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UNLABELLED: Gonorrhea, caused by the bacterium Neisseria gonorrhoeae (Gc), is characterized by neutrophilic influx to infection sites. Gc has developed mechanisms to resist killing by neutrophils that include modifications to its surface lipooligosaccharide (LOS). One such LOS modification is sialylation: Gc sialylates its terminal LOS sugars with cytidine-5-monophosphate-N-acetylneuraminic acid, which is scavenged from the host using LOS sialyltransferase (Lst) since Gc cannot make its sialic acid. Sialylation enables sensitive strains of Gc to resist complement-mediated killing in a serum-dependent manner. However, little is known about the contribution of sialylation to complement-independent, direct Gc-neutrophil interactions. In the absence of complement, we found sialylated Gc expressing opacity-associated (Opa) proteins decreased the oxidative burst and granule exocytosis from primary human neutrophils. In addition, sialylated Opa+ Gc survived better than vehicle treated or Δlst Gc when challenged with neutrophils. However, Gc sialylation did not significantly affect Opa-dependent association with or internalization of Gc by neutrophils. Previous studies have implicated sialic acid-binding immunoglobulin-type lectins (Siglecs) in modulating neutrophil interactions with sialylated Gc. Blocking neutrophil Siglecs with antibodies that bind to their extracellular domains eliminated the ability of sialylated Opa+ Gc to suppress the oxidative burst and resist neutrophil killing. These findings highlight a new role for sialylation in Gc evasion of human innate immunity, with implications for the development of vaccines and therapeutics for gonorrhea. IMPORTANCE: Neisseria gonorrhoeae, the bacterium that causes gonorrhea, is an urgent global health concern due to increasing infection rates, widespread antibiotic resistance, and its ability to thwart protective immune responses. The mechanisms by which Gc subverts protective immune responses remain poorly characterized. One way N. gonorrhoeae evades human immunity is by adding sialic acid that is scavenged from the host onto its lipooligosaccharide, using the sialyltransferase Lst. Here, we found that sialylation enhances N. gonorrhoeae survival from neutrophil assault and inhibits neutrophil activation, independently of the complement system. Our results implicate bacterial binding of sialic acid-binding lectins (Siglecs) on the neutrophil surface, which dampens neutrophil antimicrobial responses. This work identifies a new role for sialylation in protecting N. gonorrhoeae from cellular innate immunity, which can be targeted to enhance the human immune response in gonorrhea.

Mas-related G protein-coupled receptor b2 (Mrgprb2) binding to its cationic endogenous and exogenous ligands induces mast cell degranulation and promotes inflammation in mice. However, the physiological roles of its human homologue MRGPRX2 remain unclear. Here we aimed to elucidate the mechanisms by which MRGPRX2 regulates vascular permeability, and generated MRGPRX2 knock-in (MRGPRX2-KI) and Mrgprb2 knockout (Mrgprb2-KO) mice. Substance P (SP) and ciprofloxacin strongly degranulated MRGPRX2-KI peritoneal mast cells (PMCs) better than WT PMCs, whereas Dermatophagoides pteronyssinus (Der p) extract and phenol-soluble modulin α3 (PSMα3) did not degranulate PMCs. SP-stimulated MRGPRX2-KI PMCs released large amounts of histamine and mast cell protease 4 (MCPT4) chymase. Der p extract, PSMα3, and MCPT4, but not histamine, induced SP release from dorsal root ganglion (DRG) cells. However, this effect of Der p extract/PSMα3 was suppressed by a transient receptor potential vanilloid 1 (TRPV1) antagonist. SP-, ciprofloxacin-, Der p extract-, PSMα3-, and MCPT4-induced vascular permeability was highest in MRGPRX2-KI mice, which depended on SP. In addition, SP-, ciprofloxacin- and PSMα3-induced MRGPRX2-dependent vascular hyperpermeability was suppressed by antihistamine and chymase inhibitor. TRPV1 antagonist also inhibited PSMα3-induced MRGPRX2-dependent vascular hyperpermeability. Both Mrgprb2-KO and MRGPRX2-KI did not influence the histamine-induced murine vascular hyperpermeability. Overall, our results suggest that neuronal SP induces MRGPRX2-dependent mast cell degranulation, releasing histamine and chymase, which promote vascular hyperpermeability directly or indirectly via DRG cell activation. Importantly, the worsening cycle (MRGPRX2 → mast cell degranulation → chymase → DRG activation → SP → MRGPRX2) seems to play an important role in human MRGPRX2-depdendent inflammation. [ABSTRACT FROM AUTHOR]

Chronic spontaneous urticaria (CSU) is a debilitating condition characterized by mast cell activation. Platelet-activating factor (PAF) is produced by various immune cells, including mast cells, basophils, lymphocytes, and eosinophils, which play crucial roles in CSU pathogenesis. It induces mast cell degranulation, increases vascular permeability, and promotes the chemotaxis of inflammatory cells. These effects result in the release of inflammatory mediators, the development of edema, and the persistence of inflammation, which are key features of CSU. Notably, elevated PAF levels have been linked to heightened disease activity and resistance to antihistamine treatment in CSU patients. Despite these findings, the precise role of PAF in CSU pathogenesis remains unclear. Rupatadine, an antihistamine, and heat shock protein 10, a natural anti-inflammatory peptide that selectively inhibits PAF-induced mast cell degranulation, have demonstrated anti-PAF activity. Furthermore, with the molecular structure of the PAF receptor now identified, several experimental PAF receptor antagonists have been synthesized. However, there remains a significant need for the development of therapeutic options targeting PAF in CSU management. [ABSTRACT FROM AUTHOR]

We studied morphofunctional features of human platelets exposed in vitro to low-pulse laser radiation (LPLR) with different wavelengths. Platelet-rich plasma (PRP) was irradiated at wavelengths of 635, 488, and 355 nm. LPLR with λ=635 nm and 1 W power after 10 min caused degranulation of many platelets, formation of small platelet conglomerates, and accelerated platelet adhesion on glass. LPLR with λ=635 nm and 2 mW and LPLR with λ=488 nm did not cause significant changes in the morphofunctional platelet status. LPLR with λ=355 nm induced massive platelets' deformation, sharp decrease in their morphofunctional status without activation. [ABSTRACT FROM AUTHOR]

This study aimed to validate the DFS (direct fast scarlet) staining in the diagnosis of EC (eosinophilic colitis). The study included 50 patients with EC and 60 with control colons. Among the 60 control samples, 39 and 21 were collected from the ascending and descending colons, respectively. We compared the median number of eosinophils and frequency of eosinophil degranulation by HE (hematoxylin and eosin) and DFS staining between the EC and control groups. In the right hemi‐colon, eosinophil count by HE was useful in distinguishing between EC and control (41.5 vs. 26.0 cells/HPF, p < 0.001), but the ideal cutoff value is 27.5 cells/HPF (high‐power field). However, this method is not useful in the left hemi‐colon (12.5 vs. 13.0 cells/HPF, p = 0.990). The presence of degranulation by DFS allows us to distinguish between the groups even in the left hemi‐colon (58% vs. 5%, p < 0.001). DFS staining also enabled a more accurate determination of degranulation than HE. According to the current standard to diagnose EC (count by HE staining ≥20 cells/HPF), mucosal sampling from left hemi‐colon is problematic since the number of eosinophils could not be increased even in EC. Determination of degranulated eosinophils by DFS may potentiate the diagnostic performance even in such conditions. [ABSTRACT FROM AUTHOR]

Skin mast cells (MCs) mediate acute allergic reactions in the cutaneous environment and contribute to chronic dermatoses, including urticaria, and atopic or contact dermatitis. The cAMP response element binding protein (CREB), an evolutionarily well conserved transcription factor (TF) with over 4,000 binding sites in the genome, was recently found to form a feedforward loop with KIT, maintaining MC survival. The most selective MC function is degranulation with its acute release of prestored mediators. Herein, we asked whether CREB contributes to the expression and function of the degranulation-competent receptors FcεRI and MRGPRX2. Interference with CREB by pharmacological inhibition (CREBi, 666-15) or RNA interference only slightly affected the expression of these receptors, while KIT was strongly attenuated. Interestingly, MRGPRX2 surface expression moderately increased following CREB-knockdown, whereas MRGPRX2-dependent exocytosis simultaneously decreased. FcεRI expression and function were regulated consistently, although the effect was stronger at the functional level. Preformed MC mediators (tryptase, histamine, β-hexosaminidase) remained comparable following CREB attenuation, suggesting that granule synthesis did not rely on CREB function. Collectively, in contrast to KIT, FcεRI and MRGPRX2 moderately depend on unperturbed CREB function. Nevertheless, CREB is required to maintain MC releasability irrespective of stimulus, insinuating that CREB may operate by safeguarding the degranulation machinery. To our knowledge, CREB is the first factor identified to regulate MRGPRX2 expression and function in opposite direction. Overall, the ancient TF is an indispensable component of skin MCs, orchestrating not only survival and proliferation but also their secretory competence. [ABSTRACT FROM AUTHOR]

Cationic liposomes composed of cholesteryl‐3β‐carboxyamidoethylene‐N‐hydroxyethylamine (OH‐chol) and 1,2‐dioleoyl‐sn‐glycero‐3‐phosphatidylethanolamine (DOPE) inhibit mast cell degranulation mediated via crosslinking of high‐affinity IgE receptors (FcεRI). Although the inhibitory efficiency of mast cell degranulation is altered by modifying the ratio of OH‐chol and DOPE in cationic liposomes, the manner in which physicochemical properties, such as surface charge and size, influence suppression is not clear. We observed that positive surface charge, but not the size, of liposomes plays a role in suppressing rat basophilic leukemia (RBL‐2H3) cell activation. Pretreatment with middle‐ratio OH‐chol liposomes (zeta potential, 62.2 ± 0.5 mV; diameter, 325.4 ± 7.3 nm) exhibited a larger suppression of RBL‐2H3 cell degranulation evoked by FcεRI crosslinking compared with that by low‐ratio OH‐chol liposomes (zeta potential, 48.6 ± 1.9 mV; diameter, 344.4 ± 25.0 nm), although both liposomes were similarly attached to RBL‐2H3 cells. Preparation of middle‐ratio OH‐chol liposomes, classified roughly by size using an extrusion method, revealed that the liposomal size did not affect the inhibitory efficiency of RBL‐2H3 cell activation. Mechanistically, we found that middle‐ratio OH‐chol liposomes increased the inhibition of antigen‐induced Akt phosphorylation compared to low‐ratio OH‐chol liposomes. We measured the phosphorylation of linker for activation of T cells (LAT) and paxillin, which are important proteins in FcεRI‐ and focal adhesions (FAs)‐mediated signaling, respectively. Middle ratio OH‐chol liposomes significantly suppressed antigen‐induced paxillin phosphorylation, but did not affect LAT phosphorylation, suggesting that middle‐ratio OH‐chol liposomes attached to RBL‐2H3 cells suppress the degranulation by impairing FA‐mediated Akt phosphorylation evoked by FcεRI crosslinking. [ABSTRACT FROM AUTHOR]

Mas-related G protein–coupled receptor X2 (MRGPRX2) is a promiscuous receptor on mast cells that mediates IgE-independent degranulation and has been implicated in multiple mast cell–mediated disorders, including chronic urticaria, atopic dermatitis, and pain disorders. Although it is a promising therapeutic target, few potent, selective, small molecule antagonists have been identified, and functional effects of human MRGPRX2 inhibition have not been evaluated in vivo. We sought to identify and characterize novel, potent, and selective orally active small molecule MRGPRX2 antagonists for potential treatment of mast cell–mediated disease. Antagonists were identified using multiple functional assays in cell lines overexpressing human MRGPRX2, LAD2 mast cells, human peripheral stem cell–derived mast cells, and isolated skin mast cells. Skin mast cell degranulation was evaluated in Mrgprb2 em(−/−) knockout and Mrgprb2 em(MRGPRX2) transgenic human MRGPRX2 knock-in mice by assessment of agonist-induced skin vascular permeability. Ex vivo skin mast cell degranulation and associated histamine release was evaluated by microdialysis of human skin tissue samples. MRGPRX2 antagonists potently inhibited agonist-induced MRGPRX2 activation and mast cell degranulation in all mast cell types tested in an IgE-independent manner. Orally administered MRGPRX2 antagonists also inhibited agonist-induced degranulation and resulting vascular permeability in MRGPRX2 knock-in mice. In addition, antagonist treatment dose dependently inhibited agonist-induced degranulation in ex vivo human skin. MRGPRX2 small molecule antagonists potently inhibited agonist-induced mast cell degranulation in vitro and in vivo as well as ex vivo in human skin, supporting potential therapeutic utility as a novel treatment for multiple human diseases involving clinically relevant mast cell activation. [ABSTRACT FROM AUTHOR]

While several functions of the endogenous prion protein have been studied, the homeostatic function of prion protein is still debated. Notably, prion protein is highly expressed on mast cells, granular immune cells that regulate inflammation. When activated, mast cells shed prion protein, although the mechanism and consequences of this are not yet understood. First, we tested several mast cell lines and found that, while prion protein was almost always present, the total amount differed greatly. Activation of mast cells induced a cleavage of the N-terminal region of prion protein, and this was reduced by protease inhibitors. Exogenous mast cell proteases caused a similar loss of the prion protein N-terminus. Additionally, mast cells shed prion protein in an ADAM10-dependent fashion, even in the absence of activation. Our results suggest that prion protein is cleaved from resting mast cells by ADAM10 and from activated mast cells by mast cell proteases. Prion protein also appears to affect mast cell function, as Prnp −/− bone marrow–derived mast cells showed lower levels of degranulation and cytokine release, as well as lower levels of both FcεRI and CD117. Finally, we sought to provide clinical relevance by measuring the levels of prion protein in bodily fluids of asthmatic patients, a disease that involves the activation of mast cells. We found an N-terminal fragment of prion protein could be detected in human sputum and serum, and the amount of this prion protein fragment was decreased in the serum of patients with asthma. [ABSTRACT FROM AUTHOR]

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Abstract Although the cellular prion protein (PrPC) has been evolutionarily conserved, the role of this protein remains elusive. Recent evidence indicates that PrPC may be involved in neuroinflammation and the immune response in the brain, and its expression may be modified via various mechanisms. Histamine is a proinflammatory mediator and neurotransmitter that stimulates numerous cells via interactions with histamine receptors 1-4 (HRH1-4). Since microglia are the innate immune cells of the central nervous system, we hypothesized that histamine-induced stimulation regulates the expression of PrPC in human-derived microglia. The human microglial clone 3 (HMC3) cell line was treated with histamine, and intracellular calcium levels were measured via a calcium flux assay. Cytokine production was monitored by enzyme-linked immunosorbent assay (ELISA). Western blotting and quantitative reverse transcription-polymerase chain reaction were used to determine protein and gene expression of HRH1-4. Flow cytometry and western blotting were used to measure PrPC expression levels. Fluorescence microscopy was used to examine Iba-1 and PrPC localization. HMC3 cells stimulated by histamine exhibited increased intracellular calcium levels and increased release of IL-6 and IL-8, while also modifying PrPC localization. HMC3 stimulated with histamine for 6 and 24 hours exhibited increased surface PrPC expression. Specifically, we found that stimulation of the HRH2 receptor was responsible for changes in surface PrPC. Histamine-induced increases in surface PrPC were attenuated following inhibition of the HRH2 receptor via the HRH2 antagonist ranitidine. These changes were unique to HRH2 activation, as stimulation of HRH1, HRH3, or HRH4 did not alter surface PrPC. Prolonged stimulation of HMC3 decreased PrPC expression following 48 and 72 hours of histamine stimulation. HMC3 cells can be stimulated by histamine to undergo intracellular calcium influx. Surface expression levels of PrPC on HMC3 cells are altered by histamine exposure, primarily mediated by HRH2. While histamine exposure also increases release of IL-6 and IL-8 in these cells, this cytokine release is not fully dependent on PrPC levels, as IL-6 release is only partially reduced and IL-8 release is unchanged under the conditions of HRH2 blockade that prevent PrPC changes. Overall, this suggests that PrPC may play a role in modulating microglial responses.

Background: Mast cells play an important role in allergic reactions by releasing potent mediators, including histamine and platelet-activating factor (PAF), upon activation. We recently reported that a biotinylated heptapeptide (peptide 2), D-Lys(Biotinyl)-Trp-Tyr-Lys-Asp-Gly-Asp, which is highly stable in the plasma, inhibits PAF bioactivity and anaphylactic hypothermia in vivo. However, the effects of this peptide on allergy induction by mast cells have not been investigated. Methods and Results: Several biotinylated heptapeptides, including peptide 2, inhibited IgE/antigen-stimulated degranulation in RBL-2H3 cells. Peptide 2 also markedly inhibited the degranulation induced by calcium ionophore A23187. All peptides (peptide 1, 2 and 3) inhibited activating stimuli-induced increase in intracellular Ca2+ levels. The transforming growth factor-alpha shedding assay indicated that peptide 2 antagonized against the histamine H1 receptor. Furthermore, peptide 2 inhibited histamine-induced rat paw edema. Conclusion: These results highlight the potential of peptide 2 in the prevention and treatment of various allergic diseases mediated by mast cell activation. [ABSTRACT FROM AUTHOR]

The key problem of introducing nanoparticles into clinical practice for diagnostic and therapeutic purposes is their safety. Connective tissue, an important component of which are mast cells, reacts actively in response to foreign particles. The reaction of mast cells can be an indicator of the biocompatibility of foreign particles. The study was conducted on male Wistar rats. Iron-carbon nanoparticles in the FeC modification stabilized in an aqueous medium using an auxiliary substance were used. Tissue examination (liver, lungs, heart, thymus, kidneys) was performed 1, 7 and 30 days after injection. After the injection of nanoparticles, the largest quantity of them is found in the liver and lungs, a smaller quantity is found in the heart, kidneys and thymus. The liver and lungs are the main organs of excretion of nanoparticles due to phagocytes. The accumulation of nanoparticles in the liver leads to the development of destructive processes and to the activation of compensatory and adaptive mechanisms in the form of cellular and intracellular regeneration of hepatocytes. In organs with a low content of nanoparticles, structural changes are poorly expressed. The mast cells of the studied organs react differently to the introduction of nanoparticles. The first to react are the mast cells of the liver by reducing the quantity and increasing degranulation. The population of lung mast cells reacts unidirectionally by sharply activating degranulation without changing the amount. An increase in the secretory activity of mast cells in the lungs and liver indicates the participation of mast cells in the regulation of particle elimination through intercellular signaling pathways of interaction with the system of phagocytic mononuclear cells. Heart mast cells are involved in maintaining the inflammatory process in the early period of the experiment, contribute to the return of myocardial parameters to homeostatic norm in the late period of the experiment. Mast cells can be considered as indicators of biocompatibility nanoparticles. The absence of an inflammatory process and the preservation of the structural and functional characteristics of the tissues where nanoparticles accumulate, as well as the reaction of mast cells in them, indicate the relative safety of the particles under study.

Abstract Background Eosinophilic extracellular traps (EETs) are reticular complexes comprising deoxyribonucleic‐Acid (DNA) fibers and granule proteins. Aims EETs play a crucial role in antimicrobial host responses and are pathogenic when overproduced or under degraded. EETs created by eosinophils appear to enable vital immune responses against extra‐cellular pathogens, nevertheless, trap overproduction is evident in pathology. Materials & Methods As considerably research is performed, new data affirmed that EETs can alter the outcome of respiratory ailment. Results We probe into the disclosure and specificity of EETs produced in reaction to various stimuli and propose a role for those frameworks in ailment pathogenesis and the establishment of chronic, unresolved inflammation. Discussion Whether EETs can be used as a prospective brand‐new target for the diagnosis, treatment and prognosis of respiratory ailments is a scientific theme worth studying. Conclusion We probe into the disclosure and specificity of EETs produced in reaction to various stimuli and propose a role for those frameworks in ailment pathogenesis and the establishment of chronic, unresolved inflammation.

Food allergy, particularly peanut allergy (PA), is a growing health concern affecting millions globally. PA can lead to severe reactions, including fatal anaphylaxis. Despite the availability of FDA-approved therapies like Palforzia, a cure remains elusive. Current immunotherapies show promise but lack a definitive cure. This study applies an established computational biology tool to design aptamers targeting Ara h1 and Ara h2. The in silico design aims to streamline the selection process, enabling cost-effective and rapid identification of aptamer candidates. The developed aptamers (AYA22A, including AYA22AR321, AYA22AR211, and AYA22AR524), demonstrated efficacy in inhibiting degranulation of RBL-2H3 cells (rat basophilic leukemia cell line) in vitro. They showed promise in neutralizing peanut allergen-induced immune responses. The selected aptamers inhibited degranulation in RBL-2H3 cells, addressing concerns in raw peanuts. Moreover, these aptamers demonstrated stability and effectiveness in peanut plant seeds and commercial products. Our aptamers exhibited potential in modulating immune responses associated with peanut allergy. They influenced Th1/Th2 balance, indicating a role in cytokine regulation. In vitro studies also showed the aptamers’ impact on immune cell expression and cytokine production, resembling responses observed with established immunotherapies. The findings suggest AYA22A aptamers as a potential therapeutic option for peanut allergy, providing a basis for further in vivo investigations.

Upon degranulation, basophils and mast cells secrete an array of inflammatory mediators, including histamine, which leads to not only allergic inflammation but also other inflammatory diseases. We previously reported that an aqueous extract from enzyme-treated, dried sardine inhibits the degranulation of RBL-2H3 cells and attenuates the symptoms of Japanese cedar pollinosis in mice. This study evaluated an antiallergic effect of dipeptides containing acidic amino acid residue in an antigen-induced degranulation assay using RBL-2H3 cells. The result showed that acidic amino acid residue-containing dipeptides inhibit the degranulation of RBL-2H3 cells without cytotoxicity. Additionally, L-histidyl-L-glutamic acid (His-Glu), one of the acidic amino acid residue-containing dipeptides tested in this study, inhibited calcium ionophore-induced degranulation. We also found that His-Glu suppressed microtubule reorganization in RBL-2H3 cells after antigen stimulation. His-Glu slightly, but not significantly, suppressed the elevation of cytosolic calcium ion concentration leading to degranulation. Immunoblot analysis revealed that His-Glu significantly suppressed the phosphorylation of phosphoinositide 3-kinase and Akt, but not that of Syk or phospholipase Cγ. Overall results suggest that acidic amino acid residue-containing dipeptides can be used as food ingredients with an antiallergic effect. [ABSTRACT FROM AUTHOR]

BACKGROUND: Food allergy has become a global public health problem. This study aimed to explore the possible anti‐allergic effect of vitamin C (VC). A rat basophilic leukemia (RBL)‐2H3 cell degranulation model was used to assess the effect of VC on degranulation in vitro, and an ovalbumin (OVA)‐induced BALB/c mouse allergy model was used to assess the anti‐allergy effect of VC in vivo. RESULTS: In vitro, VC significantly attenuated the release of β‐hexosaminidase, tryptase and histamine, and also reduced cytokine production (interleukins 4 and 6, tumor necrosis factor α) significantly (P < 0.05), with the inhibitory effect demonstrating a positive correlation with VC dose. In vivo, compared with the OVA group, the levels of serum immunoglobulins E and G1 of the VC low‐dose (VCL) group (50 mg kg−1) and high‐dose (VCH) group (200 mg·kg−1) were significantly reduced (P < 0.05). Furthermore, the plasma histamine level was also significantly decreased (P < 0.05). Moreover, TH2 cell polarization in mice of the VCL and VCH groups was significantly inhibited (P < 0.05), promoting the TH1/TH2 cell polarization balance. Additionally, VC treatment enhanced the expression of CD80 (P < 0.05) in spleen and small intestine tissues, while significantly inhibiting the expression of CD86 (P < 0.05); notably, high‐dose VC treatment was more effective. CONCLUSION: VC exerted an anti‐allergic effect through inhibiting degranulation and regulating TH1/TH2 cell polarization balance. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Eosinophil-mediated pathophysiology is tissue destructive and tissue altering with proinflammatory, prothrombotic, and profibrotic effects. The distinctive morphology of an eosinophil reveals a cytoplasm chockfull of unique granules, and the granule proteins have numerous toxic effects on cells, tissues, and organs. Eosinophils are not found in most human tissues, and eosinophil involvement in diseased tissues generally is identified by cell infiltration on histopathologic examination. However, eosinophils characteristically lose their structural integrity and deposit granules and granule proteins at sites of inflammation. Hence, their participation in tissue damage may be underrecognized or entirely overlooked. The eosinophil major basic protein 1 is a toxic granule protein and, when deposited, persists in tissues. Major basic protein 1 deposition can be regarded as a footprint of eosinophil activity. Analyses of numerous eosinophil-related diseases have demonstrated clear-cut evidence of major basic protein 1 deposition in affected tissues where eosinophils were not recognized by hematoxylin and eosin tissue staining and light microscopy. Eosinophil granule protein deposition, as exemplified by localization of major basic protein 1, especially when disproportionately greater than cellular infiltration, emerges as a biomarker of hidden eosinophil-related pathophysiology. Consequently, current assessments of recognized eosinophils may vastly underestimate their role in disease. [ABSTRACT FROM AUTHOR]

Objectives: Mast cell (MC) degranulation is a key process in allergic reactions and inflammatory responses. Aspartate aminotransferase 1 (AAT1)-derived endogenous sulfur dioxide (SO2) is an important regulator of MC function. However, the mechanism underlying its role in MC degranulation remains unclear. This study aimed to investigate the mechanism by which endogenous SO2 controlled MC degranulation. Methods: HMC-1 and Rat basophilic leukemia cell MC line (RBL-2H3) were used in the cell experiments. SO2 content was detected by in situ fluorescent probe. MC degranulation represented by the release rate of MC β-hexosaminidase was determined using a colorimetric assay. Sulfenylation of galectin-9 (Gal-9) in MCs and purified protein was detected using a biotin switch assay. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to determine the exact sulfenylation sites of Gal-9 by SO2. Animal models of passive cutaneous anaphylaxis (PCA) and hypoxia-driven pulmonary vascular remodeling were used to investigate the effect of SO2 on mast cell activation in vivo. Site-directed mutation of Gal-9 was conducted to confirm the exact site of SO2 and support the significance of SO2/Gal-9 signal axis in the regulation of MC degranulation. Results: Degranulation was increased in AAT1-knockdowned MCs, and SO2 supplementation reversed the increase in MC degranulation. Furthermore, deficiency of endogenous SO2 contributed to IgE-mediated degranulation in vitro. Besides, SO2 inhibited IgE-mediated and hypoxia-driven MC degranulation in vivo. Mechanistically, LC-MS/MS analysis and site-directed mutation results showed that SO2 sulfenylated Gal-9 at cysteine 74. Sulfenylation of the 74th cysteine of Gal-9 protein was required in the SO2-inhibited MC degranulation under both physiological and pathophysiological conditions. Conclusion: These findings elucidated that SO2 inhibited MC degranulation via sulfenylating Gal-9 under both physiological and pathophysiological conditions, which might provide a novel treatment approach for MC activation-related diseases. [ABSTRACT FROM AUTHOR]

The anti-allergic effects of extracts prepared from two species of honeybush, Cyclopia genistoides and Cyclopia subternata, were demonstrated in vivo in a murine allergy model for inhaled antigen induced with ovalbumin (OVA) inhalation to mimic pollen allergy. Intake of the extracts increased the production of OVA-specific immunoglobulin (Ig) E (IgE), IgG1, and IgG2a antibodies in serum and significantly suppressed anaphylactic reaction-induced body temperature decline. Moreover, the extracts significantly inhibited antigen-antibodyinduced degranulation in RBL-2H3 cells. They also inhibited body temperature decline when the allergic mice were given them after antigen sensitization, indicating that anti-degranulation activity is the major mechanism underlying the anti-allergic effect of Cyclopia extracts. Despite their qualitative and quantitative differences in phenolic composition, the two extracts exhibited similar effects, suggesting that several active compounds might be involved in the activity. Therefore, oral administration of either Cyclopia extract potentially exerts a systemic anti-allergic effect, supporting the increased consumption of honeybush tea for general wellness and improved quality of life. [ABSTRACT FROM AUTHOR]

Mas-related G protein-coupled receptor b2 (Mrgprb2) binding to its cationic endogenous and exogenous ligands induces mast cell degranulation and promotes inflammation in mice. However, the physiological roles of its human homologue MRGPRX2 remain unclear. Here we aimed to elucidate the mechanisms by which MRGPRX2 regulates vascular permeability, and generated MRGPRX2 knock-in (MRGPRX2-KI) and Mrgprb2 knockout (Mrgprb2-KO) mice. Substance P (SP) and ciprofloxacin strongly degranulated MRGPRX2-KI peritoneal mast cells (PMCs) better than WT PMCs, whereas Dermatophagoides pteronyssinus (Der p) extract and phenol-soluble modulin α3 (PSMα3) did not degranulate PMCs. SP-stimulated MRGPRX2-KI PMCs released large amounts of histamine and mast cell protease 4 (MCPT4) chymase. Der p extract, PSMα3, and MCPT4, but not histamine, induced SP release from dorsal root ganglion (DRG) cells. However, this effect of Der p extract/PSMα3 was suppressed by a transient receptor potential vanilloid 1 (TRPV1) antagonist. SP-, ciprofloxacin-, Der p extract-, PSMα3-, and MCPT4-induced vascular permeability was highest in MRGPRX2-KI mice, which depended on SP. In addition, SP-, ciprofloxacin- and PSMα3-induced MRGPRX2-dependent vascular hyperpermeability was suppressed by antihistamine and chymase inhibitor. TRPV1 antagonist also inhibited PSMα3-induced MRGPRX2-dependent vascular hyperpermeability. Both Mrgprb2-KO and MRGPRX2-KI did not influence the histamine-induced murine vascular hyperpermeability. Overall, our results suggest that neuronal SP induces MRGPRX2-dependent mast cell degranulation, releasing histamine and chymase, which promote vascular hyperpermeability directly or indirectly via DRG cell activation. Importantly, the worsening cycle (MRGPRX2 → mast cell degranulation → chymase → DRG activation → SP → MRGPRX2) seems to play an important role in human MRGPRX2-depdendent inflammation.

Background: Mast cells are critically involved in IgE-mediated diseases, e.g., allergies and asthma. Human mast cells are heterogeneous, and mast cells from different anatomical sites have been shown to respond differently to certain stimuli and drugs. The origin of the mast cells is therefore of importance when setting up a model system, and human lung mast cells are highly relevant cells to study in the context of asthma. We therefore set out to optimize a protocol of IgE-mediated activation of human lung mast cells. Methods: Human lung mast cells were extracted from lung tissue obtained from patients undergoing pulmonary resection by enzyme digestion and mechanical disruption followed by CD117 magnetic-activated cell sorting (MACS) enrichment. Different culturing media and conditions for the IgE-mediated degranulation were tested to obtain an optimized method. Results: IgE crosslinking of human lung mast cells cultured in serum-free media gave a stronger response compared to cells cultured with 10% serum. The addition of stem cell factor (SCF) did not enhance the degranulation. However, when the cells were put in fresh serum-free media 30 minutes prior to the addition of anti-IgE antibodies, the cells responded more vigorously. Maximum degranulation was reached 10 minutes after the addition of anti-IgE. Both CD63 and CD164 were identified as stable markers for the detection of degranulated mast cells over time, while the staining with anti-CD107a and avidin started to decline 10 minutes after activation. The levels of CD203c and CD13 did not change in activated cells and therefore cannot be used as degranulation markers of human lung mast cells. Conclusions: For an optimal degranulation response, human lung mast cells should be cultured and activated in serum-free media. With this method, a very strong and consistent degranulation response with a low donor-to-donor variation is obtained. Therefore, this model is useful for further investigations of IgE-mediated mast cell activation and exploring drugs that target human lung mast cells, for instance, in the context of asthma. [ABSTRACT FROM AUTHOR]

Ginseng leaves are known to contain high concentrations of bioactive compounds, such as ginsenosides, and have potential as a treatment for various conditions, including fungal infections, cancer, obesity, oxidative stress, and age-related diseases. This study assessed the impact of ginseng leaf extract (GLE) on mast cell-mediated allergic inflammation and atopic dermatitis (AD) in DNCB-treated mice. GLE reduced skin thickness and lymph node nodules and suppressed the expression and secretion of histamine and proinflammatory cytokines. It also significantly lowered the production of inflammatory response mediators including ROS, leukotriene C4 (LTC4), prostaglandin E2 (PGE2), cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS). GLE inhibited the phosphorylation of MAPKs (ERK, P38, JNK) and the activation of NF-B, which are both linked to inflammatory cytokine expression. We demonstrated that GLE's inhibitory effect on mast cellmediated allergic inflammation is due to the blockade of the NF-B and inflammasome pathways. Our findings suggest that GLE can be an effective therapeutic agent for mast-cell mediated and allergic inflammatory conditions. [ABSTRACT FROM AUTHOR]

This study aims to investigate the antiallergic effects of Shiikuwasha (Citrus depressa Hayata) leaf and peel extracts by examining the regulation of degranulation and inflammatory cytokine production from rat basophilic leukemia (RBL-2H3) cells and antigen-specific antibody production in sensitized mouse spleen lymphocytes. In vivo antiallergic activity was evaluated using the passive cutaneous anaphylaxis (PCA) reaction model. Extracts of Shiikuwasha leaves and peel were prepared using 80% methanol and dissolved in dimethylsulfoxide. The dinitrophenyl-human serum albumin-induced β-hexosaminidase levels in immunoglobulin (Ig) E-sensitized RBL-2H3 cells were assessed using enzymatic assays. Cytokine production was measured by enzyme-linked immunosorbent assay. Antibody production capacity was evaluated using lymphocytes isolated from spleens of type I allergy model mice. Lymphocytes were cultured for 72 h with Shiikuwasha extracts, and ovalbumin-specific IgE, IgG1, and IgG2a levels were measured. Shiikuwasha leaf and peel extract significantly reduced β-hexosaminidase release and suppressed interleukin-4 and tumor necrosis factor-a production from RBL-2H3 cells. Ovalbumin-specific IgE and IgG1 production decreased in Shiikuwasha extracttreated lymphocytes. These extracts also significantly suppressed the PCA reaction. Shiikuwasha leaf and peel extract reduce degranulation in RBL-2H3 cells and antibody production in spleen-derived lymphocytes and therefore exhibit antiallergic effects. [ABSTRACT FROM AUTHOR]

Background: Polyoxyethylene hydrogenated castor oil (HCO ethoxylates) is a nonionic surfactant used as an excipient for ointments and injections in human and veterinary drugs. Several polyethylene glycol (PEG) derivatives can be obtained depending on the number of moles of ethylene oxide (EO). HCO ethoxylates have the potential to cause anaphylactoid reactions. There is little published information about these types of reactions in dogs. Objective: To determine the potential for HCO‐ethoxylate‐containing drugs to cause anaphylactoid reactions in dogs, employing intradermal testing (IDT) with various concentrations of HCO ethoxylates (HCO‐25, ‐40, ‐60 and ‐80). Animals: Four healthy male laboratory dogs. Materials and Methods: We performed IDT with drugs containing HCO ethoxylates and HCO ethoxylates alone to determine threshold concentrations. The IDT scores and threshold concentrations were compared. Analysis of skin biopsies from IDT sites was used to measure the percentage of degranulated mast cells. The effect of histamine at IDT sites was investigated by pre‐treatment with an antihistamine. Results: All HCO‐ethoxylate‐containing drugs caused a wheal‐and‐flare reaction. The threshold concentrations (0.001% and 0.00001%) of each HCO‐ethoxylate depended on the number of moles of EO (p < 0.05). Mast cell degranulation was enhanced by all HCO ethoxylates. The HCO‐60‐induced reaction was suppressed by an oral antihistamine. Conclusions and Clinical Relevance: The threshold concentration can serve as a consideration for developing safe new drug formulations and for clinical decision‐making around using drugs containing PEG derivatives. IDT is useful to predict the risk of adverse effects. Antihistamines could demonstrate a prophylactic effect. [ABSTRACT FROM AUTHOR]

The aim of this study was to elucidate the anti-allergic effects of polymethoxyflavonoids in combination with milk proteins and the mechanism of inhibition. Three polymethoxyflavonoids and two milk proteins were exposed to the rat basophilic leukemia cell line RBL-2H3. β-hexosaminidase was used as an indicator of degranulation inhibition. The mechanism of inhibition was examined by measuring intracellular Ca2+ levels and western blot method. In the degranulation inhibition test with polymethoxyflavonoids and milk proteins alone, nobiletin was the strongest inhibitor in the polymethoxyflavonoid group and lactoferrin in the milk protein group. Next, co-stimulation with nobiletin and lactoferrin showed stronger synergistic degranulation inhibition than treatment with nobiletin or lactoferrin alone. Western blot analysis showed that co-stimulation with nobiletin and lactoferrin significantly downregulated the induction of phospholipase Cγ 1 phosphorylation. The degranulation response in RBL-2H3 cells was synergistically suppressed by co-stimulation of nobiletin and lactoferrin acting on both Ca2+-dependent and Ca2+-independent pathways. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Food Safety & Quality is the property of Journal of Food Safety & Quality Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

SARS-CoV-2–induced excessive inflammation in brain leads to damage of blood– brain barrier, hypoxic-ischemic injury, and neuron degeneration. The production of inflammatory cytokines by brain microvascular endothelial cells and microglia is reported to be critically associated with the brain pathology of COVID-19 patients. However, the cellular mechanisms for SARS-CoV-2–inducing activation of brain cells and the subsequent neuroinflammation remain to be fully delineated. Our research, along with others’, has recently demonstrated that SARS-CoV-2–induced accumulation and activation of mast cells (MCs) in mouse lung could further induce inflammatory cytokines and consequent lung damages. Intracerebral MCs activation and their cross talk with other brain cells could induce neuroinflammation that play important roles in neurodegenerative diseases including virus-induced neuro-pathophysiology. In this study, we investigated the role of MC activation in SARSCoV-2–induced neuroinflammation. We found that (1) SARS-CoV-2 infection triggered MC accumulation in the cerebrovascular region of mice; (2) spike/RBD (receptor-binding domain) protein–triggered MC activation induced inflammatory factors in human brain microvascular endothelial cells and microglia; (3) MC activation and degranulation destroyed the tight junction proteins in brain microvascular endothelial cells and induced the activation and proliferation of microglia. These findings reveal a cellular mechanism of SARS-CoV-2– induced neuroinflammation. [ABSTRACT FROM AUTHOR]

Mast cells have long been recognized for their involvement in allergic pathology through the immunoglobulin E (IgE)-mediated degranulation mechanism. However, there is growing evidence of other "non-canonical" degranulation mechanisms activated by certain pathogen recognition receptors. Mast cells release several mediators, including histamine, cytokines, chemokines, prostaglandins, and leukotrienes, to initiate and enhance inflammation. The chemical nature of activating stimuli influences receptors, triggering mechanisms for the secretion of formed and new synthesized mediators. Mast cells have more than 30 known surface receptors that activate different pathways for direct and indirect activation by microbes. Different bacterial strains stimulate mast cells through various ligands, initiating the innate immune response, which aids in clearing the bacterial burden. Mast cell interactions with adaptative immune cells also play a crucial role in infections. Recent publications revealed another "non-canonical" degranulation mechanism present in tryptase and chymase mast cells in humans and connective tissue mast cells in mice, occurring through the activation of the Mas-related G protein-coupled receptor (MRGPRX2/b2). This receptor represents a new therapeutic target alongside antibiotic therapy. There is an urgent need to reconsider and redefine the biological role of these MASTer cells of innate immunity, extending beyond their involvement in allergic pathology. [ABSTRACT FROM AUTHOR]

The pathophysiology of severe burn injuries in the early stages involves complex emergency responses, inflammatory reactions, immune system activation, and a significant increase in vascular permeability. Neutrophils, crucial innate immune cells, undergo rapid mobilization and intricate pathophysiological changes during this period. However, the dynamic alterations and detailed mechanisms governing their biological behavior remain unclear. Stomatin protein, an essential component of the cell membrane, stabilizes and regulates the membrane and participates in cell signal transduction. Additionally, it exhibits elevated expression in various inflammatory diseases. While Stomatin expression has been observed in the cell and granule membranes of neutrophils, its potential involvement in post-activation functional regulation requires further investigation. Neutrophils were isolated from human peripheral blood, mouse peripheral blood, and mouse bone marrow using the magnetic bead separation method. Flow cytometry was used to assess neutrophil membrane surface markers, ROS levels, and phagocytic activity. The expression of the Stomatin gene and protein was examined using quantitative real-time polymerase chain reaction and western blotting methods, respectively. Furthermore, the enzyme-linked immunosorbent assay was used to evaluate the expression of neutrophil-derived inflammatory mediators (myeloperoxidase (MPO), neutrophil elastase (NE), and matrix metalloproteinase 9 (MMP9)) in the plasma. Images and videos of vascular leakage in mice were captured using in vivo laser confocal imaging technology, whereas in vitro confocal microscopy was used to study the localization and levels of the cytoskeleton, CD63, and Stomatin protein in neutrophils. This study made the following key findings: (1) Early after severe burn, neutrophil dysfunction is present in the peripheral blood characterized by significant bone marrow mobilization, excessive degranulation, and impaired release and chemotaxis of inflammatory mediators (MPO, NE, and MMP9). (2) After burn injury, expression of both the stomatin gene and protein in neutrophils was upregulated. (3) Knockout (KO) of the stomatin gene in mice partially inhibited neutrophil excessive degranulation, potentially achieved via reduced production of primary granules and weakened binding of primary granules to the cell skeleton protein F-actin. (4) In severely burned mice, injury led to notable early-stage vascular leakage and lung damage, whereas Stomatin gene KO significantly ameliorated lung injury and vascular leakage. Stomatin promotes neutrophil degranulation in the early stage of severe burn injury via increasing the production of primary granules and enhancing their binding to the cell skeleton protein F-actin in neutrophils. Consequently, this excessive degranulation results in aggravated vascular leakage and lung injury. • Neutrophil dysfunction in the early stage of severe burn injury. • The stomatin gene and protein expression were upregulated after burn injury. • Knockout of the stomatin gene partially inhibited neutrophil excessive degranulation. • Stomatin gene KO significantly ameliorated lung injury and vascular leakage. [ABSTRACT FROM AUTHOR]

Natural killer (NK) cells have the ability to lyse other cells through the release of lytic granules (LGs). This is in part mediated by the small GTPase Rab27a, which was first identified to play a crucial role in degranulation through the study of individuals harboring mutations in the gene encoding Rab27a. However, the guanine nucleotide exchange factor (GEF) regulating the activation of Rab27a in cytotoxic lymphocytes was unknown. Here, we show that knockout of MADD significantly decreased the levels of GTP-bound Rab27a in both resting and stimulated NK cells, and MADD-deficient NK cells and CD8+ T cells displayed severely reduced degranulation and cytolytic ability, similar to that seen with Rab27a deficiency. Although MADD colocalized with Rab27a on LGs and was enriched at the cytolytic synapse, the loss of MADD did not impact Rab27a association with LGs nor their recruitment to the cytolytic synapse. Together, our results demonstrate an important role for MADD in cytotoxic lymphocyte killing. [ABSTRACT FROM AUTHOR]

Mast cells originate from the CD34+/CD117+ hematopoietic progenitors in the bone marrow, migrate into circulation, and ultimately mature and reside in peripheral tissues. Microbiota/metabolites and certain immune cells (e.g., Treg cells) play a key role in maintaining immune tolerance. Cross-linking of allergen-specific IgE on mast cells activates the high-affinity membrane-bound receptor FcεRI, thereby initiating an intracellular signal cascade, leading to degranulation and release of pro-inflammatory mediators. The intracellular signal transduction is intricately regulated by various kinases, transcription factors, and cytokines. Importantly, multiple signal components in the FcεRI-mast cell-mediated allergic cascade can be targeted for therapeutic purposes. Pharmacological interventions that include therapeutic antibodies against IgE, FcεRI, and cytokines as well as inhibitors/activators of several key intracellular signaling molecues have been used to inhibit allergic reactions. Other factors that are not part of the signal pathway but can enhance an individuals susceptibility to allergen stimulation are referred to as cofactors. Herein, we provide a mechanistic overview of the FcεRI-mast cell-mediated allergic signaling. This will broaden our scope and visions on specific preventive and therapeutic strategies for the clinical management of mast cell-associated hypersensitivity reactions.

To investigate the anti-allergic activity of lactic acid bacteria, lactic acid bacterial strains were obtained from fermented vegetables by the traditional isolation technique and screened for in vitro allergic activity by the degranulation inhibition assay against RBL-2H3 cells. The results showed that all four isolates exhibited good probiotic properties and safety and had a strong inhibitory effect on β-hexosaminidase released from RBL-2H3 cells after degranulation. Moreover, they showed significant down-regulation effects on the release of histamine, tumor necrosis factor-α and interleukin-4 from RBL-2H3 cell after degranulation, suggesting their potential as anti-allergic probiotics. The four strains were identified by 16S rDNA sequencing as two strains of Lactobacillus plantarum, one strain of L. fermentum and one strain of L. paracasei.

Abstract Hundreds of thousands of polymorphonuclear neutrophils (PMNs) are collected from the ocular surface upon waking, while few are harvested during daytime. This study aimed to investigate potential factors contributing to the circadian infiltration of tear PMNs, including changes in IL-8 and C5a in tears, and their phenotypes across different time points in a 24-h cycle. Tear PMNs were collected using a gentle eyewash after 2-h and 7-h of sleep (eye closure, EC) at night, after 2-h EC during the day, and towards the end of the afternoon. Significantly fewer cells were collected after 2-h EC during the day compared to 2-h EC at night. A positive correlation between IL-8 and PMN numbers existed, but not with C5a. Tear PMNs collected after 2-h EC at night were less degranulated and possessed a larger activation potential compared to 7-h EC. Tear PMNs from 7-h EC at night exhibited hyper-segmented nuclei and more NETosis compared to 2 h EC night, indicating an aged and activated phenotype. The diurnal-nocturnal recruitment pattern of tear PMNs may be driven by increased IL-8 in nighttime tears. Higher degranulation and NETs point to the significant activation of tear PMNs on the ocular surface during prolonged eye closure at night.

Eosinophils, a type of granulocyte derived from myeloid precursors in the bone marrow, are distinguished by their cytoplasmic granules. They play crucial roles in immunoregulation, tissue homeostasis, and host defense, while also contributing to the pathogenesis of various inflammatory diseases. Although long non-coding RNAs (lncRNAs) are known to be involved in eosinophilic conditions, their specific expression and functions within eosinophils have not been thoroughly investigated, largely due to the reliance on tissue homogenates. In an effort to address this gap, we analyzed publicly available high-throughput RNA sequencing data to identify lncRNAs associated with eosinophilic conditions. Among the identified lncRNAs, ITGB2 antisense RNA 1 (ITGB2-AS1) was significantly downregulated in blood eosinophils from patients with hypereosinophilia. To further explore its role in eosinophil biology, we generated a stable ITGB2-AS1 knockdown in the HL-60 cell line. Interestingly, ITGB2-AS1 deficiency led to impaired eosinophil differentiation, as evidenced by a reduction in cytoplasmic granules and decreased expression of key eosinophil granule proteins, including eosinophil peroxidase (EPX) and major basic protein-1 (MBP-1). Additionally, ITGB2-AS1-deficient cells exhibited compromised eosinophil effector functions, with reduced degranulation and impaired production of reactive oxygen species (ROS). These findings suggest that ITGB2-AS1 plays a pivotal role in eosinophil differentiation and function, positioning it as a novel regulator in eosinophil biology.

Skin mast cells (MCs) mediate acute allergic reactions in the cutaneous environment and contribute to chronic dermatoses, including urticaria, and atopic or contact dermatitis. The cAMP response element binding protein (CREB), an evolutionarily well conserved transcription factor (TF) with over 4,000 binding sites in the genome, was recently found to form a feedforward loop with KIT, maintaining MC survival. The most selective MC function is degranulation with its acute release of prestored mediators. Herein, we asked whether CREB contributes to the expression and function of the degranulation-competent receptors FcεRI and MRGPRX2. Interference with CREB by pharmacological inhibition (CREBi, 666-15) or RNA interference only slightly affected the expression of these receptors, while KIT was strongly attenuated. Interestingly, MRGPRX2 surface expression moderately increased following CREB-knockdown, whereas MRGPRX2-dependent exocytosis simultaneously decreased. FcεRI expression and function were regulated consistently, although the effect was stronger at the functional level. Preformed MC mediators (tryptase, histamine, β-hexosaminidase) remained comparable following CREB attenuation, suggesting that granule synthesis did not rely on CREB function. Collectively, in contrast to KIT, FcεRI and MRGPRX2 moderately depend on unperturbed CREB function. Nevertheless, CREB is required to maintain MC releasability irrespective of stimulus, insinuating that CREB may operate by safeguarding the degranulation machinery. To our knowledge, CREB is the first factor identified to regulate MRGPRX2 expression and function in opposite direction. Overall, the ancient TF is an indispensable component of skin MCs, orchestrating not only survival and proliferation but also their secretory competence.

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Tumor cells rely heavily on glycolysis to meet their high metabolic demands. While this results in nutrient deprivation within the tumor microenvironment and has negative effects on infiltrating immune cells such as natural killer (NK) cells, it also creates a potential target for cancer therapies. Here we use Glupin, an inhibitor of glucose transporters, to study the effect of limited glucose uptake on NK cells and their anti-tumor functions. Glupin treatment effectively inhibited glucose uptake and restricted glycolysis in NK cells. However, acute treatment had no negative effect on NK cell cytotoxicity or cytokine production. Long-term restriction of glucose uptake via Glupin treatment only delayed NK cell proliferation, as they could switch to glutaminolysis as an alternative energy source. While IFN-γ production was partially impaired, long-term Glupin treatment had no negative effect on degranulation. Interestingly, the serial killing activity of NK cells was even slightly enhanced, possibly due to changes in NAD metabolism. This demonstrates that NK cell cytotoxicity is remarkably robust and insensitive to metabolic disturbances, which makes cellular metabolism an attractive target for immune-mediated tumor therapies. [ABSTRACT FROM AUTHOR]

Copyright of Shipin Kexue/ Food Science is the property of Food Science Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Hemophagocytic Lymphohistiocytosis (HLH) is a rare clinical condition characterized by sustained but ineffective immune system activation, leading to severe and systemic hyperinflammation. It may occur as a genetic or sporadic condition, often triggered by an infection. The multifaceted pathogenesis results in a wide range of non-specific signs and symptoms, hampering early recognition. Despite a great improvement in terms of survival in the last decades, a considerable proportion of patients with HLH still die from progressive disease. Thus, prompt diagnosis and treatment are crucial for survival. Faced with the complexity and the heterogeneity of syndrome, expert consultation is recommended to correctly interpret clinical, functional and genetic findings and address therapeutic decisions. Cytofluorimetric and genetic analysis should be performed in reference laboratories. Genetic analysis is mandatory to confirm familial hemophagocytic lymphohistiocytosis (FHL) and Next Generation Sequencing is increasingly adopted to extend the spectrum of genetic predisposition to HLH, though its results should be critically discussed with specialists. In this review, we critically revise the reported laboratory tools for the diagnosis of HLH, in order to outline a comprehensive and widely available workup that allows to reduce the time between the clinical suspicion of HLH and its final diagnosis. [ABSTRACT FROM AUTHOR]

Mast cells (MCs) are commonly recognized for their crucial involvement in the pathogenesis of allergic diseases, but over time, it has come to light that they also play a role in the pathophysiology of non-allergic disorders including atherosclerosis. The involvement of MCs in the pathology of atherosclerosis is supported by their accumulation in atherosclerotic plaques upon their progression and the association of intraplaque MC numbers with acute cardiovascular events. MCs that accumulate within the atherosclerotic plaque release a cocktail of mediators through which they contribute to neovascularization, plaque progression, instability, erosion, rupture, and thrombosis. At a molecular level, MC-released proteases, especially cathepsin G, degrade low-density lipoproteins (LDL) and mediate LDL fusion and binding of LDL to proteoglycans (PGs). Through a complicated network of chemokines including CXCL1, MCs promote the recruitment of among others CXCR2+ neutrophils, therefore, aggravating the inflammation of the plaque environment. Additionally, MCs produce extracellular traps which worsen inflammation and contribute to atherothrombosis. Altogether, evidence suggests that MCs actively, via several underlying mechanisms, contribute to atherosclerotic plaque destabilization and acute cardiovascular syndromes, thus, making the study of interventions to modulate MC activation an interesting target for cardiovascular medicine. [ABSTRACT FROM AUTHOR]

The rising global incidence of IgE-mediated allergic reactions poses a significant challenge to the quality of life of affected individuals and to healthcare systems, with current treatments being limited in effectiveness, safety, and disease-modifying capabilities. IgE acts by sensitizing the highaffinity IgE receptor FcεRI expressed by mast cells and basophils, tuning these cells for inflammatory degranulation in response to future allergen encounters. In recent years, IgG has emerged as an essential negative regulator of IgE-dependent allergic inflammation. Mechanistically, studies have proposed different pathways by which IgG can interfere with the activation of IgE-mediated inflammation. Here, we briefly summarize the major proposed mechanisms of action by which IgG controls the IgE-FceRI inflammatory axis and how those mechanisms are currently applied as therapeutic interventions for IgE-mediated inflammation. [ABSTRACT FROM AUTHOR]

MRGPRX2, the human member of the MAS-related G-protein-coupled receptors (GPCRs), mediates the immunoglobulin E (IgE)-independent responses of a subset of mast cells (MCs) that are associated with itch, pain, neurogenic inflammation, and pseudoallergy to drugs. The mechanisms underlying the responses of MRGPRX2 to its multiple and diverse ligands are still not completely understood. Given the close association between GPCR location and function, and the key role played by Rab GTPases in controlling discrete steps along vesicular trafficking, we aimed to reveal the vesicular pathways that directly impact MRGPRX2-mediated exocytosis by identifying the Rabs that influence this process. For this purpose, we screened 43 Rabs for their functional and phenotypic impacts on MC degranulation in response to the synthetic MRGPRX2 ligand compound 48/80 (c48/80), which is often used as the gold standard of MRGPRX2 ligands, or to substance P (SP), an important trigger of neuroinflammatory MC responses. Results of this study highlight the important roles played by macropinocytosis and autophagy in controlling MRGPRX2-mediated exocytosis, demonstrating a close feedback control between the internalization and post-endocytic trafficking of MRGPRX2 and its triggered exocytosis. [ABSTRACT FROM AUTHOR]

Aim: The aim of the current research is the evaluation of immunomodulatory and anti-asthmatic activity of Actinidia deliciosa on Swiss albino mice by using a number of experimental models. Materials and Methods: The immunomodulatory and anti-asthmatic activity of Actinidia deliciosa in Swiss albino mice was evaluate by using models like delayed type hypersensitivity, neutrophil adhesion test, milk induced neutrophil leucocytes in mice, clonidine induced catalepsy activity in mice, clonidine induced mast cell degranulation method. Microscopic and histopathology images also taken during the experiment for better analysis of the outcomes. Results: The result obtained after performing the various models were observed that ethanolic extract of fruit of Actinidia deliciosa significantly **p<0.001 inhibit delayed type hypersensitivity, helps to increase neutrophil migration towards foreign body which are primary mediators and playsimportant role to fight infections, it also reduces leucocytes and eosinophilia count increased by milk, hence, work as anti-asthmatic agent. It was also found that the Actinidia deliciosa fruit extract reduces cataleptic activity of mice and protects mast cells from degranulation. Conclusion: The current study findings showed that ethanolic extract of Actinidia deliciosa fruits possess immunomodulatory and anti-asthmatic activity by reducing hypersensitivity reactions, increasing neutrophil migration towards antigens, reduces leucocytes and eosinophilia counts, also helps in the reduction of cataleptogenic activity and protects mass cell degranulation. [ABSTRACT FROM AUTHOR]

Neutrophils are important players in COVID‐19, contributing to tissue damage by release of inflammatory mediators, including ROS and neutrophil elastase. Longitudinal studies on the effects of COVID‐19 on neutrophil phenotype and function are scarce. Here, we longitudinally investigated the phenotype and degranulation of neutrophils in COVID‐19 patients (28 nonhospitalized and 35 hospitalized patients) compared with 17 healthy donors (HDs). We assessed phenotype, degranulation, CXCL8 (IL‐8) release, and ROS generation within 8 days, at one or 6 month(s) after COVID‐19 diagnosis. For degranulation and ROS production, we stimulated neutrophils, either with ssRNA and TNF or granulocyte‐macrophage colony‐stimulating factor and N‐Formylmethionyl‐leucyl‐phenylalanine. During active COVID‐19, neutrophils from hospitalized patients were more immature than from HDs and were impaired in degranulation and ROS generation, while neutrophils from nonhospitalized patients only demonstrated reduced CD66b+ granule release and ROS production. Baseline CD63 expression, indicative of primary granule release, and CXCL8 production by neutrophils from hospitalized patients were elevated for up to 6 months. These findings show that patients hospitalized due to COVID‐19, but not nonhospitalized patients, demonstrated an aberrant neutrophil phenotype, degranulation, CXCL8 release, and ROS generation that partially persists up to 6 months after infection. [ABSTRACT FROM AUTHOR]

Mast cells have long been recognized for their involvement in allergic pathology through the immunoglobulin E (IgE)-mediated degranulation mechanism. However, there is growing evidence of other “non-canonical” degranulation mechanisms activated by certain pathogen recognition receptors. Mast cells release several mediators, including histamine, cytokines, chemokines, prostaglandins, and leukotrienes, to initiate and enhance inflammation. The chemical nature of activating stimuli influences receptors, triggering mechanisms for the secretion of formed and new synthesized mediators. Mast cells have more than 30 known surface receptors that activate different pathways for direct and indirect activation by microbes. Different bacterial strains stimulate mast cells through various ligands, initiating the innate immune response, which aids in clearing the bacterial burden. Mast cell interactions with adaptative immune cells also play a crucial role in infections. Recent publications revealed another “non-canonical” degranulation mechanism present in tryptase and chymase mast cells in humans and connective tissue mast cells in mice, occurring through the activation of the Mas-related G protein–coupled receptor (MRGPRX2/b2). This receptor represents a new therapeutic target alongside antibiotic therapy. There is an urgent need to reconsider and redefine the biological role of these MASTer cells of innate immunity, extending beyond their involvement in allergic pathology.

Abstract Background Human placental extract (HPE) has been documented to facilitate the healing of certain disorders including allergy. However, the effects of HPE on the functionality of mast cells, a critical cell type in allergic diseases, have not been reported. Methods To investigate the effects of HPE on the regulation of allergy with respect to the biological functions of mast cells, the mast cell line C57 or HMC-1 cells were treated with HPE followed by the assessment of cell proliferation, apoptosis, activation, chemotaxis and phagocytosis. Mouse peritoneal mast cells were also investigated for their responses to induction of apoptosis by HPE in vivo. Furthermore, the effect of HPE on mast cell degranulation was confirmed using the passive cutaneous anaphylaxis (PCA) assay, an acute allergy model. Results HPE was capable of suppressing mast cell proliferation and inducing mast cell apoptosis. Mast cell degranulation in response to compound 48/80- or anti-DNP IgE and DNP-mediated activation was suppressed. In addition, treatment with HPE compromised the production of cytokines by mast cells and cell chemotaxis. These observations were consistent with the dampened passive cutaneous anaphylaxis (PCA) assay following treatment with HPE. Conclusion This study revealed a suppressive effect of HPE on overall mast cell activities, suggesting a potential regulatory role of HPE on the alleviation of allergic diseases through mast cells.