RESUMO
Objective To investigate the autophagy level of Ana-1 cells after ingesting melanized Penicillium marneffei (PM),and to explore the feasibility of rapamycin in killing the bacteria by inducing macrophages autophagy.Methods Melanized PM was cultivated and isolated from the medium containing dopamine.The expression of microtubule-associated protein 1 light chain 3 Ⅱ (LC3 Ⅱ) in Ana-1 cells stimulated by conventional or melanized PM was detected by western blot.The expression levels of LC3 Ⅱ protein in Ana-1 cells treated with rapamycin and incubated with melanized PM was detected.Then,the localization of LC3 Ⅱ in Ana-1 cells which contained melanized PM was observed by immunofluorescence staining.Finally,the direct sterilization effect of rapamycin on melanized PM were detected,and the sterilization effect of Ana-1 cells treated with or without rapamycin on melanized PM was measured.Results No significant change was found in the LC3 Ⅱ level of Ana-1 cells after ingesting melanized PM (P＞0.05),while LC3 Ⅱ level in Ana-1 cells treated with rapamycin which contained melanized PM was significantly increased (P=0.009).The colocalization of LC3 Ⅱ with melanized PM in cytoplasm of Ana-1 cells was observed.For Ana-1 cells treated with rapamycin,3 h and 6 h after incubated with melanized PM,the survival rates of melanized PM both were significantly reduced (P=0.026,0.014).No significant sterilization effect of Ana-1 cells or rapamycin was observed under the same conditions.Conclusion Melanized PM can suppress the activation of macrophage autophagy,and rapamycin can improve sterilization effect of Ana-1 cells by inducing the activation of autophagy.