bluuebunny/crossref_metadata_2025 huggingface dataset:
Oral epithelial IL-22/STAT3 signaling licenses IL-17-mediated immunity to oral mucosal candidiasis
Authors: Aggor Felix E.Y. , Break Timothy , Trevejo-Nunez Giraldina , Whibley Natasha , Coleman Bianca M. , Bailey Rachel , Kaplan Daniel H. , Naglik Julian R. , Shan Wei , Shetty Amol C. , McCracken Carrie , Durum Scott K. , Biswas Partha S. , Bruno Vincent M. , Kolls Jay K. , Lionakis Michail S. , Gaffen Sarah L.
Published: January 2023

Oropharyngeal candidiasis (OPC) is an opportunistic infection of the oral mucosa caused by the commensal fungus Candida albicans. IL-17 and IL-22 both mediate antifungal immunity yet activate distinct downstream signaling pathways. While much is known about IL-17-dependent immunity in OPC, the activities of IL-22 are less well delineated. We show that induction of Il22 is independent of Dectin-1, CARD9 and aryl hydrocarbon receptor (AhR) and is driven by IL-23 and the C. albicans pore forming peptide candidalysin. Despite similar induction requirements and cellular sources, IL-22 and IL-17 function non-redundantly during OPC and exert opposing roles in neutrophil recruitment. The IL-22 and IL-17 receptors are required in anatomically distinct locations; loss of IL-22RA1 in the oral basal epithelial layer (BEL) but not the suprabasal epithelial layer (SEL) causes susceptibility to OPC, whereas IL-17RA is needed in the SEL. Our data reveal that IL-22 is a major activator of STAT3 in the BEL during OPC. Moreover, loss of STAT3 in the BEL but not the SEL renders mice susceptible to OPC. Transcriptional profiling of RNASeq data linked IL-22/STAT3 to oral epithelial cell proliferation and survival, but also, unexpectedly, to driving an IL-17 gene signature. We show that IL-22 acts on the BEL to replenish the IL-17RA-expressing SEL, thereby restoring the ability of the oral epithelium to respond to IL-17. Consequently, IL-22 signaling in BEL ‘licenses’ IL-17R signaling in the oral epithelium, revealing spatially distinct yet cooperative activities of IL-22 and IL-17 in oral candidiasis. This work also suggests that oral thrush in Jobs’ syndrome patients may be caused by STAT3 impairments in the oral epithelium, not just Th17 cells.
A FUNCTIONAL TL1A/DR3 SYSTEM IS CRUCIAL FOR OPTIMAL GENERATION OF PATHOGENIC TH9 CELLS IN MURINE CROHN’S DISEASE-LIKE ILEITIS
Authors: Butto Ludovica , Menghini Paola , Aladyshkina Natalia , Buela Kristine-Ann , De Salvo Carlo , Osme Abdullah , Pizarro Theresa , Cominelli Fabio
Published: January 2023

New evidence suggests the implication of IL-9-secreting TH9 cells in the pathogenesis of Crohn’s disease (CD). The role of death receptor 3 (DR3) in the differentiation and function of TH9 cells in CD is not fully explored. Thus, here, we investigated the role of a functional DR3 receptor in disease progression and activation of downstream signaling pathways relevant to TH9 cells. We found that lack of DR3 ameliorated ileitis in SAMP/YitFc (SAMP,WT) mice, reduced by 40% Baft3, PU.1, il-9 mRNAs and IL-9 protein, increased by 3-fold Id3 mRNA in ileal tissues (P≤0.003). TH9 cells from DR3−/−×SAMP (KO) mice harbored 2-fold less il-9 mRNA and IL-9 and TNF proteins (P≤0.0002), 5-fold higher IL-10 protein than WT TH9 cells (P<0.0001). RNA-seq analysis of WT and KO TH9 cells is currently being analyzed. Rag2−/− recipients of KO TH9 cells developed less severe colitis (P=0.005). Histology score of ilea and colons from SAMP×Rag2−/− is currently being evaluated. The frequency of KO TH9 cells producing IL-9 in Rag2−/− MLNs was 2-fold lower than that of mice reconstituted with WT TH9 cells (P=0.02). The frequency of IL-9-producing KO TH9 cells in SAMP×Rag2−/− SPLs and MLNs was 2-fold lower than that of mice reconstituted with WT TH9 cells (P≤0.006). In conclusion, DR3 receptor heavily contributes to IL-9-signaling pathway activation and it is required for the secretion of higher amount of IL-9 protein by TH9 cells in SAMP mice. KO TH9 cells were less colitogenic that those isolated from WT mice, indicating a crucial role of a functional DR3 receptor in TH9 pathogenicity. Collectively, our results hold great translational significance by showing that modulation of DR3 signaling may be a novel therapeutic target for the treatment of CD.
Disturbances in Chemokine Pathways as a Mechanism for Poor Immune Containment of MTB in the setting of HIV Co-infection
Authors: Martinez-Martinez Yazmin Berenice , Huante Matthew B. , Chauhan Sadhana , Golovko George , Fofanov Yuriy , Gelman Benjamin , Endsley Janice
Published: January 2023

Tuberculosis (TB) is one of the ten top causes of death worldwide and the leading cause in people living with Human Immunodeficiency Virus (HIV). The loss of function by diverse leukocyte populations and poor organization of the protective granuloma structure due to HIV infection is associated with poor immunity and increased proliferation of Mtb bacilli in co-infection. We employed our humanized mouse model of Mtb and HIV co-infection to investigate the HIV-mediated disturbances in the chemokines and chemokine receptors that regulate the recruitment, activation, and localization of inflammatory leukocytes at sites of pulmonary infection. Lung tissue of mice that were non-infected or infected with Mtb, HIV, or Mtb/HIV was used for multiplex ELISA of chemokine profiles in supernatants and for high throughput RNA sequencing to identify differential transcription of chemokine and chemokine receptor genes. Our results show that Mtb infection of humanized mouse lung activates human chemokine pathways important for recruitment and effector function of leukocytes at sites of infection such as CXCL1, CXCL5, and CCL2. Co-infection with HIV, however, activated differential transcription of several chemokines and chemokine receptor families, compared to Mtb mono-infection. These outcomes were associated with greater bacterial and viral burden in co-infected animals and may identify candidate mechanisms for host directed therapies to reduce aggressive TB in those living with HIV.
Early B cell protein λ5 affects skeletal homeostasis
Authors: Khass Mohamed , Rashid Harunur , Burrows Peter D , Bridges Louis S , Javed Amjad , Schroeder Harry
Published: January 2023

There are strong indications that B lineage cells play a key role in bone homeostasis. The preB cell receptor is composed of an immunoglobulin mu H chain, VpreB and immunoglobulin λ5. Young λ5 deficient mice have reduced numbers of preB cells and mature B cells, moreover; their bone appears to be normal. With time, they achieve normal numbers of mature B cells while exhibiting major bone loss. To test whether expression of λ5 is playing a critical role maintaining bone homeostasis, we evaluated bone in: mice lacking immunoglobulin JH (JH −/− λ5+/+). These mice lack both preB and mature B cells, but maintain expression of λ5 in progenitor B cells. Mice deficient in both JH and λ5 (JH −/− λ5−/−) have no preB or mature B cells, and also no λ5. Using both μCT and histologic evaluation, we compared femur and skull at six months of age in these three mutant strains to normal controls. JH −/− λ5+/+ femur and skull were similar to WT. Both JH +/+λ5−/− and JH −/− λ5−/− mice showed significant decrease in femoral trabecular bone and increased loss of skull bone. Irradiated WT mice transplanted with JH −/− λ5−/− bone marrow cells developed defective femoral and cranial bones. In contrast, JH −/−λ5−/− mice that received WT bone marrow cells recovered bone architecture. In vitro culture of bone progenitors from JH +/+λ5−/− and JH −/− λ5−/− mice showed decreased ability of bone formation. These data underscore the importance of immunoglobulin λ5 in skeletal development.
The aged thymus is impaired in supporting T cell central tolerance
Authors: Lancaster Jessica N , Srinivasan Jayashree , Selden Hilary J. , Nam Seohee , Ehrlich Lauren Ilyse Richie
Published: January 2023

During aging, the thymus undergoes involution, decreasing dramatically in size and mass, and becoming increasingly disorganized. Notably, thymic epithelial cells decline with age, especially within the compartment of mature medullary thymic epithelial cells (mTECs) that express and present tissue-restricted antigens to thymocytes. Age-associated changes in the composition of hematopoietic antigen-presenting cells (APCs) have also been observed, but are not well characterized. Given that both mTECs and APCs present diverse self-antigens to thymocytes, we hypothesized that central tolerance is impaired in the aged thymus. Using live-cell two-photon microscopy, we determined that CD4-single positive (CD4SP) thymocytes, regardless of thymocyte age, had reduced motility and accumulation within the medulla of an aged thymus. We report age-associated changes in both the localization and composition of thymic APC subsets. To test whether this altered thymic environment functionally impacted central tolerance, we used thymic slice assays to measure tolerance induction of CD8− and CD4SP thymocytes encountering a range of model self-antigens. These assays indicated that central tolerance in the aged thymus was impaired, particularly against low-avidity auto-antigens. Our findings indicate that newly generated T cells selected in an aged thymic environment have not undergone a rigorous screening process against autoimmunity.
Histone Deacetylase 3 controls lung alveolar macrophage development and homeostasis by regulation of PPAR-γ expression through deacetylating PU.1
Authors: Yao Yi , Liu Queping , Adrianto Indra , Yin Congcong , Geissmann Frederic , Zhou Li , Mi Qing-Sheng
Published: January 2023

Alveolar macrophages (AMs) are derived from embryonic precursors that seed the lung before birth and self-maintain locally throughout adulthood but can be regenerated by bone marrow (BM) under stress conditions. However, how to precisely regulate AM development still remains unknown. Here we report that histone deacetylase 3 (HDAC3) is a key epigenetic factor required for AM embryonic development, postnatal homeostasis, maturation, and regeneration from BM. Loss of HDAC3 in early embryonic development affects AM development starting at E14.5, while loss of HDAC3 after birth affects AM homeostasis and maturation. HDAC3-deficient AMs exhibit severe mitochondrial oxidative dysfunction and deteriorative cell death. Single-cell RNA sequencing analyses further identified four distinct AM sub-clusters and a lack of HDAC3 dysregulated AM transcriptome in a cluster-specific manner. Mechanistically, HDAC3 serves as a co-activator via deacetylation of PU.1 to promote Pparg transcription. Our findings identify HDAC3 as a key epigenetic regulator of lung AM development and homeostasis.
Siglec-F-negative resident eosinophils represent an unexpectedly dominant population in homeostatic murine bone marrow
Authors: Jeong Brian M , Walker Matthew , Coden Mackenzie , Nam Kiwon , Nagasaka Reina , Abdala-Valencia Hiam , Berdnikovs Sergejs
Published: January 2023

Bone marrow, while known to be a hematopoietic site for eosinophils, also harbors resident populations of these cells. Identification of such resident eosinophils typically requires flow cytometry-assisted sorting, with Siglec-F as a key marker for identification of murine eosinophils. At baseline, we found that bone marrow resident eosinophils described as Siglec-F(+) constitute about 5% of CD45 cells. Although this identifies mature eosinophils, it may not be sufficient to represent all resident phenotypes. Using sequencing and FACS sorting approaches, we found that the Siglec-F(+) population represented only a minority of the resident eosinophil pool. Rather, we identified a lineage-positive population of IL-5Ra(+)Ly6G(+) but Siglec-F(−) cells distinct from lineage-negative EoPs (eosinophil progenitors). At 40% of all CD45(+) hematopoietic cells in adult mice, this bone marrow resident population dwarfs the Siglec-F(+) eosinophil pool. When sorted from bone marrow homogenates and treated with IL-5, these resident cells acquired all characteristics of mature eosinophils, including expression of Siglec-F and granular proteins (EPX, MPO). These tissue residents may play critical homeostatic roles in normal murine bone marrow and should be accounted for in studies of eosinophil biology.
IL7R regulates fetal tissue resident macrophage development by facilitating cell survival
Authors: Leung Gabriel , Valencia Clint , Beaudin Anna
Published: January 2023

Tissue-resident macrophages (TRMs) play critical roles in tissue homeostasis and disease. Many populations of TRMs derive from fetal progenitors and independently self-maintain across the lifespan through in situ proliferation. Previously, we have identified the interleukin-7 receptor (IL7R) as a novel regulator of TRM development. We have shown that antibody blockade of the IL7R during gestation impaired liver, lung, and epidermal TRM cellularity at birth. Here we show that in vivo fetal rIL-7 stimulation of the IL7R increased neonatal liver and lung TRM. In order to determine how IL7R signaling regulates fetal macrophage development, apoptosis was measured after late gestation IL7R blockade. Apoptosis in the macrophages and monocytes of the liver and lung was dramatically increased after IL7R blockade, suggesting that increases in neonatal cellularity after rIL-7 treatment may be due to increased survival signaling in these tissues. Our previous analysis also revealed dynamic regulation of IL7R mRNA surface protein expression as fetal monocytes differentiate into CD64+ macrophages during fetal development. We performed intracellular staining for IL7Ra protein to determine that monocytes contain intracellular IL7R protein that is not expressed on the surface until differentiation, suggesting that regulation of IL7R expression occurs at the level of surface expression. These data reveal dynamic regulation of IL7Ra expression in TRMs and TRM precursors during late gestation and provide evidence that IL-7/IL7R signaling regulates fetal TRM development by facilitating cell survival. Ongoing work addresses downstream signaling and other developmental processes regulated by IL7R signaling during fetal TRM development.
Cannabidiol alters macrophage differentiation resulting in accumulation of immunosuppressive myeloid cells and dendritic cells
Authors: Miranda Kathryn , Culpepper Courtney , Wilson Kiesha , Nagarkatti Prakash S. , Nagarkatti Mitzi
Published: January 2023

Cannabidiol (CBD) is a non-psychoactive phytocannabinoid that has anti-inflammatory properties; however the precise immunological mechanisms involved are not clear. Research from our laboratory has established that CBD promotes expansion of immunosuppressive myeloid-derived suppressor cells (MDSC), which are immature leukocytes thought to have stalled their differentiation towards mature myeloid lineages such as macrophages and dendritic cells (DC). In the current study, we aimed to better understand the mechanisms of CBD-altered hematopoiesis. We performed flow cytometry and single cell RNA sequencing (sc-RNAseq) on peritoneal cells from C57BL/6 mice treated with Vehicle (Veh) or CBD (20mg/kg, i.p.). Consistent with our previous studies, CBD promoted accumulation of MDSCs after 24h, which dissipated after 48h. Strikingly, MDSC accumulation was preceded by ablation of macrophages, which was maintained beyond 48h. In addition, accumulation of DCs started at 12h and accelerated through 48h. Similarly, sc-RNAseq of peritoneal cells harvested at 24h showed two distinct populations of cells expressing Adgre1, which encodes F4/80. These two clusters were distinct between Veh- and CBD-treated mice due to their drastically altered transcriptomes. The Veh-distinct cluster was comprised of macrophages expressing complement genes, while the CBD-distinct cluster was comprised of MDSCs expressing Arg1. Furthermore, the DC cluster expressing Ccr2 was enlarged in CBD-treated mice. Together, our results demonstrate that CBD drastically alters hematopoiesis, depleting macrophages and promoting MDSCs and DCs. These findings suggest CBD treatment may be beneficial in various macrophage-driven inflammatory disorders.
Inflammasome driven cell death in response to DNA damage is required for normal brain development and function
Authors: Lammert Catherine Rose , Lukens John R
Published: January 2023

Neurodevelopment is characterized by rapid rates of neural cell proliferation and differentiation followed by a period of massive cell death where over half of all recently generated brain cells are removed. Large amounts of DNA damage, cellular debris, and byproducts of cellular stress are generated during these neurodevelopmental events, all of which can potentially activate immune signaling. How the immune response to this damage influences brain maturation and function currently remains poorly understood. Here we show that the AIM2 inflammasome contributes to proper brain development and that disruptions in this immune sensor of genotoxic stress lead to behavioral abnormalities. We find that neurodevelopment is characterized by pronounced AIM2 inflammasome activation and that defects in this DNA damage surveillance sensor result in anxiety-related behaviors. We show that the AIM2 inflammasome contributes to CNS homeostasis specifically through its regulation of the cell death executioner Gasdermin-D, and not via its involvement in IL-1/IL-18 production. Consistent with a role for this dsDNA sensor in the purging of genetically compromised central nervous system cells, we find that defective AIM2 inflammasome signaling results in decreased neural cell death both in response to DNA damage-inducing agents and during neurodevelopment. Moreover, we report that disruptions in DNA damage surveillance by the AIM2 inflammasome lead to excessive DNA damage accumulation in neurons as well as increased numbers of neurons that incorporate into the adult brain. Our findings identify the inflammasome as a critical player in establishing a properly formed CNS through its role in the removal of genetically compromised cells.
Role of CD137L in lumbar spinal cord microglial responses following sciatic nerve crush
Authors: OHARA KINUYO , Nicholas David , Cao Ling
Published: January 2023

Neuropathic pain is one of the most devastating kinds of chronic pain, which is still largely treated sub-optimally in part due to incomplete understanding. Using murine sciatic nerve crush (SNC) model, we have shown that spinal cord CD137 ligand (CD137L; AKA 4-1BBL) contributes to the development of SNC-induced pain-like behaviors and functional recovery following SNC. Here, we further investigated the potential involvement of lumbar spinal cord microglial responses in CD137L-mediated behavioral changes following SNC. Mononuclear cells were isolated from lumbar spinal cords of both wild type (WT) B6 and B6_CD137L knockout (KO) mice via Percoll gradient followed by flow cytometric analysis at days 0 (naïve), 3, 7, 14, 28 and 56 following SNC or sham surgery. Microglia were identified as CD45lowCD11b+ populations. In WT mice, the total number of microglia peaked at day 14 and then gradually reduced following SNC. The increase of microglial numbers was minimal in all other groups compared to the WT SNC mice. Similar pattern of changes were also observed with the numbers of CD86+ microglia (pro-inflammatory phenotype) and CD206+ microglia (anti-inflammatory phenotype), however the numbers of CD206+ microglia were much less. Our results support the notion that reduced lumbar spinal cord microglial responses contributes to the reduction of pain-like behaviors and improved functional recovery in CD137L deficient mice. CD137L-mediated microglial responses will be further investigated to elucidate the role of spinal cord CD137L in the development of neuropathic pain. (Supported by NIH/NINDS R01NS098426 (Cao))
α-Synuclein-specific T cell reactivity is associated with preclinical and early Parkinson’s disease
Authors: Arlehamn Cecilia S Lindestam , Dhanwani Rekha , Pham John , Kuan Rebecca , Frazier April , Dutra Juliana Rezende , Phillips Elizabeth J , Mallal Simon Alexander , Roederer Mario , Marder Karen S , Amara Amy W , Standaert David G , Goldman Jennifer G , Litvan Irene , Peters Bjoern , Sulzer David , Sette Alessandro
Published: January 2023

A clinical diagnosis of motor Parkinson’s disease (PD) is preceded by a prolonged premotor phase with accumulating neuronal damage. α-Synuclein (α-syn)-specific T cell reactivity is preferentially associated with PD patients. Here we examined the temporal relation between α-syn T cell reactivity and PD. Analysis of PBMCs from a longitudinal case study revealed that elevated α-syn-specific T cell responses were detected prior to the diagnosis of motor PD, and declined after. These results suggest that α-syn-specific proinflammatory T cell reactivity could serve as an early predictor of PD. The relationship between T cell reactivity and early PD was further examined in two independent cohorts, which showed that α-syn-specific T cell responses were highest shortly after diagnosis of motor PD and then decreased. Additional analysis of clinical variables revealed significant association of α-syn-specific T cell responses with age and lower levodopa equivalent dose, and no correlation with motor and cognitive deficits. These results confirm the presence of α-syn-reactive T cells in PD and show that they are most abundant immediately after diagnosis of motor PD. Our single case study suggests that these cells may be present years before the diagnosis of motor PD, suggesting new avenues of investigation into PD pathogenesis, potential early diagnosis, and treatment options.
Conventional DC specific CSF1R depletion represents the reduction of the alveolar cDC2 and attenuation of allergic lung inflammation
Authors: Moon Hyung-Geun , Kim Seung-Jae , Park Gye Young
Published: January 2023

Dendritic cells play a key role in sensing aeroallergen and initiating Th2 immune reaction in asthma by bridging between innate and adaptive immune responses. Among the DC subtypes, conventional DC2 (cDC2) has been known to be a major player in developing allergic asthma. We recently reported that airway epithelial cell-derived CSF1 significantly promotes allergic sensitization through activating alveolar CSF1R+ cDC2 resulting in enhanced allergen-specific IgE production and subsequent Th2 immune response. This finding highlights the fundamental role of cDC2 in allergen sensitization. However, studying cDC2 has been hindered by the lack of available cDC2 specific-Cre mouse. Since CSF1R promoter activity is relatively enhanced in cDC2, we generated Zbtb46-cre;Csf1rfl/fl mice (hereafter CSF1RΔcDC) by crossbreeding cDC specific Zbtb46-cre and CSF1R flox mice. Of interest, we found that CSF1RΔcDC has a significant reduction in cDC2 population in alveolar space compared to Zbtb46-cre mice, whereas cDC1 and cDC2 in the lung and cDC1 in alveolar space were not affected in the steady state. When the mice were subjected to the allergen challenge, the number of cDC2 migrating to regional lymph node with carrying the allergen was markedly diminished in CSF1RΔcDC mice. Subsequently, the production of total and allergen-reactive IgE was blocked and allergic lung inflammation including eosinophil recruitment was attenuated in CSF1RΔcDC mice, compared to control littermate mice. These data suggest that CSF1RΔcDC mice are suitable to investigate the specific role of cDC2 in alveolar space and CSF1R+ cDC2 in alveolar space plays a key role in allergen sensitization and asthma development.
The role of IL-9 secreting CD4+ T helper cells in promoting mast cell expansion in pulmonary models of inflammation
Authors: Pajulas Abigail , Kaplan Mark H
Published: January 2023

Mast cells are tissue resident cells with granules that contain histamine and other proinflammatory mediators. Mast cells are important effectors in immediate type I hypersensitivity reactions. In severe allergic reactions, immunoglobulin E (IgE)-mediated mast cell degranulation triggers localized and systemic anaphylaxis. In allergic asthma, mast cells and T cells interact in the generation of long-lasting inflammation. The molecular mechanisms responsible for mast cell expansion has not been fully elucidated. Because T cells are an important source of the mast cell growth factor IL-9, we are interested in understanding the role of IL-9, and IL-9-producing T cells in mast cell expansion and recruitment. We have identified a role of T helper 9 (Th9) cells in influencing mast cell number and function in pulmonary models of inflammation. Using adoptive transfer models, we have defined a contribution of Th9 cells in immediate hypersensitivity. We have demonstrated that IL-9 expands mast cell progenitor numbers in bone marrow and lung, suggesting that there are systemic effects of local IL-9 production. We hypothesize that increased mature mast cells in the pulmonary system are from recruitment and expansion of progenitor cells rather than expansion of mature cells in situ. Ongoing studies will directly examine the relative contribution and function of the two populations during models of allergic airway inflammation.
Therapeutic approach to steroid-resistant asthma
Authors: Mori Akio , Kouyama Satoshi , Yamaguchi Miyako , Kumitani Chiemi , Ohtomo-Abe Akemi , Fujita Norihiko , Iwata Maki , Nagayama Kisako , Ryu Kai , Nakamura Yuto , Hamada Yuto , Hayashi Hiroaki , Watai Kentaro , Kamide Yosuke , Sekiya Kiyoshi , Fukutomi Yuma , Taniguchi Masami , Ohtomo Takayuki , Kaminuma Osamu
Published: January 2023

Background

To control therapy-resistant eosinophilia, synergistic effects of CTLA4-Ig and glucocorticoid was investigated on T cell-induced asthma model.

Methods

Ovalbumin (OVA) specific murine helper T cell (Th) clones were established from DO11.10 transgenic mice. To analyze steroid responsiveness in vitro, Th clones were cultured with antigen presenting cells and OVA in the presence of various concentration of dexamethasone (DEX). Proliferative responses were measured by incorporation of either 3H-thymidine or BrdU. For in vivo analysis, unprimed Balb/c mice were transferred with Th clones, challenged with OVA, and administered with DEX subcutaneously. CTLA4-Ig was administered intravenously. BALF was obtained 48 hours after the challenge, and the number of infiltrating cells was differentially counted.

Results

Steroid-sensitive (SS) and -resistant (SR) clones were selected based on the effect of DEX on the proliferative responses of antigen-stimulated Th clones. Airway infiltration of eosinophils of mice transferred with SS clones were effectively inhibited by the administration of DEX. In contrast, those of mice transferred with SR clones were not significantly inhibited by DEX. Addition of CTLA4-Ig into the culture significantly suppressed the proliferation of DEX-treated SR clones in vitro. Administration of CTLA4-Ig significantly suppressed eosinophil infiltration of SR asthma model transferred with SR clones in vivo. CTLA4-Ig and DEX synergistically suppressed in vitro proliferation of SS clones and in vivo BALF eosinophilia of mice transferred with SS clones.

Conclusion

Blocking costimulatory signal mediated through CD28 is a promising target to treat therapy-resistant eosinophilia.
Respiratory Syncytial Virus exacerbates acute lung damage during ovalbumin-induced asthmatic inflammation
Authors: Muralidharan Abenaya , Bauer Christopher D , Sun Keer
Published: January 2023

With more than 235 million people affected, asthma is the most common chronic disease in the world. In addition, respiratory viral infections, particularly respiratory syncytial virus (RSV) infection, has been associated with asthma development and significant exacerbation of asthmatic symptoms. While there are several theories exploring the causal role of RSV infection in asthma development, the pathogenic mechanism underlying the effect of RSV on asthma exacerbation remains to be understood. In this study, we investigated the effects of RSV infection on airway immune responses and lung damage in ovalbumin (OVA)-induced asthmatic mice. We found that RSV infection exacerbates acute lung damage in asthmatic mice, as indicated by their significantly increased protein and LDH levels in the airway. Although asthma is considered an allergic disease with increased Th2 responses, the levels of IL-4 and IL-5, as well as eosinophil recruitment, were unaffected in OVA-induced asthmatic mice with or without RSV super-infection. Moreover, pro-inflammatory responses, such as TNF-alpha, KC and MIP-2 production, and neutrophil infiltration, were also comparable between RSV-infected asthmatic mice and controls that only received RSV infection. Interestingly, the number of resident alveolar macrophages significantly decreased in RSV-infected asthmatic mice, as compared with RSV-infected non-asthmatic and uninfected asthmatic controls. Considering the crucial role of alveolar macrophages in resolution of airway inflammation, our future studies will determine whether RSV impairs alveolar macrophage self-maintenance, thereby prolonging airway inflammation and exacerbating lung damage during asthma.
Airway Microbiota-Host Interactions Regulate Secreted Leukocyte Proteinase Inhibitor Levels and Influence Allergic Airway Inflammation
Authors: Jaeger Natalia , McDonough Ryan , Hernandez-Leyva Ariel , Rosen Anne , Wilson Naomi G. , Russler-Germain Emilie V. , Chai Jiani N. , Bacharier Leonard B. , Hsieh Chyi-Song , Kau Andrew L.
Published: January 2023

Mucosal immune responses fine-tuned by naturally evolved interactions with native microbes influence airway inflammation but are not routinely integrated into experimental models. Here, we leverage a recently discovered murine-adapted airway microbe, Bordetella pseudohinzii (Bph), to investigate how chronic colonization impacts mucosal immunity and the development of allergic airway inflammation (AAI). Despite the persistent colonization, we did not observe significant weight loss in Bph colonized mice. Microscopic examination of the lungs demonstrated the formation of lymphoid aggregates, consistent with inducible Bronchus Associated Lymphoid Tissue (iBALT). Airway colonization of mice with Bph elicits an antigen-specific Th17 immune response that aids in controlling bacterial abundance. Remarkably, when animals undergo allergic airway challenge, mice previously colonized with Bph demonstrate protection from AAI. This phenotype corresponded to a reduction in type 2 cytokines, eosinophilic infiltration, goblet cell metaplasia, and airway hyper responsiveness. Colonization with Bph does not appear to impact the degree of allergic sensitization nor the presentation of allergen in the lung. Further, colonization and the Th17 response was associated with increased expression of Secreted Leukocyte Proteinase Inhibitor (SLPI), an antimicrobial peptide with anti-inflammatory properties. We confirmed these findings in humans by showing that higher levels of SLPI correlate both to improved asthma control and the presence of the asthma-associated bacterium, Haemophilus influenzae. We propose that SLPI could be used as a biomarker of beneficial host-commensal relationships in the airway.
A single cell approach to identify the role of IL-9 secreting tissue-resident CD4+ T cells in allergic airway recall responses
Authors: Kharwadkar Rakshin P , Ulrich Benjamin J , Fu Yongyao , Koh Byunghee , Nelson Andrew Scott , Kaplan Mark H
Published: January 2023

Allergic asthma is a chronic intermittent inflammatory lung disease where patients experience relatively few symptoms between exacerbations precipitated by allergen-challenge of allergic memory responses. Allergen-specific CD4+ T cells are believed to be a major mediator of allergic memory responses through the production of type 2 cytokines including IL-4, IL-5, IL-13 and IL-9. Recently, tissue-resident memory (Trm) cells have been identified in peripheral tissues that mediate mucosal barrier immunity. However, the precise role of Trm CD4+ T cells in allergic exacerbations is not clearly defined. In studies to examine the role of IL-9 in allergen-specific Trm responses to Aspergillus fumigatus we observed that all IL-9-secreting T cells had a resident memory phenotype. Inhibition of circulating T cells by administering FTY720 in the last month of rest, diminished the total CD4 T cells population in the lung, while the Th9 cells remained stable. Blockade of IL-9 prior to recall challenge phase reduced overall allergic lung inflammation as assessed both by flow cytometry and single cell RNA-seq. Within eight hours after challenge, IL-9 is one of the top genes identified by scRNA-seq that distinguishes ST2+ and ST2− CD4+ T cell populations in the lung. Using IL-9-reporter mice we also compared gene expression in isolated IL-9+ T cells and other type 2 T cells. These findings demonstrate that IL-9-producing Trms play an important role in mediating allergic memory responses. IL-9 could be a promising therapeutic target specifically in patients showing symptoms of intermittent allergic response.
IL-6 deficiency exacerbates allergic asthma and promotes Streptococcus pneumoniae pathogenesis
Authors: Schmit Taylor , Ambigapathy Ganesh , Ghosh Sumit , Mathur Ramkumar , Wu Min , Combs Colin , Khan Nadeem
Published: January 2023

Allergic asthma consists of diverse immune phenotypes exhibiting differential lung pathology, remodeling of the respiratory tract and mucociliary bronchial clearance. These diverse inflammatory immune mechanisms may promote or resist microbial infections in allergic human and mouse models. IL-6 governs optimal T-cell and inflammatory response leading to the resolution of bacterial and viral infections in respiratory airways. Additionally, IL-6 plays an important role in maintaining the integrity of airway barrier response during microbial infections. We hypothesized that lung IL-6 responses alleviate Streptococcus pneumoniae (Spn) pathogenesis during fungal allergic asthma caused by Aspergillus fumigatus.

Using a murine Aspergillus fumigatus asthma model, we show that IL-6 deficiency exacerbates lung inflammation and promotes Spn disease pathogenesis in allergic host. IL-6 deficiency was associated with increased lung damage, which correlated with higher levels of lung eosinophilia, inflammatory cytokines and chemokines. Additionally, IL-6 deficiency correlated with increased TGF-β1 cytokine levels, increased epithelial-mesenchymal transition and dysregulated epithelial barrier proteins in asthmatic mice.

We conclude that IL-6 offers a significant protective role in the control of Spn pathogenesis during allergic asthma by regulation of immune cell recruitment and maintenance of lung epithelial barrier integrity.
The pseudo-allergy receptor, MrgprB2/X2 is controlled by neurokinin A and the neurokinin 2 receptor in human and mouse skin
Authors: Li Yuting , Zhang Jiying , Falo Louis D , Larregina Adriana T , Sumpter Tina L
Published: January 2023

Within the skin, mast cells (MCs) and sensory neurons form a cohesive unit that plays an important role in initiating immune responses. Neuropeptides such as neurokinin A and substance P direct MC function by initiating signaling through neurokinin (NK) receptors, and Mas related G-protein receptors (Mrgpr)s. Neurokinin A, in particular, is thought to signal through the NK2R. Recent studies highlight the importance of the MrgprB2 (mouse) and MrgprX2 (human) in the MC response to pseudo-allergens, secretagogues and substance P. To date, a relationship between neurokinin receptors and the MrgprB2/X2 has not been investigated. In this study, we hypothesize that MrgprB2/X2 expression is controlled by the NK2R and its high affinity ligand, neurokinin A. In mice, we show that administration of neurokinin A diminishes MrgprB2 expression. Surprisingly, antagonism of the NK2R also downregulates MrgprB2 expression and in NK2R-deficient mice, MrgprB2 expression is markedly diminished. In contrast, co-administration of neurokinin A and a NK2R antagonist markedly increases MrgprB2 expression. In human skin explants, NK2R antagonism has minimal effect on MrgprX2 expression, but co-administration of neurokinin A and a NK2R antagonist upregulates MrgprX2 expression, as seen in murine skin. These data demonstrate that NK2R-signaling influences MrgprB2/X2 expression and, in absence of the NK2R, neurokinin A interacts with an unknown receptor to increase MrpgrB2/X2 expression. Collectively, these data uncover a novel role for NK2R signaling in the regulation of MrgprB2/X2. These important findings have implications for patients with dysregulated mast cell function initiated through the MrgprX2.
Decoding the role of CBLB for innate immune responses regulating systemic dissemination during Non-Tuberculous Mycobacteria infection
Authors: Nanjappa Som G. , Mudalagiriyappa Srinivasu , Sharma Jaishree , Vieson Miranda
Published: January 2023

Non-Tuberculous Mycobacteria (NTM) are ubiquitous in nature, present in soil and water, and cause primary and disseminated infections in the immune-compromised individuals. NTM infections are surging in recent years at 8% annually, due to an increase in immune-suppressed population, medical interventions, and underlying lung diseases. The innate immune responses, frontiers for controlling infections and dissemination, are poorly defined during NTM infections. Here, we define the role of CBLB, an E3-ubiquitin ligase, for innate immune responses and disease progression in a mouse model of NTM infection in the absence of antigen-specific T-cell responses. We found that NTM was readily disseminated into the tissues in the Cblb KO mice in a time- and infection route-dependent manner, compared with the WT group. We uncovered defects in many innate immune cells in Cblb KO mice: poor responses with NK cells, inflammatory monocytes, neutrophils, and activation of conventional dendritic cell responses. Strikingly, Cblb-deficient macrophages were able to produce higher ROS in both in vivo & in vitro compared with the WT group. Histopathological studies buttressed the role of Cblb for disseminated NTM infection. Collectively, Cblb plays a dichotomous role in innate immunity, and may help prevent the dissemination of NTM in the infected host. Our studies illuminate the translational implications for controlling NTM infections by modulating the innate immune immunity.
The role of high-mobility group box 1 protein in respiratory syncytial virus-induced autophagy
Authors: Kempaiah Kempaiah Rayavara , Hosakote Yashoda Madaiah
Published: January 2023

Respiratory syncytial virus (RSV) is the most common cause of acute lower respiratory tract infections in infants and young children worldwide, for which no efficacious treatments or vaccines are currently available. Although the pathogenic mechanisms of RSV-induced acute airway disease are still unclear, lung immune and inflammatory responses are key. High-mobility group box 1 (HMGB1) is a redox-sensitive multifunctional protein that serves as both a DNA regulatory protein and an extracellular signaling molecule that mediates the activation of inflammatory responses to infection/injury. HMGB1 has been implicated in mitochondrial function and quality control. The objective of this study is to investigate the role of HMGB1 in autophagosome formation and mitochondrial quality control in RSV infection. Studies were conducted in lung epithelial cells including A549 and normal small alveolar epithelial cells. Our studies show that RSV induces HMGB1 secretion in lung epithelial cells via NFκB and TLR4 pathways, and activates innate immune cells to promote inflammation. We show that RSV-induces LC3B and promote autophagy whereas recombinant HMGB1 (rHMGB1) treatment inhibits RSV-induced autophagy. We also show that RSV infection inhibits the inflammasome pathway in lung epithelial cells but rHMGB1 treatment activates inflammasomes. Our results indicate that RSV-induced epithelial HMGB1 secretion triggers inflammasome activation in innate immune cells to promote inflammation. Understanding HMGB1 and associated cellular signaling in RSV-induced lung inflammation may pave the way for the development of novel therapeutic interventions to treat not only RSV, but also other respiratory infections that affect humans of all ages.
Interferon-mediated Reprogramming of Intracellular Cholesterol Metabolism in Response to Secreted Bacterial Toxins
Authors: Lee Minsub S , Zhou Quan , Chi Xun , Hsieh Wei Yuan , Fang An-Chieh , Scumpia Philip O. , Williams Kevin , Damoiseaux Robert , Bensinger Steven
Published: January 2023

Macrophages undergo profound metabolic reprogramming in response to activation signals. However, the purpose and the mechanism of this reprogramming remains poorly understood. Cholesterol is an essential lipid molecule that modulates biochemical and biophysical properties of cellular membranes. Microbes have evolved strategies which specifically exploit the presence of membrane cholesterol to facilitate pathogenesis. Cholesterol-dependent cytolysin (CDC) are pore-forming toxins produced by gram-positive microorganisms that results in loss of membrane integrity, cellular dysfunction, and cell death. Herein, we delineate a unappreciated immune cell strategy that relies on rapid reprogramming of cholesterol homeostasis to avoid the deleterious effects CDCs. We show that interferon signals convey resistance to CDCs in myeloid cells by altering the availability of cholesterol in the plasma membrane (PM). Mechanistic studies reveal that resistance to CDCs occurs as a result of the movement of cholesterol from the PM to the endoplasmic reticulum (ER), and ultimately into an intracellular esterified pool. We also find that IFN-mediated inhibition of cholesterol synthesis is required to ensure maximal resistance to CDC challenge. Finally, we demonstrate that the IFN-mediated inhibition of cholesterol synthesis is dependent on production of the oxysterol, 25-hydroxycholesterol, and that the loss of CH25H renders mice more susceptible to CDC-mediated tissue damage. Together, these data provide evidence for remodeling of membrane lipid composition as a host defense strategy to bacterial toxins, and suggests that metabolic reprogramming of lipid architecture has significance beyond that of supporting immune function.
Internal N6-methyladenosine modification controls the innate immune response to viral RNA
Authors: xue miaoge , Lu Mijia , Li Anzhong , Zhang Zijie , He Chuan , Li Jianrong
Published: January 2023

Internal N6-methyladenosine (m6A) modification is one of the most prevalent and abundant modifications of RNA. The m6A methylation is catalyzed by host methyltransferases (METTL3 and METTL14) and can be reversibly removed by host demethylases (FTO and ALKBH5). Recent studies have revealed that RNA of many DNA and RNA viruses contain m6A methylation and that viral m6A can play a pro-viral or anti-viral role. However, the biological functions of viral RNA m6A methylation remain poorly understood. In this study, we found that genome and antigenome of human respiratory syncytial virus (RSV), a non-segmented negative-sense RNA virus, are m6A methylated and that viral m6A methylation positively regulates RSV replication and gene expression. Interestingly, virion RNAs purified from RSV grown in METTL3-knockout cells induced significantly higher type I interferon responses than virion RNAs purified from RSV grown in wild type cells, demonstrating that viral RNA m6A methylation inhibits innate immunity. Similarly, inactivation of m6A methylation in RSV genome and/or antigenome by site-directed mutagenesis induced significantly higher innate immune responses than wild type RSV RNA. Mechanistically, these m6A-unmodified or m6A-deficient RSV RNAs trigger a higher expression of pattern recognition receptors (such as RIG-I and MDA5), enhance their binding affinity to the innate immune RNA sensors, and enhance the phosphorylation of downstream transcription factor IRF3, leading to an enhanced activation of type I interferon signaling pathway. Taken together, our study identified a novel biological function of viral m6A methylation, which is to serves as a molecular marker for host innate immunity to discriminate self from nonself RNAs.
NLRC3 Localizes to the Endoplasmic Reticulum via Interactions with a Novel ER-Resident Protein
Authors: Davis Beckley K , Baker Taylor , Rechnitzer Alma , Hamby Matthew , Troiani Zachary , Stenske Sarah , Davis Samuel , Bauer Anna
Published: January 2023

Sensing of cytosolic nucleotides is a critical initial step in the elaboration of type I interferon. One of several upstream receptor cGAS (cyclic-GMP-AMP synthase) binds to cytosolic DNA and generates di-cyclic nucleotides that act as secondary messengers. These secondary messengers bind directly to Stimulator of Interferon Genes (STING). STING recruits TANK binding kinase 1 (TBK1) which acts as a critical node that allows for efficient activation of interferon regulatory factors (IRFs) to drive the anti-viral transcriptome. NLRC3 is a recently characterized nucleotide-binding domain, leucine rich repeat containing protein (NLR) that negatively regulates the type I interferon pathway by inhibiting subcellular redistribution and effective signaling of STING, thus blunting the transcription of type I interferons. NLRC3 is predominantly expressed in lymphoid and myeloid cells. IQGAP1 was identified as a putative interacting partner of NLRC3 through yeast two hybrid screening. Here we show that a novel ER-resident protein associates with NLRC3 in human cells. This interaction occurs at the ER. This data provides a mechanism by which NLRC3 localizes to the ER to affect STING signaling in response to cytosolic nucleotides.
Olfactory receptor 2 (Olfr2) and its human ortholog OR6A2 expressed in macrophages drive NLRP3 inflammasome activation and exacerbate atherosclerosis in mice
Authors: Orecchioni Marco , Kobiyama Kouji , Winkels Holger , Ghosheh Yanal , McArdle Sara , Mikulski Zbigniew , Fan Zhichao , Wen Lai , Jung Yunmin , Roy Payel , Myamoto Yukiko , Matsunami Hiroaki , Eckmann Lars , Latz Eicke , Wang Zeneng , Hazen Stanley L , Ley Klaus
Published: January 2023

Atherosclerosis is an inflammatory disease of the arterial wall driven by macrophages and other immune cells. Olfactory receptors (Olfrs) are G-protein coupled receptors expressed in olfactory epithelium and are responsible for the sense of smell. We found that macrophages in the wall of atherosclerotic mouse aortas express some olfactory receptors including Olfr2, a specific receptor for an 8 carbon fatty-aldehyde called octanal. Octanal is detectable in food, mouse and human blood plasma, elevated by western diet, and partially derived from gut microbiota. Ligation of Olfr2 or its human orthologue OR6A2, expressed in human atherosclerotic plaque and in human monocyte-derived macrophages, is strongly pro-inflammatory, activates the NLRP3 inflammasome and, in synergy with LPS, induces secretion of IL-1β. Knockdown of Olfr2/OR6A2 in macrophages significantly reduced inflammatory cytokine secretion. To test the role of Olfr2 in atherosclerosis, we treated Apoe−/− mice with octanal or the Olfr2 antagonist citral. Octanal significantly exacerbated while citral significantly inhibited atherosclerosis. We generated Olfr2−/− mice by Crispr-Cas9. Ldlr−/− mice reconstituted with Olfr2−/− bone marrow developed ~50% smaller lesions on high cholesterol diet than littermate controls reconstituted with Olfr2+/+ bone marrow. Our findings suggest that small molecule inhibitors of Olfrs like OR6A2 are promising targets for drug development to prevent and treat atherosclerosis-based cardiovascular diseases.