Preparation of chloroquine liposomes and its influence on exosome secretion
Objective:To establish a highly efficient exosomal secretion promoting technology system by preparing chloroquine liposomes(CQ-Lips)and exploring the effects of CQ-Lips on the secretion of exosomes.Methods:Using soybean lecithin and cholesterol as membrane materials,the best prescription of CQ-Lips was screened by single factor experiment.The effects of CQ-Lips loaded with different concentrations of CQ on cell viability were explored through CCK-8 experiments.CQ-Lips loaded with chloroquine of 5～100 μmol·L-1 were incubated with cells,taking exosome surface markers CD9 and TSG101 as indicators to detect the secretion of exosomes to determine the optimal liposome drug loading concentration.CQ-Lips loaded with chloroquine of 5 and 10 μmol·L-1 were incubated with cells for different time,then the secretion of exosomes was detected to determine the optimal time point for collecting supernatant.Results:CQ-Lips were prepared under the optimum condition which was as follows:phospholipid mass ratio 1∶80,phospholipid cholesterol mass ratio 3.75∶1,incubated at 20℃for 10 min,freeze-thawed for twice.The resulting product had uniformly sized particles with the encapsulation rate reaching to 72.71%;CCK-8 results showed that CQ of 20 μmol·L-1 alone had no effect on cell activity.For different concentration groups of CQ in CQ-Lips,the cell activity decreased gradually with the increasing of CQ concentration.The cell activity was higher for 5 and 10 μmol·L-1 CQ-loaded liposomes,but gradually decreased with the prolongation of incubation time.The effect of CQ-Lips on exosome secretion was better than that of CQ alone,CQ-Lips containing CQ of 5 μmol·L-1 had the best effect.The secretion of exosomes decreased with the prolonging of incubation time,and 48 h was the best time point for collecting cell supernatant.Conclusion:In this study,more chloroquine was promoted to enter the lysosome through preparation of chloroquine liposomes with the best prescription,so the production of exosomes could be increased by the action of reducing the exosome degradation by lysosome,which provided an experimental basis for the basic research and clinical use of exosomes.
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