Construction Of Quercetin-loed Biomimetic Nanoparticles And Evaluation Of Blood-brain Barrier Permeability

Diseases of the central nervous system account for 35%of all human diseases and affect approximately 1 billion people worldwide.Among the many neurocentral system disorders Alzheimer’s disease,Parkinson’s disease,stroke,and brain tumors are more prevalent.However,few drugs are available for the treatment of CNS disorders,and therapeutic efficacy is limited by many factors,with inefficient drug transport across the blood-brain barrier being the greatest challenge.The blood-brain barrier is selectively permeable and plays an important role in maintaining the stability of the physiological environment of brain tissue and protecting the CNS from harmful substances or microorganisms in the blood,but the presence of the blood-brain barrier can also limit the access of therapeutic drugs to the brain for the treatment of diseases.Biomimetic nanoparticles are prepared from drug-laden nanoparticles wrapped in biomimetic cell membranes,which have the characteristics of high drug loading capacity,good biocompatibility,long circulation time,strong targeting and low immunogenicity,showing great advantages in intracerebral drug delivery.Erythrocyte membranes are also used to modify the surface of nanoparticles,as theyhas the characteristics of large number.Quercetin（Qu）has been shown to have protective effects on neurons,as well as inhibiting reactive oxygen species and scavenging free radicals,making it a potential drug for the treatment of many neurodegenerative diseases and neuronal damage.However,the hydrophobic nature of Qu and its lower blood-brain barrier permeability limit its use in the treatment of central nervous system diseases.In this study,polymeric nanoparticles loaded with Qu were prepared using erythrocyte membranes to build a biomimetic drug delivery system.The main research of this paper includes the following three parts:（1）Preparation and characterization of Qu nanoparticles（Qu NP）.（2）preparation and characterization of red blood cell membrane-modified Qu nanoparticles（RBCm-Qu NP）.（3）brain targeting study of RBCm-Qu NP.Firstly,Qu NP was prepared by a modified emulsification solvent evaporation method,and the content of Qu was determined by high performance liquid chromatography.In the process of optimal prescription screening,four single factors with high influence were analyzed experimentally,and the best preparation process of Qu NP was obtained by orthogonal experiments with encapsulation rate and drug loading rate as indicators.Through extensive experimental studies,the optimal preparation conditions were obtained as A2B2C3D2,i.e.,a mass concentration of 0.75mg/m L of posoxam 188,a dosage of 4 mg of Qu,a dosage of 56 mg of PCL and a volume ratio of 1:8 of oil phase to water phase.The particle size of RBCm-Qu NP was177.66±5.19 nm,the potential was-10.44±0.85 m V,the entrapment efficiency was63.37%,and the drug loading rate was 4.33%.RBCm-Qu NP was spherical and well dispersed.Secondly,on the basis of the best preparation process of Qu NP screened,the surface of Qu NP was modified by wrapping the red blood cell membrane.The particle size of RBCm-Qu NP was 177.66±5.19 nm,the potential was-10.44±0.85 m V,the entrapment efficiency was 63.37%,and the drug loading rate was 4.33%.RBCm-Qu NP was spherical and well dispersed.Finally,nanoparticles were found to have no significant toxic effects on rat adrenal medullary pheochromocytoma differentiated cell line（PC12 cells）,human brain microvascular endothelial cells（h CMEC/D3 cells）,and human brain astrocytes（U-118MG cells）,and all nanoparticles increased the cell proliferation rate of PC12cells with Aβ_25-35-induced injury.Nanoparticles were prepared using isothiocyanate（FITC）as a fluorescent probe instead of Qu.Quantitative and qualitative analyses of cellular uptake revealed significant values of 0.0004 and 0.0001 for the FITC-NP and RBCm-FITC-NP groups,respectively,compared with the blank group for PC12 cells,and for U-118MG cells 0.0021 and 0.0007,and the significant values for the uptake of h CMEC/D3 cells were 0.0025 and 0.0005,demonstrating that the FITC-NP group and the RBCm-FITC-NP group were significant for the uptake of all three cells.It was also demonstrated that RBCm could increase the uptake of nanoparticles by cells.An in vitro blood-brain barrier model was established to examine the permeability of FITC-NP and RBCm-FITC-NP,respectively,and it was found that the permeability of RBCm-FITC-NP to the blood-brain barrier model was greater compared with that of FITC-NP.The experimental results showed that RBCm-Qu NP could facilitate the entry of Qu into the blood-brain barrier.In recent years,the application of bionic drug delivery systems has become more and more widespread,but there are relatively few applications for the preparation of Qu into bionic nanoparticles for the treatment of brain diseases.The RBCm-Qu NP prepared in this experiment has stronger blood-brain barrier permeability and better therapeutic effect on brain diseases,which provides a new direction and method for the treatment of central nervous system diseases.