Electrolytic hydrogen-generating bottle supplies drinking water with free/combined chlorine and ozone repressed within safety standard under hydrogen-rich conditions.

Hydrogen molecules have attracted attention as a new antioxidant, but are left to be confirmedly verified whether the oral administration is highly safe or not, concurrently with retention of abundant hydrogen. When electrolysis was performed for 10 minutes using a direct-current electrolytic hydrogen-water generating bottle with tap water, "residual free chlorine" concurrently upon the production of molecular hydrogen (444 µg/L) could be appreciably decreased from 0.18 mg/L to 0.12 mg/L as quantified by a N,N-diethyl-p-phenylenediamine-dye colorimetric method. Moreover, the total chlorine concentration (residual bound chlorine plus free chlorine) was estimated to be decreased from 0.17 mg/L to 0.11 mg/L. Although a merit of electrolytic hydrogen-generating bottles exists in electrolysis for periods as short as 10 minutes, the 30-minute electrolysis brought about the more abundant hydrogen (479 µg/L) together with an oxidation-reduction potential of -245 mV; even upon this long-term electrolysis, the gross amounts of chlorine, hypochlorous acid and chloramine were shown not to be increased (0.09-0.10 mg/L from 0.11 mg/L for tap water) as detected by orthotolidine colorimetry. Above-mentioned levels of diverse-type chlorines might fulfill the World Health Organization guideline for drinking water below 5 mg/L. In addition, the dissolved ozone upon electrolytic generation of hydrogen-water was below the detection limit (< 0.05 mg/L) or undetectable, which fulfilled the official safety standards in Japan and the USA for drinking water below 0.1 mg/L, as evaluated by three methods such as an electrode-type ozone checker, indigo dye-utilizing ozone detector capillaries and potassium iodide-based colorimetry. Importantly, even when half the amount of tap water was poured into the tank of the apparatus and electrolyzed, both the residual chlorine and ozone concentrations measured were also below the safety standard. Thus, major potently harmful substances, such as residual free/bound chlorine, or hypochlorous-acid/chloramine, respectively, and dissolved ozone, as the drinking hydrogen-water was direct-current-electrolytically generated, were estimated to be repressed within safety concentration ranges with achievements of abundant hydrogen generation.

Activation of PPARbeta/delta induces endothelial cell proliferation and angiogenesis.

OBJECTIVE: The role of the nuclear receptor peroxisome-proliferator activated receptor (PPAR)-beta/delta in endothelial cells remains unclear. Interestingly, the selective PPARbeta/delta ligand GW501516 is in phase II clinical trials for dyslipidemia. Here, using GW501516, we have assessed the involvement of PPARbeta/delta in endothelial cell proliferation and angiogenesis. METHODS AND RESULTS: Western blot analysis indicated PPARbeta/delta was expressed in primary human umbilical and aortic endothelial cells, and in the endothelial cell line, EAHy926. Treatment with GW501516 increased human endothelial cell proliferation and morphogenesis in cultures in vitro, endothelial cell outgrowth from murine aortic vessels in vitro, and angiogenesis in a murine matrigel plug assay in vivo. GW501516 induced vascular endothelial cell growth factor mRNA and peptide release, as well as adipose differentiation-related protein (ADRP), a PPARbeta/delta target gene. GW501516-induced proliferation, morphogenesis, vascular endothelial growth factor (VEGF), and ADRP were absent in endothelial cells transfected with dominant-negative PPARbeta/delta. Furthermore, treatment of cells with cyclo-VEGFI, a VEGF receptor1/2 antagonist, abolished GW501516-induced endothelial cell proliferation and tube formation. CONCLUSIONS: PPARbeta/delta is a novel regulator of endothelial cell proliferation and angiogenesis through VEGF. The use of GW501516 to treat dyslipidemia may need to be carefully monitored in patients susceptible to angiogenic disorders.

Attenuation of interferon-gamma mRNA expression in activated Jurkat T cells by exogenous zinc via down-regulation of the calcium-independent PKC-AP-1 signaling pathway.

Zinc is known to modulate a wide variety of cellular functions including anti-inflammatory responses. We examined the intracellular signaling pathways that contribute to the regulation of interferon-gamma (IFN-gamma) by zinc in activated human Jurkat T cells. Zinc significantly reduced IFN-gamma expression and activator protein-1 (AP-1) signaling in cells activated by phorbol 12-myristate 13-acetate (PMA) and phytohemagglutinin (PHA) without affecting cell viability. Moreover, partial inhibition of AP-1 activity by SP600125, a c-Jun N-terminal kinase (JNK) inhibitor, resulted in marked reduction of IFN-gamma transcription. We also found that this inhibitory effect of zinc on AP-1 signaling was abolished by treatment with rottlerin, a selective inhibitor of calcium-independent protein kinase C (PKC). These results suggest a novel target of zinc in the calcium-independent protein kinase C-AP-1 pathway to regulate endogenous IFN-gamma gene expression in activated T cells.

Prostacyclin-deficient mice develop ischemic renal disorders, including nephrosclerosis and renal infarction.

BACKGROUND: Prostacyclin (PGI2) is a short-lived endogenous inhibitor of platelet aggregation and a potent vasodilator and regulator of the growth of vascular smooth muscle cells. To study the role of PGI2 in the vascular system in vivo, PGI2-deficient (PGID) mice were established by genetic disruption of the PGI2 synthase gene. METHODS AND RESULTS: PGI2 synthase-null mice were generated by replacing the exons of PGI2 synthase gene that encodes for the catalytic site of the enzyme with a neomycin resistance gene. In these mice, PGI2 levels in the plasma, kidneys, and lungs were reduced, whereas thromboxane and prostaglandin E2 levels became elevated. Blood pressure and the amounts of urea nitrogen and creatinine in plasma of the PGID mice were significantly higher than those of wild-type mice (P<0.05). They developed progressive morphological abnormalities in the kidneys, accompanied by atrophy, surface irregularity, fibrosis, cyst, arterial sclerosis, and hypertrophy of vessel walls. Thickening of the thoracic aortic media and adventitia were observed in aged PGID mice. Importantly, these phenotypes have not been reported in PGI2 receptor-deficient mice. CONCLUSIONS: PGI2 deficiency resulted in the development of vascular disorders with the thickening of vascular walls and interstitial fibrosis, especially in mouse kidneys. The findings demonstrated in vivo that PGI2 is important in the homeostasis of blood vessels. Our established PGID mice are useful for studies on the initiation and development of vascular diseases, such as ischemic renal disorders with arterial sclerosis and infarction, and also for studies on the novel signaling pathway of PGI2.

Purification and characterization of recombinant human prostacyclin synthase.

Prostacyclin synthase (PGIS), which catalyzes the conversion of prostaglandin (PG) H(2) to prostacyclin (PGI(2)), is a member of the cytochrome P-450 (P450) superfamily, CYP8A1. To study the enzymatic and protein characteristics of human PGIS, the enzyme was overexpressed in Spodoptera frugiperda 21 (Sf21) cells using the baculovirus expression system. PGIS was expressed in the microsomes of the infected Sf21 cells after culture in 5 microg/ml hematin-supplemented medium for 72 h. The holoenzyme was isolated from the solubilized microsomal fraction by calcium phosphate gel absorption and purified to homogeneity by DEAE-Sepharose and hydroxyapatite column chromatography. The K(m) and V(max) values of the purified human PGIS for PGH(2) were 30 microM and 15 micromol/min/mg of protein at 24 degrees C, respectively. The optical absorption and EPR spectra of the enzyme revealed the characteristics of a low-spin form of P450 in the oxidized state. The carbon monoxide-reduced difference spectrum, however, exhibited a peak at 418 nm rather than 450 nm. The addition of a PGH(2) analogue, U46619, to the enzyme produced an oxygen-ligand type of the difference spectrum with maximum absorption at 407 nm and minimum absorption at 430 nm. Treatment with another PGH(2) analogue, U44069, produced a peak at 387 nm and a trough at 432 nm in the spectrum (Type I), while treatment with tranylcypromine, a PGIS inhibitor, produced a peak at 434 nm and a trough at 412 nm (Type II). A Cys441His mutant of the enzyme possessed no heme-binding ability or enzyme activity. Thus, we succeeded in obtaining a sufficient amount of the purified recombinant human PGIS from infected insect cells for spectral analyses that has high specific activity and the characteristics of a P450, indicating substrate specificity.