The Research Of Tat Peptide-Assisting USPIO Crossing The Blood Brain Barrier Model

Objective To structure blood brain barrier in vitro using brain microvascular endothelial cells and astrocytes.Methods 1.primary culture of astrocytes:cerebral cortex was abtained from Kunming white old-fetal mice,treated with trypsin,and filter.Then remove fibroblasts with differential attachment,and eliminate oligodentrocytes and microglial cells with orbital shaker.Check the cells with immunostaining for glial fibrillary acidic protein(GFAP);2.primary culture of brain microvascular endothelial cells:cerebral cortex was obtained from 1 to 3 days new-born Kunming white mice,digested by collagenaseâ…¡,and treated with BSA gradient centrifugation and second enzyme digestion of collagenase/dispase.Then change the culture solution in the next 24h to pury the cells. Checked by immunostaining forâ…§factor associated antigen;3.co-culture passageâ…¡brain microvascular endothelial cells and passageâ…£astrocytes by transwell inserts.Evaluate the Brain Blood Barrier model by AKP transpeptidase activity,4h permeation test,and the permeation of Horseradish Peroxidase(HRP).Results passageâ…£Astrocytes was characterized by GFAP immunostaining, and positive rate was 95%;passageâ…¡brain brain microvascular endothelial cells was characterized byâ…§factor immunostaining,and positive rate was 90%.The AKP activity of microvascular endothelial cells was higher in co-culture model than monolayers.Conclusion We can establish stable blood brain barrier model in vitro,which lay the foundations for further research of opening the BBB.And it's also a good model to tentatively identify the permeation of drugs for diagnosis and treatment. Objective To detect whether tat peptide-assisting superparamagnetic iron oxide particles (Tat-MNPs) can pass the blood-brain barrier.Methods Using transwell co-culture construct blood-brain barrier model in vitro.With a different dilution of FITC labeled Tat peptide-mediated superparamagnetic iron oxide particles (FITC-Tat-MNPs) joined the transwell for the pool, in the different time periods, using inverted fluorescence microscope by the pool if there are fluorescent display, to determine the FITC-Tat-MNPs can pass the blood-brain barrier model.Results different levels of green fluorescence were visible on PET membrane after the 4h FITC-Tat-MNPs joined in, and relatived with FITC-Tat-MNPs diluted multiples,and 4d after the green fluorescence can be seen in the cells at the bottom of six plates.Conclusion FITC-Tat-MNPs may be through the blood-brain barrier model, and maybe crossed by penetrating cells, it provided a good carrier of nuclear magnetic resonance technology for early diagnosis of Alzheimer disease.