Document Detail

Polyplex-mediated gene transfer and cell cycle: effect of carrier on cellular uptake and intracellular kinetics, and significance of glycosaminoglycans.
MedLine Citation:
PMID:  17410614     Owner:  NLM     Status:  MEDLINE    
BACKGROUND: Here we report on studies that probe whether the intracellular kinetics of plasmid DNA (pDNA) and cell surface glycosaminoglycans (GAGs) are modified during the cell cycle in a way that can be correlated with changes in gene transfer efficiency with poly(ethyleneimine) (PEI) and poly-L-lysine (PLL) polyplexes. METHODS: Synchronized D407 retinal cells were transfected with PEI and PLL polyplexes using a luciferase reporter. The free and/or loosely complexed nuclear pDNA was determined by real-time PCR, and compared with transgene expression, the rate of pinocytosis by FITC-dextran uptake and the content of cell surface GAGs. RESULTS: The amount of free and/or loosely complexed nuclear pDNA between cell cycle phases varied approximately 4-20 times (G1 < S < G2/M). Both carriers delivered pDNA in a similar way into the nucleus (PLL vs. PEI < or = 3.5-fold), but PEI was approximately 10-100 times more efficient in gene expression than PLL (G1 < G2/M < S). The rate of pinocytosis increased up to 70-fold from G1 to middle S phase. Cell surface heparan and chondroitin sulfate increased 50-80%, and hyaluronan decreased 50% when the cells went from G1 through S to G2/M. CONCLUSIONS: The data obtained indicates that no single parameter (pinocytosis, cell surface GAGs, nuclear uptake) solely accounts for the differential pDNA uptake or expression during cell cycle, and that the main difference in PLL- and PEI-mediated transfections seems to be at the nuclear level.
Marjo Männistö; Mika Reinisalo; Marika Ruponen; Paavo Honkakoski; Markku Tammi; Arto Urtti
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  The journal of gene medicine     Volume:  9     ISSN:  1099-498X     ISO Abbreviation:  J Gene Med     Publication Date:  2007 Jun 
Date Detail:
Created Date:  2007-06-04     Completed Date:  2007-09-27     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9815764     Medline TA:  J Gene Med     Country:  England    
Other Details:
Languages:  eng     Pagination:  479-87     Citation Subset:  IM    
Department of Pharmaceutics, University of Kuopio, Kuopio, Finland.
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MeSH Terms
Cell Cycle / drug effects*
Cell Nucleus / drug effects,  metabolism
Cytoplasm / drug effects,  metabolism
DNA / metabolism
Glycosaminoglycans / metabolism*
Pinocytosis / drug effects
Plasmids / metabolism
Polyethyleneimine / pharmacology*
Polylysine / pharmacology*
Transfection / methods*
Reg. No./Substance:
0/Glycosaminoglycans; 25104-18-1/Polylysine; 3326-32-7/Fluorescein-5-isothiocyanate; 9002-98-6/Polyethyleneimine; 9007-49-2/DNA

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