Histone H1-mediated transfection: serum inhibition can be overcome by Ca2+ ions.

Abstract:

PURPOSE:One of the drawbacks of polycationic and cationic liposomal gene transfer is its sensitivity to serum. Gene therapy requires the transfectant-DNA complex to be resistant to serum as well as blood. Since Ca2+ has proved to be an efficient cofactor of polycationic gene transfer, we decided to investigate its effects on transfection in the presence of serum. METHODS:We studied transgene expression of luciferase gene (pCMV Luc) on ECV 304 human endothelial cells using H1 histone and DOSPER as transfectants in the presence of 0-100% fetal calf serum. RESULTS:H1-and DOSPER-mediated transfection was found to be inhibited by serum above the concentration of 10%. If 2 mM Ca2+ or 2 mM Ca2+/0.1 mM chloroquine was included in the culture medium which replace the transfection mixture and was left on the cells for 24 hours postincubation, the inhibiting effect of even 100% serum was overcome. CONCLUSIONS:A high serum level does not interfere with binding and uptake of H1- and DOSPER-DNA complexes, but inhibits subsequent steps such as endosomal escape. Ca2+ in the form of nascent calcium phosphate microprecipitates and other lysosomolytical agents facilitate endosomal/lysosomal release by their fusigenic and membranolytic activity.

journal_name

Pharm Res

journal_title

Pharmaceutical research

authors

Haberland A,Knaus T,Zaitsev SV,Buchberger B,Lun A,Haller H,Böttger M

doi

10.1023/a:1007581700996

subject

Has Abstract

pub_date

2000-02-01 00:00:00

pages

229-35

issue

2

eissn

0724-8741

issn

1573-904X

journal_volume

17

pub_type

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