Li Wang, David A. Dean and Robert C. MacDonald Pages 93 - 96 ( 4 )
As previously shown, vinblastine, when incorporated into a cationic lipid prior to generation of lipoplexes, increases by ∼30-fold the extent of transfection of pβ-Gal with a cytomegalovirus promoter (pCMV-β- Gal) to vascular smooth muscle cells (VSMC) by 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (EDOPC)-pCMV- β-Gal complexes. To test if this increase is limited to VSMC and EDOPC, or is general, we examined three other cell types, human umbilical artery endothelial cells (HUAEC), baby hamster kidney (BHK) cells and 293 cells derived from human kidney, as well as a different cationic lipid, 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP). In addition, to determine the contribution of the NF-kB transcription factor to the vinblastine effect, pCMV was replaced with a smooth muscle g-actin gene promoter, SMGA, which, unlike pCMV, does not respond to NF-kB. It was found that on all cell types we tested, the transfection efficiency increased with vinblastine incorporation; however, the magnitude depended greatly on the cell type, e.g. whereas the transfection of VSMC increased ∼30- fold, that of 293 cells increased only ∼2-fold. The cationic phospholipid could be replaced with DOTAP with no loss of effect. In contrast, the promoter was critical and the stimulation was lost if pCMV was replaced with pSMGA. It is concluded that the positive effect of vinblastine on transfection is general and the stimulation of the transcription factor NF-kB is involved in this action. The activation of NF-kB by anti-microtubule agents should thus allow for transfection of specific cell types by vinblastine lipoplexes.
vinblastine, transfection, smooth muscle actin gene promoter, intracellular transport
Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanston, IL 60208, USA.