Эффективность доставки генов в клетки мозга рыб при помощи рекомбинантных аденоассоциированных вирусов гиппокампа мыши

Abstract

Viral vectors constitute a flexible system that can efficiently deliver an exogenous gene to a variety of target cells and, thus, is a potentially powerful tool for genetic manipulation. To deliver the viral genes, an injection of recombinant adeno-associated viruses was performed according to the Wallace method [1]. The use of recombinant adeno-associated vectors allows viral particles to diffuse relatively easily from the injection area and quickly reproduce themselves in the brain cells. The work objective was to study the effectiveness of infecting various brain regions of chum salmon Oncorhynchus keta aged 2 years old with mammalian viral vectors with a short-term (4 and 8 days) and a longer-term (10 weeks) time intervals with subsequent ultrastructural analysis of the nerve tissue in the injection area. We have found that the mouse hippocampal AAV-1 incorporated in vivo brains of a chum salmon Oncorhynchus keta. Injection of an adeno-associated vector into various areas of the brain was made: telencephalon, cerebellum, mesencephalic tectum, and tegmentum of Oncorhynchus keta, and led to the expression of the reporter genes in pallial (Dd and Dl) regions of the telencephalon, the periventricular region of the masencephalon, in the dorsal part of the brain stem, and in the cerebellum (valvula and corpus cerebellum). Using fluorescence microscopy, we have shown that the presence of green fluorescence is associated with the expression of green fluorescence protein (GFP) in chum salmon cells. The ultrastructural analysis confirmed the presence of subcellular virus particles in the cells of chum salmon brain and intercellular space, which indicates the ability of the adeno-associated vector to diffuse in the nervous tissue of the chum salmon brain.