| 1. |
- Che, Karlhans F., et al.
(författare)
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Complex Involvement of Interleukin-26 in Bacterial Lung Infection
- 2021
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Ingår i: Frontiers in Immunology. - : Frontiers Media SA. - 1664-3224. ; 12, s. 1-18
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Tidskriftsartikel (refereegranskat)abstract
- Pneumonia is a global cause of mortality, and this provides a strong incentive to improve the mechanistic understanding of innate immune responses in the lungs. Here, we characterized the involvement of the cytokine interleukin (IL)-26 in bacterial lung infection. We observed markedly increased concentrations of IL-26 in lower airway samples from patients with bacterial pneumonia and these correlated with blood neutrophil concentrations. Moreover, pathogen-associated molecular patterns (PAMPs) from both Gram-negative and -positive bacteria increased extracellular IL-26 concentrations in conditioned media from human models of alveolar epithelial cells, macrophages, and neutrophils in vitro. Stimulation with IL-26 inhibited the inherent release of neutrophil elastase and myeloperoxidase in unexposed neutrophils. This stimulation also inhibited the expression of activity makers in neutrophils exposed to Klebsiella pneumoniae. In addition, priming of human lung tissue ex vivo with exogenous IL-26 potentiated the endotoxin-induced increase in mRNA for other cytokines involved in the innate immune response, including the master Th17-regulator IL-23 and the archetype inhibitory cytokine IL-10. Finally, neutralization of endogenous IL-26 clearly increased the growth of Klebsiella pneumoniae in the macrophage culture. These findings suggest that IL-26 is involved in bacterial lung infection in a complex manner, by modulating critical aspects of innate immune responses locally and systemically in a seemingly purposeful manner and by contributing to the killing of bacteria in a way that resembles an antimicrobial peptide. Thus, IL-26 displays both diagnostic and therapeutic potential in pneumonia and deserves to be further evaluated in these respects.
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| 2. |
- Johnsson, Anna-Karin, et al.
(författare)
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COX-1 dependent biosynthesis of 15-hydroxyeicosatetraenoic acid in human mast cells
- 2021
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Ingår i: Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids. - : Elsevier. - 1388-1981 .- 1879-2618. ; 1866:5
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Tidskriftsartikel (refereegranskat)abstract
- 15-hydroxyeicosatetraenoic acid (15-HETE) is an arachidonic acid derived lipid mediator which can originate both from 15-lipoxygenase (15-LOX) activity and cyclooxygenase (COX) activity. The enzymatic source determines the enantiomeric profile of the 15-HETE formed. 15-HETE is the most abundant arachidonic acid metabolite in the human lung and has been suggested to influence the pathophysiology of asthma. Mast cells are central effectors in asthma, but there are contradictory reports on whether 15-HETE originates from 15-LOX or COX in human mast cells. This prompted the current study where the pathway of 15-HETE biosynthesis was examined in three human mast cell models; the cell line LAD2, cord blood derived mast cells (CBMC) and tissue isolated human lung mast cells (HLMC). Levels and enantiomeric profiles of 15-HETE and levels of the downstream metabolite 15-KETE, were analyzed by UPLC-MS/MS after stimulation with anti-IgE or calcium ionophore A23187 in the presence and absence of inhibitors of COX isoenzymes. We found that 15-HETE was produced by COX-1 in human mast cells under these experimental conditions. Unexpectedly, chiral analysis showed that the 15(R) isomer was predominant and gradually accumulated, whereas the 15(S) isomer was metabolized by the 15hydroxyprostaglandin dehydrogenase. We conclude that during physiological conditions, i.e., without addition of exogenous arachidonic acid, both enantiomers of 15-HETE are produced by COX-1 in human mast cells but that the 15(S) isomer is selectively depleted by undergoing further metabolism. The study highlights that 15-HETE cannot be used as an indicator of 15-LOX activity for cellular studies, unless chirality and sensitivity to pharmacologic inhibition is determined.
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| 3. |
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| 4. |
- Liu, Jielu, et al.
(författare)
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Monensin inhibits mast cell mediated airway contractions in human and guinea pig asthma models
- 2022
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Ingår i: Scientific Reports. - : Springer Nature. - 2045-2322. ; 12
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Tidskriftsartikel (refereegranskat)abstract
- Asthma is a common respiratory disease associated with airway hyperresponsiveness (AHR), airway inflammation and mast cell (MC) accumulation in the lung. Monensin, an ionophoric antibiotic, has been shown to induce apoptosis of human MCs. The aim of this study was to define the effect of monensin on MC responses, e.g., antigen induced bronchoconstriction, and on asthmatic features in models of allergic asthma. Tracheal segments from house dust mite (HDM) extract sensitized guinea pigs were isolated and exposed to monensin, followed by histological staining to quantify MCs. Both guinea pig tracheal and human bronchi were used for pharmacological studies in tissue bath systems to investigate the monensin effect on tissue viability and antigen induced bronchoconstriction. Further, an HDM-induced guinea pig asthma model was utilized to investigate the effect of monensin on AHR and airway inflammation. Monensin decreased MC number, caused MC death, and blocked the HDM or anti-IgE induced bronchoconstriction in guinea pig and human airways. In the guinea pig asthma model, HDM-induced AHR, airway inflammation and MC hyperplasia could be inhibited by repeated administration of monensin. This study indicates that monensin is an effective tool to reduce MC number and MCs are crucial for the development of asthma-like features.
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| 5. |
- Manson, Martijn L, et al.
(författare)
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Bitter taste receptor agonists mediate relaxation of human and rodent vascular smooth muscle.
- 2014
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Ingår i: European Journal of Pharmacology. - : Elsevier BV. - 1879-0712 .- 0014-2999. ; 740:Jul 15, s. 302-311
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Tidskriftsartikel (refereegranskat)abstract
- Taste-sensing type 2 receptors (TAS2Rs) have been implicated in extraoral functions. Airway smooth muscle expresses TAS2Rs and is strongly relaxed by TAS2R agonists. We hypothesised that TAS2R agonists might affect vascular smooth muscle as well. Moreover, the general pharmacological profile of TAS2R agonists, which are used to investigate the functions of TAS2R׳s, are undefined. The aim of this study was to pharmacologically characterise the effects of five prototype TAS2R agonists in vascular smooth muscle. Responses to the TAS2R agonists were investigated in guinea-pig aorta and taenia coli, mouse aorta (wild-type and caveolin-1(-/-) mice) and human pulmonary arteries. Chloroquine, denatonium, dextromethorphan, noscapine and quinine, agonists for TAS2R3, TAS2R4, TAS2R10 and TAS2R14, induced strong endothelium-independent relaxations (responses between 82-96% of maximal relaxations) in phenylephrine pre-contracted guinea-pig aorta that persisted in the presence of L-type Ca(2+) and KCa1.1-channel blockers. Experiments in guinea-pig taenia coli revealed that denatonium and quinine also inhibited relaxations to phenylephrine, indicating antagonism of α-adrenoceptors. Only chloroquine and noscapine mediated relaxations when the guinea pig aorta was pre-contracted by U-46619 or PGF2α. Relaxations to chloroquine and noscapine after U-46619 pre-contractions were however markedly impaired in aortae from caveolin-1(-/-) mice. Chloroquine and noscapine mediated relaxations of human pulmonary arteries that expressed also mRNA for TAS2R3, TAS2R4, TAS2R10 and TAS2R14, at levels similar to that of the α1A adrenoceptor. Notwithstanding whether TAS2Rs are involved or not, TAS2R agonists have profound effects on vascular smooth muscle. Chloroquine and noscapine are of special interest as their effects cannot be accounted for by conventional pathways.
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| 6. |
- Mazzurana, Luca, et al.
(författare)
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Tissue-specific transcriptional imprinting and heterogeneity in human innate lymphoid cells revealed by full-length single-cell RNA-sequencing
- 2021
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Ingår i: Cell Research. - : Springer Science and Business Media LLC. - 1748-7838 .- 1001-0602. ; 31:5, s. 554-568
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Tidskriftsartikel (refereegranskat)abstract
- The impact of the microenvironment on innate lymphoid cell (ILC)-mediated immunity in humans remains largely unknown. Here we used full-length Smart-seq2 single-cell RNA-sequencing to unravel tissue-specific transcriptional profiles and heterogeneity of CD127+ ILCs across four human tissues. Correlation analysis identified gene modules characterizing the migratory properties of tonsil and blood ILCs, and signatures of tissue-residency, activation and modified metabolism in colon and lung ILCs. Trajectory analysis revealed potential differentiation pathways from circulating and tissue-resident naïve ILCs to a spectrum of mature ILC subsets. In the lung we identified both CRTH2+ and CRTH2− ILC2 with lung-specific signatures, which could be recapitulated by alarmin-exposure of circulating ILC2. Finally, we describe unique TCR-V(D)J-rearrangement patterns of blood ILC1-like cells, revealing a subset of potentially immature ILCs with TCR-δ rearrangement. Our study provides a useful resource for in-depth understanding of ILC-mediated immunity in humans, with implications for disease.
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| 7. |
- Ravindran, Avinash, et al.
(författare)
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An Optimized Protocol for the Isolation and Functional Analysis of Human Lung Mast Cells
- 2018
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Ingår i: Frontiers in Immunology. - : FRONTIERS MEDIA SA. - 1664-3224. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Background: Mast cells are tissue-resident inflammatory cells defined by their high granularity and surface expression of the high-affinity IgE receptor, Fc + RI, and CD117/KIT, the receptor for stem cell factor (SCF). There is a considerable heterogeneity among mast cells, both phenotypically and functionally. Human mast cells are generally divided into two main subtypes based on their protease content; the mucosa-associated MCT (tryptase positive and chymase negative mast cell) and the connective tissue associated-residing MCTC (tryptase and chymase positive mast cell). Human lung mast cells exhibit heterogeneity in terms of cellular size, expression of cell surface receptors, and secreted mediators. However, knowledge about human lung mast cell heterogeneity is restricted to studies using immunohistochemistry or purified mast cells. Whereas the former is limited by the number of cellular markers that can be analyzed simultaneously, the latter suffers from issues related to cell yield.Aim: To develop a protocol that enables isolation of human lung mast cells at high yields for analysis of functional properties and detailed analysis using single-cell based analyses of protein (flow cytometry) or RNA (RNA-sequencing) expression.Methods: Mast cells were isolated from human lung tissue by a sequential combination of washing, enzymatic digestion, mechanical disruption, and density centrifugation using Percoll (WEMP). As a comparison, we also isolated mast cells using a conventional enzyme-based protocol. The isolated cells were analyzed by flow cytometry.Results: We observed a significant increase in the yield of total human lung CD45(+) immune cells and an even more pronounced increase in the yield of CD117(+) mast cells with the WEMP protocol in comparison to the conventional protocols. In contrast, the frequency of the rare lymphocyte subset innate lymphoid cells group 2 (ILC2) did not differ between the two methods.Conclusion: The described WEMP protocol results in a significant increase in the yield of human lung mast cells compared to a conventional protocol. Additionally, the WEMP protocol enables simultaneous isolation of different immune cell populations such as lymphocytes, monocytes, and granulocytes while retaining their surface marker expression that can be used for advanced single-cell analyses including multi-color flow cytometry and RNA-sequencing.
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| 8. |
- Rönnberg, Elin, et al.
(författare)
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Divergent Effects of Acute and Prolonged Interleukin 33 Exposure on Mast Cell IgE-Mediated Functions
- 2019
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Ingår i: Frontiers in Immunology. - : FRONTIERS MEDIA SA. - 1664-3224. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- Background: Epithelial cytokines, including IL-33 and Thymic stromal lymphopoietin (TSLP), have attracted interest because of their roles in chronic allergic inflammation-related conditions such as asthma. Mast cells are one of the major targets of IL-33, to which they respond by secreting cytokines. Most studies performed thus far have investigated the acute effects of IL-33 on mast cells. In the current study, we investigated how acute vs. prolonged exposure of mast cells to IL-33 and TSLP affects mediator synthesis and IgE-mediated activation.Methods: Human lung mast cells (HLMCs), cord blood-derived mast cells (CBMCs), and the ROSA mast cell line were used for this study. Receptor expression and the levels of mediators were measured after treatment with IL-33 and/or TSLP.Results: IL-33 induced the release of cytokines. Prolonged exposure to IL-33 increased while TSLP reduced intracellular levels of tryptase. Acute IL-33 treatment strongly potentiated IgE-mediated activation. In contrast, 4 days of exposure to IL-33 decreased IgE-mediated activation, an effect that was accompanied by a reduction in Fc epsilon RI expression.Conclusion: We show that IL-33 plays dual roles in mast cells, in which its acute effects include cytokine release and the potentiation of IgE-mediated degranulation, whereas prolonged exposure to IL-33 reduces IgE-mediated activation. We conclude that mast cells act quickly in response to the alarmin IL-33 to initiate an acute inflammatory response, whereas extended exposure to IL-33 during prolonged inflammation reduces IgE-mediated responses. This negative feedback effect suggests the presence of a novel regulatory pathway that modulates IgE-mediated human mast cell responses.
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| 9. |
- Säfholm, Jesper
(författare)
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Prostaglandin E2 as mediator and modulator of airway smooth muscle responses
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Prostaglandin E2 (PGE2) is a lipid mediator produced by virtually every cell of the human body. Because common non-steroidal anti-inflammatory drugs (NSAIDs) inhibit its biosynthesis, PGE2 is usually considered to be a ‘pro-inflammatory’ mediator. The role of PGE2 in the lung and airways has however always been unclear. In particular, the airway responses caused by activation of its four different EP receptors have been debated. Research on the mechanisms involved in the actions of PGE2 has previously been limited by the low potency and selectivity of available pharmacological tools. Recently, a number of potent receptor antagonists and enzyme inhibitors have become available. The aim of this thesis was therefore to characterise airway responses to PGE2 in greater detail, focusing on the role of its receptors on baseline smooth muscle function and during antigen-induced contractions. Alongside investigating PGE2 responses, the newly discovered relaxant effects of bitter tasting substances acting at their respective receptors (TAS2Rs) were examined. The project mainly involved analysis of isometric contractions and relaxations in isolated airways from guinea pigs and humans in organ baths. In addition, mRNA expression of receptors and enzymes was analysed by PCR and prostanoid release was measured by chemical or immunological methods. It was found that all four EP receptors, the two cyclooxygenases (COX-1 and COX-2) and three PGE synthases were expressed in the guinea pig trachea and lung. Exogenous PGE2 induced a bell-shaped concentration-response curve, causing contraction at lower concentrations mediated by the EP1 receptor and relaxation mediated by the EP2 receptor at higher concentrations. The spontaneous airway tone was maintained by biosynthesis of endogenous PGE2, mainly catalysed by COX-2 in the airway epithelium. The level of tone in the preparation was determined by the balance between activation of EP1 and EP2 receptors. The EP1 receptor, but not the EP2 receptor, displayed homologous desensitisation to endogenous PGE2. When the antigen-induced contractions of the guinea pig trachea were studied, it was found that the primary effect of PGE2 was again to maintain the spontaneous airway tone. When this effect was blocked by EP1 and EP2 receptor antagonists, it was revealed that a component of the antigen-response was mediated by prostaglandin D2 and thromboxane A2, acting at the thromboxane TP receptor. In human small airways, PGE2 induced relaxations mediated by EP4 receptors at low concentrations and contractions through TP receptors at higher concentrations. In addition, it was discovered that the IgE-dependent contraction of human bronchi could be abolished by the action of exogenous PGE2 at the EP2 receptor, an effect presumably involving inhibition of mast cell mediator release. The bitter tasting substances chloroquine, denatonium, thiamine, and noscapine caused relaxations of human and guinea pig airways with a greater efficacy than beta-adrenergic bronchodilators. TAS2R3, TAS2R4 and TAS2R10 were expressed in the guinea pig trachea. The EP receptor-mediated tone could be relaxed by chloroquine and noscapine, but not by denatonium and thiamine. Although the mechanisms underlying these powerful relaxations remain unknown, the data support the involvement of several different pathways. In summary, PGE2 causes contractions and relaxations of guinea pig and human airways, but the receptors involved differ between the two species. Furthermore, the data gathered suggest that EP, TP and TAS2R receptors may be potential targets for the development of drugs to treat asthma and other forms of airway obstruction.
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| 10. |
- Tebroke, Julia, et al.
(författare)
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Wnt-3a Induces Cytokine Release in Human Mast Cells
- 2019
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Ingår i: Cells. - : MDPI. - 2073-4409. ; 8:11
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Tidskriftsartikel (refereegranskat)abstract
- Mast cells are well known for their detrimental effects in allergies and asthma, and Wnt signaling has recently been implicated in asthma and other airway diseases. However, it is not known if or how Wnts affect human mast cells. Since Wnt expression is elevated in individuals with asthma and is linked to a Th2 profile, we hypothesized that mast cells could be affected by Wnts in the context of asthma. We therefore sought to investigate the role of Wnt signaling in human mast cell development and activation. We first examined the expression of the 10 main Wnt receptors, Frizzled 1-10 (FZD(1-10)), and found expression of several FZDs in human mast cells. Treatment with purified recombinant Wnt-3a or Wnt-5a did not affect the proliferation or maturation of CD34(+) progenitors into mast cells, as indicated by cellular expression of CD117 and Fc epsilon RI, activation by Fc epsilon RI crosslinking, and histamine and tryptase release. Furthermore, Wnt treatment did not change the phenotype from MCT to MCTC, since MrgX2 expression, compound 48/80-mediated activation, and carboxypeptidase A3 content were not affected. However, Wnt-3a activated WNT/beta-catenin signaling in mature human mast cells, as revealed by stabilization of beta-catenin, upregulation of IL-8 and CCL8 mRNA expression, and release of IL-8 protein. Thus, our data suggest that Wnt-3a activation of mast cells could contribute to the recruitment of immune cells in conditions associated with increased Wnt-3a expression, such as asthma.
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