RNA-Seq Revealed Expression of Many Novel Genes Associated With Leishmania donovani Persistence and Clearance in the Host Macrophage

Abstract

Host- as well as parasite-specific factors are equally crucial in allowing either the Leishmania parasites to dominate, or host macrophages to resist infection. To identify such factors, we infected murine peritoneal macrophages with either the virulent (vAG83) or the non-virulent (nvAG83) parasites of L. donovani. Then, through dual RNA-seq, we simultaneously elucidated the transcriptomic changes occurring both in the host and the parasites. Through Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the differentially expressed (DE) genes, we showed that the vAG83-infected macrophages exhibit biased anti-inflammatory responses compared to the macrophages infected with the nvAG83. Moreover, the vAG83-infected macrophages displayed suppression of many important cellular processes, including protein synthesis. Further, through protein-protein interaction study, we showed significant downregulation in the expression of many hubs and hub-bottleneck genes in macrophages infected with vAG83 as compared to nvAG83. Cell signaling study showed that these two parasites activated the MAPK and PI3K-AKT signaling pathways differentially in the host cells. Through gene ontology analyses of the parasite-specific genes, we discovered that the genes for virulent factors and parasite survival were significantly upregulated in the intracellular amastigotes of vAG83. In contrast, genes involved in the immune stimulations, and those involved in negative regulation of the cell cycle and transcriptional regulation, were upregulated in the nvAG83. Collectively, these results depicted a differential regulation in the host and the parasite-specific molecules during in vitro persistence and clearance of the parasites.

Gene expression patterns associated with Leishmania panamensis infection in macrophages from BALB/c and C57BL/6 mice

“…panamensis ( S6 Fig ). Several of the DE genes observed in the transcriptomic signatures for both mouse strains have been previously reported as differentially expressed in studies conducted with other Leishmania species [ 8 – 10 , 14 – 16 , 18 ], although the direction and intensity of fold change varies. Similarities and differences concerning DE genes reported in this study for L .…”

“…C57BL/6 mice often display a resistant phenotype to infection with most Leishmania species, due to a predominant proinflammatory macrophage activation profile (M1), often resulting in parasite clearance [ 4 , 8 – 10 ]. Conversely, BALB/c mice tend to exhibit a susceptible phenotype, possibly associated with initial alternative macrophage activation profile (M2), characterized by inhibition of proinflammatory signals and leading to sustained parasite growth and tissue damage [ 2 – 4 , 11 – 14 ]. However, there is conflicting evidence regarding the direction of polarization of macrophage activation, based on studies considering different combinations of Leishmania species and host mouse strains, which highlights the importance of studying species-specific interactions between Leishmania parasites and different host systems [ 8 , 9 , 23 , 10 , 12 – 18 ].…”

“…Conversely, BALB/c macrophages showed a non-classical and possibly intermediate activation profile, reflected by the simultaneous upregulation of anti-inflammatory cytokine IL-10 and M2 macrophage-secreted arginase 1, together with milder induction and/or downregulation of both inflammatory cytokines and pro-apoptotic genes. Although several studies conducted in Leishmania -infected BALB/c macrophages have reported downregulation of many genes involved in inflammatory processes [ 12 – 14 ], the occurrence of an intermediate macrophage activation profile has also been suggested [ 15 ]. Since these studies were conducted with different L .…”

“…C57BL/6 mice show resistance to Leishmania infections, due to a predominant Th1 cytokine response and M1 macrophage activation [ 4 , 8 – 10 ]. Conversely, most Leishmania infections in BALB/c mice trigger an initial Th2 response conferring a susceptible phenotype that results in sustained parasite growth and tissue damage [ 2 – 4 , 11 – 14 ].…”

Of Drugs and Trypanosomatids: New Tools and Knowledge to Reduce Bottlenecks in Drug Discovery

“…These models have great advantages such as the direct evaluation of drug penetration in the host cell, as well as drug activity in the phagolysosome milieu, among others [ 29 , 30 ]. Moreover, intracellular amastigotes are generally more sensitive than promastigotes against most of the drugs currently used in clinic, such as antimony or miltefosine [ 31 , 32 ], which could be a consequence of genes differentially regulated in the two developmental stages of the parasite [ 31 , 33 , 34 ]. The activity of candidate compounds against intracellular amastigotes is determined by microscopic automatic/manual counting of infected macrophages and the number of parasites per macrophage (parasitic index) or by spectrophotometric (e.g., optical density or staining) and fluorometric methods.…”