Bienzyme-Locked Activatable Fluorescent Probes for Specific Imaging of Tumor-Associated Mast Cells.

Tumor-associated mast cells (TAMCs) have been recently revealed to play a multifaceted role in the tumor microenvironment. Noninvasive optical imaging of TAMCs is thus highly desired to gain insights into their functions in cancer immunotherapy. However, due to the lack of a single enzyme that is specific to mast cells, a common probe design approach based on single-enzyme activation is not applicable. Herein, we reported a bienzyme-locked molecular probe (THCMC) based on a photoinduced electron transfer-intramolecular charge-transfer hybrid strategy for in vivo imaging of TAMCs. The bienzyme-locked activation mechanism ensures that THCMC exclusively turns on near-infrared (NIR) fluorescence only in the presence of both tryptase and chymase specifically coexpressed by mast cells. Thus, THCMC effectively distinguishes mast cells from other leukocytes, including T cells, neutrophils, and macrophages, a capability lacking in single-locked probes. Such a high specificity of THCMC allows noninvasive tracking of the fluctuation of TAMCs in the tumor of living mice during cancer immunotherapy. The results reveal that the decreased intratumoral signal of THCMC after combination immunotherapy correlates well with the reduced population of TAMCs, accurately predicting the inhibition of tumor growth. Thus, this study not only presents the first NIR fluorescent probe specific for TAMCs but also proposes a generic bienzyme-locked probe design approach for in vivo cell imaging.

Histochemical and immunohistochemical investigation of the number and localization of mast cells in the feline tongue.

This is the first study to describe the subtypes, number and distribution of mast cells (MC) in cat tongue by histochemical and immunohistochemical methods. Six male adult felines' tongue tissue samples consist of the study's material. Samples were fixed in 10% formaldehyde. MC number and distribution in the feline tongue were assessed using toluidine blue. Also, sections taken from blocks were stained in alcian blue/safranin O (AB/SO) combined dyes to determine the MC subtypes. The Streptavidin biotin complex method using anti-chymase and anti-tryptase primary antibodies was used for immunohistochemistry. Metachromatic MCs were mainly observed in the lamina propria close to the multilayered keratinized stratified squamous epithelium. The high number of MCs in this region may be because the dorsal surface of the tongue plays an essential role in the defence system of tongue tissue and, thus, of the body as a whole. Additionally, the number of MCs stained with AB (+) (1.7 ± 0.08) in the feline tongue was statistically higher than those with SO (+) (0.18 ± 0.02). This might be interpreted as an indication that MC heterogeneity may be due not only to their staining properties but also to their localization. It is also conceivable that the high histamine content may be a factor in this. Tryptase-positive MCs were found in the loose connective tissue around blood vessels, between the glands, as solitary cells, or in groups of several cells. Chymase-positive MCs were observed more individually rather than in groups. Moreover, chymase-positive MCs were detected to be located in the filiform papillae subepithelial and in the blood vessels' immediate vicinity. Animals often lick themselves to clean themselves and promote healing. For this reason, it is very important to protect the tongue, which is in direct contact with the external environment, against foreign agents. Considering both the functional and protective properties of the tongue, we concluded that MCs may play a role in oral cavity immunity and protective effect.

Involvement of Mast Cells in the Pathology of COVID-19: Clinical and Laboratory Parallels.

Recent studies suggested the potential role of mast cells (MCs) in the pathology of coronavirus disease 2019 (COVID-19). However, the precise description of the MCs' activation and the engagement of their proteases is still missing. The objective of this study was to further reveal the importance of MCs and their proteases (chymase, tryptase, and carboxypeptidase A3 (CPA3)) in the development of lung damage in patients with COVID-19. This study included 55 patients who died from COVID-19 and 30 controls who died from external causes. A histological analysis of the lung parenchyma was carried out to assess the protease profiles and degranulation activity of MCs. In addition, we have analyzed the general blood test, coagulogram, and C-reactive protein. The content of tryptase-positive MCs (Try-MCs) in the lungs of patients with COVID-19 was higher than in controls, but their degranulation activity was lower. The indicators of chymase-positive MCs (Chy-MCs) were significantly lower than in the controls, while the content of CPA3-positive MCs (CPA3-MCs) and their degranulation activity were higher in patients with COVID-19. In addition, we have demonstrated the existence of correlations (positive/negative) between the content of Try-MCs, Chy-MCs, and CPA3-MCs at different states of their degranulation and presence (co-adjacent/single) and the levels of various immune cells (neutrophils, eosinophils, basophils, and monocytes) and other important markers (blood hemoglobin, activated partial thromboplastin time (aPTT), international normalized ratio (INR), and fibrinogen). Thus, the identified patterns suggest the numerous and diverse mechanisms of the participation of MCs and their proteases in the pathogenesis of COVID-19, and their impact on the inflammatory process and coagulation status. At the same time, the issue requires further study in larger cohorts of patients, which will open up the possibility of using drugs acting on this link of pathogenesis to treat lung damage in patients with COVID-19.

Diagnostic and prognostic significance of mast cell markers in HIV/AIDS: Current insights and future directions.

Human immunodeficiency virus (HIV) infection continues to pose significant global health challenges, necessitating advancements in diagnostic and prognostic approaches to optimize disease management. While primarily recognized for their roles in allergic responses, mast cells have emerged as potential markers with diagnostic and prognostic significance in the context of HIV/AIDS. This paper aims to synthesize current insights and delineate future directions regarding the utility of mast cell markers in diagnosing HIV infection, predicting disease progression, and guiding therapeutic strategies. Mast cells, equipped with distinct markers such as tryptase, chymase, carboxypeptidase A3, and c-kit/CD117 receptors, exhibit tissue-specific expression patterns that offer potential as diagnostic indicators for HIV infection. Understanding the dynamics of these markers in different tissues and body fluids holds promise for accurate HIV diagnosis, disease staging, and monitoring treatment responses. Moreover, the prognostic significance of mast cell markers in HIV/AIDS lies in their potential to predict disease progression, immune dysregulation, and clinical outcomes. The integration of mast cell markers into clinical applications offers promising avenues for refining diagnostic assays, patient monitoring protocols, and therapeutic strategies in HIV/AIDS. Future research directions involve the development of novel diagnostic tools and targeted therapies based on mast cell-specific markers, potentially revolutionizing clinical practice and enhancing patient care in the management of HIV/AIDS. Continued investigations into mast cell markers' diagnostic and prognostic implications hold immense potential to advance our understanding and improve outcomes in HIV/AIDS management.

Vagus nerve stimulation (VNS) inhibits cardiac mast cells activation and improves myocardial atrophy after ischemic stroke.

BACKGROUND: Ischemic stroke is one of the leading causes of chronic disability worldwide, and stroke-induced heart damage can lead to death. According to research, patients with a variety of brain disease have good clinical results after vagus nerve stimulation (VNS). After ischemic stroke, mast cells (MCs) degranulate and release a large number of mediators, which may cause systemic inflammation. Chymase secreted by MCs can increase the levels of pathological angiotensin II (AngⅡ), which plays a crucial role in the deterioration of heart disease. Our goal was to develop a minimally invasive, targeted, and convenient VNS approach to assess the impact of VNS and to clarify the relationship between VNS and MCs in the prognosis of patients with myocardial atrophy after acute ischemic stroke. METHODS: In this study, we verified the role of VNS in the treatment of myocardial atrophy after stroke and its molecular mechanism using a rat model of middle cerebral artery occlusion (MCAO/r). Behavioral studies were assessed using neurobehavioral deficit scores. Enzyme-linked immunosorbent assays, immunofluorescence staining, Western blotting and qRT-PCR were used to analyze the expression levels of myocardial atrophy, MC and inflammatory markers in rat hearts. RESULTS: VNS improved myocardial atrophy in MCAO/r rats, inhibited MC activation, reduced the expression of chymase and AngⅡ, and inhibited the expression of proinflammatory factors. The chymase activator C48/80 reversed these effects of VNS. Chymase activation inhibited the effect of VNS on myocardial atrophy in MCAO/r rats, increased AngⅡ expression and aggravated inflammation and autophagy. The myocardial atrophy of MCAO/r rats was improved after chymase inhibition, and AngⅡ expression, inflammation and autophagy were reduced. Our results suggest that VNS may reduce the expression of chymase and AngⅡ by inhibiting MC activation, thereby improving myocardial atrophy and reducing inflammation and autophagy in MCAO/r rats. Inhibition of MC activation may be an effective strategy for treating myocardial atrophy after stroke. CONCLUSIONS: VNS inhibits MC activation and reduces the expression of chymase and AngII, thereby alleviating myocardial atrophy, inflammation and autophagy after stroke.

Oral administration of egg ovalbumin allergen induces dysregulation of tryptophan metabolism in sensitized BALB/c mice.

Food allergy (FA), triggered by specific dietary allergens, has emerged as a substantial global concern for food safety and public health. While studies have elucidated changes in immune cells and cytokines associated with allergen exposure, a comprehensive analysis of the host's metabolic features and the interaction between metabolites and the gut microbiota has not been conducted. In this study, egg allergen ovalbumin (OVA) was administered by the oral route to sensitized BALB/c mice to faithfully replicate key aspects of human FA, including severe allergic diarrhea, mast cell infiltration, and elevated levels of serum IgE, mMCPT-1, and Th2 cell hallmark cytokines (such as IL-4, IL-5, and IL-13). Furthermore, the untargeted and targeted metabolomic analyses indicated that FA in mice precipitated a substantial decrease in the tryptophan metabolites indole-3-acrylic acid (IA) and indole-3-lactic acid (ILA). The integration of shotgun metagenome and metabolome data further unveiled that the dysregulation of indole metabolism is related to a decline in the abundance of beneficial bacteria such as Lactobacillus and Bifidobacterium. Additionally, disruption of the tryptophan indole derivative pathway compromises the maintenance of intestinal mucosal function through the AHR signaling pathway, manifested by decreased expression of Reg3g and IL22. Taken together, this study demonstrated that the anaphylaxis triggered by oral ingestion of food allergens can lead to disruptions in tryptophan metabolism, consequently impairing intestinal immune homeostasis.

Mast Cell Infiltration and Subtype Promote Malignant Transformation of Oral Precancer and Progression of Oral Cancer.

The role of mast cell (MC), a common myeloid-derived immune cell, in the development of oral squamous cell carcinoma (OSCC) is unclear. The aim of this study was to investigate MC infiltration in oral precancer and oral cancer. The evaluation of immune cell infiltration and its association with prognosis in OSCC used RNA sequencing and multiple public datasets. Multiplex immunofluorescence was used to explore the infiltration of MC in the microenvironment of OSCC and oral precancer and the interaction with CD8+ cells. The role of MC in OSCC progression was verified by in vivo experiments. The resting MC infiltration was mainly present in oral precancer, whereas activated MC infiltration was significantly higher in OSCC. Activated MC was associated with malignant transformation of oral precancer and poor prognosis of OSCC. In vivo studies showed that MC promoted the growth of OSCC. The infiltration of activated MC was negatively correlated with the infiltration of CD8+ T cells. The subtype of MC containing tryptase without chymase (MCT) was significantly higher in OSCC compared with oral precancer and was associated with poor survival. Furthermore, spatial distance analysis revealed a greater distance between MCT and CD8+ cells, which was also linked to poor prognosis in OSCC. Cox regression analysis showed that MCT could be a potential diagnostic and prognostic biomarker. This study provides new insights into the role of MC in the immune microenvironment of OSCC. It might enhance the immunotherapeutic efficacy of OSCC by developing targeted therapies against MC. SIGNIFICANCE: In this study, we investigated the role of mast cells (MC) in oral precancer and oral cancer and demonstrated that MCs are involved in oral cancer progression and may serve as a potential diagnostic and prognostic marker. It might improve the immunotherapeutic efficacy through developing targeted therapies against MCs.

Reducing the Allergenicity of ß-Lactoglobulin by Covalent Modification with Different Contents of Epigallocatechin Gallate (EGCG): In Vitro and In Vivo Studies.

ß-Lactoglobulin (ßLG) is a major allergen in bovine milk protein. This study was designed to investigate changes in ßLG structure, digestibility, and allergenicity induced by covalent binding modification with different contents of (-)-epigallocatechin 3-gallate (EGCG). The reaction of EGCG conjugation with ßLG reached saturation at a molar ratio of 1:60 ßLG:EGCG. Conjugation with EGCG altered the ßLG structure, decreased IgE-binding capacity, and increased digestibility in a dose-dependent manner. In vivo studies showed that covalent conjugation with EGCG can reduce ßLG-induced allergic symptoms with reducing levels of IgE, histamine, and mast cell protease-1 (mMCP-1) and the percentage of sensitized mast cells. Allergenicity was reduced more effectively in saturated ßLG-EGCG conjugates compared to semisaturated conjugates. Observed changes in IFN-γ, IL-4, IL-5, IL-10, and TGF-ß levels suggested that ßLG-EGCG conjugates were able to promote Th1/Th2 immune balance. These findings further our understanding of the relationship between the degree of polyphenol conjugation and the allergenicity of food allergens.

Enzyme-linked immunosorbent assay and immunohistochemical analysis of mast cell related biochemicals in oral submucous fibrosis.

Oral submucous fibrosis (OSMF), a potentially malignant disorder, is developed by progressive fibrous tissue deposition in connective tissue along with atrophy of oral mucosa. Histological sections also show the mast cell infiltration in submucosa which may indicate their possible role in this entity. Abundant availability of biochemicals in mast cells like histamine and serine proteases like chymase may be released and play specific pathways in the disease pathophysiology. Possibly, if the histamine release has some part to play, diamine oxidase may also be found to have a relationship as it metabolizes histamine. The present study is proposed to identify the presence of chymase, histamine, and diamine oxidase in both, serum as well as tissue by enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry (IHC) respectively. This study may provide probable insight into the mast cell-related chemicals and their association with OSMF.

Alternative pathways for angiotensin II generation in the cardiovascular system

The classical renin-angiotensin system (RAS) consists of enzymes and peptides that regulate blood pressure and electrolyte and fluid homeostasis. Angiotensin II (Ang II) is one of the most important and extensively studied components of the RAS. The beneficial effects of angiotensin converting enzyme (ACE) inhibitors in the treatment of hypertension and heart failure, among other diseases, are well known. However, it has been reported that patients chronically treated with effective doses of these inhibitors do not show suppression of Ang II formation, suggesting the involvement of pathways alternative to ACE in the generation of Ang II. Moreover, the finding that the concentration of Ang II is preserved in the kidney, heart and lungs of mice with an ACE deletion indicates the important role of alternative pathways under basal conditions to maintain the levels of Ang II. Our group has characterized the serine protease elastase-2 as an alternative pathway for Ang II generation from Ang I in rats. A role for elastase-2 in the cardiovascular system was suggested by studies performed in heart and conductance and resistance vessels of normotensive and spontaneously hypertensive rats. This mini-review will highlight the pharmacological aspects of the RAS, emphasizing the role of elastase-2, an alternative pathway for Ang II generation.