Study Of The Endocytic Pathway Of Heparosan And Heparosan-based Nanocarries For Drug Delivery

The difference of cellular internalization influences the drug delivery efficiency of nano drug delivery systems(NDDSs),as well as the efficacy.In the previous study,we found that the cellular uptake of heparosan-based NDDSs by cancer cells was selective,rapid and efficient.The results showed this phenomenon was related to the endocytic pathway of the NDDSs.To illustrate the cause,this thesis studies the endocytic pathway,uptake efficiency,drug distribution and fate of both heparosan and heparosan-based NDDSs,which was showed as follows:(1)Endocytic pathway and intracellular distribution of heparosanHeparosan was labeled via fluorescent probe to obtain HB,and dextran was used as the control.The experimental results showed that the internalizations of HB by MCF-7,A549 and B16 cancerous cell-lines were specific as compared with that of COS7 normal cell-line.Moreover,the uptake efficiency was correlated with the concentration and incubation periods of HB.The main endocytic pathway of HB by three kinds of cancer cells was caveolae-mediated endocytosis and micropinocytosis,which was energy-dependent.Fluorescent observation showed the HB was distributed in lysosomes.(2)Endocytic pathway and cellular distribution of heparosan-based nanoparticlesHeparosan-based nanoparticles(HD)were obtained by conjugating HB and deoxycholic acid.The average size of HD was determined to be 170 nm,which dispersed well in aqueous medium.The cellular internalization results showed that both normal cell and cancer cells were able to internalized HD efficiently.Moreover,the uptake of HD by cancer cells was more capable than that by normal cells.The main endocytic pathway of HD by three kinds of cancer cells turns into clathrin-mediated endocytosis and micropinocytosis.Yet,the endocytosis of HD was energy-dependent.The internalized HD distributed in lysosomes of cells.(3)Drug delivery of heparosan-based NDDSsHeparosan was modified with deoxycholic acid and tocopherol(Vitamin E),respectively,to obtain two kinds of amphiphilic drug carriers(i.e.,HepD and HepV).These carriers could encapsulate anticancer drug doxorubicin(DOX),which is hydrophobic.The drug loading amount of HDD and HVD was determined to be 4.3% and 4.7%.The cellular experiments showed that both HDD and HVD could rapidly deliver DOX into B16 cells.The accumulated drug amount increased with the extention of incubation period.In addition,the growth of B16 cancer cells was efficiently inhibited by HDD and HVD.In conclusion,it had different endocytic pathways and uptake efficacy between heparosan and heparosan-based nanoparticles,but both distributed and accumulated in lysosomes of normal cells and cancer cells,which provide an experimental reference for constructing heparosan-based nano drug delivery systems that are sensitive to lysosomes of cells.