Hypo-osmotic stress enhances the uptake of polyethylenimine/oligonucleotide complexes in A549 cells via Ca(2+) mobilization from intracellular stores.
To determine the mechanism of osmolarity involved in polyethylenimine (PEI)/oligonucleotide (ON) complex transfection in cells, we measured the fluorescence intensities of fluorescein isothiocyanate-labeled ONs complexed with PEI and the changes in cytosolic Ca(2+) concentration ([Ca(2+)](c)) in A549 cells, and we found that uptake of PEI/ON complexes was improved in the cells along with a rise of [Ca(2+)](c) in A549 cells challenged by 50% hypotonic medium. Further experiments showed that the enhanced uptake efficiency and the rise in [Ca(2+)](c) in A549 cells were almost completely abolished from cells loaded with the intracellular calcium chelator 1,2-bis(2-aminophenoxy)-N,N,N,N-tetraacetic acid-acetoxymethyl ester. 2-Aminoethoxydiphenyl borate or 8-(N,N-diethylamino) octyl-3,4,5-trimethoxybenzoate, two potent antagonists of inositol 1,4,5-trisphosphate-mediated Ca(2+) release that blunt [Ca(2+)](c) elevation via Ca(2+) release from endoplasmic reticulum, inhibited the enhanced uptake of PEI/ON complexes induced by Ca(2+)-free hypo-osmotic stress. In summary, the results strongly suggest that calcium-dependent transfection is responsible for the uptake of PEI/ON complexes into A549 cells under hypotonic conditions.
Sun
Wenwu
W
Department of Respiratory Medicine, The Northern Hospital of ShenYang, Shenyang, China.
Pan
Leiting
L
Ma
Zhuang
Z
D002118:Calcium / Q000378:metabolism*; D045744:Cell Line, Tumor; D002614:Chelating Agents / Q000737:chemistry; D006801:Humans; D009841:Oligonucleotides / Q000378:metabolism*; D009997:Osmotic Pressure; D011094:Polyethyleneimine / Q000378:metabolism*
D002614:Chelating Agents; D009841:Oligonucleotides; D011094:Polyethyleneimine; D002118:Calcium