Activity-dependent FosB gene expression negatively regulates mast cell functions.

Mast cells are innate immune cells that play a crucial role in numerous physiological processes across tissues by releasing pre-stored and newly synthesized mediators in response to stimuli, an activity largely driven by changes in gene expression. Given their widespread influence, dysfunction in mast cells can contribute to a variety of pathologies including allergies, long COVID, and autoimmune and neuroinflammatory disorders. Despite this, the specific transcriptional mechanisms that control mast cell mediator release remain poorly understood, significantly hindering the development of effective therapeutic strategies. We found that the two proteins encoded by the transcription factor FosB, FOSB and the highly stable variant ΔFOSB, are robustly expressed upon stimulation in both murine and human mast cell progenitors. Motivated by these findings, we generated a novel mouse model with targeted ablation of FosB gene expression specifically in mast cells (MC FosB- ) by crossing a mast cell-specific Cre reporter line (Mcpt5-Cre) with a Cre-dependent floxed FosB mouse lines. We found that mast cell progenitors derived from MC FosB- mice, compared to wild types (WT), exhibit baseline increased histamine content and vesicle numbers. Additionally, they show enhanced calcium mobilization, degranulation, and histamine release following allergy-related IgE-mediated stimulation, along with heightened IL-6 release in response to infection-like LPS stimulation. In vivo experiments with IgE- mediated and LPS challenges revealed that MC FosB- mice experience greater drops in body temperature, heightened activation of tissue-resident mast cells, and increased release of pro-inflammatory mediators compared to their WT counterparts. These findings suggest that FosB products play a crucial regulatory role in moderating stimulus-induced mast cell activation in response to both IgE and LPS stimuli. Lastly, by integrating CUT&RUN and RNAseq data, we identified several genes targeted by ΔFOSB that could mediate these observed effects, including Mir155hg, CLCF1, DUSP4, and Trib1. Together, this study provides the first evidence that FOSB/ΔFOSB modulate mast cell functions and provides a new possible target for therapeutic interventions aimed at ameliorating mast cell-related diseases.

Early life adversity drives sex-specific anhedonia and meningeal immune gene expression through mast cell activation.

Exposure to early life adversity (ELA) in the form of physical and/or psychological abuse or neglect increases the risk of developing psychiatric and inflammatory disorders later in life. It has been hypothesized that exposure to ELA results in persistent, low grade inflammation that leads to increased disease susceptibility by amplifying the crosstalk between stress-processing brain networks and the immune system, but the mechanisms remain largely unexplored. The meninges, a layer of three overlapping membranes that surround the central nervous system (CNS)- dura mater, arachnoid, and piamater - possess unique features that allow them to play a key role in coordinating immune trafficking between the brain and the peripheral immune system. These include a network of lymphatic vessels that carry cerebrospinal fluid from the brain to the deep cervical lymph nodes, fenestrated blood vessels that allow the passage of molecules from blood to the CNS, and a rich population of resident mast cells, master regulators of the immune system. Using a mouse model of ELA consisting of neonatal maternal separation plus early weaning (NMSEW), we sought to explore the effects of ELA on sucrose preference behavior, dura mater expression of inflammatory markers and mast cell histology in adult male and female C57Bl/6 mice. We found that NMSEW alone does not affect sucrose preference behavior in males or females, but it increases the dura mater expression of the genes coding for mast cell protease CMA1 (cma1) and the inflammatory cytokine TNF alpha (tnf alpha) in females. When NMSEW is combined with an adult mild stress (that does not affect behavior or gene expression in NH animals) females show reduced sucrose preference and even greater increases in meningeal cma1 levels. Interestingly, systemic administration of the mast cell stabilizer Ketotifen before exposure to adult stress prevents both, reduction in sucrose preference an increases in cma1 expression in NMSEW females, but facilitates stress-induced sucrose anhedonia in NMSEW males and NH females. Finally, histological analyses showed that, compared to males, females have increased baseline activation levels of mast cells located in the transverse sinus of the dura mater, where the meningeal lymphatics run along, and that, in males and females exposed to adult stress, NMSEW increases the number of mast cells in the interparietal region of the dura mater and the levels of mast cell activation in the sagittal sinus regions of the dura mater. Together, our results indicate that ELA induces long-term meningeal immune gene changes and heightened sensitivity to adult stress-induced behavioral and meningeal immune responses and that these effects could mediated via mast cells.

Histamine-dependent interactions between mast cells, glia, and neurons are altered following early-life adversity in mice and humans.

Early-life adversity contributes to the development of functional bowel disorders later in life through unresolved mechanisms. Here, we tested the hypothesis that early-life adversity alters anatomical and functional interactions between mast cells and enteric glia. The effects of early-life stress were studied using the neonatal maternal separation (NMS) stress mouse model. Anatomical relationships between mast cells and enteric glia were assessed using immunohistochemistry and mast cell reporter mice (Mcpt5Cre;GCaMP5g-tdT). Immunohistochemistry was used to assess the expression of histamine, histamine 1 (H1) receptors, and glial fibrillary acidic protein. Functional responses of glia to mast cell mediators were assessed in calcium imaging experiments using Sox10CreERT2;GCaMP5g-tdT mice and cultured human enteric glial cells. NMS increases mast cell numbers at the level of the myenteric plexus and their proximity to myenteric ganglia. Myenteric glia respond to mediators released by activated mast cells that are blocked by H1 receptor antagonists in mice and humans and by blocking neuronal activity with tetrodotoxin in mouse tissue. Histamine replicates the effects of mast cell supernatants on enteric glia, and NMS increases histamine production by mast cells. NMS reduces glial responses to mast cell mediators in mouse tissue, while potentiating responses in cultured human enteric glia. NMS increases myenteric glial fibrillary acidic protein expression and reduces glial process length but does not cause neurodegeneration. Histamine receptor expression is not altered by NMS and is localized to neurons in mice, but glia in humans. Early-life stress increases the potential for interactions between enteric glia and mast cells, and histamine is a potential mediator of mast cell-glial interactions through H1 receptors. We propose that glial-mast cell signaling is a mechanism that contributes to enteric neuroplasticity driven by early-life adversity.NEW & NOTEWORTHY Early-life adversity places an individual at risk for developing functional gastrointestinal disorders later in life through unknown mechanisms. Here, we show that interactions between mast cells and glia are disrupted by early-life stress in mice and that histamine is a potential mediator of mast cell-glial interactions.

Impacts of environmental complexity on respiratory and gut microbiome community structure and diversity in growing pigs.

The limited understanding of the interaction between rearing environment of the growing pig and the pig's microbial community impedes efforts to identify the optimal housing system to maximize animal health and production. Accordingly, we characterized the impact of housing complexity on shaping the respiratory and gut microbiota of growing pig. A total of 175 weaned pigs from 25 litters were randomly assigned within liter to either simple slatted-floor (S) or complex straw-based rearing ecosystem (C). Beside the floor swabs samples, fecal swabs and mucosal scraping samples from bronchus, ileum, and colon were collected approximately 164 days post-weaning at the time of slaughter. The S ecosystem seems to increase the α-diversity of respiratory and gut microbiota. Moreover, the C-raised pigs showed 35.4, 89.2, and 60.0% reduction in the Firmicutes/Bacteroidetes ratio than the S-raised pigs at bronchus, ileum, and colon, respectively. The unfavorable taxa Psychrobacter, Corynebacterium, Actinobacteria, and Neisseria were the signature taxa of C environment-associated microbial community. Therefore, the microbiota of S-raised pigs seems to show higher density of the most essential and beneficial taxa than the C-raised pigs. We preliminarily conclude that increasing the physical complexity of rearing environment seems to provide suboptimal conditions for establishing a healthy microbial community in the growing pigs.

Gastrointestinal microbiota and mucosal immune gene expression in neonatal pigs reared in a cross-fostering model.

Cross fostering is employed to equalize the number of piglet between litters ensuring colostrum intake for their survival and growth. However, little is known about the impact of cross fostering on the intestinal microbiota and mucosal immune gene expression of the neonatal pig. The objective of this study was to determine the influence of maternal microbial communities on the gastrointestinal (GI) microbiota and mucosal immune gene expression in young pigs reared in a cross-fostering model. Piglets were given high quality colostrum from birth dam or foster dam upon birth. Twenty-four piglets were randomly assigned at birth to 1 of 3 treatments according to colostrum source and postcolostral milk feeding during, as follow: treatment 1 (n = 8), received colostrum and post-colostral milk feeding from their own dam; treatment 2 (n = 8), received colostrum from foster dam and returned to their own dam for post-colostral milk feeding; and treatment 3 (n = 8), received colostrum and post-colostral milk feeding from foster dam. Genomic DNA was extracted, and the V1-V3 hypervariable region of the bacterial 16S rRNA gene was amplified and sequenced using the Illumina MiSeq platform. Quantitative real-time PCR analysis was also performed to quantify the expression of toll-like receptors (TLR) 2, TLR 4, TLR 10, tumor necrosis factor alpha (TNFα), interferon gamma (IFNγ), and interleukin (IL) 4 and IL 10. Data analysis revealed that microbial communities were varied according to the GI biogeographical location, with colon being the most diverse section. Bacterial communities in both maternal colostrum and vaginal samples were significantly associated with those present in the fecal samples of piglets. Cross-fostering did not affect bacterial communities present in the piglet GI tract. However, the mRNA expression of TLR and inflammatory cytokines changed (P < 0.05) with biogeographical location in the GI tract. Higher mRNA expression of TLR and inflammatory cytokines was observed in ileum and ileum associated lymph tissues. This study suggests an impact of colostrum and maternal microbial communities on the microbiota development and mucosal immune gene expression in the newly born piglet. This study revealed novel information about the distribution and expression patterns of TLR and inflammatory cytokines in the GI tract of the young pig. Future studies are needed to determine the role and clinical importance of the mucosal microbiota and mucosal gene expression in health, productivity, and susceptibility to the development of GI disease, in piglets.

Disparity in the nasopharyngeal microbiota between healthy cattle on feed, at entry processing and with respiratory disease.

Bovine respiratory disease (BRD) is one of the most serious causes of health and economic problems in the beef production industry, especially in recently weaned, intensely raised and newly transported feedlot cattle. While the importance of upper airway structure and function in the susceptibility of the lower respiratory tract to colonization with potential pathogens is well established, the role of the mucosal microbiota in respirtatory health is less well defined. The objective of this study was to characterize the nasopharyngeal microbiota of feedlot cattle at entry into a commercial feedlot, during initial management processing, and to compare the dynamics of change in these microbial communities between clinically healthy calves and those that develop BRD within the first month after entry. Deep nasopharyngeal swabs were collected from randomly selected healthy calves (n=66) during initial handling and processing at the feedlot, and again at the initial diagnosis of BRD (n=22). Clinically healthy pen matched controls calves (n=10) were sampled at the same time as the BRD affected animals. Genomic DNA was extracted from each sample, and the 16S rRNA gene V1-V3 hypervariable region was amplified and sequenced using the Illumina MiSeq platform. Across all the samples, the predominant bacterial phyla were Proteobacteria, Firmicutes and Actinobacteria. While the predominant genera were Moraxella, Mycoplasma and Acinetobacter. Linear discriminant analysis (LDA) effect size (LEfSe) revealed significant differences in bacterial taxa between healthy and BRD affected calves. Discriminant analysis revealed that the nasopharyngeal microbiota in feedlot calves at entry and in BRD affected calves were distinct from pen matched healthy calves. While the temporal dynamics of this shift were not examined in this study, it is possible that the observed changes in mucosal microbiota are linked to the increased susceptibility of calves to BRD during the first month after entry in to the feedlot. Additional studies are needed to examine the trajectory of change in nasopharyngeal microbial communities from entry to disease onset, and to explore the impact of other factors such as diet transition, commingling, vaccination and housing on the nasopharyngeal microbiota of growing cattle.