Methods for Analyzing Sphingosine-1-Phosphate Signaling in Human and Mouse Primary Mast Cells

  • Alena P. Chumanevich
  • Piper A. Wedman
  • Carole A. Oskeritzian
Part of the Methods in Molecular Biology book series (MIMB, volume 1697)


Mast cells produce a potently bioactive sphingolipid metabolite sphingosine-1-phosphate (S1P) constitutively and upon activation. The ligation of S1P to its type 2 receptor on mast cells triggers a novel downstream signaling pathway that we discovered links activation of transcription factor signal transducer and activator of transcription 3 to mast cell-derived chemokine release in both humans and mice. In this chapter, we describe the methods used to study S1P signaling in human and mouse primary mast cells.


Primary mast cells Sphingosine-1-phosphate Signaling Protein phosphorylation Signal transducer and activator of transcription 3 Activation Chemokines Inflammation Western blot Quantitative PCR 



Mouse bone marrow-derived mast cell


Bovine serum albumin


Threshold cycle




Glyceraldehyde-3-phosphate dehydrogenase


4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid


Mast cell


Phosphate-buffered saline


Phosphorylated Stat3


Real-time quantitative PCR


Recombinant human stem cell factor


Recombinant murine




S1P receptor


Human skin-derived mast cell

Stat3 or STAT3

Signal transducer and activator of transcription 3


Western blot



This work was supported by grants from the US National Institutes of Health (NIH)/National Institute of Allergy and Infectious Diseases R01 AI095494 and NIH/National Institute of Arthritis and Musculoskeletal and Skin Diseases R21 AR067996 to C.A.O.


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Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Alena P. Chumanevich
    • 1
  • Piper A. Wedman
    • 1
  • Carole A. Oskeritzian
    • 1
  1. 1.Department of Pathology, Microbiology and ImmunologyUniversity of South Carolina School of MedicineColumbiaUSA

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