MrgX2‑mediated internalization of LL‑37 and degranulation of human LAD2 mast cells

Murakami,Taisuke; Suzuki,Kaori; Niyonsaba,Francois; Tada,Hiroyuki; Reich,Johannes; Tamura,Hiroshi; Nagaoka,Isao

doi:10.3892/mmr.2018.9532

MrgX2‑mediated internalization of LL‑37 and degranulation of human LAD2 mast cells

Authors:
- Taisuke Murakami
- Kaori Suzuki
- Francois Niyonsaba
- Hiroyuki Tada
- Johannes Reich
- Hiroshi Tamura
- Isao Nagaoka
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Affiliations: Department of Host Defense and Biochemical Research, Juntendo University Graduate School of Medicine, Tokyo 113‑8421, Japan, Atopy (Allergy) Research Center, Juntendo University Graduate School of Medicine, Tokyo 113‑8421, Japan, Division of Oral Microbiology, Tohoku University Graduate School of Dentistry, Sendai‑shi, Miyagi 980‑8575, Japan, Endotoxin Test Service, Microcoat Biotechnologie GmbH, D‑82347 Bernried, Germany
Published online on: October 2, 2018 https://doi.org/10.3892/mmr.2018.9532
Pages: 4951-4959
Copyright : © Murakami et al. This is an open access article distributed under the terms of Creative Commons Attribution License [CC BY 4.0].

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Abstract

LL‑37 is the sole antimicrobial peptide of human cathelicidin comprising 37 amino acids, which is expressed mainly in epithelial cells and neutrophils, and activates mast cells. In the present study, in order to elucidate the mechanism of mast cell activation by LL‑37, the associations between the internalization of LL‑37 and Mas‑related gene X2 (MrgX2)‑mediated mast cell activation (degranulation) was investigated using the human mast cell line, LAD2. LL‑37 was rapidly internalized into the cells, and induced degranulation, as assessed by the extracellular release of β‑hexosaminidase. Pertussis toxin, a G‑protein inhibitor, significantly suppressed the internalization of LL‑37 and the degranulation of LAD2 cells. Furthermore, small interfering (si)‑RNA‑mediated knockdown of MrgX2, a putative G protein‑coupled receptor for LL‑37, inhibited the internalization of LL‑37 and degranulation of LAD2 cells. Notably, LL‑37 internalization was enhanced by the stable expression of MrgX2 in HMC‑1 and 293 cells. In addition, the internalized LL‑37 mainly colocalized with MrgX2 in the perinuclear region of LAD2 cells. Furthermore, neuraminidase treatment, which removes negatively charged sialic acid from the cell surface, markedly reduced the internalization of LL‑37 and degranulation of LAD2 cells, and clathrin‑mediated endocytosis inhibitors (dynasore and chlorpromazine) inhibited the internalization and degranulation of LAD2 cells. Taken together, these observations indicated that LL‑37 may bind the negatively charged cell surface molecules, rapidly internalize into the cells via clathrin‑mediated endocytosis and interact with MrgX2 to activate mast cells (LAD2 cells).

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