Mast Cell Granules Research Articles

Neutrophil serine protease 4 is required for mast cell-dependent vascular leakage

Vascular leakage, or edema, is a serious complication of acute allergic reactions. Vascular leakage is triggered by the release of histamine and serotonin from granules within tissue-resident mast cells. Here, we show that expression of Neutrophil Serine Protease 4 (NSP4) during the early stages of mast cell development regulates mast cell-mediated vascular leakage. In myeloid precursors, the granulocyte–macrophage progenitors (GMPs), loss of NSP4 results in the decrease of cellular levels of histamine, serotonin and heparin/heparan sulfate. Mast cells that are derived from NSP4-deficient GMPs have abnormal secretory granule morphology and a sustained reduction in histamine and serotonin levels. Consequently, in passive cutaneous anaphylaxis and acute arthritis models, mast cell-mediated vascular leakage in the skin and joints is substantially reduced in NSP4-deficient mice. Our findings reveal that NSP4 is required for the proper storage of vasoactive amines in mast cell granules, which impacts mast cell-dependent vascular leakage in mouse models of immune complex-mediated diseases.

Development of a specific immunoassay to selectively measure active tryptase in airway samples.

Aim: Tryptase is a tetrameric trypsin-like serine protease contained within the secretory granules of mast cells and is an important mediator of allergic inflammatory responses in respiratory diseases. Detection of active tryptase in the airway may provide important information about asthma and other respiratory diseases. Materials & Methods: An activity based probe has been incorported within an immunoassay to allow for measurement of active tryptase in human tissues. Results: A specific Simoa immunoassay to measure active tryptase in nasosorption samples was developed and qualified using an activity-based probe label and a specific antitryptase capture antibody. Conclusion: The assay was capable of measuring active tryptase in human samples, which will enable evaluation of the role of tryptase proteolytic activity in human disease.

Differential expression of mast cell granules in samples of metastatic and non-metastatic colorectal cancer in patients

Various cell types participate in the tumor process, in which the mast cells have been described; however, the role they play in colorectal adenocarcinoma has not yet been fully understood. Therefore, the present work aimed to compare employing histochemistry and immunohistochemistry, the number of mast cells and the content of some cytoplasmic granules in moderately differentiated non-metastatic and metastatic colorectal adenocarcinoma, analyzing tissue samples from patients. Histochemical techniques with Toluidine Blue (TBO), Periodic Schiff Acid (PAS), Alcian Blue/Periodic Acid-Schiff (PAB) and Alcian Blue/Safranin (ABS); as well as immunohistochemical reactions with anti-antibodies anti-Tryptase and anti-Chymase were applied to quantify total mast cells and content of some cytoplasmic granules. Statistical analysis was performed using SPSS V22.0 software (p ≤ 0.05). The degree of positivity of the reaction and degranulation of mast cells was reported in percentages. In our results, we observed that there are differences in the quantity and histochemical composition of the granules of mast cells (metastatic group PAS and ABS comparing the TBO reaction), as well as in the immunohistochemical composition between Tryptase and Chymase and the number of degranulated cells in both study groups (74 % degranulated mast cells in the metastatic group, 66 % integrate mast cells in the non-metastatic group). Therefore, we consider that the differences may be some of the probable factors that lead to metastasis of colorectal adenocarcinoma.

Directional Mast Cell Degranulation of TNF into Blood Vessels Primes Neutrophil Extravasation

Tissue resident mast cells (MCs) rapidly initiate neutrophil infiltration upon inflammatory insult, yet the molecular mechanism is still unknown. Here, we demonstrate that MC-derived TNF is crucial for neutrophil extravasation to sites of sterile skin inflammation by promoting intraluminal adhesion and crawling. Strikingly, MC-TNF directly primes circulating neutrophils via TNFR1, while being dispensable for chemokine production and endothelial cell activation. The MC-TNF is fused into the bloodstream by directional degranulation of perivascular MCs that are part of the vascular unit with access to the vessel lumen. Consistently, intravenous administration of MC granules can boost neutrophil extravasation. Consequently, the directional MC degranulation of pro-inflammatory mediators into the bloodstream may represent an important target for therapeutic approaches aimed at dampening cytokine storm syndromes or shock symptoms, or intentionally pushing immune defense.

Mast Cell β-Tryptase Is Enzymatically Stabilized by DNA.

Tryptase is a tetrameric serine protease located within the secretory granules of mast cells. In the secretory granules, tryptase is stored in complex with negatively charged heparin proteoglycans and it is known that heparin is essential for stabilizing the enzymatic activity of tryptase. However, recent findings suggest that enzymatically active tryptase also can be found in the nucleus of murine mast cells, but it is not known how the enzmatic activity of tryptase is maintained in the nuclear milieu. Here we hypothesized that tryptase, as well as being stabilized by heparin, can be stabilized by DNA, the rationale being that the anionic charge of DNA could potentially substitute for that of heparin to execute this function. Indeed, we showed that double-stranded DNA preserved the enzymatic activity of human β-tryptase with a similar efficiency as heparin. In contrast, single-stranded DNA did not have this capacity. We also demonstrated that DNA fragments down to 400 base pairs have tryptase-stabilizing effects equal to that of intact DNA. Further, we showed that DNA-stabilized tryptase was more efficient in degrading nuclear core histones than heparin-stabilized enzyme. Finally, we demonstrated that tryptase, similar to its nuclear localization in murine mast cells, is found within the nucleus of primary human skin mast cells. Altogether, these finding reveal a hitherto unknown mechanism for the stabilization of mast cell tryptase, and these findings can have an important impact on our understanding of how tryptase regulates nuclear events.

Mast Cell Tryptase Promotes Inflammatory Bowel Disease-Induced Intestinal Fibrosis.

Intestinal fibrosis is the final pathological outcome of chronic intestinal inflammation without specific therapeutic drugs, which leads to ileus and surgical intervention. Intestinal fibrosis is characterized by excessive deposition of extracellular matrix (ECM). The role of mast cells (MCs), which are members of the sentinel immune cell population, is unknown in intestinal fibrosis. In this study, we analyzed changes in MCs, tryptase proteins, and ECM components in human fibrotic and control patient intestines. We constructed dextran sodium sulfate-induced intestinal fibrosis models using wild-type mice, MC-reconstituted mice, and MC-deficient mice to explore the role of MCs and tryptase in intestinal fibrosis. The roles and mechanisms of MCs and tryptase on fibroblasts were evaluated using human MCs (HMC-1 and LAD-2), commercial tryptase proteins, human colon fibroblasts (CCD-18Co fibroblasts), the tryptase inhibitor APC366, and the protease-activated receptor-2 (PAR-2) antagonist ENMD-1068. Regardless of whether the colon was a human colon or a mouse colon, the fibrotic intestinal tissue had increased MC infiltration and a higher expression of ECM proteins or genes than that of the control group. The dextran sodium sulfate-induced intestinal fibrosis in MC-deficient mice was alleviated compared with that in wild-type mice. After MC reconstruction in MC-deficient mice, the alleviating effect disappeared. Tryptase, as a content stored in MC granules, was released into fibrotic intestinal tissues in the form of degranulation, resulting in an increased expression of tryptase. Compared with the control group, the tryptase inhibition group (the APC366 group) had reduced intestinal fibrosis. The CCD-18Co fibroblasts, when cocultured with MCs or treated with tryptase proteins, were activated to differentiate into myofibroblasts and secrete more ECM proteins (such as collagen and fibronectin). The underlying mechanism of fibroblast activation by tryptase was the activation of the PAR-2/Akt/mTOR pathway. We found that MC tryptase promotes inflammatory bowel disease-induced intestinal fibrosis. The underlying mechanism is that tryptase promotes the differentiation of fibroblasts into fibrotic-phenotype myofibroblasts by activating the PAR-2/Akt/ mTOR pathway of fibroblasts.

Mast cell histamine mediates intestinal immune response to early life stress in piglets via histamine 1 receptor (H1R)

Abstract Mast cells are important immune modulators of stress-related gastrointestinal (GI) disorders. Our previous studies investigating early life stress in piglets showed that histamine, a major mast cell granule mediator is released within minutes of early weaning stress and is followed by increased intestinal mucosal expression of histamine receptor subtypes. The precise contribution of histamine to stress-induced GI immune responses is unknown. The objective of this study was to test the hypothesis that stress-induced intestinal immune response is mediated through the histamine 1 receptor (H1R). Fifteen-day-old female Yorkshire piglets were administered either saline vehicle or the H1R antagonist, desloratadine (2 mg/kg; intramuscular), 30 minutes prior to early weaning stress. At weaning, piglets were weaned from their dams and housed in nursery pens with ad libitum access to water. At 24 hours post-weaning, mid-jejunum and mesenteric lymph nodes (MLN) were collected for subsequent qPCR gene expression as well as IHC localization and protein expression analyses. Unweaned control piglets remained with the sow and were collected immediately at weaning. Gene expression for H1R was not influenced by weaning; however, immunohistochemical analysis of H1R revealed increased expression and localization in lamina propria cells and myofibroblasts in jejunal villi and expression in the longitudinal and circular muscle layers in early weaned piglets. In the MLN, weaning increased the expression of TNFα, TGFβ, and β-Integrin. This response was reduced in the group treated with the H1R antagonist. Together, these data demonstrate that histamine via H1R plays an important role in early weaning stress-induced intestinal immune responses.

Quantitative In-Depth Analysis of the Mouse Mast Cell Transcriptome Reveals Organ-Specific Mast Cell Heterogeneity.

Mast cells (MCs) are primarily resident hematopoietic tissue cells that are localized at external and internal surfaces of the body where they act in the first line of defense. MCs are found in all studied vertebrates and have also been identified in tunicates, an early chordate. To obtain a detailed insight into the biology of MCs, here we analyzed the transcriptome of MCs from different mouse organs by RNA-seq and PCR-based transcriptomics. We show that MCs at different tissue locations differ substantially in their levels of transcripts coding for the most abundant MC granule proteins, even within the connective tissue type, or mucosal MC niches. We also demonstrate that transcript levels for the major granule proteins, including the various MC-restricted proteases and the heparin core protein, can be several orders of magnitude higher than those coding for various surface receptors and enzymes involved in protease activation, as well as enzymes involved in the synthesis of heparin, histamine, leukotrienes, and prostaglandins. Interestingly, our analyses revealed an almost complete absence in MCs of transcripts coding for cytokines at baseline conditions, indicating that cytokines are primarily produced by activated MCs. Bone marrow-derived MCs (BMMCs) are often used as equivalents of tissue MCs. Here, we show that these cells differ substantially from tissue MCs with regard to their transcriptome. Notably, they showed a transcriptome indicative of relatively immature cells, both with respect to the expression of granule proteases and of various enzymes involved in the processing/synthesis of granule compounds, indicating that care should be taken when extrapolating findings from BMMCs to the in vivo function of tissue-resident MCs. Furthermore, the latter finding indicates that the development of fully mature tissue-resident MCs requires a cytokine milieu beyond what is needed for in vitro differentiation of BMMCs. Altogether, this study provides a comprehensive quantitative view of the transcriptome profile of MCs resident at different tissue locations that builds nicely on previous studies of both the mouse and human transcriptome, and form a solid base for future evolutionary studies of the role of MCs in vertebrate immunity.

Physiological responses to central and peripheral injections of compound 48/80 and histamine in chicks

Mast cells are a type of immune cell widely distributed in the body of vertebrates. Mast cells have many granules that contain several bioactive molecules such as histamine, and these molecules are released through degranulation when the mast cell receives certain stimuli. Because the number of mast cells increases during infection in chickens (Gallus gallus), the activity of mast cells might be related to non-specific symptoms such as anorexia under an infectious condition. Therefore, the purpose of the present study was to investigate whether intraperitoneal (IP) and intracerebroventricular (ICV) injections of compound 48/80, which induces degranulation of mast cells, affects feeding, voluntary activity, cloacal temperature, and the concentrations of plasma corticosterone (CORT) and glucose in chicks. The effect of histamine, which is found in mast cell granules, on these parameters was also investigated. IP injection of compound 48/80 significantly decreased food intake, voluntary activity, and cloacal temperature, and increased plasma CORT concentration in the chicks. While ICV injection of compound 48/80 also decreased food intake, it increased cloacal temperature and plasma glucose concentration. Both IP and ICV injections of histamine significantly decreased food intake, cloacal temperature, and plasma CORT concentration. However, only IP injection of histamine significantly decreased voluntary activity and increased plasma glucose concentration. The results suggest that degranulation of mast cells is related to non-specific symptoms in chicks, although the mechanism seems to be different between peripheral and central tissues. In addition, the effect of peripherally-injected compound 48/80 may be partly mediated by histamine.

Dr James Riley, pioneer in histamine and mast cell studies.

Dr James Riley, an honours graduate of Edinburgh University, had to give up a surgical career because of an affliction of his hands, and instead became a radiotherapist in Dundee. He had always been fascinated by past heroes of medical research, and set out to carve his own niche in the study of mast cells. In the early 1950s, no one knew what their function was other than the mast cell granules being the storehouses of heparin. Riley and pharmacologist Dr Geoffrey West went on to galvanise this area of study by showing in a notable series of experiments that the mast cell granules are also the main repository of histamine, a key participant in allergic and anaphylactic reactions. Riley's story is a prime example of the ability to make cutting-edge discoveries even in the face of modest facilities.

Exosome-mediated uptake of mast cell tryptase into the nucleus of melanoma cells: a novel axis for regulating tumor cell proliferation and gene expression

It is well established that mast cell accumulation accompanies most malignancies. However, the knowledge of how mast cells functionally impact on tumors is still rudimentary. Here we addressed this issue and show that mast cells have anti-proliferative activity on melanoma cells and that this effect is dependent on tryptase, a tetrameric protease stored in mast cell granules. Mechanistically, tryptase was found to be endocytosed by melanoma cells as cargo of DNA-coated exosomes released from melanoma cells, followed by transport to the nucleus. In the nucleus, tryptase executed clipping of histone 3 and degradation of Lamin B1, accompanied by extensive nuclear remodeling. Moreover, tryptase degraded hnRNP A2/B1, a protein involved in mRNA stabilization and interaction with non-coding RNAs. This was followed by downregulated expression of the oncogene EGR1 and of multiple non-coding RNAs, including oncogenic species. Altogether, these findings establish a new principle for regulation of tumor cell proliferation.

Endoglin is Highly Expressed in Human Mast Cells.

Endoglin, known to be expressed in proliferating vessels, is of worth when evaluating microvessel density as a prognostic factor in many types of malignancies, including some subtypes of leukemia cells. In childhood acute lymphoblastic leukemia, endoglin is associated with adverse outcome. In bone marrow, endoglin identifies the repopulating hematopoietic stem cells. Mast cells are a component of normal tissue and play an important role in the regulation of several processes, including inflammation and neoplasia. The aim of this study was to evaluate the use of endoglin as a biological marker of mast cells compared with the gold standard stains. We studied 15 specimens of neurofibroma, 9 of mastocytosis, and 6 of fibrous scar tissue through immunohistochemistry (for endoglin and mast cell tryptase) and histochemical staining using toluidine blue. Quantitative analysis of the cells was performed by counting 5 hotspots. The validity of endoglin as a mast cell marker was assessed by intraclass correlation coefficient. The Kruskal-Wallis test was used to compare mast cell count for each marker. A strong endoglin expression was found in the cytoplasmic granules of mast cells within the 3 groups. Similar results were observed with mast cell tryptase as well as toluidine blue. The intraclass correlation coefficient revealed that endoglin is a highly reliable biomarker of mast cells when compared with mast cell tryptase and toluidine blue. In conclusion, endoglin may assist in the diagnosis and pathogenesis study of various processes associated with mast cells. An endoglin-neutralizing treatment for solid cancers and leukemia could also affect mastocytes and the immunologic system.

Anti-degranulation activity of caffeine: In vitro and in vivo study

Abstract Based on some studies, moderate caffeine consumption provides several benefits to human health. However, there are few reports on the anti-degranulation activity of caffeine. Allergy is a global healthcare problem that is drastically increasing in both developed and developing countries. Meanwhile, allergic reactions are triggered by the presence of chemical mediators released from the granules in mast cells, which are called the degranulation process. In this study, caffeine was evaluated for anti-degranulation activity using basophilic cell line RBL-2H3 cells as the in vitro model and the passive cutaneous anaphylaxis (PCA) model mice in vivo. As a result, caffeine effectively suppressed antigen-induced degranulation by RBL-2H3 cells in a dose-dependent manner without cytotoxicity. Besides, caffeine also inhibited FceRI-mediated intracellular signaling pathways, such as phosphorylation of Syk, Btk, PLCγ1, PI3K, and Akt in antigen-stimulated RBL-2H3 cells. Caffeine also suppressed PCA response in mice.

Polymyxin B and polymyxin E induce anaphylactoid response through mediation of Mas-related G protein–coupled receptor X2

Polymyxin B (PMB) and polymyxin E (PME) are cyclic, peptide antibiotics which derived from various species of Paenibacillus (Bacillus) polymyxa. They are decapeptide antibiotics with an antimicrobial spectrum that includes Gram-negative bacteria, and reused as therapeutic agents due to the emergence of multidrug-resistant (MDR) Gram-positive bacteria. PMB or PME-induced anaphylactoid reactions in the clinic have been documented. However, the mechanism underlying anaphylactoid reaction induced by polymyxin has not yet been reported. Here, we report that human Mas-related G protein-coupled receptor X2 (MRGPRX2) and its mouse homologue Mas-related G protein-coupled receptor B2 (MrgprB2) are the receptors mediating the anaphylactoid response provoked by PMB and PME. We firstly investigated the anaphylactoid reactions induced by PMB and PME in LAD2 cells in vitro and in vivo, and found that treatment with PMB and PME led to significant release of mast cell granules such as histamine and β-hexosaminidase, secretion of pro-inflammatory cytokines, such as TNF-α and PGD2, and provocation of calcium flux in LAD2 cells. Furthermore, treatment with PMB and PME led to reduced release of β-hexosaminidase in MRGPRX2 knockdown-LAD2 cells, and obvious increased calcium release in MRGPRX2 overexpressing HEK293 cells, which suggested that MRGPRX2 are involved in mast cell activation provoked by PMB or PME. In vivo, MRGPRB2 knockout mice exhibited lower pseudo-allergic reactions than wild type mice. Activation of MrgprB2 also triggers increased capillary permeability and paw swelling. Our results elucidated the role of MRGPRX2 in PMB and PME-induced anaphylactoid response and suggested that MRGPRX2 as a potential therapeutic target to control the anaphylactoid reactions which triggered by PMB or PME.

Electron microscopic characteristic of local immune processes during regression of developed experimental tumors under the influence of magnetite nanoparticles.

e14179 Background: Previously, regression of the developed transplanted tumors under the influence of magnetite nanoparticles (NPs) as monofactor was shown. The purpose of the study was to reveal signs of intercellular interactions in the zone of tumors regressed under the influence of magnetite NPs. Methods: Experiments were carried out on 14 white outbred male rats (180-200 g) with transplanted sarcoma 45 of an initial volume of 0.7-1.3 cm3. After 6 peritumoral injections of magnetite NPs (10 ± 2 nm) in the form of magnetic fluid AM-01 ("AM-Kub", Ekaterinburg) twice a week in a single dose of 17.7 mg/kg, 5 out of 7 rats of the main group demonstrated a regression of the tumor from 1.5-3 cm3 to 0.7 ± 0.2 cm3 (in the control group – 9.7 ± 1.9 cm3, p < 0.01). Ultrathin sections of the tumor tissue of 12 animals (including 7 rats of the main group) were examined in electron microscope JEOL JEM-1011 (Japan). Results: Under effective action of magnetite NPs in tumor tissue, in addition to necrosis, apoptosis and autophagy were marked. In the case of apoptosis, chromatin condensation in compact masses and separation of apoptotic bodies were observed. Significantly more common autophagic cell death was identified by violation of the integrity of the cell membrane and the presence of autophagosomes. Numerous signs of activation of cell-cell interactions with participation of immune cells were observed. Diverse groups of 2-4 contacting cells included macrophages, lymphocytes, plasmocytes, degranulated mast cells, neutrophils in various combinations with distinct signs of the metabolic activity. Invagination of macrophages cytoplasm into the cytoplasm of tumor cells, close adjacency of the cytoplasmic membranes of neutrophils and lymphocytes at considerable length, tight intertwining of mast cell cytoplasmic processes and engulfing of the mast cell granules by macrophages were noted. Conclusions: The results supplement the characteristics of immune processes during the self-dependent antitumor action of magnetite NPs.

Mast cell involvement in human cervical ripening

ObjectiveCervical ripening resembles an inflammatory process in many aspects, involving invasion of inflammatory cells, collagen breakdown and remodelling of the extracellular matrix. Mast cells produce a variety of inflammatory agents and are attributed a functional role in cervical ripening. The aim of this study was to examine if cervical mast cells are increased in number and stimulated during pregnancy. Study designCervical biopsies were obtained with a biopsy needle prior to surgical termination of pregnancy in the first trimester, surgery for first-trimester miscarriage, elective caesarean section, and benign gynaecological surgery in non-pregnant women. After fixation, semithin sections were prepared and stained with toluidine blue. The number of mast cells was counted under a light microscope and their secretory activity was scored (0.5–4) according to specified criteria and further visualised with electron microscopy. For pairwise comparison between groups Fisher’s nonparametric permutation test was used. ResultsThe number of mast cells was increased from 3.4 ± 1.65 mast cells per 10 visual fields in non-pregnant women to 7.70 ± 0.35 per 10 visual fields in first trimester control women (p < 0.05). The highest number of mast cells was observed at term with 10.8 ± 2.1 per 10 visual fields, a number that was significantly higher than in first trimester control women (p < 0.05). At term mast cell activity scores were 3.39 ± 0.37 compared with 2.69 ± 0.27 in control first trimester women and 2.21 ± 0.86 in women with missed miscarriage (p < 0.05). The percentage of mast cells with activity score 4 was significantly higher at term compared with in the first trimester. Free mast cell granules were predominantly observed in areas with disorganized collagen fibres. ConclusionThe findings confirm that an increased influx of mast cells to the cervix occurs during pregnancy. The stimulated mast cell secretory activity in conditions associated with cervical tissue remodelling, such as term pregnancy and symptomatic miscarriage, provides further evidence that mast cells play a physiological role in cervical ripening.

Dietary cholesterol is essential to mast cell activation and associated obesity and diabetes in mice

Mast cell (MC) deficiency in KitW-sh/W-sh mice and inhibition with disodium chromoglycate (DSCG) or ketotifen reduced obesity and diabetes in mice on a high-cholesterol (1.25%) Western diet. Yet, Kit-independent MC-deficient mice and mice treated with DSCG disproved MC function in obesity and diabetes when mice are fed a high-fat diet (HFD) that contains no cholesterol. This study reproduced the obesity and diabetes inhibitory activities of DSCG and ketotifen from mice on a Western diet. Yet, such inhibitory effects were diminished in mice on the HFD. DSCG and ketotifen MC inhibitory activities were recovered from mice on the HFD supplemented with the same amount of cholesterol (1.25%) as that in the Western diet. DSCG and ketotifen effectively blunted the high-cholesterol diet-induced elevations of blood histamine and adipose tissue MC degranulation. Pearson's correlation test demonstrated significant and positive correlations between plasma histamine and total cholesterol or low-density lipoprotein-cholesterol (LDL). In cultured bone marrow-derived MCs, plasma from mice following a Western diet or a cholesterol-supplemented HFD, but not those from HFD-fed mice, induced MC degranulation and the release of β-hexosaminidase, histamine, and serotonin. IgE, LDL, very low-density lipoprotein, and high-density lipoprotein also induced MC activation, which can be inhibited by DSCG and ketotifen depending on the doses and types of MC inhibitors and cholesterol, and also the MC granule molecules of interest. DSCG or ketotifen lost their activities in inhibiting LDL-induced activation of MCs from LDL receptor-deficient mice. These results indicate that dietary cholesterol critically influences the function of mouse MCs.

Structure-based virtual screening for novel chymase inhibitors by in silico fragment mapping

The term chymase refers to a family of chymotrypsin-like serine proteases stored within the secretory granules of mast cells. Recently, a variety of small molecule inhibitors for chymase have been developed with a primary focus on the treatment of cardiovascular diseases. Despite the expected therapeutic benefit of these chymase inhibitors, they have not been used clinically. Here, we attempted to identify new chymase inhibitors using a multistep structure-based virtual screening protocol combined with our knowledge-based in silico fragment mapping technique. The mapping procedure identified fragments with novel modes of interaction at the oxyanion hole of chymase. Next, we constructed a three-dimensional (3D) pharmacophore model and retrieved eight candidate chymase inhibitors from a commercial database that included approximately five million compounds. This selection was achieved using a multistep virtual screening protocol, which combined a 3D pharmacophore-based search, docking calculations, and analyses of binding free energy. One of the eight compounds exhibited concentration-dependent chymase inhibitory activity, which could be further optimized to develop more potent chymase inhibitors.

Esculetin을 담지한 ZnO microsphere 외용제의 광노화 차단 상가 효능

This study was aimed to examine the synergetic effects of photochemopreventive external agents composed of inorganic ZnO and esculetin. Zinc oxide (ZnO) is widely used in sunblocks because of its excellent biostability and little toxicity. Esculetin derived from Citrus Peel has an anti-oxidative effect. We made the hollow-shaped ZnO microsphere (MS), soaked it with esculetin (EZnO). We used SKH-1 mice to measure the photodamaging effects of UVB. The mice were divided into five groups as follows; UVB nontreated group (N), vehicle (C), esculetin (E), ZnO MS (ZnO), esculetin + ZnO MS(EZno) group. Each group of samples was topically applied to the dorsal skins before the UVB irradiation. The changes of collagen fibers in the skin tissues were observed by H & E staining and Van Gieson staining. The expression of mast cells in skin tissue was observed by immunohistochemical staining of tryptase present in the mast cell granules. Expression of inflammatory cytokines such as TNF-α, IL-1β, and MMP-9, which plays an essential role in wrinkle formation, was measured by RT-PCR. Interestingly, the composition of collagen fibers was better in the EZnO applied group than in the E or ZnO group. Moreover, mast cell expression and the expression of TNF-α, IL-1β, and MMP-9 mRNA were markedly suppressed in the EZnO group, indicating that the synergetic effects of esculetin and ZnO were excellent.

Characterization of mast cell-derived rRNA-containing microvesicles and their inflammatory impact on endothelial cells.

Tissue-resident mast cells (MCs) are well known for their role in inflammatory responses and allergic and anaphylactic reactions, but they also contribute to processes of arterial remodeling. Although ribosomes and cytosolic RNAs are located around secretory granules in mature MCs, their functional role in MC responses remains unexplored. Previous studies by our group characterized extracellular RNA (eRNA) as an inflammatory and pathogenetic factor in vitro and in vivo. In the present study, RNA-containing MCs and eRNA were located in close proximity to growing collateral arteries in vivo. In vitro, various agonists were found to induce the degranulation of MCs and the concomitant release of eRNA in association with microvesicles (MVs). The liberation of eRNA from MCs was abolished by MC stabilizers or by preventing the increase of intracellular Ca2+ in MCs. eRNA was found to be mainly contained inside MVs, as demonstrated by electron microscopy and immunocytochemistry. The exposure to and the uptake of MC-released MVs by cultured endothelial cells increased their expression of cytokines, such as monocyte chemoattractant protein or IL-6, in a dose- and time-dependent manner. These results indicate that RNA-containing MC-derived MVs are likely to be involved in inflammatory responses, relevant, for example, to processes of vascular remodeling.-Elsemüller, A.-K., Tomalla, V., Gärtner, U., Troidl, K., Jeratsch, S., Graumann, J., Baal, N., Hackstein, H., Lasch, M., Deindl, E., Preissner, K. T., Fischer, S. Characterization of mast cell-derived rRNA-containing microvesicles and their inflammatory impact on endothelial cells.