Despite active research, so far the detailed mechanisms of TBEV pathogenesis have not been fully disclosed. Recently, extracellular vesicles, especially small-sized vesicles, which have been shown to play an important role in the pathogenesis of many viruses, have attracted the attention of scientists. In this study, we investigated the effect of the expression of nonstructural protein 1 (NS1) of tick-borne encephalitis virus on the release of extracellular vesicles by cells and assessed the possibility of these vesicles affecting other cells. NS1 expression by TBEV was found to increase the release of extracellular vesicles by HEK293T cells; however, no changes in the size profile of released vesicles were detected. In addition, NS1 is detected in both large and small vesicle size fractions. It was found that NS1 TBEV is not present inside the vesicles, but is associated with their outer surface. Small-sized vesicles derived from the culture medium of NS1-expressing HEK293T cells are able to induce an increase in the mRNA content and interleukin-1β(IL-1β) secretion in human neuroblastoma SHSY5Y cells. The results indicate the involvement of the NS1 protein and vesicles in the development of neuroinflammation and are important for understanding the mechanisms of pathogenesis of tick-borne encephalitis virus.