The Cell Trafficking And Autophagy Pathway Of Microgel And Its Application On Cancer Treatment

Microgels(also known as hydrogel)have been widely used in the treatment of cancer as an efficient tools for drug delivery.Inner membrane vesicle system which mainly includes endocytosis,exocytosis and autophagy is a complex transport system.Autophagy is an important mechanism for the degradeation of cellular protein aggregates,damaged organelles and the invaded pathogenic microorganisms,which is responsible for maintaining the stability of the intracellular environment.Deficiency of autophagy,may lead to a variety of diseases,such as cancer,neurodegenerative diseases and diabetes.However,the details of the intracellular trafficking pathway of the microgel is poorly investigated and understood.In the present study,we synthesized microgels by free radical precipitation polymerization method,predetermined amount of SDS,NIPAM and PolyFluor,in this mixture added BIS as crossliker,KPS was added to initiate the polymerization.The microgels were characterized(size and size distribution,surface morphology,drug loading content,n vitro drug release profile).The diameters of the three types of Rhodamine-loaded microgels are 52.5 nm(PN1),101.3 nm(PN2),299.8 nm(PN3).The diameters was satisfied to EPR effect.The in vitro drug release profile of the drug loaded microgel was determined.The CQ-loaded microgel released about 50% drug in the first 2 hours,and accumulative up to 95% in 12 hours.In contrast,the DOX-loaded microgel just released only 47% drug in 12 hours.Additionally,we applied the Rhodamine-loaded microgels to MCF-7 cells and investigated the cellular uptake via transfection,immunofluorescence,immunoprecipitation and other means.We found that Rhodamine-loaded microgels were internalized by the cells through multiple endocytosis.Transported to EEA1 and Sorting nexins 3(SNX3)positive early endosome(EEs),and then transported to Rab7 positive late endosome(LEs),followed by transportation to LAMP1 positive lysosome.Except this canonical pathway,we also found the microgels were degraded through the autophagy pathway.The microgels were found in autophagosome(LC3 positive)and finally to lysosome(LAMP1 positive).Finally,in order to investigate the curative effect of chemotherapeutic drug-loadedmicrogel combination with autophagy inhibitors,the xenograft SCID nude mice model with the MCF-7 cell line implantation was also constructed.Co-delivery of autophagy inhibitor chloroquine(CQ)and chemotherapeutic drug doxorubicin(DOX)by microgel greatly decreased both the volume and weight of the tumors in severe combined immunodeficient nude mice,which exhibited effective therapeutic effects on tumors.These findings provide valuable evidences for the development of nanomedicine for clinical application.