Lowering of brain endothelial cell barrier function by exposure to 4'-iodo-α-pyrrolidinononanophenone.

Overuse of pyrrolidinophenones (PPs) is known to cause damage to vascular and central nervous systems, but little is known about its effect on brain endothelial barrier function. In this study, we found that exposure to 4'-iodo-α-pyrrolidinononanophenone (I-α-PNP), one of the most potently cytotoxic PPs, at sublethal concentrations decreases trans-endothelial electrical resistance and increases paracellular permeability across a monolayer of human brain microvascular endothelial cells. Treatment with I-α-PNP also elevated the production of superoxide anion. Furthermore, the treatment reduced the expression and plasma membrane localization of a tight junction protein claudin-5 (CLDN5), which was almost restored by pretreatment with an antioxidant N-acetyl-l-cysteine. These results indicate that I-α-PNP treatment may down-regulate the plasma membrane-localized CLDN5 by elevating the production of reactive oxygen species (ROS). The treatment with I-α-PNP increased the nuclear translocation of Forkhead box protein O1 (FoxO1), an oxidative stress-responsive transcription factor, and pretreating with a FoxO1 inhibitor ameliorated the decrease in CLDN5 mRNA. In addition, I-α-PNP treatment up-regulated the expression and secretion of matrix metalloproteinase-2 (MMP2) and MMP9, and the addition of an MMP inhibitor reversed the degradation of CLDN5 by I-α-PNP. Moreover, I-α-PNP treatment facilitated the activation of 26S proteasome-based proteolytic activity and pretreatment with an inhibitor of 26S proteasome, but not autophagy, suppressed the CLDN5 degradation by I-α-PNP. Accordingly, it is suggested that the down-regulation of CLDN5 by exposure to I-α-PNP is ascribable to suppression of the gene transcription due to FoxO1 nuclear translocation through ROS production and to acceleration both of the MMPs (MMP2 and MMP9)- and 26S proteasome-based proteolysis.

Poly-phenylene jacketed tailor-made dendritic phenylazomethine ligand for nanoparticle synthesis.

Synthesizing metal clusters with a specific number of atoms on a preparative scale for studying advanced properties is still a challenge. The dendrimer templated method is powerful for synthesizing size or atomicity controlled nanoparticles. However, not all atomicity is accessible with conventional dendrimers. A new tailor-made phenylazomethine dendrimer (DPA) with a limited number of coordination sites (n = 16) and a non-coordinating large poly-phenylene shell was designed to tackle this problem. The asymmetric dendron and adamantane core four substituted dendrimer (PPDPA16) were successfully synthesized. The coordination behavior confirmed the accumulation of 16 metal Lewis acids (RhCl3, RuCl3, and SnBr2) to PPDPA16. After the reduction of the complex, low valent metal nanoparticles with controlled size were obtained. The tailor-made dendrimer is a promising approach to synthesize a variety of metal clusters with desired atomicity.

Apoptotic mechanism in human brain microvascular endothelial cells triggered by 4'-iodo-α-pyrrolidinononanophenone: Contribution of decrease in antioxidant properties.

In this study, we newly synthesized four α-pyrrolidinononanophenone (α-PNP) derivatives [4'-halogenated derivatives and α-pyrrolidinodecanophenone (α-PDP)], and then performed the structure-cytotoxicity relationship analyses. The results showed the rank order for the cytotoxic effects, α-PNP < α-PDP < 4'-fluoro-α-PNP < 4'-chrolo-α-PNP < 4'-bromo-α-PNP < 4'-iodo-α-PNP (I-α-PNP), and suggest that cytotoxicities of 4'-halogenated derivatives were more intensive than that of elongation of the hydrocarbon chain (α-PDP). We also surveyed the apoptotic mechanism of I-α-PNP in brain microvascular endothelial (HBME) cells that are utilized as the in vitro model of the blood-brain barrier. HBME cell treatment with I-α-PNP facilitated the apoptotic events (caspase-3 activation, externalization of phosphatidylserine, and DNA fragmentation), which were almost completely abolished by pretreating with antioxidants. In addition, the immunofluorescent staining revealed the enhanced production of hydroxyl radical in mitochondria by the I-α-PNP treatment, inferring that the I-α-PNP treatment triggers the apoptotic mechanism dependent on the enhanced ROS production in mitochondria. The treatment with I-α-PNP increased the production of cytotoxic aldehyde 4-hydroxy-2-nonenal and decreased the amount of reduced glutathione. Additionally, the treatment decreased the 26S proteasome-based proteolytic activities and aggresome formation. These results suggest that decrease in the antioxidant properties is also ascribable to HBME cell apoptosis elicited by I-α-PNP.

9,10-Phenanthrenequinone provokes dysfunction of brain endothelial barrier through down-regulating expression of claudin-5.

Chronic exposure to diesel exhaust particle (DEP) is considered to provoke dysfunction of the blood-brain barrier, but the detailed molecular mechanism remains unclear. In this study, we investigated the toxic effects of five DEP components against human vascular cells and found that, among them, 9,10-phenanthrenequinone (9,10-PQ), a major tricyclic quinone in DEP, most potently elicits the cellular toxicities. Additionally, treatment with 9,10-PQ at its cytolethal concentrations (more than 2 µM) facilitated the production of reactive oxygen species (ROS), caspase activation, and DNA fragmentation in human brain microvascular endothelial (HBME) cells, inferring that high concentrations of 9,10-PQ elicit the cell apoptosis through the ROS-dependent mechanism. Measurement of trans-endothelial electrical resistance and paracellular permeability showed that treatment with sublethal concentrations (less than 1 µM) of 9,10-PQ elevates permeability across HBME cell monolayer. Immunofluorescence observation and Western blotting analysis also revealed that the 9,10-PQ treatment remarkably down-regulated the intercellular localization and expression of claudin-5 (CLDN5), a tight junctional protein that plays a key role in function of the blood-brain barrier, and the down-regulation was markedly recovered by pretreatment with a proteasome inhibitor Z-Leu-Leu-Leu-CHO. This result may indicate that sublethal concentrations of 9,10-PQ facilitate the dysfunction of the endothelial cell barrier through lowering in the expression and proteasomal proteolysis of CLDN5. The treatment with 9,10-PQ promoted nitric oxide (NO) production presumably through the induction of inducible NO synthase. In addition, the 9,10-PQ-mediated down-regulation of CLDN5 was ameliorated and deteriorated by pretreating with a scavenger and donor, respectively, of NO. Similarly to the 9,10-PQ treatment, treatment with a donor of peroxynitrite, a highly reactive oxidant formed by the reaction of NO and superoxide anion, resulted in the marked reduction of CLDN5 expression and elevation of 26S proteasome-based proteolytic activities. Thus, it is suggested that the formation of NO and peroxynitrite participates in the mechanism of brain endothelial cell barrier dysfunction elicited by 9,10-PQ.

Photo-patternable electroluminescence based on one-way photoisomerization reaction of tetraoxidized triangle terarylenes.

Oxidized triangle terarylenes with asymmetric side-aryl units are synthesized, which show photo-induced turn-on yellow luminescence based on photochemical intramolecular ring-cyclization reaction in the solid state. A direct photo-patterning process for organic electroluminescent devices is successfully demonstrated with selective light irradiation through an appropriate mask-pattern.

Efficient oxidative cycloreversion reaction of photochromic dithiazolythiazole.

Electrochemical response of photochromic tearylenes was surveyed by means of cyclic voltammetry, DFT calculations, and spectroelectrochemistry. 4,5-Bis(2-phenyl-5-methylthiazolyl)-2-phenylthiazole was found to show electrochemical oxidative ring-cycloreversion reaction. The net current efficiency of the cycloreversion reaction under constant potential electrolysis was as high as 900%, which is ascribed to an electrochemical local-cell mechanism and a chain reaction mechanism. Electron transfer stopped-flow study using a chemical oxidant successfully identified radical cation intermediates of both closed- and open-ring isomers, involved in the oxidative cycloreversion process. The significantly long-lived radical cation of open-ring isomer with the lifetime of 33 s takes part in the indirect electron transfer process from the neutral closed-ring isomer to the radical cation of open-ring isomer in the chain reaction manner.