| Abstrakt: |
In the Americas, L. infantum (syn. chagasi) is the main cause of human visceral leishmaniasis. The role of neutrophils as part of the innate response to Leishmania spp. infection is dubious and varies according to the species causing the infection. Global expression of coding RNAs, microRNAs and long non-coding RNAs changes as part of the immune response against pathogens. Changes in mRNA and non-coding RNA expression resulting from infection by Leishmania spp. are widely studied in macrophages, but scarce in neutrophils, the first cell to encounter the trypanosomatid, especially following infection by L. infantum. Herein, we aimed to understand the expression patterns of coding and non-coding transcripts during acute in vitro infection of human neutrophils by L. infantum. We isolated neutrophils from whole blood of healthy male donors (n = 5) and split into groups: 1) infected with L. infantum (MOI = 5:1), and 2) uninfected controls. After 3 hours of exposure of infected group to promastigotes of L. infantum, followed by 17 hours of incubation, total RNA was extracted and total RNA-Seq and miRNA microarray were performed. A total of 212 genes were differentially expressed in neutrophils following RNA-Seq analysis (log2(FC)±0.58, FDR≤0.05). In vitro infection with L. infantum upregulated the expression of 197 and reduced the expression of 92 miRNAs in human neutrophils (FC±2, FDR≤0.01). Lastly, 5 downregulated genes were classified as lncRNA, and of the 10 upregulated genes, there was only 1 lncRNA. Further bioinformatic analysis indicated that changes in the transcriptome and microtranscriptome of neutrophils, following in vitro infection with L. infantum, may impair phagocytosis, apoptosis and decrease nitric oxide production. Our work sheds light on several mechanisms used by L. infantum to control neutrophil-mediated immune response and identifies several targets for future functional studies, aiming at the development of preventive or curative treatments for this prevalent zoonosis. Author summary: Visceral leishmaniasis is a neglected tropical disease that causes fever, weight loss, anemia and swelling of liver and spleen. About 2500 cases are reported annually in the Americas, with a high mortality rate. Understanding how the immune system of people with visceral leishmaniasis responds to this parasite is essential for the development of preventive and curative methods. In order to understand how gene expression is modulated during visceral leishmaniasis, we infected in vitro cultured human neutrophils, the first immune cells to be recruited in this infection, with Leishmania infantum, the protozoan that causes visceral leishmaniasis in the Americas. Next, we measured the expression of coding RNAs, responsible for the production of proteins required for an effective immune response, and of non-coding RNAs, able to control these coding RNAs, thus helping or hindering host response to infection. Analysis of coding and non-coding RNAs points to an attempt by the parasite to modulate the transcriptome of host cells, influencing the host's response to infection. Our work identifies several targets for future functional studies, aiming at the development of preventive or curative treatments for this prevalent zoonosis. [ABSTRACT FROM AUTHOR] |
