leucotriene; Th2, T-helper 2
Epidemiological and animal studies suggest that besides genetic factors, environmental exposures early in life are important determinants of health and disease later in life(Reference Barker1). Since the effects of the early exposures can be long-lasting, even persisting until adulthood, this phenomenon has been termed ‘early life programming’, ‘early life origins of health and disease’ or ‘developmental origins of heath and disease’(Reference Gluckman, Hanson and Cooper2). Nutrition has been identified as one source of early exposures that might influence early development and later phenotype(Reference Jackson3). There is substantial immune development in human subjects in utero and in the weeks and months after birth(Reference Jones, Holloway and Warner4–Reference Levy6), and it is possible that such development can be influenced by nutritional factors(Reference Calder, Krauss-Etschmann and de Jong7). However, relatively little attention has been devoted to the potential for early life programming of the immune system by dietary factors. Nevertheless a body of mainly epidemiological literature has developed, which associates temporal changes in the patterns of intake of n-6 and n-3 PUFA with temporal changes in the incidence and prevalence of atopic sensitisation or its clinical manifestations (allergies, atopic eczema, hayfever and allergic asthma), and there is a proposed molecular and cellular mechanism to explain the observed association(Reference Hodge, Peat and Salome8, Reference Black and Sharp9). In this article, the proposed mechanism that relates early exposure to n-6 or n-3 PUFA to increased or decreased risk of developing atopy will be described as will the literature relating early exposure to n-6 or n-3 PUFA or their main dietary sources to atopy or its manifestations or immune outcomes relevant to atopy.
Fatty acids: nomenclature, sources and intakes
Fatty acid structure and nomenclature have been described elsewhere(Reference Calder, Burdge, Nicolaou and Kafatos10). There are two principal families of PUFA, the n-6 and the n-3 families. The simplest members of each family, linoleic acid (LA; 18:2n-6) and α-linolenic acid (ALA; 18:3n-3), cannot be synthesised by mammals. LA is found in significant quantities in many vegetable oils, including corn, sunflower and soyabean oils, and in products made from such oils, such as margarines. ALA is found in green plant tissues, in some common vegetable oils, including soyabean and rapeseed oils, in some nuts, and in flaxseed (also known as linseed) and flaxseed oil. Between them, LA and ALA contribute over 95%, and perhaps as much as 98% of dietary PUFA intake in most Western diets,(Reference Calder, Burdge, Nicolaou and Kafatos10) with LA intake being in excess of that of ALA. The intake of LA in Western countries increased greatly over the second half of the 20th century, following the introduction and marketing of cooking oils and margarines(Reference Calder, Burdge, Nicolaou and Kafatos10, 11). ALA intake probably changed little over this time. Typical intakes of both essential fatty acids are in excess of requirements. However, the changed pattern of consumption of LA has resulted in a marked increase in the ratio of n-6 to n-3 PUFA in the diet. This ratio is currently between 5 and 20 in most Western populations(Reference Calder, Burdge, Nicolaou and Kafatos10, Reference Burdge and Calder12).
Although LA and ALA cannot be synthesised by human subjects they can be metabolised to other fatty acids (Fig. 1). This is achieved by the insertion of additional double bonds into the acyl chain (i.e. unsaturation) and by elongation of the acyl chain. Thus, LA can be converted via γ-linolenic acid (18:3n-6) and di-homo-γ-linolenic acid (20:3n-6) to arachidonic acid (AA; 20:4n-6) (Fig. 1). By an analogous set of reactions catalysed by the same enzymes, ALA can be converted to EPA (20:5n-3). Both AA and EPA can be further metabolised, EPA giving rise to docosapentaenoic acid (22:5n-3) and DHA (22:6n-3) (Fig. 1). Dietary intakes of the longer-chain, more unsaturated PUFA are much lower than those of LA and ALA(Reference Calder, Burdge, Nicolaou and Kafatos10, 11, 13). AA is found in meat and offal and intakes are estimated at 50–500 mg/d. EPA and DHA are found in fish, especially so-called ‘oily’ fish (tuna, salmon, mackerel, herring and sardine). One oily fish meal can provide between 1·5 and 3·5 g of these long-chain n-3 PUFA(13). Commercial products known as fish oils also contain these long-chain n-3 PUFA, which typically will contribute about 30% of the fatty acids present. Thus, consumption of a typical one gram fish oil capsule per day can provide about 300 mg of these fatty acids. In the absence of oily fish or fish oil consumption, intake of long-chain n-3 PUFA is likely to be <100 mg/d(Reference Calder, Burdge, Nicolaou and Kafatos10, 11, 13), although foods fortified with these fatty acids are now available in many countries.
n-6 PUFA, eicosanoids, inflammatory processes and atopy
PUFA play roles ensuring the correct environment for membrane protein function, maintaining membrane fluidity and regulating cell signalling, gene expression and cellular function(Reference Calder, Burdge, Nicolaou and Kafatos10). Through these actions PUFA can influence the functioning of immune cells(Reference Yaqoob and Calder14–Reference Calder16) and so could impact on the development and manifestations of atopy(Reference Calder and Miles17, Reference Calder18). However, the key link between PUFA and immunological processes related to atopy is that the eicosanoid family of mediators is derived from 20-C PUFA. Because immune cells typically contain a high proportion of the n-6 PUFA AA and low proportions of other 20-C PUFA, AA is usually the major substrate for eicosanoid synthesis. Eicosanoids, which include PG, thromboxanes (TX), leucotrienes (LT) and other oxidised derivatives, are generated from AA by the action of cyclooxygenase (COX) and lipoxygenase (LOX) enzymes (Fig. 2). These enzymes are expressed in inflammatory and epithelial cells and give rise to a mix of mediators, depending upon the nature of cell types present and the nature, timing and duration of the stimulus(Reference Nicolaou, Nicolaou and Kafatos19,20–Reference Tilley, Coffman and Koller22). Eicosanoid mediators are involved in modulating the intensity and duration of inflammatory responses. Through actions on dendritic cells, T cell differentiation and Ig class switching in B cells, some eicosanoids (e.g. PGE2) are believed to play a role in promoting sensitisation to allergens. Through their actions on inflammatory cells, smooth muscles and epithelial cells, some eicosanoids are strongly implicated in different immunologic features and clinical manifestations of atopic disease. Indeed, allergic inflammation in animal models is associated with increased PG and LT production. However, inhibition of COX-1 or COX-2 or knockout of either COX results in augmented allergic inflammation with increased T-helper 2 (Th2)-type cytokine production and increased airway reactivity (see Moore et al.(Reference Moore and Peebles23) and Park & Christman(Reference Park and Christman24)). This suggests that the overall effect of PG is to restrain allergic inflammation. However, individual PG might enhance or inhibit allergic inflammation, depending upon their specific action. One current view is that PGD2, PGF2α and thromboxane A2 increase allergic inflammation, whereas PGE2 and PGI2 inhibit it (see Moore et al.(Reference Moore and Peebles23) and Park & Christman(Reference Park and Christman24)). PGD2 is produced mainly by mast cells and activated macrophages. It is a potent bronchoconstrictor, promotes vascular permeability, and activates eosinophils and a Th2-type response. Thromboxane A2 is a bronchoconstrictor and stimulates acetylcholine release. PGE2 is a vasodilator, increases vascular permeability, inhibits the production of T-helper 1-type cytokines and primes naïve T cells to produce IL-4 and IL-5. PGE2 also promotes Ig class switching in uncommitted B cells towards the production of IgE. Despite these effects of PGE2, it is now considered that this eicosanoid is protective towards airway inflammation(Reference Moore and Peebles23, Reference Park and Christman24). It is possible that PGE2 promotes sensitisation via its effects on T cell phenotype and B cells, but is protective against the subsequent manifestations of inflammation upon re-exposure to allergen. PGI2 appears to suppress Th2 lymphocyte activity and eosinophil recruitment. LTB4 is chemotactic for leucocytes, increases vascular permeability, induces the release of lysosomal enzymes and reactive oxygen species by neutrophils and of inflammatory cytokines (e.g. TNF-α) by macrophages, and promotes IgE production by B cells. The cysteinyl LT (LTC4, D4 and E4) may be either vasoconstrictors or vasodilators, depending upon the situation and the location of their synthesis. They cause smooth muscle contraction and bronchoconstriction, increase vascular permeability and eosinophil recruitment, and promote mucus secretion. PGE2 inhibits 5-LOX activity, so down-regulating LT production(Reference Levy, Clish and Schmidt25). Furthermore PGE2 induces 15-LOX leading to production of lipoxin A4, which is anti-inflammatory(Reference Vachier, Chanez and Bonnans26–Reference Gewirtz, Collier-Hyams and Young28). These effects highlight the antagonist nature of eicosanoids and may underlie, at least in part, the protective effect of PGE2 in allergic inflammation.
The foregoing discussion has led to suggestions that there is a causal link between the increased intake of the n-6 PUFA LA over the second half of the 20th century and the incidence and prevalence of atopy and its clinical manifestations and that the link is mediated via increased potential to produce pro-atopic and pro-allergic eicosanoids from AA(Reference Hodge, Peat and Salome8, Reference Black and Sharp9). This link is shown in Fig. 3.
Evidence relating high early n-6 PUFA exposure to increased risk of atopic outcomes in infancy or childhood
There is some ecological, epidemiological and case v. control evidence associating high LA intake with atopy or its manifestations. Differences in the prevalence of asthma and allergic rhinitis and in blood concentrations of allergen-specific IgE between former East and West Germany accorded with differences in butter and margarine consumption(Reference von Mutius, Martinez and Fritzsch29). Differences in the prevalence of bronchial asthma, allergic rhinitis and atopic dermatitis among Finnish schoolchildren were related to levels of LA in plasma cholesteryl esters, an indicator of dietary LA intake(Reference Poysa, Korppi and Pietikainen30). Margarine consumption among German schoolchildren was associated with a greater risk of hayfever compared with not consuming margarine (OR=2·0 after adjustment for other factors)(Reference von Mutius, Weiland and Fritzsch31). Margarine consumption was higher in Australian schoolchildren with atopic dermatitis or with other manifestations of atopic disease compared with controls(Reference Dunder, Kuikka and Turtinen32). High PUFA consumption was associated with increased risk of recent asthma compared with low PUFA consumption among Australian schoolchildren (OR=2·03)(Reference Haby, Peat and Marks33). Margarine consumption was associated with increased risks of allergic sensitisation and of allergic rhinitis v. no margarine, although this effect was seen in boys only(Reference Bolte, Frye and Hoelscher34). Polyunsaturated oil consumption was associated with increased risk of wheeze in Swedish children (OR=1·91)(Reference Kim, Elfman and Mi35). A high dietary ratio of n-6 to n-3 PUFA was associated with increased risk of asthma in Australian schoolchildren (OR=1·93; after adjustment for other factors=2·89)(Reference Oddy, de Klerk and Kendall36). These studies all associate dietary intake and disease at the same point in time and none attempt to associate early LA exposure with later disease. However, some studies report that LA is higher in breast milk consumed by infants who go on to develop atopy in infancy, although not all such studies have found this (reviewed in Sala-Vila et al.(Reference Sala-Vila, Miles and Calder37). Umbilical cord lipids from neonates who go on to develop atopy in early childhood contain a higher amount of LA than normal (see Sala-Vila et al.(Reference Sala-Vila, Miles and Calder37)). Thus there is some evidence to support some aspects of the chain of events shown in Fig. 3.
n-3 PUFA eicosanoids and inflammatory processes
Increased consumption of long-chain n-3 PUFA such as EPA and DHA (usually given as fish oil) results in increased proportions of those fatty acids in inflammatory cell phospholipids(Reference Yaqoob, Pala and Cortina-Borja38–Reference Rees, Miles and Banerjee40). The incorporation of EPA and DHA into human inflammatory cells occurs in a dose–response fashion and is partly at the expense of AA(Reference Yaqoob, Pala and Cortina-Borja38–Reference Rees, Miles and Banerjee40). Since there is less substrate available for synthesis of eicosanoids from AA, fish oil supplementation of the human diet has been shown to result in decreased production of AA-derived eicosanoids by inflammatory cells (see Calder(Reference Calder41) for references) (Fig. 4). EPA is also able to act as a substrate for COX and LOX enzymes, giving rise to eicosanoids with a slightly different structure to those formed from AA. Thus, fish oil supplementation of the human diet has been shown to result in increased production of 5-series LT by inflammatory cells(Reference Calder41). The functional significance of this is that the mediators formed from EPA are believed to be less potent than those formed from AA. For example, LTB5 is 10- to 100-fold less potent as a neutrophil chemotactic agent than LTB4 (see Calder(Reference Calder41)). In addition to long-chain n-3 PUFA modulating the generation of eicosanoids from AA and to EPA acting as substrate for the generation of alternative eicosanoids, recent studies have identified a novel group of mediators, termed E- and D-series resolvins, formed from EPA and DHA, respectively, that appear to exert anti-inflammatory and inflammation-resolving actions (see Serhan et al.(Reference Serhan, Arita and Hong42) and Serhan(Reference Serhan43) for reviews). In recent studies in ovalbumin-sensitised Balb/C mice, administration of resolvin E1 was found to decrease airway eosinophil and lymphocyte recruitment, production of the Th2 cytokine IL-13, circulating ovalbumin-specific IgE and airway hyper-responsiveness to inhaled methacholine(Reference Aoki, Hisada and Ishizuka44) and to promote the resolution of inflammatory airway responses by directly suppressing the production of IL-23 and IL-6 in the lung(Reference Haworth, Cernadas and Yang45).
The above considerations have led to the idea that while a high exposure to n-6 PUFA (or low exposure to n-3 PUFA) will promote atopy (both sensitisation and manifestations), a high exposure to n-3 PUFA will be protective(Reference Hodge, Peat and Salome8, Reference Black and Sharp9).
Evidence relating high early n-3 PUFA exposure to decreased risk of atopic outcomes in infancy or childhood
A small number of studies report that EPA and DHA are lower in breast milk consumed by infants who go on to develop atopy in infancy, although not all studies find this (reviewed in Sala-Vila et al.(Reference Sala-Vila, Miles and Calder37)). Umbilical cord blood lipids from neonates who go on to develop atopy in early childhood appear to contain lower than normal amounts of EPA and DHA (see Sala-Vila et al.(Reference Sala-Vila, Miles and Calder37)). All five studies investigating the effect of maternal fish intake during pregnancy on atopic or allergic outcomes in infants/children of those pregnancies concluded protective associations(Reference Sausenthaler, Koletzko and Schaaf46–Reference Calvani, Alessandri and Sopo50). The protective effect varied between 25 and 95%, which might be attributed to differences in study design, exposure and outcome measure classification and assessment. Nine studies observed a beneficial effect of fish intake during infancy/childhood and atopic outcomes in those infants/children(Reference Dunder, Kuikka and Turtinen32,35,51–Reference Alm, Aberg and Erdes57). The reduction in atopy/allergy risk among these studies ranged between 22 and 80%. It is important to note that two studies observed a negative effect of fish intake on childhood atopy(Reference Huang, Lin and Pan58, Reference Takemura, Sakurai and Honjo59), and three studies observed no associations(Reference Hijazi, Abalkhail and Seaton60–Reference Wijga, Smit and Kerkhof62).
Studies of maternal fish oil supplementation during pregnancy report effects on umbilical cord blood immune markers (blood cytokine mRNA(Reference Krauss-Etschmann, Hartl and Rzehak63), plasma cytokines(Reference Dunstan, Mori and Barden64), LTB4 production from neutrophils(Reference Prescott, Barden and Mori65), cytokine production by mononuclear cells(Reference Dunstan, Mori and Barden66)) and an altered cord blood haemopoietic progenitor phenotype(Reference Denburg, Hatfield and Cyr67). These immunologic effects might be expected to impact on allergic sensitisation and on the development of atopic disease. Indeed, Dunstan et al.(Reference Dunstan, Mori and Barden66) reported beneficial effects on atopic outcomes in one-year-old infants as a result of maternal fish oil supplementation during pregnancy (less severe atopic dermatitis, lower risk of positive skin prick test to egg). Olsen et al.(Reference Olsen, Osterdal and Salvig68) identified that fish oil supplementation in late pregnancy is associated with a marked reduction in atopic manifestations in the offspring at age 16 years, suggesting a long-term effect of any immunologic changes that occurred in pregnancy and early life of those children. A study of fish oil supplementation during both pregnancy and lactation showed expected effects on n-3 PUFA status and these were associated with differences in PGE2 production by stimulated maternal blood(Reference Warstedt, Furuhjelm and Duchen69). The latter might be expected to influence Th2 polarisation. Indeed, infants from mothers in the fish oil group had a reduced risk of developing allergic sensitisation to egg, IgE-associated eczema and food allergy during the first year of life(Reference Furuhjelm, Warstedt and Larsson70). A study of maternal fish oil supplementation during lactation(Reference Lauritzen, Kjaer and Fruekilde71) is the only one of these studies investigating immune outcomes in the offspring beyond birth. Infants of lactating mothers who received fish oil supplements had a higher n-3 PUFA status at 4 months of age and interferon-γ production at 2·5 years of age was higher in the fish oil group, an observation that may reflect faster maturation of the immune system. This study did not assess clinical outcomes.
One study has examined the long-term effect of fish oil supplementation of infants on atopy and its manifestations(Reference Mihrshahi, Peat and Marks72–Reference Almqvist, Garden and Xuan76). Fish oil supplementation from 6 months of age increased plasma n-3 PUFA status and decreased n-6 PUFA status at 18 months, 3 years and 5 years of age. At 18 months of age there was a decreased prevalence of wheeze in the fish oil group and higher plasma n-3 PUFA levels were associated with lower bronchodilator use(Reference Mihrshahi, Peat and Marks72, Reference Mihrshahi, Peat and Webb73). Follow-up at 3 years of age suggested that fish oil supplementation from infancy to childhood could reduce allergic sensitisation and airway disease at this early age, as the fish oil group had reduced cough, but not wheeze(Reference Peat, Mihrshahi and Kemp74). However, no effect of fish oil was seen on the other end points measured such as eczema, serum IgE concentration or doctor diagnosis of asthma. At 5 years of age there was no significant effect of fish oil on any of the clinical outcomes relating to lung function(Reference Marks, Mihrshahi and Kemp75), allergy(Reference Marks, Mihrshahi and Kemp75) or asthma(Reference Almqvist, Garden and Xuan76). Possible reasons for the lack of beneficial effects of long-chain n-3 PUFA at 5 years of age may be related to suboptimal adherence to and/or implementation of the intervention (50% and 56% compliance in the intervention and control group, respectively), as well as to the dose of fish oil used, loss to follow-up and lack of power.
Thus, there is quite good evidence that early exposure to n-3 PUFA induces immune effects that may be associated with reduced atopic sensitisation and with a reduction in allergic manifestations. However, data available from existing studies, which are of many different types, are not entirely consistent and so it is not possible to draw a firm conclusion at this stage. Clearly more studies in this area are needed and, where these are interventions, it is important that they be sufficiently powered, that they measure both immune and clinical outcomes where possible, and that the dose of n-3 PUFA and duration are carefully considered.
There are two main families of PUFA, the n-6 and the n-3 families. Intake of the n-6 PUFA LA increased over the second half of the 20th century, this increase coinciding with increased prevalence of atopy and its clinical manifestations. 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 AA, that could explain this. There is some evidence from association studies that high LA intake is linked with increased risk of atopic sensitisation and allergic manifestations. However, there is little evidence that early exposure to LA increases later risk, although there are supportive observations using cord blood and later outcomes. Fish and fish oils are sources of long-chain n-3 PUFA and these fatty acids act to oppose the actions of n-6 PUFA. Thus, it is considered that n-3 PUFA will protect against atopic sensitisation and against the clinical manifestations of atopy. Evidence to examine this has been acquired from epidemiological studies investigating associations between fish intake in pregnancy, infancy and childhood and atopic outcomes in infants and children and from intervention studies with fish oil supplements in pregnancy, lactation and infancy 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. 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 (9/14) showed a protective effect of fish intake during infancy or childhood on atopic outcomes in those infants/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 sensitisation to common food allergens and reduce the prevalence and severity of atopic dermatitis in the first year of life, with a possible persistence until adolescence with a reduction in eczema, hayfever and asthma. 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. Further studies of increased long-chain n-3 PUFA provision in pregnancy, lactation and infancy are needed to more clearly identify the immunologic and clinical effects in infants and children and to identify protective effects and their persistence.
P.C.C. has funding from the European Commission under Framework 6 for research in the area of seafood and human health (FOOD-CT-2006-16249; Sustainable aquafeeds to maximize the health benefits of farmed fish for consumers (Aquamax)) and L.-S.K., M.V., P.S.N. and E.A.M. are supported by this funding. P.C.C. has funding for research in the area of n-3 fatty acids from Abbott Nutrition and from Vifor Pharma and serves on the Scientific Advisory Board on Baby Nutrition of the Danone Research Centre for Specialised Nutrition. All authors had input into the writing of the manuscript.