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  • 1.
    Alim, Md. Abdul
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine. Department of Molecular Medicine and Surgery, Karolinska Institutet.
    Ackermann, Paul W
    Eliasson, Pernilla
    Blomgran, Parmis
    Kristiansson, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Peterson, Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine.
    Increased mast cell degranulation and co-localization of mast cells with the NMDA receptor-1 during healing after Achilles tendon rupture2017In: Cell and Tissue Research, ISSN 0302-766X, E-ISSN 1432-0878, Vol. 370, no 3, p. 451-460Article in journal (Refereed)
    Abstract [en]

    The role of inflammation and the mechanism of tendon healing after rupture has historically been a matter of controversy. The purpose of the present study is to investigate the role of mast cells and their relation to the NMDA receptor-1 (a glutamate receptor) during healing after Achilles tendon rupture. Eight female Sprague Dawley rats had their right Achilles tendon transected. Three weeks after rupture, histological quantification of mast cell numbers and their state of degranulation was assessed by histochemistry. Co-localization of mast cell tryptase (a mast cell marker) and NMDA receptor-1 was determined by immunofluorescence. The intact left Achilles tendon was used as control. An increased number of mast cells and a higher proportion of degranulated mast cells were found in the healing Achilles tendon compared to the intact. In addition, increased co-localization of mast cell tryptase and NMDA receptor-1 was seen in the areas of myotendinous junction, mid-tendon proper and bone tendon junction of the healing versus the intact tendon. These findings introduce a possible role for mast cells in the healing phase after Achilles tendon rupture.

  • 2.
    Alim, Md Abdul
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine. Department of Molecular Medicine and Surgery, Karolinska Institutet.
    Ackermann, Paul W
    Eliasson, Pernilla
    Blomgran, Parmis
    Kristiansson, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Peterson, Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine.
    Increased mast cell degranulation and co-localization of mast cells with the NMDA receptor-1 during healing after Achilles tendon rupture2017In: Cell and Tissue Research, ISSN 0302-766X, E-ISSN 1432-0878, Vol. 370, no 3, p. 451-460Article in journal (Refereed)
    Abstract [en]

    The role of inflammation and the mechanism of tendon healing after rupture has historically been a matter of controversy. The purpose of the present study is to investigate the role of mast cells and their relation to the NMDA receptor-1 (a glutamate receptor) during healing after Achilles tendon rupture. Eight female Sprague Dawley rats had their right Achilles tendon transected. Three weeks after rupture, histological quantification of mast cell numbers and their state of degranulation was assessed by histochemistry. Co-localization of mast cell tryptase (a mast cell marker) and NMDA receptor-1 was determined by immunofluorescence. The intact left Achilles tendon was used as control. An increased number of mast cells and a higher proportion of degranulated mast cells were found in the healing Achilles tendon compared to the intact. In addition, increased co-localization of mast cell tryptase and NMDA receptor-1 was seen in the areas of myotendinous junction, mid-tendon proper and bone tendon junction of the healing versus the intact tendon. These findings introduce a possible role for mast cells in the healing phase after Achilles tendon rupture.

  • 3.
    Alim, Md Abdul
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine.
    Grujic, Mirjana
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Ackerman, Paul W.
    Karolinska Institutet, Department of Molecular Medicine and Surgery, Stockholm, Sweden.
    Kristiansson, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine.
    Peterson, Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish University of Agricultural Sciences, Department of Anatomy, Physiology and Biochemistry, Uppsala, Sweden.
    Glutamate Triggers the Expression of Functional Ionotropic and Metabotropic Glutamate Receptors in Mast CellsManuscript (preprint) (Other academic)
  • 4.
    Bradding, Peter
    et al.
    Univ Leicester, Inst Lung Hlth, Dept Infect Immun & Inflammat, Leicester, Leics, England..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    The controversial role of mast cells in fibrosis2018In: Immunological Reviews, ISSN 0105-2896, E-ISSN 1600-065X, Vol. 282, no 1, p. 198-231Article, review/survey (Refereed)
    Abstract [en]

    Fibrosis is a medical condition characterized by an excessive deposition of extracellular matrix compounds such as collagen in tissues. Fibrotic lesions are present in many diseases and can affect all organs. The excessive extracellular matrix accumulation in these conditions can often have serious consequences and in many cases be life-threatening. A typical event seen in many fibrotic conditions is a profound accumulation of mast cells (MCs), suggesting that these cells can contribute to the pathology. Indeed, there is now substantialv evidence pointing to an important role of MCs in fibrotic disease. However, investigations from various clinical settings and different animal models have arrived at partly contradictory conclusions as to how MCs affect fibrosis, with many studies suggesting a detrimental role of MCs whereas others suggest that MCs can be protective. Here, we review the current knowledge of how MCs can affect fibrosis.

  • 5.
    Desbiens, Louisane
    et al.
    Univ Sherbrooke, Med Sch, Dept Pharmacol & Physiol, Sherbrooke, PQ, Canada.
    Lapointe, Catherine
    Univ Sherbrooke, Med Sch, Dept Pharmacol & Physiol, Sherbrooke, PQ, Canada.
    Gendron, Louis
    Univ Sherbrooke, Med Sch, Dept Pharmacol & Physiol, Sherbrooke, PQ, Canada.
    Gharagozloo, Marjan
    Univ Sherbrooke, Med Sch, Dept Pharmacol & Physiol, Sherbrooke, PQ, Canada.
    Vincent, Laurence
    Univ Sherbrooke, Med Sch, Dept Pharmacol & Physiol, Sherbrooke, PQ, Canada.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Gris, Denis
    Univ Sherbrooke, Med Sch, Dept Pharmacol & Physiol, Sherbrooke, PQ, Canada.
    D'Orleans-Juste, Pedro
    Univ Sherbrooke, Med Sch, Dept Pharmacol & Physiol, Sherbrooke, PQ, Canada.
    Experimental Autoimmune Encephalomyelitis Potentiates Mouse Mast Cell Protease 4-Dependent Pressor Responses to Centrally or Systemically Administered Big Endothelin-12019In: Journal of Pharmacology and Experimental Therapeutics, ISSN 0022-3565, E-ISSN 1521-0103, Vol. 370, no 3, p. 437-446Article in journal (Refereed)
    Abstract [en]

    Multiple sclerosis is a neurodegenerative disease affecting predominantly female patients between 20 and 45 years of age. We previously reported the significant contribution of mouse mast cell protease 4 (mMCP-4) in the synthesis of endothelin-1 (ET-1) in healthy mice and in a murine model of experimental autoimmune encephalomyelitis (EAE). In the current study, the cardiovascular effects of ET-1 and big endothelin-1 (big-ET-1) administered systemically or intrathecally were assessed in the early preclinical phase of EAE in telemetry instrumented/conscious mice. Chymase-specific enzymatic activity was also measured in the lung, brain, and mast cell extracts in vitro. Finally, the impact of EAE immunization was studied on the pulmonary and brain mRNA expression of different genes of the endothelin pathway, interleukin-33 (IL-33), and monitoring of immunoreactive tumor necrosis factor-α (TNF-α). Systemically or intrathecally administered big-ET-1 triggered increases in blood pressure in conscious mice. One week post-EAE, the pressor responses to big-ET-1 were potentiated in wild-type (WT) mice but not in mMCP-4 knockout (KO) mice. EAE triggered mMCP-4–specific activity in cerebral homogenates and peritoneal mast cells. Enhanced pulmonary, but not cerebral preproendothelin-1 and IL-33 mRNA were found in KO mice and further increased 1 week post-EAE immunization, but not in WT animals. Finally, TNF-α levels were also increased in serum from mMCP-4 KO mice, but not WT, 1 week post-EAE. Our study suggests that mMCP-4 activity is enhanced both centrally and systemically in a mouse model of EAE.

  • 6.
    Frisk, Jun Mei Hu
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Kjellén, Lena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Melo, Fabio R.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Öhrvik, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Mitogen-Activated Protein Kinase Signaling Regulates Proteoglycan Composition of Mast Cell Secretory Granules2018In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 9, article id 1670Article in journal (Refereed)
    Abstract [en]

    Mast cells (MCs) are characterized by an abundance of lysosome-like secretory granules filled with immunomodulatory compounds including histamine, cytokines, lysosomal hydrolases, MC-restricted proteases, and serglycin proteoglycans. The latter are essential for promoting the storage of other granule compounds and are built up of the serglycin core protein to which highly sulfated and thereby negatively charged glycosaminoglycan (GAG) side chains of heparin or chondroitin sulfate type are attached. In the search for mechanisms operating in regulating MC granule homeostasis, we here investigated the role of mitogen-activated protein kinase (MAPK) signaling. We show that inhibition of MEK1/2 (a MAPK kinase) leads to increased metachromatic staining of MC granules, indicative of increased proteoglycan content. Indeed, MEK1/2 inhibition caused a profound increase in the expression of the gene coding for the serglycin core protein and of genes coding for various enzymes involved in the biosynthesis/sulfation of the GAGs attached to the serglycin core protein. This was accompanied by corresponding increases in the levels of the respective GAGs. Deletion of the serglycin core protein abrogated the induction of enzymes operative in proteoglycan synthesis, indicating that availability of the serglycin proteoglycan core protein has a regulatory function impacting on the expression of the various serglycin-modifying enzymes. MEK1/2 inhibition also caused a substantial increase in the expression of granule-localized, proteoglycan-binding proteases. Altogether, this study identifies a novel role for MAPK signaling in regulating the content of secretory granules in MCs.

  • 7. Galli, Stephen J
    et al.
    Tsai, Mindy
    Marichal, Thomas
    Tchougounova, Elena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology.
    Reber, Laurent L
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Approaches for analyzing the roles of mast cells and their proteases in vivo2015In: Advances in Immunology, ISSN 0065-2776, E-ISSN 1557-8445, Vol. 126, p. 45-127Article in journal (Refereed)
    Abstract [en]

    The roles of mast cells in health and disease remain incompletely understood. While the evidence that mast cells are critical effector cells in IgE-dependent anaphylaxis and other acute IgE-mediated allergic reactions seems unassailable, studies employing various mice deficient in mast cells or mast cell-associated proteases have yielded divergent conclusions about the roles of mast cells or their proteases in certain other immunological responses. Such "controversial" results call into question the relative utility of various older versus newer approaches to ascertain the roles of mast cells and mast cell proteases in vivo. This review discusses how both older and more recent mouse models have been used to investigate the functions of mast cells and their proteases in health and disease. We particularly focus on settings in which divergent conclusions about the importance of mast cells and their proteases have been supported by studies that employed different models of mast cell or mast cell protease deficiency. We think that two major conclusions can be drawn from such findings: (1) no matter which models of mast cell or mast cell protease deficiency one employs, the conclusions drawn from the experiments always should take into account the potential limitations of the models (particularly abnormalities affecting cell types other than mast cells) and (2) even when analyzing a biological response using a single model of mast cell or mast cell protease deficiency, details of experimental design are critical in efforts to define those conditions under which important contributions of mast cells or their proteases can be identified.

  • 8.
    Garcia-Faroldi, Gianni
    et al.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, BMC, Uppsala, Sweden..
    Melo, Fabio R.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, BMC, Uppsala, Sweden..
    Bruemmer, Dennis
    Univ Kentucky, Coll Med, Saha Cardiovasc Res Ctr, Wethington, KY USA..
    Conneely, Orla M.
    Baylor Coll Med, Dept Mol & Cellular Biol, Houston, TX 77030 USA..
    Pejler, Gunnar
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, BMC, Uppsala, Sweden..
    Lundequist, Anders
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, BMC, Uppsala, Sweden..
    Nuclear Receptor 4a3 (Nr4a3) Regulates Murine Mast Cell Responses and Granule Content2014In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 9, no 2, article id e89311Article in journal (Refereed)
    Abstract [en]

    Nuclear receptor 4a3 (Nr4a3) is a transcription factor implicated in various settings such as vascular biology and inflammation. We have recently shown that mast cells dramatically upregulate Nuclear receptor 4a3 upon activation, and here we investigated the functional impact of Nuclear receptor 4a3 on mast cell responses. We show that Nuclear receptor 4a3 is involved in the regulation of cytokine/chemokine secretion in mast cells following activation via the high affinity IgE receptor. Moreover, Nuclear receptor 4a3 negatively affects the transcript and protein levels of mast cell tryptase as well as the mast cell's responsiveness allergen. Together, these findings identify Nuclear receptor 4a3 as a novel regulator of mast cell function.

  • 9.
    Garcia-Faroldi, Gianni
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Rönnberg, Elin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Grujic, Mirjana
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Department of Anatomy, Physiology and Biochemistry, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Inhibition of the BET family of epigenetic reader proteins: A novel principle for modulating gene expression in IgE-activated mast cells2017In: Immunity, Inflammation and Disease, E-ISSN 2050-4527, Vol. 5, no 2, p. 141-150Article in journal (Refereed)
    Abstract [en]

    Introduction: The BET family of bromodomain-containing proteins constitute epigenetic readers that bind to acetylated lysine residues of core histones, thereby translating epigenetic histone marks to effects on gene expression. BET inhibitors are currently emerging as promising therapeutic agents for treatment of various pathological conditions. Here, we explored the potential of using BET inhibition to modulate IgE-mediated responses in mast cells.

    Methods: We assessed the effects of BET inhibitors PFI-1, I-BET151, and I-BET762 on responses downstream of mast cell activation through IgE receptor cross-linking.

    Results: BET inhibitors were neither toxic for mast cells (at doses up to 20M), nor did they prevent IgE-mediated mast cell degranulation. However, we found that BET inhibition, in particular by I-BET151, suppressed IL-6 gene expression and IL-6 protein release in response to IgE-mediated mast cell activation. This was observed in both bone marrow-derived mast cells (BMMCs) and in mature peritoneal-cell derived mast cells. Further analysis showed that BET inhibition also suppressed the expression of a number of additional genes of those that were upregulated by IgE receptor cross-linking, including IL-3, IL-7R, CCR1, and ADAMTS9. However, BET inhibition was selective, i.e., several genes that were upregulated by IgE receptor cross-linking were not affected by BET inhibitors.

    Conclusions: These findings suggest that BET inhibition can interfere with the upregulated expression of selected genes in mast cells activated by IgE receptor cross-linking. Further, our findings introduce the concept of utilizing epigenetic mechanisms for modulating mast cell function in the context of IgE-driven disease.

  • 10. Garcia-Vilas, Javier A.
    et al.
    Medina, Miguel A.
    Melo, Fabio R.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Garcia-Faroldi, Gianni
    Damnacanthal inhibits IgE receptor-mediated activation of mast cells2015In: Molecular Immunology, ISSN 0161-5890, E-ISSN 1872-9142, Vol. 65, no 1, p. 86-93Article in journal (Refereed)
    Abstract [en]

    Damnacanthal, an anthraquinone obtained from the noni fruit (Morinda citrifolia L), has been described to possess anti-cancer and anti-inflammatory properties. Since mast cells are key players in various inflammatory conditions as well as in cancer, we considered the possibility that the biological actions of damnacanthal, at least partly, could be due to effects on mast cells. Many of the biological activities of mast cells are mediated by IgE receptor cross-linking, which results in degranulation with release of preformed granule mediators, as well as de nova synthesis and release of additional compounds. Here we show that damnacanthal has profound inhibitory activity on mast cell activation through this pathway. The release of the granule compounds beta-hexosaminidase and tryptase release was completely abrogated by damnacanthal at doses that were non-toxic to mast cells. In addition, damnacanthal inhibited activation-dependent pro-inflammatory gene induction, as well as cytokine/chemokine release in response to mast cell stimulation. The mechanism underlying damnacanthal inhibition was linked to impaired phosphorylation of Syk and Akt. Furthermore, damnacanthal inhibited mast cell activation in response to calcium ionophore A23187. Altogether, the data presented here demonstrate that damnacanthal inhibits mast cell activation induced by different stimuli and open a new window for the use of this compound as a mast cell stabilizer. 

  • 11. Gendrin, Claire
    et al.
    Shubin, Nicholas J
    Boldenow, Erica
    Merillat, Sean
    Clauson, Morgan
    Power, Danial
    Doran, Kelly S
    Abrink, Magnus
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Department of Anatomy, Physiology and Biochemistry, the Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Rajagopal, Lakshmi
    Piliponsky, Adrian M
    Mast cell chymase decreases the severity of group B Streptococcus infections2018In: Journal of Allergy and Clinical Immunology, ISSN 0091-6749, E-ISSN 1097-6825, Vol. 142, no 1, p. 120-129Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Group B Streptococcus (GBS) or Streptococcus agalactiae are β-hemolytic gram-positive bacteria that colonize the lower genital tracts of women and are frequently associated with infections during pregnancy. Innate immune defenses are critical for controlling GBS dissemination and systemic infection. Mast cells are resident sentinel cells that come into contact with pathogens early during colonization and infection.

    OBJECTIVE: We aimed to investigate the contribution of chymase to systemic GBS infection and rates of preterm birth.

    METHODS: Pharmacologic and genetic approaches using mice deficient in mast cell protease (MCPT) 4, the mouse functional homologue of human chymase, were used.

    RESULTS: Our studies show that mast cells release a protease with chymotrypsin-like cleavage specificity in response to GBS. Additionally, increased GBS systemic infection and preterm births were observed in MCPT4-deficient mice versus MCPT4-sufficient mice. Furthermore, we observed that proteolytic cleavage of the host extracellular matrix protein fibronectin by peritoneal cell-derived mast cell lysates diminished GBS adherence. Consistent with this observation, the increase in GBS dissemination and preterm births observed in MCPT4-deficient mice was abolished when GBS was deficient in expression of the fibronectin-binding protein SfbA.

    CONCLUSIONS: Taken together, our results suggest that the protective effect of MCPT4 against GBS dissemination and preterm labor can be attributed in part to MCPT4-mediated proteolysis of fibronectin. Our studies reveal a novel role of mast cells in defense against bacterial infections.

  • 12.
    Grujic, Mirjana
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Paivandy, Aida
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Gustafson, Ann-Marie
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Thomsen, Allan R.
    Univ Copenhagen, Dept Immunol & Microbiol, Copenhagen, Denmark..
    Öhrvik, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    The combined action of mast cell chymase, tryptase and carboxypeptidase A3 protects against melanoma colonization of the lung2017In: OncoTarget, ISSN 1949-2553, E-ISSN 1949-2553, Vol. 8, no 15, p. 25066-25079Article in journal (Refereed)
    Abstract [en]

    Mast cell secretory granules are densely packed with various bioactive mediators including proteases of chymase, tryptase and CPA3 type. Previous studies have indicated that mast cells can affect the outcome of melanoma but the contribution of the mast cell granule proteases to such effects has not been clear. Here we addressed this issue by assessing mice lacking either the chymase Mcpt4, the tryptase Mcpt6 or carboxypeptidase A3 (Cpa3), as well as mice simultaneously lacking all three proteases, in a model of melanoma dissemination from blood to the lung. Although mice with individual deficiency in the respective proteases did not differ significantly from wildtype mice in the extent of melanoma colonization, mice with multiple protease deficiency (Mcpt4/Mcpt6/Cpa3-deficient) exhibited a higher extent of melanoma colonization in lungs as compared to wildtype animals. This was supported by higher expression of melanoma-specific genes in lungs of Mcpt4/Mcpt6/CPA3-deficient vs. wildtype mice. Cytokine profiling showed that the levels of CXCL16, a chemokine with effects on T cell populations and NKT cells, were significantly lower in lungs of Mcpt4/Mcpt6/Cpa3-deficient animals vs. controls, suggesting that multiple mast cell protease deficiency might affect T cell or NKT cell populations. In line with this, we found that the Mcpt4/Mcpt6/Cpa3-deficiency was associated with a reduction in cells expressing CD1d, a MHC class 1-like molecule that is crucial for presenting antigen to invariant NKT (iNKT) cells. Together, these findings indicate a protective role of mast cell-specific proteases in melanoma dissemination, and suggest that this effect involves a CXCL16/CD1d/NKT cell axis.

  • 13. Hagforsen, E
    et al.
    Lampinen, M
    Paivandy, A
    Weström, S
    Velin, H
    Öberg, S
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Rollman, O
    Siramesine causes preferential apoptosis of mast cells in skin biopsies from psoriatic lesions.2017In: British Journal of Dermatology, ISSN 0007-0963, E-ISSN 1365-2133, Vol. 177, no 1, p. 179-187Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Skin mast cells are implicated as detrimental effector cells in various inflammatory skin diseases such as contact eczema, atopic dermatitis and psoriasis. Selective reduction of cutaneous mast cells, e.g. by inducing targeted apoptosis, might prove a rational and efficient therapeutic strategy in dermatoses negatively influenced by mast cells.

    OBJECTIVES: The objective of the present study was to evaluate whether a lysosomotropic agent such as siramesine can cause apoptosis of mast cells present in psoriatic lesions.

    MATERIALS AND METHODS: Punch biopsies were obtained from lesional and uninvolved skin in 25 patients with chronic plaque psoriasis. After incubation with siramesine, the number of tryptase-positive mast cells and their expression of interleukin (IL)-6 and IL-17 was analysed. Skin biopsies were digested to allow flow cytometric analysis of the drug's effect on cutaneous fibroblasts and keratinocytes.

    RESULTS: Siramesine caused a profound reduction in the total number of mast cells in both lesional and uninvolved psoriatic skin biopsies without affecting the gross morphology of the tissue. The drug reduced the density of IL-6- and IL-17-positive mast cells, and showed antiproliferative effects on epidermal keratinocytes but had no apparent cytotoxic effect on keratinocytes or dermal fibroblasts.

    CONCLUSIONS: Considering the pathophysiology of psoriasis, the effects of siramesine on cutaneous mast cells may prove favourable from the therapeutic aspect. The results encourage further studies to assess the usefulness of siramesine and other lysosomotropic agents in the treatment of cutaneous mastocytoses and inflammatory skin diseases aggravated by dermal mast cells.

  • 14.
    Hagforsen, Eva
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Dermatology and Venereology.
    Paivandy, Aida
    Lampinen, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Gastroenterology/Hepatology.
    Weström, Simone
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Dermatology and Venereology.
    Calounova, Gabriela
    Melo, Fabio R.
    Rollman, Ola
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Dermatology and Venereology.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Ablation of human skin mast cells in situ by lysosomotropic agents2015In: Experimental dermatology, ISSN 0906-6705, E-ISSN 1600-0625, Vol. 24, no 7, p. 516-521Article in journal (Refereed)
    Abstract [en]

    Mast cells are known to have a detrimental impact on numerous types of inflammatory skin diseases such as contact dermatitis, atopic eczema and cutaneous mastocytosis. Regimens that dampen skin mast cell-mediated activities can thus offer an attractive therapeutic option under such circumstances. As mast cells are known to secrete a large array of potentially pathogenic compounds, both from preformed stores in secretory lysosomes (granules) and after de novo synthesis, mere inhibition of degranulation or interference with individual mast cell mediators may not be sufficient to provide an effective blockade of harmful mast cell activities. An alternative strategy may therefore be to locally reduce skin mast cell numbers. Here, we explored the possibility of using lysosomotropic agents for this purpose, appreciating the fact that mast cell granules contain bioactive compounds prone to trigger apoptosis if released into the cytosolic compartment. Based on this principle, we show that incubation of human skin punch biopsies with the lysosomotropic agents siramesine or Leu-Leu methyl ester preferably ablated the mast cell population, without causing any gross adverse effects on the skin morphology. Subsequent analysis revealed that mast cells treated with lysosomotropic agents predominantly underwent apoptotic rather than necrotic cell death. In summary, this study raises the possibility of using lysosomotropic agents as a novel approach to targeting deleterious mast cell populations in cutaneous mastocytosis and other skin disorders negatively influenced by mast cells.

  • 15.
    Henningsson, Frida
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology.
    Ledin, Johan
    Department of Physiology and Developmental Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology.
    Lunderius, Carolina
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology. Molekylär immunologi.
    Wilén, M
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology.
    Hellman, Lars
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology. Molekylär Immunologi.
    Pejler, Gunnar
    Department of Physiology and Developmental Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology.
    Altered storage of proteases in mast cells from mice lacking heparin: a possible role for heparin in carboxypeptidase A processing.2002In: Biological Chemistry, Vol. 383, p. 793-801Article in journal (Refereed)
  • 16.
    Houde, Martin
    et al.
    Univ Sherbrooke, Fac Med & Sci Sante, Dept Pharmacol Physiol, Sherbrooke, PQ, Canada; Leiden Univ, Leiden Acad, Ctr Drug Res, Div BioTherapeut, Leiden, Netherlands.
    Schwertani, Adel
    McGill Univ, Dept Med, Montreal, PQ, Canada.
    Touil, Hanene
    Univ Sherbrooke, Fac Med & Sci Sante, Dept Pharmacol Physiol, Sherbrooke, PQ, Canada.
    Desbiens, Louisane
    Univ Sherbrooke, Fac Med & Sci Sante, Dept Pharmacol Physiol, Sherbrooke, PQ, Canada.
    Sarrhini, Otman
    Univ Sherbrooke, CRCHUS, Sherbrooke Mol Imaging Ctr, Dept Nucl Med & Radiobiol, Sherbrooke, PQ, Canada.
    Lecomte, Roger
    Univ Sherbrooke, CRCHUS, Sherbrooke Mol Imaging Ctr, Dept Nucl Med & Radiobiol, Sherbrooke, PQ, Canada.
    Lepage, Martin
    Univ Sherbrooke, CRCHUS, Sherbrooke Mol Imaging Ctr, Dept Nucl Med & Radiobiol, Sherbrooke, PQ, Canada.
    Gagnon, Hugo
    PhenoSwitch Biosci Inc, Sherbrooke, PQ, Canada.
    Takai, Shinji
    Osaka Med Coll, Dept Innovat Med, Osaka, Japan.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Jacques, Danielle
    Univ Sherbrooke, Fac Med & Sci Sante, Dept Anat & Cell Biol, Sherbrooke, PQ, Canada.
    Gobeil, Fernand, Jr.
    Univ Sherbrooke, Fac Med & Sci Sante, Dept Pharmacol Physiol, Sherbrooke, PQ, Canada.
    Day, Robert
    Univ Sherbrooke, Fac Med & Sci Sante, Dept Surg, Sherbrooke, PQ, Canada.
    D'Orleans-Juste, Pedro
    Univ Sherbrooke, Fac Med & Sci Sante, Dept Pharmacol Physiol, Sherbrooke, PQ, Canada.
    Mouse Mast Cell Protease 4 Deletion Protects Heart Function and Survival After Permanent Myocardial Infarction2018In: Frontiers in Pharmacology, ISSN 1663-9812, E-ISSN 1663-9812, Vol. 9, article id 868Article in journal (Refereed)
    Abstract [en]

    Chymase, a mast cell serine protease involved in the generation of multiple cardiovascular factors, such as angiotensin II and endothelin-1 (ET-1), is elevated and participates in tissue degeneration after permanent myocardial infarction (PMI). Anesthetized 4-month old male wild-type (WT) C57BL/6J mice and mouse mast cell protease-4 knockout (mMCP-4 KO) congeners were subjected to ligation of the left anterior descending (LAD) coronary artery. A group of mice was then subjected to Kaplan-Meier 28-day survival analysis. In another group of mice, F-18-fluorodeoxyglucose positron emission tomography (PET) was performed to evaluate heart function and the infarcted zone 3 days post-PMI surgery. Cardiac morphology following PMI was evaluated on formalin-fixed heart slices and glycoproteomic analysis was performed using mass spectrometry. Finally, cardiac and lung tissue content of immunoreactive ET-1 was determined. PMI caused 60% mortality in WT mice, due to left ventricular wall rupture, and 7% in mMCP-4 KO mice. Cardiac PET analysis revealed a significant reduction in left ventricular volume (systolic and diastolic) and preserved the ejection fraction in mMCP-4 KO compared to WT animals. The infarcted area, apoptotic signaling and wall remodeling were significantly decreased in mMCP-4 KO mice compared to their WT congeners, while collagen deposition was increased. Glycoproteomic analysis showed an increase in apolipoprotein A1, an established chymase substrate in mMCP-4 KO mice compared to WT mice post-PMI. ET-1 levels were increased in the lungs of WT, but not mMCP-4 KO mice, 24 h post-PMI. Thus, the genetic deletion of mMCP-4 improved survival and heart function post-PMI.

  • 17.
    Hu Frisk, Jun Mei
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Kjellén, Lena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Kaler, Stephen G
    Eunice Kennedy Shriver Natl Inst Child Hlth & Hum, Sect Translat Neurosci, Mol Med Branch, NIH, Bethesda, MD 20892 USA.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75007 Uppsala, Sweden.
    Öhrvik, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Copper Regulates Maturation and Expression of an MITF: Tryptase Axis in Mast Cells2017In: Journal of Immunology, ISSN 0022-1767, E-ISSN 1550-6606, Vol. 199, no 12, p. 4132-4141Article in journal (Refereed)
    Abstract [en]

    Copper has previously been implicated in the regulation of immune responses, but the impact of this metal on mast cells is poorly understood. In this article, we address this issue and show that copper starvation of mast cells causes increased granule maturation, as indicated by higher proteoglycan content, stronger metachromatic staining, and altered ultrastructure in comparison with nontreated cells, whereas copper overload has the opposite effects. In contrast, copper status did not impact storage of histamine in mast cells, nor did alterations in copper levels affect the ability of mast cells to degranulate in response to IgER cross-linking. A striking finding was decreased tryptase content in mast cells with copper overload, whereas copper starvation increased tryptase content. These effects were associated with corresponding shifts in tryptase mRNA levels, suggesting that copper affects tryptase gene regulation. Mechanistically, we found that alterations in copper status affected the expression of microphthalmia-associated transcription factor, a transcription factor critical for driving tryptase expression. We also found evidence supporting the concept that the effects on microphthalmia-associated transcription factor are dependent on copper-mediated modulation of MAPK signaling. Finally, we show that, in MEDNIK syndrome, a condition associated with low copper levels and a hyperallergenic skin phenotype, including pruritis and dermatitis, the number of tryptase-positive mast cells is increased. Taken together, our findings reveal a hitherto unrecognized role for copper in the regulation of mast cell gene expression and maturation.

  • 18.
    Johnzon, Carl-Fredrik
    et al.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Dahlberg, Josef
    Swedish Univ Agr Sci, Dept Anim Nutr & Management, Uppsala, Sweden.
    Gustafson, Ann-Marie
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Waern, Ida
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Moazzami, Ali A.
    Swedish Univ Agr Sci, Dept Mol Sci, Uppsala, Sweden.
    Ostensson, Karin
    Swedish Univ Agr Sci, Dept Clin Sci, Uppsala, Sweden.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden .
    The Effect of Lipopolysaccharide-Induced Experimental Bovine Mastitis on Clinical Parameters, Inflammatory Markers, and the Metabolome: A Kinetic Approach2018In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 9, article id 1487Article in journal (Refereed)
    Abstract [en]

    Mastitis is an inflammatory condition of the mammary tissue and represents a major problem for the dairy industry worldwide. The present study was undertaken to study how experimentally induced acute bovine mastitis affects inflammatory parameters and changes in the metabolome. To this end, we induced experimental mastitis in nine cows by intramammary infusion of 100 pg purified Escherichia call lipopolysaccharide (LPS) followed by kinetic assessments of cytokine responses (by enzyme-linked immunosorbent assay), changes in the metabolome (assessed by nuclear magnetic resonance), clinical parameters (heat, local pain perception, redness, swelling, rectal temperature, clot formation, and color changes in the milk), and milk somatic cell counts, at several time points post LPS infusion. Intramammary LPS infusion induced clinical signs of mastitis, which started from 2 h post infusion and had returned to normal levels within 24-72 h. Milk changes were seen with a delay compared with the clinical signs and persisted for a longer time. In parallel, induction of IL-6 and TNF-alpha were seen in milk, and there was also a transient elevation of plasma IL-6 whereas plasma TNF-alpha was not significantly elevated. In addition, a robust increase in CCL2 was seen in the milk of LPS-infused cows, whereas G-CSF, CXCL1, and histamine in milk were unaffected. By using a metabolomics approach, a transient increase of plasma lactose was seen in LPS-induced cows. In plasma, significant reductions in ketone bodies (3-hydroxybutyrate and acetoacetate) and decreased levels of short-chain fatty acids, known to be major products released from the gut microbiota, were observed after LPS infusion; a profound reduction of plasma citrate was also seen. Intramammary LPS infusion also caused major changes in the milk metabolome, although with a delay in comparison with plasma, including a reduction of lactose. We conclude that the LPS-induced acute mastitis rapidly affects the plasma metabolome and cytokine induction with similar kinetics as the development of the clinical signs, whereas the corresponding effects in milk occurred with a delay.

  • 19.
    Johnzon, Carl-Fredrik
    et al.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Ronnberg, Elin
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    The Role of Mast Cells in Bacterial Infection2016In: American Journal of Pathology, ISSN 0002-9440, E-ISSN 1525-2191, Vol. 186, no 1, p. 4-14Article, review/survey (Refereed)
    Abstract [en]

    Mast cells (MCs) are particularly abundant at host-environment interfaces, such as skin and intestinal mucosa. Because of their Location, it has been hypothesized that MCs can act as sentinel cells that sense microbial attacks and initiate a protective immune response. Several studies have suggested that animals deficient in MCs exhibit a worsened pathology in various experimental models of bacterial infection. However, other studies have indicated that MCs under certain circumstances may have a detrimental impact on bacterial disease, and there are also recent studies indicating that MCs are dispensable for the clearance of bacterial pathogens. Herein, we review the current knowledge of the role of MCs in bacterial infection.

  • 20.
    Johnzon, Carl-Fredrik
    et al.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Rönnberg, Elin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Guss, Bengt
    Swedish Univ Agr Sci, Dept Biomed Sci & Vet Publ Hlth, Uppsala, Sweden..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Live Staphylococcus aureus Induces Expression and Release of Vascular Endothelial Growth Factor in Terminally Differentiated Mouse Mast Cells2016In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 7, article id 247Article in journal (Refereed)
    Abstract [en]

    Mast cells have been shown to express vascular endothelial growth factor (VEGF), thereby implicating mast cells in pro-angiogenic processes. However, the mechanism of VEGF induction in mast cells and the possible expression of VEGF in fully mature mast cells have not been extensively studied. Here, we report that terminally differentiated peritoneal cell-derived mast cells can be induced to express VEGF in response to challenge with Staphylococcus aureus, thus identifying a mast cell-bacteria axis as a novel mechanism leading to VEGF release. Whereas live bacteria produced a robust upregulation of VEGF in mast cells, heat-inactivated bacteria failed to do so, and bacteria-conditioned media did not induce VEGF expression. The induction of VEGF was not critically dependent on direct cell-cell contact between bacteria and mast cells. Hence, these findings suggest that VEGF can be induced by soluble factors released during the co-culture conditions. Neither of a panel of bacterial cell-wall products known to activate toll-like receptor (TLR) signaling promoted VEGF expression in mast cells. In agreement with the latter, VEGF induction occurred independently of Myd88, an adaptor molecule that mediates the downstream events following TLR engagement. The VEGF induction was insensitive to nuclear factor of activated T-cells inhibition but was partly dependent on the nuclear factor kappa light-chain enhancer of activated B cells signaling pathway. Together, these findings identify bacterial challenge as a novel mechanism by which VEGF is induced in mast cells.

  • 21.
    Karlsson, I
    et al.
    Double Bond Pharmaceut AB, Uppsala, Sweden.
    Veevnik, D.
    Minsk City Emergency Clin Hosp, Hlth Care Facil, Minsk, BELARUS.
    Fedulov, A.
    Belarusian State Med Univ, Dept Neurol & Neurosurg, Minsk, BELARUS.
    Yurkshtovich, N.
    Belarusian State Univ, Res Inst Phys Chem Problems, Minsk, BELARUS.
    Yurkshtovich, T.
    Belarusian State Univ, Res Inst Phys Chem Problems, Minsk, BELARUS.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Lokot, I
    Double Bond Pharmaceut AB, Uppsala, Sweden.
    Local delivery of temozolomide via a biologically inert carrier (Temodex) prolongs survival in glioma patients, irrespectively of the methylation status of MGMT2019In: Neoplasma (Bratislava), ISSN 0028-2685, E-ISSN 1338-4317, Vol. 66, no 2, p. 288-293Article in journal (Refereed)
    Abstract [en]

    Glioma is the most common brain malignancy. Standard first-line therapy for glioma includes surgery, radiotherapy and systemic administration of temozolomide. However, temozolomide does not reach the brain in sufficient doses when administered orally and has poor efficiency in more than half of the patients. Strategies to improve the treatment of glial malignancies are therefore needed. We have recently developed a system (Temodex) for local administration of temozolomide by encapsulating the drug in a biologically inert matrix. Here, we assessed the effect of Temodex in combination with standard therapy in a small-scale clinical study. Since the efficacy of temozolomide therapy is known to depend on the methylation status of the O-6-methylguanine-DNA methyltransferase gene (MGMT) promoter, we also analyzed whether the effect of Temodex was influenced by the methylation status of MGMT. Our data show that the combination of standard therapy and Temodex was more efficient than standard therapy alone, promoting the overall patient survival by up to 33 weeks. Moreover, the efficacy of Temodex was not dependent on the methylation status of MGMT. Local Temodex administration in combination with standard therapy thereby emerges as a novel therapeutic option, with applicability that is independent on the methylation status of the MGMT promoter.

  • 22.
    Lind, Thomas
    et al.
    Univ Uppsala Hosp, Clin Pharmacol Sect, Dept Med Sci, Uppsala, Sweden..
    Gustafson, Ann-Marie
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Univ Uppsala Hosp, Clin Pharmacol Sect, Dept Med Sci, Uppsala, Sweden.
    Calounova, Gabriela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Hu, Lijuan
    Univ Uppsala Hosp, Clin Pharmacol Sect, Dept Med Sci, Uppsala, Sweden..
    Rasmusson, Annica
    Univ Uppsala Hosp, Clin Pharmacol Sect, Dept Med Sci, Uppsala, Sweden..
    Jonsson, Kenneth B
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Orthopaedics.
    Wernersson, Sara
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Abrink, Magnus
    Swedish Univ Agr Sci, Dept Biomed Sci & Vet Publ Hlth, Uppsala, Sweden..
    Andersson, Göran
    Karolinska Univ Hosp Huddinge, Karolinska Inst, Div Pathol, Dept Lab Med, Stockholm, Sweden..
    Larsson, Sune
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Orthopaedics.
    Melhus, Håkan
    Univ Uppsala Hosp, Clin Pharmacol Sect, Dept Med Sci, Uppsala, Sweden..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Increased Bone Mass in Female Mice Lacking Mast Cell Chymase2016In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 11, no 12, article id e0167964Article in journal (Refereed)
    Abstract [en]

    Here we addressed the potential impact of chymase, a mast-cell restricted protease, on mouse bone phenotype. We show that female mice lacking the chymase Mcpt4 acquired a persistent expansion of diaphyseal bone in comparison with wild type controls, reaching a 15% larger diaphyseal cross sectional area at 12 months of age. Mcpt4(-/-) mice also showed increased levels of a bone anabolic serum marker and higher periosteal bone formation rate. However, they were not protected from experimental osteoporosis, suggesting that chymase regulates normal bone homeostasis rather than the course of osteoporosis. Further, the absence of Mcpt4(-/-) resulted in age-dependent upregulation of numerous genes important for bone formation but no effects on osteoclast activity. In spite of the latter, Mcpt4(-/-) bones had increased cortical porosity and reduced endocortical mineralization. Mast cells were found periosteally and, notably, bone-proximal mast cells in Mcpt4(-/-) mice were degranulated to a larger extent than in wild type mice. Hence, chymase regulates degranulation of bone mast cells, which could affect the release of mast cell-derived factors influencing bone remodelling. Together, these findings reveal a functional impact of mast cell chymase on bone. Further studies exploring the possibility of using chymase inhibitors as a strategy to increase bone volume may be warranted.

  • 23.
    Lind, Thomas
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical pharmacogenomics and osteoporosis.
    Öhman, Caroline
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Applied Materials Sciences.
    Calounova, Gabriela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Rasmusson, Annica
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical pharmacogenomics and osteoporosis.
    Andersson, Goran
    Karolinska Univ Hosp Huddinge, Karolinska Inst, Div Pathol, Dept Lab Med, Stockholm, Sweden..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Department of Biomedical Science and Veterinary Public Health, Swedish University of Agricultural Sciences, Uppsala.
    Melhus, Håkan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical pharmacogenomics and osteoporosis.
    Excessive dietary intake of vitamin A reduces skull bone thickness in mice2017In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 12, no 4, article id e0176217Article in journal (Refereed)
    Abstract [en]

    Calvarial thinning and skull bone defects have been reported in infants with hypervitaminosis A. These findings have also been described in humans, mice and zebrafish with loss-of-function mutations in the enzyme CYP26B1 that degrades retinoic acid (RA), the active metabolite of vitamin A, indicating that these effects are indeed caused by too high levels of vitamin A and that evolutionary conserved mechanisms are involved. To explore these mechanisms, we have fed young mice excessive doses of vitamin A for one week and then analyzed the skull bones using micro computed tomography, histomorphometry, histology and immunohistochemistry. In addition, we have examined the effect of RA on gene expression in osteoblasts in vitro. Compared to a standard diet, a high dietary intake of vitamin A resulted in a rapid and significant reduction in calvarial bone density and suture diastasis. The bone formation rate was almost halved. There was also increased staining of tartrate resistant acid phosphatase in osteocytes and an increased perilacunar matrix area, indicating osteocytic osteolysis. Consistent with this, RA induced genes associated with bone degradation in osteoblasts in vitro. Moreover, and in contrast to other known bone resorption stimulators, vitamin A induced osteoclastic bone resorption on the endocranial surfaces.

  • 24.
    Lützelschwab, Claudia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Immunology and Microbiology.
    Pejler, Gunnar
    Aveskogh, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Immunology and Microbiology.
    Hellman, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Immunology and Microbiology.
    Secretory granule proteases in rat mast cells: Cloning of 10 different serine proteases and a carboxypeptidase A from various rat mast cell populations1997In: Journal of Experimental Medicine, ISSN 0022-1007, E-ISSN 1540-9538, Vol. 185, no 1, p. 13-29Article in journal (Refereed)
    Abstract [en]

    Two of the major rat mast cell proteases, rat mast cell protease 1 (RMCP-1) and RMCP-2, have for many years served as important phenotypic markers for studies of various aspects of mast cell (MC) biology. However, except for these proteases only fragmentary information has been available on the structure and complexity of proteases expressed by different subpopulations of rat MCs. To address these questions, cDNA libraries were constructed from freshly isolated rat peritoneal MCs and from the rat mucosal MC line RBL-1. cDNA clones for 10 different serine proteases (RMCP- 1-10), and the MC carboxypeptidase A were isolated and characterized. Six of these proteases have not been isolated previously. Based on their protease content, three separate subpopulations of MCs were identified. Connective tissue MCs (CTMCs) from the ear and peritoneum express the chymases RMCP-1 and -5, the tryptases RMCP-6, and -7 and the carboxypeptidase A. However, based on a large difference in the level of expression of RMCP-7, CTMCs of these two organs may be regarded as two separate subpopulations. RMCP-2 and the three closely related proteases of the RMCP-8 subfamily were identified as the major mucosal MC proteases in rat. In contrast to what has been reported for human MCs, no expression of cathepsin G or cathepsin G-like proteases was detected in any of the rat MC populations. To determine mRNA frequencies for the various proteases expressed by normal tissue MCs, an unamplified peritoneal MC cDNA library was screened with a panel of monospecific cDNA probes. These results showed that peritoneal MCs are highly specialized effector cells with mRNA frequencies for the major proteases in the range of several percent of the total mRNA pool.

  • 25.
    Magnúsdóttir, Elín Ingibjörg
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Grujic, Mirjana
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Bergman, Jessica
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Lagerström, Malin C.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Mouse mast cell tryptase and carboxypeptidase A3 play a protective role in itch induced by endothelin-1Manuscript (preprint) (Other academic)
  • 26.
    Magnúsdóttir, Elín Ingibjörg
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Grujic, Mirjana
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Lagerström, Malin C.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Compound 48/80-induced itch is attenuated by mouse mast cell tryptase and carboxypeptidase A3Manuscript (preprint) (Other academic)
  • 27.
    Magnúsdóttir, Elín Ingibjörg
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Genetics.
    Grujic, Mirjana
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Roers, Axel
    Univ Technol Dresden, Inst Immunol, Dresden, Germany.
    Hartmann, Karin
    Univ Lubeck, Dept Dermatol, Lubeck, Germany.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Lagerström, Malin C.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Genetics.
    Mouse mast cells and mast cell proteases do not play a significant role in acute tissue injury pain induced by formalin2018In: Molecular Pain, ISSN 1744-8069, E-ISSN 1744-8069, Vol. 14, article id 1744806918808161Article in journal (Refereed)
    Abstract [en]

    Subcutaneous formalin injections are used as a model for tissue injury-induced pain where formalin induces pain and inflammation indirectly by crosslinking proteins and directly through activation of the transient receptor potential A1 receptor on primary afferents. Activation of primary afferents leads to both central and peripheral release of neurotransmitters. Mast cells are found in close proximity to peripheral sensory nerve endings and express receptors for neurotransmitters released by the primary afferents, contributing to the neuro/immune interface. Mast cell proteases are found in large quantities within mast cell granules and are released continuously in small amounts and upon mast cell activation. They have a wide repertoire of proposed substrates, including Substance P and calcitonin gene-related peptide, but knowledge of their in vivo function is limited. We evaluated the role of mouse mast cell proteases (mMCPs) in tissue injury pain responses induced by formalin, using transgenic mice lacking either mMCP4, mMCP6, or carboxypeptidase A3 (CPA3), or mast cells in their entirety. Further, we investigated the role of mast cells in heat hypersensitivity following a nerve growth factor injection. No statistical difference was observed between the respective mast cell protease knockout lines and wild-type controls in the formalin test. Mast cell deficiency did not have an effect on formalin-induced nociceptive responses nor nerve growth factor-induced heat hypersensitivity. Our data thus show that mMCP4, mMCP6, and CPA3 as well as mast cells as a whole, do not play a significant role in the pain responses associated with acute tissue injury and inflammation in the formalin test. Our data also indicate that mast cells are not essential to heat hypersensitivity induced by nerve growth factor.

  • 28. Meen, Astri J
    et al.
    Drevon, Christian A
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Jenssen, Trond G
    Olstad, Ole Kristoffer
    Åbrink, Magnus
    Kolset, Svein O
    Serglycin protects against high fat diet-induced increase in serum LDL in mice2015In: Glycoconjugate Journal, ISSN 0282-0080, E-ISSN 1573-4986, Vol. 32, no 9, p. 703-714Article in journal (Refereed)
    Abstract [en]

    Proteoglycans have been implicated in regulation of lipoprotein metabolism. However, the impact of serglycin, the major proteoglycan expressed by many hematopoietic- and endothelial cells, on lipoprotein metabolism has not been explored. Here we addressed this issue by comparing several parameters of lipid metabolism in wild type (WT) and serglycin-/- mice, both at baseline and after feeding mice the Paigen diet. We show that, after feeding this diet for 20 weeks, serglycin deficient mice exhibited elevated concentrations of serum LDL in comparison with WT mice, thus suggesting that serglycin protects against an elevation of serum LDL levels after intake of a high-fat diet. Body weight increased in both groups, but only significantly in the serglycin-/- group. To explore the mechanism underlying this phenotype, genome-wide expression analysis was performed on liver tissues from WT and serglycin-/- mice. This analysis showed that serglycin-deficiency is associated with differential expression of numerous genes involved in the regulation of lipid metabolism, suggesting that the impact of serglycin on LDL levels may be related to effects at the gene expression level. In particular, several members of the CYP gene family were differently regulated in serglycin-/- compared with WT mice. Moreover, upstream regulator analysis suggested that several pro-inflammatory pathways, including the NFκB pathway, could contribute to the impact of serglycin on LDL. Hence, the elevation of serum LDL seen in serglycin-/- mice may be linked to dysregulated inflammatory responses. Taken together, our findings introduce serglycin as a novel player in processes that regulate lipid metabolism.

  • 29.
    Melo, Fabio R.
    et al.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, BMC, S-75123 Uppsala, Sweden..
    Grujic, Mirjana
    Spirkoski, Jane
    Calounova, Gabriela
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Serglycin Proteoglycan Promotes Apoptotic versus Necrotic Cell Death in Mast Cells2012In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 287, no 22, p. 18142-18152Article in journal (Refereed)
    Abstract [en]

    The mechanisms that govern whether a cell dies by apoptosis or necrosis are not fully understood. Here we show that serglycin, a secretory granule proteoglycan of hematopoietic cells, can have a major impact on this decision. Wild type and serglycin(-/-) mast cells were equally sensitive to a range of cell death-inducing regimens. However, whereas wild type mast cells underwent apoptotic cell death, serglycin(-/-) cells died predominantly by necrosis. Investigations of the underlying mechanism revealed that cell death was accompanied by leakage of secretory granule compounds into the cytosol and that the necrotic phenotype of serglycin(-/-) mast cells was linked to defective degradation of poly(ADP-ribose) polymerase-1. Cells lacking mouse mast cell protease 6, a major serglycin-associated protease, exhibited similar defects in apoptosis as observed in serglycin(-/-) cells, indicating that the pro-apoptotic function of serglycin is due to downstream effects of proteases that are complex-bound to serglycin. Together, these findings implicate serglycin in promoting apoptotic versus necrotic cell death.

  • 30.
    Melo, Fabio R.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Martin, Sebastin Santosh
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Sommerhoff, Christian P.
    Ludwig Maximilians Univ Munchen, Univ Hosp, Inst Lab Med, Munich, Germany.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Exosome-mediated uptake of mast cell tryptase into the nucleus of melanoma cells: a novel axis for regulating tumor cell proliferation and gene expression2019In: Cell Death and Disease, ISSN 2041-4889, E-ISSN 2041-4889, Vol. 10, article id 659Article in journal (Refereed)
    Abstract [en]

    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.

  • 31.
    Melo, Fabio R.
    et al.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75123 Uppsala, Sweden..
    Vita, Francesca
    Univ Trieste, Dept Life Sci, I-34127 Trieste, Italy..
    Berent-Maoz, Beata
    Hebrew Univ Jerusalem, Fac Med, Inst Drug Res, Dept Pharmacol, IL-91120 Jerusalem, Israel..
    Levi-Schaffer, Francesca
    Hebrew Univ Jerusalem, Fac Med, Inst Drug Res, Dept Pharmacol, IL-91120 Jerusalem, Israel..
    Zabucchi, Giuliano
    Univ Trieste, Dept Life Sci, I-34127 Trieste, Italy..
    Pejler, Gunnar
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75123 Uppsala, Sweden..
    Proteolytic Histone Modification by Mast Cell Tryptase, a Serglycin Proteoglycan-dependent Secretory Granule Protease2014In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 289, no 11, p. 7682-7690Article in journal (Refereed)
    Abstract [en]

    Background: Tryptase is a serglycin proteoglycan-dependent protease localized in mast cell granules. Results: Tryptase was found to degrade core histones, both during apoptosis and in viable cells. Conclusion: A serglycin-tryptase axis is identified as a novel mechanism for histone modification. Significance: Secretory granule compounds are implicated in the regulation of nuclear events. A hallmark feature of mast cells is their high content of cytoplasmic secretory granules filled with various preformed compounds, including proteases of tryptase-, chymase-, and carboxypeptidase A3 type that are electrostatically bound to serglycin proteoglycan. Apart from participating in extracellular processes, serglycin proteoglycan and one of its associated proteases, tryptase, are known to regulate cell death by promoting apoptosis over necrosis. Here we sought to outline the underlying mechanism and identify core histones as primary proteolytic targets for the serglycin-tryptase axis. During the cell death process, tryptase was found to relocalize from granules into the cytosol and nucleus, and it was found that the absence of tryptase was associated with a pronounced accumulation of core histones both in the cytosol and in the nucleus. Intriguingly, tryptase deficiency resulted in defective proteolytic modification of core histones even at baseline conditions, i.e. in the absence of cytotoxic agent, suggesting that tryptase has a homeostatic impact on nuclear events. Indeed, tryptase was found in the nucleus of viable cells and was shown to cleave core histones in their N-terminal tail. Moreover, it was shown that the absence of the serglycin-tryptase axis resulted in altered chromatin composition. Together, these findings implicate histone proteolysis through a secretory granule-derived serglycin-tryptase axis as a novel principle for histone modification, during both cell homeostasis and cell death.

  • 32.
    Melo, Fabio R.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Wallerman, Ola
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Paivandy, Aida
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Calounova, Gabriela
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Gustafson, Ann-Marie
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Sabari, Benjamin R.
    Rockefeller Univ, Lab Chromatin Biol & Epigenet, 1230 York Ave, New York, NY 10021 USA..
    Zabucchi, Giuliano
    Univ Trieste, Dept Life Sci, Trieste, Italy..
    Allis, C. David
    Rockefeller Univ, Lab Chromatin Biol & Epigenet, 1230 York Ave, New York, NY 10021 USA..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Tryptase-catalyzed core histone truncation: A novel epigenetic regulatory mechanism in mast cells2017In: Journal of Allergy and Clinical Immunology, ISSN 0091-6749, E-ISSN 1097-6825, Vol. 140, no 2, p. 474-485Article in journal (Refereed)
    Abstract [en]

    Background: Mast cells are key effector cells in allergic reactions. When activated to degranulate, they release a plethora of bioactive compounds from their secretory granules, including mast cell-restricted proteases such as tryptase. In a previous study, we showed that tryptase, in addition to its intragranular location, can be found within the nuclei of mast cells where it truncates core histones at their N-terminal ends. Objective: Considering that the N-terminal portions of the core histones constitute sites for posttranslational modifications of major epigenetic impact, we evaluated whether histone truncation by tryptase could have an impact on epigenetic events in mast cells. Methods: Mast cells were cultured from wild-type and tryptase null mice, followed by an assessment of their profile of epigenetic histone modifications and their phenotypic characteristics. Results: We show that tryptase truncates nucleosomal histone 3 and histone 2B (H2B) and that its absence results in accumulation of the epigenetic mark, lysine 5-acetylated H2B. Intriguingly, the accumulation of lysine 5-acetylated H2B was cell age-dependent and was associated with a profound upregulation of markers of non-mast cell lineages, loss of proliferative control, chromatin remodeling as well as extensive morphological alterations. Conclusions: These findings introduce tryptase-catalyzed histone clipping as a novel epigenetic regulatory mechanism, which in the mast cell context may be crucial for maintaining cellular identity.

  • 33.
    Melo, Fabio R
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Wernersson, Sara
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Induction of mast cell apoptosis by a novel secretory granule-mediated pathway.2015In: Methods in Molecular Biology, ISSN 1064-3745, E-ISSN 1940-6029, Vol. 1220, p. 325-37Article in journal (Refereed)
    Abstract [en]

    Mast cells (MCs) have detrimental functions in the context of numerous pathologies, and regimens aimed at neutralizing MCs or individual MC products can thus be of therapeutic value. One way to target MCs in disease is to selectively induce MC apoptosis, but there is so far no agent available that selectively induces apoptosis in MCs. Mast cells are heavily loaded with secretory granules containing large amounts of fully active proteases bound to serglycin proteoglycan. Damage to the secretory granules will thus lead to the release of serglycin-protease complexes into the cytosol. A potential consequence of this would be that the unleashed granular proteases cause apoptosis by proteolytic activation of proapoptotic compounds located in the cytosol. Indeed, we have recently found that MCs are highly sensitive to apoptosis induced by permeabilization of the secretory granules. In this chapter, we describe the methods used to study MC apoptosis induced by this novel, secretory granule-mediated pathway.

  • 34.
    Nyekel, Flavie Ngo
    et al.
    INSERM, UMRS 1149, Paris, France;CNRS, ERL8252, Paris, France;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.
    Pacreau, Emeline
    INSERM, UMRS 1149, Paris, France;CNRS, ERL8252, Paris, France;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.
    Benadda, Samira
    INSERM, UMRS 1149, Paris, France;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.
    Msallam, Rasha
    Univ Paris 05, Sorbonne Paris Cite, Fac Med, Inst Necker Enfants Malad,INSERM,U1151,CNRS,UMR82, Paris, France;ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore.
    Abrink, Magnus
    Swedish Univ Agr Sci, VHC, Dept Biomed Sci & Vet Publ Hlth, Immunol Sect, Uppsala, Sweden.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Davoust, Jean
    Univ Paris 05, Sorbonne Paris Cite, Fac Med, Inst Necker Enfants Malad,INSERM,U1151,CNRS,UMR82, Paris, France.
    Benhamou, Marc
    INSERM, UMRS 1149, Paris, France;CNRS, ERL8252, Paris, France;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.
    Charles, Nicolas
    INSERM, UMRS 1149, Paris, France;CNRS, ERL8252, Paris, France;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.
    Launay, Pierre
    INSERM, UMRS 1149, Paris, France;CNRS, ERL8252, Paris, France;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.
    Blank, Ulrich
    INSERM, UMRS 1149, Paris, France;CNRS, ERL8252, Paris, France;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.
    Gautier, Gregory
    INSERM, UMRS 1149, Paris, France;CNRS, ERL8252, Paris, France;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.
    Mast Cell Degranulation Exacerbates Skin Rejection by Enhancing Neutrophil Recruitment2018In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 9, article id 2690Article in journal (Refereed)
    Abstract [en]

    Recent evidences indicate an important role of tissue inflammatory responses by innate immune cells in allograft acceptance and survival. Here we investigated the role of mast cells (MC) in an acute male to female skin allograft rejection model using red MC and basophil (RMB) mice enabling conditional MC depletion. Kinetic analysis showed that MCs markedly accelerate skin rejection. They induced an early inflammatory response through degranulation and boosted local synthesis of KC, MIP-2, and TNF. This enhanced early neutrophil infiltration compared to a female-female graft-associated repair response. The uncontrolled neutrophil influx accelerated rejection as antibody-mediated depletion of neutrophils delayed skin rejection. Administration of cromolyn, a MC stabilizer and to a lesser extent ketotifen, a histamine type I receptor antagonist, and absence of MCPT4 chymase also delayed graft rejection. Together our data indicate that mediators contained in secretory granules of MC promote an inflammatory response with enhanced neutrophil infiltration that accelerate graft rejection.

  • 35.
    Paivandy, Aida
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Eriksson, Jens
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Melo, Fabio R.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Sellin, Mikael E.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Lysosomotropic challenge of mast cells causes intra-granular reactive oxygen species production2019In: CELL DEATH DISCOVERY, ISSN 2058-7716, Vol. 5, article id 95Article in journal (Refereed)
    Abstract [en]

    Mast cells contribute to the pathology of allergic and other disorders. Strategies to interfere with harmful mast cell-related activities are therefore warranted. Previously we established a principle for inducing selective apoptosis of mast cells, by the use of lysosomotropic agents that cause secretory granule permeabilization, leading to production of reactive oxygen species (ROS). However, the mechanism of ROS production has not been known. Here we addressed this issue. Live microscopy analysis showed that the secretory granules comprise major subcellular compartments for ROS production in response to mefloquine. As further signs for the primary involvement of secretory granules, both ROS production and cell death was blunted in mast cells lacking serglycin, a secretory granule-restricted proteoglycan. Inhibition of granule acidification caused an essentially complete blockade of granule permeabilization, ROS production and cell death in response to mefloquine. ROS production was also attenuated in the presence of an iron chelator, and after inhibition of either granzyme B or the ERK1/2 MAP kinase signaling pathway. Together, our findings reveal that the mast cell secretory granules constitute major sites for ROS production in mast cells subjected to lysosomotropic challenge. Moreover, this study reveals a central role for granule acidification in ROS generation and the pro-apoptotic response triggered downstream of secretory granule permeabilization.

  • 36.
    Paivandy, Aida
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Sandelin, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Igelström, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Physiotherapy.
    Landelius, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Thoracic Surgery.
    Janson, Christer
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Lung- allergy- and sleep research.
    Melo, Fabio R.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Department of Anatomy, Physiology and Biochemistry, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Induction of Human Lung Mast Cell Apoptosis by Granule Permeabilization: A Novel Approach for Targeting Mast Cells2017In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 8, article id 1645Article in journal (Refereed)
    Abstract [en]

    Mast cells are implicated as detrimental players in inflammatory lung diseases, particularly asthma. Mast cells respond to activating stimuli by releasing a wide panel of pro-inflammatory compounds that can contribute profoundly to the pathology, and there is currently an unmet need for strategies that efficiently ameliorate harmful effects of mast cells under such conditions. Here, we sought to evaluate a novel concept for targeting human lung mast cells, by assessing the possibility of selectively depleting the lung mast cells by induction of apoptosis. For this purpose, we used lysosomotropic agents, i.e., compounds that are known to permeabilize the secretory granules of mast cells, thereby releasing the contents of the granules into the cytosol. Either intact human lung tissue, purified human lung mast cells or mixed populations of human lung cells were incubated with the lysosomotropic agents mefloquine or siramesine, followed by measurement of apoptosis, reactive oxygen species (ROS) production, and release of cytokines. We show that human lung mast cells were highly susceptible to apoptosis induced by this strategy, whereas other cell populations of the lung were largely refractory. Moreover, we demonstrate that apoptosis induced by this mode is dependent on the production of ROS and that the treatment of lung tissue with lysosomotropic agents causes a decrease in the release of pathogenic cytokines. We conclude that selective apoptosis of human lung mast cells can be accomplished by administration of lysosomotropic agents, thus introducing the possibility of using such drugs as novel therapeutics in the treatment of inflammatory lung disorders such as asthma.

  • 37.
    Palm, Anna-Karin E.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Chemical Biology.
    Garcia-Faroldi, Gianni
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Lundberg, Marcus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Kleinau, Sandra
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Chemical Biology.
    Activated mast cells promote differentiation of B cells into effector cells2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, article id 20531Article in journal (Refereed)
    Abstract [en]

    Based on the known accumulation of mast cells (MCs) in B cell-dependent inflammatory diseases, including rheumatoid arthritis, we hypothesized that MCs directly modulate B cells. We show here that degranulated, and to a lesser extent naive or IgE-sensitized, MCs activate both naive and B cell receptor-activated B cells. This was shown by increased proliferation, blast formation, and expression of CD19, MHC class II and CD86 in the B cells. Further, MCs stimulated the secretion of IgM and IgG in IgM(+) B cells, indicating that MCs can induce class-switch recombination in B cells. We also show that coculture of MCs with B cells promotes surface expression of L-selectin, a homing receptor, on the B cells. The effects of MCs on B cells were partly dependent on cell-cell contact and both follicular and marginal zone B cells could be activated by MCs. Our findings suggest that degranulated MCs support optimal activation of B cells, a finding that is in line with in vivo studies showing that MCs frequently degranulate in the context of B-cell driven pathologies such as arthritis. Together, our findings show that MCs have the capacity to differentiate B cells to effector cells.

  • 38.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    MAST CELL GRANULES: ARMED WITH HISTAMINE AND FRIENDS2015In: Inflammation Research, ISSN 1023-3830, E-ISSN 1420-908X, Vol. 64, no S1, p. S8-S8Article in journal (Other academic)
  • 39.
    Pejler, Gunnar
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Frisk, Jun Mei Hu
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Sjöström, Daniel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Paivandy, Aida
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Öhrvik, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Acidic pH is essential for maintaining mast cell secretory granule homeostasis2017In: Cell Death and Disease, ISSN 2041-4889, E-ISSN 2041-4889, Vol. 8, article id e2785Article in journal (Refereed)
    Abstract [en]

    It has been recognized for a long time that the secretory granules of mast cells are acidic, but the functional importance of maintaining an acidic pH in the mast cell granules is not fully understood. Here we addressed this issue by examining the effects of raising the pH of the mast cell secretory granules. Mast cells were incubated with bafilomycin A1, an inhibitor of the vacuolar-type ATPase proton pump. Supporting a role of vacuolar-type ATPase in mast cell granule acidification, bafilomycin A1 treatment caused a robust increase in granule pH. This was accompanied by marked effects on mast cell granules, including swelling and acquisition of vacuole-like morphology. Moreover, bafilomycin A1 caused extensive, yet selective effects on the granule content. These included aberrant processing of pro-carboxypeptidase A3 and a reduction in the level of intracellular histamine, the latter being accompanied by an increase in extracellular histamine. In contrast, the storage of beta-hexosaminidase, a prototype lysosomal hydrolase known to be stored in mast cell granules, was not affected by abrogation of granule acidification. Moreover, bafilomycin A1 caused a reduction of tryptase enzymatic activity and appearance of tryptase degradation products. Tryptase inhibition prevented the formation of such degradation products, suggesting that the pH elevation causes tryptase to undergo autoproteolysis. Taken together, our findings reveal that mast cell secretory granule homeostasis is critically dependent on an acidic milieu.

  • 40.
    Pons, Maguelonne
    et al.
    INSERM, UMRS 1149, Paris, France.;CNRS, ERL8252, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Hop Robert Debre, APHP,Dept Pediat Surg & Urol, Paris, France..
    Ali, Liza
    INSERM, UMRS 1149, Paris, France.;CNRS, ERL8252, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Hop Robert Debre, APHP,Dept Pediat Surg & Urol, Paris, France..
    Beghdadi, Walid
    INSERM, UMRS 1149, Paris, France.;CNRS, ERL8252, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France..
    Danelli, Luca
    CNRS, ERL8252, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France..
    Alison, Marianne
    Univ Paris Diderot, Sorbonne Paris Cite, Hop Robert Debre, APHP,Dept Pediat Radiol, Paris, France..
    Madjene, Lydia Celia
    INSERM, UMRS 1149, Paris, France.;CNRS, ERL8252, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France..
    Calvo, Jessica
    Univ Paris Diderot, Sorbonne Paris Cite, Hop Robert Debre, APHP,Dept Pathol, Paris, France..
    Claver, Julien
    INSERM, UMRS 1149, Paris, France.;CNRS, ERL8252, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France..
    Vibhushan, Shamila
    INSERM, UMRS 1149, Paris, France.;CNRS, ERL8252, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France..
    Åbrink, Magnus
    Swedish Univ Agr Sci, Dept Biomed Sci & Vet Publ Hlth, Sect Immunol, Uppsala, Sweden..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Poli-Merol, Marie-Laurence
    Univ Reims Champagne Ardennes, Amer Mem Hosp, Pediat Surg Unit, Reims, France..
    Peuchmaur, Michel
    Univ Paris Diderot, Sorbonne Paris Cite, Hop Robert Debre, APHP,Dept Pathol, Paris, France..
    El Ghoneimi, Alaa
    INSERM, UMRS 1149, Paris, France.;CNRS, ERL8252, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Hop Robert Debre, APHP,Dept Pediat Surg & Urol, Paris, France..
    Blank, Ulrich
    INSERM, UMRS 1149, Paris, France.;CNRS, ERL8252, Paris, France.;Univ Paris Diderot, Sorbonne Paris Cite, Lab Excellence INFLAMEX, Paris, France..
    Mast cells and McPT4 chymase Promote renal impairment after Partial Ureteral Obstruction2017In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 8, article id 450Article in journal (Refereed)
    Abstract [en]

    Obstructive nephropathy constitutes a major cause of pediatric renal progressive disease. The mechanisms leading to disease progression are still poorly understood. Kidney fibrotic lesions are reproduced using a model of partial unilateral ureteral obstruction (pUUO) in newborn mice. Based on data showing significant mast cell (MC) infiltration in patients, we investigated the role of MC and murine MCPT4, a MC-released chymase, in pUUO using MC- (W-sh/sh), MCPT4-deficient (Mcpt4(-/-)), and wild-type (WT) mice. Measurement of kidney length and volume by magnetic resonance imaging (MRI) as well as postmortem kidney weight revealed hypotrophy of operated right kidneys (RKs) and compensatory hypertrophy of left kidneys. Differences between kidneys were major for WT, minimal for Wsh/sh, and intermediate for Mcpt4(-/-) mice. Fibrosis development was focal and increased only in WT-obstructed kidneys. No differences were noticed for local inflammatory responses, but serum CCL2 was significantly higher in WT versus Mcpt4(-/-) and Wsh/sh mice. Alpha-smooth muscle actin (alpha SMA) expression, a marker of epithelial-mesenchymal transition (EMT), was high in WT, minimal for W-sh/sh, and intermediate for Mcpt4(-/-) RK. Supernatants of activated MC induced aSMA in co-culture experiments with proximal tubular epithelial cells. Our results support a role of MC in EMT and parenchyma lesions after pUUO involving, at least partly, MCPT4 chymase. They confirm the importance of morphologic impairment evaluation by MRI in pUUO.

  • 41.
    Ronnberg, Elin
    et al.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75123 Uppsala, Sweden..
    Johnzon, Carl-Fredrik
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75123 Uppsala, Sweden..
    Calounova, Gabriela
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75123 Uppsala, Sweden..
    Faroldi, Gianni Garcia
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75123 Uppsala, Sweden..
    Grujic, Mirjana
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75123 Uppsala, Sweden..
    Hartmann, Karin
    Univ Hosp Cologne, Dept Dermatol, Cologne, Germany..
    Roers, Axel
    Tech Univ Dresden, Inst Immunol, Dresden, Germany..
    Guss, Bengt
    Swedish Univ Agr Sci, Dept Microbiol, S-75123 Uppsala, Sweden..
    Lundequist, Anders
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75123 Uppsala, Sweden..
    Pejler, Gunnar
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, S-75123 Uppsala, Sweden..
    Mast cells are activated by Staphylococcus aureus in vitro but do not influence the outcome of intraperitoneal S. aureus infection in vivo2014In: Immunology, ISSN 0019-2805, E-ISSN 1365-2567, Vol. 143, no 2, p. 155-163Article in journal (Refereed)
    Abstract [en]

    Staphylococcus aureus is a major pathogen that can cause a broad spectrum of serious infections including skin infections, pneumonia and sepsis. Peritoneal mast cells have been implicated in the host response towards various bacterial insults and to provide mechanistic insight into the role of mast cells in intraperitoneal bacterial infection we here studied the global effects of S. aureus on mast cell gene expression. After co-culture of peritoneal mast cells with live S. aureus we found by gene array analysis that they up-regulate a number of genes. Many of these corresponded to pro-inflammatory cytokines, including interleukin-3, interleukin-13 and tumour necrosis factor-a. The cytokine induction in response to S. aureus was confirmed by ELISA. To study the role of peritoneal mast cells during in vivo infection with S. aureus we used newly developed Mcpt5-Cre(+) x R-DTA mice in which mast cell deficiency is independent of c-Kit. This is in contrast to previous studies in which an impact of mast cells on bacterial infection has been proposed based on the use of mice whose mast cell deficiency is a consequence of defective c-Kit signalling. Staphylococcus aureus was injected intraperitoneally into mast-cell-deficient Mcpt5-Cre(+) x R-DTA mice using littermate mast-cell-sufficient mice as controls. We did not observe any difference between mast-cell-deficient and control mice with regard to weight loss, bacterial clearance, inflammation or cytokine production. We conclude that, despite peritoneal mast cells being activated by S. aureus in vitro, they do not influence the in vivo manifestations of intraperitoneal S. aureus infection.

  • 42. Semaan, Walid
    et al.
    Desbiens, Louisane
    Houde, Martin
    Labonte, Julie
    Gagnon, Hugo
    Yamamoto, Daisuke
    Takai, Shinji
    Laidlaw, Tanya
    Bkaily, Ghassan
    Schwertani, Fadel
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Levesque, Christine
    Desjardins, Roxane
    Day, Robert
    D'Orleans-Juste, Pedro
    Chymase inhibitor-sensitive synthesis of endothelin-1 (1-31) by recombinant mouse mast cell protease 4 and human chymase2015In: Biochemical Pharmacology, ISSN 0006-2952, E-ISSN 1356-1839, Vol. 94, no 2, p. 91-100Article in journal (Refereed)
    Abstract [en]

    Important structural differences imply that human and mouse mast cell chymases may differ with respect to their enzymatic properties. We compared in this study the catalytic efficiencies of recombinant human chymase (rCMA1) and its functional murine homologue recombinant mouse mast cell protease-4 (rmMCP-4) toward a fluorogenic chymase substrate (Suc-Ala-Ala-Pro-Phe-7-amino-4-methylcoumarin (AMC) and by their ability to convert Big-endothelin (ET)-1 into ET-1 (1-31) using a LC/MS/MS system. Activities toward a fluorogenic substrate (Suc-Leu-Leu-Val-Tyr-AMC) and Big ET-1 were also measured in extracts from mouse peritoneal mast cells, LUVA human mast cell-like cells and human aortas. The specificity of these activities was assessed with the chymase inhibitor TY-51469 (2-[4-(5-fluoro-3-methylbenzo[b]thiophen-2-yl)sulfonamido-3-methanesulfonyl-phenyl]thiazole-4-carboxylic acid). For similar affinities, rmMCP-4 showed a higher activity toward the fluorogenic substrate and a higher ability to process Big ET-1 as compared to recombinant CMA1 (chymase activity (k(cat)/K-M in mu M-1 s(-1)): 2.29 x 10(-4) vs. 6.41 x 10(-6); ET-1 (1-31) production: 2.19 x 10(-3) vs. 6.57 x 10(-5)), and both of these activities of mouse and human chymase were sensitive to TY-51469. Furthermore, extracts from mouse peritoneal mast cells, LUVA cells and human aorta homogenates contained processing activities toward the fluorogenic chymase substrate as well as Big ET-1, all of which were sensitive to TY-51469. Finally, the pressor responses to Big ET-1 but not to ET-1 were significantly reduced in conscious and free moving mMCP-4 KO mice when compared to wild type congeners. Our results suggest that both mouse and human chymases have potent ET-1 (1-31)-producing abilities, with the murine isoform being more efficient.

  • 43.
    Shubin, Nicholas J.
    et al.
    Seattle Childrens Res Inst, Ctr Immun & Immunotherapies, 1900 9th Ave,Rm 721, Seattle, WA 98101 USA..
    Glukhova, Veronika A.
    Seattle Childrens Res Inst, Ctr Immun & Immunotherapies, 1900 9th Ave,Rm 721, Seattle, WA 98101 USA..
    Clauson, Morgan
    Seattle Childrens Res Inst, Ctr Immun & Immunotherapies, 1900 9th Ave,Rm 721, Seattle, WA 98101 USA..
    Truong, Phuong
    Seattle Childrens Res Inst, Ctr Immun & Immunotherapies, 1900 9th Ave,Rm 721, Seattle, WA 98101 USA..
    Abrink, Magnus
    Swedish Univ Agr Sci, Dept Biomed Sci & Vet Publ Hlth, Uppsala, Sweden..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    White, Nathan J.
    Univ Washington, Dept Med, Div Emergency Med, Seattle, WA USA..
    Deutsch, Gail H.
    Seattle Childrens Res Inst, Dept Labs, Seattle, WA USA..
    Reeves, Stephen R.
    Seattle Childrens Res Inst, Ctr Immun & Immunotherapies, 1900 9th Ave,Rm 721, Seattle, WA 98101 USA.;Univ Washington, Dept Pediat, Seattle, WA 98195 USA..
    Vaisar, Tomas
    Univ Washington, Dept Med, Div Metab Endocrinol & Nutr, Seattle, WA USA..
    James, Richard G.
    Seattle Childrens Res Inst, Ctr Immun & Immunotherapies, 1900 9th Ave,Rm 721, Seattle, WA 98101 USA..
    Piliponsky, Adrian M.
    Seattle Childrens Res Inst, Ctr Immun & Immunotherapies, 1900 9th Ave,Rm 721, Seattle, WA 98101 USA.;Univ Washington, Dept Pediat, Seattle, WA 98195 USA..
    Proteome analysis of mast cell releasates reveals a role for chymase in the regulation of coagulation factor XIIIA levels via proteolytic degradation2017In: Journal of Allergy and Clinical Immunology, ISSN 0091-6749, E-ISSN 1097-6825, Vol. 139, no 1, p. 323-334Article in journal (Refereed)
    Abstract [en]

    Background: Mast cells are significantly involved in IgE-mediated allergic reactions; however, their roles in health and disease are incompletely understood. Objective: We aimed to define the proteome contained in mast cell releasates on activation to better understand the factors secreted by mast cells that are relevant to the contribution of mast cells in diseases. Methods: Bone marrow-derived cultured mast cells (BMCMCs) and peritoneal cell-derived mast cells were used as "surrogates'' for mucosal and connective tissue mast cells, respectively, and their releasate proteomes were analyzed by mass spectrometry. Results: Our studies showed that BMCMCs and peritoneal cell-derived mast cells produced substantially different releasates following IgE-mediated activation. Moreover, we observed that the transglutaminase coagulation factor XIIIA (FXIIIA) was one of the most abundant proteins contained in the BMCMC releasates. Mast cell-deficient mice exhibited increased FXIIIA plasma and activity levels as well as reduced bleeding times, indicating that mast cells are more efficient in their ability to downregulate FXIIIA than in contributing to its amounts and functions in homeostatic conditions. We found that human chymase and mouse mast cell protease-4 (the mouse homologue of human chymase) had the ability to reduce FXIIIA levels and function via proteolytic degradation. Moreover, we found that chymase deficiency led to increased FXIIIA amounts and activity, as well as reduced bleeding times in homeostatic conditions and during sepsis. Conclusions: Our study indicates that the mast cell protease content can shape its releasate proteome. Moreover, we found that chymase plays an important role in the regulation of FXIIIA via proteolytic degradation.

  • 44.
    Succar, Julien
    et al.
    Harvard Med Sch, Brigham & Womens Hosp, Dept Surg, Tissue Engn & Wound Healing Lab,Div Plast Surg, 75 Francis St, Boston, MA 02115 USA.
    Giatsidis, Giorgio
    Harvard Med Sch, Brigham & Womens Hosp, Dept Surg, Tissue Engn & Wound Healing Lab,Div Plast Surg, 75 Francis St, Boston, MA 02115 USA.
    Yu, Nanze
    Harvard Med Sch, Brigham & Womens Hosp, Dept Surg, Tissue Engn & Wound Healing Lab,Div Plast Surg, 75 Francis St, Boston, MA 02115 USA.
    Hassan, Kazi
    Harvard Med Sch, Brigham & Womens Hosp, Dept Surg, Tissue Engn & Wound Healing Lab,Div Plast Surg, 75 Francis St, Boston, MA 02115 USA.
    Khouri, Roger, Jr.
    Harvard Med Sch, Brigham & Womens Hosp, Dept Surg, Tissue Engn & Wound Healing Lab,Div Plast Surg, 75 Francis St, Boston, MA 02115 USA.
    Gurish, Michael F.
    Harvard Med Sch, Brigham & Womens Hosp, Div Rheumatol Immunol & Allergy, Human Immunol Ctr, Boston, MA 02115 USA.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Abrink, Magnus
    Swedish Univ Agr Sci, Dept Biomed Sci & Vet Publ Hlth, Sect Immunol, Uppsala, Sweden.
    Orgill, Dennis Paul
    Harvard Med Sch, Brigham & Womens Hosp, Dept Surg, Tissue Engn & Wound Healing Lab,Div Plast Surg, 75 Francis St, Boston, MA 02115 USA.
    Mouse Mast Cell Protease-4 Recruits Leukocytes in the Inflammatory Phase of Surgically Wounded Skin2019In: ADVANCES IN WOUND CARE, ISSN 2162-1918, Vol. 8, no 10, p. 469-475Article in journal (Refereed)
    Abstract [en]

    Objective: Mouse mast cell protease-4 (mMCP-4, also known as chymase) has both pro- and anti-inflammatory roles depending on the disease model. However, its effects have not been studied in surgically wounded skin. Given the significant clinical applications of modulating the inflammatory response in wound healing, we examined the role of mMCP-4 and the effect of its inhibitor chymostatin on leukocyte and polymorphonuclear cell (PMN) recruitment in our skin model. Approach: Recruitment was assessed on day-1 postwounding of three groups of mice (n = 10 each): mMCP-4 null mice, wild-type (WT) mice treated with the mMCP-4 inhibitor chymostatin, and WT with no other intervention. Leukocytes were stained with CD-45 cell marker, and PMN cells were stained with chloroacetate esterase. Results: The WT mice had 27 +/- 9 leukocytes per field compared with 11 +/- 6 for the mMCP-4 nulls, a decrease of 60% (p = 0.03), whereas the chymostatin-injected group had a count comparable with the uninjected WT controls at 24 +/- 9. The WT group had a PMN count of 96 +/- 12 cells, compared with just 24 +/- 8 in the mMCP-4 null group, a decrease of 75% (p = 0.001), whereas the chymostatin-treated group had 60 +/- 18 cells, a decrease of 38% compared with the WT group (p = 0.03). Innovation: We showed that the inflammatory process can be influenced by impeding the arrival of PMNs into the surgically injured site using the mMCP-4 inhibitor chymostatin. Conclusion: Chymase contributes to the recruitment of white blood cells in surgically wounded skin.

  • 45.
    Sutton, Vivien R.
    et al.
    Peter MacCallum Canc Ctr, Canc Cell Death Killer Cell Biol Labs, East Melbourne, Vic, Australia..
    Brennan, Amelia J.
    Peter MacCallum Canc Ctr, Canc Cell Death Killer Cell Biol Labs, East Melbourne, Vic, Australia..
    Ellis, Sarah
    Peter MacCallum Canc Ctr, Microscopy & Histol, East Melbourne, Vic, Australia..
    Danne, Jill
    Peter MacCallum Canc Ctr, Microscopy & Histol, East Melbourne, Vic, Australia..
    Thia, Kevin
    Peter MacCallum Canc Ctr, Canc Cell Death Killer Cell Biol Labs, East Melbourne, Vic, Australia..
    Jenkins, Misty R.
    Peter MacCallum Canc Ctr, Canc Cell Death Killer Cell Biol Labs, East Melbourne, Vic, Australia..
    Voskoboinik, Ilia
    Peter MacCallum Canc Ctr, Canc Cell Death Killer Cell Biol Labs, East Melbourne, Vic, Australia..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Johnstone, Ricky W.
    Peter MacCallum Canc Ctr, Canc Cell Death Killer Cell Biol Labs, East Melbourne, Vic, Australia.;Univ Melbourne, Sir Peter MacCallum Dept Oncol, Melbourne, Vic 3010, Australia..
    Andrews, Daniel M.
    Peter MacCallum Canc Ctr, Canc Cell Death Killer Cell Biol Labs, East Melbourne, Vic, Australia..
    Trapani, Joseph A.
    Peter MacCallum Canc Ctr, Canc Cell Death Killer Cell Biol Labs, East Melbourne, Vic, Australia.;Univ Melbourne, Sir Peter MacCallum Dept Oncol, Melbourne, Vic 3010, Australia..
    Serglycin determines secretory granule repertoire and regulates natural killer cell and cytotoxic T lymphocyte cytotoxicity2016In: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 283, no 5, p. 947-961Article in journal (Refereed)
    Abstract [en]

    The anionic proteoglycan serglycin is a major constituent of secretory granules in cytotoxic T lymphocyte (CTL)/natural killer (NK) cells, and is proposed to promote the safe storage of the mostly cationic granule toxins, granzymes and perforin. Despite the extensive defects of mast cell function reported in serglycin gene-disrupted mice, no comprehensive study of physiologically relevant CTL/NK cell populations has been reported. We show that the cytotoxicity of serglycin-deficient CTL and NK cells is severely compromised but can be partly compensated in both cell types when they become activated. Reduced intracellular granzyme B levels were noted, particularly in CD27(+)CD11b(+) mature NK cells, whereas serglycin(-/-) TCR-transgenic (OTI) CD8 T cells also had reduced perforin stores. Culture supernatants from serglycin(-/-) OTI T cells and interleukin-2-activated NK contained increased granzyme B, linking reduced storage with heightened export. By contrast, granzyme A was not significantly reduced in cells lacking serglycin, indicating differentially regulated trafficking and/or storage for the two granzymes. A quantitative analysis of different granule classes by transmission electronmicroscopy showed a selective loss of dense-core granules in serglycin(-/-) CD8(+) CTLs, although other granule types were maintained quantitatively. The findings of the present study show that serglycin plays a critical role in the maturation of dense-core cytotoxic granules in cytotoxic lymphocytes and the trafficking and storage of perforin and granzyme B, whereas granzyme A is unaffected. The skewed retention of cytotoxic effector molecules markedly reduces CTL/NK cell cytotoxicity, although this is partly compensated for as a result of activating the cells by physiological means.

  • 46.
    Tejada, Thor
    et al.
    Emory Univ, Sch Med, Dept Med Cardiol, Atlanta, GA 30322 USA..
    Tan, Lin
    Emory Univ, Sch Med, Dept Med Cardiol, Atlanta, GA 30322 USA..
    Torres, Rebecca A.
    Emory Univ, Sch Med, Dept Med Cardiol, Atlanta, GA 30322 USA..
    Calvert, John W.
    Emory Univ, Sch Med, Dept Surg, Carlyle Fraser Heart Ctr, Atlanta, GA 30322 USA..
    Lambert, Jonathan P.
    Emory Univ, Sch Med, Dept Surg, Carlyle Fraser Heart Ctr, Atlanta, GA 30322 USA..
    Zaidi, Madiha
    Emory Univ, Sch Med, Dept Med Cardiol, Atlanta, GA 30322 USA..
    Husain, Murtaza
    Emory Univ, Sch Med, Dept Med Cardiol, Atlanta, GA 30322 USA..
    Berce, Maria D.
    Emory Univ, Sch Med, Dept Med Cardiol, Atlanta, GA 30322 USA..
    Naib, Hussain
    Emory Univ, Sch Med, Dept Med Cardiol, Atlanta, GA 30322 USA..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Abrink, Magnus
    Swedish Univ Agr Sci, Dept Biomed Sci & Vet Publ Hlth, Uppsala, Sweden..
    Graham, Robert M.
    Victor Chang Cardiac Res Inst, Sydney, NSW 2010, Australia..
    Lefer, David J.
    Emory Univ, Sch Med, Dept Surg, Carlyle Fraser Heart Ctr, Atlanta, GA 30322 USA..
    Naqvi, Nawazish
    Emory Univ, Sch Med, Dept Med Cardiol, Atlanta, GA 30322 USA..
    Husain, Ahsan
    Emory Univ, Sch Med, Dept Med Cardiol, Atlanta, GA 30322 USA..
    IGF-1 degradation by mouse mast cell protease 4 promotes cell death and adverse cardiac remodeling days after a myocardial infarction2016In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 113, no 25, p. 6949-6954Article in journal (Refereed)
    Abstract [en]

    Heart disease is a leading cause of death in adults. Here, we show that a few days after coronary artery ligation and reperfusion, the ischemia-injured heart elaborates the cardioprotective polypeptide, insulin-like growth factor-1 (IGF-1), which activates IGF-1 receptor prosurvival signaling and improves cardiac left ventricular systolic function. However, this signaling is antagonized by the chymase, mouse mast cell protease 4 (MMCP-4), which degrades IGF-1. We found that deletion of the gene encoding MMCP-4 (Mcpt4), markedly reduced late, but not early, infarct size by suppressing IGF-1 degradation and, consequently, diminished cardiac dysfunction and adverse structural remodeling. Our findings represent the first demonstration to our knowledge of tissue IGF-1 regulation through proteolytic degradation and suggest that chymase inhibition may be a viable therapeutic approach to enhance late cardioprotection in post-ischemic heart disease.

  • 47.
    Vangansewinkel, Tim
    et al.
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium..
    Geurts, Nathalie
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium..
    Quanten, Kirsten
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium..
    Nelissen, Sofie
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium..
    Lemmens, Stefanie
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium..
    Geboes, Lies
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium..
    Dooley, Dearbhaile
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium..
    Vidal, Pia M.
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium.;Univ Hlth Network, Div Genet & Dev, Krembil Neurosci Ctr, Toronto, ON, Canada..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden..
    Hendrix, Sven
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium..
    Mast cells promote scar remodeling and functional recovery after spinal cord injury via mouse mast cell protease 62016In: The FASEB Journal, ISSN 0892-6638, E-ISSN 1530-6860, Vol. 30, no 5, p. 2040-2057Article in journal (Refereed)
    Abstract [en]

    An important barrier for axon regeneration and recovery after traumatic spinal cord injury (SCI) is attributed to the scar that is formed at the lesion site. Here, we investigated the effect of mouse mast cell protease (mMCP) 6, a mast cell (MC)-specific tryptase, on scarring and functional recovery after a spinal cord hemisection injury. Functional recovery was significantly impaired in both MC-deficient and mMCP6-knockout (mMCP6(-/-)) mice after SCI compared with wild-type control mice. This decrease in locomotor performance was associated with an increased lesion size and excessive scarring at the injury site. Axon growth-inhibitory chondroitin sulfate proteoglycans and the extracellular matrix components fibronectin, laminin, and collagen IV were significantly up-regulated in MC-deficient and mMCP6(-/-) mice, with an increase in scar volume between 23 and 32%. A degradation assay revealed that mMCP6 directly cleaves fibronectin and collagen IV in vitro. In addition, gene expression levels of the scar components fibronectin, aggrecan, and collagen IV were increased up to 6.8-fold in mMCP6(-/-) mice in the subacute phase after injury. These data indicate that endogenous mMCP6 has scar-suppressing properties after SCI via indirect cleavage of axon growth-inhibitory scar components and alteration of the gene expression profile of these factors.-Vangansewinkel, T., Geurts, N., Quanten, K., Nelissen, S., Lemmens, S., Geboes, L., Dooley, D., Vidal, P. M., Pejler, G., Hendrix, S. Mast cells promote scar remodeling and functional recovery after spinal cord injury via mouse mast cell protease 6. www.fasebj.org

  • 48.
    Vangansewinkel, Tim
    et al.
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium.
    Lemmens, Stefanie
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium.
    Geurts, Nathalie
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium.
    Quanten, Kirsten
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium.
    Dooley, Dearbhaile
    Univ Coll Dublin, Sch Med, Hlth Sci Ctr, Dublin, Ireland.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Hendrix, Sven
    Hasselt Univ, Biomed Res Inst, Dept Morphol, Diepenbeek, Belgium.
    Mouse mast cell protease 4 suppresses scar formation after traumatic spinal cord injury2019In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 9, article id 3715Article in journal (Refereed)
    Abstract [en]

    Spinal cord injury (SCI) triggers the formation of a glial and fibrotic scar, which creates a major barrier for neuroregenerative processes. Previous findings indicate that mast cells (MCs) protect the spinal cord after mechanical damage by suppressing detrimental inflammatory processes via mouse mast cell protease 4 (mMCP4), a MC-specific chymase. In addition to these immunomodulatory properties, mMCP4 also plays an important role in tissue remodeling and extracellular matrix degradation. Therefore, we have investigated the effects of mMCP4 on the scarring response after SCI. We demonstrate that the decrease in locomotor performance in mMCP4(-/-) mice is correlated with excessive scar formation at the lesion. The expression of axon-growth inhibitory chondroitin sulfate proteoglycans was dramatically increased in the perilesional area in mMCP4(-/-) mice compared to wild type mice. Moreover, the fibronectin-, laminin-, and collagen IV-positive scar was significantly enlarged in mMCP4(-/-) mice at the lesion center. A degradation assay revealed that mMCP4 directly cleaves collagen IV in vitro. On the gene expression level, neurocan and GFAP were significantly higher in the mMCP4(-/-) group at day 2 and day 28 after injury respectively. In contrast, the expression of fibronectin and collagen IV was reduced in mMCP4(-/-) mice compared to WT mice at day 7 after SCI. In conclusion, our data show that mMCP4 modulates scar development after SCI by altering the gene and protein expression patterns of key scar factors in vivo. Therefore, we suggest a new mechanism via which endogenous mMCP4 can improve recovery after SCI.

  • 49.
    von Beek, Christopher
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Waern, Ida
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Eriksson, Jens
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Melo, Fabio R.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Robinson, Carl
    Anim Hlth Trust, Dept Bacteriol, Newmarket, Suffolk, England.
    Waller, Andrew S.
    Anim Hlth Trust, Dept Bacteriol, Newmarket, Suffolk, England.
    Sellin, Mikael E.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Guss, Bengt
    Swedish Univ Agr Sci, Dept Biomed Sci & Vet Publ Hlth, Uppsala, Sweden.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Sweden;Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Streptococcal sagA activates a proinflammatory response in mast cells by a sublytic mechanism2019In: Cellular Microbiology, ISSN 1462-5814, E-ISSN 1462-5822, Vol. 21, no 9, article id e13064Article in journal (Refereed)
    Abstract [en]

    Mast cells are implicated in the innate proinflammatory immune defence against bacterial insult, but the mechanisms through which mast cells respond to bacterial encounter are poorly defined. Here, we addressed this issue and show that mast cells respond vividly to wild type Streptococcus equi by up-regulating a panel of proinflammatory genes and by secreting proinflammatory cytokines. However, this response was completely abrogated when the bacteria lacked expression of sagA, whereas the lack of a range of other potential virulence genes (seeH, seeI, seeL, seeM, hasA, seM, aroB, pyrC, and recA) had no effect on the amplitude of the mast cell responses. The sagA gene encodes streptolysin S, a lytic toxin, and we next showed that the wild type strain but not a sagA-deficient mutant induced lysis of mast cells. To investigate whether host cell membrane perturbation per se could play a role in the activation of the proinflammatory response, we evaluated the effects of detergent- and pneumolysin-dependent lysis on mast cells. Indeed, exposure of mast cells to sublytic concentrations of all these agents resulted in cytokine responses of similar amplitudes as those caused by wild type streptococci. This suggests that sublytic membrane perturbation is sufficient to trigger full-blown proinflammatory signalling in mast cells. Subsequent analysis showed that the p38 and Erk1/2 signalling pathways had central roles in the proinflammatory response of mast cells challenged by either sagA-expressing streptococci or detergent. Altogether, these findings suggest that sagA-dependent mast cell membrane perturbation is a mechanism capable of activating the innate immune response upon bacterial challenge.

  • 50.
    Wernersson, S.
    et al.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Box 7011, S-75007 Uppsala, Sweden..
    Riihimäki, M.
    Swedish Univ Agr Sci, Sect Large Anim Surg & Med, Dept Clin Sci, Equine Internal Med, Uppsala, Sweden..
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Box 7011, S-75007 Uppsala, Sweden..
    Waern, I.
    Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Box 7011, S-75007 Uppsala, Sweden..
    Equine Airway Mast Cells are Sensitive to Cell Death Induced by Lysosomotropic Agents2017In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 85, no 1, p. 30-34Article in journal (Refereed)
    Abstract [en]

    Mast cells are known for their detrimental effects in various inflammatory conditions. Regimens that induce selective mast cell apoptosis may therefore be of therapeutic significance. Earlier studies have demonstrated that murine-and human-cultured mast cells are highly sensitive to apoptosis induced by the lysosomotropic agent LeuLeuOMe (LLME). However, the efficacy of lysosomotropic agents for inducing apoptosis of in vivo-derived airway mast cells and the impact on mast cells in other species have not been assessed. Here we addressed whether lysosomotropic agents can induce cell death of equine in vivo-derived mast cells. Bronchoalveolar lavage (BAL) fluids from horses were incubated with LLME at 15-100 lM for up to 48 h. The overall cell viability was unaffected by 15 lM LLME up to 48 h, whereas a relatively modest drop in total cell counts (similar to 30%) was seen at the highest LLME dose used. In contrast to the relatively low effect on total cell counts, LLME efficiently and dose dependently reduced the number of mast cells in BAL fluids, with an almost complete depletion (96%) of mast cells after 24 h of incubation with 100 lM LLME. A significant but less dramatic reduction (up to similar to 45%) of lymphocytes was also seen, whereas macrophages and neutrophils were essentially resistant. The appearance of apoptotic bodies suggested a mechanism involving apoptosis rather than necrosis. These findings suggest that equine airway mast cells are highly sensitive to lysosomotropic agents. Possibly, lysosomotropic agents could be of therapeutic value to treat disorders involving harmful accumulation of mast cells in the airways.

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