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Serotonin and histamine storage in mast cell secretory granules is dependent on serglycin proteoglycan  Maria Ringvall, PhD, Elin Rönnberg, MSc, Sara.

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Presentation on theme: "Serotonin and histamine storage in mast cell secretory granules is dependent on serglycin proteoglycan  Maria Ringvall, PhD, Elin Rönnberg, MSc, Sara."— Presentation transcript:

1 Serotonin and histamine storage in mast cell secretory granules is dependent on serglycin proteoglycan  Maria Ringvall, PhD, Elin Rönnberg, MSc, Sara Wernersson, PhD, Annette Duelli, MSc, Frida Henningsson, PhD, Magnus Åbrink, PhD, Gianni García-Faroldi, MSc, Ignacio Fajardo, PhD, Gunnar Pejler, PhD  Journal of Allergy and Clinical Immunology  Volume 121, Issue 4, Pages (April 2008) DOI: /j.jaci Copyright © 2008 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2 Fig 1 Reduced levels of histamine and serotonin in BMMCs from serglycin null animals. Bone marrow precursor cells were cultured in vitro into BMMCs. At the time points indicated, cells (2 × 106) were taken and analyzed for cellular content of histamine (A) and serotonin (B). The results displayed are from 1 cell culture experiment (n = 2; mean of duplicates ± SD; Student t test; ∗∗P < .01) and are representative of 3 individual experiments. C, RT-PCR analysis of BMMCs for expression of the mMCP-4 and mMCP-6 genes. HPRT, Housekeeping control. D, May-Grünwald/Giemsa staining of WT and serglycin–/– BMMCs at different time points of cell culture. ko, Knockout. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2008 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3 Fig 2 Histamine, serotonin, and polyamines levels in BMMCs. Serglycin+/+ (A; WT) or serglycin–/– (B; knockout [KO]) bone marrow precursor cells were cultured in vitro into BMMCs. After 18 days, cells (4 × 106) were taken and analyzed for cellular contents of histamine, serotonin, putrescine, spermidine, and spermine by HPLC. 1,8-Diaminooctane was used as an internal standard added to the cell extract before derivatization with dansyl chloride. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2008 American Academy of Allergy, Asthma & Immunology Terms and Conditions

4 Fig 3 Expression of HDC, TPH-1, and serglycin in BMMCs. WT or knockout (ko) bone marrow precursor cells were cultured in vitro into BMMCs and were analyzed for levels of transcript for HDC (A; the levels of expression relative to that of WT cells at day 19), TPH-1 (B; the levels of expression relative to that of WT cells at day 35), and serglycin (C; the levels of expression relative to that of cells at day 28) by qPCR. The results displayed are from 1 cell culture experiment (n = 2; mean of duplicates ± SD) and are representative of 2 individual experiments. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2008 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5 Fig 4 Release of histamine and serotonin from BMMCs. BMMCs, taken at the time points of culture indicated, were stimulated with calcium ionophore A23187 (n = 2; mean of duplicates ± SD; Student t test; ∗P < .05). Conditioned media were collected and analyzed for histamine (A) and serotonin (B) by EIA. As controls, media from nonstimulated cells were analyzed. The results are representative of 3 individual experiments. ko, Knockout. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2008 American Academy of Allergy, Asthma & Immunology Terms and Conditions

6 Fig 5 Reduced storage and release of histamine and serotonin in peritoneal cells from serglycin null animals. Peritoneal cells from WT or knockout (KO) mice were stimulated in vitro with A23187 for 60 minutes. Conditioned medium from stimulated and control cells were analyzed for content of histamine (A) and serotonin (B). Cellular levels (cell) of histamine (A) and serotonin (B) were analyzed in A23187-activated or nontreated cells (n = 4; mean values ± SD; Student t test; ∗∗∗P < .001). C, The percentage of CD117+ cells in peritoneum was determined by flow cytometry (n = 5). D, Cell complexity (granularity) of CD117+ cells is demonstrated by side scatter (SSC) detection (representative dot blots; n = 5). ns, Not significant. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2008 American Academy of Allergy, Asthma & Immunology Terms and Conditions

7 Fig 6 Extracellular levels of histamine and serotonin in the peritoneum and serum of WT and serglycin–/– animals. Peritoneal lavage fluid and serum were collected from WT and serglycin–/– (knockout [KO]) animals (n = 5 for peritoneal fluid; n = 3 for serum) and were analyzed for histamine and serotonin content by EIA. A, Histamine levels in the peritoneum. B, Histamine levels in serum. C, Serotonin levels in the peritoneum. D, Serotonin levels in serum. ∗∗P < .01 (Student t test). Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2008 American Academy of Allergy, Asthma & Immunology Terms and Conditions

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