title: Dynamic colocalization microscopy to characterize intracellular trafficking of nanomedicines.
abstract: To gain a better understanding of intracellular processing of nanomedicines, we employed quantitative live-cell fluorescence colocalization microscopy to study endosomal trafficking of polyplexes in retinal pigment epithelium cells. A new, dynamic colocalization algorithm was developed, based on particle tracking and trajectory correlation, allowing for spatiotemporal characterization of internalized polyplexes in comparison with endosomal compartments labeled with EGFP constructs. This revealed early trafficking of the polyplexes specifically to Rab5- and flotillin-2-positive vesicles and subsequent delivery to Rab7 and LAMP1-labeled late endolysosomes where the major fraction of the polyplexes remains entrapped for days, suggesting the functional loss of these nanomedicines. Colocalization of polyplexes with the autophagy marker LC3 suggests for the first time that the process of xenophagy could play an important role in the persistent endosomal entrapment of nanomedicines.
authors.affiliation: Laboratory of General Biochemistry and Physical Pharmacy, Ghent University, Harelbekestraat 72, B-9000 Ghent, Belgium.
authors.forename: Dries
authors.initials: D
authors.lastname: Vercauteren
authors.forename: Hendrik
authors.initials: H
authors.lastname: Deschout
authors.forename: Katrien
authors.initials: K
authors.lastname: Remaut
authors.forename: Johan F J
authors.initials: JF
authors.lastname: Engbersen
authors.forename: Arwyn T
authors.initials: AT
authors.lastname: Jones
authors.forename: Jo
authors.initials: J
authors.lastname: Demeester
authors.forename: Stefaan C
authors.initials: SC
authors.lastname: De Smedt
authors.forename: Kevin
authors.initials: K
authors.lastname: Braeckmans
mesh_terms: D001343:Autophagy; D001692:Biological Transport; D002460:Cell Line; D002470:Cell Survival; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003911:Dextrans / Q000378:metabolism; D004337:Drug Carriers / Q000737:chemistry* / Q000378:metabolism*; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism; D005786:Gene Expression Regulation; D015316:Genetic Therapy; D042541:Intracellular Space / Q000378:metabolism*; D007700:Kinetics; D008856:Microscopy, Fluorescence / Q000379:methods*; D050997:Nanomedicine / Q000379:methods*; D053758:Nanoparticles / Q000737:chemistry*; D011108:Polymers / Q000737:chemistry / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D055213:Retinal Pigment Epithelium / Q000166:cytology / Q000378:metabolism
chemical_list: D003911:Dextrans; D004337:Drug Carriers; D011108:Polymers; D012333:RNA, Messenger; D004247:DNA