Lysophosphatidylcholine aggravates contact hypersensitivity by promoting neutrophil infiltration and IL17 expression

Abstract

Lysophosphatidylcholine (LPC) is a bioactive lysolipid known to contribute to the development of lung allergic diseases. However, it remains unknown whether LPC possesses proinflammatory properties in the skin as well. Here, we investigated this issue by injection of LPC into the murine contact hypersensitivity (CHS) model induced by 2,4-dinitrofluorobenzene (DNFB). LPC increased the expression of IL17, recruited more neutrophils, and eventually aggravated the CHS in the skins. Moreover, the effects of LPC diminished after neutralizing IL17 or depleting neutrophils. Mechanistically, LPC upregulated not only IL17 but also CXCL1 and CXCL2 in a G2A-dependent manner. Taken together, our study demonstrated that the upregulation of LPC could contribute to allergic skin inflammation by increasing IL17 expression and neutrophil recruitment via G2A receptor. [BMB Reports 2021; 54(4): 203-208].

Fig. 1

LPC aggravated DNFB-induced skin inflammation. (A) Experimental design. We sensitized WT mice with 0.5% DNFB on d0 and d1, and then challenged them with 0.2% DNFB on d5. During the whole period, we injected LPC s.c. into the mice. The extent of CHS is shown as the increase of the ear thickness (ear swelling, Δ), which we calculated by subtracting the ear thickness of the treated mice (DNFB sensitization and DNFB challenge) from that of the control mice (acetone sensitization and DNFB challenge: representing the non-specific irritation). (B) Ear thickness results over time. We pooled data from five independent experiments. The difference of ear swelling between BSA and LPC was statistically significant on d7 and d8. (C) Ear thickness results on d7. We pooled data; each circle represents a single mouse. (D) FACS analysis of the ear skin. The percentages of hematopoietic cells (CD45+, left), macrophages (CD11b+F4/80+, middle) and neutrophils (CD11b+Ly6G+, right) are shown. We pooled data; each circle represents a single mouse. (E) RT-qPCR analysis on the expressions of CXCL1 and CXCL2 in ear skin. We pooled data; each circle represents a single mouse. (F) We treated naïve mice with anti-Ly6G mAb to deplete neutrophils and then sensitized and challenged them with DNFB. The ear thickness results are shown. Data are representative of two independent experiments, and each circle represents a single mouse. Data are presented as the mean ± SD. NS, not significant; *P < 0.05; **P < 0.01; ***P < 0.001.

Fig. 2

G2A mediated the effects of LPC on neutrophils. (A) RT-qPCR analysis on the expressions of G2A. We subjected the whole ear skin tissues to RNA extraction and used it for RT-qPCR study. Data are representative of two independent experiments. (B) RT-qPCR analysis on the expressions of CXCL1 and CXCL2 in ear skin. We pooled data; each circle represents a single mouse. (C) FACS analysis of the ear skins of WT and G2A KO mice in DNFB-induced CHS. The percentages of neutrophils (CD11b+Ly6G+) in CD45+ population are shown. Statistical analysis is shown in the right panel. We pooled data; each circle represents a single mouse. (D) Ear thickness results of WT and G2A KO mice. Statistical analysis is shown. We pooled data; each circle represents a single mouse. Data are presented as the mean ± SD. NS, not significant; *P < 0.05; **P < 0.01; ***P < 0.001.

Fig. 3

LPC upregulated IL17. (A) The FACS analysis of IL17 and IFN-γ in CD45+ cells of the ears. Statistical analysis is shown in the right. We pooled data; each circle represents a single mouse. (B) RT-qPCR analysis on the expressions of IL17 and IFN-γ. We subjected the whole skin tissues to RNA extraction and used them for RT-qPCR study. Data are presented as the mean ± SD. NS, not significant; *P < 0.05; **P < 0.01; ***P < 0.001.

Fig. 4

LPC exacerbated CHS in an IL17-dependent manner. (A) Ear thickness results after anti-IL17 mAb treatment. We pooled data; each circle represents a single mouse. (B, C) FACS analysis of the ear skin after anti-IL17 mAb treatment. The percentages of neutrophils (CD11b+ Ly6G+) in CD45+ population are shown in (B). (D) RT-qPCR analysis on the expressions of CXCL1 (left) and CXCL2 (right). We subjected the whole skin tissues to RNA extraction and used them for RT-qPCR study. Data are representative of two independent experiments. (E) FACS analysis on the expressions of IL17 and IFN-γ in the WT and G2A KO skins. The frequency of the cytokine-expressing cells in the CD45+ population is shown. Statistical analysis is shown in the right panel. Data are representative of two independent experiments and each circle represents one mouse. Data are presented as the mean ± SD. NS, not significant; *P < 0.05; **P < 0.01; ***P < 0.001.