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. 2017 Jan 18;18(1):190. doi: 10.3390/ijms18010190

Figure 3.

Figure 3

Schematic diagram showing in vitro experimental schedule (A) and representative images of toluidine blue-stained HMC-1 cells treated with vehicle as a control (CON, (B)); corticotrophin-releasing factor (CRF, (C)), substance P (SP, (D)); CRF + SP (E); CRF + Polygala tenuifolia Willd. (PTW, (F)); SP + PTW (G) and CRF + SP + PTW (H); their tryptase levels (I) and cell viability (J) measured in the culture medium during degranulation. In the experiments of (B–D), cells were harvested for assay 24 h after vehicle (medium, CON), CRF and SP treatments, respectively. In (F–H), PTW was added 30 min before SP and/or CRF treatment, and the cells were harvested 24 h after SP or CRF treatment. The cells in H were harvested according to the in vitro experimental schedule in A with PTW treatment. Arrows in (C–E) indicated the degranulated cells of HMC-1. CRF: corticotropin-releasing factor; SP: substance P; SB: SB203580; PTW: Polygala tenuifolia Willd.; O.D: optical density. SB20358 (10 µM), a p38 mitogen-activated protein kinase (MAPK) inhibitor, was used as a positive control of inhibiting HMC-1 degranulation. Plus (+) and minus (−) in the X-axis description of I indicate ‘treated’ and ‘non-treated’, respectively. Scale bar = 200 μm. *** p < 0.001 vs. non-treated HMC-1 cells (CON); ## p < 0.01 vs. SP-treated HMC-1 cells; a p < 0.05, b p < 0.05, ccc p < 0.001, and ddd p < 0.001 vs. CRF + SP-treated HMC-1 cells; # p < 0.05 vs. vehicle-treated naïve HMC-1 cells (CON).