RESUMO
Multiple sclerosis is a disease caused by autoantigen-responsive immune cells that disrupt the myelin in the central nervous system (CNS). Although immunosuppressive drugs are used to suppress symptoms, no definitive therapy exists. As in the experimental autoimmune encephalitis (EAE) model of multiple sclerosis, a partial sequence of the myelin oligodendrocyte glycoprotein (MOG35-55) was identified as a causative autoantigen. This suggests that the induction of immune tolerance that is specific to MOG35-55 would be a fundamental treatment for EAE. We previously reported that lipid nanoparticles (LNPs) containing an anionic phospholipid, phosphatidylserine (PS), in their lipid composition, can be used to deliver mRNA and that this leads to proteins of interest to be expressed in the spleen. In addition to the targeting capability of PS, PS molecules avoid activating the immune system. Physiologically, the recognition of PS on apoptotic cells suppresses immune activation against these cells by releasing cytokines, such as interleukin-10 (IL-10) and transforming growth factor (TGF)-ß that negatively regulate immunity. In this study, we tested whether mRNA delivery of autoantigens to the spleen by PS-LNPs causes the expression of MOG35-55 antigens with minimal immune stimulation and whether this could be used to treat an EAE model by inducing immune tolerance.
RESUMO
The blood-brain barrier (BBB), which is comprised of brain capillary endothelial cells, plays a pivotal role in the transport of drugs from the blood to the brain. Therefore, an analysis of proteins in the endothelial cells, such as transporters and tight junction proteins, which contribute to BBB function, is important for the development of therapeutics for the treatment of brain diseases. However, gene transfection into the vascular endothelial cells of the BBB is fraught with difficulties, even in vitro. We report herein on the development of lipid nanoparticles (LNPs), in which mRNA is encapsulated in a nano-sized capsule composed of a pH-activated and reductive environment-responsive lipid-like material (ssPalm). We evaluated the efficiency of mRNA delivery into non-polarized human brain capillary endothelial cells, hCMEC/D3 cells. The ssPalm LNPs permitted marker genes (GFP) to be transferred into nearly 100% of the cells, with low toxicity in higher concentration. A proteomic analysis indicated that the ssPalm-LNP had less effect on global cell signaling pathways than a Lipofectamine MessengerMAX/GFP-encoding mRNA complex (LFN), a commercially available transfection reagent, even at higher mRNA concentrations.
