Transfection of Myoblasts in Primary Culture with Isomeric Cationic Cholesterol Derivatives

Bischoff, Rainer; Cordier, Yves; Perraud, Frédéric; Thioudellet, Christine; Braun, Serge; Pavirani, Andréa

doi:10.1006/abio.1997.2401

Cited by 13 publications

(14 citation statements)

References 28 publications

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“…Microscope magnification: 100X. values are similar to those of cell lines such as A549 cells (Bischoff et al, 1997). No differences in gene transfer efficiency between healthy and dystrophic dog myoblasts were found (not shown).…”

Section: Resultssupporting

confidence: 66%

“…Most of the molecules are formulated together with the zwitterionic phospholipid dioleoyl-phosphatidylethanolamine (DOPE) to enhance transfection. Differences in efficiency among cationic lipids have been reported depending on the cell type and the ratio between lipid and DNA (Felgner et al, 1994, Bischoff et al, 1997. In a more detailed study (Bischoff et al, 1997), we found that slight structural differences between cationic lipids led to significantly different transfection efficiencies for myoblasts in primary cultures, with implications for cell or organ targeting.…”

Section: Resultssupporting

confidence: 51%

“…Synthesis of spermidine-cholesterol or DC-Cholesterol and preparation of DNA/lipid complexes (charge ratio 1/1) were performed as described elsewhere (Bischoff et al, 1997).…”

Section: Vectorsmentioning

confidence: 99%

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et al. 1999

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We have developed and characterized cultures of healthy and dystrophic canine myoblasts for the evaluation of various gene transfer protocols. The number of desmin-positive myoblasts was elevated (>>80%) in cultures of myoblasts obtained from different muscle territories, the diaphragm muscle giving rise to the purest cultures. Myoblasts from dogs turned out to be a very convenient source of well transfectable and transducible cells. Transfection with plasmid DNA allowed efficient transgene expression (50% of beta-galactosidase positive cells and about 375 ng luciferase/mg protein after transfection with a calcium phosphate-precipitated plasmid). Infection with high concentrations of adenoviral and retroviral vectors allowed transgene (beta-galactosidase or mini-dystrophin) detection in about 75 to 90% of the canine cells. Therefore, primary dog myoblast cultures represent a useful in vitro model for viral and non-viral gene delivery, as well as for functional evaluation and cell grafting with applications in genetic diseases, vaccination or production of circulating therapeutic proteins.

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Section: Resultssupporting

confidence: 66%

Section: Resultssupporting

confidence: 51%

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et al. 1999

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“…As the literature contains conflicting reports in this area ( , ), accurate, unambiguous spectroscopic 1 H and 13 C assignments of our lipo-polyamines are essential. Therefore, we investigated the NMR of spermine 1 , and compared our data to the literature values and also to those calculated using additivity rules ( , ).…”

Section: Resultsmentioning

confidence: 99%

“…Consequently C1, C2, and C3 are under the influence of a carbamate rather than a (protonated) primary amine, are relatively less deshielded, and therefore, come into resonance further upfield than their respective counterparts C12, C11, and C10 on the other propylene chain. During these studies, two other research groups have independently synthesized and published NMR assignments of this carbamate 13 (Table ) as its free base in CDCl 3 , but their assignments disagree with each other ( , ). Our assignments compare favorably with those of Bischoff et al (), with the differences in chemical shifts being attributable to a change in solvent and the protonation state of the polyamine.…”

Section: Resultsmentioning

confidence: 99%

Synthesis of Cholesteryl Polyamine Carbamates: pK_a Studies and Condensation of Calf Thymus DNA

et al. 2000

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Novel polyamine carbamates have been designed and prepared from cholesterol. Our synthesis uses an orthogonal protection strategy based upon trifluoroacetyl and Boc-protecting groups. These unsymmetrical polyamine carbamates have been prepared from symmetrical (e.g., spermine and thermine) polyamines. Detailed interpretations of (1)H and (13)C NMR spectroscopic data led to the unambiguous assignment of these polyamine carbamates. These target conjugates contain a variety of positive charges distributed along methylene chains. Their pK(a)s have been determined potentiometrically for conjugates substituted with up to five amino functional groups. Condensation of calf thymus DNA into particles was monitored using light scattering at 320 nm. Salt-dependent binding affinity for calf thymus DNA was determined using an ethidium bromide fluorescence quenching assay. These cholesteryl polyamine carbamates are models for lipoplex formation with respect to gene delivery (lipofection), a key first step in gene therapy.

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Serum Compatible Spermine‐Based Cationic Lipids with Nonidentical Hydrocarbon Tails Mediate High Transfection Efficiency

et al. 2022

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Cationic lipids are widely used as nonviral synthetic vectors for gene delivery as a safer alternative to viral vectors. In this work, a library of L-shaped spermine-based cationic lipids with identical and nonidentical hydrophobic chains having variable carbon lengths (from C10 to C18) was designed and synthesized. These lipids were characterized and the structure-activity relationships of these compounds were determined for DNA binding and transfection ability when formulated as cationic liposomes. The liposomes were then used successfully for the transfection of HEK293T, HeLa, PC3, H460, HepG2, SH-SY5Y and Calu'3 cell lines. The transfection efficiency of lipids with nonidentical hydrocarbon chains was greater than the identical analogue. These reagents exhibited superior efficiency to the commercial reagent, Lipofectamine3000, under both serum-free and 10-40 % serum conditions in HEK293T, HeLa and H460 cell lines. The lipids were not toxic to the tested cell line. The results suggest that L-shaped spermine-based cationic lipids with nonidentical hydrocarbon tails could serve as efficient and safe nonviral vector gene carriers in further in vivo studies.

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Transfection of Myoblasts in Primary Culture with Isomeric Cationic Cholesterol Derivatives

Cited by 13 publications

References 28 publications

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Untitled

Synthesis of Cholesteryl Polyamine Carbamates: pK_a Studies and Condensation of Calf Thymus DNA

Serum Compatible Spermine‐Based Cationic Lipids with Nonidentical Hydrocarbon Tails Mediate High Transfection Efficiency

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Transfection of Myoblasts in Primary Culture with Isomeric Cationic Cholesterol Derivatives

Cited by 13 publications

References 28 publications

Untitled

Untitled

Synthesis of Cholesteryl Polyamine Carbamates: pKa Studies and Condensation of Calf Thymus DNA

Serum Compatible Spermine‐Based Cationic Lipids with Nonidentical Hydrocarbon Tails Mediate High Transfection Efficiency

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Synthesis of Cholesteryl Polyamine Carbamates: pK_a Studies and Condensation of Calf Thymus DNA