Listening In: Plasmacytoid DC, Monocyte-Derived DC, and Neutrophil Crosstalk in Antifungal Defense

Abstract

Plasmacytoid DCs (pDCs) are typically thought to be key in antiviral defense. In this issue of Cell Host & Microbe, Guo, Kasahara et al. (2020) reveal a critical role for pDCs in antifungal immunity. Aspergillus-infected monocyte-derived DCs and neutrophils recruit pDCs, which promote neutrophil fungicidal activity.

Aspergillus fumigatus is an opportunistic fungal pathogen found ubiquitously throughout the environment. Humans constantly inhale fungal spores, termed conidia, which normally do not pose a problem to healthy individuals. However, A. fumigatus can cause invasive aspergillosis (IA) in immunocompromised hosts and is the most common and lethal cause of mold pneumonia. Host defense against A. fumigatus relies on recruitment of myeloid cells, including neutrophils (NΦs), monocytes, and monocyte-derived dendritic cells (Mo-DCs), into the lung. NADPH oxidase activity in phagocytes is critical for fungal clearance in both mice and humans, as patients with chronic granulomatous disease are highly susceptible to IA. Upon exposure to reactive oxygen species (ROS) within neutrophils, conidia undergo regulated cell death, which prevents fungal dissemination and allows for sterilizing immunity (Shlezinger et al., 2017).

During A. fumigatus infection, CCR2+ inflammatory monocytes (iMo) are recruited into the lung, where they serve as potent proinflammatory cells and differentiate into Mo-DCs. CCR2+ iMo and Mo-DCs are required for NΦ fungicidal activity (Espinosa et al., 2017; Espinosa et al., 2014), but how they regulate NΦ function has been unknown. One possibility is that CCR2+ iMo and Mo-DCs produce inflammatory mediators that upregulate antifungal effectors within NΦs. Alternatively, CCR2+ iMo and Mo-DCs could act through a third cell type that subsequently regulates NΦ function.

In this issue of Cell Host & Microbe, Guo, Kasahara, and colleagues first examined the contributions of CCR2+ iMo and Mo-DCs to the production of inflammatory mediators during A. fumigatus infection to distinguish between the above two possibilities (Guo et al., 2020). They observed that these cell types are required for production of the chemokines CXCL9 and CXCL10. Using CXCL9- and CXCL10-reporter mice, they found that Mo-DCs and NΦs are the predominant sources of CXCL9 and CXCL10 (Figure 1). Interestingly, fungus-engaged Mo-DCs and NΦs produced much higher levels of CXCL9 and CXCL10 than uninfected bystander cells. They implicated two distinct pathways—Dectin-1/Card9-signaling and type I/III interferon (IFN) receptor signaling—in regulating CXCL9 and CXCL10 production, respectively (Figure 1). The fungal pattern recognition receptor Dectin-1 binds to β-glucan exposed during conidia swelling, the first step of germination (Hohl et al., 2005), thus indicating that Dectin-1 recognition of β-glucan in Mo-DCs and NΦs directly regulates CXCL9 production. Deficiency in type I IFN receptor (IFNAR) or type III IFN receptor (IFNLR1) signaling led to a reduction in CXCL10, a known IFN-stimulated gene. Whether cell-intrinsic Dectin-1 or IFNAR/IFNLR1 signaling in Mo-DCs and NΦs regulates chemokine production is unclear. CCR2+ iMo is a critical source of type I IFNs, and type I IFNs are required for type III IFN production (Espinosa et al., 2017). The innate immune sensing pathways that direct production of type I and III IFNs and the cellular source of type III IFN during infection are not yet known.

Figure 1. Mo-DCs and NΦs Mediate CXCR3-Dependent Recruitment of pDCs, which Enhance NΦ Fungicidal Activity.

During A. fumigatus infection, pDCs egress from the bone marrow in a CCR2-dependent manner and are recruited into the lung in a CXCR3-dependent manner. The CXCR3 ligands CXCL9 and CXCL10 are predominantly produced by Mo-DCs and NΦs upon Dectin-1 recognition of conidia β-glucan and type I/III IFN (IFN-α/β and IFN-λ) signaling; whether Mo-DC- and NΦ-intrinsic Dectin-1 and IFNR signaling regulates chemokine production remains unclear. Once within the lung, pDCs enhance NΦ-specific ROS production, resulting in conidial killing; the mechanism underlying how pDCs regulate ROS production in NΦs remains to be determined. Abbreviations are as follows: pDCs, plasmacytoid dendritic cells; Mo-DCs, monocyte-derived dendritic cells; NΦs, neutrophils; IFNs, interferons; IFNRs, interferon receptors; ROS, reactive oxygen species.

CXCL9 and CXCL10 are ligands for CXCR3, and the authors found that Cxcr3^−/− mice are more susceptible to A. fumigatus. They next sought to identify the cellular target of CXCR3 required for antifungal defense. A variety of immune cell types, including T cells, NK cells, and pDCs, express CXCR3 during A. fumigatus infection. Lymphocytes are dispensable for A. fumigatus clearance (Espinosa et al., 2014), and pDCs have been previously implicated in antifungal defense (Ramirez-Ortiz et al., 2011). Thus, the authors hypothesized that CXCR3-dependent pDC trafficking to the lung promotes antifungal immunity. They observed that pDCs are recruited into the lung during A. fumigatus infection and that pDCs express both CXCR3 and CCR2. They found that CCR2 regulates pDC egress from the bone marrow (BM), while CXCR3 primarily mediates pDC trafficking into the lung at steady state and during infection (Figure 1).

Although pDCs constitute a small population of leukocytes, they have a well-appreciated role as potent producers of type I IFNs during viral infection. However, the role of pDCs in immune defense against bacterial or fungal infections is relatively poorly understood. Previous studies reporting a role for pDCs in host defense against A. fumigatus used a monoclonal antibody (anti-PDCA-1) that targets pDCs (Ramirez-Ortiz et al., 2011) but that likely also targets other immune cell types. Here, the authors took advantage of a transgenic mouse model in which the human BDCA2 (also known as CLEC4C, a pDC-specific type II C-type lectin) promoter drives expression of the diphtheria toxin receptor (DTR) to enable acute and specific depletion of pDCs (Swiecki et al., 2010). pDC-depleted mice exhibited higher mortality associated with increased fungal burdens and enhanced fungal germination in the lung than non-depleted controls, thus demonstrating that pDCs are indeed essential for control of A. fumigatus infection.

NΦs are essential for antifungal defense due to NADPH oxidase-dependent production of reactive oxygen species (ROS) (Espinosa et al., 2014), which induces regulated cell death of conidia engulfed by NΦs (Shlezinger et al., 2017). Intriguingly, the authors demonstrate that pDCs are required for NΦ ROS production and fungicidal activity (Figure 1). How pDCs specifically enhance NΦ fungicidal activity remains to be determined. Given the relatively small number of pDCs present within the lung, one possible scenario is that, upon sensing fungal or host-derived signals, pDCs produce inflammatory mediators that upregulate antifungal effector mechanisms within NΦs. As GM-CSF, type I IFN, and type III IFN are critical regulators of NΦ NADPH oxidase activity (Espinosa et al., 2017; Kasahara et al., 2016), it would be interesting to see if pDC-mediated production of these cytokines or additional mediators regulate NΦ antifungal activity.

Overall, this study elegantly illustrates a three-cell circuit involving pDCs, Mo-DCs, and NΦs that participates in a feed-forward loop to mediate anti-fungal immunity. This study also provides insight into pDC trafficking, as the authors demonstrate CCR2-mediated egress of pDCs from the BM and subsequent CXCR3-mediated recruitment into the lung tissue during infection. Given that pDCs play a role in host defense against a variety of viral, bacterial, and fungal pathogens, do similar mechanisms operate for pDC trafficking and enhancement of NΦ effector function during other infections? Are these findings translatable to human immunity? Intriguingly, a Cxcl10 polymorphism has been implicated in invasive aspergillus susceptibility in hematopoietic cell transplant recipients (Mezger et al., 2008), and donor pDC reconstitution in bone marrow transplant patients is associated with favorable clinical outcomes (Waller et al., 2014). Thus, understanding precisely how pDCs regulate antifungal activity and whether pDCs can be therapeutically harnessed to promote antifungal responses in immunocompromised patients warrant future study.