Regulatory effect of Candida albicans hyphae on the key autophagy-related molecule microtubule-associated protein 1 light chain 3 in murine bone marrow-derived macrophages / 中华皮肤科杂志

ABSTRACT

Objective:To evaluate the effect of Candida albicans ( C. albicans) hyphae on autophagic flux in murine bone marrow-derived macrophages (BMDM) . Methods:BMDM were in vitro stimulated with C. albicans hyphae for 0.5, 4 and 12 hours, and the 0-hour group treated without hyphae served as a control. Western blot analysis was performed to detect the conversion of microtubule-associated protein 1 light chain 3 (LC3) -Ⅰto LC3-Ⅱ, and determine the expression of phosphorylated mechanistic target of rapamycin (p-mTOR) at each time point. Some BMDM were divided into several groups control group receiving no treatment, hyphae group treated with C. albicans hyphae, lysosomal inhibitor groups treated with different lysosomal inhibitors, including E-64d (a cysteine proteinase inhibitor) + pepstatin (a pepsin inhibitor) , bafilomycin-A1 (BAF-A1) , ammonium chloride and chloroquine, and hyphae combined with lysosomal inhibitor groups treated with lysosomal inhibitors immediately followed by C. albicans hyphae. After 4- or 12-hour treatment, the effect of C. albicans hyphae on basal autophagic flux in murine BMDM was evaluated. Statistical analysis was carried out by using unpaired t test, factorial design analysis of variance and least significant difference- t test. Results:After 0.5-, 4- and 12-hour in vitro treatment with C. albicans hyphae, the conversion of LC3-Ⅰ to LC3-Ⅱ significantly increased in murine BMDM (1.254±0.118, 1.629±0.391, 1.598±0.379, respectively) compared with the 0-hour group (0.983±0.030; t=3.875, 2.856, 2.804, respectively, all P< 0.05) , while there was no significant difference in the protein expression of p-mTOR among the 0-, 0.5-, 4- and 12-hour groups. After 4- and 12-hour in vitro treatment with C. albicans hyphae combined with lysosomal inhibitors E-64d and pepstatin, the accumulation level of LC3-Ⅱ significantly increased in BMDM compared with those treated with E-64d and pepstatin alone ( t=3.691, 6.648, respectively, both P< 0.05) . Compared with the corresponding lysosomal inhibitor groups, the accumulation level of LC3-Ⅱsignificantly increased in BMDM treated with C. albicans hyphae combined with BAF-A1, ammonium chloride or chloroquine for 4 and 12 hours (all P< 0.05) . Conclusion:In vitro treatment with C. albicans hyphae can increase the conversion of LC3-Ⅰto LC3-Ⅱ in the basal autophagic flux in murine BMDM.