Presentation Information
[10a-PB5-8]Local Fusion of Exosomes using Micro Laminar-Flow Confinement System
〇Nahoko Kasai1,2, Luo Huan3, Kiyoshi Sato2, Hizuru Nakajima2, Hiroyoshi Kawakami2, Shungo Kato2, Sifeng Mao3 (1.Tokyo Metropolitan Univ, 2.Grad. Tokyo Metropolitan Univ, 3.Soochow Univ)
Keywords:
exosomes,fusion,laminar flow
Introduction
Exosomes are nanosized extracellular vesicles that mediate intercellular communication. Among the mechanisms of cellular uptake, direct membrane fusion between exosomes and the plasma membrane is particularly important because it enables efficient cytosolic delivery of cargo while avoiding endocytic degradation pathways. However, conventional approaches to study exosome–cell fusion often require global changes in the culture environment, which can damage cells.
Methods
We employed a hydrodynamically confined laminar flow system composed of three glass capillaries (two inlets and one outlet). Under appropriate injection (QI), aspiration (QA), and gap conditions, two solution streams form beneath the nozzle, creating a narrow diffusion-based mixing zone. In this study, a low-pH solution (pH 5.0) and an exosome suspension (pH 7.4) were locally introduced to generate a fusion region. Exosomes isolated from A549 cells were applied, and their localized fusion with supported lipid bilayers or cells was observed using inverted fluorescence microscopy.
Results and Discussion
Application of the system to supported lipid membranes revealed exosome fusion within the mixing region. Similar results were obtained with living cells, demonstrating spatially controlled fusion. These findings indicate that this laminar flow system is a promising tool for precise investigation of exosome–cell membrane fusion.
Exosomes are nanosized extracellular vesicles that mediate intercellular communication. Among the mechanisms of cellular uptake, direct membrane fusion between exosomes and the plasma membrane is particularly important because it enables efficient cytosolic delivery of cargo while avoiding endocytic degradation pathways. However, conventional approaches to study exosome–cell fusion often require global changes in the culture environment, which can damage cells.
Methods
We employed a hydrodynamically confined laminar flow system composed of three glass capillaries (two inlets and one outlet). Under appropriate injection (QI), aspiration (QA), and gap conditions, two solution streams form beneath the nozzle, creating a narrow diffusion-based mixing zone. In this study, a low-pH solution (pH 5.0) and an exosome suspension (pH 7.4) were locally introduced to generate a fusion region. Exosomes isolated from A549 cells were applied, and their localized fusion with supported lipid bilayers or cells was observed using inverted fluorescence microscopy.
Results and Discussion
Application of the system to supported lipid membranes revealed exosome fusion within the mixing region. Similar results were obtained with living cells, demonstrating spatially controlled fusion. These findings indicate that this laminar flow system is a promising tool for precise investigation of exosome–cell membrane fusion.