Photopolymerized micelles of diacetylene amphiphile: physical characterization and cell delivery properties.

A series of polydiacetylene (PDA) - based micelles were prepared from diacetylenic surfactant bearing polyethylene glycol, by increasing UV-irradiation times. These polymeric lipid micelles were analyzed by physicochemical methods, electron microscopy and NMR analysis. Cellular delivery of fluorescent dye suggests that adjusting the polymerization state is vital to reach the full in vitro potential of PDA-based delivery systems.

A FRET-based probe with a chemically deactivatable quencher.

A new concept of a chemically deactivatable quencher is proposed for a FRET-based probe that turns-on its fluorescence by either an enzymatic cleavage or a chemical reagent (sodium dithionite). This concept allowed us to quantify the caspase-3 cleavage activity in solution and to reveal unreacted probes in cell experiments.

Antiangiogenic effects of spironolactone and other potassium-sparing diuretics in human umbilical vein endothelial cells and in fibrin gel chambers implanted in rats.

OBJECTIVE: Potassium-sparing diuretics have different effects on angiogenesis that may mediate some abilities to treat cardiovascular diseases. The aim of the current study was to compare the effects of spironolactone and an active metabolite, canrenone, or a derivative, eplerenone, and amiloride, a diuretic without affecting mineralocorticoid activity, on the proliferation of human umbilical vein endothelial cells (HUVEC) and on angiogenesis in fibrin gel chambers implanted in rats. MATERIALS AND METHODS: We measured the effects of spironolactone, canrenone, eplerenone, and amiloride on the proliferation of HUVEC in the presence or absence of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). We also examined the effects of these compounds on migration and capillary tube formation by HUVEC. Finally, the effects of the compounds on neovessel formation in vivo were investigated by implanting Wistar rats for 14 days with perforated Plexiglas chambers containing rat fibrin. RESULTS: Spironolactone and amiloride inhibited the proliferation of HUVEC, but canrenone and eplerenone had no effect. The inhibitory effect of spironolactone was not prevented by VEGF or bFGF. Aldosterone had no effect on spironolactone-induced inhibition of HUVEC proliferation. Spironolactone induced a dose-dependent reduction of both cell chemotaxis and capillary tube formation. In fibrin gel chambers, spironolactone and amiloride significantly reduced the numbers of both peripheral and central neovessels. Canrenone and eplerenone, in contrast, had no antiangiogenic effect. CONCLUSION: Spironolactone and amiloride significantly inhibited angiogenesis in vitro and in the fibrin gel chamber in vivo. Spironolactone antiangiogenic effects are unrelated to antimineralocorticoid activity.

Angiogenesis and arteriogenesis are increased in fibrin gel chambers implanted in prehypertensive spontaneously hypertensive rats.

OBJECTIVE: Microvascular rarefaction by an unbalanced angiogenesis could promote the onset of hypertension in spontaneously hypertensive rats and in hypertensive patients. We studied the angiogenic potency in the fibrin gel chamber model in prehypertensive spontaneously hypertensive rats and their controls, Wistar-Kyoto rats. METHODS: Four-week-old prehypertensive spontaneously hypertensive rats (n = 9) and Wistar-Kyoto rats (n = 9) were implanted with four fibrin gel chambers located in the dorsal subcutaneous space. After 14 days, vasculoconjunctive buds had invaded the fibrin gel through the 10 hole-perforated bottom slip of the chamber. The intact vascular buds were studied using optical microscopy, alpha-actin and von Willebrand factor stainings. Capillaries and arterialized vessels were counted in three peripheral and one central field in each bud. The immunodetection of vascular endothelial growth factor and fibroblast growth factor 2 was performed on the neovascular buds. RESULTS: In fibrin chambers implanted in spontaneously hypertensive rats, the number of peripheral vessels was significantly higher than in Wistar-Kyoto rats. There were significantly more arterialized vessels in spontaneously hypertensive rats compared with Wistar-Kyoto rats. The number of immunostained cells for fibroblast growth factor 2 was significantly greater in spontaneously hypertensive rats compared with Wistar-Kyoto rats. There was no significant difference in vascular endothelial growth factor staining between the two strains of rats. CONCLUSION: Angiogenesis and arteriogenesis are increased in fibrin chambers implanted in prehypertensive spontaneously hypertensive rats compared with Wistar-Kyoto rats. These results argue against microvascular rarefaction as a cause of hypertension using this model of angiogenesis.

Effective polyethylenimine-mediated gene transfer into human endothelial cells.

BACKGROUND: The major advantage in choosing non-viral vectors such as cationic polymers for in vitro and in vivo transfection is their higher biosafety than viral ones. Among the cationic polymers, polyethylenimines (PEIs) are promising molecules for gene delivery to a variety of cells. Efficient transfection of primary endothelial cells using PEIs could be regarded as an interesting strategy of treatment in some ischemic cardiovascular diseases. METHODS: Efficacies of a 22-kDa linear PEI (L-PEI) and its glucose-grafted derivative (L-PEI-Glc(4)) were compared for gene transfer into human umbilical vein endothelial cells (HUVEC) using the reporter gene luciferase. Cells were incubated for 2, 4 and 24 h with PEI/DNA complexes made in 150 mM sodium chloride (NaCl) or in 5% glucose solution. Luciferase activity was measured 24 h after the onset of transfection. The effects of low (2%) and high (30%) concentrations of serum on transfection efficacy were assessed as well. We then studied the intracellular fate of the PEI/DNA complexes labelled with the DNA intercalator YOYO-1 using flow cytometry analysis (FACS) and confocal microscopy. RESULTS: PEI/DNA complexes formed in NaCl led to a higher transfection efficacy than those made in glucose. The optimal formulation, depending on the incubation time and the presence of serum in the medium, was obtained using DNA complexed to L-PEI-Glc(4) and incubated for 4 h with the cells. This condition led to 50% fluorescent cells after GFP transfection. A high serum concentration diminished the L-PEI associated toxicity but decreased L-PEI-Glc(4) transfection efficiency. FACS analysis using both vectors showed that almost 90% of the cells had internalized the DNA complexes. Confocal microscopic observations showed a fast attachment of the complexes to the cell surface followed by inclusion into vesicles that migrated to the perinuclear region. CONCLUSIONS: In this work, we defined the optimal conditions for gene delivery in HUVEC. These conditions were obtained when using derivatives L-PEI and L-PEI-Glc(4) complexed with DNA in 150 mM NaCl and added to cells for 2 and 4 h, respectively. Cellular trafficking of the complexes suggested that cell entry was not a limiting factor for gene delivery using PEI. This study underlined the interest in PEIs as efficient vectors for gene transfer into human endothelial cells.

Effects of combinations of 7beta-hydroxycholesterol and anticancer drugs or ionizing radiation on the proliferation of cultured tumor cells.

Oxysterols compose a large class of natural substances endowed in a number of cases with marked antiproliferative and cytotoxic activities. The consequences of treatments combining 7beta-hydroxycholesterol (7beta-OHC) or XG-142 (a galactose-linked hydrosoluble derivative of 7beta-OHC) with drugs used in cancer chemotherapy or gamma radiation has been evaluated upon a variety of tumor cell lines: Hep-G2, U937, K562 cells, its adriamycin-resistant variant K562 Adr+ and RDM4. Proliferation was assessed by the Uptiblue assay and the [3H]Thymidine incorporation test. Results indicated that 7beta-OHC increased the sensitivity of tumor cells to adriamycin, VP-16, 5-FU and bleomycin to various degrees. 7beta-OHC was found to reinforce the susceptibility of K562 adriamycin-resistant cells to this drug. In RDM4 cells, an enhanced radiosensitivity by 7beta-OHC was also obtained, whereas XG-142 was less efficient in provoking such an effect.