TY - JOUR
T1 - In Vitro Delivery of Cell Impermeable Phallotoxin Using Cationic Liposomes Composed of Lipids Bearing Lysine Headgroup
AU - Sarker, Satya Ranjan
AU - Takikawa, Masato
AU - Takeoka, Shinji
N1 - Funding Information:
This project was partially supported by the GCOE "Practical chemical wisdom" and "High-Tech Research Centre" project for Waseda University: matching fund subsidy from MEXT, Japan. We thank Dr. Ueda (RIKEN Cluster for Pioneering Research) and the support by the Materials Characterization Support Unit, RIKEN CEMS, for TEM observation and the discussion. Also, this project was partially supported by the Leading Graduate Program in Science and Engineering, Waseda University, from MEXT, Japan. We appreciate the Edanz Group (www.edanzediting.com/ac) for editing the manuscript draft.
Funding Information:
This project was partially supported by the GCOE “Practical chemical wisdom” and “High-Tech Research Centre” project for Waseda University: matching fund subsidy from MEXT, Japan. We thank Dr. Ueda (RIKEN Cluster for Pioneering Research) and the support by the Materials Characterization Support Unit, RIKEN CEMS, for TEM observation and the discussion. Also, this project was partially supported by the Leading Graduate Program in Science and Engineering, Waseda University, from MEXT, Japan. We appreciate the Edanz Group ( www.edanzediting.com/ac ) for editing the manuscript draft.
Publisher Copyright:
Copyright © 2020 American Chemical Society.
PY - 2020/4/20
Y1 - 2020/4/20
N2 - Bioactive peptides, which act as biologically active regulators, often require intracellular delivery systems to access their therapeutic targets in the cytosolic space maintaining their bioactivity. Here, we report on the delivery of a polar cell impermeable bioactive peptide, phalloidin, into living HeLa cells with cationic liposomes prepared from lysine-based lipids. Liposome/Alexa Fluor 594 phalloidin complexes were characterized regarding their size and zeta potential, which were 85 ± 38 nm and +24.5 ± 4.21 mV, respectively. The delivery of Alexa Fluor 594 phalloidin into live HeLa cells with K3C14 liposomes was evaluated using a fluorescence activated cell sorter and confocal laser scanning microscopy. The highest Alexa Fluor 594 phalloidin delivery efficiency was 92% when using 200 μg of the cationic lipid/1 × 105 cells seeded at 37 °C. The cellular uptake mechanism for the cationic liposome/Alexa Fluor 594 phalloidin complexes was investigated using various endocytosis inhibitors. We confirmed the complexes were mainly taken up through caveolae-mediated endocytosis. Incubation with bafilomycin A1, which inhibits the acidification of lysosomes, revealed that Alexa Fluor 594 phalloidin did not pass through the lysosomal pathway. Rather, Alexa Fluor 594 phalloidin was released from early endosomes or caveosomes to the cytosol to exhibit its bioactive effects including the multinucleation of HeLa cells.
AB - Bioactive peptides, which act as biologically active regulators, often require intracellular delivery systems to access their therapeutic targets in the cytosolic space maintaining their bioactivity. Here, we report on the delivery of a polar cell impermeable bioactive peptide, phalloidin, into living HeLa cells with cationic liposomes prepared from lysine-based lipids. Liposome/Alexa Fluor 594 phalloidin complexes were characterized regarding their size and zeta potential, which were 85 ± 38 nm and +24.5 ± 4.21 mV, respectively. The delivery of Alexa Fluor 594 phalloidin into live HeLa cells with K3C14 liposomes was evaluated using a fluorescence activated cell sorter and confocal laser scanning microscopy. The highest Alexa Fluor 594 phalloidin delivery efficiency was 92% when using 200 μg of the cationic lipid/1 × 105 cells seeded at 37 °C. The cellular uptake mechanism for the cationic liposome/Alexa Fluor 594 phalloidin complexes was investigated using various endocytosis inhibitors. We confirmed the complexes were mainly taken up through caveolae-mediated endocytosis. Incubation with bafilomycin A1, which inhibits the acidification of lysosomes, revealed that Alexa Fluor 594 phalloidin did not pass through the lysosomal pathway. Rather, Alexa Fluor 594 phalloidin was released from early endosomes or caveosomes to the cytosol to exhibit its bioactive effects including the multinucleation of HeLa cells.
KW - amino lipid
KW - bioactive peptide
KW - cationic liposomes
KW - endosomal/lysosomal escape
KW - intracellular delivery
KW - liposomal complex
KW - multinucleation
KW - phalloidin
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U2 - 10.1021/acsabm.9b01167
DO - 10.1021/acsabm.9b01167
M3 - Article
AN - SCOPUS:85085842109
VL - 3
SP - 2048
EP - 2057
JO - ACS Applied Bio Materials
JF - ACS Applied Bio Materials
SN - 2576-6422
IS - 4
ER -