Silencing of IL-32 in human THP-1-derived macrophages increases <i>Leishmania</i> species infection.

<p>PMA-differentiated THP-1 cells (2.5 x 10⁶ cells/800 μL) were electroporated by using Amaxa Nucleofector Technology with IL-32 siRNA (for knockdown IL-32) and control siRNA according with protocol described by [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0005413#pntd.0005413.ref038" target="_blank">38</a>]. The final concentration of cells per well were 3 x 10⁵ cells/100 μL. After 24 h of transfection, cells were infected with promastigotes forms (1.5 x 10⁶ parasites) in the growth stationary phase of <i>L</i>. (<i>L</i>.) <i>amazonensis</i> (<i>L</i>. <i>amaz</i>), metacyclic promastigote forms (1.5 x 10⁶ parasites) of <i>L</i>. (<i>V</i>.) <i>braziliensis</i> (<i>L</i>. <i>braz</i>). After 4 h, cells were washed to remove non-internalized parasites and incubated for 24 h or 48 h. Cells were stained and percentage of infected macrophages, number of parasites per infected cells, and infection index were evaluated. (A) infection with <i>L</i>. <i>amazonensis</i>; (B) infection with <i>L</i>. <i>braziliensis</i>. iNOS (C—left panel), cathelidicin (D—left panel) and β-defensin 2 (E) mRNA expression were determined by quantitative real-time PCR (24 h). Production of nitrite (C—right panel) and LL-37 (D—right panel) was determined by Griess reagent and ELISA in supernatants, respectively (24 h). Values are expressed as means ± SEM of three independent experiments. *p < 0.05 (Control SiRNA vs IL-32 SiRNA).