Real-Time qPCR Array of miRNAs in Leishmania-infected THP-1 macrophages

Leishmania amazonensis is a protozoan that primarily infects macrophages and causes cutaneous leishmaniasis in humans. MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression at posttranscriptional levels. Previous work demonstrated changes in miRNA profile of host cells favoring parasite survivel. Thus, here we demonstrate that human macrophages upregulate several miRNAs on the initial time points of infection, including the hsa-miR-372, hsa-miR-373, and hsa-miR-520d, which present the same seed. Further functional analysis demonstrated that inhibition of the miR-372 impaired Leishmania survival in THP-1 macrophages and the effect was further enhanced with combinatorial inhibition of the miR-372/373/520d family, pointing to a cooperative mechanism. Our study demonstrated miRNA-dependent modulation of polyamines production, establishing permissive conditions for intracellular parasite survival.Our findings suggest that the miR-372/373/520d family may represent a potential target for the development of new therapeutic strategies against cutaneous leishmaniasis. qPCR array for miRNA expression profile. Differentiated THP-1 macrophages where either infected with Leishmania amazonensis promastigotes (MOI 5) or left non-infected as indicated and miRNAs were quantified after 4 or 24 hours of infection (N=4 for each group). qPCR array was performed using the Qiagen Human Inflammatory Response & Autoimmunity, RT-PCR Array platform and all data were analyzed using QIAGEN's web-based software (https://geneglobe.qiagen.com/us/analyze). Unfortunately, the resulting analysis table does not give the normalized values for each sample, and instead provides averaged normalized value per group.