Allergy: when does it begin and where will it end?
Authors: S. L. Prescott
Journal: Allergy (2003)

Although established allergic immune responses have been well characterized, relatively little is known about how or why these develop. There is also still no definitive explanation for the enormous increase in affected individuals. The manifestation of allergic symptoms within months (or even weeks) of life strongly suggests that early events (possibly during gestation) are contributing to this. However, although expanding research in this area has provided some important information about early immune development, the most critical questions remain unanswered. These issues are discussed here, in the context of our existing knowledge and the new observation by Kihlström et al. published in this issue (1) that sensitization to inhaled allergen (birch pollen) is not related to antenatal exposure to this allergen. There is still no clear answer to this, and immune development is likely to vary between individuals according to both genetic and environmental influences. Collectively, the literature supports intuitive notions that early events are important particularly during gestation and early postnatal life when immune response to the environment are presumably initiated [as discussed recently (2)]. There has been the assumption that, because allergic (Type 2) immune responses are directed towards allergens, that events during first allergen encounter are the most critical. This has also focused great attention on the detection of allergen-specific immune responses as early in life as possible, without a clear understanding of normal processes or the significance of these responses. While allergen-specific immune responses have been detected as early as 22 weeks (3), it is likely that other early immunological processes, independent of early allergen-specific events, may contribute or predispose to subsequent allergic disease. Potential environmental exposures and maternal influences during gestation and perinatal life may exert effects through yet poorly defined immunological pathways. This emphasises the need to explore this issue widely, and think beyond the presence or absence of allergen-specific responses. Many potential influences could operate well before the ‘22-week’ time-point, influencing immune development and susceptibility before specific responses to allergens are evident. Clearly the expression of disease is regulated and possibly even initiated in the organs subsequently affected (and associated lymphoid tissue). This highlights the limitations of using cord blood (CBMC) or peripheral blood (PBMC) mononuclear cell responses to assess complex immunological processes, many of which are likely to be organ-specific. Although allergens could enter the fetal circulation through the placenta, exposure through the fetal gut (and lung) has also been proposed (4). Events at mucosal surface are known to be important for initiating and informing immune development in the postnatal period, and may also be initiated in utero. Allergens have been detected in amniotic fluid (5) and cells capable of presenting antigens are present in the fetal gut (4). Proteins involved in the activation of APC and pro-Th1 signaling (such as sCD14) have also been detected in amniotic fluid and are reduced in those who go on to develop atopic dermatis (6). Although studies of cord blood have been of great value, they cannot provide information about these events, and it may also be difficult to extrapolate animal models to the human dilemma. Firstly, it has not been clear if the fetal T cells responding to allergen in vitro have been primed by allergen exposure in vivo. Fetal responses to allergens have been documented extensively by many groups (reviewed recently by 7) and appear to be of fetal origin (8). This has lead to obvious speculation that allergens encountered by the mother during gestation may be reaching the fetus in some way. While this seems feasible for food allergens, which can cross the placenta in animals (9), transfer of aeroallergens has been more difficult to conceptualise given the much lower levels of these proteins. To address whether allergens can cross the placenta in humans, Szepfalusi et al. (10) used a novel ex vivo placenta perfusion model to demonstrate that both food and inhaled allergens can pass from the maternal to the fetal circulation. This is not necessarily physiological and the transfer of aeroallergens was not as consistent. Thus, the issue of whether the fetus is directly exposed to levels of allergens sufficient to induce T cell priming in vivo remains unresolved. Secondly, the significance of fetal responses has been unclear. Because of controversy about whether the fetus is exposed to allergens, other explanations for these responses have been considered. In most cases, CBMC responses are to allergens to which mother have almost universal exposure (such as food allergens and perennial aeroallergens). However, in a number of situations CBMC responses to specific stimuli have been observed when there was no previous documented exposure. This includes responses to tetanus (11) and diphtheria toxoid (12) when there was no apparent exposure to these antigens in pregnancy. It has been proposed that these responses may therefore be nonspecific or because of cross-reactivity to other antigens (13). There is also no direct evidence for this. It is not clear what ‘cross-reacting’ antigens could initiate these responses, or why these would be generated. A further explanation for this observation has been that these responses are not true ‘memory’ responses, but some form of primary response by reactive clones. In many ways this also seems unlikely, as the precursor frequency of un-primed T cells is likely to be extremely low and generate responses below the threshold of most assays. Given the relative immaturity of neonatal T cells and the susceptibility to tolerance, strong allergen-specific primary responses also seem unlikely. Furthermore, T cell cytokine responses are evident (albeit lower) within a similar time-frame as adult memory responses. Although the strongest neonatal responses are seen when measuring cytokines [such as interleukin-6 (IL-6) and IL-10] derived from the non-T cell fraction which do not require previous exposure, T cell specific cytokines (such as IL-13 and IL-5) are also readily detectable within 24–48 h of allergen stimulation [(8) and others]. Regardless of exposure, there has also been controversy about whether the fetus is capable of generating classical ‘memory’ responses as detected in adults using traditional lymphoproliferation assays. There are many clear differences between adult and neonatal T cells, raising concerns that ‘positive’ responses in the fetus may not have the same meaning. The fact that most (more than 90%) of fetal T cells are of the CD45RA+ ‘naïve’ phenotype (14) has raised doubts about the likelihood of a significant repertoire of T cell ‘memory’. However, one recent study (15) found that 50% of CBMC responses to grass pollen were mediated by T cells of the CD45RO+ ‘memory’ phenotype, which are presumed to have been previously stimulated. More significantly, it is not clear why the remaining 50% of responses were mediated by CD45RA+ cells which are assumed to be ‘naïve’. As we previously discussed (16), for many reasons it is probably inappropriate to assume that neonatal CD4 + CD45RO + T cells are equivalent to adult T cells with the same ‘memory’ markers. Thus, further studies are required to determine the origin and significance of neonatal immune responses to allergens. It is not clear if these responsive clones subsequently contribute to the memory pool. Whatever their origin, these responses are commonly observed and appear to be part of a normal developmental process. Rather than indicating sensitization these early responses may be part of the development of normal tolerance. There are no studies which show a convincing relationship between levels of maternal allergen exposure (and presumed fetal exposure) and the presence, magnitude or character of fetal immune responses to these allergens. Smilie et al. (11) did not find any relationship between house dust mite (HDM) exposure in pregnancy and CBMC lymphoproliferation. Marks et al. also found no relationship between HDM levels in maternal beds at 36-week gestation and neonatal IL-4, IL-5, IL-10 and interferon γ (IFNγ) responses to HDM (17). Others have even found negative associations between neonatal HDM allergen-induced proliferation and maternal HDM exposure (18). While there are many studies linking allergic sensitization with early postnatal allergen exposure (with the recent exception of pets), there is still considerable controversy about associations with antenatal allergen exposure. In this issue, a retrospective study by Kihlström et al. (1) addressed this question by comparing clinical outcomes (allergic disease and sensitization) in two small cohorts of children with different levels of maternal birch pollen in pregnancy. Taking advantage of an unusually high pollen count in Stockholm during the spring of 1993, Kihlström et al. compared outcomes in infants of mothers exposed to high levels of birch pollen while pregnant during this season (born autumn 1993) to those with significantly lower maternal exposure the following season (born autumn 1994). They did not observe any significant differences in allergic symptoms and sensitisation at 4 years of age. Although there were some trends for more frequent sensitisation to birch with ‘high’ maternal exposure, this did not approach statistical significance. There is no way of determining that these seasonal differences resulted in major differences in fetal exposure. Even if birch pollen allergens reach the fetus, these would presumably be at extremely low levels, and it is unclear if seasonal fluctuations could change this substantially. This situation cannot be extrapolated to animal models of ‘high dose’ or ‘low dose’ antigen exposure in which tolerance in which high antigen exposure induces tolerance. It is also not clear whether there are ‘more critical periods’ during gestation when exposures are potentially more significant. It is also important to recognize that numerous potential confounding lifestyle factors and other environmental exposures, can make interpretation difficult in population-based studies of this nature. This is likely to be compounded by retrospective data collection. However these limitations are extremely difficult to overcome in human studies, which can still provide some valuable information. These issues aside, this study supports a growing body of evidence that antenatal allergen exposure is not a key factor in the development of atopic sensitization [(18) and others]. In keeping with earlier studies, Kihlström et al. also observed that postnatal allergen exposure was more strongly related to clinical outcomes than antenatal exposure (1). This reinforces the notion that antenatal allergen exposure is not a detrimental process, and allergen avoidance during pregnancy may have little benefit in preventing allergy. It is not clear when Type 2 responses, which are normal in the neonate, become pathological and lead to sensitization and disease. It is also not clear when (or if) these immune responses become ‘committed’ to a particular pattern of reactivity. Furthermore, although a number of subtle differences have been observed when comparing high risk neonates (of atopic parents) and low risk neonates (of nonatopic parents), these have been conflicting, and are not of predictive value. More recent studies suggest that atopic risk (maternal atopy) is associated with higher numbers of Type 2 helper cell CD4+IL13+ cells in cord blood (19) and stronger neonatal IL-13 responses to milk allergen (β lactoglobulin) (20). Atopic disease (at 1 year) has also been associated with higher neonatal polyclonal IL-13 responses (21) or increased numbers of CD4+IL13+ cells in cord blood (19). Reduced capacity for Th1 IFNγ responses is another key functional difference in high risk neonates (22–24 and others), but the significance of this is still unclear. Although, this may be because of APC immaturity resulting in reduced pro-Th1 signaling, there is no direct proof for this. There may also be function differences in the capacity of T cells of high risk infants to respond to IL-12 and other pro-Type 1 signals from APC, as observed recently by Janefjord et al. (25). A great deal more information is needed to understand how these early processes contribute to established allergic responses. Although allergen exposure in pregnancy does not appear to be a major factor determining the development of subsequent allergic disease, it is likely that other early environmental influences have important effects on immune development. This is in keeping with the growing sense that allergens may not play a major causal role in the recent increase in allergic disease, but that other environmental changes are contributing to this process. Many potential exposures are likely to have greater effects during formative periods of development when response patterns are being established. Thus, although allergen exposure in utero may not be a major issue, immune interactions in this early period remain fundamentally important in defining allergy pathogenesis.