AUTHOR=Muxel Sandra Marcia , Acuña Stephanie Maia , Aoki Juliana Ide , Zampieri Ricardo Andrade , Floeter-Winter Lucile Maria 

TITLE=Toll-Like Receptor and miRNA-let-7e Expression Alter the Inflammatory Response in Leishmania amazonensis-Infected Macrophages

JOURNAL=Frontiers in Immunology

VOLUME=9

YEAR=2018

URL=https://www.frontiersin.org/journals/immunology/articles/10.3389/fimmu.2018.02792

DOI=10.3389/fimmu.2018.02792

ISSN=1664-3224

ABSTRACT=<p>Parasite recognition by Toll-like receptors (TLRs) contributes to macrophage activation and subsequent control of <italic>Leishmania</italic> infection through the coordinated production of pro-inflammatory and microbicidal effector molecules. The modulation of microRNA (miRNA) expression by <italic>Leishmania</italic> infection potentially mediates the post-transcriptional regulation of the expression of genes involved in leishmanicidal activity. Here, the contribution of TLR signaling to the miRNA profile and gene expression was evaluated in <italic>Leishmania amazonensis</italic>-infected murine macrophages. The infectivity of <italic>L. amazonensis</italic> was higher in murine bone marrow-derived macrophages from mice knockout for myeloid differentiation factor 88 (MyD88<sup>−/−</sup>), TLR2 (TLR2<sup>−/−</sup>), or TLR4 (TLR4<sup>−/−</sup>) than wild type C57BL/6 (WT). <italic>L. amazonensis</italic> infection of WT macrophages modulated the expression of 32% of the miRNAs analyzed, while 50% were upregulated. The absence of MyD88, TLR2, and TLR4 altered the percentage of miRNAs modulated during <italic>L. amazonensis</italic> infection, including the downregulation of let-7e expression. Moreover, the absence of signals mediated by MyD88, TLR2, or TLR4 reduced nitric oxide synthase 2 (<italic>No</italic>s2) mRNA expression during infection. Indeed, the inhibition of let-7e increased levels of the <italic>Nos</italic>2 mRNA and NOS2 (or iNOS) protein and nitric oxide (NO) production in <italic>L. amazonensis</italic>-infected macrophages (4–24 h). The absence of TLR2 and inhibition of let-7e increased the expression of the arginase 1 (<italic>Arg</italic>1) mRNA but did not alter the protein level during infection. However, higher levels of the L-arginine transporters <italic>Cat</italic>2B and <italic>Cat</italic>1 were detected in the absence of Myd88 signaling during infection but were not altered following let-7e inhibition. The inhibition of let-7e impacted the global expression of genes in the TLR pathway by upregulating the expression of recognition and adaptors molecules, such as <italic>Tlr</italic>6, <italic>Tlr</italic>9, <italic>Ly</italic>96, <italic>Tirap, Traf</italic> 6, <italic>Ticam</italic>1, <italic>Tollip, Casp</italic>8, <italic>Map</italic>3<italic>k</italic>1, <italic>Mapk</italic>8, <italic>Nfkbib, Nfkbil</italic>1, <italic>Ppara, Mapk8i</italic>p3, <italic>Hspd</italic>1, and <italic>Ube2</italic>n, as well as immunomodulators, such as <italic>Ptgs</italic>2/<italic>Cox</italic>2, <italic>Csf</italic> 2, <italic>Csf</italic> 3, <italic>Ifnb</italic>1, <italic>Il</italic>6<italic>ra</italic>, and <italic>Ilr</italic>1, impacting NOS2 expression, NO production and parasite infectiveness. In conclusion, <italic>L. amazonensis</italic> infection alters the TLR signaling pathways by modulating the expression of miRNAs in macrophages to subvert the host immune responses.</p>