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Quantification of Intracellular Growth Inside Macrophages is a Fast and Reliable Method for Assessing the Virulence of Leishmania Parasites
Published on: March 16, 2018
Transcriptional signatures of BALB/c mouse macrophages housing multiplying Leishmania amazonensis amastigotes
José Osorio y Fortéa1, Emilie de La Llave, Béatrice Regnault
1Institut Pasteur, Unité d'Immunophysiologie et Parasitisme Intracellulaire, Département de Parasitologie et Mycologie, Paris, France. josorio@pasteur.fr
Background:
Mammal macrophages (MPhi) display a wide range of functions which contribute to surveying and maintaining tissue integrity. One such function is phagocytosis, a process known to be subverted by parasites like Leishmania (L). Indeed, the intracellular development of L. amazonensis amastigote relies on the biogenesis and dynamic remodelling of a phagolysosome, termed the parasitophorous vacuole, primarily within dermal MPhi.
Results:
Using BALB/c mouse bone marrow-derived MPhi loaded or not with amastigotes, we analyzed the transcriptional signatures of MPhi 24 h later, when the amastigote population was growing. Total RNA from MPhi cultures were processed and hybridized onto Affymetrix Mouse430_2 GeneChips, and some transcripts were also analyzed by Real-Time quantitative PCR (RTQPCR). A total of 1,248 probe-sets showed significant differential expression. Comparable fold-change values were obtained between the Affymetrix technology and the RTQPCR method. Ingenuity Pathway Analysis software pinpointed the up-regulation of the sterol biosynthesis pathway (p-value = 1.31e-02) involving several genes (1.95 to 4.30 fold change values), and the modulation of various genes involved in polyamine synthesis and in pro/counter-inflammatory signalling.
Conclusion:
Our findings suggest that the amastigote growth relies on early coordinated gene expression of the MPhi lipid and polyamine pathways. Moreover, these MPhi hosting multiplying L. amazonensis amastigotes display a transcriptional profile biased towards parasite-and host tissue-protective processes.
Insights
Leishmania amazonensis amastigotes subvert macrophage functions, hijacking lipid and polyamine pathways for growth. Macrophages hosting these parasites develop a protective transcriptional profile, aiding parasite survival and host tissue integrity.
Area of Science:
- Immunology
- Molecular Biology
- Parasitology
Background:
- Mammalian macrophages (MPhi) are crucial for tissue integrity and defense.
- The parasite Leishmania (L.) amazonensis subverts MPhi functions, particularly phagocytosis.
- Intracellular development of L. amazonensis amastigotes depends on phagolysosome remodeling within MPhi.
Purpose of the Study:
- To investigate the transcriptional changes in MPhi during L. amazonensis amastigote intracellular growth.
- To identify host pathways modulated by L. amazonensis infection.
Main Methods:
- Transcriptional profiling of BALB/c mouse bone marrow-derived MPhi infected with L. amazonensis amastigotes using Affymetrix GeneChips.
- Validation of gene expression using Real-Time quantitative PCR (RTQPCR).
- Pathway analysis using Ingenuity Pathway Analysis software.
Main Results:
- A total of 1,248 probe-sets exhibited significant differential expression in infected MPhi.
- Upregulation of the sterol biosynthesis pathway (p-value = 1.31e-02) with fold changes from 1.95 to 4.30.
- Modulation of genes involved in polyamine synthesis and inflammatory signaling.
Conclusions:
- L. amazonensis amastigote growth is supported by early, coordinated MPhi gene expression in lipid and polyamine pathways.
- Infected MPhi display a transcriptional profile promoting parasite survival and host tissue protection.
