Interactions of new synthesized fluorescent cationic amphiphiles bearing pyrene hydrophobe with plasmid DNA: binding affinities, aggregation and intracellular uptake.
In this work, we prepared a novel series of cationic amphiphiles denoted as the Py-cations (Py-Gly, Py-Ala, Py-Cap, Py-G(1)-Lys and Py-G(2)-Lys) bearing fluorescent pyrene and various hydrocarbon linkers between the pyrene hydrophobe and cationic block. Employing these new cationic amphiphiles with pyrene as the fluorescent probe, the interactions between these Py-cations and plasmid DNA (pDNA) in distilled water and 0.1 M PBS buffer solution have been explored by means of UV-vis and fluorescent spectrometers, and ethidium bromide dye displacement and agarose-gel retardant assays were also implemented to evaluate their pDNA binding affinities in aqueous solution. Furthermore, the average sizes and morphologies of self-assembled Py-cation/pDNA lipoplex aggregates were examined by dynamic laser light scattering (DLS) and atomic force microscopy (AFM). It was found that these fluorescent cationic amphiphiles showed blue fluorescence emission of pyrene probe at λ = 340 nm in distilled water while their interactions with pDNA led to new strong green emission at λ = 490 nm, and this may be due to the stacking of pyrene and new formation of excimers via the rigid pDNA templated self-assembly. It was also revealed that the binding between new Py-cations and pDNA in aqueous solution was strongly influenced by the Py-cation hydrophobicity, charges of the cation and the presence of electrolytes. With respect to the Py-cation/pDNA aggregate morphologies, very interesting 1-D hybrid nanofibers were predominantly observed by AFM for the Py-Cap/pDNA aggregates. In addition, utilizing a COS-7 cell-line, in-vitro cellular uptakes of new cationic amphiphiles with pyrene probe were studied and visualized by fluorescent microscopy. As a result, this may provide a new approach to investigate the interactions between synthetic cationic lipids and nucleic acids, and pave an alternative clue to design new organic gene delivery carriers.
Sheng
Ruilong
R
Laboratory of Polymer Materials, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, 345 Lingling Road, Shanghai 200032, China.
Luo
Ting
T
Zhu
Yingdan
Y
Li
Hui
H
Cao
Amin
A
D002412:Cations; D004247:DNA / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D010957:Plasmids; D011721:Pyrenes / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet
D002412:Cations; D005456:Fluorescent Dyes; D011721:Pyrenes; D004247:DNA; C030984:pyrene