Mast Cell Chymase/Mcpt4 Suppresses the Host Immune Response to Plasmodium yoelii, Limits Malaria-Associated Disruption of Intestinal Barrier Integrity and Reduces Parasite Transmission to Anopheles stephensi

An increase in mast cells (MCs) and MCs mediators has been observed in malaria-associated bacteremia, however, the role of these granulocytes in malarial immunity is poorly understood. Herein, we studied the role of mouse MC protease (Mcpt) 4, an ortholog of human MC chymase, in malaria-induced bacteremia using Mcpt4 knockout (Mcpt4(-/-)) mice and Mcpt4(+/+) C57BL/6J controls, and the non-lethal mouse parasite Plasmodium yoelii yoelii 17XNL. Significantly lower parasitemia was observed in Mcpt4(-/-) mice compared with Mcpt4(+/+) controls by day 10 post infection (PI). Although bacterial 16S DNA levels in blood were not different between groups, increased intestinal permeability to FITC-dextran and altered ileal adherens junction E-cadherin were observed in Mcpt4(-/-) mice. Relative to infected Mcpt4(+/+) mice, ileal MC accumulation in Mcpt4(-/-) mice occurred two days earlier and IgE levels were higher by days 8-10 PI. Increased levels of circulating myeloperoxidase were observed at 6 and 10 days PI in Mcpt4(+/+) but not Mcpt4(-/-) mice, affirming a role for neutrophil activation that was not predictive of parasitemia or bacterial 16S copies in blood. In contrast, early increased plasma levels of TNF-alpha, IL-12p40 and IL-3 were observed in Mcpt4(-/-) mice, while levels of IL-2, IL-10 and MIP1 beta (CCL4) were increased over the same period in Mcpt4(+/+) mice, suggesting that the host response to infection was skewed toward a type-1 immune response in Mcpt4(-/-) mice and type-2 response in Mcpt4(+/+) mice. Spearman analysis revealed an early (day 4 PI) correlation of Mcpt4(-/-) parasitemia with TNF-alpha and IFN-gamma, inflammatory cytokines known for their roles in pathogen clearance, a pattern that was observed in Mcpt4(+/+) mice much later (day 10 PI). Transmission success of P. y. yoelii 17XNL to Anopheles stephensi was significantly higher from infected Mcpt4(-/-) mice compared with infected Mcpt4(+/+) mice, suggesting that Mcpt4 also impacts transmissibility of sexual stage parasites. Together, these results suggest that early MCs activation and release of Mcpt4 suppresses the host immune response to P. y. yoelii 17XNL, perhaps via degradation of TNF-alpha and promotion of a type-2 immune response that concordantly protects epithelial barrier integrity, while limiting the systemic response to bacteremia and parasite transmissibility

Nonlethal Plasmodium yoelii Infection Drives Complex Patterns of Th2-Type Host Immunity and Mast Cell-Dependent Bacteremia.

Malaria strongly predisposes to bacteremia, which is associated with sequestration of parasitized red blood cells and increased gastrointestinal permeability. The mechanisms underlying this disruption are poorly understood. Here, we evaluated the expression of factors associated with mast cell activation and malaria-associated bacteremia in a rodent model. C57BL/6J mice were infected with Plasmodium yoelii yoelli 17XNL, and blood and tissues were collected over time to assay for circulating levels of bacterial 16S DNA, IgE, mast cell protease 1 (Mcpt-1) and Mcpt-4, Th1 and Th2 cytokines, and patterns of ileal mastocytosis and intestinal permeability. The anti-inflammatory cytokines (interleukin-4 [IL-4], IL-6, and IL-10) and MCP-1/CCL2 were detected early after P. yoelii yoelii 17XNL infection. This was followed by the appearance of IL-9 and IL-13, cytokines known for their roles in mast cell activation and growth-enhancing activity as well as IgE production. Later increases in circulating IgE, which can induce mast cell degranulation, as well as Mcpt-1 and Mcpt-4, were observed concurrently with bacteremia and increased intestinal permeability. These results suggest that P. yoelii yoelii 17XNL infection induces the production of early cytokines that activate mast cells and drive IgE production, followed by elevated IgE, IL-9, and IL-13 that maintain and enhance mast cell activation while disrupting the protease/antiprotease balance in the intestine, contributing to epithelial damage and increased permeability

Mast Cell Chymase/Mcpt4 Suppresses the Host Immune Response to Plasmodium yoelii, Limits Malaria-Associated Disruption of Intestinal Barrier Integrity and Reduces Parasite Transmission to Anopheles stephensi.

An increase in mast cells (MCs) and MCs mediators has been observed in malaria-associated bacteremia, however, the role of these granulocytes in malarial immunity is poorly understood. Herein, we studied the role of mouse MC protease (Mcpt) 4, an ortholog of human MC chymase, in malaria-induced bacteremia using Mcpt4 knockout (Mcpt4 -/-) mice and Mcpt4 +/+ C57BL/6J controls, and the non-lethal mouse parasite Plasmodium yoelii yoelii 17XNL. Significantly lower parasitemia was observed in Mcpt4 -/- mice compared with Mcpt4 +/+ controls by day 10 post infection (PI). Although bacterial 16S DNA levels in blood were not different between groups, increased intestinal permeability to FITC-dextran and altered ileal adherens junction E-cadherin were observed in Mcpt4 -/- mice. Relative to infected Mcpt4 +/+ mice, ileal MC accumulation in Mcpt4 -/- mice occurred two days earlier and IgE levels were higher by days 8-10 PI. Increased levels of circulating myeloperoxidase were observed at 6 and 10 days PI in Mcpt4 +/+ but not Mcpt4 -/- mice, affirming a role for neutrophil activation that was not predictive of parasitemia or bacterial 16S copies in blood. In contrast, early increased plasma levels of TNF-α, IL-12p40 and IL-3 were observed in Mcpt4 -/- mice, while levels of IL-2, IL-10 and MIP1β (CCL4) were increased over the same period in Mcpt4 +/+ mice, suggesting that the host response to infection was skewed toward a type-1 immune response in Mcpt4 -/- mice and type-2 response in Mcpt4 +/+ mice. Spearman analysis revealed an early (day 4 PI) correlation of Mcpt4 -/- parasitemia with TNF-α and IFN-γ, inflammatory cytokines known for their roles in pathogen clearance, a pattern that was observed in Mcpt4 +/+ mice much later (day 10 PI). Transmission success of P. y. yoelii 17XNL to Anopheles stephensi was significantly higher from infected Mcpt4 -/- mice compared with infected Mcpt4 +/+ mice, suggesting that Mcpt4 also impacts transmissibility of sexual stage parasites. Together, these results suggest that early MCs activation and release of Mcpt4 suppresses the host immune response to P. y. yoelii 17XNL, perhaps via degradation of TNF-α and promotion of a type-2 immune response that concordantly protects epithelial barrier integrity, while limiting the systemic response to bacteremia and parasite transmissibility

The Basophil IL-18 Receptor Precisely Regulates the Host Immune Response and Malaria-Induced Intestinal Permeability and Alters Parasite Transmission to Mosquitoes without Effect on Gametocytemia

We have recently demonstrated that basophils are protective against intestinal permeability during malaria and contribute to reduced parasite transmission to mosquitoes. Given that IL-18 is an early cytokine/alarmin in malaria and has been shown to activate basophils, we sought to determine the role of the basophil IL-18R in this protective phenotype. To address this, we infected control [IL18r flox/flox or basoIL-18R (+)] mice and mice with basophils lacking the IL-18R [IL18r flox/flox × Basoph8 or basoIL-18R (-)] with Plasmodium yoelii yoelii 17XNL, a nonlethal strain of mouse malaria. Postinfection (PI), intestinal permeability, ileal mastocytosis, bacteremia, and levels of ileal and plasma cytokines and chemokines were measured through 10 d PI. BasoIL-18R (-) mice exhibited greater intestinal permeability relative to basoIL-18R (+) mice, along with increased plasma levels of proinflammatory cytokines at a single time point PI, day 4 PI, a pattern not observed in basoIL-18R (+) mice. Surprisingly, mosquitoes fed on basoIL-18R (-) mice became infected less frequently than mosquitoes fed on basoIL-18R (+) mice, with no difference in gametocytemia, a pattern that was distinct from that observed previously with basophil-depleted mice. These findings suggest that early basophil-dependent protection of the intestinal barrier in malaria is mediated by IL-18, and that basophil IL-18R-dependent signaling differentially regulates the inflammatory response to infection and parasite transmission

Basophil Depletion Alters Host Immunity, Intestinal Permeability, and Mammalian Host-to-Mosquito Transmission in Malaria

Malaria-induced bacteremia has been shown to result from intestinal mast cell (MC) activation. The appearance of MCs in the ileum and increased intestinal permeability to enteric bacteria are preceded by an early Th2-biased host immune response to infection, characterized by the appearance of IL-4, IL-10, mast cell protease (Mcpt)1 and Mcpt4, and increased circulating basophils and eosinophils. Given the functional similarities of basophils and MCs in the context of allergic inflammation and the capacity of basophils to produce large amounts of IL-4, we sought to define the role of basophils in increased intestinal permeability, in MC influx, and in the development of bacteremia in the context of malaria. Upon infection with nonlethal Plasmodium yoelii yoelii 17XNL, Basoph8 × ROSA-DTα mice or baso (-) mice that lack basophils exhibited increased intestinal permeability and increased ileal MC numbers, without any increase in bacterial 16S ribosomal DNA copy numbers in the blood, relative to baso (+) mice. Analysis of cytokines, chemokines, and MC-associated factors in the ileum revealed significantly increased TNF-α and IL-13 at day 6 postinfection in baso (-) mice compared with baso (+) mice. Moreover, network analysis of significantly correlated host immune factors revealed profound differences between baso (-) and baso (+) mice following infection in both systemic and ileal responses to parasites and translocated bacteria. Finally, basophil depletion was associated with significantly increased gametocytemia and parasite transmission to Anopheles mosquitoes, suggesting that basophils play a previously undescribed role in controlling gametocytemia and, in turn, mammalian host-to-mosquito parasite transmission

Multi-omic brain and behavioral correlates of cell-free fetal DNA methylation in macaque maternal obesity models.

Maternal obesity during pregnancy is associated with neurodevelopmental disorder (NDD) risk. We utilized integrative multi-omics to examine maternal obesity effects on offspring neurodevelopment in rhesus macaques by comparison to lean controls and two interventions. Differentially methylated regions (DMRs) from longitudinal maternal blood-derived cell-free fetal DNA (cffDNA) significantly overlapped with DMRs from infant brain. The DMRs were enriched for neurodevelopmental functions, methylation-sensitive developmental transcription factor motifs, and human NDD DMRs identified from brain and placenta. Brain and cffDNA methylation levels from a large region overlapping mir-663 correlated with maternal obesity, metabolic and immune markers, and infant behavior. A DUX4 hippocampal co-methylation network correlated with maternal obesity, infant behavior, infant hippocampal lipidomic and metabolomic profiles, and maternal blood measurements of DUX4 cffDNA methylation, cytokines, and metabolites. We conclude that in this model, maternal obesity was associated with changes in the infant brain and behavior, and these differences were detectable in pregnancy through integrative analyses of cffDNA methylation with immune and metabolic factors

Baseline immunoreactivity before pregnancy and poly(I:C) dose combine to dictate susceptibility and resilience of offspring to maternal immune activation

Despite the potential of rodent models of maternal immune activation (MIA) to identify new biomarkers and therapeutic interventions for a range of psychiatric disorders, current approaches using these models ignore two of the most important aspects of this risk factor for human disease: (i) most pregnancies are resilient to maternal viral infection and (ii) susceptible pregnancies can lead to different combinations of phenotypes in offspring. Here, we report two new sources of variability-the baseline immunoreactivity (BIR) of isogenic females prior to pregnancy and differences in immune responses in C57BL/6 dams across vendors-that contribute to resilience and susceptibility to distinct combinations of behavioral and biological outcomes in offspring. Similar to the variable effects of human maternal infection, MIA in mice does not cause disease-related phenotypes in all pregnancies and a combination of poly(I:C) dose and BIR predicts susceptibility and resilience of pregnancies to aberrant repetitive behaviors and alterations in striatal protein levels in offspring. Even more surprising is that the intermediate levels of BIR and poly(I:C) dose are most detrimental to offspring, with higher BIR and poly(I:C) doses conferring resilience to measured phenotypes in offspring. Importantly, we identify the BIR of female mice as a biomarker before pregnancy that predicts which dams will be most at risk as well as biomarkers in the brains of newborn offspring that correlate with changes in repetitive behaviors. Together, our results highlight considerations for optimizing MIA protocols to enhance rigor and reproducibility and reveal new factors that drive susceptibility of some pregnancies and resilience of others to MIA-induced abnormalities in offspring

Sequential perturbations to mouse corticogenesis following in utero maternal immune activation.

In utero exposure to maternal immune activation (MIA) is an environmental risk factor for neurodevelopmental and neuropsychiatric disorders. Animal models provide an opportunity to identify mechanisms driving neuropathology associated with MIA. We performed time-course transcriptional profiling of mouse cortical development following induced MIA via poly(I:C) injection at E12.5. MIA-driven transcriptional changes were validated via protein analysis, and parallel perturbations to cortical neuroanatomy were identified via imaging. MIA-induced acute upregulation of genes associated with hypoxia, immune signaling, and angiogenesis, by 6 hr following exposure. This acute response was followed by changes in proliferation, neuronal and glial specification, and cortical lamination that emerged at E14.5 and peaked at E17.5. Decreased numbers of proliferative cells in germinal zones and alterations in neuronal and glial populations were identified in the MIA-exposed cortex. Overall, paired transcriptomic and neuroanatomical characterization revealed a sequence of perturbations to corticogenesis driven by mid-gestational MIA

Sequential perturbations to mouse corticogenesis following in utero maternal immune activation.

In utero exposure to maternal immune activation (MIA) is an environmental risk factor for neurodevelopmental and neuropsychiatric disorders. Animal models provide an opportunity to identify mechanisms driving neuropathology associated with MIA. We performed time-course transcriptional profiling of mouse cortical development following induced MIA via poly(I:C) injection at E12.5. MIA-driven transcriptional changes were validated via protein analysis, and parallel perturbations to cortical neuroanatomy were identified via imaging. MIA-induced acute upregulation of genes associated with hypoxia, immune signaling, and angiogenesis, by 6 hr following exposure. This acute response was followed by changes in proliferation, neuronal and glial specification, and cortical lamination that emerged at E14.5 and peaked at E17.5. Decreased numbers of proliferative cells in germinal zones and alterations in neuronal and glial populations were identified in the MIA-exposed cortex. Overall, paired transcriptomic and neuroanatomical characterization revealed a sequence of perturbations to corticogenesis driven by mid-gestational MIA