Biomimetic Membrane Vesicles: Evaluation of Inflammatory Response In Vivo

Introduction: Biomimetic membrane vesicles are produced from live cells using cytochalasin B which disrupts the structure of the cytoskeleton and facilitate generation of membrane vesicles under subsequent vortexing. Membrane vesicles are pinched off from the cell surface, surrounded by a cytoplasmic membrane and contain the cytoplasm of parental cell. It is known that mesenchymal stem cells (MSCs) are immunoprivileged. The aim of our study was to determine whether Cytochalasin B-induced membrane vesicles (CIMVs) derived from mesenchymal stem cells retain the immunoprivileged properties of parental cells.

Method: All experiments were carried out in compliance with the procedure protocols approved by Kazan Federal University and local ethics committee (protocol #5, date 27.05.2014). To analyze the immunogenicity, murine MSCs (7.5x10⁴ cells) either CIMVs-MSCs (15µg) were injected i.v. in 8 week old mice (Mus musculus, C57Bl/6). CIMVs were used at a concentration equivalent to 7.5×104 MSCs based on total protein concentration. Serum isolation was performed after 2 hours post-administration. Secretion of inflammatory cytokines was evaluated using multiplex analysis BioPlex Pro Mouse 23 Plex kit. (BioRad, USA).

Results: We detected all investigated cytokines in serum of control and experimental mice: Eotaxin, G-CSF, GM-CSF, IFN-g, IL-10, IL-13, IL-17A, IL-1a, IL-1b, IL-2, IL-3, IL-4, IL-5, IL-6, IL-9, IL12p40, KC, MCP-1, MIP-1a, MIP-1b, RANTES, TNFa. Allogenic MSCs but not CIMVs increased concentration of Eotaxin, G-CSF, IL-17A and IL-9. The level of GM-CSF, IFN-g, IL-10, IL-1a, IL-1b, IL-2, IL-3, IL-4, IL-5, IL-6, IL12p40, KC, MCP-1, MIP-1a, MIP-1b, RANTES, TNFa in mice serum were not affected by murine MSCs injection. Injection of CIMVs did not induce any statistically significant changes in cytokines level.

Conclusion: Elevated levels of Eotaxin, G-CSF, IL-17A and IL-9 cytokines after the i.v.injection of murine MSCs suggest that moderate allergy inflammation was developed after the MSCs allotransplantation. CIMVs injection did not induced increase of cytokines level in mice serum indicating absence of immunogenicity. Taken together our results demonstrate that CIMVs show less/no immunogenicity compared to parental MSCs. We believe that small diameter, better biodistribution and fusion with host cells lead to the non-immunogenicity of CIMVs. Thus, CIMVs are confirmed to be a perspective, new biomimetic vector system. This study was funded by the grant of the President of the Russian Federation for state support of the leading scientific schools of the Russian Federation НШ-2603.2020.4. Kazan Federal University was supported by the Russian Government Program of Competitive Growth.