Mast Cell Apoptosis Induced by Cyclosporin A

Roh, Hwan Jung; Jang, Hyeong Jun; Cho, Kyu Sub; Kim, Yu Soen; Yoo, Young Hyun; Chon, Kyong Myong

Abstract

BACKGROUND AND OBJECTIVES: Mast cell is a key cell in the pathogenesis of allergic rhinitis. It is expected that apoptosis of mast cell is an important step that can lead to treatment of allergic rhinitis. Meanwhile, the cyclosporin A (CsA) is immunosuppresant agent to inhibit the action of various immune cells. The purpose of this study is to identify whether CsA directly induces apoptosis of mast cells in vitro. MATERIALS AND METHOD: After the culture of p815 cells, mouse mastocytoma cells, the cells were treated with 1 micrometer, 2 micrometer, 5 micrometer, and 10 micrometer CsA, and then LD50 of p815 cells were calculated by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) assay. For identification of apoptosis of p815 cells, electrophoresis and flow cytometry after CsA treatment were done and morphological changes in light microscope was observed. We also quantified apoptotic cells by TUNEL assay and Hoechst stain.
RESULTS
The LD50 of p815 cells is 9.87 micrometer after CsA treatment during 24 hours, 6.11 micrometer during 48 hours and 6.21 micrometer during 72 hours. With the higher concentration of CsA treatment, the greater effect on apoptosis of p815 cells was revealed. We observed laddering pattern for DNA fragmentation in electrophoresis. Nuclear fragmentation and chromatin condensation of p815 cells was observed under the light microscope. Results of flow cytometry were similar to the MTT assay results. Quantification of apoptotic p185 cells by TUNEL assay and Hoechst stain were calculated.
CONCLUSION
Apoptosis of mast cells can be induced by CsA treatment in vitro.