Sorting of Leishmania-bearing dendritic cells reveals subtle parasite-induced modulation of host-cell gene expression

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• 作者：Hervé ; Lecoeur ; Emilie de La Llave ; José ; Osorio Y Forté ; a ; Sophie Goyard ; Hé ; lè ; ne Kiefer-Biasizzo ; Anne-Marie Balazuc ; Geneviè ; ve Milon ; Eric Prina ; Thierry Lang
• 关键词：Leishmania ; Mouse dendritic cells ; Flow-sorting ; BSL-2 ; DsRed2
• 刊名：Microbes and Infection
• 出版年：2010
• 出版时间：January 2010
• 年：2010
• 卷：12
• 期：1
• 页码：46-54
• 全文大小：1080 K

文摘

Once in the mouse skin, Leishmania (L) amazonensis amastigotes are hosted by professional mononuclear phagocytes such as dendritic cells (DCs). When monitored after parasite inoculation, the frequency of amastigote-hosting DCs is very low (<1 % ) in both the skin and skin-draining lymph nodes. Therefore, we designed and validated an efficient procedure to purify live amastigotes-hosting DCs with the objective to facilitate quantitative and qualitative analysis of such rare cells. To this end, a L. amazonensis transgenic parasite expressing DsRed2 fluorescent protein was generated and added to mouse bone marrow-derived DC cultures. Then, a high speed sorting procedure, performed in BSL-2 containment, was setup to pick out only DCs hosting live amastigotes. This study reveals, for the first time, a unique transcript pattern from sorted live amastigotes-hosting DCs that would have been undetectable in unsorted samples. It was indeed possible to highlight a significant and coordinated up-regulation of l-arginine transporter and arginase2 transcripts in Leishmania-hosting DCs compared to un-parasitized DCs. These results indicate that arginine catabolism for polyamine generation is dominating over L-arginine catabolism for NO generation. In conclusion, this approach provides a powerful method for further characterisation, of amastigote-hosting DCs in the skin and the skin-draining lymph nodes.