Generation of reactive oxygen species accounts for cytotoxicity of an endogenous dopaminergic neurotoxin, (R)-N-methylsalsolinol, to differentiated dopaminergic SH-SY5Y cells

The mechanism of the cytotoxicity of endogenous dopamine-derived (R)-1,2-dimethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline [(R)-N-methylsalsolinol] to differentiated human dopaminergic neuroblastoma SH-SY5Y cells was studied using a reduction-oxidation indicator, Alamar Blue. N-Methylsalsolinol and its oxidation product, 1,2-dimethyl-6,7-dihydroxyisoquinolinium ion, were found to inhibit oxidative phosphorylation, as shown by the Redox capacity. Antioxidants, such as reduced glutathione, catalase, Tris and n-propyl gallate, reduced the cytotoxicity of N-methylsalsolinol, suggesting that hydroxyl radical was the major reactive oxygen species for the cytotoxicity. Deprenyl also protected the cells from the decrease of the Redox capavity by N-methylsalsolinol. However, antioxidants did not protect the cells from the cytotoxicity of the catechol isoquinolinium ion. The results suggest that oxidative stress induced by hydroxyl radical may be involved in the cell death of dopaminergic neurons by N-methylsalsolinol.     

Effect of docosahexaenoic acid on smooth muscle cell functions

There is increasing evidence that fish oil-enriched diets attenuate the progression of several types of human and experimental renal, intestinal and cardiovascular disorders, including hypertension. Docosahexaenoic acid (DHA), may be one of the active biological component. We previously reported that dietary DHA suppressed the progression of hypertension in stroke-prone spontaneously hypertensive rats (SHRSP). The purpose of this study is to clarify the in vitro effect of DHA on cultured smooth muscle cell functions such as cell growth, hypertrophy, NO release, and intracellular Ca2+ metabolism, which are involved in the regulatory mechanisms of vascular tone. Addition of DHA to the culture medium of aortic smooth muscle cells isolated from SHRSP and normotensive Wistar Kyoto rats (WKY) had no significant effects on cell growth or on cell hypertrophy induced by angiotensin II as measured by flow cytometry. DHA had no stimulatory effect on interleukin-1beta (10 ng/ml)-induced nitric oxide release from smooth muscle cells of SHRSP, but rather slightly inhibited it. However, the treatment of smooth muscle cells with DHA (30 microM) for 2 days significantly suppressed the increase in intracellular Ca2+ concentration induced by angiotensin II, but not by thapsigargin. This was due to the suppression of Ca2+ influx, as determined by Mn2+ influx experiment. These results indicate that DHA specifically suppresses Ca2+ mobilization into smooth muscle cells. This may be one of the mechanisms by which dietary DHA prevents the development of hypertension in SHRSP.     

Potent and selective thrombin inhibitors featuring hydrophobic, basic P3-P4-aminoalkyllactam moieties

Crystal structure and evolving SAR considerations of potent, selective benzylsulfonamide lactam thrombin inhibitors and related serine protease inhibitors have led to the design of novel thrombin inhibitors 1a-g, featuring hydrophobic, basic, P4-alkylaminolactam scaffolds that serve as novel types of P3-P4 dipeptide mimics. The design, synthesis, and biological activity of these targets is presented.     

Effect of 5-hydroxytryptamine on intracellular calcium dynamics in cultured rat vascular smooth muscle cells

The present study elucidated the precise mechanism of 5-hydroxytryptamine (5-HT)-induced increase of intracellular Ca2+ concentration ([Ca2+]i) in cultured vascular smooth muscle cells isolated from rat aortic media. [Ca2+]i was measured using fluorescent Ca2+ indicator, fura-2. 5-HT caused a dose-dependent increase in [Ca2+]i, which was completely inhibited by ketanserin. alpha-Methyl-5-HT had an equipotent effect to 5-HT. Diltiazem at 10 microM partially suppressed the 5-HT-induced increase in [Ca2+]i. 5-HT also augmented Mn2+ influx, when monitored by Mn2+ quenching of fura-2 fluorescence. When extracellular Ca2+ (1.3 mM) was removed, a decrease in resting level and a small, transient increase in [Ca2+]i were observed. 5-HT stimulation also induced an increase in the production of inositol triphosphate. 5-HT-induced increase in [Ca2+]i was significantly, but partially inhibited by staurosporin and H-7. Phorbol 12-myristate 13-acetate induced an increase in [Ca2+]i, which was abolished by removal of extracellular Ca2+. 5-HT-induced increase in [Ca2+]i was not affected by the pretreatment with pertussis toxin (PTX), and was not accompanied by a change in cyclic AMP content. These results suggest that, in cultured rat aortic smooth muscle cells, 5-HT increases [Ca2+]i via 5-HT2 receptor subtype by inducing influx of extracellular Ca2+ partially through L-type voltage-dependent Ca2+ channel, as well as by mobilizing Ca2+ from its intracellular stores. Activation of protein kinase C may be positively involved in the regulatory mechanism of Ca2+ influx, but PTX-sensitive G protein and cyclic AMP seem to be not involved.     

Stability of nano-thick transparent conducting oxide films for use in a moist environment

The stability of nano-thick transparent conducting oxide thin films in a high humidity environment was investigated. The stability of ITO and impurity-doped ZnO thin films prepared with a thickness in the range from approximately 20 to 100 nm on glass substrates at a temperature below 200 °C by a pulsed laser deposition was evaluated in air at a relative humidity of 90% and a temperature of 60 °C. The resistivity of all Al- and Ga-doped ZnO thin films tested was found to increase markedly with test time, whereas that of ITO remained relatively stable; the stability (resistivity increase) of the doped ZnO thin films was considerably affected by film thickness but was relatively independent of the deposition substrate temperature. In particular, doped ZnO thin films with a thickness below approximately 50 nm were very unstable under the test conditions. The resistivity increase of doped ZnO films is mainly attributed to the grain boundary scattering resulting from the adsorption of oxygen on the grain boundary.     

Down-regulation of the noradrenaline transporter by interferon-alpha in cultured bovine adrenal medullary cells

The aim of this study was to investigate the effect of long-term treatment with interferon (IFN)-alpha on the noradrenaline transporter of bovine adrenal medullary cells. Treatment of cultured adrenal medullary cells with IFN-alpha caused a decrease in uptake of [3H]noradrenaline by the cells in time (4-48 h)- and concentration (300-1,000 U/ml)-dependent manners. IFN-beta also inhibited [3H]noradrenaline uptake to a lesser extent than did IFN-alpha, whereas IFN-gamma had little effect. An anti-IFN-alpha antibody reduced the effect of IFN-alpha on [3H]noradrenaline uptake. Saturation analysis of [3H]noradrenaline uptake showed that the inhibitory effect of IFN-alpha was due to a reduction in the maximal uptake velocity (Vmax) values without altering apparent Michaelis constant (Km) values. Incubation of cells with IFN-alpha caused a translocation of protein kinase C from the soluble to the particulate fraction in the cells. The effect of IFN-alpha on [3H]noradrenaline uptake was diminished in protein kinase C-down-regulated cells. Incubation of cells with IFN-alpha for 48 h significantly reduced the specific binding of [3H]desipramine to crude plasma membranes isolated from cells. Scatchard analysis of [3H]desipramine binding revealed that IFN-alpha decreased the maximal binding (Bmax) values without any change in the dissociation constant (K(D)) values. These findings suggest that IFN-alpha suppresses the function of noradrenaline transporter by reducing the density of the transporter in cell membranes through, at least in part, a protein kinase C pathway.     

Heparin inhibits human coronary artery smooth muscle cell migration

Heparin, an anticoagulant, has been shown to reduce neointimal proliferation and restenosis following vascular injury in experimental studies, but the clinical trials of heparin in coronary balloon angioplasty have been negative. The current study, therefore, examined the effect of heparin on basal or stimulated migration by serum and platelet-derived growth factor (PDGF)-BB in cultured human coronary artery smooth muscle cells (SMCs) by Boyden's chamber method. In addition, the reversibility of the heparin effect on human coronary artery SMC migration was examined. Fetal calf serum (FCS) and PDGF-BB stimulated SMC migration in a concentration-dependent manner. Heparin in moderate to high concentration (10 to 100 U/mL) exhibited concentration-related inhibition of FCS- and PDGF-BB-stimulated SMC migration; however, a low concentration (1 U/mL) of heparin had no inhibitory effects. Heparin also had weak inhibitory effects on nonstimulated SMC migration. The SMCs that were exposed to a high concentration (100 U/mL) of heparin for 6 hours were capable of migrating after a short lag period of removal of heparin from the culture medium. These SMCs also showed recovery of responses to FCS and PDGF-BB by migrating significantly greater than the nonstimulated level. Furthermore, heparin-containing medium did not contain detached cells. These results indicate that heparin inhibits human coronary artery SMC migration, especially when stimulated by FCS or PDGF-BB, and that this inhibitory effect of heparin is reversible and not simply a function of killing cells.     

Highly conductive and transparent ZnO:Al thin films prepared on high-temperature substrates by d.c. magnetron sputtering

Highly conductive and transparent aluminium-doped ZnO (AZO) thin films have been prepared on high-temperature substrates using d.c. magnetron sputtering. In AZO films, the spatial distribution of resistivity across substrates placed parallel to the target was improved by deposition at substrate temperatures above 300 °C. AZO films with resistivities of 2−5 x 10⁻⁴ Ω cm were prepared under sputter gas pressures of between 0.6 and 3.0 Pa and at a substrate temperature of 350 °C. In addition, milky AZO films with a textured surface were prepared on high-temperature substrates under sputtering conditions which suppressed the c-axis orientation. A total transmittance of 72% and a haze factor of 63% at a wavelength of 500 nm and a sheet resistance as low as 2.0 Ω sq⁻¹ were obtained in milky AZO films 3 μm thick prepared at a sputter gas pressure of 12 Pa and a substrate temperature of 350 °C.     

Preparation of B2O3-P2O5-SiO2 coating films by the sol-gel method

ThexB₂O₃ · (20-x) P₂O₅ · 80SiO₂ (in mol%) glass films withx=0, 10 and 20 have been prepared from metal alkoxides by carrying out the coating in a dry atmosphere. These coating films have shown a larger value of load at scratch and a smaller shrinkage during heat-treatment by replacing P₂O₅ in the films with B₂O₃. It has been found that B₂O₃ more effectively reduces the glass transition temperature of SiO₂ glass than P₂O₅. The concentrations of sodium ions, which migrated from soda-lime-silica glass substrates during the film formation, were higher in phosphosilicate and borophosphosilicate films than in borosilicate and pure silica films. This finding should be ascribed to the gettering effects of phosphorus for sodium ions.