Basophils in Antihelminth Immunity

Abstract

It has been appreciated that basophilia is a common feature of helminth infections for approximately 50 years. The ability of basophils to secrete IL-4 and other type 2 cytokines has supported the prevailing notion that basophils contribute to antihelminth immunity by promoting optimal type 2 T helper (Th2) cell responses. While this appears to be the case in several helminth infections, emerging studies are also revealing that the effector functions of basophils are extremely diverse and parasite-specific. Further, new reports now suggest that basophils can restrict type 2 inflammation in a manner that preserves the integrity of helminth-affected tissue. Finally, exciting data has also demonstrated that basophils can regulate inflammation by participating in neuro-immune interactions. This article will review the current state of basophil biology and describe how recent studies are transforming our understanding of the role basophils play in the context of helminth infections.

1. Introduction

Since Norman Stoll’s landmark paper that estimated the global burden of helminthiasis in 1947, many systemic- and meta-analyses have reported that more than a quarter of the global population is infected with helminth parasites (de Silva, Brooker et al. 2003, Steinmann, Zhou et al. 2007, Jia, Melville et al. 2012, Organization 2016, Jourdan, Lamberton et al. 2018). A rich diversity of parasites contributes to these high rates of infection including the most prevalent helminths: the roundworms (Ascaris lumbricoides and Strongyloides stercoralis), the whipworms (Trichuris trichiura and Trichinella spiralis) and the hookworms (Ancylostoma duodenale and Necator americanus). Long considered as neglected tropical diseases, these chronic helminth infections often result in malnutrition, anemia, growth delay, cognitive deficiencies, and immunopathy (Cox 2002, Dean, Crump et al. 2012, Jia, Melville et al. 2012, Gause, Wynn et al. 2013). Even though antihelminth treatments are employed in heavily affected areas, reinfection rates remain extremely high. For example, studies have shown that up to 60% of individuals will be reinfected within 6-12 months of receiving treatment (Jia, Melville et al. 2012, Zawawi and Else 2020). This has resulted in the need to implement frequent treatment regimens to reduce infection burdens more effectively. Importantly, the need to consistently treat heavily infected areas has resulted in an increased risk of selecting for drug resistant parasites. In fact, similar drug resistance issues have already been identified in livestock populations that are subjected to similar treatment regimens (Charlier, van der Voort et al. 2014, de Oliveira, Leite et al. 2017, Pena-Espinoza, Valente et al. 2018). These concerns highlight the need for more long-lasting treatment strategies such as effective vaccines. Even though recent advances in the genomic and transcriptomic analysis of helminths have helped to inform vaccine strategies (Jex, Smith et al. 2011, Foth, Tsai et al. 2014, Tang, Gao et al. 2014, Liu, Wang et al. 2021), attempts at vaccine development have been largely unfruitful. This is likely a result of the extremely complex nature of helminth life cycles and our poor understanding of host-parasite interactions. It is essential that we begin to fill these critical gaps in knowledge to assist with the development of long-lasting and immune-based therapeutic strategies. Given that studying helminth infections in humans is extremely difficult, animal models employing Trichuris muris, Trichinella spiralis, Nippostrongylus brasiliensis, Heligmosomoides polygyrus, Strongyloides ratti, Strongyloides venezuelensis, Brugia malayi, and Schistosoma mansoni have been widely employed to study the mammalian immune response to helminths (summarized in Table 1, reviewed by (Eberle and Voehringer 2016, Obata-Ninomiya, Domeier et al. 2020)). In nature these diverse parasites are transmitted by the consumption of food or water that is contaminated with eggs, via insect bite, or by the larvae directly penetrating the skin (Organization 2016, Jourdan, Lamberton et al. 2018). The use of animal model systems has allowed us to more effectively study these early aspects of infection and have dramatically informed our understanding of the innate immune responses initiated upon initial exposure to these parasites. Collectively, these studies have revealed that the activation and population expansion of innate immune cells, including eosinophils, mast cells, dendritic cells, group 2 innate lymphoid cells (ILC2s), and basophils, are common features of helminth infections. Further, it is now appreciated that these cell lineages play diverse roles in promoting type 2 immunity and host protective responses to helminths, reviewed by (Gause, Wynn et al. 2013, Inclan-Rico and Siracusa 2018, Kumar, Jeong et al. 2019, Moyat, Coakley et al. 2019).

Table 1.

The Animal Experimental Models of Human Helminth Infections

Animal Model	Infection Stage	Natural Route of Infection	Experimental Route of Inoculation	Affected Compartment1	Human Pathogen Mimic
Trichuris muris	Eggs	Oral ingestion	p.o.	Intestinal tract	Trichuris trichiura
Trichinella spiralis	L1 larvae	Oral ingestion	p.o.	Intestinal tract, skeletal muscle	Trichinella spiralis
Heligmosomoides polygyrus	L3 larvae	Oral ingestion	p.o	Intestinal tract	Ascaris lumbricoides, Ancylostoma duodenale, Necator americanus
Nippostrongylus brasiliensis	L3 larvae	Skin penetration	i.d., s.c.	Skin, lungs, intestinal tract
Strongyloides ratti	L3 larvae	Skin penetration	s.c.	Skin, intestinal tract	Strongyloides stercolaris
Strongyloides venezuelensis
Litomosoides sigmodontis	L3 larvae	Mosquito Blackflies	s.c., mite	Blood, pleural cavity	Brugia malayi, Wuchereria bancrofti, Onchocerca volvulus
Schistosoma mansoni	Cercariae	Skin penetration	Percutaneous exposure s.c.	Skin, liver, lungs	Schistosoma spp.

¹Affected compartment: refers to the experimental models of infection. Intestinal tract changes generally include both host and commensal alterations that occur across both the small and large intestines.

This article will highlight emerging studies that have substantially increased our understanding of helminth-induced basophil responses and emphasize the distinct functions of these cells in the context of antihelminth immunity. We will first provide a brief overview of type 2 immunity and the mechanisms through which it promotes host protective responses to helminths. We will then review the nearly 50 years of reports that have defined peripheral basophilia as a common feature of antihelminth immunity. Next, we will provide a brief overview of basophil development, activation, and effector functions in the context of helminth infections and describe how these pathways promote host protection. Finally, we will discuss emerging studies that are beginning to define the role that basophils play in regulating neuro-immune interactions and highlight how this work has advanced the field of basophil biology and our understanding of how these enigmatic cells contribute to antihelminth immunity.

1.1. Type 2 Immunity

It is well established that type 2 inflammation is required to promote immunity to helminth parasites. During infection helminths migrate throughout different host organs, including the skin, lungs, liver, and intestines. This results in substantial tissue damage and the release of cytokine alarmins by both hematopoietic and non-hematopoietic cells (Figure 1, reviewed by (McSorley and Maizels 2012, Loser, Smith et al. 2019, Maizels 2020, Oyesola, Fruh et al. 2020)). The early production of alarmins (interleukin-25 (IL-25), IL-33 and thymic stromal lymphopoietin (TSLP)) results in the activation and expansion of innate immune cells, including eosinophils, mast cells, dendritic cells, ILC2s and basophils. The activation of these diverse innate cells results in the release of the type 2 cytokines IL-4, IL-5, IL-9 and IL-13 (Figure 1, (Gause, Wynn et al. 2013, Harris and Loke 2017, Webb and Tait Wojno 2017, Maizels 2020)). These early cytokine responses along with presentation of antigen result in the robust activation and differentiation of type 2 T helper (Th2) cells. These responses also result in the activation of B cells that class switch to produce immunoglobulin (Ig)E, a hallmark of type 2 inflammation (Wu and Zarrin 2014, Eberle and Voehringer 2016, Oyesola, Fruh et al. 2020). Altogether, these immunological changes serve to initiate host protection by stimulating smooth muscle contraction, mucus production from goblet cells, and epithelial cell turnover (Zhao, Urban et al. 2008, Eberle and Voehringer 2016, Nakayama, Hirahara et al. 2017, Zhang and Wu 2020). Collectively, type 2 responses simultaneously promote both resistance and tolerance mechanisms with alternatively activated or M2 macrophages playing a central role (Coakley and Harris 2020). For instance, M2 macrophages have been shown to be crucial for rapid worm killing and clearance (Bonne-Annee, Kerepesi et al. 2014, Chen, Wu et al. 2014), are key regulators of wound healing processes (Chen, Liu et al. 2012, Ferrante and Leibovich 2012) and play critical roles in the formation of granulomas that serve to trap the parasites and limit their migration (Nascimento, Huang et al. 2014, Marino, Cilfone et al. 2015, De Santis, Locati et al. 2018, Shamaei, Mortaz et al. 2018).

Figure 1. Anti-helminth Immunity at Mucosal Surface.

Helminths cause substantial tissue damage as they migrate through different organs, such as the lungs, gut, and skin. Both hematopoietic and non-hematopoietic cells release cytokine alarmins (IL-25, IL-33, and TSLP), such as from macrophages (Macs) and epithelial cells, leading to the activation and expansion of innate immune cells, including eosinophils (Eos), mast cells (MCs), dendritic cells (DCs), ILC2s and basophils (Baso), etc. Furthermore, hematopoietic stem/progenitor (HSPCs) are activated and contribute to inflammation by developing into mast cells and basophils. Innervating neurons also respond to helminth-derived products and promote inflammation via their release of neuromedin U (NMU). Collectively, these mechanisms result in the robust production of the type 2 cytokines that promote the polarization of type 2 T helper (Th2) and the induction of M2 macrophages that serve to expel the worm and initiate the healing of wounded tissue. Reviewed in (McSorley and Maizels 2012, Gause, Wynn et al. 2013, Harris and Loke 2017, Webb and Tait Wojno 2017, Loser, Smith et al. 2019, Maizels 2020, Oyesola, Fruh et al. 2020).

1.2. Basophilia & Helminth Infections

Peripheral basophilia has been recognized as a conserved feature of helminth infections in animal models for ~50 years. For example, in the 1960s and 1970s, dramatic increases in basophil numbers were reported to occur in guinea pigs injected with Ascaris suis body fluid (Chan 1965) and Nippostrongylus brasiliensis infected Mongolian gerbils (Smith, Termer et al. 1976, Okada, Nawa et al. 1997). These early findings were further supported by subsequent studies describing basophil responses upon rodent infections by Trichuris muris, Trichinella spiralis, Strongyloides venezuelensis, and filarial nematode Litomosoides sigmodontis (Giacomin, Siracusa et al. 2012, Mukai, BenBarak et al. 2012, Mukai, Karasuyama et al. 2017, Hartmann, Linnemann et al. 2018).

Consistent with observations in animal models, it has also been shown that human basophils from Toxocara-, Ascaris-, Strongyloides-, and Schistosoma-infected patients were able to release histamine in response to parasite antigen (Genta, Ottesen et al. 1983, Hofstetter, Fasano et al. 1983, Nielsen, Lind et al. 1994). Despite these facts, the contributions of basophils to antihelminth immunity and host protection remain poorly defined. The sections below will review what is known about basophil development, their mediators of action, their recognized effector functions and highlight gain- and loss-of-function studies demonstrating a role for basophils in promoting protective immunity to helminths.

1.3. Basophil Development

Basophil progenitors

Basophils are the least abundant granulocyte and constitute less than 1% of leukocytes in the peripheral blood (Siracusa, Comeau et al. 2011). Using flow cytometric techniques, murine basophils can be identified as FcεRlα⁺, CD49b⁺, CD69⁺, Thy-1.2⁺, CD123⁺, CD200R⁺, CD117⁻, CD3⁻, CD19⁻, CD14⁻, CD122⁻, CD11c⁻, Gr-1⁻, NK1.1⁻, B220⁻, αβTCR⁻, γδTCR⁻ cells (Siracusa, Comeau et al. 2011, Eberle and Voehringer 2016). Although mast cells and basophils share common characteristics, basophils can be easily separated from mast cells by their positive expression of CD49b and CD90 and their lack of the mast cell marker CD117 (c-Kit) (Ohnmacht, Schwartz et al. 2010). Moreover, basophils can be distinguished by staining for mouse mast cell protease 8 (Mcpt8) or Mcpt11 (Ugajin, Kojima et al. 2009). Human basophils can also be identified by flow cytometric analysis as CD123⁺ FcεRIa⁺ cells that are c-Kit⁻ and Lin⁻ (lineage markers CD3, CD4, CD19, CD14, CD34, and CD56) (Wang and Kim 2020). Additionally, the activation state of human basophils can also be evaluated by their expression of various surface markers such as CD203c (Hauswirth, Natter et al. 2002, Boumiza, Monneret et al. 2003). Intracellular and histological staining with the basophil-specific antibody 2D7 can also be used to help distinguish human basophils from other cell lineages including mast cells (Kepley, Pfeiffer et al. 1998).

In addition to their divergent expression of surface markers and proteases, basophils and mast cells also develop via distinct pathways and progenitor cells. It was originally reported that basophils primarily develop in the bone marrow from a common granulocyte-monocyte precursor (GMP). These CD34⁺ CD117⁺ GMPs were shown to develop into multiple cell lineages, including macrophages, eosinophils, mast cells and basophils (Arinobu, Iwasaki et al. 2005) depending on their expression patterns of C/EBPα and GATA-2 (Iwasaki, Mizuno et al. 2006). However, recent single-cell transcriptomic analyses in both mouse and humans have begun to reveal that progenitor cells such as GMPs are more heterogeneous than originally appreciated (Olsson, Venkatasubramanian et al. 2016, Dahlin, Hamey et al. 2018, Drissen, Thongjuea et al. 2019, Pellin, Loperfido et al. 2019). Further, emerging studies have begun to reveal that mouse basophil and mast cell fates are more closely related to erythroid and megakaryocytes lineages than was previously thought (Dahlin, Hamey et al. 2018, Tusi, Wolock et al. 2018, Drissen, Thongjuea et al. 2019, Pellin, Loperfido et al. 2019, Grootens, Ungerstedt et al. 2020, Inclan-Rico, Hernandez et al. 2020). Importantly, several reports have also described a unique erythrocyte-mast cell progenitor that appears to have extremely little basophil potential (Gentek, Ghigo et al. 2018, Zheng, Papalexi et al. 2018, Inclan-Rico, Hernandez et al. 2020). These studies strongly suggest that basophil and mast cell development may not be as tightly linked as previously thought (Gentek, Ghigo et al. 2018, Zheng, Papalexi et al. 2018, Inclan-Rico, Hernandez et al. 2020). In addition to bone marrow-resident progenitors, studies have also highlighted the contribution of extramedullary hematopoiesis (EMH) to basophil development (Saenz, Siracusa et al. 2010, Huang and Qi 2013, Siracusa, Saenz et al. 2013). For example, it is reported that Lin⁻ c-Kit⁺ FcγRII/III^hi β7 integrin^hi progenitors with basophil potential reside in the spleen (Arinobu, Iwasaki et al. 2005, Huang and Qi 2013). Moreover, it was demonstrated that Trichinella spiralis-induced TSLP promotes the accumulation of a GMP-like Lin⁻ CD34⁺ c-Kit⁺ FcεRIα⁻ cell population in the spleen with substantial potential to develop into basophils (Siracusa, Saenz et al. 2013). Collectively, these studies indicate that basophils can develop from both bone marrow-resident and peripheral progenitor cell populations depending on the nature of the stimuli. However, further studies are required to better understand the contributions of these various progenitor populations to helminth-induced basophilia.

1.4. Regulators of Basophil Development and Activation

Development

It is well established that T cell-derived IL-3 operates as a potent growth and survival factor of basophils (Lantz, Boesiger et al. 1998, Shen, Kim et al. 2008, Voehringer 2012, Oetjen, Noti et al. 2016, Benard, Jacobsen et al. 2021). For example, Lantz et al. demonstrated that Nippostrongylus- and Strongyloides-induced basophil responses were critically dependent on IL-3-IL3R signaling (Lantz, Boesiger et al. 1998). This work has been supported by subsequent studies showing that IL-3 induces basophil expansion in vivo by directing GMPs to differentiate into basophil lineage-restricted progenitors in the bone marrow (Ohmori, Luo et al. 2009). Moreover, Kim et al. also showed that basophil recruitment to the draining lymph nodes post Nippostrongylus brasiliensis infection is dependent on IL-3 (Kim, Prout et al. 2010). More recently, it is recognized that basophil development can be affected by commensal signals via IL-3-IL-3R signaling (Herbst, Esser et al. 2012, Hill, Siracusa et al. 2012).

In addition to IL-3, granulocyte–macrophage colony-stimulating factor (GM-CSF), Toll-like receptors, Notch signaling and TSLP have also been shown to regulate basophil development (Denburg, Woolley et al. 1994, Reece, Baatjes et al. 2013, Siracusa, Saenz et al. 2013, Webb, Oyesola et al. 2019). Critically, TSLP can cooperate with IL-3 to promote optimal basophil responses and is also sufficient to promote basophil development and peripheral basophilia in an IL3R-deficient environment (Siracusa 2011). Importantly, basophils developed in a TSLP-rich environment display distinct transcriptional profiles and exhibited different states of activation compared to basophils elicited with IL-3 alone (Siracusa, Saenz et al. 2011). Collectively, these studies helped advance the concept of heterogeneity within the basophil compartment and suggest that basophils may possess differing effector functions depending on the signals that promote their development. The section below will highlight the known mediators of basophil activation and discuss the different effector functions they initiate.

Activation

It is well recognized that Toll-like receptor ligands, complement factors, allergen-bound IgE, proteases, various cytokines (IL-3, GM-CSF, TSLP, IL-18 and IL-33) and helminth-derived antigens can promote basophil activation (Siracusa, Saenz et al. 2011, Willart, Deswarte et al. 2012, Reece, Gauvreau et al. 2014, Galeotti, Karnam et al. 2020). Specifically, it has been demonstrated that human and mouse basophils can be activated via either IgE-dependent or IgE-independent pathways, resulting in the release of basophil-derived effector molecules, including the cytokines IL-4, IL-13, IL-6 and TNFα; chemokines, histamine, and lipid mediators such as leukotriene C4 (LTC4) and prostaglandins (Schroeder 2009). Importantly, whether basophils are activated in IgE-dependent or IgE-independent pathways result in varying kinetics of activation and distinct transcriptional profiles. For example, it was recently reported that basophils differentially engage stromal interaction molecules (STIMs), localized primarily to the endoplasmic reticulum (ER), in response to IgE-dependent or IgE-independent stimulation. Once stimulated, stored Ca²⁺ is released from the ER to the cytoplasm. STIM1 or STIM2 sense the Ca²⁺ depletion in the ER and thereby promote an influx of extracellular Ca²⁺ via Orai (Calcium release-activated calcium channel protein 1) calcium channels on the cell surface (Yoshikawa, Oh-Hora et al. 2019). Yoshikawa et al. demonstrated that the Ca²⁺ influx in response to IgE-dependent activation (STIM1-utilized) can be observed within 1 hour, while IL-3-mediated Ca²⁺ influx (STIM2-utilized) occurs after 6 hours. These differences are consistent with the differential kinetics of basophil-derived IL-4 production in response to IgE- or IL-3-stimulation (Yoshikawa, Oh-Hora et al. 2019). Critically, in the studies highlighted above it was shown that basophils developing in a TSLP-rich environment are more responsive to IgE-independent activation than basophils elicited with IL-3 alone (Siracusa, Saenz et al. 2011). In contrast, basophils elicited with IL-3 are more responsive to IgE-dependent signals. However, it remains unclear whether these differences are a result of unique calcium sensing mechanisms. Better elucidating the factors that promote basophil heterogeneity will increase our understanding of the functional characteristics of basophils in the context of various helminth infections.

2. Basophil Effector Functions on Anti-Helminth Immunity

2.1. Animal models of basophil depletion

Several studies have demonstrated that MAR-1 (anti-FcεRI) or Ba103 (anti-CD200R3) monoclonal antibodies (mAbs) can be used to deplete basophils to study their functional importance. Although these antibodies effectively deplete basophils, it is important to note that these approaches, like many cell depletion strategies, can result in off-target effects such as the activation or partial depletion of mast cells (Denzel, Maus et al. 2008). To address this, several basophil-specific mouse models have also been developed in the past decade. For example, an inducible mouse model of basophil depletion was generated by inserting a diphtheria toxin receptor (DTR)-encoding cassette (IRES-DTR-EGFP) into the 3’ untranslated region of the Mcpt8 gene. Injecting these Mcpt8-DTR mice with DT leads to the transient and selective depletion of >90% of basophils (Wada, Ishiwata et al. 2010). Another DTR-based transgenic model, termed toxin receptor-mediated conditional cell knockout (TRECK), takes advantage of basophil-specific IL-4 enhancer elements to selectively deplete basophils following DT administration (Sawaguchi, Tanaka et al. 2012). Moreover, different approaches have also been used to introduce Cre recombinase under the control of the regulatory elements of Mcpt8 to target basophils. In one of these mouse models the Cre levels were found to be toxic to the basophils and resulted in the constitutive loss of more than 90% of basophils (Ohnmacht, Schwartz et al. 2010). In the case of Basoph8 mice, an IRES-YFP-Cre cassette was used to replace the gene coding for Mcpt8. When Basoph8 mice are crossed with Rosa-DTa mice, up to 96% of basophils are depleted (Sullivan, Liang et al. 2011). In summary, there are a number of effective antibody-mediated and genetic approaches to constitutively or temporally deplete basophils in vivo. These experimental approaches have proven invaluable in investigating the importance of basophils in promoting immunity to a wide range of helminth parasites including many of those highlighted in Table 1.

2.2. The effects of basophil depletion on helminth-induced inflammation

As mentioned previously, activated basophils are robust producers of IL-4 and IL-13 which play important roles in regulating Th2 cell priming. Given that basophilia is a common feature of helminth infections, it was hypothesized that their activation promoted the development of optimal type 2 inflammation required for antihelminth immunity. This hypothesis was confirmed by-loss-of-function studies done in the context of Trichinella spiralis (Ts), and Heligmosomoides polygyrus (Hp) infections (Giacomin, Siracusa et al. 2012, Schwartz, Turqueti-Neves et al. 2014)). Specifically, Giacomin et al. found that TSLP-elicited basophils promote optimal Th2 cytokine-associated inflammation during Ts infection (Figure 2A). Further, Schwartz et al. showed that during an Hp infection basophils play an important role in promoting M2 macrophages and worm clearance in an FcR-, IL-4-dependent manner (Schwartz, Turqueti-Neves et al. 2014). While similar loss-of-function studies have suggested that basophils are not required to promote type 2 responses following a primary Nippostrongylus brasiliensis (Nb) infection, studies have demonstrated that they promote host protection during a secondary challenge. Specifically, Ohnmacht et al. showed that during a secondary Nb infection, antibody-mediated depletion of basophils by targeting Thy1.2 or CD200R3, led to reduced type 2 cytokine responses and increased worm burden (Figure 2B, (Ohnmacht, Schwartz et al. 2010)). Subsequent studies by Obata-Ninomiya et al. showed that basophil-derived IL-4 promotes M2 macrophage polarization, and the production of the enzyme arginase-1, resulting in the trapping of Nb larvae in the skin during secondary challenge (Figure 2B, (Obata-Ninomiya, Ishiwata et al. 2013)). In contrast to Nb, Mukai et al. observed that targeting basophils by employing Mcpt8-DTR mice following Strongyloides venezuelensis (Sv) challenge resulted in significantly lower Sv egg production during primary infection but had no detectable effect during a secondary Sv challenge (Mukai, Karasuyama et al. 2017). Similar studies also suggested that basophils are dispensable during primary and secondary infections with Strongyloides ratti (Reitz, Brunn et al. 2017, Reitz, Brunn et al. 2018), further highlighting the parasite-specific role of helminth-induced basophils.

Figure 2. The role of basophils in primary and secondary helminth infections.

A. Basophils are recruited to the mesenteric lymph nodes (mLN), secrete IL-4 and promote optimal Th2 cytokine-mediated inflammation during Trichinella Spiralis (Ts) primary infection. B. In the context of secondary Nippostrongylus brasiliensis (Nb) infection, IL-4 made by basophils promotes the polarization of Arginase 1-expresseing (Arg1) M2 macrophages and the trapping of Nb larvae in the skin (Giacomin, Siracusa et al. 2012, Obata-Ninomiya, Ishiwata et al. 2013, Schwartz, Turqueti-Neves et al. 2014).

In the context of Trichuris muris (Tm) infection, peripheral basophilia was found to be dependent on infection-induced TSLP. Further, it was demonstrated that the adoptive transfer of TSLP-elicited basophils was sufficient to promote worm clearance in highly susceptible TSLPR-deficient mice (Siracusa, Saenz et al. 2011). More recently, Webb et al. demonstrated that basophil-intrinsic Notch signaling promotes optimal worm clearance, type 2 inflammation, and basophil localization to the intestine during Tm infection (Webb, Oyesola et al. 2019). Collectively, these studies suggest that the functions and contributions of basophils are highly dependent on the infections being studied. Further, the relatively minor differences seen in several of the loss-of-function experiments is somewhat surprising and provokes the hypothesis that basophils may also promote host protection independently of their ability to promote type 2 inflammation.

2.3. Basophils & Neuro-Immune Crosstalk in Anti-Helminth Responses

As new technologies emerge, such as novel transgenic models, or precise activation techniques like chemogenetics and optogenetics, more research is shedding light on the importance of neuro-immune crosstalk in promoting immunity and inflammation (Chu, Artis et al. 2020). Considering that helminths have coevolved with mammals for millennia, it is not surprising that the host-parasite relationship extends to include the central nervous system (CNS). In fact, the ability of parasites to alter host behavior is a well described phenomenon (Poulin and Maure 2015, Iritani and Sato 2018). However, the ability of the CNS to communicate with the immune system in a manner that regulates antihelminth immunity is just beginning to be appreciated. The paragraphs below will highlight emerging research demonstrating a role for basophils in regulating neuro-immune interactions.

The lung is now recognized as a critical checkpoint for the development of protective immunity to several helminth parasites including hookworms (Harvie, Camberis et al. 2010, Allen and Sutherland 2014, Craig and Scott 2014). Despite its important role in antihelminth immunity, it is critical that inflammation in the lung is tightly regulated to assure proper tissue function (Robb, Regan et al. 2016, Branchett and Lloyd 2019). Similar to human hookworms, Nb larvae burrow through the lung of their host and promote substantial tissue damage. The host combats this challenge by initiating wound healing responses that are promoted by the specialized actions of N2 neutrophils, ILC2s, subset of T and B cells and M2 macrophages (Chen, Liu et al. 2012, Gause, Wynn et al. 2013, Castellanos and Longman 2019). Importantly, a recent study demonstrated that basophils migrate into the lung tissue during this wound healing phase of Nb infection. Further, it was demonstrated that lung ILC2 responses are exaggerated in the absence of basophils, resulting in increased local inflammation, and declined pulmonary function (Inclan-Rico, Ponessa et al. 2020). Additional analysis of ILC2s from basophil-depleted mice revealed that they expressed significantly lower levels of the receptor for the neuropeptide neuromedin B (NMB). Moreover, NMB stimulation was sufficient to inhibit ILC2 responses from control but not basophil-depleted mice (Inclan-Rico, Ponessa et al. 2020). Finally, Inclan-Rico et al. showed that prostaglandin E2 (PGE2) was sufficient to promote NMBR expression by ILC2s and thereby primed them for neuropeptide-mediated inhibition (Figure 3A). This work suggests that basophils can operate as negative regulators of type 2 immune responses and promote a rethinking about the roles that basophils play in promoting host protection. Given that basophils can also produce PGD2 (Ugajin, Satoh et al. 2011), it is possible that basophils participate in the recently defined ability of PGD2 and its receptor CRTH2 to restrict type 2 cytokine-driven epithelial responses following an Nb challenge (Oyesola, Shanahan et al. 2021). However, additional studies are required to thoroughly evaluate the ability of basophil-derived prostaglandins to regulate helminth-induced inflammation. Further, more work is required to better determine how these pathways fit with a growing body of literature highlighting the importance of basophils in regulating neuro-immune interactions, such as recent work defining these pathways in the context of pruritus (Wang and Kim 2020, Wang, Trier et al. 2021).

Figure 3. Basophils regulate Neuro-Immune Crosstalk.

A. After the passage Nippostrongylus brasiliensis (Nb) larvae through the lung, basophils are recruit to the damaged tissue and promote NMBR expression by ILC2s and thereby prime them for neuropeptide-mediated inhibition (Inclan-Rico, Ponessa et al. 2020). B. In acute itch exacerbation, basophil-derived leukotriene C4 (LTC4) can interact with sensory nerves. LTC4 binds to its receptor CysLTR2 on non-peptidergic 3 –itch-sensory neurons and triggers itch sensation (Wang, Trier et al. 2021). However, more work is required to determine the importance of this pathway in antihelminth immunity.

Pruritus, or itch, is considered an evolutionarily protective behavioral extension of type 2 immunity that serves to expel parasites from the host (Garcovich, Maurelli et al. 2021). The most studied itch mechanism is the canonical IgE-mast cell-histamine axis, where allergens are recognized by IgE bound to FcεRI and subsequently leads to the release of effector molecules including serotonin and histamine (Yang and Kim 2019). Histamine can directly stimulate sensory neurons and thereby promotes the itch sensation and neuroinflammation (Wang and Kim 2020). Importantly, basophils are potent producers of histamine and other serine proteases such as Mcpt8 and Mcpt11 which play important roles in promoting skin inflammation (Iki, Tanaka et al. 2016, Tsutsui, Yamanishi et al. 2017). Further, studies by Kim et al. demonstrated that basophil-derived IL-4 amplifies ILC2 responses in the context of atopic dermatitis (Kim, Wang et al. 2014). Collectively, these studies suggest that basophils play important roles in regulating skin inflammation. This concept has been further supported by recent work by Wang et al. revealing a previously unrecognized basophil-intrinsic leukotriene C4 (LTC4) axis required for acute itch flare in skin inflammation (Wang, Trier et al. 2021). Wang and colleagues found that the LTC4 receptor cysteinyl LT receptor 2 is exclusively expressed on non-peptidergic 3 sensory neurons and serves as a downstream target in a novel basophil-neuronal circuit (Figure 3B). Collectively, these advances in our understanding of itch and skin inflammation further shed light on the contribution of basophils in regulating neuro-immune crosstalk. While the skin has been shown as an important site for host protection to helminths (Obata-Ninomiya, Ishiwata et al. 2013, Garcovich, Maurelli et al. 2021), more work is needed to evaluate how these pathways operate in the context of parasitic immunity.

Concluding Remarks

While it was initially hypothesized that basophils regulate antihelminth immunity by producing type 2 cytokines and promoting optimal Th2 cell responses, our appreciation for these dynamic cells continues to grow. It is now recognized that basophils can come from several progenitor cell compartments that reside in the bone marrow or periphery. Further, it’s been shown that the signals regulating basophil develop can influence their transcriptional profiles and effector functions. Collectively, these studies suggest the nature of basophil development may fine tune the cells to more efficiently respond to either IgE-dependent or IgE-independent signals. This work could help to explain why basophils participate in primary immunity that occurs before antigen-specific antibodies are generated in some contexts but in secondary responses when the cells are loaded with parasite-specific-IgE in other cases. Additionally, recent studies are now beginning to reveal that basophils can function as negative regulators of helminth-induced inflammation and can also influence neuro-immune interactions. Further investigating these exciting advances in basophil biology is likely to lead to an even greater appreciation for how these specialized cells regulate inflammation, influence neuronal circuits and maintain tissue functions. Gaining a better understanding of these complex pathways will also inform strategies to therapeutically target basophils in a manner that promotes optimal outcomes in the context of helminth infections.