Sang-In Kim

Decursin Inhibits Induction of Inflammatory Mediators by Blocking Nuclear Factor-κB Activation in Macrophages

In the course of screening inhibitors of matrix metalloproteinase (MMP)-9 induction in macrophages, we isolated decursin, a coumarin compound, from the roots of Angelicae gigas. As a marker for the screening and isolation, we tested expression of MMP-9 in RAW264.7 cells and THP-1 cells after treatment with bacterial lipopolysaccharide (LPS), the TLR-4 ligand. Decursin suppressed MMP-9 expression in cells stimulated by LPS in a dose-dependent manner at concentrations below 60 μM with no sign of cytotoxicity. The suppressive effect of decursin was observed not only in cells stimulated with ligands for TLR4, TLR2, TLR3, and TLR9 but also in cells stimulated with interleukin (IL)-1β, and tumor necrosis factor (TNF)-α, indicating that the molecular target of decursin is common signaling molecules induced by these stimulants. In addition to the suppression of MMP-9 expression, decursin blocked nitric oxide production and cytokine (IL-8, MCP-1, IL-1β, and TNF-α) secretion induced by LPS. To find out the molecular mechanism responsible for the suppressive effect of decursin, we analyzed signaling molecules involved in the TLR-mediated activation of MMP-9 and cytokines. Decursin blocked phosphorylation of IκB and nuclear translocation of NF-κB in THP-1 cells activated with LPS. Furthermore, expression of a luciferase reporter gene under the promoter containing NF-κB binding sites was blocked by decursin. These data indicate that decursin is a novel inhibitor of NF-κB activation in signaling induced by TLR ligands and cytokines.

Decursin inhibits induction of inflammatory mediators by blocking NF-κB activation in macrophages

In the course of screening inhibitors of matrix metalloproteinase (MMP)-9 induction in macrophages, we isolated decursin, a coumarin compound, from the roots of Angelicae gigas. As a marker for the screening and isolation, we tested expression of MMP-9 in RAW264.7 cells and THP-1 cells after treatment with bacterial lipopolysaccharide (LPS), the TLR-4 ligand. Decursin suppressed MMP-9 expression in cells stimulated by LPS in a dose-dependent manner at concentrations below 60 μM with no sign of cytotoxicity. The suppressive effect of decursin was observed not only in cells stimulated with ligands for TLR4, TLR2, TLR3, and TLR9 but also in cells stimulated with interleukin (IL)-1β, and tumor necrosis factor (TNF)-α, indicating that the molecular target of decursin is common signaling molecules induced by these stimulants. In addition to the suppression of MMP-9 expression, decursin blocked nitric oxide production and cytokine (IL-8, MCP-1, IL-1β, and TNF-α) secretion induced by LPS. To find out the molecular mechanism responsible for the suppressive effect of decursin, we analyzed signaling molecules involved in the TLR-mediated activation of MMP-9 and cytokines. Decursin blocked phosphorylation of IκB and nuclear translocation of NF-κB in THP-1 cells activated with LPS. Furthermore, expression of a luciferase reporter gene under the promoter containing NF-κB binding sites was blocked by decursin. These data indicate that decursin is a novel inhibitor of NF-κB activation in signaling induced by TLR ligands and cytokines.

Syringin from stem bark of Fraxinus rhynchophylla protects Aβ(25–35)-induced toxicity in neuronal cells

The medicinal herb Jinpi, derived from the dried stem barks of Fraxinus rhynchophylla belonging to Oleaceae is widely used as a variety of Korean folk remedies for anti-inflammatory, febricide, antidiarrhea, and antileukorrhea diseases. In the course of screening antidementia agents from natural products, F. rhynchophylla showed significant inhibitory activity toward Aβ(25–35)-induced neuronal cell death. An active principle was isolated and identified as syringin. When the neuroblastoma cells were exposed to 50 μM Aβ(25–35), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction rate (survival rate) decreased to 60.21 ± 2.16% over control while syringin treated ones recovered cell viability up to 79.12 ± 1.39% at 20 μM. In addition, 20 μM syringin almost completely removed Aβ(25–35)-induced reactive oxygen species. The neuroprotective effect of syringin seemed to be originated from the reduction of apoptosis since decrease in caspase-3 activity and expression, reduction in cleaved PARP, and DNA fragmentation were observed. These results suggest that F. rhynchophylla and syringin are expected to be useful for preventing Aβ(25–35)-induced neuronal cell damage.

Prolyl endopeptidase inhibitors from green tea.

Three prolyl endopeptidase (PEP) inhibitors were isolated from the methanolic extract of green tea leaves. They were identified as (-)-epigallocatechin gallate, (-)-epicatechin gallate, and (+)-gallocatechin gallate with the IC50 values of 1.42 × 10-4 mM, 1.02 × 10-2 mM, and 1.09 × 10-4 mM, respectively. They were non-competitive with a substrate in Dixon plots and did not show any significant effects against other serine proteases such as elastase, trypsin, and chymotrypsin, suggesting that they were relatively specific inhibitors against PEP. The isolated compounds are expected to be useful for preventing and curing of Alzheimer’s disease.

A prolyl endopeptidase-inhibiting antioxidant from Phyllanthus ussurensis.

A prolyl endopeptidase inhibitor was isolated from the ethyl acetate soluble fraction ofPhyllanthus ussurensis. The active compound was identified as an ellagitannin, corilagin. It was shown to non-competitively inhibit prolyl endopeptidase (PEP) with the IC50 value of 1.17x10-6 μM. TheKi value was 6.70x10-7 M. Corilagin was less inhibitory to other serine proteases such as chymotrypsin, trypsin, and elastase, indicating that it was relatively a specific inhibitor of PEP. Corilagin also effectively inhibited reactive oxygen species such as hydroxide and superoxide anion radical, hydrogen peroxide, and DPPH. Especially, corilagin showed potent scavenging activity on the superoxide anion radical in the ESR method (IC50 = 3.79x10-6 M) as well as xanthine oxidase system.

β-Secretase (BACE1)-inhibiting C-methylrotenoids from Abronia nana suspension cultures.

Suspension cultures of Abronia nana were established to produce C-methylisoflavones. A new C-methylrotenoid, named abronione A (2), was isolated along with three known rotenoids, boeravinone D (1), boeravinone A methyl ether (3), and mirabijalone D (4). The IC50 values of compounds 1, 2, and 4 on β-secretase (BACE1) were 4.77, 62.21, and 4.24 μM, respectively, whereas 3 was inactive. At concentrations up to 1.0 mM, the compounds did not inhibit other proteases such as trypsin, chymotrypsin, and elastase, indicating that they were specific inhibitors of β-secretase. Compounds 1 and 4 were non-competitive inhibitors based on the Dixon plot and with Ki values of 5.01 and 4.28 μM, respectively. At 50 μM, compound 4 inhibited Aβ1-42 production by 43.7% in APPSW-N2a cells.