Effects of the yeast form of Sporothrix schenckii on activation of p38MAPK and expression of interleukin-6 in human THP-1 macrophage-like cells

Abstract

Objective To evaluate effects of the yeast form of Sporothrix schenckii on activation of p38 mitogen-activated protein kinase (p38MAPK) and expression of interleukin-6 (IL-6) in macrophage-like THP-1 cells, which were differentiated from the human acute monocytic leukemia cell line THP-1. Methods THP-1 macrophage-like cells were divided into 3 groups to be treated with the yeast form of Sporothrix schenckii at a concentration of 2 × 106 colony-forming units (CFU) /ml (yeast form group) , 100 mg/L curdlan (curdlan group) and RPMI 1640 medium (blank control group) respectively. Real-time fluorescence-based quantitative PCR was performed to measure the mRNA expression of IL-6 in THP-1 macrophage-like cells in the above 3 groups after 3-and 6-hour treatment separately, and enzyme-linked immunosorbent assay (ELISA) to detect the level of IL-6 in the culture supernatant of THP-1 macrophage-like cells after 24-hour treatment. Western blot analysis was conducted to determine the protein expression of p38MAPK and phosphorylated p38MAPK (p-p38MAPK) in the above 3 groups after 30-and 60-minute treatment separately. Other THP-1 macrophage-like cells were pretreated with 100 nmol/L dexamethasone (a p38MAPK inhibitor) for 30 minutes, and then were divided into 3 groups to be treated with the yeast form of Sporothrix schenckii, curdlan and RPMI 1640 medium respectively, and changes in the level of p-p38MAPK and mRNA expression of IL-6 were also detected. Statistical analysis was carried out with SPSS19.0 software by using one-way or multi-way analysis of variance and least significant difference (LSD) test. Results Significant differences in the mRNA expression of IL-6 in THP-1 macrophage-like cells were observed among the yeast form group, curdlan group and blank control group (F= 5 552.22, P < 0.001) after 3-hour treatment (56.81 ± 7.36, 26.69 ± 1.22 and 0.97 ± 0.05, respectively) and 6-hour treatment (378.03 ± 16.67, 276.24 ± 39.13 and 1.02 ± 0.04, respectively) . Additionally, the yeast form group showed significantly higher mRNA expression of IL-6 after 6-hour treatment than that after 3-hour treatment (q= 16.74, P < 0.001) . After 24-hour treatment, the level of IL-6 in the culture supernatant of THP-1 macrophage-like cells also significantly differed among the yeast form group, curdlan group and blank control group (59.96 ± 18.16 pg/L, 91.01 ± 17.27 pg/L, 5.50 ± 2.30 pg/L, respectively; F= 26.62, P < 0.01) , and was significantly higher in the yeast form group than in the blank control group (P < 0.01) . After 30-and 60-minute treatment, the protein expression of p-p38MAPK was significantly higher in the yeast form group than in the blank control group (both P < 0.01) . Moreover, the mRNA expression of IL-6 (4.46 ± 1.03 vs. 493.52 ± 113.87, P < 0.001) and protein expression of p-p38MAPK (2.29 ± 0.37 vs. 4.55 ± 0.46, q= 10.81, P < 0.01) were both significantly lower in the yeast form group with dexamethasone pretreatment than in that without dexamethasone pretreatment. Conclusion In vitro treatment with the yeast form of Sporothrix schenckii can enhance the expression of IL-6 in human THP-1 macrophage-like cells by activating the p38MAPK signaling pathway. Key words: Sporothrix; Macrophages; Interleukin-6; p38 Mitogen-activated protein kinases