation with moderate levels of ALNA, γ-linolenic acid (GLA), arachidonic acid (ARA), DHA, or FO on inflammatory cell numbers and functions and on circulating levels of soluble adhesion molecules. Healthy subjects aged 55 to 75 yr consumed nine capsules per day for 12 wk. The capsules contained placebo oil (an 80∶20 mix of palm and sunflowerseed oils) or blends of placebo oil with oils rich in ALNA, GLA, ARA, or DHA or FO. Subjects in these groups consumed 2 g ALNA; approximately 700 mg GLA, ARA, or DHA; or 1 g EPA plus DHA (720 mg EPA+280 mg DHA) daily from the capsules. Total fat intake from the capsules was 4 g per day. None of the treatments affected inflammatory cell numbers in the bloodstream; neutrophil and monocyte phagocytosis or respiratory burst in response to E. coli; production of tumor necrosis factor-α, interleukin-1β, and interleukin-6 in response to bacterial lipopolysaccharide; or plasma concentrations of soluble intercellular adhesion molecule-1. In contrast, the ALNA and FO treatments decreased the plasma concentrations of soluble vascular cell adhesion molecule-1 (16 and 28% decrease, respectively) and soluble E-selectin (23 and 17% decrease, respectively). It is concluded that, in contrast to previous reports using higher amounts of these fatty acids, a moderate increase in consumption of long-chain n−6 or n−3 polyunsaturated fatty acids does not significantly affect inflammatory cell numbers or neutrophil and monocyte responses in humans and so would not be expected to cause immune impairment. Furthermore, we conclude that moderate levels of ALNA and FO, which could be incorporated into the diet, can decrease some markers of endothelial activation and that this mechanism of action may contribute to the reported health benefits of n−3 fatty acids.

Influence of marine n-3 polyunsaturated fatty acids on immune function and a systematic review of their effects on clinical outcomes in rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease of the joints and bones. The n-6 polyunsaturated fatty acid (PUFA) arachidonic acid (ARA) is the precursor of inflammatory eicosanoids which are involved in RA. Some therapies used in RA target ARA metabolism. Marine n-3 PUFAs (eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)) found in oily fish and fish oils decrease the ARA content of cells involved in immune responses and decrease the production of inflammatory eicosanoids from ARA. EPA gives rise to eicosanoid mediators that are less inflammatory than those produced from ARA and both EPA and DHA give rise to resolvins that are anti-inflammatory and inflammation resolving, although little is known about these latter mediators in RA. Marine n-3 PUFAs can affect other aspects of immunity and inflammation relevant to RA, including dendritic cell and T cell function and production of inflammatory cytokines and reactive oxygen species, although findings for these outcomes are not consistent. Fish oil has been shown to slow the development of arthritis in animal models and to reduce disease severity. A number of randomised controlled trials of marine n-3 PUFAs have been performed in patients with RA. A systematic review included 23 studies. Evidence is seen for a fairly consistent, but modest, benefit of marine n-3 PUFAs on joint swelling and pain, duration of morning stiffness, global assessments of pain and disease activity, and use of non-steroidal anti-inflammatory drugs.

Dietary fatty acids influence the production of Th1- but not Th2-type cytokines

C57Bl6 mice were fed for 6 weeks on a low‐fat diet or on high‐fat diets containing coconut oil (rich in saturated fatty acids), safflower oil [rich in n‐6 polyunsaturated fatty acids (PUFAs)], or fish oil (rich in n‐3 PUFAs) as the main fat sources. The fatty acid composition of the spleen lymphocytes was influenced by that of the diet fed. Thymidine incorporation into concanavalin A‐stimulated spleen lymphocytes and interleukin (IL)‐2 production were highest after feeding the coconut oil diet. Interferon (IFN)‐γ production was decreased by safflower oil or fish oil feeding. IL‐4 production was not significantly affected by diet, although production was lowest by lymphocytes from fish oil‐fed mice. The ratio of production of Th1‐ to Th2‐type cytokines (determined as the IFN‐γ/IL‐4 ratio) was lower for lymphocytes from mice fed the safflower oil or fish oil diets. After 4 h of culture, IL‐2 mRNA levels were higher in cells from mice fed coconut oil, and IFN‐γ mRNA levels were higher in cells from mice fed coconut oil or safflower oil. After 8 h of culture, IL‐2, IFN‐γ, and IL‐4 mRNA levels were lowest in cells from mice fed fish oil. The ratio of the relative levels of IFN‐γ mRNA to IL‐4 mRNA was highest in cells from mice fed coconut oil and was lowest in cells of mice fed fish oil. The influence of individual fatty acids on IL‐2 production by murine spleen lymphocytes was examined in vitro. Although all fatty acids decreased IL‐2 production in a concentration‐dependent manner, saturated fatty acids were the least potent and n‐3 PUFAs the most potent inhibitors, withn‐6 PUFAs falling in between in terms of potency. It is concluded that saturated fatty acids have minimal effects on cytokine production. In contrast, PUFAs act to inhibit production of Th1‐type cytokines with little effect on Th2‐type cytokines; n‐3 PUFAs are particularly potent. The effects of fatty acids on cytokine production appear to be exerted at the level of gene expression.

Effect of low-to-moderate amounts of dietary fish oil on neutrophil lipid composition and function

Although essential to host defense, neutrophils are also involved in numerous inflammatory disorders including rheumatoid arthritis. Dietary supplementation with relatively large amounts of fish oil [containing >2.6 g eicosapentaenoic acid (EPA) plus 1.4 g docosahexaenoic acid (DHA) per day] can attenuate neutrophil functions such as chemotaxis and superoxide radical production. In this study, the effects of more moderate supplementation with fish oil on neutrophil lipid composition and function were investigated. The rationale for using lower supplementary doses of fish oil was to avoid adverse gastrointestinal problems, which have been observed at high supplementary concentrations of fish oil. Healthy male volunteers aged <40 yr were randomly assigned to consume one of six dietary supplements daily for 12 wk (n=8 per treatment group). The dietary supplements included four different concentrations of fish oil (the most concentrated fish oil provided 0.58 g EPA plus 1.67 g DHA per day), linseed oil, and a placebo oil. The percentages of EPA and DHA increased (both P<0.05) in neutrophil phospholipids in a dose-dependent manner after 4 wk of supplementation with the three most concentrated fish oil supplements. No further increases in EPA or DHA levels were observed after 4 wk. The percentage of arachidonic acid in neutrophil phospholipids decreased (P<0.05) after 12 wk supplementation with the linseed oil supplement or the two most concentrated fish oil supplements. There were no significant changes in N-formyl-met-leu-phe-induced chemotaxis and superoxide radical production following the dietary supplementations. In conclusion, low-to-moderate amounts of dietary fish oil can be used to manipulate neutrophil fatty acid composition. However, this may not be accompanied by modulation of neutrophil functions such as chemotaxis and superoxide radical production.

Differential effects of short-chain fatty acids on proliferation and production of pro- and anti-inflammatory cytokines by cultured lymphocytes

Short-chain fatty acids (SCFA) are produced by fermentation of water-soluble fiber by anaerobic bacteria in the large bowel. Fiber-rich diets decrease the risk of developing inflammatory bowel disease (IBD) and butyrate enemas are effective as a therapy in some patients. Crohns disease, one form of IBD, appears to involve an exagerated T helper-1 (Th1) lymphocyte phenotype, characterised by production of interleukin (IL)-2 and interferon (IFN)-gamma, that drives the inflammation. To examine whether SCFA influence pro- and anti-inflammatory cytokine production, rat mesenteric lymph node lymphocytes were cultured in the presence of acetate (10 mM), butyrate (1.5 mM) or propionate (2 mM) and the production of cytokines in response to concanavalin A determined. Butyrate, but not acetate or propionate, inhibited lymphocyte proliferation and IL-2 production. Acetate and propionate were able to partly prevent the inhibitory effect of butyrate on IL-2 production. Acetate and propionate increased IFN-gamma production, whereas butyrate inhibited it. Acetate and propionate in combination were able to prevent the inhibitory effect of butyrate on IFN-gamma production. IL-4 was not detected in any cultures. Acetate and propionate increased IL-10 production, which was not affected by butyrate. It is concluded that butyrate significantly inhibits Th1-type responses and that this might explain the therapeutic effect of butyrate in IBD patients. Acetate and propionate have less marked modulatory actions, and in some cases have effects that oppose those of butyrate. A combination of the three SCFA causes a shift in the T helper lymphocyte phenotype towards a more anti-inflammatory phenotype and this might explain the protective effects of fiber.

Comparison of the effects of linseed oil and different doses of fish oil on mononuclear cell function in healthy human subjects

Studies on animal and human subjects have shown that greatly increasing the amount of linseed (also known as flaxseed) oil (rich in the n-3 polyunsaturated fatty acid (PUFA) alpha-linolenic acid (ALNA)) or fish oil (FO; rich in the long-chain n-3 PUFA eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)) in the diet can decrease a number of markers of immune function. The immunological effects of more modest doses of n-3 PUFA in human subjects are unclear, dose-response relationships between n-3 PUFA supply and immune function have not been established and whether ALNA has the same effects as its long-chain derivatives is not known. Therefore, the objective of the present study was to determine the effect of enriching the diet with different doses of FO or with a modest dose of ALNA on a range of functional responses of human monocytes and lymphocytes. In a randomised, placebo-controlled, double-blind, parallel study, forty healthy males aged 18-39 years were randomised to receive placebo or 3.5 g ALNA/d or 0.44, 0.94 or 1.9 g (EPA+DHA)/d in capsules for 12 weeks. The EPA:DHA ratio in the FO used was 1.0:2.5. ALNA supplementation increased the proportion of EPA but not DHA in plasma phospholipids. FO supplementation decreased the proportions of linoleic acid and arachidonic acid and increased the proportions of EPA and DHA in plasma phospholipids. The interventions did not alter circulating mononuclear cell subsets or the production of tumour necrosis factor-alpha, interleukin (IL) 1beta, IL-2, IL-4, IL-10 or interferon-gamma by stimulated mononuclear cells. There was little effect of the interventions on lymphocyte proliferation. The two higher doses of FO resulted in a significant decrease in IL-6 production by stimulated mononuclear cells. It is concluded that, with the exception of IL-6 production, a modest increase in intake of either ALNA or EPA+DHA does not influence the functional activity of mononuclear cells. The threshold of EPA+DHA intake that results in decreased IL-6 production is between 0.44 and 0.94 g/d.

Atopy risk in infants and children in relation to early exposure to fish, oily fish, or long-chain omega-3 fatty acids: a systematic review.

There are two main families of polyunsaturated fatty acids (PUFAs), the n−6 and the n−3 families. It has been suggested that there is a causal relationship between n−6 PUFA intake and allergic disease, and there are biologically plausible mechanisms, involving eicosanoid mediators of the n−6 PUFA arachidonic acid, that could explain this. Fish and fish oils are sources of long-chain n−3 PUFAs and these fatty acids act to oppose the actions of n−6 PUFAs. Thus, it is considered that n−3 PUFAs will protect against atopic sensitization and against the clinical manifestations of atopy. Evidence to examine this has been acquired from epidemiologic studies investigating associations between fish intake in pregnancy, lactation, infancy, and childhood, and atopic outcomes in infants and children and from intervention studies with fish oil supplements in pregnancy, lactation, infancy, and childhood, and atopic outcomes in infants and children. All five epidemiological studies investigating the effect of maternal fish intake during pregnancy on atopic or allergic outcomes in infants/children of those pregnancies concluded protective associations. One study investigating the effects of maternal fish intake during lactation did not observe any significant associations. The evidence from epidemiological studies investigating the effects of fish intake during infancy and childhood on atopic outcomes in those infants or children is inconsistent, although the majority of the studies (nine of 14) showed a protective effect of fish intake during infancy or childhood on atopic outcomes in those infants/children. Fish oil supplementation during pregnancy and lactation or during infancy or childhood results in a higher n−3 PUFA status in the infants or children. Fish oil provision to pregnant women is associated with immunologic changes in cord blood and such changes may persist. Studies performed to date indicate that provision of fish oil during pregnancy may reduce sensitization to common food allergens and reduce prevalence and severity of atopic dermatitis in the first year of life, with a possible persistence until adolescence with a reduction in eczema, hay fever, and asthma. Fish oil provision to infants or children may be associated with immunologic changes in the blood but it is not clear if these are of clinical significance and whether they persist. Fish oil supplementation in infancy may decrease the risk of developing some manifestations of allergic disease, but this benefit may not persist as other factors come into play. It is not clear whether fish oil can be used to treat children with asthma as the two studies conducted to date give divergent results. Further studies of increased long-chain n−3 PUFA provision in during pregnancy, lactation, and infancy are needed to more clearly identify the immunologic and clinical effects in infants and children and to identify protective and therapeutic effects and their persistence.