Preparation Of Liposome Fusion Vesicles Of Stem Cell Membrane And Its Application In The Treatment Of Vascular Obstruction

The size,shape,chemical composition,surface structure and charge of nanomaterials can significantly affect their interactions with biomolecules and cells,so they have received extensive attention from researchers.However,due to the short circulation time,insufficient driving force,poor targeting and other problems,nanomaterials have not been widely used in the clinical biomedical field.Especially in vascular obstructive diseases,because of the blood clearance effect and the obstruction of complex physiological structures and tissue physiological barriers,the delivery rate of nano carriers targeting lesions has been greatly limited.To solve the above problems,this paper designed a nano drug loading platform for the fusion of liposome vesicles on the stem cell membrane based on the biomimetic strategy of the cell membrane,which not only inherits the functions of immune escape and targeting inflammation of stem cells,but also has the characteristics of easy drug loading and surface modification of lipid vesicles,which is expected to improve the drug delivery efficiency of vascular obstruction disease.In view of the physiological characteristics of retinal thrombosis and tissue ischemia and the demand for drug delivery,two types of liposome fusion vesicles of stem cell membrane were designed and developed in this paper.The research work is as follows:（1）To solve the problem of low delivery efficiency of traditional drugs for retinal thrombosis,a vitreous injection delivery strategy based on magnetic stem cell membrane liposome fusion vesicles was designed,which can overcome the vitreous barrier under the effect of external magnetic field,and target the inflammatory site of blocked blood vessels through the stem cell membrane to improve the drug delivery efficiency.First,magnetic stem cell membrane liposome fusion vesicles（MFL）were prepared by liquid nitrogen freeze-thaw and ultrasonic emulsification,and Fe₃O₄ nanoparticles,L-arginine and perfluorohexane were successfully loaded.The size of MFL is uniform,and the average particle size is about 350 nm.Western Blot and cell experiment results show that the stem cell membrane and liposome are successfully fused,which can produce nitric oxide under the effect of ultrasound for dilating and blocking blood vessels.The animal model of retinal vein thrombosis in rats showed that the migration ability of MFL was 2.3 times higher than that of the liposome fusion vesicle of nonmagnetic drug loaded stem cell membrane;The targeting ability of MFL is 2.0 times higher than that of magnetic liposomes.Compared with MFL without ultrasound,MFL with ultrasound reduced the density of new blood vessels and the number of nodes by 2.5 times and 2.0times,respectively.This strategy can significantly indicate the therapeutic efficiency of retinal vein thrombosis.（2）To solve the problem of low drug delivery efficiency in treating tissue ischemia caused by vascular obstruction,a stem cell membrane liposome fusion vesicle（FLPV）loaded with vascular endothelial growth factor（VEGF）was designed.It can target the inflammatory site of tissue ischemia through the stem cell membrane surface protein CXCR4,and release VEGF via ultrasound to promote angiogenesis,thereby relieving tissue ischemia.First,the drug loaded liposome fusion vesicles were prepared by liquid nitrogen freeze-thaw followed by ultrasonic emulsification,and were successfully loaded with perfluorohexane and VEGF,which were applied to the treatment of ischemic thrombosis.TEM images of FLPV and DLS show that FLPV has uniform size,and the average particle size is about 330 nm.Fluorescence resonance energy transfer diagram and Western Blot experiment showed that the FLPV surface had a chemokine CXCR4 and successfully loaded VEGF.Cell experiment,H&E staining sections of various tissues and blood routine analysis showed that FLPV had low cytotoxicity and good biocompatibility.In addition,after ultrasound was applied,FLPV achieved the effect of endothelial cell proliferation in vitro by releasing VEGF,and its endothelial proliferation ability was 2.7 times and 2.5 times that of the liposome fusion vesicle group of stem cell membrane and the FLPV group without ultrasound respectively.At the same time,FLPV can also use the chemokine CXCR4 on the fusion membrane to achieve tissue ischemia caused by vascular occlusion in vitro through CXCR4-SDF-1 axis.