CD11c+ dendritic cells PlexinD1 deficiency exacerbates airway hyperresponsiveness, IgE and mucus production in a mouse model of allergic asthma.

Dendritic cells (DCs) are pivotal in regulating allergic asthma. Our research has shown that the absence of Sema3E worsens asthma symptoms in acute and chronic asthma models. However, the specific role of PlexinD1 in these processes, particularly in DCs, remains unclear. This study investigates the role of PlexinD1 in CD11c+ DCs using a house dust mite (HDM) model of asthma. We generated CD11c+ DC-specific PlexinD1 knockout (CD11cPLXND1 KO) mice and subjected them, alongside wild-type controls (PLXND1fl/fl), to an HDM allergen protocol. Airway hyperresponsiveness (AHR) was measured using FlexiVent, and immune cell populations were analyzed via flow cytometry. Cytokine levels and immunoglobulin concentrations were assessed using mesoscale and ELISA, while collagen deposition and mucus production were examined through Sirius-red and periodic acid Schiff (PAS) staining respectively. Our results indicate that CD11cPLXND1 KO mice exhibit significantly exacerbated AHR, characterized by increased airway resistance and tissue elastance. Enhanced mucus production and collagen gene expression were observed in these mice compared to wild-type counterparts. Flow cytometry revealed higher CD11c+ MHCIIhigh CD11b+ cell recruitment into the lungs, and elevated total and HDM-specific serum IgE levels in CD11cPLXND1 KO mice. Mechanistically, co-cultures of B cells with DCs from CD11cPLXND1 KO mice showed significantly increased IgE production compared to wild-type mice.These findings highlight the critical regulatory role of the plexinD1 signaling pathway in CD11c+ DCs in modulating asthma features.

Targeting the Semaphorin3E-plexinD1 complex in allergic asthma.

Asthma is a heterogenous airway disease characterized by airway inflammation and remodeling. It affects more than 300 million people worldwide and poses a significant burden on society. Semaphorins, discovered initially as neural guidance molecules, are ubiquitously expressed in various organs and regulate multiple signaling pathways. Interestingly, Semaphorin3E is a critical molecule in lung pathophysiology through its role in both lung development and homeostasis. Semaphorin3E binds to plexinD1, mediating regulatory effects on cell migration, proliferation, and angiogenesis. Recent in vitro and in vivo studies have demonstrated that the Semaphorin3E-plexinD1 axis is implicated in asthma, impacting inflammatory and structural cells associated with airway inflammation, tissue remodeling, and airway hyperresponsiveness. This review details the Semaphorin3E-plexinD1 axis in various aspects of asthma and highlights future directions in research including its potential role as a therapeutic target in airway allergic diseases.

PlexinD1 Deficiency in Lung Interstitial Macrophages Exacerbates House Dust Mite-Induced Allergic Asthma.

Interstitial macrophages (IMs) are key regulators of allergic inflammation. We previously showed that the absence of semaphorin 3E (Sema3E) exacerbates asthma features in both acute and chronic asthma models. However, it has not been studied whether Sema3E, via its receptor plexinD1, regulates IM function in allergic asthma. Therefore, we investigated the role of plexinD1 deficiency on IMs in allergic asthma. We found that the absence of plexinD1 in IMs increased airway hyperresponsiveness, airway leukocyte numbers, allergen-specific IgE, goblet cell hyperplasia, and Th2/Th17 cytokine response in the house dust mite (HDM)-induced allergic asthma model. Muc5ac, Muc5b, and α-SMA genes were increased in mice with Plxnd1-deficient IMs compared with wild-type mice. Furthermore, plexinD1-deficient bone marrow-derived macrophages displayed reduced IL-10 mRNA expression, at both the baseline and following HDM challenge, compared with their wild-type counterpart mice. Our data suggest that Sema3E/plexinD1 signaling in IMs is a critical pathway that modulates airway inflammation, airway resistance, and tissue remodeling in the HDM murine model of allergic asthma. Reduced IL-10 expression by plexinD1-deficient macrophages may account for these enhanced allergic asthma features.

PTX3 Deficiency Promotes Enhanced Accumulation and Function of CD11c+CD11b+ DCs in a Murine Model of Allergic Inflammation.

PTX3 is a unique member of the long pentraxins family and plays an indispensable role in regulating the immune system. We previously showed that PTX3 deletion aggravates allergic inflammation via a Th17 -dominant phenotype and enhanced CD4 T cell survival using a murine model of ovalbumin (OVA) induced allergic inflammation. In this study, we identified that upon OVA exposure, increased infiltration of CD11c+CD11b+ dendritic cells (DCs) was observed in the lungs of PTX3-/- mice compared to wild type littermate. Further analysis showed that a short-term OVA exposure led to an increased number of bone marrow common myeloid progenitors (CMP) population concomitantly with increased Ly6Chigh CCR2high monocytes and CD11c+CD11b+ DCs in the lungs. Also, pulmonary CD11c+CD11b+ DCs from OVA-exposed PTX3-/- mice exhibited enhanced expression of maturation markers, chemokines receptors CCR2, and increased OVA uptake and processing compared to wild type controls. Taken together, our data suggest that PTX3 deficiency heightened lung CD11c+CD11b+DC numbers and function, hence exacerbating airway inflammatory response.

Semaphorin 3E deficiency dysregulates dendritic cell functions: In vitro and in vivo evidence.

Regulation of dendritic cell functions is a complex process in which several mediators play diverse roles as a network in a context-dependent manner. The precise mechanisms underlying dendritic cell functions have remained to be addressed. Semaphorins play crucial roles in regulation of various cell functions. We previously revealed that Semaphorin 3E (Sema3E) contributes to regulation of allergen-induced airway pathology partly mediated by controlling recruitment of conventional dendritic cell subsets in vivo, though the underlying mechanism remained elusive. In this study, we investigate the potential regulatory role of Sema3E in dendritic cells. We demonstrated that bone marrow-derived dendritic cells differentiated from Sema3e-/- progenitors have an enhanced migration capacity both at the baseline and in response to CCL21. The enhanced migration ability of Sema3E dendritic cells was associated with an overexpression of the chemokine receptor (CCR7), elevated Rac1 GTPase activity and F-actin polymerization. Using a mouse model of allergic airway sensitization, we observed that genetic deletion of Sema3E leads to a time dependent upregulation of CCR7 on CD11b+ conventional dendritic cells in the lungs and mediastinal lymph nodes. Furthermore, aeroallergen sensitization of Sema3e-/- mice lead to an enhanced expression of PD-L2 and IRF-4 as well as enhanced allergen uptake in pulmonary CD11b+ DC, compared to wild type littermates. Collectively, these data suggest that Sema3E implicates in regulation of dendritic cell functions which could be considered a basis for novel immunotherapeutic strategies for the diseases associated with defective dendritic cells in the future.

Semaphorin 3E Protects against Chlamydial Infection by Modulating Dendritic Cell Functions.

Recent studies have identified semaphorin 3E (Sema3E) as a novel mediator of immune responses. However, its function in immunity to infection has yet to be investigated. Using a mouse model of chlamydial lung infection, we show that Sema3E plays a significant role in the host immune response to the infection. We found that Sema3E is induced in the lung after chlamydial infection, and Sema3E deficiency has a detrimental impact on disease course, dendritic cell (DC) function, and T cell responses. Specifically, we found that Sema3E knockout (KO) mice exhibited higher bacterial burden, severe body weight loss, and pathological changes after Chlamydia muridarum lung infection compared with wild-type (WT) mice. The severity of disease in Sema3E KO mice was correlated with reduced Th1/Th17 cytokine responses, increased Th2 response, altered Ab response, and a higher number of regulatory CD4 T cells. Moreover, DCs isolated from Sema3E KO mice showed lower surface expression of costimulatory molecules and production of IL-12, but higher expression of PD-L1, PD-L2, and IL-10 production. Functional DC-T cell coculture studies revealed that DCs from infected Sema3E KO mice failed to induce Th1 and Th17 cell responses compared with DCs from infected WT mice. Upon adoptive transfer, mice receiving DCs from Sema3E KO mice, unlike those receiving DCs from WT mice, were not protected against challenge infection. In conclusion, our data evidenced that Sema3E acts as a critical factor for protective immunity against intracellular bacterial infection by modulating DC functions and T cell subsets.

Semaphorin 3E Promotes Susceptibility to Leishmania major Infection in Mice by Suppressing CD4+ Th1 Cell Response.

Protective immunity to cutaneous leishmaniasis is mediated by IFN-γ-secreting CD4+ Th1 cells. IFN-γ binds to its receptor on Leishmania-infected macrophages, resulting in their activation, production of NO, and subsequent destruction of parasites. This study investigated the role of Semaphorin 3E (Sema3E) in host immunity to Leishmania major infection in mice. We observed a significant increase in Sema3E expression at the infection site at different timepoints following L. major infection. Sema3E-deficient (Sema3E knockout [KO]) mice were highly resistant to L. major infection, as evidenced by significantly (p < 0.05-0.01) reduced lesion sizes and lower parasite burdens at different times postinfection when compared with their infected wild-type counterpart mice. The enhanced resistance of Sema3E KO mice was associated with significantly (p < 0.05) increased IFN-γ production by CD4+ T cells. CD11c+ cells from Sema3E KO mice displayed increased expression of costimulatory molecules and IL-12p40 production following L. major infection and were more efficient at inducing the differentiation of Leishmania-specific CD4+ T cells to Th1 cells than their wild-type counterpart cells. Furthermore, purified CD4+ T cells from Sema3E KO mice showed increased propensity to differentiate into Th1 cells in vitro, and this was significantly inhibited by the addition of recombinant Sema3E in vitro. These findings collectively show that Sema3E is a negative regulator of protective CD4+ Th1 immunity in mice infected with L. major and suggest that its neutralization may be a potential therapeutic option for treating individuals suffering from cutaneous leishmaniasis.

The Long Pentraxin 3 (PTX3) Suppresses Immunity to Cutaneous Leishmaniasis by Regulating CD4+ T Helper Cell Response.

The long pentraxin 3 (PTX3) plays a critical role in inflammation, tissue repair, and wound healing. Here, we show that PTX3 regulates disease pathogenesis in cutaneous leishmaniasis (CL). PTX3 expression increases in skin lesions in patients and mice during CL, with higher expression correlating with severe disease. PTX3-deficient (PTX3-/-) mice are highly resistant to L. major and L. braziliensis infections. This enhanced resistance is associated with increases in Th17 and IL-17A responses. The neutralization of IL-17A abolishes this enhanced resistance, while rPTX3 treatment results in decrease in Th17 and IL-17A responses and increases susceptibility. PTX3-/- CD4+ T cells display increased differentiation to Th17 and expression of Th17-specific transcription factors. The addition of rPTX3 suppresses the expression of Th17 transcription factors, Th17 differentiation, and IL-17A production by CD4+ T cells from PTX3-/- mice. Collectively, our results show that PTX3 contributes to the pathogenesis of CL by negatively regulating Th17 and IL-17A responses.

Semaphorin 3E Regulates the Response of Macrophages to Lipopolysaccharide-Induced Systemic Inflammation.

Semaphorin 3E (Sema3E) is a secreted protein that was initially discovered as a neuronal guidance cue. Recent evidence showed that Sema3E plays an essential role in regulating the activities of various immune cells. However, the exact role of Sema3E in macrophage function, particularly during inflammation, is not fully understood. We studied the impact of Sema3E gene deletion on macrophage function during the LPS-induced acute inflammatory response. We found that Sema3E-deficient (Sema3e-/- ) mice were better protected from LPS-induced acute inflammation as exemplified by their superior clinical score and effective temperature control compared with their wild-type littermates. This superior control of inflammatory response in Sema3e-/- mice was associated with significantly lower phosphorylation of ERK1/2, AKT, STAT3, and NF-κB, and a concomitant reduction in inducible NO synthase expression and production of TNF and IL-6 compared with their Sema3e+/+ littermates. Sema3e-/- mice also contained significantly higher numbers of activated macrophages compared with their Sema3e+/+ littermates at both baselines and after LPS challenge. In vivo-specific deletion of the Sema3E high-affinity receptor, plexinD1, on macrophages led to the improvement in clinical disease following exposure to a lethal dose of LPS. Collectively, our data show that Sema3E plays an essential role in dampening the early inflammatory response to LPS by regulating macrophage function, suggesting an essential role of this pathway in macrophage inflammatory response.

Glucocorticoids regulate pentraxin-3 expression in human airway smooth muscle cells.

Pentraxin-3 (PTX3) is a multifunctional protein involved in both innate and adaptive immunity. Glucocorticoid (GC) is the first-line therapy to mitigate airway inflammation in asthma. Previous pieces of evidence showed that GC has divergent effects on PTX3 production in various cell types. The molecular mechanisms controlling PTX3 expression in HASMC are, however, not yet characterized. In this study, we demonstrate that the synthetic GC, dexamethasone (DEX) increases the expression of PTX3 both at the protein and mRNA levels. We also found that such an effect of DEX was dependent on de novo protein synthesis and the GC receptor (GR). While DEX increases PTX3 mRNA stability, it did not affect its promoter activity. Interestingly, HASMC pre-treated with p42/p44 ERK inhibitor, but not with p38 or JNK-MAPK inhibitors, significantly interfered with DEX-induced PTX3 secretion. Taken together, our data suggest that GC regulates PTX3 expression in HASMC through transcriptional and post-transcriptional mechanisms in a GR and ERK-dependent manner.

Semaphorin 3E Inhibits House Dust Mite-Induced Angiogenesis in a Mouse Model of Allergic Asthma.

Increased angiogenesis is a characteristic feature of remodeling in asthmatic airways and stems from the imbalance between pro-angiogenic and anti-angiogenic factors. Surprisingly, the factors regulating this process in allergic asthma are poorly defined. Previously, we showed an important role of semaphorins 3E (Sema3E) in growth factor-induced airway smooth muscle proliferation and migration in vitro, and in down-regulating airway inflammation, T helper 2/T helper 17 cytokine response, mucus cell hyperplasia, and airway hyperresponsiveness in vivo. However, the role of Sema3E in airway angiogenesis is not fully understood. Here, we investigated the role of Sema3E in airway angiogenesis using a house dust mite (HDM) murine model of allergic asthma. Intranasal treatment with recombinant Sema3E significantly reduced the expression of angiogenesis markers within the airways of HDM-challenged mice compared with untreated mice. HDM-induced expression of vascular endothelial growth factor (VEGF) and VEGF receptor 2 protein were diminished substantially on Sema3E treatment. Interestingly, Sema3E-treated mice showed an enhanced expression of the negative regulator of angiogenesis, soluble VEGF receptor 1, compared with the untreated mice. These events were reversed in Sema3E-deficient mice at baseline or on HDM challenge. Taken together, this study provides the first evidence that Sema3E modulates angiogenesis in allergic asthmatic airways via modulating pro- and anti-angiogenic factors.

The regulatory role of semaphorin 3E in allergic asthma.

Semaphorins were originally discovered as essential mediators involved in regulation of axonal growth during development of the nervous system. Ubiquitously expressed on various organs, they control several cellular functions by regulating essential signaling pathways. Among them, semaphorin3E binds plexinD1 as the primary receptor and mediates regulatory effects on cell migration, proliferation, and angiogenesis considered major physiological and pathological features in health and disease. Recent in vitro and in vivo experimental evidence demonstrate a key regulator role of semaphorin3E on airway inflammation, hyperresponsivenss and remodeling in allergic asthma. Herein, we aim to provide a broad overview of the biology of semaphorin family and review the recently discovered regulatory role of semaphorin3E in modulating immune cells and structural cells function in the airways. These findings support the concept of semaphorin3E/plexinD1 axis as a therapeutic target in allergic asthma.

Downregulation of semaphorin 3E promotes hallmarks of experimental chronic allergic asthma.

Guidance cues such as semaphorins are attractive novel therapeutic targets for allergic disorders. We have previously described an inhibitory effect of semaphorin 3E (Sema3E) on human airway smooth muscle cell function. We have further addressed a canonical role for Sema3E in acute model of allergic asthma in vivo. Considering the chronic nature of the disease, the potential implication of Sema3E to alleviate long-lasting deficits should be investigated. Expression of Sema3E in a chronic model of allergic asthma was assessed after exposure to house dust mite (HDM) as a clinically relevant allergen. Chronic features of allergic asthma including airway hyper-responsiveness (AHR), inflammation, and remodeling were studied in Sema3E-deficient mice. Additionally, the effect of exogenous Sema3E treatment was evaluated in prophylactic and therapeutic experimental models. We have demonstrated that expression of Sema3E is robustly suppressed in the airways upon chronic HDM exposure. Chronic allergic airway disease was significantly augmented in Sema3E-deficient mouse model which was associated with an increased AHR, remodeling, and Th2/Th17 inflammation. Intranasal Sema3E administration restored chronic deficits of allergic asthma in mice. Data from this study unveil a key regulatory role of Sema3E in chronic course of asthma via orchestration of impaired inflammatory and remodeling responses.

Semaphorin 3E Alleviates Hallmarks of House Dust Mite-Induced Allergic Airway Disease.

Semaphorins are an essential family of guidance cues ubiquitously expressed in various organs, which play diverse developmental, homeostatic, and pathological roles. Semaphorin 3E (Sema3E), initially identified as a neuronal chemorepellent, is involved in the regulation of cell migration, proliferation, and angiogenesis. However, expression and function of Sema3E in allergic asthma has not been extensively investigated. We determined the expression of Sema3E in the airways and its effect on airway inflammation, hyperresponsiveness, and remodeling as pathological features of allergic asthma provoked by house dust mite in vivo. Our data indicate that exposure to house dust mite markedly reduces Sema3E expression in mouse airways. More important, replenishment of Sema3E by intranasal administration of exogenous Sema3E protects mice from allergic asthma by reducing eosinophilic inflammation, serum IgE level, and T helper cell 2/T helper cell 17 cytokine response. The regulatory effect of Sema3E on cytokine response was sustained on allergen recall response in the lymph nodes and spleen. Furthermore, goblet cell hyperplasia, collagen deposition, and airway hyperresponsiveness were significantly diminished on Sema3E treatment. The inhibitory effect of Sema3E was associated with a reduction of pulmonary CD11b+ conventional dendritic cells and regulation of CD4+ T-cell cytokine response. Collectively, our data represent a novel approach to treating allergic asthma via regulation of immune response to house dust mite.

Semaphorin 3E Deficiency Exacerbates Airway Inflammation, Hyperresponsiveness, and Remodeling in a Mouse Model of Allergic Asthma.

Semaphorin 3E (Sema3E) plays a crucial role in axon guidance, vascular patterning, and immune regulation. Nevertheless, the role of Sema3E in asthma is still elusive. In this study, we show that genetic ablation of Sema3E in mice results in increased lung granulocytosis, airway hyperresponsiveness, mucus overproduction, collagen deposition, and Th2/Th17 inflammation. Transfer of Sema3e-/- bone marrow progenitor cells to irradiated wild-type (WT) recipients exacerbates airway hyperresponsiveness and inflammation, whereas transfer of WT bone marrow progenitor cells ameliorates asthma pathology in Sema3e-/- recipients. Sema3e-/- mice display a higher frequency of CD11b+ pulmonary dendritic cells than their WT controls at the baseline and after sensitization with house dust mite. Adoptive transfer of CD11b+ pulmonary dendritic cells from Sema3e-/- mice into WT recipients increases house dust mite-induced Th2/Th17 inflammation in the airway. Together, these findings identify Sema3E as a novel regulatory molecule in allergic asthma that acts upstream of proallergic events and suggest that targeting this molecule could be a novel approach to treat allergic asthma.

Pentraxin 3 deletion aggravates allergic inflammation through a TH17-dominant phenotype and enhanced CD4 T-cell survival.

BACKGROUND: Pentraxin 3 (PTX3) is a multifunctional molecule that plays a nonredundant role at the crossroads between pathogen clearance, innate immune system, matrix deposition, female fertility, and vascular biology. It is produced at sites of infection and inflammation by both structural and inflammatory cells. However, its role in allergen-induced inflammation remains to be tested. OBJECTIVE: We sought to determine the effect of Ptx3 deletion on ovalbumin (OVA)-induced allergic inflammation in a murine model of asthma. METHODS: Bronchoalveolar lavage fluid was collected from patients with severe asthma and healthy subjects, and the level of PTX3 was determined by using ELISA. Ptx3+/+ and Ptx3-/- mice were sensitized and challenged with OVA and bronchoalveolar lavage fluid, and the lungs were collected for assessing inflammation. Lung tissue inflammation and mucus production were assessed by means of flow cytometry and hematoxylin and eosin and periodic acid-Schiff staining, respectively. flexiVent was used to determine airway resistance to methacholine in these mice. RESULTS: Here we report that mice with severe asthma and OVA-sensitized/challenged mice had increased PTX3 levels in the lungs compared with healthy control mice. Mice lacking PTX3 have exaggerated neutrophilic/eosinophilic lung inflammation, mucus production, and airway hyperresponsiveness in an experimental model of OVA-induced asthma. Furthermore, OVA-exposed lung Ptx3-/- CD4 T cells exhibit an increased production of IL-17A, an effect that is accompanied by an increased signal transducer and activator of transcription 3 phosphorylation, reduced IL-2 production, and enhanced activation and survival. Also, we observed an increase in numbers of IL-6- and IL-23-producing dendritic cells in OVA-exposed Ptx3-/- mice compared with those in wild-type control mice. CONCLUSION: Altogether, PTX3 deficiency results in augmented airway hyperresponsiveness, mucus production, and IL-17A-dominant pulmonary inflammation, suggesting a regulatory role of PTX3 in the development of allergic inflammation.

Chemorepellent Semaphorin 3E Negatively Regulates Neutrophil Migration In Vitro and In Vivo.

Neutrophil migration is an essential step in leukocyte trafficking during inflammatory responses. Semaphorins, originally discovered as axon guidance cues in neural development, have been shown to regulate cell migration beyond the nervous system. However, the potential contribution of semaphorins in the regulation of neutrophil migration is not well understood. This study examines the possible role of a secreted chemorepellent, Semaphorin 3E (Sema3E), in neutrophil migration. In this study, we demonstrated that human neutrophils constitutively express Sema3E high-affinity receptor, PlexinD1. Sema3E displayed a potent ability to inhibit CXCL8/IL-8-induced neutrophil migration as determined using a microfluidic device coupled to real-time microscopy and a transwell system in vitro. The antimigratory effect of Sema3E on human neutrophil migration was associated with suppression of CXCL8/IL-8-mediated Ras-related C3 botulinum toxin substrate 1 GTPase activity and actin polymerization. We further addressed the regulatory role of Sema3E in the regulation of neutrophil migration in vivo. Allergen airway exposure induced higher neutrophil recruitment into the lungs of Sema3e-/- mice compared with wild-type controls. Administration of exogenous recombinant Sema3E markedly reduced allergen-induced neutrophil recruitment into the lungs, which was associated with alleviation of allergic airway inflammation and improvement of lung function. Our data suggest that Sema3E could be considered an essential regulatory mediator involved in modulation of neutrophil migration throughout the course of neutrophilic inflammation.

Human airway smooth muscle cell proliferation from asthmatics is negatively regulated by semaphorin3A.

Airway smooth muscle (ASM) hyperplasia is a key feature of airway remodeling in development of lung diseases such as asthma. Anomalous proliferation of ASM cells directly contributes to ASM hyperplasia. However, the molecular mechanisms controlling ASM cell proliferation are not completely understood. Semaphorins are versatile regulators of various cellular processes including cell growth and proliferation. The role of semaphorins in ASM cell proliferation has remained to be addressed. Here, we report that semaphorin 3A (Sema3A) receptor, neuropilin 1 (Nrp1), is expressed on human ASM cells (HASMC) isolated from healthy and asthmatic donors and treatment of these cells with exogenous Sema3A inhibits growth factor-induced proliferation. Sema3A inhibitory effect on HASMC proliferation is associated with decreased tyrosine phosphorylation of PDGFR, downregulation of Rac1 activation, STAT3 and GSK-3ß phosphorylation. Bronchial sections from severe asthmatics displayed immunoreactivity of Nrp1, suggestive of functional contribution of Sema3A-Nrp1 axis in airway remodeling. Together, our data suggest Sema3A-Nrp1 signaling as a novel regulatory pathway of ASM hyperplasia.