Allergies are Th2 cytokine–mediated pathologies, involving IL-4–, IL-5– and IL-13–dependent amplification of innate effector cell populations acting together with antibodies and inflammatory mediators (50
). As Th1 and Th2 cell responses are mutually antagonistic, it has been argued that declining microbial stimulation of Th1 cell responses in the developed world has led to over-vigorous allergic Th2 cell reactions (51
). However, allergen-specific Th1 cells can in fact exacerbate airway inflammation (52
). Moreover, epidemiological reports increasingly link Th2 cell–inducing helminth parasite infections with reduced allergic disease in humans (53
), whereas nematode-infected mice display attenuated allergic responses (28
). Current theories postulate that pathogen-induced T reg cells control both Th1 and Th2 effector populations (2
). The possibility that T reg cells inhibit allergic disease has received growing support from both animal (13
) and human (14
We now provide direct experimental evidence to support the hypothesis that helminth infection down-regulates allergic reactions through the action of regulatory T cells rather than by altering the Th1–Th2 balance. To do this, we selected the murine intestinal nematode H. polygyrus
as a model of helminthiasis (28
), because it follows a purely enteric infective cycle and establishment; has the ability to establish stable, chronic infections; and evokes a Th2 cell–dominated immune response similar to that observed in general with gastrointestinal nematodes. We chose an allergic airway inflammation model that permited us to observe modifications in cellular recruitment, cytokine and chemokine expression, and tissue pathology. We also opted to study two allergens in parallel, which induce airway inflammation in different strains of mice: the environmental house dust mite allergen Der p 1, in C57BL/6 mice; and the experimental allergen, OVA in BALB/c mice.
Mice carrying an established helminth infection are shown here to be less prone to allergic airway inflammation, whether measured by the inflammatory infiltrate in BALF or by peribronchial and perivascular inflammation. Our results extend earlier work with H. polygyrus
that demonstrate a reduced allergy to food antigens in mice infected with this parasite (28
). More broadly, these findings can be translated to the human situation, in which individuals carrying active helminth infections appear, in epidemiological studies, to be less responsive to allergen provocation than coresidents free of infection (17
To address the mechanisms involved in helminth-induced suppression, we first established that the Th2-driving cytokine IL-4 was unaffected, as was the overall Th1–Th2 balance. However, the Th2 effector cytokines IL-5 and IL-13 were diminished, as was the inflammatory phase within the lung. Thus, Th2 cell–associated pathological reactions were suppressed within the context of a Th2 cell–inducing helminth infection. The fact that Th2 cell priming occurs normally is shown by the similar allergen-specific IgE responses in infected and uninfected mice, and the ability of the suppressive mechanism to target the effector phase is further demonstrated by the impact of infection on a previously sensitized allergy-prone host ( F). The hypothesis that T reg cell activity may be responsible for this phenomenon was supported by our finding that anti-CD25 antibody treatment reversed suppression. However, allergic inflammation was not restored by the administration of anti–IL-10R antibody, indicating that T reg cell activity in this setting is not IL-10 dependent.
We then used the adoptive transfer system of MLNC from infected mice into sensitized, but uninfected, hosts to demonstrate that down-regulation is mediated by T cells, primarily those with the CD4+
phenotype associated with T reg cells (61
). Several distinct regulatory phenotypes of T reg cell cells have been proposed, including “Th3” cells, primarily acting through TGF-β, which are most closely associated with the gut mucosal environment, and “Tr1” cells, which are capable of suppressing airway allergy through the action of IL-10 (13
). Interestingly, the H. polygyrus
-elicited T reg cells are generated in the Th3 environment, but manifest their function in the Tr1 environment.
Although the CD4+
subset was the most potent at suppressing allergic inflammation, some activity could also be observed in the CD4−
population, particularly in the C57BL/6 system. Although this may represent a CD8+
regulatory T cell (63
), we also have evidence that CD19+
B cell populations are able, on transfer, to exert a dampening effect on immunopathology. Perhaps, in the down-regulatory milieu of a chronic infection, additional non–T reg cell types are recruited to curtail pathology more completely.
The cells transferred from infected mice express elevated levels of IL-10 and TGF-β, the two principal T reg cell–associated down-regulatory cytokines, and contain a significantly higher number of CD4+
cells. However, despite the prominence of IL-10 production by H. polygyrus
-exposed T cells, their ability to suppress airway allergy is not mediated by IL-10, as MLNC from infected IL-10–/–
mice can mediate suppression, which is consistent with the failure of anti–IL-10R antibodies to reverse suppression in infected animals. In contrast, TGF-β remains a strong candidate for immune suppression by T reg cells from helminth-infected mice, particularly as this cytokine is known to alleviate experimental airway allergy (37
) and has the capacity to instruct peripheral T cells to develop regulatory capacity (46
T cells are generally associated with self-reactive regulatory cells that prevent autoimmune reactivities (61
). Hence, the expansion of CD4+
cell–mediated regulatory activity in infection raises the issue of the origin and antigen specificity of these cells (65
). Are there preexisting “natural” T reg phenotype cells that expand in response to infection, or can parasite antigen-specific regulatory cells arise from naive Th0 precursors? In this respect, the fact that transfer of suppression did not require renewed antigen stimulation was striking. One possibility is that the T reg cell population is self-reactive rather than parasite specific, and is restimulated by ligands in the recipient host. However, it has been shown that T reg cells specific for exogenous antigen (alloantigen or HGG), reactivated in a donor immediately before transfer, suppressed bystander allograft responses in a recipient without the need for antigen restimulation in the new host (67
). Thus, it is possible that chronic parasite infection maintains a high level of activation in T reg cells, sufficient for their function in our short-term experiments. As with other instances of T reg cell responses to pathogens (66
), these issues are currently under active investigation.
The transfer model also allows us to exclude changes in antibody production as a major mechanism for abatement of allergy. For example, the production of allergen-specific-IgE is comparable in mice who have received naive or infected MLNC, which show respectively normal and suppressed allergic reactions. There is also no rise in polyclonal IgE in recipients of infected CD4+
T cells, arguing that changes in either absolute allergen-specific IgE, or in the ratio of nonspecific to specific IgE, are not responsible for the diminution of allergic responses in infected animals. A similar conclusion was drawn from measurements of specific and total IgE in atopic and nonatopic humans harboring chronic schistosome infections (21
A final intriguing question is why helminth parasites such as H. polygyrus
and Schistosoma mansoni
) induce T reg cells. As argued elsewhere (27
), parasites that can exploit host down-regulatory networks are likely to gain advantage in the battle for long-term survival in the host. Much of the pathology encountered in helminth infection is immune mediated (69
), and a dampening of responsiveness would not necessarily compromise the host. However, the immune system may have evolved to operate optimally in the regulated environment of infection, and in our more hygienic environment, we are prone to overzealous reactions to innocuous targets, generating the rapidly increasing levels of allergy and autoimmunity being experienced in the developed world.