关键词: 细胞系, 肿瘤

Abstract:

Duan Zhimin*, Du Leilei, Zeng Rong, Shen Yongnian, Hu Suquan, Liu Weida, Chen Qing, Li Min. *Institute of Dermatology, Chinese Academy of Medical Sciences and Peking Union Medical College; Jiangsu Key Laboratory of Molecular Biology for Skin Diseases and STIs, Nanjing 210042, China Corresponding authors: Li Min, Email: minli08@gmail.com; Chen Qing, Email: qngchen@hotmail.com 【Abstract】 Objective To investigate the effects of Candida albicans on the expression of tumor necrosis factor-α （TNF-α） and activation of the intracellular signaling molecule p38 mitogen-activated protein kinase （p38MAPK） in a human acute monocytic leukemia cell line THP-1. Methods Some THP-1 cells were divided into several groups in vitro: two C. albicans groups treated with 105 CFU/ml and 106 CFU/ml heat-killed C. albicans respectively, a lipopolysaccharide （LPS） group treated with 100 μg/L LPS, a blank control group treated with RPMI 1640 medium, two dexamethasone-inhibited groups pretreated with 40 μg/L dexamethasone for 30 minutes followed by treatment with 106 CFU/ml heat-killed C. albicans and LPS respectively. After treatment for 1, 3 and 6 hours, real-time fluorescence-based quantitative PCR was performed to measure TNF-α mRNA expression in THP-1 cells in the above groups. Enzyme-linked immunosorbent assay （ELISA） was conducted to determine the level of TNF-α protein in the supernatant of THP-1 cells treated with 106 CFU/ml heat-killed C. albicans, 100 μg/L LPS or RPMI 1640 medium （blank control group） for 24 hours. Western blot was performed to measure the protein expression of p38MAPK and phosphorylated p38MAPK in THP-1 cells after treatment with 106 CFU/ml heat-killed C. albicans or RPMI 1640 medium （blank control group） for 30 and 60 minutes. Statistical analysis was carried out by using two-way analysis of variance, one-way analysis of variance and the least significant difference （LSD）-t test. Results Significant differences were observed in the mRNA expression level of TNF-α among the C. albicans groups, LPS group and blank control group （F = 110.98, P < 0.001）. The mRNA expression level of TNF-α in THP-1 cells increased over time in a time-dependent manner after C. albicans treatment, with significant differences among different time points （F = 701.680, P < 0.001）. Compared with the blank control group, both 106-CFU/ml C. albicans group and LPS group showed a significant increase in TNF-α protein expression （6385.70 ± 533.99 ng/L and 3212.06 ± 353.00 ng/L vs. 147.10 ± 0.53 ng/L, P < 0.001 and 0.005, respectively）. An obvious increase was observed in the expression level of phosphorylated p38MAPK protein, but no significant changes were noted in that of p38MAPK protein, in THP-1 cells treated with 106 CFU/ml C. albicans for 30 and 60 minutes compared with the blank control group. The mRNA expression level of TNF-α significantly decreased in dexamethasone-pretreated 106-CFU/ml C. albicans group and LPS group compared with those without dexamethasone pretreatment （3.77 ± 0.62 vs. 208.50 ± 10.50, 6.20 ± 1.93 vs. 161.35 ± 1.65, both P < 0.001）. Conclusions Heat-killed C. albicans can induce the activation of p38MAPK in and secretion of TNF-α by human THP-1 cells, which then participate in the innate immune response against C. albicans.