We examined the inhibition of the expression of some inflammatory genes associated with ischemia-reperfusion (I/R) injury by anthocyanins isolated from black soybean seed coat in tumor necrosis factor-alpha (TNF-a)-treated bovine aortic endothelial cells. In addition, its potential use on I/R-injury was investigated using rats subjected to 30-min occlusion of left descending coronary artery followed by 24-h reperfusion. Western blot analysis and luciferase activity assay showed that anthocyanins inhibited TNF-a-induced vascular cell adhesion molecule-1, intracellular adhesion molecule-1, and cyclooxygenase-2 levels, which is through NF-jB-dependent pathway. Further, anthocyanins protected myocardiac injury from I/R in rats. It is suggested that anthocyanins from black soybean seed coat can be used as a useful drug to modulate cardiovascular disorder.

Nitric oxide (NO) is a potent inducer of heme oxygenase (HO)-1, and NO-induced HO-1 expression is dependent on the cGMPsignaling pathway. Sodium nitroprusside (SNP) produces NO and iron. However, it is unclear whether NO is exclusively responsible for induction of HO-1 by SNP in RAW 264.7 cells. We tested our hypothesis that iron may contribute more to the SNP induction of HO-1 than does NO by comparing the HO-1 protein level and the production of NO in RAW 264.7 cells treated with SNP and S-nitroso-N-acetyl-DL-penicillamine (SNAP). Although SNP induced less NO production than SNAP, SNP induced the production of more HO-1 protein than did SNAP. Deferoxamine (DFO) decreased SNP-but not SNAPinduced HO-1 expression but did not decrease the production of NO. SNP-induced HO-1 was significantly inhibited by specific protein kinase A (PKA) inhibitors or an antagonist of cAMP but not by guanylyl cyclase inhibitors. Exogenous iron (ferric ammonium citrate or ferricyanide) and forskolin increased the level of HO-1, which was inhibited by PKA inhibitor N-[2-(4-bromocinnamylamino)ethyl]-5-isoquinoline (H89). These results indicate that iron and cAMP, but not cGMP, play crucial roles in the induction of HO-1 in RAW 264.7 cells. Moreover, DFO and inhibitors of extracellular signal-related kinases 1/2 or c-Jun NH 2 -terminal kinase inhibited HO-1 production induced by SNP. This study illustrates that iron rather than NO from SNP contributes to HO-1 induction. Therefore, studies on the effects of SNP should consider the role of iron in some biological functions. We concluded that iron released by SNP contributes to HO-1 induction via the cAMP-PKA-mitogen-activated protein kinase pathway.Heme oxygenase (HO) is an essential enzyme in heme catabolism that cleaves heme to release carbon monoxide, iron, and biliverdin (Tenhunen et al., 1968(Tenhunen et al., , 1969. HO-1 is induced by a variety of physiological stimuli, including heme, heavy metals, inflammatory cytokines, endotoxins, and nitric oxide (NO) (Durante et al., 1997b;Maines, 1997;Yet et al., 1997). Recent studies have shown that HO-1 expression plays a critical role in mediating antioxidant, anti-inflammatory, and antiapoptotic effects (Brouard et al., 2000;Otterbein et al., 2000). The beneficial effects of HO-1 induction might occur via several postulated mechanisms. Increased HO-1 activity results in the degradation of the heme moiety, a potentially toxic prooxidant, and generates bilirubins, an antioxidant capable of scavenging peroxy radicals and inhibiting lipid peroxidation (Stocker et al., 1987;Llesuy and Tomaro, 1994;Nath et al., 1998).NO is a free radical involved in the regulation of many physiological functions, including endothelium-dependent vasodilation, neurotransmission, and the cell-mediated im-

We investigated anti-invasive effects of the anthocyanins from fruits of Vitis coignetiae Pulliat (known as meoru in Korea) on human hepatoma Hep3B cells. The anthocyanins inhibited cell invasion in a dose-dependent manner as measured by Matrigel (BD Biosciences, San Jose, CA, USA) invasion assays. They also inhibited expression of matrix metalloproteinase (MMP)-2 and MMP-9 and activation of nuclear factor kappaB (NF-kappaB) stimulated by tumor necrosis factor alpha. Taken together, the results of this study indicate that the anthocyanins isolated from fruits of V. coignetiae Pulliat have anti-invasive effects on human hepatoma Hep3B cells and inhibit MMP-2 and MMP-9 gene expression at least in part through the inhibition of NF-kappaB activation.

We hypothesized that catecholamines through b-adrenoceptor might modulate macrophage function. We showed that isoproterenol concentration-dependently induced HO-1 production through b 1 -but not b 2 -adrenoceptor. Production was increased by forskolin and inhibited by pretreatment with the PKA inhibitor, H-89. Furthermore, induction of HO-1 by isoproterenol effectively protected RAW264.7 cells from effects of glucose oxidase treatment, which was abrogated either by HO-1 inhibitor, ZnPP IX and b-adenoceptor antagonist, propranolol. Thus, stimulation of HO-1 production through b 1 -adenoceptors, and via the PKA pathways by isoproterenol, can enable RAW264.7 cells to resist oxidant stress, suggesting that catecholamine hormones may be necessary, at least, to maximize defending role of macrophages.

Infection with Helicobacterpylori increases apoptosis and the activity of the inducible form of NO synthase (;NOS) in gastric mucosa. However, whether NO triggers apoptosis or exerts a defensive role in such circumstances is unclear. The purpose of this work was therefore firstly to examine apoptotic responses of rat gastric mucosal cells subjected to long term exposure to exogenous NO in vitro, and secondly to determine whether induction of iNOS in vivo, by injection of lipopolysaccharide (LPS), caused similar changes to those obtained in vitro. The specific involvement of NO in effects of LPS were examined by using the selective iNOS inhibitor 1400W. Apoptotic activity was assessed by assay of caspase )-like activity and from fragmentation of DNA. Caspase Mike activity in isolated rat gastric mucosal cells at 24, 36, 48 and 60 h after plating of cells was substantially greater in the detached cells than in those that were attached to the culture plate. For both attached and detached cells the addition of the NO donor S-nitroso-Nacetyl-penicillamine (SNAP, 0.05 -2 mM) on plating produced a doserelated inhibition of caspase 3-like activity. Also at 60 h after plating the presence of SNAP (2 mM) prevented the appearance of DNA ladders (apoptotic DNA fragmentation) in cells which had detached from the plate. Injection of LPS (3 mg/kg, i.v.) to rats in vivo increased caspase 3-like activity measured 5 and 24 h later in rat gastric mucosa. Co-administration of the selective iNOS inhibitor, 1400 W (5 mglkg, i.v.), with LPS enhanced caspase 3-like activity above that with LPS alone. In conclusion data obtained both in vitro and zn vivo suggest that NO exerts an anti-apoptotic effect in rat gastric mucosa. VP5, 5'teminal small open reading frame (small ORF) of the A segment of the aquatic birnavirus (infectious pancreatic necroaia virus, IPNV) genome, encodes a 17 kDa non-structure protein.In recently, we reported that apoptosis induced by IPNV in a fish cell line. In the present study, we cloned and identified the VP5 and test its function. Compared the amino acids sequence of VP5 with well known Bcl-2 family member proteins that found the VP5 protein also contains the Bcl-2 homology (BH) domains BH1, BH2, BH3 and BH4 but the absence of the transmembrane region (TM). In addition VP5 stable clone either CHSE-VP5 or ZLE-VP5 also showed delay cell death and DNA internucleoso-ma1 cleavage during IPNV infection. O n the other hand, VP5 could both up-regulation of the survival factor Mcl-1 for enhance host survival and could to limit-regulate the specific viral proteins expression in early replication cycle. Taken together, these results suggest that the aquatic birnavirus may use a notable strategy with VP5 (a novel viral Bcl-2 family member protein) to against the host defense and enhance the progeny production.

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