N-acetyl serotonin (NAS) as a melatonin precursor has neuroprotective actions. Nonetheless, it is not clarified how NAS protects neuronal cells against oxidative stress. Recently, we have reported that N-palmitoyl serotonins possessed properties of antioxidants and neuroprotection. Based on those, we hypothesized that NAS, a N-acyl serotonin, may have similar actions in oxidative stress-induced neuronal cells, and examined the effects of NAS based on in vitro and in vivo tests. NAS dose-dependently inhibited oxidative stress-induced cell death in HT-22 cells. Moreover, NAS suppressed glutamate-induced apoptosis by suppressing expression of AIF, Bax, calpain, cytochrome c and cleaved caspase-3, whereas it enhanced expression of Bcl-2. Additionally, NAS improved phosphorylation of tropomyosin-related kinase receptor B (TrkB) and cAMP response element-binding protein (CREB) as well as expression of brain-derived neurotrophic factor (BDNF), whereas the inclusion of each inhibitor of JNK, p38 or Akt neutralized the neuroprotective effect of NAS, but not that of ERK. Meanwhile, NAS dose-dependently reduced the level of reactive oxygen species, and enhanced the level of glutathione in glutamate-treated HT-22 cells. Moreover, NAS significantly increased expression of heme oxygenase-1, NAD(P)H quinine oxidoreductase-1 and glutamate-cysteine ligase catalytic subunit as well as nuclear translocation of NF-E2-related factor-2. Separately, NAS at 30 mg/kg suppressed scopolamine-induced memory impairment and cell death in CA1 and CA3 regions in mice. In conclusion, NAS shows actions of antioxidant and anti-apoptosis by activating TrkB/CREB/BDNF pathway and expression of antioxidant enzymes in oxidative stress-induced neurotoxicity. Therefore, such effects of NAS may provide the information for the application of NAS against neurodegenerative diseases.

Haginin A, an isoflav-3-ens isolated from the branch of Lespedeza cyrtobotrya, is almost unknown. Here, we report that haginin A exhibits a strong hypopigmentary effect in Melan-a cells and significantly inhibits melanin synthesis. Haginin A shows potent inhibitory effects with an IC(50) (half-maximal inhibitory concentration) value of 5.0 microM on mushroom tyrosinase activity, and functioned as a noncompetitive inhibitor. Also, haginin A decreased microphthalmia-associated transcription factor (MITF), tyrosinase, and tyrosinase-related protein-1 (TRP-1) protein production. To identify the signaling pathway of haginin A, the ability of haginin A to influence extracellular signal-regulated protein kinase (ERK) and Akt/protein kinase B (PKB) activation was investigated. Apparently, haginin A induced ERK and Akt/PKB in a dose-dependent manner. In addition, the specific inhibition of the ERK and the Akt/PKB signaling pathways by PD98059 and LY294002, respectively, increased melanin synthesis. Furthermore, haginin A decreased UV-induced skin pigmentation in brown guinea-pigs. Also, haginin A presented remarkable inhibition on the body pigmentation in the zebrafish model system and decreased tyrosinase activity. Together, haginin A is an effective inhibitor of hyperpigmentation caused by UV irradiation or by pigmented skin disorders through downregulation via ERK and Akt/PKB activation, MITF, and also by the subsequent downregulation of tyrosinase and TRP-1 production.

No abstract

Some endocannabinoids have been known to express anti-inflammatory and antioxidant actions independent of cannabinoid receptors. In this respect, we investigated whether N-acyl 5-hydroxytryptamines (5-HTs) might prevent against glutamate-induced oxidative cytotoxicity in HT-22 cells, and attempted to elucidate the mechanism for their cytoprotective action. N-acyl 5-HTs with palmitoyl, stearoyl, arachidonoyl or docosahexaenoyl chain expressed a remarkable protective effect on glutamate-induced cytotoxicity. Additionally, glutamate-induced oxidative stress, represented by the increase of reactive oxygen species level and the reduction of glutathione level, was prevented markedly by N-acyl 5-HTs at submicromolar levels. Further, N-palmitoyl 5-HT, the most cytoprotective, enhanced antioxidant defense by up-regulating the expression of antioxidant enzymes such as heme oxygenase-1, glutamate-cysteine ligase catalytic subunit or NAD(P)H quinine oxidoreductase-1 in the presence or absence of glutamate. Consistent with this, N-palmitoyl 5-HT stimulated nuclear translocation of Nrf2 in early phase (2 h), and this effect was remarkably suppressed by inhibitors of PI3K, PDK-1, Akt or p38 MAPK. Additionally, N-palmitoyl 5-HT suppressed glutamate-induced activation of ERK in late phase (12 h), but not in early phase (2 h), presumably supporting the implication of MEK/ERK pathway in glutamate-induced cytotoxicity. Collectively, it is suggested that N-acyl 5-HTs may attenuate glutamate-induced cytotoxicity via the activation of PI3K/PDK-1/Akt- and p38 MAPK-dependent Nrf2 signaling in early phase as well as the suppression of MEK/ERK pathway in late phase.

The dopamine precursor L-3,4-dihydroxyphenylalanine (L-DOPA) is widely used as a therapeutic choice for the treatment of patients with Parkinson's disease. However, the long-term use of L-DOPA leads to the development of debilitating involuntary movements, called L-DOPA-induced dyskinesia (LID). The cAMP/protein kinase A (PKA) signaling in the striatum is known to play a role in LID. However, from among the nine known adenylyl cyclases (ACs) present in the striatum, the AC that mediates LID remains unknown. To address this issue, we prepared an animal model with unilateral 6-hydroxydopamine lesions in the substantia nigra in wild-type and AC5-knock-out (KO) mice, and examined behavioral responses to short-term or long-term treatment with L-DOPA. Compared with the behavioral responses of wild-type mice, LID was profoundly reduced in AC5-KO mice. The behavioral protection of long-term treatment with L-DOPA in AC5-KO mice was preceded by a decrease in the phosphorylation levels of PKA substrates ERK (extracellular signalregulated kinase) 1/2, MSK1 (mitogen-and stress-activated protein kinase 1), and histone H3, levels of which were all increased in the lesioned striatum of wild-type mice. Consistently, FosB/⌬FosB expression, which was induced by long-term L-DOPA treatment in the lesioned striatum, was also decreased in AC5-KO mice. Moreover, suppression of AC5 in the dorsal striatum with lentivirusshRNA-AC5 was sufficient to attenuate LID, suggesting that the AC5-regulated signaling cascade in the striatum mediates LID. These results identify the AC5/cAMP system in the dorsal striatum as a therapeutic target for the treatment of LID in patients with Parkinson's disease.

Lysophosphatidylcholine is known to be a lipid mediator in various cellular responses. In this study, we examined the anti-inflammatory actions of lysophosphatidylcholine containing docosahexaenoic acid esterified at the sn-1 position. First, in RAW 264.7 cells, DHA-lysoPtdCho suppressed the LPS-induced formation of NO concentration-dependently. However, ARA-lysoPtdCho showed a partial suppression, and LNA-lysoPtdCho had no significant effect. Additionally, DHA-lysoPtdCho also reduced the level of TNF-alpha or IL-6, but not PGE(2). In animal experiments, the i.v. administration of ARA-lysoPtdCho (150 or 500 mug/kg) prevented zymosan A-induced plasma leakage remarkably with a maximal efficacy (Emax) of 50%, in contrast to no effect with LNA-lysoPtdCho. Remarkably, DHA-lysoPtdCho suppressed zymosan A-induced plasma leakage with an ED(50) value of 46 mug/kg and an Emax value of around 95%. Additionally, mechanistic studies indicated that the anti-inflammatory action of DHA-lysoPtdCho was partially related to the reduced formation of LTC(4,) TNF-alpha, and IL-6. When the interval time between lysoPtdCho administration and zymosan A challenge was extended up to 2 h, such a suppressive action of DHA-lysoPtdCho was augmented, suggesting that a DHA-lysoPtdCho metabolite is important for anti-inflammatory action. In support of this, 17-HPDHA-lysoPtdCho showed a greater anti-inflammatory action than DHA-lysoPtdCho. Furthermore, a similar anti-inflammatory action was also observed with i.p. administration of DHA-lysoPtdCho or a 17(S)-hydroperoxy derivative. Additionally, oral administration of DHA-lysoPtdCho also expressed a significant anti-inflammatory action. Taken together, it is proposed that DHA-lysoPtdCho and its metabolites may be anti-inflammatory lipids in vivo systems.

In this study, the anti-inflammatory and antisepticemic activities of a water extract of aged black garlic (AGE), which is not pungent, were compared with those of raw garlic extract (RGE). The methyl thiazolyl tetrazolium (MTT) assay showed that AGE was not toxic up to 1000 μg/mL and was at least four times less cytotoxic than RGE. AGE significantly suppressed the production of nitric oxide (NO), tumor-necrosis factor-α (TNF-α), and prostaglandin (PG)-E2 in a dose-dependent manner in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. Furthermore, the inhibitory effect of AGE on LPS-induced inflammation was confirmed by downregulation of inducible NO synthase and TNF-α mRNA expression, as well as cyclooxygenase-2 protein expression. The anti-inflammatory activities of AGE were similar to those of RGE at nontoxic concentrations up to 250 μg/mL. Signal transduction pathway studies further indicated that both garlic extracts inhibited activation of mitogen-activated protein kinase and nuclear factor-κB induced by LPS stimulation. Treatment with both AGE and RGE in an in vivo experiment of LPS-induced endotoxemia significantly reduced the level of TNF-α and interleukin-6 in serum and completely protected against LPS-induced lethal shock in C57BL/6 mice. The results suggest that AGE is a more promising nutraceutical or medicinal agent to prevent or cure inflammation-related diseases for safety aspects compared with RGE.

SummaryThe anti-atherogenic effects of spirulina ( Spirulina platensis ) were investigated in the New Zealand White (NZW) rabbit model. The animal had hypercholesterolemia induced by being fed a high cholesterol diet (HCD) containing 0.5% cholesterol for 4 wk, and then fed a HCD supplemented with 1 or 5% spirulina (SP1 or SP5) for an additional 8 wk. Spirulina supplementation lowered intimal surface of the aorta by 32.2 to 48.3%, compared to HCD. Serum triglyceride (TG) and total cholesterol (TC) significantly were reduced in SP groups. After 8 wk, serum low density lipoprotein cholesterol (LDL-C) remarkably decreased by 26.4% in SP1 and 41.2% in SP5, compared to HCD. On the other hand, high density lipoprotein cholesterol (HDL-C) was markedly increased in SP1 and SP5 compared with that in the HCD group from 2 to 8 wk. These results suggest that spirulina intake can cause the reduction of hypercholesterolemic atherosclerosis, associated with a decrease in levels of serum TC, TG and LDL-C, and an elevation of HDL-C level. Spirulina may, therefore, be beneficial in preventing atherosclerosis and reducing risk factors for cardiovascular diseases.