Trypanosoma cruzi Exploits E- and P-Selectins To Migrate Across Endothelial Cells and Extracellular Matrix Proteins.

The Chagas disease parasite Trypanosoma cruzi must extravasate to home in on susceptible cells residing in most tissues. It remains unknown how T. cruzi undertakes this crucial step of its life cycle. We hypothesized that the pathogen exploits the endothelial cell programming leukocytes use to extravasate to sites of inflammation. Transendothelial migration (TEM) starts after inflammatory cytokines induce E-selectin expression and P-selectin translocation on endothelial cells (ECs), enabling recognition by leukocyte ligands that engender rolling cell adhesion. Here, we show that T. cruzi upregulates E- and P-selectins in cardiac ECs to which it binds in a ligand-receptor fashion, whether under static or shear flow conditions. Glycoproteins isolated from T. cruzi (TcEx) specifically recognize P-selectin in a ligand-receptor interaction. As with leukocytes, binding of P-selectin to T. cruzi or TcEx requires sialic acid and tyrosine sulfate, which are pivotal for downstream migration across ECs and extracellular matrix proteins. Additionally, soluble selectins, which bind T. cruzi, block transendothelial migration dose dependently, implying that the pathogen bears selectin-binding ligand(s) that start transmigration. Furthermore, function-blocking antibodies against E- and P-selectins, which act on endothelial cells and not T. cruzi, are exquisite in preventing TEM. Thus, our results show that selectins can function as mediators of T. cruzi transendothelial transmigration, suggesting a pathogenic mechanism that allows homing in of the parasite on targeted tissues. As selectin inhibitors are sought-after therapeutic targets for autoimmune diseases and cancer metastasis, they may similarly represent a novel strategy for Chagas disease therapy.

Trypanosoma cruzi Neurotrophic Factor Facilitates Cardiac Repair in a Mouse Model of Chronic Chagas Disease.

Most patients acutely infected with Trypanosoma cruzi undergo short-term structural and functional cardiac alterations that heal without sequelae. By contrast, in patients whose disease progresses to chronic infection, irreversible degenerative chronic Chagas cardiomyopathy (CCC) may develop. To account for the contrast between cardiac regeneration in high-parasitism acute infection and progressive cardiomyopathy in low-parasitism CCC, we hypothesized that T. cruzi expresses repair factors that directly facilitate cardiac regeneration. We investigated, as one such repair factor, the T. cruzi parasite-derived neurotrophic factor (PDNF), known to trigger survival of cardiac myocytes and fibroblasts and upregulate chemokine chemokine C-C motif ligand 2, which promotes migration of regenerative cardiac progenitor cells (CPCs). Using in vivo and in vitro models of Chagas disease, we tested whether T. cruzi PDNF promotes cardiac repair. Quantitative PCR and flow cytometry of heart tissue revealed that stem-cell antigen-1 (Sca-1+) CPCs expand in acute infection in parallel to parasitism. Recombinant PDNF induced survival and expansion of ex vivo CPCs, and intravenous administration of PDNF into naïve mice upregulated mRNA of cardiac stem-cell marker Sca-1. Furthermore, in CCC mice, a 3-week intravenous administration of PDNF protocol induced CPC expansion and reversed left ventricular T-cell accumulation and cardiac remodeling including fibrosis. Compared with CCC vehicle-treated mice, which developed severe atrioventricular block, PDNF-treated mice exhibited reduced frequency and severity of conduction abnormalities. Our findings are in support of the novel concept that T. cruzi uses PDNF to promote mutually beneficial cardiac repair in Chagas disease. This could indicate a possible path to prevention or treatment of CCC.

The novel flavone tetramethoxyluteolin is a potent inhibitor of human mast cells.

BACKGROUND: Mast cells (MCs) are hematopoietic cells that mature in tissues and are involved in allergy, immunity, and inflammation by secreting multiple mediators. The natural flavone luteolin has anti-inflammatory actions and inhibits human mast cells (MCs). OBJECTIVE: We sought to investigate the ability of luteolin and its novel structural analog 3',4',5,7-tetramethoxyluteolin (methlut) to inhibit human MC mediator expression and release in vitro and in vivo. METHODS: Human LAD2 cells and umbilical primary human cord blood-derived cultured mast cells were stimulated with substance P (SP) or IgE/anti-IgE with or without preincubation with luteolin, methlut, or cromolyn (1-100 µmol/L) for 2 or 24 hours, after which mediator secretion was measured. The effect of the compounds on MC intracellular calcium levels and nuclear factor κB activation was also investigated. Pretreatment with methlut was also studied in mice passively sensitized with dinitrophenol-human serum albumin and challenged intradermally. RESULTS: Methlut is a more potent inhibitor than luteolin or cromolyn for ß-hexosaminidase and histamine secretion from LAD2 cells stimulated by either SP or IgE/anti-IgE, but only methlut and luteolin significantly inhibit preformed TNF secretion. Methlut is also a more potent inhibitor than luteolin of de novo-synthesized TNF from LAD2 cells and of CCL2 from human cord blood-derived cultured MCs. This mechanism of action for methlut might be due to its ability to inhibit intracellular calcium level increases, as well as nuclear factor κB induction, at both the transcriptional and translational levels in LAD2 cells stimulated by SP without affecting cell viability. Intraperitoneal treatment with methlut significantly decreases skin vascular permeability of Evans blue dye in mice passively sensitized to dinitrophenol-human serum albumin and challenged intradermally. CONCLUSION: Methlut is a promising MC inhibitor for the treatment of allergic and inflammatory conditions.

Mast cells in meningiomas and brain inflammation.

BACKGROUND: Research focus in neuro-oncology has shifted in the last decades towards the exploration of tumor infiltration by a variety of immune cells and their products. T cells, macrophages, B cells, and mast cells (MCs) have been identified. METHODS: A systematic review of the literature was conducted by searching PubMed, EMBASE, Google Scholar, and Turning Research into Practice (TRIP) for the presence of MCs in meningiomas using the terms meningioma, inflammation and mast cells. RESULTS: MCs have been detected in various tumors of the central nervous system (CNS), such as gliomas, including glioblastoma multiforme, hemangioblastomas, and meningiomas as well as metastatic brain tumors. MCs were present in as many as 90 % of all high-grade meningiomas mainly found in the perivascular areas of the tumor. A correlation between peritumoral edema and MCs was found. INTERPRETATION: Accumulation of MCs in meningiomas could contribute to the aggressiveness of tumors and to brain inflammation that may be involved in the pathogenesis of additional disorders.

Targeting IL-33 in autoimmunity and inflammation.

Interleukin-33 (IL-33) belongs to the IL-1 family of cytokines. Whereas IL-1 is processed and released by live immune cells in response to infection or other triggers, IL-33 is mostly released as a danger signal ("alarmin") from damaged cells. IL-33 may also be processed and released from activated mast cells (MCs) with subsequent autocrine and paracrine actions. IL-33 augments the stimulatory effects of IgE and substance P on MCs but can also trigger release of cytokines from MCs on its own. Blood IL-33 levels are increased in asthma, atopic dermatitis, multiple sclerosis, rheumatoid arthritis, and Sjögren's syndrome. However, prolonged elevation of IL-33 downregulates FcεRI and may be protective in atherosclerosis, suggesting different roles in immune-regulated diseases. Even though neutralizing IL-33, knocking-down its receptor, or using its soluble "decoy" receptor has resulted in anti-inflammatory effects, there appear to be different outcomes in different tissues. Hence, selective regulation of IL-33 synthesis, release, and signaling may be required to provide effective treatment options.

Fibromyalgia syndrome in need of effective treatments.

Fibromyalgia syndrome (FMS) is a chronic, idiopathic condition of widespread musculoskeletal pain, affecting primarily women. It is clinically characterized by chronic, nonarticular pain and a heightened response to pressure along with sleep disturbances, fatigue, bowel and bladder abnormalities, and cognitive dysfunction. The diagnostic criteria have changed repeatedly, and there is neither a definitive pathogenesis nor reliable diagnostic or prognostic biomarkers. Clinical and laboratory studies have provided evidence of altered central pain pathways. Recent evidence suggests the involvement of neuroinflammation with stress peptides triggering the release of neurosenzitizing mediators. The management of FMS requires a multidimensional approach including patient education, behavioral therapy, exercise, and pain management. Here we review recent data on the pathogenesis and propose new directions for research and treatment.

Trigeminal nerve stimulation triggers oral mast cell activation and vascular permeability.

BACKGROUND: The nervous system contributes to the pathophysiology of allergic and inflammatory diseases, including oral inflammation. Mast cells (MCs) are involved in their pathogenesis through proinflammatory mediator release. OBJECTIVE: To investigate the effect of trigeminal nerve (TN) stimulation compared with sham operation on MC activation and oral vascular permeability in the gingiva, palate, buccal mucosa, and tongue of the rat and to examine the possible role of substance P using rats treated with capsaicin as neonates to deplete substance P. METHODS: Six male Sprague-Dawley rats (250 g) were anesthetized and injected intravenously with Evans Blue (EB). Six other rats were injected neonatally with capsaicin (n = 3) or solvent (n = 3) and then injected with EB when they reached 250 g. The mandibular branch of the TN was stimulated for 1 minute (n = 3), and the remaining rats (n = 3) were subjected to sham operation. The ipsilateral and contralateral sides of the mouth were examined for EB extravasation, and tissue sections were removed for light and electron microscopy. RESULTS: TN stimulation resulted in EB extravasation in the ipsilateral side compared with the contralateral side or the ipsilateral side of sham-operated rats. Significant degranulation of MCs also was evident only on the ipsilateral side (P < .0001). There was no difference in MC degranulation between the vehicle- and capsaicin-treated rats, implying that neuropeptides other than substance P may be involved. CONCLUSION: This is the first time that TN stimulation has been shown to result in MC activation and oral vascular permeability, suggesting that MC inhibitors may be used for the treatment of oral inflammatory diseases.

Stress triggers coronary mast cells leading to cardiac events.

OBJECTIVE: Stress precipitates and worsens not only asthma and atopic dermatitis but also acute coronary syndromes (ACSs), which are associated with coronary inflammation. Evidence linking stress to ACS was reviewed and indicated that activation of coronary mast cells (MCs) by stress, through corticotropin-releasing hormone (CRH) and other neuropeptides, contributes to coronary inflammation and coronary artery disease. DATA SOURCES: PubMed was searched (2005-2013) for articles using the following keywords: allergies, anaphylaxis, anxiety, coronary arteries, coronary artery disease, C-reactive protein, cytokines, chymase, histamine, hypersensitivity, interleukin-6 (IL-6), inflammation, mast cells, myocardial ischemia, niacin, platelet-activating factor, rupture, spasm, statins, stress, treatment, tryptase, and uroctortin. STUDY SELECTIONS: Articles were selected based on their relevance to how stress affects ACS and how it activates coronary MCs, leading to coronary hypersensitivity, inflammation, and coronary artery disease. RESULTS: Stress can precipitate allergies and ACS. Stress stimulates MCs through the activation of high-affinity surface receptors for CRH, leading to a CRH-dependent increase in serum IL-6. Moreover, neurotensin secreted with CRH from peripheral nerves augments the effect of CRH and stimulates cardiac MCs to release IL-6, which is elevated in ACS and is an independent risk factor for myocardial ischemia. MCs also secrete CRH and uroctortin, which induces IL-6 release from cardiomyocytes. The presence of atherosclerosis increases the risk of cardiac MC activation owing to the stimulatory effect of lipoproteins and adipocytokines. Conditions such as Kounis syndrome, mastocytosis, and myalgic encephalopathy/chronic fatigue syndrome are particularly prone to coronary hypersensitivity reactions. CONCLUSION: Inhibition of cardiac MCs may be a novel treatment approach.

Antigen presentation by cardiac fibroblasts promotes cardiac dysfunction.

Heart failure (HF) is a leading cause of morbidity and mortality. Studies in animal models and patients with HF revealed a prominent role for CD4+ T cell immune responses in the pathogenesis of HF and highlighted an active crosstalk between cardiac fibroblasts and IFNγ producing CD4+ T cells that results in profibrotic myofibroblast transformation. Whether cardiac fibroblasts concomitantly modulate pathogenic cardiac CD4+ T cell immune responses is unknown. Here we show report that murine cardiac fibroblasts express major histocompatibility complex type II (MHCII) in two different experimental models of cardiac inflammation. We demonstrate that cardiac fibroblasts take up and process antigens for presentation to CD4+ T cells via MHCII induced by IFNγ. Conditional deletion of MhcII in cardiac fibroblasts ameliorates cardiac remodelling and dysfunction induced by cardiac pressure overload. Collectively, we demonstrate that cardiac fibroblasts function as antigen presenting cells (APCs) and contribute to cardiac fibrosis and dysfunction through IFNγ induced MHCII.

Topical Application of a Mast Cell Stabilizer Improves Impaired Diabetic Wound Healing.

Impaired wound healing in the diabetic foot is a major problem often leading to amputation. Mast cells have been shown to regulate wound healing in diabetes. We developed an indole-carboxamide type mast cell stabilizer, MCS-01, which proved to be an effective mast cell degranulation inhibitor in vitro and can be delivered topically for prolonged periods through controlled release by specifically designed alginate bandages. In diabetic mice, both pre- and post-wounding, topical MCS-01 application accelerated wound healing comparable to that achieved with systemic mast cell stabilization. Moreover, MCS-01 altered the macrophage phenotype, promoting classically activated polarization. Bulk transcriptome analysis from wounds treated with MCS-01 or placebo showed that MCS-01 significantly modulated the mRNA and microRNA profile of diabetic wounds, stimulated upregulation of pathways linked to acute inflammation and immune cell migration, and activated the NF-κB complex along with other master regulators of inflammation. Single-cell RNA sequencing analysis of 6,154 cells from wounded and unwounded mouse skin revealed that MCS-01 primarily altered the gene expression of mast cells, monocytes, and keratinocytes. Taken together, these findings offer insights into the process of diabetic wound healing and suggest topical mast cell stabilization as a potentially successful treatment for diabetic foot ulceration.

Mast cells, brain inflammation and autism.

Increasing evidence indicates that brain inflammation is involved in the pathogenesis of neuropsychiatric diseases. Mast cells (MCs) are located perivascularly close to neurons and microglia, primarily in the leptomeninges, thalamus, hypothalamus and especially the median eminence. Corticotropin-releasing factor (CRF) is secreted from the hypothalamus under stress and, together with neurotensin (NT), can stimulate brain MCs to release inflammatory and neurotoxic mediators that disrupt the blood-brain barrier (BBB), stimulate microglia and cause focal inflammation. CRF and NT synergistically stimulate MCs and increase vascular permeability; these peptides can also induce each other׳s surface receptors on MCs leading to autocrine and paracrine effects. As a result, brain MCs may be involved in the pathogenesis of "brain fog," headaches, and autism spectrum disorders (ASDs), which worsen with stress. CRF and NT are significantly increased in serum of ASD children compared to normotypic controls further strengthening their role in the pathogenesis of autism. There are no clinically affective treatments for the core symptoms of ASDs, but pilot clinical trials using natural-antioxidant and anti-inflammatory molecules reported statistically significant benefit.

Mast Cells Regulate Wound Healing in Diabetes.

Diabetic foot ulceration is a severe complication of diabetes that lacks effective treatment. Mast cells (MCs) contribute to wound healing, but their role in diabetes skin complications is poorly understood. Here we show that the number of degranulated MCs is increased in unwounded forearm and foot skin of patients with diabetes and in unwounded dorsal skin of diabetic mice (P < 0.05). Conversely, postwounding MC degranulation increases in nondiabetic mice, but not in diabetic mice. Pretreatment with the MC degranulation inhibitor disodium cromoglycate rescues diabetes-associated wound-healing impairment in mice and shifts macrophages to the regenerative M2 phenotype (P < 0.05). Nevertheless, nondiabetic and diabetic mice deficient in MCs have delayed wound healing compared with their wild-type (WT) controls, implying that some MC mediator is needed for proper healing. MCs are a major source of vascular endothelial growth factor (VEGF) in mouse skin, but the level of VEGF is reduced in diabetic mouse skin, and its release from human MCs is reduced in hyperglycemic conditions. Topical treatment with the MC trigger substance P does not affect wound healing in MC-deficient mice, but improves it in WT mice. In conclusion, the presence of nondegranulated MCs in unwounded skin is required for proper wound healing, and therapies inhibiting MC degranulation could improve wound healing in diabetes.

Dysregulated brain immunity and neurotrophin signaling in Rett syndrome and autism spectrum disorders.

Rett syndrome is a neurodevelopmental disorder, which occurs in about 1:15,000 females and presents with neurologic and communication defects. It is transmitted as an X-linked dominant linked to mutations of the methyl-CpG-binding protein (MeCP2), a gene transcription suppressor, but its definitive pathogenesis is unknown thus hindering development of effective treatments. Almost half of children with Rett syndrome also have behavioral symptoms consistent with those of autism spectrum disorders (ASDs). PubMed was searched (2005-2014) using the terms: allergy, atopy, brain, brain-derived neurotrophic factor (BDNF), corticotropin-releasing hormone (CRH), cytokines, gene mutations, inflammation, mast cells (MCs), microglia, mitochondria, neurotensin (NT), neurotrophins, seizures, stress, and treatment. There are a number of intriguing differences and similarities between Rett syndrome and ASDs. Rett syndrome occurs in females, while ASDs more often in males, and the former has neurologic disabilities unlike ASDs. There is evidence of dysregulated immune system early in life in both conditions. Lack of microglial phagocytosis and decreased levels of BDNF appear to distinguish Rett syndrome from ASDs, in which there is instead microglia activation and/or proliferation and possibly defective BDNF signaling. Moreover, brain mast cell (MC) activation and focal inflammation may be more prominent in ASDs than Rett syndrome. The flavonoid luteolin blocks microglia and MC activation, provides BDNF-like activity, reverses Rett phenotype in mouse models, and has a significant benefit in children with ASDs. Appropriate formulations of luteolin or other natural molecules may be useful in the treatment of Rett syndrome.

Gut-Microbiota-Brain Axis and Its Effect on Neuropsychiatric Disorders With Suspected Immune Dysregulation.

PURPOSE: Gut microbiota regulate intestinal function and health. However, mounting evidence indicates that they can also influence the immune and nervous systems and vice versa. This article reviews the bidirectional relationship between the gut microbiota and the brain, termed the microbiota-gut-brain (MGB) axis, and discusses how it contributes to the pathogenesis of certain disorders that may involve brain inflammation. METHODS: Articles were identified with a search of Medline (starting in 1980) by using the key words anxiety, attention-deficit hypersensitivity disorder (ADHD), autism, cytokines, depression, gut, hypothalamic-pituitary-adrenal (HPA) axis, inflammation, immune system, microbiota, nervous system, neurologic, neurotransmitters, neuroimmune conditions, psychiatric, and stress. FINDINGS: Various afferent or efferent pathways are involved in the MGB axis. Antibiotics, environmental and infectious agents, intestinal neurotransmitters/neuromodulators, sensory vagal fibers, cytokines, and essential metabolites all convey information to the central nervous system about the intestinal state. Conversely, the hypothalamic-pituitary-adrenal axis, the central nervous system regulatory areas of satiety, and neuropeptides released from sensory nerve fibers affect the gut microbiota composition directly or through nutrient availability. Such interactions seem to influence the pathogenesis of a number of disorders in which inflammation is implicated, such as mood disorder, autism-spectrum disorders, attention-deficit hypersensitivity disorder, multiple sclerosis, and obesity. IMPLICATIONS: Recognition of the relationship between the MGB axis and the neuroimmune systems provides a novel approach for better understanding and management of these disorders. Appropriate preventive measures early in life or corrective measures such as use of psychobiotics, fecal microbiota transplantation, and flavonoids are discussed.

Exosomes in neurologic and psychiatric disorders.

PURPOSES: The purposes of this review were to discuss the role of exosomes in neurologic and psychiatric diseases and to propose future therapeutic approaches. METHODS: PubMed was searched (2000-2014) using the terms exosomes, microvesicles, neurological disorders, psychiatric disorders, multivesicular bodies, Alzheimer's disease, Parkinson's disease, prion disease, multiple sclerosis, schizophrenia, glioblastoma multiforme, and flavonoids. FINDINGS: Many cells of the nervous system have been reported to release exosomes that could have an active role in the function, development, and diseases of the CNS, such as Alzheimer disease, Parkinson disease, prion diseases, multiple sclerosis, brain tumors, and schizophrenia. In all of these diseases, exosomes are involved in the spread of "toxic" proteins that are mutated or "misfolded" and serve as templates for the formation of disease-producing oligomers. IMPLICATIONS: Exosomes' simple structure and abilities to be incorporated into plasma membrane and to cross the blood-brain barrier allow for the opportunity to utilize them as delivery vehicles of drugs and genetic elements in the treatment of immune, psychiatric, and neurologic disorders. Flavonoids have emerged as unique, natural molecules with antioxidant and antiinflammatory properties. It would, therefore, be of interest to design flavonoid-containing exosomes.

Spectrum of mast cell activation disorders.

Mast cell (MC) activation disorders present with multiple symptoms including flushing, pruritus, hypotension, gastrointestinal complaints, irritability, headaches, concentration/memory loss and neuropsychiatric issues. These disorders are classified as: cutaneous and systemic mastocytosis with a c-kit mutation and clonal MC activation disorder, allergies, urticarias and inflammatory disorders and mast cell activation syndrome (MCAS), idiopathic urticaria and angioedema. MCs are activated by IgE, but also by cytokines, environmental, food, infectious, drug and stress triggers, leading to secretion of multiple mediators. The symptom profile and comorbidities associated with these disorders, such as chronic fatigue syndrome and fibromyalgia, are confusing. We propose the use of the term 'spectrum' and highlight the main symptoms, useful diagnostic tests and treatment approaches.

The "missing link" in autoimmunity and autism: extracellular mitochondrial components secreted from activated live mast cells.

Autoimmune diseases continue to increase, but the reason(s) remain obscure and infections have not proven to be major contributors. Mast cells are tissue immune cells responsible for allergies, but have been increasingly shown to be involved in innate and acquired immunity, as well as inflammation. This involvement is possible because of their ability to release multiple mediators in response to a great variety of triggers. We recently published that activation of mast cells is accompanied by mitochondrial fission and translocation to the cell surface from where they secrete at least ATP and DNA outside the cell without cell damage. These extracellular mitochondrial components are misconstrued by the body as "innate pathogens" leading to powerful autocrine and paracrine auto-immune/auto-inflammatory responses. We also showed that mitochondrial DNA is increased in the serum of young children with autism spectrum disorders (ASD), a condition that could involve "focal brain allergy/encephalitits". Blocking the secretion of extracellular mitochondrial components could present unique possibilities for the therapy of ASD and other autoimmune diseases. Unique formulation of the flavonoid luteolin offers unique advantages.