The cytoplasmic escape and nuclear accumulation of endocytosed and microinjected HPMA copolymers and a basic kinetic study in hep G2 cells
Journal Title: The AAPS Journal - Year 2001, Vol 3, Issue 4
Abstract
The development of macromolecules as drugs and drug carriers requires knowledge of their fate in cells. To this end, we studied the internalization and subcellular fate of N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers in Hep G2 (human hepatocellular carcinoma) cells. Semiquantitative fluorometry confirmed that galactose was an effective ligand for receptor-mediated endocytosis for Hep G2 cells. The rate of internalization of a galactose-targeted copolymer was almost 2 orders of magnitude larger than that of the nontargeted copolymer. Confocal fluorescent microscopy of both fixed and live cells revealed that the polymer entered the cells by endocytosis. After longer incubation times (typically >8 hours), polymer escaped from small vesicles and distributed throughout the cytoplasm and nuclei of the cells. Polymer that entered the cytoplasm tended to accumulate in the nucleus. Microinjection of the HPMA copolymers into cells' cytoplasm and nuclei indicated that the polymers partitioned to the nucleus. The data from fixed cells was confirmed by microscopy of live, viable cells. To examine the effect of the fluorescent dye on the intracellular fate, polymers with fluorescein, Oregon Green 488, Lissamine rhodamine B, and doxorubicin were tested; no significant differences were observed.
Authors and Affiliations
Keith D. Jensen, Pavla Kopečková, John H. B. Bridge, Jindřich Kopeček
Keywords
Related Articles
- EP ID EP682070
- DOI 10.1208/ps030432
- Views 93
- Downloads 0
A Platform for Characterizing Therapeutic Monoclonal Antibody Breakdown Products by 2D Chromatography and Top-Down Mass Spectrometry
The online version of this article (doi:10.1208/s12248-012-9361-6) contains supplementary material, which is available to authorized users.
Population Pharmacokinetic Modeling of LY2189102 after Multiple Intravenous and Subcutaneous Administrations
Interleukin-1 beta (IL-1β) is an inflammatory mediator which may contribute to the pathophysiology of rheumatoid arthritis (RA) and type 2 diabetes mellitus (T2DM). Population pharmacokinetics (PK) of LY2189102,...
Retrospective Data Analysis and Proposal of a Practical Acceptance Criterion for Inter-laboratory Cross-validation of Bioanalytical Methods Using Liquid Chromatography/Tandem Mass Spectrometry
The purpose of this study is to conduct a retrospective data analysis for inter-laboratory cross-validation studies to set a reasonable and practical acceptance criterion based on a number of cross-validation results. Fr...
Variation of Stratum Corneum Biophysical and Molecular Properties with Anatomic Site
Several serine protease enzymes are known to be involved in both normal desquamation and the inflammatory processes of the skin. Alteration in the activity of these proteases should also affect corneocyte maturity and si...
Effect of rise in simulated inspiratory flow rate and carrier particle size on powder emptying from dry powder inhalers
The purpose of this study was to evaluate the effect of carrier particle size and simulated inspiratory flow increase rate on emptying from dry powder inhalers (DPIs). Several flow rate ramps were created using a compute...