Development of In Vitro Macrophage System to Evaluate Phagocytosis and Intracellular Fate of Penicillium marneffei Conid
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Development of In Vitro Macrophage System to Evaluate Phagocytosis and Intracellular Fate of Penicillium marneffei Conidia Sha Lu • Yongxuan Hu • Changming Lu Junmin Zhang • Xiqing Li • Liyan Xi
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Received: 26 May 2012 / Accepted: 11 April 2013 / Published online: 4 May 2013 Ó Springer Science+Business Media Dordrecht 2013
Abstract Penicillium marneffei is a pathogenic fungus that can cause a life-threatening systemic mycosis in the immunocompromised hosts. We established the model for the phagocytosis of P. marneffei conidia by RAW264.7 murine macrophages and designated the fate of P. marneffei in RAW264.7 cells with respect to persistence, phagosome–lysosome-fusion. And we impaired the immune status of mouse and compared the fate and phagosome–lysosome-fusion of P. marneffei in immunocompetent and immunosuppressed mouse peritoneal macrophages cells. We found that conidia could germinate and survive in macrophages. Within 30 min and up to 2 h of heat-killed conidia internalization, the majority of all phagosome types were labeled for the EEA1 (endosomal markers) and LAMP-1 (lysosomal markers), respectively. But both the percentages of LAMP1 and EEA1 that associated with live conidia were significantly lower than that with heat-killed conidia. Administration of cyclophosphamide resulted in a significant suppression of macrophages function (phagocytic and fungicidal) against P. marneffei that were not apparently seen. Our data provide the evidence that (i) intracellular conversion of P. marneffei conidia into yeast cells still could be observed in
S. Lu Y. Hu C. Lu J. Zhang X. Li L. Xi (&) Department of Dermatology, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, 107 West Yanjiang Rd., Guangzhou 510120, China e-mail: [email protected]
macrophages. (ii) Phagosomes containing live Penicillium marneffei conidia might inhibit the phagosome–lysosome-fusion and result to no acidification surrounding the organisms. (iii) Immunity impaired by cyclophosphamide could not influence the function, including phagocytosis, fungicidal activity and phagosome–lysosome-fusion, of macrophages against P. marneffei. Keywords Penicillium marneffei Macrophage Phagolysosome Immunosuppression
Introduction Penicilliosis marneffei, an infection caused by a thermally dimorphic fungus Penicillium marneffei (P. marneffei), is an endemic disease in areas of South East Asia. Most infections occurred in immunocompromised individuals, especially those with HIV infection. With the HIV pandemic, cases have been increased and now Penicilliosis marneffei become one of the common AIDS-defining illnesses in endemic areas, but more and more cases are being recognized in non-endemic countries [1]. Despite the importance of the disease, the routes and mechanisms of infection by P. marneffei are poorly understood. Since P. marneffei appears to be a primary pulmonary pathogen, infection is presumably via inhalation of conidia from the environment [2]. Phagocytic cells are likely to be the primary line of the
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