In Vitro Inhibition of Replication of Dengue Virus Serotypes 1–4 by siRNAs Bound to Non-Toxic Liposomes

Abstract

Dengue virus is a ssRNA+ flavivirus, which produces the dengue disease in humans. Currently, no specific treatment exists. siRNAs regulate gene expression and have been used systematically to silence viral genomes; however, they require controlled release. Liposomes show favorable results encapsulating siRNA for gene silencing. The objective herein was to design and evaluate in vitro siRNAs bound to liposomes that inhibit DENV replication. siRNAs were designed against DENV1–4 from conserved regions using siDirect2.0 and Web-BLOCK-iT™ RNAiDesigner; the initial in vitro evaluation was carried out through transfection into HepG2 cells. siRNA with silencing capacity was encapsulated in liposomes composed of D-Lin-MC3-DMA, DSPC, Chol. Cytotoxicity, hemolysis, pro-inflammatory cytokine release and antiviral activity were evaluated using plaque assay and RT-qPCR. A working concentration of siRNA was established at 40 nM. siRNA1, siRNA2, siRNA3.1, and siRNA4 were encapsulated in liposomes, and their siRNA delivery through liposomes led to a statistically significant decrease in viral titers, yielded no cytotoxicity or hemolysis and did not stimulate release of pro-inflammatory cytokines. Finally, liposomes were designed with siRNA against DENV, which proved to be safe in vitro.

PD1 CD44 antiviral peptide as an inhibitor of the protein-protein interaction in dengue virus invasion

“…It was incubated at 37 °C for 2 h. BCIP ® /NBT Liquid Substrate System (Sigma Aldrich Cat. B1911) was used as the substrate, and the reaction incubated for 30 min at 37 °C and washed with abundant water [ 7 ]. A photographic record of the results was made, and subsequently, an intensity analysis per pixel area was carried out using Fiji software [ 38 ].…”

“…It was incubated for 2 h at 37 °C and an alkaline phosphatase-conjugated anti-mouse IgG was used as a secondary antibody (Sigma Aldrich Ref. A4656) [ 7 ].…”

“…The first exposure generates a protective immune response against the infecting serotype but subsequent exposure with heterologous serotypes can lead to severe disease. The clinical disease spectrum ranges from asymptomatic to severe hemorrhage or death [ 3 – 7 ].…”

Applications of Green Synthesized Metal Nanoparticles — a Review

“…The novel vaccine with hybrid coated gold nanoparticles and domain III of the viral envelope protein (EDIII) response antiviral activity against the dengue virus. The complex coated hybrid gold nanoparticle response differs, depending on the size and concentration [235]. The ethanolic and hydroalcoholic extract of gold nanoparticles showed antitubercular activity, and two different types of concentrations are identified as the response such as MIC (2.5 μg/mL and 20 μg/mL) and the highest concentration level (50 μg/mL and 75 μg/mL), respectively [236].…”