Extracellular Delivery Induced by Ultrasound and Microbubbles in Cells
Ultrasound and microbubble treatment (USMB) can enhance the intracellular uptake of molecules, which otherwise would be excluded from the cell, through USMB-mediated transient membrane disruption and through enhanced endocytosis. However, the effect of USMB on the outward movement of molecules from cells is not well understood. This study investigates the effects of USMB on the release of molecules from various cellular compartments including cytoplasm, lysosomes, and recycling endosomes. In vitro ARPE-19 (RPE henceforth) cells were loaded with Alexa fluor- labeled transferrin as a marker for recycling endosomes, LAMP-1 antibody was used to detect the fusion of lysosomes with the plasma membrane, GFP-transfected RPE cells were used to examine the release of GFP from the cytoplasm, and 7-AAD was used to assess cell viability. Subsequently, cells were exposed to USMB (106 cells/mL, 300 kPa peak negative pressure, 1 min treatment duration, and 20 μL/mL Definity microbubbles). Following USMB, the release of the fluorescent markers was examined at 1.5, 11.5, and 21.5 minutes from the start of USMB. The mean fluorescent intensity (MFI) of untreated and USMB treated samples were measured using flow cytometry. USMB increased the extracellular delivery of GFP molecules from the cytoplasm; the MFI in USMB treated GFP-transfected RPE cells decreased by 17% in viable cells and this MFI decreased by 70% in non-viable cells. This could be due to diffusion of GFP through the membrane disruptions induced by USMB. Additionally, the MFI of viable cells stained with LAMP-1 antibody increased by 50% and this increase was 15 folds in the non-viable cells indicating lysosome exocytosis as a mechanism for membrane repair. Furthermore, the MFI of cells loaded with fluorescent transferrin decreased by 22% after USMB treatment in viable cells, indicating a significant increase in transferrin recycling to the cell membrane. However, the increased recycling was not statistically significant in the non-viable cells. This indicates that the increase in transferrin recycling was through an active mechanism that was triggered or enhanced by USMB. It was concluded from this study that USMB enhances the release of molecules from the cytoplasm, lysosomes, and recycling endosomes.
Faculty of Applied Science and Technology (FAST)
AIP Conference Proceedings
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Original Publication Citation
Hussein, F., Antonescu, C., & Karshafian, R. (2017). Extracellular delivery induced by ultrasound and microbubbles in cells. AIP Conference Proceedings, 1821(1). https://doi.org/10.1063/1.4977636
Hussein, Farah; Andonescu, Costin; and Karshafian, Raffi, "Extracellular Delivery Induced by Ultrasound and Microbubbles in Cells" (2017). Publications and Scholarship. 82.