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High School Poster

HS Poster

Poster 59 - Effects of Inflammation on Blood-Brain Barrier Permeability

Saturday, October 26, 2024
10:00 AM - 12:00 PM EST
Location: Exhibit Hall E, F & G

Introduction: The blood-brain barrier (BBB) is a selectively permeable membrane that protects the brain from foreign molecules. This barrier is made up of mostly brain endothelial cells, and junction proteins hold these cells together to block unwanted particles from passing through the membrane. While this membrane is meant to protect the brain from harm, it also stops many treatment molecules from reaching the brain and treating brain diseases. This can lead to off-target accumulation of drugs, which can cause further damage to one’s health. However, inflammation can cause greater permeability of the BBB, making it easier for molecules to pass through it. This is essential for effective drug delivery to the brain to properly treat brain diseases. In addition to inflammation, nanoparticles (NPs) can also increase the amount of drugs that can pass through the BBB and reach the brain. Because of their small size, NPs are able to pass through barriers such as the BBB more easily than other molecules. NPs can also be modified to target specific disease sites, further increasing the efficiency of drug delivery. However, the BBB is still a significant obstacle that stops many NPs from reaching the brain and their targeted disease sites. Thus, this research study will examine the question: to what extent does inflammation of the BBB impact its permeability and the ability of cancer nanomedicine to cross this barrier? By pairing inflammation with nanoparticle drug delivery, treatment of brain diseases, such as cancer, may become more successful.

Materials and

Methods: To quantify the level of permeability in inflamed cells, hCMEC/D3 brain endothelial cells were grown until confluent on chamber slides and treated with TNF-ɑ, an inflammatory agent, for 24 hours. The cells were then stained using DAPI and VE-cadherin to visualize the cell nuclei and adherens junctions, respectively. Images of each well were taken using an EVOS M5000 microscope. The computer program ImageJ was used to quantify the number of cells in each well as well as the total VE-cadherin signal in each image to quantify the average VE-cadherin signal per cell. To quantify the effects of inflammation on the ability of various molecules to migrate across the endothelial cell barrier, hCMEC/D3 cells were grown in transwell inserts, particles were added into the inner wells, and samples from the outer wells were collected at specific timepoints. Two particle types were tested: FITC-dextran and Cy5-PLGA nanoparticles. A plate reader was used to measure either the FITC or Cy5 fluorescence signals of the particles present in each sample. The results were then compared to a known amount of either FITC or Cy5 to quantify relative signal.

Results, Conclusions, and Discussions: The results of the VE-cadherin quantification assay demonstrated that the cells treated with TNF-ɑ expressed significantly less VE-cadherin by approximately 25% (Figure 1B). Adherens junctions are proteins that hold endothelial cells together, so less VE-cadherin signal indicates an increase in the permeability of the BBB due to a decreased presence of adherens junctions. The transwell migration assay demonstrated that inflammation of the cells increased the amount of particles that passed through the inserts, further proving that inflammation can increase the permeability of the BBB. In the FITC-dextran assay, the cells treated with inflammation had around 1.5% more particle flowthrough than the cells without at both thirty and sixty minutes (Figure 1C). Similarly, the Cy5-PLGA NP assay demonstrated that the inflamed cells had around 1% (one hour), 3% (four hours), and 7% (24 hours) more flowthrough than the untreated cells (Figure 1D). These findings can help to optimize treatment of brain diseases, specifically brain cancer, by increasing BBB permeability and allowing more therapeutics to be delivered directly to the brain. The PLGA NPs used in the transwell assay are often used to deliver drugs to treat brain tumors, so the results of this study indicate a potential avenue to improve their delivery to diseased sites in the brain. Furthermore, inflammation in the BBB could be a less invasive method of drug delivery to the brain due to it being a naturally occurring immune response. However, inflammation can hinder the overall BBB functionality and cause further health issues, so further research must be done to test the viability of this drug delivery method. Future studies can be performed to test the optimal level of inflammation in the BBB to increase the effectiveness of drug delivery but not cause further health issues. In conclusion, the results of this study indicate that inflammation increases BBB permeability and the ability of cancer nanomedicine to cross the BBB. These findings can be applied to clinical needs through the development of a more effective and less invasive method of delivering drugs to treat brain tumors and other maladies.

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