Targeting brain inflammation with bioconjugated nanoparticles
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Brain inflammation has been implicated with the pathogenesis of neurodegenerative diseases. Activated microglia and endothelial cells induce production of reactive oxygen species (ROS) and overexpress pro-inflammatory mediators that perpetuate tissue damage. Current treatments are not effective against progressive stages of neurodegenerative diseases and more advanced therapies need to be developed. Recently, nanomaterials have been investigated for therapeutic applications. Nanoparticles can increase efficiency of drug delivery due to increased tissue distribution and the ability to modify surface chemistry to increase biocompatibility and incorporate targeting moieties. In the present study, we established in vitro and in vivo brain inflammation models by administering lipopolysaccharide to mouse brain endothelial cells, microglia, macrophage cells and C57BL/6 male mice. Changes in mRNA expression of pro-inflammatory mediators were analyzed by real-time reverse transcriptase-polymerase chain reaction (RT-PCR). Tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP-1), E-selectin, and intercellular adhesion molecule-1 (ICAM-1) displayed significant overexpression when compared to the control. Additionally, folate receptor-α (FR-α) was also overexpressed, confirming that our model will function appropriately for specific targeting experiments. Cellulose nanocrystals are rod-like particles, approximately 5 nm wide and 100-150 nm long. The surface area consists of extended hydroxyl groups and the structure is hydrophilic in nature. These characteristics make cellulose nanocrystals ideal for surface modification and ensuring long blood circulation half-life. Cell viability was determined using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] conversion assay and a Lactate Dehydrogenase (LDH) Cytotoxicity Detection Kit. At each concentration of cellulose nanocrystals (10, 25, 50 μg/mL), both assays showed the nanoparticles to be non-toxic. Binding/uptake experiments utilizing a fluorescence plate reader and fluorescence microscope showed no non-specific uptake of untargeted cellulose nanocrystals. In contrast, when conjugated to folic acid, cellulose nanocrystals were selectively incorporated to folate receptor-overexpressing cells. These results indicate that both in vitro and in vivo brain inflammation models can be utilized to assess therapeutic efficacy of folate receptor-targeted bioconjugated nanoparticles.
- Masters Theses