Early growth-responsive-1-dependent manganese superoxide dismutase gene transcription mediated by platelet-derived growth factor.

Manganese superoxide dismutase Mn-SOD plays a major role in protecting mitochondria from oxidative damage. Overexpression of Mn-SOD maintains cell survival under conditions that lead to apoptotic death. In addition to the antioxidative enzyme, platelet-derived growth factor (PDGF) is a principal survival factor that inhibits apoptosis and promotes proliferation by activating survival signaling pathways in various cells. Here we show that PDGF induced the expression of the Mn-SOD gene in NIH3T3 cells, and its induction was associated with early growth response-1 (Egr-1), a transcription factor. An electrophoretic mobility shift assay demonstrated that Egr-1 bound to the proximal promoter of the Mn-SOD gene in response to PDGF. The proximal promoter region of Mn-SOD was shown to be transcriptionally responsive to both basal and PDGF stimulation by transfection studies. Forced expression of Egr-1 in the cells activated Mn-SOD transcription in a dose-dependent manner. The pathway by which PDGF induced Egr-1 involved the mitogen-activated protein kinase kinase-1 (MEK1) and extracellular signal-regulated kinases 1 and 2 (ERK1/2), because the effect of PDGF on the induction of Egr-1 was blocked by U0126, a specific MEK1 inhibitor. These findings indicate that the induction of Mn-SOD is part of the anti-apoptotic properties mediated by PDGF.

Peroxynitrite induces GADD34, 45, and 153 VIA p38 MAPK in human neuroblastoma SH-SY5Y cells.

Peroxynitrite, one of the most reactive radicals, is produced from superoxide anion and nitric oxide. A peroxynitrite generator, 3-morpholinosydonimine (SIN-1), was found to induce the expression of three different growth arrest and DNA damage-inducible (GADD) mRNA, GADD34, GADD45, and GADD153, at the early phase during cell death in human neuroblastoma SH-SY5Y cells. In addition, peroxynitrite activated p38 MAPK just before induction of three GADD mRNA. A specific inhibitor of p38 MAPK, SB202190, markedly suppressed peroxynitrite-induced expression of three GADD mRNA in SH-SY5Y cells. The expression of three GADD genes and also p38 MAPK phosphorylation were suppressed by treatment with radical scavengers, superoxide dismutase plus catalase and glutathione. Glutathione depletion by L-buthionine-S, R-sulfoximine (BSO), increased the vulnerability of the cells to peroxynitrite. These findings indicate that peroxynitrite-mediated oxidative stress activated p38 MAPK to induce three GADD genes.

Peroxynitrite induces neuronal cell death in aging and age-associated disorders: A review.

Peroxynitrite produced from nitric oxide and superoxide has been proposed to cause neuronal dysfunction and cell death in aging and age-related degenerative diseases. 3-Nitrotyosine, an oxidation product of tyrosine by peroxynitrite, was reported to increase in degenerating brains. In this paper, involvement of peroxynitrite in neuronal cell death was studied by analyses of human brains and in vitro experiments on cell death induced by a peroxynitrite-generating agent, SIN-1. 3-Nitrotyrosine-containing proteins were detected in lipofuscin, a typical aging-related pigment in human brains. The cytotoxicity of peroxynitrite was examined in human dopaminergic SH-SY5Y cells by use of SIN-1. SIN-1 induced apoptotic cell death in the cells, and increased the level of 3-nitrotyrosine-containing proteins. The intracellular transduction of death signal was studied in apoptosis induced by peroxynitrite. Apoptosis was induced by sequential death cascade, collapse of mitochondrial membrane potential, activation of caspases and fragmentation of nuclear DNA. In addition, phosphorylation of p38 mitogen activated phosphokinase (MAPK) was found to be associated with apoptosis by SIN-1, as shown by inhibition of apoptotic process by SB202190, a p38 inhibitor. Involvement of peroxynitrite in the cell death is discussed in relation to neuronal degeneration in aging and age-associated diseases.

Mitogen-activated protein kinase pathway mediates peroxynitrite-induced apoptosis in human dopaminergic neuroblastoma SH-SY5Y cells.

Peroxynitrite, a product of nitric oxide and superoxide, is one of the most potent oxidants and it has been suggested to be involved in many neurodegenerative disorders. The mechanism of the cytotoxicity by peroxynitrite was examined using 3-morpholinosydonimine (SIN-1) as a peroxynitrite donor and SH-SY5Y cells as a model of dopamine neurons. SIN-1 was found to induce apoptotic cell death with typical nucleosomal DNA fragmentation with activation of caspase 3-like proteases. The signal transduction of apoptosis was studied in concern to mitogen-activated protein kinases (MAPKs). After SIN-1 treatment, phosphorylation of p38 was detected, followed by that of Erk. SB202190, an inhibitor of p38, suppressed Erk phosphorylation to the basal level and partially reduced the activation of caspase 3-like proteases and also the cell death. These results suggest that peroxynitrite may activate p38 MAPK pathway to induce apoptosis in dopamine cells via activation of caspase 3-like proteases.

Dietary alpha-linolenate suppresses endotoxin-induced platelet-activating factor production in rat kidney.

In comparison with dietary high-linoleate safflower oil, high alpha-linolenate perilla oil decreased alkylacyl- and alkenylacyl-glycerophosphocholine (GPC) content in rat kidney by roughly 30 and 25%, respectively. The fatty acid composition was also modified by high alpha-linolenate oil; arachidonic acid (AA) level in alkylacyl-GPC, a platelet-activating factor (PAF) precursor, decreased by 30% along with concomitant increases in the n-3 fatty acid levels. PAF contents under resting conditions were similarly low in the two dietary groups. Fifteen minutes after endotoxin administration, PAF and lyso-PAF contents increased significantly, and the PAF content in the high alpha-linolenate group was 60% lower than in the high linoleate group; the lyso-PAF contents also tended to be lower. Lyso-PAF acetyltransferase and CoA-independent transacylase activities in kidney microsomes increased significantly after endotoxin administration, while PAF acetylhydrolase activity in the cytosol was relatively unchanged. The lyso-PAF acetyltransferase and PAF acetylhydrolase activities did not differ between the two dietary groups, but the CoA-independent transacylase activity was roughly 30% lower in the high alpha-linolenate group. In agreement with in vitro study, our present study demonstrates that dietary high alpha-linolenate suppresses PAF production in rat kidney during systemic endotoxemia, and which is mainly due to the decrease in alkylacyl-GPC content, altered fatty acid compositions of the precursor lipids and lower CoA-independent transacylase activity.

Reevaluation of the effect of a high alpha-linolenate and a high linoleate diet on antigen-induced antibody and anaphylactic responses in mice.

Previously, we reported that a high alpha-linolenate [18:3(n-3)] diet compared with a high linoleate [18:2(n-6)] diet suppressed the anti-egg albumin (EA) immunoglobulin E (IgE) antibody response in mice. Because of relatively high background values obtained with the method used previously, we used an improved ELISA and once again determined serum IgE levels. In contrast to our previous results, the serum level of anti-dinitrophenyl specific (anti-DNP) as well as total IgE in mice immunized with DNP-antigen was slightly but significantly higher in the high alpha-linolenate diet group than in the high linoleate diet group. Anti-DNP IgG1 and IgG2a antibody responses were not significantly different in mice fed these diets. Indomethacin administration during immunization tended to enhance the IgE antibody responses. The mortality of mice from antigen-induced anaphylactic shock was significantly lower in the high alpha-linolenate diet group than in the high linoleate diet group; however, there was no difference between the groups in terms of vascular permeability and histamine levels. Thus, the high alpha-linolenate diet enhances the IgE antibody response slightly without affecting either the IgG antibody response, vascular permeability or histamine release. The high alpha-linolenate diet possibly suppresses anaphylactic shock by reducing the synthesis of lipid mediators such as eicosanoids and platelet-activating factor.

Possible mechanisms for the differential effects of high linoleate safflower oil and high alpha-linolenate perilla oil diets on platelet-activating factor production by rat polymorphonuclear leukocytes.

As compared with high dietary linoleate safflower oil, high dietary alpha-linolenate perilla oil decreased platelet-activating factor (PAF) production by nearly half in calcium ionophore (CaI)-stimulated rat polymorphonuclear leukocytes (PMN). In the CaI-stimulated PMN from the perilla oil group, the accumulated amount of arachidonate (AA) plus eicosapentaenoate (EPA) was 30% less and that of lyso-PAF was 50% less, indicating that the decreased availability of lyso-PAF is a factor contributing to the relatively low PAF production. Consistently, eicosatetraynoic acid (ETYA), a dual inhibitor of cyclooxygenase and lipoxygenase, increased free fatty acids (FFA) and decreased PAF production possibly by decreasing the availability of lyso-PAF. Although, leukotrienes (LTs) have been proposed to stimulate PAF production synergistically, a potent LTB4 receptor antagonist, ONO-4057, decreased the formation of free fatty acids and LTB4, but stimulated PAF production somewhat, indicating that LTB4 may not stimulate PAF production in PMN. Lysophospholipid-induced transacylase (CoA-independent transacylase) activity in PMN homogenates was 25-30% lower in the perilla oil group but no significant differences were observed in the lyso-PAF acetyltransferase and PAF acetylhydrolase activities between the two dietary groups. Thus, decreased transacylase activity is another factor associated with the relatively low PAF production in the perilla oil group.