Pyeongjae Lee

Apigenin inhibits the production of NO and PGE2 in microglia and inhibits neuronal cell death in a middle cerebral artery occlusion-induced focal ischemia mice model

Flavonoids have been intensively studied on their pharmacological activities such as anti-cancer, anti-oxidant and anti-inflammation. However, little is known about their neuroprotective effects. Recent studies suggest that inflammation mediated by microglia may play a role in neurodegenerative diseases. In this study, we evaluated the anti-inflammatory effect of various flavonoid compounds by using BV-2, a murine microglia cell line. Of the compounds that were evaluated, apigenin inhibited the production of nitric oxide and prostaglandin E(2) by suppressing the expression of inducible nitric oxide synthase and cyclooxygenase-2 protein, respectively. Moreover, apigenin suppressed p38 mitogen-activated protein kinase (MAPK), c-Jun N-terminal kinase (JNK) phosphorylation without affecting the activity of extracellular signal-regulated kinase (ERK). Apigenin was also found to protect neuronal cells from injury in middle cerebral artery occlusion.

Cytoprotective effect of Scutellaria baicalensis in CA1 hippocampal neurons of rats after global cerebral ischemia

Based on the use of Scutellaria baicalensis for the treatment of stroke in traditional Oriental medicine, the current study was carried out to evaluate neuroprotective effects of S. baicalensis after transient global ischemia using rat 4-vessel occlusion model. Methanol extracts from the dried roots of S. baicalensis (0.1-10 mg/kg) administered intra-peritoneally significantly protected CA1 neurons against 10 min transient forebrain ischemia as demonstrated by measuring the density of neuronal cells stained with Cresyl violet. Methanol extract of S. baicalensis inhibited microglial tumor necrosis factor-alpha (TNF-alpha) and nitric oxide production, and protected PC12 cells from hydrogen peroxide-induced toxicity in vitro.

Ischemia-activated microglia induces neuronal injury via activation of gp91phox NADPH oxidase

Although glial cells play a major role in the pathogenesis of many neurological diseases by exacerbating neuronal and non-neuronal cell death, the mechanisms involved are unclear. We examined the effects of microglia-(MCM) or astrocyte-(ACM) conditioned media obtained by chemical ischemia on the neuronal injury in SH-SY5Y cells. Chemical ischemia was induced by the treatment with NaN(3) and 2-deoxy-d-glucose for 2h. MCM-treated SH-SY5Y cells showed reduced the viability, increased caspase-3 activity, decreased Bcl-2/Bax ratio, and increased cytochrome c release, increased inflammatory cytokines, and increased reactive oxygen species (ROS) generation. MCM also increased gp91phox nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, which was inhibited by NADPH oxidase inhibitor, apocynin, and gp91phox siRNA. However, ACM did not show any significant changes. The results suggest that microglia activated by ischemic insult may increase reactive oxygen species generation via activation of gp91phox NADPH oxidase, resulting in neuronal injury.

Neurite outgrowth induced by spicatoside A, a steroidal saponin, via the tyrosine kinase A receptor pathway

Although nerve growth factor (NGF) therapy is an available option for the treatment of Alzheimers disease, several limitations exist in its medical application. In the present study, we examined the neurotrophic effects of spicatoside A isolated from Liriope platyphylla on PC12 cells as well as the mechanisms involved in this process. Spicatoside A (10 microg/mL) induced neurite outgrowth similar to NGF (50 ng/mL). Furthermore, spicatoside A, a steroidal saponin, activated extracellular signal-regulated kinase 1/2 (ERK 1/2) and phosphatidylinositol 3-kinase (PI 3-kinase/Akt) via tyrosine receptor kinase A (TrkA), which is responsible for the induction of the neuritic process. The effects of NGF and spicatoside A on neurite outgrowth disappeared in TrkA knockdown PC12 cells by siRNA. In conclusion, neuritogenic effects resulting from spicatoside A may be involved in TrkA activation.

Alkaloid fraction of Uncaria rhynchophylla protects against N-methyl-d-aspartate-induced apoptosis in rat hippocampal slices

Uncaria rhynchophylla is a medicinal herb which has sedative and anticonvulsive effects and has been applied in the treatment of epilepsy in Oriental medicine. In this study, the effect of alkaloid fraction of U. rhynchophylla against N-methyl-D-aspartate (NMDA)-induced neuronal cell death was investigated. Pretreatment with an alkaloid fraction of U. rhynchophylla for 1 h decreased the degree of neuronal damage induced by NMDA exposure in cultured hippocampal slices and also inhibited NMDA-induced enhanced expressions of apoptosis-related genes such as c-jun, p53, and bax. In the present study, the alkaloid fraction of U. rhynchophylla was shown to have a protective property against NMDA-induced cytotoxicity by suppressing the NMDA-induced apoptosis in rat hippocampal slices.

15,16-Dihydrotanshinone I suppresses the activation of BV-2 cell, a murine microglia cell line, by lipopolysaccharide

Microglial activation has been implicated in neurodegenerative diseases. Therefore, inhibition of inflammation mediated by microglia is a strategy in neurodegenerative disease therapy. In this study, we isolated cryptotanshinone and 15,16-dihydrotanshinone I from Salvia miltiorrhiza, a traditional Korean herb medicine, by bioactivity-guided fractionation based on inhibitory effect on nitric oxide in a lipopolysaccharide-stimulated BV-2 cells, a murine microglial cell line. 15,16-Dihydotanshinoe I suppressed the expression of not only inducible nitric oxide synthase but also of interleukin-1beta, tumor necrosis factor-alpha, and of TNF-alpha converting enzyme.

Excessive production of nitric oxide induces the neuronal cell death in lipopolysaccharide-treated rat hippocampal slice culture.

In this study, we investigated the role of nitric oxide (NO) in neuronal cell death in the lipopolysaccharide (LPS)-treated rat hippocampal slice culture. Neuronal cell death was induced after a LPS treatment in a rat hippocampal slice. The region and time course of the cell death caused by LPS were different from those by N-methyl-D-aspartate. Cell death was inhibited when the slice was co-treated with both LPS and N-monomethyl-L-arginine, a nitric oxide synthase inhibitor. On the other hand, sodium nitroprusside, a NO donor, induced cell death significantly. A reverse transcription-polymerase chain reaction and Western analysis showed that inducible nitric oxide synthase (iNOS) was induced in the LPS-treated hippocampal slice culture. These results suggest that NO produced by iNOS is a major mediator of cell death in LPS-treated hippocampal slice culture.

Protective effects of standardized Thuja orientalis leaves against 6-hydroxydopamine-induced neurotoxicity in SH-SY5Y cells.

Although the etiology of Parkinsons disease (PD) remains unknown, recent studies have suggested that oxidative stress (OS) and apoptosis, as a result of mitochondrial defects, may play important roles in its pathogenesis. 6-Hydroxydopamine (6-OHDA), a neurotoxin commonly used in models of PD, induces selective catecholaminergic cell death, mediated by reactive oxygen species (ROS) and mitochondrial defects. This study investigated the protective effect of Thuja orientalis leaves (TOFE), a well-known oriental traditional medicine, on 6-OHDA-induced neurotoxicity in SH-SY5Y cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and Hoechst staining showed that TOFE attenuated the cell damage caused by 6-OHDA stress. TOFE showed strong radical scavenging effects in 2,2-diphenyl-2-picrylhydrazyl and 2,2-azinobis-(3-ethyl-benzthiazoline-6-sulphonic acid) assays, and it reduced the intracellular ROS and extracellular nitric oxide production induced by 6-OHDA. Additionally, TOFE blocked the reduction in the mitochondrial membrane potential, the release of cytochrome c, and the activation of caspase-3. Moreover, TOFE decreased the phosphorylation of extracellular signal-regulated kinase (pERK), which has pro-apoptotic functions. Taken together, TOFE might protect SH-SY5Y cells from 6-OHDA through the downregulation of OS and mitochondrial-mediated apoptosis, and regulation of pERK.

Regulatory Effect of 25-hydroxyvitamin D3 on Nitric Oxide Production in Activated Microglia.

Microglia are activated by inflammatory and pathophysiological stimuli in neurodegenerative diseases, and activated microglia induce neuronal damage by releasing cytotoxic factors like nitric oxide (NO). Activated microglia synthesize a significant amount of vitamin D3 in the rat brain, and vitamin D3 has an inhibitory effect on activated microglia. To investigate the possible role of vitamin D3 as a negative regulator of activated microglia, we examined the effect of 25-hydroxyvitamin D3 on NO production of lipopolysaccharide (LPS)-stimulated microglia. Treatment with LPS increased the production of NO in primary cultured and BV2 microglial cells. Treatment with 25-hydroxyvitamin D3 inhibited the generation of NO in LPS-activated primary microglia and BV2 cells. In addition to NO production, expression of 1-α-hydroxylase and the vitamin D receptor (VDR) was also upregulated in LPS-stimulated primary and BV2 microglia. When BV2 cells were transfected with 1-α-hydroxylase siRNA or VDR siRNA, the inhibitory effect of 25-hydroxyvitamin D3 on activated BV2 cells was suppressed. 25-Hydroxyvitamin D3 also inhibited the increased phosphorylation of p38 seen in LPS-activated BV2 cells, and this inhibition was blocked by VDR siRNA. The present study shows that 25-hydroxyvitamin D3 inhibits NO production in LPS-activated microglia through the mediation of LPS-induced 1-α-hydroxylase. This study also shows that the inhibitory effect of 25-hydroxyvitamin D3 on NO production might be exerted by inhibiting LPS-induced phosphorylation of p38 through the mediation of VDR signaling. These results suggest that vitamin D3 might have an important role in the negative regulation of microglial activation.

Neochlorogenic Acid Inhibits Lipopolysaccharide-induced Activation and Pro-inflammatory Responses in BV2 Microglial Cells

Microglia is the resident innate immune cells that sense pathogens and tissue injury in the central nervous system. Microglia becomes activated in response to injury, infection, and other stimuli that threaten neuronal survival. Microglia activation plays an important role in neurodegenerative diseases. Neochlorogenic acid (NCA) is a natural polyphenolic compound found in dried fruits and other plants. Although previous studies have shown that phenolic acids including NCA have outstanding antioxidant, antibacterial, antiviral, and antipyretic activities, there has not yet been investigated for anti-inflammatory effects. Therefore, for the first time we have examined the potential of NCA to inhibit microglial activation and pro-inflammatory responses in the brain. We found that lipopolysaccharide-induced inducible nitric oxide synthase, and cyclooxygenase-2 expression, and nitric oxide formation was suppressed by NCA in a dose-dependent manner in BV2 microglia. NCA also inhibited the production of pro-inflammatory mediators, tumor necrosis factor-α and interleukin-1 beta. Furthermore, phosphorylated nuclear factor-kappa B p65 and p38 mitogen-activated protein kinase activation were blocked by NCA. Taken together, these results suggest that NCA exerts neuroprotective effects through the inhibition of pro-inflammatory pathways in activated microglia.