Журнал микробиологии, эпидемиологии и иммунобиологии (Jun 2015)
EFFECTIVENESS OF POLYCATIONIC NANOPARTICLES OF POLYETHYLENEIMINE-POLYHYDROZIDE-CHITOSAN (PEI-PG-OCHG) AS A VECTOR FOR SMALL INTERFERING RNA, DIRECTED TO SUPPRESS HERPES SIMPLEX TYPE 2 VIRUS REPLICATION
Abstract
Aim. Evaluation of an antiviral effect of miRNA in the nanoparticles of a polycationic compound against mRNA of vp16 protein (UL48 gene) of herpes simplex virus type 2 (HSV-2) in vitro. Materials and methods. 50% aqueous solution of polyethyleneimine (BDH, Great Britain), chitosan, containing approximately 15% of N-acetylated glucosamine chains (Sonat, Russia), hydrazine-hydrate and other chemical reagents (Chimmed, Russia); Vero continuous cell line, MS HSV-2 virus were used. Vero cells were cultivated in DMEM medium supplemented by 10% fetal bovine serum at 37°C in the atmosphere of 5% CO2. Cell viability was evaluated by using Neutral Red vital stain and MTT-test. Primers and probes for RT-PCR were modeled in Vector NTI 8.0 computer program according to the mRNA sequences of the studied genes (the sequences were obtained from GenBank) and synthesized in Sintol (Russia). RT-PCR tests were set using a standard procedure. Synthesis of PEI-PG-chitosan was carried out by Krivtsov G.G. et al. (2010). Results. A design and synthesis of nucleotide sequences, that have interfering activity against this virus, was carried out to study the effect of siRNA on HSV-2 virus replication. During simultaneous addition of HSV-2 and specific siRNA to Vero cells in cell culture, a significant (by 4 lg) reduction of virus yield was observed. A level of UL48 mRNA expression level was determined after the influence of various siRNA variants. A S2 siRNA variant was shown to cause the most pronounced virus-inhibiting effect, aiming for the center of RNA-target (the level of expression of the studied gene decreased by 0.5 lg). Conclusion. siRNA in the PEI-PG-chitosan complexes were established to possess in vitro pronounced suppressive HSV-2 replication activity. The results obtained could be used in creation of new therapeutic preparation against herpes viruses.
