Many helminth parasites migrate through multiple host organs during infection but how immunity is regulated across these tissues is still poorly understood. To investigate the cellular and molecular aspects of inter-tissue communication during infection we established a percutaneous infection model with the tissue-migrating nematode Nippostrongylus brasiliensis. High-dimension profiling of the initial cutaneous immune response revealed that dermal gamma delta T cells become activated, engage cell motility-associated transcriptional pathways and leave the skin after parasite invasion. Chemical and genetic inhibition of leukocyte migration prevents the accumulation of IL-17-producing gamma delta T cells in the lungs. Notably, bypassing the skin phase of infection, and therefore preventing dermal gamma delta T cell migration, dampens the increase in early IL-17 production in the lungs, and, instead, leads to enhanced IFN-gamma responses together with increased lung damage. Collectively, our data highlights a critical skin-lung axis regulating host-parasite interactions and safeguarding lung health. ### Competing Interest Statement The authors have declared no competing interest.

Chitinase-like proteins (CLPs) are established biomarkers of inflammation and airway remodeling in asthma, yet their direct contribution toward disease pathogenesis is unknown. In a mouse model of type 2/type 17 airway inflammation induced by house dust mite, ragweed, and Aspergillus fumigatus (DRA) allergens, we demonstrate that murine CLPs Ym1 (Chil3) and Ym2 (Chil4) exert distinct and potent effects on airway extracellular matrix (ECM) composition during chronic lung pathology. Using both Ym1-knockout (Chil3-/-) and Ym2-knockdown (Chil4KD/KD) mice, we found that these CLPs have fundamental roles in airway remodeling that are independent of interleukin-13 (IL-13) and IL-17A signaling pathways. Antibody-mediated inhibition of CLPs after pathology has developed reverses airway remodeling independently of chronic inflammation. However, this inhibition is not sufficient to reduce airway hyperresponsiveness (AHR) in allergic animals. Instead, the absence of CLPs increases epithelial damage and leads to a loss of bronchial epithelial integrity. This work disentangles chronic IL-13 and IL-17A signaling from the development of allergic airway pathology and reveals CLPs as orchestrators of airway remodeling that can produce both protective and adverse outcomes.

Fungal spores are abundant in the environment and a major cause of asthma. Originally characterised as a type 2 inflammatory disease, allergic airway inflammation that underpins asthma can also involve type 17 inflammation, which can exacerbate disease causing failure of treatments tailored to inhibit type 2 factors. However, the mechanisms that determine the host response to fungi, which can trigger both type 2 and type 17 inflammation in allergic airway disease, remain unclear. Here we find that CD11c+ DCs and CD4+ T cells are essential for development of both type 2 and type 17 airway inflammation in mice repeatedly exposed to inhaled spores. Single cell RNA-sequencing with further multi-parameter cytometry shows that allergic inflammation dramatically alters the proportion of numerous DC clusters in the lung, but that only two of these (Mgl2+ cDC2s and CCR7+ DCs) migrate to the dLNs. Targeted removal of several DC subsets shows that Mgl2+ cDC2 depletion reduces type 2, but not type 17, fungal allergic airway inflammation. These data highlight distinct DC subsets as potential therapeutic targets for the treatment of pulmonary fungal disease.

Stimulation of macrophages via CD40 promotes their classical activation. Therefore, CD154 (CD40 ligand) can be expected to oppose macrophage polarization and proliferation induced by IL-4. However, there are limited experimental data to support this, which is additionally complicated by the possibility of differential effects of CD40 agonists in different formats/contexts. Whereas canonically CD4+ T cells upregulate CD154 strongly following exposure to cognate antigen, naïve CD4+ cells constitutively express significant levels of CD154, which could be a tonic signal. Soluble CD154 and agonistic CD40 antibodies also trigger CD40 signaling. We explored these questions in a reductionist model of IL-4 delivery to mouse peritoneal cavity cells in vitro and in vivo. Soluble CD40 agonists inhibited M(IL-4) polarization, with a stronger effect on RELM-α than on Ym1 (Chil3), as well as inhibiting IL-4-induced proliferation. CD154 provided by CD4+ cells in the context of an antigen-specific interaction blunted macrophage RELM-α expression but did not affect Ym1. Macrophages negatively regulated, via CD40, constitutive cell-surface CD154 on naïve CD4+ cells, both in vitro and in vivo. The large peritoneal macrophages of CD40 KO mice showed a moderately enhanced RELM-α response to IL-4, but this was not a cell-autonomous effect. No differences between WT and CD40 KO mice were detected in IL-4-induced macrophage proliferation. We conclude that strong CD40 stimulation, including stimulation by CD154 expressed by antigen-specific CD4+ cells, blunts selected macrophage responses to IL-4, and that constitutive CD4+-cell CD154, in spite of interacting with CD40 on macrophages, does not directly influence macrophage responses to IL-4.

Filarial nematodes infect over 200 million people, predominantly stimulating a Type 2 immune response. Protective immunity takes decades to become effective due to dominant immune suppression that develops during infection. Using Litomosoides sigmodontis infection as a murine model of filariasis, we previously demonstrated that PD-1 co-inhibition causes Th2 cells to become intrinsically dysfunctional or hypo-responsive during infection, resulting in impaired protective immunity. Τhis study demonstrates that Th2 cell-intrinsic hypo-responsiveness is associated with a loss of GATA-3 expression by CD4+ T cells from WT, but not PD-L2-/- mice. PD-L2-/- mice were more resistant to L. sigmodontis and had increased Th2 cell-intrinsic functional quality. CD19+ B cells expressed PD-L2, and Jh-/- B cell-deficient mice were more resistant to infection. However, Th2 cell-intrinsic hypo-responsiveness still developed in Jh-/- mice, and restricting PD-L2 deficiency to B cells using bone marrow chimaeras did not alter resistance to L. sigmodontis or Th2 cell-intrinsic functional quality. Together, these data indicate that PD-L2 inhibits protective immunity to L. sigmodontis, downregulates GATA-3 in CD4+ T cells, and induces Th2 cell-intrinsic hypo-responsiveness. Whilst B cells play a suppressive role during infection, this is independent of PD-L2 and B cells do not instigate Th2 cell-intrinsic hypo-responsiveness.

Objectives: Clostridioides difficile infection (CDI) is the leading hospital-acquired infection in North America. We have previously discovered that antibiotic disruption of the gut microbiota decreases intestinal IL-33 and IL-25 and increases susceptibility to CDI. We further found that IL-33 promotes protection through type 2 Innate Lymphoid Cells (ILC2s), which produce IL-13. However, the contribution of IL-13 to disease has never been explored. Methods: We used a validated model of CDI in mice, in which we neutralized via blocking antibodies, or administered recombinant protein, IL-13 to assess the role of this cytokine during infection using weight and clinical scores. Fluorescent activated cell sorting (FACS) was used to characterize myeloid cell population changes in response to IL-13 manipulation. Results: We found that administration of IL-13 protected, and anti-IL-13 exacerbated CDI. Additionally, we observed alterations to the monocyte/macrophage cells following neutralization of IL-13 as early as day three post infection. We also observed elevated accumulation of myeloid cells by day four post-infection following IL13 neutralization. Neutralization of the decoy receptor, IL-13R alpha 2, resulted in protection from disease, likely through increased available endogenous IL-13. Conclusions: Our data highlight the protective role of IL-13 in protecting from more severe CDI and the association of poor responses with a dysregulated monocyte-macrophage compartment. These results increase our understanding of type 2 immunity in CDI and may have implications for treating disease in patients.

Background We previously conducted a phase 2a randomized placebo-controlled trial of 40 subjects to assess the efficacy and safety of dupilumab use in people hospitalized with coronavirus disease 2019 (COVID-19) (NCT04920916). Based on our preclinical data suggesting that downstream pulmonary dysfunction with COVID-19 induced type 2 inflammation, we contacted patients from our phase 2a study at 1 year for assessment of post-COVID-19 conditions.Methods Subjects at 1 year after treatment underwent pulmonary function tests, high-resolution computed tomographic imaging, symptom questionnaires, neurocognitive assessments, and serum immune biomarker analysis, with subject survival also monitored. The primary outcome was the proportion of abnormal diffusion capacity for carbon monoxide (DLCO) or 6-minute walk test (6MWT) at the 1-year visit.Results Of those survivors who consented to 1-year visits (n = 16), subjects who had originally received dupilumab were less likely than those who received placebo to have an abnormal DLCO or 6MWT (Fisher exact P = .011; adjusted P = .058). As a secondary endpoint, we saw that 16% of subjects in the dupilumab group died by 1 year compared to 38% in the placebo group, though this was not statistically significant (log-rank P = .12). We did not find significant differences in neurocognitive testing, symptoms, or chest computed tomography between treatment groups but observed a larger reduction in eotaxin levels in those who received dupilumab.Conclusions In this observational study, subjects who received dupilumab during acute COVID-19 hospitalization were less likely to have a reduced DLCO or 6MWT, with a nonsignificant trend toward reduced mortality at 1 year compared to placebo. In this follow-up study, subjects who had received dupilumab during their acute COVID-19 hospitalization through a prior phase 2a randomized placebo-controlled trial were less likely to have reduced lung diffusion or abnormal 6-minute walk testing at 1 year postenrollment.

Macrophages activated via the IL-4 receptor possess non-immune functions that support tissue homeostasis, but their specific role in aging is unknown. In this issue of Immunity , Zhou et al. show that IL-4 extends lifespan by inducing DNA repair pathways that protect macrophages from cellular senescence.

Cutaneous T cell lymphoma (CTCL) is a potentially fatal clonal malignancy of T cells primarily affecting the skin. The most common form of CTCL, mycosis fungoides, can be difficult to diagnose, resulting in treatment delay. We performed single-cell and spatial transcriptomics analysis of skin from patients with mycosis fungoides-type CTCL and an integrated comparative analysis with human skin cell atlas datasets from healthy and inflamed skin. We revealed the co-optation of T helper 2 (T(H)2) cell-immune gene programs by malignant CTCL cells and modeling of the tumor microenvironment to support their survival. We identified MHC-II+ fibroblasts and dendritic cells that can maintain T(H)2 cell-like tumor cells. CTCL tumor cells are spatially associated with B cells, forming tertiary lymphoid structure-like aggregates. Finally, we validated the enrichment of B cells in CTCL and its association with disease progression across three independent patient cohorts. Our findings provide diagnostic aids, potential biomarkers for disease staging and therapeutic strategies for CTCL.

In mouse peritoneal and other serous cavities, the transcription factor GATA6 drives the identity of the major cavity resident population of macrophages, with a smaller subset of cavity-resident macrophages dependent on the transcription factor IRF4. Here we showed that GATA6+ macrophages in the human peritoneum were rare, regardless of age. Instead, more human peritoneal macrophages aligned with mouse CD206+ LYVE1+ cavity macrophages that represent a differentiation stage just preceding expression of GATA6. A low abundance of CD206+ macrophages was retained in C57BL/6J mice fed a high-fat diet and in wild-captured mice, suggesting that differences between serous cavity-resident macrophages in humans and mice were not environmental. IRF4-dependent mouse serous cavity macrophages aligned closely with human CD1c+CD14+CD64+ peritoneal cells, which, in turn, resembled human peritoneal CD1c+CD14−CD64− cDC2. Thus, major populations of serous cavity-resident mononuclear phagocytes in humans and mice shared common features, but the proportions of different macrophage differentiation stages greatly differ between the two species, and dendritic cell (DC2)-like cells were especially prominent in humans. Randolph and colleagues analyze the immune cells in the human and mouse peritoneum and show that the major populations of serous cavity-resident macrophages in humans and mice represent distinct differentiation stages of an overlapping differentiation program.

Background:We previously conducted a Phase IIa randomized placebo-controlled trial of 40 subjects to assess the efficacy and safety of dupilumab use in those hospitalized with COVID-19 (NCT04920916). Based on our pre-clinical data suggesting downstream pulmonary dysfunction with COVID-19 induced type 2 inflammation, we contacted patients from our Phase IIa study at 1 year for assessment of Post Covid-19 Conditions (PCC).Methods:Subjects at 1 year after treatment underwent pulmonary function testing (PFTs), high resolution computed tomography (HRCT) imaging, symptom questionnaires, neurocognitive assessments, and serum immune biomarker analysis, with subject survival also monitored. The primary outcome was the proportion of abnormal PFTs, defined as an abnormal diffusion capacity for carbon monoxide (DLCO) or 6-minute walk testing (6MWT) at the 1-year visit.Results:Sixteen of the 29 one-year survivors consented to the follow up visit. We found that subjects who had originally received dupilumab were less likely to have abnormal PFTs compared to those who received placebo (Fisher's exact p=0.011, adjusted p=0.058). We additionally found that 3 out of 19 subjects (16%) in the dupilumab group died by 1 year compared to 8 out of 21 subjects (38%) in the placebo group (log rank p=0.12). We did not find significant differences in neurocognitive testing, symptoms or CT chest imaging between treatment groups but observed evidence of reduced type 2 inflammation in those who received dupilumab.Conclusions:We observed evidence of reduced long-term morbidity and mortality from COVID-19 with dupilumab treatment during acute hospitalization when added to standard of care regimens.

Abstract In murine peritoneal and other serous cavities, Gata6 drives identity of the major resident cavity macrophage population, with a smaller subset of cavity macrophages dependent upon Irf4. Here we show that Gata6+ macrophages in the human peritoneum are rare, regardless of age. Instead, many aligned with CD206 + LYVE1 + mouse cavity macrophages from a differentiation stage just preceding expression of Gata6. The comparatively low abundance of CD206 + converting macrophages in healthy C57BL/6J mice was sustained after high-fat diet feeding or in mice capture from the wild, suggesting that the major resident macrophages between the species share overlapping differentiation programs but differing predispositions to distinct differentiation stages. Irf4-dependent mouse serous cavity macrophages aligned closely with CD1c + CD14 + CD64 + human peritoneal dendritic cells (DCs) that, in turn, resembled CD1c + CD14 - CD64 - peritoneal cDC2. Peritoneal DC1 and plasmacytoid DCs were rare in both species. This cross-species resource comparing mouse and human serous cavity immune cells will facilitate translational research.

The larval stages of the cestode parasites belonging to the genus Echinococcus grow within internal organs of humans and a range of animal species. The resulting diseases, collectively termed echinococcoses, include major neglected tropical diseases of humans and livestock. Echinococcus larvae are outwardly protected by the laminated layer (LL), an acellular structure that is unique to this genus. The LL is based on a fibrillar meshwork made up of mucins, which are decorated by galactose-rich O-glycans. In addition, in the species cluster termed E. granulosus sensu lato, the LL features nano-deposits of the calcium salt of myo-inositol hexakisphosphate (Insp6). The main purpose of our article is to update the immunobiology of the LL. Major recent advances in this area are (i) the demonstration of LL "debris" at the infection site and draining lymph nodes, (ii) the characterization of the decoy activity of calcium Insp6 with respect to complement, (iii) the evidence that the LL mucin carbohydrates interact specifically with a lectin receptor expressed in Kupffer cells (Clec4F), and (iv) the characterization of what appear to be receptor-independent effects of LL particles on dendritic cells and macrophages. Much information is missing on the immunology of this intriguing structure: we discuss gaps in knowledge and propose possible avenues for research.

For decades, immunologists have studied the role of circulating immune cells in host protection, with a more recent appreciation of immune cells resident within the tissue microenvironment and the intercommunication between nonhematopoietic cells and immune cells. However, the extracellular matrix (ECM), which comprises at least a third of tissue structures, remains relatively underexplored in immunology. Similarly, matrix biologists often overlook regulation of complex structural matrices by the immune system. We are only beginning to understand the scale at which ECM structures determine immune cell localization and function. Additionally, we need to better understand how immune cells dictate ECM complexity. This review aims to highlight the potential for biological discovery at the interface of immunology and matrix biology.

Chitinase-like proteins (CLPs) are biomarkers of inflammation and airway remodelling in asthma, yet their direct contribution towards disease pathogenesis is unknown. Using a mouse model of allergen-induced type 2/type 17 airway inflammation we sought to directly investigate the role of the murine CLPs Ym1 and Ym2 during chronic lung pathology. Data demonstrated distinct chronic inflammatory roles for Ym2, IL-13, and IL-17a signalling pathways. Notably, only CLPs were key for initiating the pathogenic accumulation and re-organisation of the pulmonary extracellular matrix (ECM) environment. Furthermore, inhibition of CLPs after chronic pathology developed, reversed airway remodelling independently of chronic inflammation. These studies disentangle chronic IL-13 and IL-17a signalling from the development of allergic airway remodelling and instead highlight a central role for CLPs, which provides new avenues to therapeutically target aberrant ECM accumulation.

Immune system development is greatly influenced by vertically transferred cues. However, beyond antibody-producing B cells, little is known about the role of other cell subsets of the maternal immune system in regulating offspring immunity. We reasoned γδ T cells to be attractive candidates based on their tissue distribution pattern: abundant in the skin, mammary glands and female reproductive tract. Here we found that mice born from γδ T cell-deficient (TCRδ-/-) dams display, early after birth, a pulmonary milieu selectively enriched in type-2 cytokines such as IL-33, IL-4, IL-5, and IL-13, and type-2-polarized immune cells, when compared to the progeny of γδ T cell-sufficient dams. In addition, upon helminth infection, mice born from TCRδ-/- dams sustained an increased type-2 inflammatory response. Critically, this was independent of the genotype of the pups. Despite similar levels of circulating antibodies in mothers and progeny, the intestinal microbiota in the offspring of TCRδ-/- and TCRδ+/- dams harbored distinct bacterial communities acquired during birth and fostering. These differences were accompanied by changes in the intestinal short-chain fatty acids (SCFA) profile. Importantly, antibiotic treatment abrogated the differences observed in the pulmonary milieu, and exogenous SCFA supplementation suppressed first-breath- and infection-induced inflammation. In summary, maternal γδ T cells control the establishment of a neonatal gut–lung axis by conditioning the postnatal microbial colonization of the off-spring and bacterial-derived metabolite availability; ultimately impacting on the development of pulmonary type-2 immunity in the offspring.

Acquired immunity to gastrointestinal nematodes reduces during late pregnancy and lactation which is known as periparturient relaxation of immunity (PPRI). Protein supplementation reduces the degree of PPRI in a rat model re-infected with Nippostrongylus brasiliensis, but the underlying molecular mechanisms have yet to be elucidated. Here, we hypothesized that protein supplementation will enhance T helper type 2 immunity (Th2) in the lung and small intestine. Nulliparous Sprague-Dawley rats were given a primary infection of N. brasiliensis prior to mating and restrictedly fed diets with either low protein (LP) or high protein (HP) during pregnancy and lactation. Dams were secondary infected with N. brasiliensis on day 2 post-parturition, and histology and gene expression were analysed for tissue samples collected at days 5, 8 and 11. Genes related to Th2 immunity in the lung, Retnla, Il13 and Mmp12, and in the intestine, Retnlb, were upregulated in HP dams compared to LP dams, which indicates the effect of dietary protein on Th2 immunity. HP dams also had increased splenic CD68(+) macrophage populations compared to LP dams following secondary infection, suggesting enhanced immunity at a cellular level. Our data assist to define strategic utilization of nutrient supply in mammals undergoing reproductive and lactational efforts.

Peritoneal adhesions commonly occur after abdominal or pelvic surgery. These scars join internal organs to each other or to the cavity wall and can present with abdominal or pelvic pain, and bowel obstruction or female infertility. The mechanisms underlying adhesion formation remain unclear and thus, effective treatments are not forthcoming. Peritoneal macrophages accumulate after surgery and previous studies have attributed either pro- or anti-scarring properties to these cells. We propose that there are complex and nuanced responses after surgery with respect to both resident and also monocyte-derived peritoneal macrophage subpopulations. Moreover, we contend that differences in responses of specific macrophage subpopulations in part explain the risk of developing peritoneal scars. We characterized alterations in peritoneal macrophage subpopulations after surgery-induced injury using two strains of mice, BALB/c and C57BL/6, with known differences in macrophage response post-infection. At 14 days post-surgery, BALB/c mice displayed more adhesions compared with C57BL/6 mice. This increase in scarring correlated with a lower influx of monocyte-derived macrophages at day 3 post-surgery. Moreover, BALB/c mice showed distinct macrophage repopulation dynamics after surgery. To confirm a role for monocyte-derived macrophages, we used Ccr2 -deficient mice as well as antibody-mediated depletion of CCR2 expressing cells during initial stages of adhesion formation. Both Ccr2 -deficient and CCR2-depleted mice showed a significant increase in adhesion formation associated with the loss of peritoneal monocyte influx. These findings revealed an important protective role for monocyte-derived cells in reducing adhesion formation after surgery.