title: Investigation of microsphere-mediated cellular delivery by chemical, microscopic and gene expression analysis.
abstract: Amino functionalized cross-linked polystyrene microspheres of well defined sizes (0.2-2 mum) have been prepared and shown to be efficient and controllable delivery devices, capable of transporting anything from small dye molecules to bulky proteins into cells. However, the specific mechanism of cellular entry is largely unknown and widely variant from study to study. As such, chemical, biological and microscopic methods are used to elucidate the mechanism of cellular uptake for polystyrene microspheres of 0.2, 0.5 and 2 mum in mouse melanoma cells. Uptake is found to be wholly unreliant upon energetic processes, while lysosomal and endosomal tracking agents failed to show co-localisation with lysosomes/endosomes, suggesting a non-endocytic uptake pathway. To further explore the consequences of microsphere uptake, gene expression profiling is used to determine if there is a transcriptional response to "beadfection" in both murine and human cells. None of the common transcriptional responses to enhanced endocytosis are observed in beadfected cells, further supporting a non-endocytic uptake mechanism. Furthermore, the microspheres are noted to have a limited interaction with cells at a transcriptional level, supporting them as a non-toxic delivery vehicle.
authors.affiliation: Chemical Biology Section, School of Chemistry, University of Edinburgh, West Mains Road, Edinburgh, UK.
authors.forename: Lois M
authors.initials: LM
authors.lastname: Alexander
authors.forename: Salvatore
authors.initials: S
authors.lastname: Pernagallo
authors.forename: Alessandra
authors.initials: A
authors.lastname: Livigni
authors.forename: Rosario M
authors.initials: RM
authors.lastname: Sánchez-Martín
authors.forename: Joshua M
authors.initials: JM
authors.lastname: Brickman
authors.forename: Mark
authors.initials: M
authors.lastname: Bradley
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D021941:Caveolae / Q000378:metabolism; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002784:Cholesterol / Q000378:metabolism; D002966:Clathrin / Q000378:metabolism; D016503:Drug Delivery Systems / Q000379:methods*; D011992:Endosomes / Q000378:metabolism; D019793:Fluorescein / Q000378:metabolism; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D008247:Lysosomes / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal; D008863:Microspheres; D008870:Microtubules / Q000378:metabolism; D010873:Pinocytosis; D011137:Polystyrenes / Q000008:administration & dosage / Q000737:chemistry / Q000493:pharmacokinetics* / Q000494:pharmacology; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013696:Temperature
chemical_list: D002966:Clathrin; D011137:Polystyrenes; D000255:Adenosine Triphosphate; D002784:Cholesterol; D019793:Fluorescein