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4-Hydroxybenzyl alcohol (4-HBA) is an important phenolic constituent of Gastrodia elata Blume (GEB), a traditional herbal medicine used in East Asia. Many activities have been reported to underlie the beneficial effects of 4-HBA in the brain, and in particular, its anti-inflammatory, anti-oxidative, and anti-zinc-toxic effects have been implicated in the postischemic brain. Here, the authors investigated the anti-oxidative effect of 4-HBA on astrocytes and sought to identify the underlying molecular mechanisms involved. 4-HBA dose-dependently suppressed H2O2-induced astrocyte cell death. More specifically, pre-incubation of C6 cells (an astrocyte cell line) with 100 μM 4-HBA for 6 hrs increased survival when cells were treated with H2O2 (100 μM, 1 hr) from 54.2±0.7% to 85.9±1.5%. In addition, 4-HBA was found to up-regulate and activate Nrf2, and subsequently, to induce the expressions of several anti-oxidative genes, such as, HO-1, NQO1, and GCLM. Notably, HO-1 was induced by 3.4-fold in 4-HBA-treated C6 cells, and siRNA-mediated HO-1 knockdown demonstrated that Nrf2 activation and HO-1 induction were responsible for the observed cytoprotective effect of 4-HBA. ERK and Akt signaling pathways were activated by 4-HBA in C6 cells, suggesting their involvements in protective effect of 4-HBA. In addition, 4-HBA-conditioned astrocyte culture medium was found to have neuroprotective effects on primary neuronal cultures or fresh C6 cells exposed to oxidative stress, and these effects seemed to be mediated by glial cell line-derived neurotrophic factor (GDNF) and vascular endothelial growth factor (VEGF), which both accumulated in 4-HBA-treated astrocyte culture media. Thus, the 4-HBA-mediated activation of Nrf2 and induction of HO-1 in astrocytes were found to act via autocrine and paracrine mechanisms to confer protective effects. Furthermore, given the pleiotropic effects of 4-HBA with respect to its targeting of various brain cell types and functions, it would appear that 4-HBA has therapeutic potential for the prevention and amelioration of various brain diseases.

Fluoxetine is a selective serotonin reuptake inhibitor (SSRI) and one of the commonly prescribed antidepressants. Numerous clinical observations and animal studies indicate that fluoxetine enhances the anticonvulsant potencies of several antiepileptic drugs. In the previous report, we showed that fluoxetine strongly protects against delayed cerebral ischemic injury. In the present study, the authors investigated whether fluoxetine has a beneficial effect on KA-induced neuronal cell death. An intracerebroventricular (i.c.v.) injection of 0.94 nmol (0.2 microg) of KA produced typical neuronal cell death both in CA1 and CA3 regions of the hippocampus. Although, there was no significant difference in the time course or severity of epileptic behavior, the systemic administration of fluoxetine 30 min before KA administration significantly attenuated this neuronal cell death. Fluoxetine was found to suppress neuronal cell loss when injected at 10 mg/kg and the effect was enhanced at 50 mg/kg. Furthermore, this fluoxetine-induced neuroprotection was accompanied by marked improvements in memory impairment, as determined by passive avoidance tests. KA-induced gliosis and proinflammatory marker (COX-2, IL-1beta, and TNF-alpha) inductions were also suppressed by fluoxetine administration. It is interesting to note here that fluoxetine treatment suppressed NF-kappaB activity dose-dependently in KA-treated mouse brains, suggesting that this explains in part its anti-inflammatory effect. Together, these results suggest that fluoxetine has therapeutic potential in terms of suppressing KA-induced pathogenesis in the brain, and that these neuroprotective effects are associated with its anti-inflammatory effects.

The activation of p38 mitogen-activated protein kinase (MAPK) has been implicated in the pathological changes accompanying inflammatory and apoptotic processes of various cell types including neurons. In a kainic acid (KA)-induced mouse seizure model, p38 MAPK is induced in reactive astrocytes in the CA3 region of the hippocampus where severe neuronal loss occurs. Here we report the delayed and protracted activation of p38 MAPK in the CA3 region of the hippocampus of mice treated with KA. In this model, the inhibition of p38 MAPK isoforms by SB203580, a specific inhibitor, attenuated neuronal loss in the CA3 and CA1 regions of the hippocampus, which was accompanied by the suppression of the p38 MAPK activation as well as astrogliosis. Thus, the delayed and sustained induction of p38 MAPK plays a crucial role in the neuronal damage of KA-induced brain seizures.

Platycodin D (PD) is a major active component of the roots of Platycodon grandiflorum (Jacq.) A.DC. and possesses multiple biological and pharmacological properties, including anti-cancer activity. The aim of this study was to characterize PD-induced cytoplasmic vacuolation in human cancer cells and investigate the underlying mechanisms. PD-induced cancer cell death was associated with cytoplasmic pinocytic and autophagic vacuolation. Cellular energy levels were decreased by this compound, leading to the activation of AMP-activated protein kinase (AMPK). Additionally, compound C, an inhibitor of AMPK, completely prevented PD-induced vacuolation. These results suggest that PD induces cancer cell death, associated with excessive vacuolation through AMPK activation when cellular energy levels are low. Therefore, our findings provide a mechanistic rationale for a novel combinatorial approach using PD to treat cancer. [ABSTRACT FROM AUTHOR]

Ethyl pyruvate (EP) is a simple aliphatic ester of pyruvic acid and has been shown to have robust protective effect in various pathological conditions. A variety of mechanisms have been reported to underlie the protective effects of EP, which include anti-inflammatory, anti-oxidative, and anti-apoptotic functions. Recently, we reported that EP suppressed high mobility group box 1 (HMGB1) release from primary microglial cells via direct Ca 2+ chelation. In the present study, we investigated whether and how EP chelates Zn 2+ in neurons when it is present at toxic levels. In cortical neurons treated with 40 μM of Zn 2+ for 24 h, both EP and pyruvate significantly suppressed neuronal cell death, although the potency of pyruvate was greater than that of EP, and that NAD replenishment contributed to the neuroprotective effects of both pyruvate and EP. However, when cortical neurons were exposed to acute treatment of Zn 2+ (400 μM, 15 min), EP, but not pyruvate, significantly suppressed neuronal death, despite the fact that NAD replenishment by EP was weaker than that by pyruvate. Spectrophotometric studies revealed that EP directly chelates Zn 2+ , and this was confirmed in physiological contexts, such as, NMDA-treated primary cortical cultures and OGD-subjected hippocampal slice cultures, in which EP suppressed intracellular Zn 2+ elevation and neuronal cell death. In addition, EP markedly reduced the expressions of PARP-1 and of the NADPH oxidase subunit in Zn 2+ -treated primary cortical neurons, well known Zn 2+ -induced downstream processes. Together, these results show EP suppresses Zn 2+ induced neurotoxicity via dual functions, chelating Zn 2+ and promoting NAD replenishment. [ABSTRACT FROM AUTHOR]

High mobility group box 1 (HMGB1) was originally identified as ubiquitously expressed nonhistone DNA-binding protein, but recently, it was found to act as an endogenous danger molecule, which signals danger and traumatic cell death. Previously, the authors showed that HMGB1 is massively released immediately after an ischemic insult and that it subsequently activates microglia and induces inflammation in the postischemic brain. Here, we showed the endogenous danger molecule-like function of HMGB1 in primary cortical cultures. HMGB1 was found to be accumulated in NMDA-treated primary cortical culture media, and media collected from these cultures were able to induce neuronal cell death when added to fresh primary cortical cultures. However, HMGB1-depleted NMDA-conditioned media produced by HMGB1 siRNA transfection or by preincubation with anti-HMGB1 antibody or with HMGB1 A box failed to induce neuronal cell death. Furthermore, siRNA-mediated HMGB1 knockdown substantially suppressed NMDA- or Zn-induced cell death. It was interesting to find that extracellular HMGB1-induced neuronal apoptosis, as evidenced by TUNEL staining and caspase 3 assay in combination with double immunofluorescence staining. A series of RAGE and HMGB1 co-immunoprecipitation experiments in the presence of SB203580 and PD98059 (p38 MAPK and ERK inhibitors, respectively) demonstrated that RAGE-p38 MAPK and RAGE-ERK pathway might underlie extracellular HMGB1-mediated neuronal apoptosis. These results together with our previous reports regarding microglial activation by extracellular HMGB1 indicate that HMGB1 functions as a novel danger signal, which aggravates brain damage via autocrine and paracrine manners. [ABSTRACT FROM AUTHOR]

Abstract: αB-crystallin is a member of the small heat shock proteins, which is abundantly expressed in various vertebrate tissues including the central nervous system. In our previous report, we showed αB-crystallin induction in activated astrocytes in the postischemic brain and in H2O2-treated primary astrocyte cultures. To investigate the functional significance of αB-crystallin induction in astrocytes, we generated a stable C6 astroglioma cell line overexpressing αB-crystallin. In these cells, hydrogen peroxide-induced apoptosis was reduced by 60% compared to parent cells. Furthermore, the repression of αB-crystallin expression by αB-crystallin siRNA transfection suppressed this protective effect, indicating that αB-crystallin is responsible for the protection against H2O2-induced apoptosis in C6 astroglioma cells. Similar level of aggravation in H2O2-induced apoptosis was observed in primary astrocyte cultures when αB-crystallin expression was suppressed by αB-crystallin siRNA transfection, confirming the importance of αB-crystallin. In addition, the induction of caspase-3 activity after H2O2 treatment was markedly suppressed in αB-crystallin-overexpressing cells, and immunoprecipitation proved binding between αB-crystallin and partially processed caspase-3 (a p24 intermediate). These results indicate that αB-crystallin confers protection against hydrogen peroxide-induced astrocytes apoptosis in part by inhibiting caspase-3 activation. [Copyright &y& Elsevier]

It has been reported that neutrophil extracellular traps (NETs) play important roles in non-infectious diseases. In ischemic stroke, neutrophils infiltrate damaged brain tissue soon after injury and aggravate inflammation. Using a rat permanent MCAO model, we showed citrullinated histone H3+ (CitH3, a marker of NETosis) induction in neutrophils in leptomeninges and in peripheral blood soon after MCAO. Entry of CitH3+ cells occurred through leptomeninges after 6 h of MCAO and these cells were observed in cerebral cortex from 12 h and subsequently in striatum. It is interesting to note that CitH3+ induction began in circulating neutrophils before they migrated to brain parenchyma and they were detected as intact or lysed form. High mobility group box 1 (HMGB1), a danger associated molecular pattern (DAMP) molecule, was accumulated massively in serum after permanent MCAO and plays a critical role in CitH3 inductions in neutrophils in brain parenchyma and in peripheral blood. Both the all-thiol and disulfide types of HMGB1 induced CitH3 via their specific receptors, CXCR4 and TLR4, respectively. Importantly, HMGB1 not only induced NETosis but was included as a part of the extruded NETs, and contribute to NETosis-mediated neuronal death. Therefore, it would appear a vicious cycle exists between neuronal cell death and NETosis and HMGB1 mediates detrimental effects exerted by this cycle. When NETosis was suppressed by a PAD inhibitor in MCAO animals, delayed immune cell infiltrations were markedly suppressed and damages in blood vessels were significantly mitigated. The study shows NETosis with the involvement of HMGB1 as a mediator in a vicious cycle aggravates inflammation and subsequent damage in the ischemic brain. [ABSTRACT FROM AUTHOR]

Neuromyelitis optica (NMO) is an inflammatory demyelinating disease of the central nervous system and is mediated by complement-dependent cytotoxicity (CDC) of NMO-specific immunoglobulin G (IgG) antibodies (NMO-IgG). Glycyrrhizic acid (GA) has numerous pharmacological effects including inhibition of the complement pathway. We aimed to study the influence of GA on NMO-IgG-induced CDC. NMO-IgG samples from 7 patients with NMO, together with human complement, induced CDC in an aquaporin 4 M23-overexpressing glial cell line, an in vitro NMO model. GA attenuated NMO-IgG-induced CDC in a dose-dependent manner. The mechanism of the GA-related CDC inhibition was sequentially dissected and found to involve inhibition of C1q binding to NMO-IgG. Consequently, GA attenuates NMO-IgG-induced CDC and may be a promising novel therapeutic agent against NMO. [ABSTRACT FROM AUTHOR]