Type 2 immunity and wound healing: evolutionary refinement of adaptive immunity by helminths

Epithelial alarmins elicit type 2 immunity

Although helminth molecules with direct T_H2 cell-inducing capacity might exist^17,21, the large size of the parasite and the consequent tissue damage that it causes during invasion might be the most important factor in the initiation of a type 2 immune response. This idea is supported by studies showing that helminth invasion, and the associated tissue damage, induces the release of several epithelium-derived cytokine alarmins that can initiate type 2 immunity^{22– 24}. One of the first such alarmins that was shown to have an important role in helminth-induced type 2 immunity was thymic stromal lymphopoietin (TSLP)²⁵; it was suggested in a recent study that TSLP directs type 2 immunity partly by eliciting a population of functionally distinct IL-4-producing basophils²⁶. TSLP also promotes type 2 immunity by actively suppressing the production of IL-12 by dendritic cells²⁷. The role of TSLP in eliciting type 2 immunity varies in different helminth infections because some parasites bypass the need for TSLP by directly suppressing IL-12 expression²⁷. In these cases, the type 2 immune response might be a default response or it might rely on other alarmins.

IL-33 is a cytokine alarmin that is released from the nucleus of necrotic epithelial cells, endothelial cells and fibroblasts. The IL-33 receptor complex — interleukin-1 receptor-like 1 (IL-1RL1; also known as ST2)–IL-1R accessory protein (IL-1RAP) — has also been shown to initiate type 2 responses following helminth infection^28,29. A similar type 2-inducing role has been identified for the IL-17 family cytokine IL-25 (also known as IL-17E)^30,31. Thus, multiple cytokine alarmins that are released by epithelial cells and other cells are involved in the initiation of protective type 2 immune responses during macroparasite infection; the relative importance of each mediator is probably dictated by the pathogen and by its mode of entry into the host.

Mechanisms that mimic helminth invasion

Recent studies have shown that type 2 immunity is activated by large inert particulate structures that can cause direct cell and tissue damage. In particular, inert silica³² and titanium microparticles³³ induce innate type 2 immune responses and have been successfully used as adjuvants to promote antigen-specific T_H2 cell responses through pathways that are independent of Toll-like receptor 4 (TLR4) and the signalling adaptor molecule myeloid differentiation primary-response protein 88 (MYD88)³³. One explanation for why these relatively large particulate structures induce type 2 immunity is that they share essential features with helminths. They are both large structures that might induce cell and tissue damage in the absence of type 1-inducing PAMPs; for example, ingested microparticles might induce cellular damage in phagocytes through the release of lysosomal enzymes following lysosome rupture and, consequently, this might activate endogenous danger signals that mediate tissue damage³².

In addition, the T_H2-inducing adjuvant aluminium hydroxide (alum) might induce the release of the damage-associated molecular pattern (DAMP) uric acid³⁴. Uric acid crystals that are released from damaged tissues drive type 2 immunity through inflammasome-independent pathways^32,34. Furthermore, extracellular ATP, which is presumably released by damaged cells, binds to P2 purinergic receptors that induce the release of IL-33 in the lungs³⁵, resulting in the generation of type 2 immunity. Thus, in this case, the ATP released from damaged tissues functions as a DAMP to induce the secretion of cytokine alarmins that, in turn, stimulate a type 2 immune response. Importantly, no single cellular DAMP has yet been shown to be essential for the development of helminth-induced responses, which suggests that a range of redundant pathways are involved in sensing helminth-induced danger. Indeed, multiple causes of tissue damage, including helminth-derived proteases and insect venoms, might contribute to the activation of type 2 immune responses (BOX 2).

Strong support for the importance of injury alone in the induction of type 2 immunity comes from the evidence that innocuous antigens, when exposed to the immune system in the context of experimentally induced cutaneous tissue damage, can stimulate potent type 2 immune responses³⁶. Therefore, injury and type 2 immunity are closely linked. The capacity of helminths to induce injury might explain an ancient association of metazoan parasites with the type 2 response, leading to the evolution of an adaptive type 2 immune response that protects against the consequences of helminth infection.

Mechanisms that amplify type 2 immunity

The data above show that tissue stress and damage, in the absence of strong TLR signalling, rapidly stimulates type 2 immunity. The combination of tissue stress and damage might be one mechanism by which the host recognizes helminth invasion and responds with an appropriate protective response. The anti-helminth response is further enhanced and polarized by the addition of individual helminth excretory–secretory products that condition dendritic cells to suppress TLR signals and the associated protein machinery that is required to prime T_H1 cells^37–39. This potent combination might be essential to induce the highly polarized and potent adaptive type 2 immune response that develops following helminth infection.

The cytokine alarmins are important for inducing the recruitment of group 2 innate lymphoid cells (ILC2s), which secrete large quantities of IL-5 and IL-13 (REFS 40–42), that might then activate anti-parasite effector responses^43,44. ILC2s, along with multipotent progenitor type 2 cells²², eosinophils, mast cells and basophils, are rapidly recruited to the site of infection and function as important innate sources of IL-4, IL-5 and IL-13. Eosinophils and basophils in particular are important producers of IL-4, which further amplifies and regulates both the innate and adaptive branches of type 2 immunity^45,46.

Memory responses augment type 2 immunity

Effector memory T cells (which rapidly secrete cytokines at high levels) and the production of IgE and IgG by B cells can increase resistance to many helminths^50–52. Shortly after infection, many of these parasites migrate through tissues, such as the lungs, where they are actively targeted by immune cells, before they enter the lumen of the small intestines⁵³. Fcε receptors expressed by mast cells bind to parasite-specific IgE, and the subsequent crosslinking of bound IgE by helminth antigens induces degranulation and the release of mediators from mast cells that, in some cases, contribute to parasite expulsion, as has been suggested by the immune response to Trichinella spiralis⁵⁴. The IgE-mediated rapid degranulation of mast cells might also be important in the defence against other macroparasites such as ticks, as localized tissue oedema that is mediated by the molecules released during degranulation might impede parasite invasion^55,56. Amphiregulin is also produced by mast cells following signalling from the high-affinity Fc receptor for IgE (FcεR1)⁵⁷; this highlights a potential link between the IgE-mediated memory response and wound repair. Thus, a very rapid and sometimes complex immune response is needed to induce expulsion or killing and simultaneously to repair the damage caused by these large macroparasites.

Control of helminth-induced tissue damage

Effective wound repair requires both the direct reconstruction of the injured tissue and the suppression of pro-inflammatory responses¹⁵. Recent studies have shown that the type 2 immune response has a direct role in wound healing through the production of mediators that directly enhance the tissue repair process and through the control of inflammation. In the experimental model of schistosomiasis, the adaptive type 2 immune response protects the host during infection by inducing granulomas that sequester the tissue-damaging toxins that are released by parasite eggs. If the CD4⁺ T_H2 cell response is impaired, infected animals develop defective granulomas and quickly succumb to the infection because they develop uncontrolled type 1 and type 17 inflammatory responses in the liver and intestines^58,59. The specific blockade of T_H2-induced arginase 1 in mice that are severely infected with Schistosoma mansoni caused increased egg-associated intestinal tissue damage, haemorrhaging, and disruption and leakage of the mucosal barrier, all of which were dependent on signalling through IL-12 receptor subunit β1 (IL-12RB1; which is a receptor subunit shared by the cytokines IL-12p40 and IL-23)¹¹. Thus, in this system, arginase 1 production controls the harmful inflammation that is associated with a type 1 immune response, which consequently contributes to tissue repair and to the maintenance of the mucosal barrier.

In another example, infection with the nematode parasite Nippostrongylus brasiliensis induces acute pulmonary haemorrhaging as parasites traffic through the lungs towards to the intestines^60,61. This damage is caused by both physical trauma and by IL-17-driven neutrophil inflammation⁹. The type 2 immune response, which emerges shortly afterwards, reduces the haemorrhaging and the inflammation through multiple IL-4Rα-dependent mechanisms⁹. In particular, macrophages that have been stimulated through IL-4Rα express IGF1 (REFS 9,62), which has been associated with tissue regeneration, collagen synthesis and fibroblast activation^63–65. IGF1 has been shown to be essential for the enhanced acute woundhealing response during parasite migration through the lungs⁹, which indicates that the helminth-induced type 2 immune response can directly enhance wound healing.

A role for IL-10 and regulatory macrophages

In the N. brasiliensis lung infection model, M2 macrophages engulf and remove damaged red blood cells⁹; this is consistent with other studies that have implicated M2 macrophages in the phagocytosis and clearance of neutrophils^48,66. Although recent in vitro studies have suggested that M2 macrophages might have impaired phagocytosis⁶⁷, in vivo studies in N. brasiliensis⁹ suggest that M2 macrophages retain efferocytosis properties, which is a process that is crucial for resolving inflammation and, thus, it is an important aspect of the wound-healing response.

Some studies have suggested that IL-10-secreting macrophages also have an important protective role in the immune response to helminths by suppressing inflammation^68–70. However, more recent studies of N. brasiliensis infection have shown that IL-10 is primarily produced by CD4⁺ T cells rather than by macrophages⁹, and RNA-sequencing analysis of macrophages during filarial nematode infection showed that there was an absence of IL-10 expression in M2 macrophages⁶². It is probable that in the absence of additional signals, such as TLR ligands or type I interferons⁷¹, helminth-induced M2 macrophages do not secrete substantial quantities of IL-10. These findings are consistent with the proposed wound-healing macrophage phenotype⁷² and indicate that the majority of macrophages that are induced during these nematode infections may be distinct from the regulatory macrophages that secrete IL-10. Nonetheless, helminthinduced M2 macrophages are fundamentally non-inflammatory; they mediate the IL-4Rα-dependent suppression of all proinflammatory mediators⁶², in addition to the production of the pro-repair molecules discussed above.

Taken together, these recent studies show that type 2 immunity mediates tissue repair both by limiting inflammation and by directly activating mediators that facilitate wound healing. Thus, as helminths migrate through vital tissues, sometimes causing extensive tissue damage, the potent type 2 immune response that is rapidly evoked by the infection will enhance tolerance to these large multicellular organisms by inducing wound-healing mechanisms that include the suppression of the inflammatory response.

Helminths regulate inflammatory disease

The potent anti-inflammatory and direct wound-healing pathways that are induced by helminths might greatly affect the overall immune function. The constant presence of helminths probably promotes a homeostatic set point where immunoregulatory mediators, such as IL-10 and arginase 1, are elevated and are therefore available to control harmful inflammatory responses that might otherwise result in disease (FIG. 2). Indeed, the marked reduction in helminth infections in industrialized countries has been causally linked with the rapid rise in the incidence of many inflammatory diseases; this is hypothesized to be due to a loss of immunoregulatory and wound-healing activity. Consistent with this model, the severity of several inflammatory diseases, including inflammatory bowel disease and type 1 diabetes, is considerably reduced in different experimental models of helminth infection^18,73. As a result, much effort has been made to understand helminth-induced immunoregulation, with the aim of therapeutically harnessing these mechanisms of regulation in the future.

The results so far indicate that the immunomodulatory effects of live helminth infections are superior to those of helminth-derived products. The large structure of a living mobile parasite might be necessary to induce multiple regulatory pathways, particularly those that are associated with suppressing injury-induced inflammatory tissue damage. Indeed, multiple independent regulatory pathways are induced during helminth infection and all of these pathways have been implicated in the downregulation of harmful type 1 inflammation⁷⁴. Although these mechanisms often work together to suppress tissuedamaging inflammation, recent studies have suggested that helminth-induced IL-10 and TGFβ work independently of conventional T_H2 cells and forkhead box P3 (FOXP3)⁺ regulatory T cells in controlling inflammation and disease severity in non-obese diabetic (NOD) mice^75,76. This multicomponent response, which has built-in redundancy, might partly be an explanation for the robustness of the helminth-induced immune response in controlling tissue damage in numerous harmful inflammatory diseases.

Persistent type 2 immunity promotes fibrosis

As described above, type 2 cytokines maintain host fitness during helminth infection by downregulating inflammation, by promoting wound healing and by reducing parasite numbers. However, helminth infection is often chronic with a persistent production of type 2 cytokines. If type 2 immunity itself is not appropriately regulated, it can become pathogenic and can contribute to the development of lethal fibrotic pathology, which results from overzealous or persistent wound-healing responses^47,77. In schistosomiasis, hepatic fibrosis leads to the development of portal hypertension, which induces the formation of gastric and oesophageal varices that are prone to rupture⁷⁸. Indeed, the development of fibrosis and portal hypertension are the major causes of morbidity and mortality in chronic schistosomiasis. Whereas IL-4 was identified as the principal inducer of the protective type 2 response during acute S. mansoni infection⁵⁹, IL-13 was identified as the key driver of hepatic fibrosis in mice chronically infected with S. mansoni^77,79. In mice, the progression of hepatic fibrosis in schistosomiasis correlates with the intensity of the IL-13 response⁸⁰, and immunological interventions that selectively impair IL-13 activity have been shown to reduce collagen deposition and to improve survival, as long as IL-4 production is preserved⁷⁷. Thus, type 2 cytokines have both protective and pathogenic activity in chronic schistosome infections, and each infection outcome is directly linked with either beneficial or aberrant wound-healing responses.

Helminth-induced responses control allergy

The need to prevent the consequences of uncontrolled type 2, as well as type 1, immune responses might partly explain the counter-intuitive observation that helminth infection attenuates the harmful type 2 immune responses that are associated with allergy in experimental models⁸¹. This is consistent with epidemiological observations that indicate that the incidence of allergy is reduced in helminth-infected individuals^2,18.

The findings from both experimental and clinical trials are consistent with a model in which the absence of chronic parasite infections can result in dysregulated immune responses to allergens. However, these data are not consistent with alternative models that suggest that allergic responses might be beneficial because they elicit adverse responses to potentially noxious substances, thereby stimulating avoidance behaviour³. This immunosuppressive effect of helminth infection on allergic responses might be a consequence of evasion mechanisms that have evolved in the parasite that enhance parasite survival not only by impairing type 2 immunity but also by protecting the host from damage caused by overzealous repair. IL-10 — which can be produced by effector T cells, FOXP3⁺ regulatory T cells, T regulatory type 1 cells and various innate immune cell populations following helminth infection — might be a particularly potent regulator of harmful type 2 immune responses. In addition, a recent study has suggested that basophil-derived IL-4 converts inflammatory monocytes into M2 macrophages that suppress allergic inflammation⁸². Arginase 1 and RELMα that are produced by M2 macrophages, as well as the IL-13 decoy receptor, are potent negative regulators of type 2 immune responses^12,83–85. These helminth-induced regulators of type 2 immunity might therefore control overzealous wound healing and type 2 allergic responses; this would provide a possible explanation for the increase in severe allergic responses that has been observed in industrialized countries. If this hypothesis is correct, it will be important in future studies to elucidate the mechanisms that preferentially induce regulatory molecules during the type 2 immune response to helminths; these mechanisms could then be used to control harmful type 2 inflammation and to promote tissue repair.