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Drug Delivery

Drug Delivery - Poster Session E

Poster X10 - Quantifying Propidium Iodide Uptake by Electroporation

Saturday, October 26, 2024
10:00 AM - 11:00 AM EST
Location: Exhibit Hall E, F & G

Introduction:  Endovascular therapies, such as balloon angioplasty and stent deployment, have enhanced blood vessel treatment, but their ability to maintain long-term channel patency remains limited. These treatments, which often require administering therapeutic drugs directly to the artery walls, depends on vessel patency. Current techniques rely on balloon catheters that release medication gradually or use stents coated with therapeutic substances. However, these methods leave foreign objects within the vessel or cause the therapeutic to gradually wash out because of blood flow. Better technologies that increase drug delivery without leaving behind permanent materials are required to address these limitations.
We have created a novel method that combines current balloon catheter technology with electroporation. By using electrical pulses, cell membrane permeability is increased during electroporation, enhancing molecular interaction between the external and intracellular environments. Though promising, little is known about the use of electroporation in the vascular system. By investigating the relationship between electroporation voltage and propidium iodide (PI) uptake in both ex vivo and in vitro models, this study aims to address this gap. The following investigations begins to lay the groundwork for better therapeutic drug distribution to vessel walls through electroporation parameter optimization.

Materials and

Methods:  In vitro experiments: Human vascular smooth muscle cells (VSMCs) were plated on fibrin-coated 2-well chamber slides. Propidium iodide (PI) was added to a concentration of 1 mg/mL to the wells, while control wells were treated with Tyrode solution. Electroporation was conducted using needle electrodes spaced 5 mm apart, mounted on a pressure detector and vertical micromanipulator to ensure accurate placement and avoid contact pressure on the well floor. A square wave pulse train with 8 repetitions, 50 µs on, 100 ms off, was applied using an ECM® 630 electroporation system at voltages of 160V, 320V, and 480V. Control conditions were processed first to avoid cross-contamination. Post-electroporation, the slides were fixed in paraformaldehyde and subsequently imaged using blue-fluorescent DAPI and PI to assess cellular uptake.
Ex vivo experiments: Thoracic pig aorta segments were carefully harvested and sectioned, minimizing trauma during transport to the chamber slide. A duplicate mold with parallel electrodes (5 mm apart, 12 mm in length) was used, and a pressure of 1N was applied to ensure consistent contact with the vessel segment. After electroporation at the specified voltages, the sections were washed in phosphate-buffered saline, embedded in Optimal Cutting Temperature (OCT) compound, and frozen. Frozen sections (10 µm) were mounted on microscope slides, covered, and then imaged using DAPI and PI fluorescence to evaluate PI uptake in the vascular tissue.

Results, Conclusions, and Discussions: Our experiments demonstrate a clear relationship between electroporation voltage and increased propidium iodide (PI) uptake in ex vivo vascular tissues, suggesting that electroporation enhances molecular delivery to the vessel walls. In the thoracic pig aorta segments, the mean gray values for PI uptake increased significantly with voltage: from 17.997 at 0V to 36.44 at 480V. This indicates that higher voltages lead to greater permeability and enhanced uptake of the extracellular marker PI.
These findings are consistent with our in vitro results using human vascular smooth muscle cells (VSMCs). Observing the graph from 0V with a max below 1000 to 480V surpassing this threshold along the entire electrode gap suggest a similar trend. This supports the notion that electroporation effectively enhances cell membrane permeability in a consistent manner across different experimental setups.
The increased PI uptake observed in both ex vivo and in vitro models highlights the potential of electroporation to improve therapeutic delivery. By modulating the voltage, we can enhance the permeability of the vessel walls or cell membranes, facilitating more effective drug transfer. This advancement addresses a key challenge in endovascular treatments, where optimizing drug delivery is crucial for maintaining vessel patency and achieving successful therapeutic outcomes.
Despite these promising results, it is essential to consider the impact of electroporation on cell and tissue health. Elevated PI uptake may indicate increased permeability, but it is crucial to ensure that this does not adversely affect cellular viability or tissue integrity. To address these concerns, we will perform additional assessments, including TUNEL assays to evaluate apoptosis and Ki-67 staining to measure cell proliferation. These analyses will provide a more comprehensive understanding of the effects of electroporation on cell and tissue health, ensuring that enhanced drug delivery does not compromise overall tissue viability.
In conclusion, our study supports the potential of electroporation as a valuable tool for improving drug delivery in endovascular therapies. Future research will focus on refining electroporation parameters and assessing long-term effects to establish its clinical viability and optimize its application for vascular treatments.

Acknowledgements (Optional):  I would like to thank our senior researcher and my co-fellow for their help in conducting these experiments and analyzing the data. I would also like to thank my supervisor for their guidance and support