Highly efficient liposome-mediated gene transfer of inducible nitric oxide synthase in vivo and in vitro in vascular smooth muscle cells

Veit K, Boissel J-P, Buerke M, Grosser T, Meyer J, Darius H. Highly efficient liposome-mediated gene transfer of inducible nitric oxide synthase in vivo and in vitro in vascular smooth muscle cells. Cardiovascular Research. 1999;43(3):808-822.Objective: The efficient introduction of regulatory genes into vascular smooth muscle cells (SMCs) is one of the most promising options for gene therapy of cardiovascular diseases. Cationic liposome-mediated gene transfer may become a favorable transfection technique with regard to patient’s safety for in vivo administration. However, this method until now has its limitation in a low transfection efficiency. Therefore, the present study was designed to improve cationic liposome-mediated transfection of rabbit vascular SMCs in vitro and in vivo, in order to enhance transfection efficiency and present an optimized system which may offer a potential therapeutic benefit for in vivo application. Methods and results: Optimized lipofection of rabbit SMCs with the mammalian expression vector pE-N1 and the reporter gene green fluorescent protein resulted in a mean transfection efficiency of about 50%. The unique transfection of rabbit SMCs in vitro and in vivo with the inducible isoform of human nitric oxide synthase (NOSII), using the same vector, resulted in a successful transient transcription and translation of a functionally active human NOSII in rabbit SMC, persisting 5–6 days. We could further demonstrate that the transfection procedure and the transgene product did neither induce necrosis nor apoptosis under the conditions chosen and did not result in the induction of endogenous NOSII of transfected SMCs. Conclusion(s): These findings indicate potential therapeutic relevance for this nonviral gene transfer system for in vivo gene therapy for cardiovascular diseases

Metallisation des plastiques haute temperature

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Involvement of PKC and NF-κB in Nitric Oxide Induced Apoptosis in Human Coronary Artery Smooth Muscle Cells

Ibe W, Bartels W, Lindemann S, et al. Involvement of PKC and NF-κB in Nitric Oxide Induced Apoptosis in Human Coronary Artery Smooth Muscle Cells. Cellular Physiology and Biochemistry. 2001;11(5):231-240.Apoptosis of vascular smooth muscle cells is critically involved in progression of atherosclerosis and may prevent intimal hyperplasia in restenosis and vascular remodeling. Nitric oxide (NO) is known to induce apoptosis, but the signaling pathways still remain unclear. We investigated p53 accumulation, protein kinase C (PKC) activation and nuclear transcription factor (NF-ĸB) binding activity as possible signaling mechanisms of NO-induced apoptosis. Apoptosis was induced dose-dependently with the NO-donors sodiumnitroprusside (SNP: 232±48%) and SIN-1 (241±90% of actinomycin D induced apoptosis; means ± SEM, * p£0.05 vs. control) in HSMC. Inhibition of PKC significantly attenuated NO-induced apoptosis. Staurosporine reduced SIN-1/SNP-mediated DNA fragmentation by 55.3±13.8% and 38.3±13.9% respectively. Comparable results were obtained for calphostin C. However, NO-mediated induction of apoptosis was not preceded by p53 accumulation. SNP decreased NF-ĸB binding activity in HSMC. These results suggest that induction of apoptosis by exogenous NO in HSMC is not dependent on p53 accumulation but involves protein kinase C signaling and regulation of NF-ĸB binding activity. This opens a new therapeutical approach in preventing restenosis after angioplasty

Inhibition of folic acid uptake by catechins and tea extracts in Caco-2 cells

In this present study it was aimed to determine whether the catechins contained in green tea and the whole extracts of Camellia sinensis (Theaceae) inhibit the uptake of folic acid by Caco-2 cell monolayers. Our results indicate that (-)-epigallocatechin 3-gallate (EGCG) and (-)-epicatechin 3-gallate (ECG) inhibit cellular folic acid uptake with IC50 values of 34.8 micromol/L and 30.8 micromol/L, respectively. Furthermore, green and black tea extracts were also found to inhibit folic acid uptake with IC50 values of approximately 7.5 and 3.6 mg/mL, respectively. According to these results, simultaneous intake of tea and folic acid may inhibit intestinal folic acid absorption. The consequences with respect to the folate status of the body will need to be examined in vivo

Introduction to MIOSOTYS: a multiple-object, high-speed photometer

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