Eunjung Moon

Quinic acid derivatives from Pimpinella brachycarpa exert anti-neuroinflammatory activity in lipopolysaccharide-induced microglia.

Five new quinic acid derivatives (1-5), together with 10 known quinic acid derivatives (6-15), were isolated from the MeOH extract of Pimpinella brachycarpa (Umbelliferae). Their structures were established on the basis of spectroscopic analyses including extensive 2D NMR studies (COSY, HMQC and HMBC). Isolated compounds 1-15 were evaluated for their inhibitory activities on nitric oxide (NO) production in an activated murine microglial cell line. Compounds 2, 3, 8 and 11 significantly inhibited NO production without high cell toxicity in lipopolysaccharide (LPS)-activated BV-2 cells, a microglia cell line (IC50=4.66, 12.52, 9.04 and 12.11 μM, respectively).

4-Methylthio-butanyl derivatives from the seeds of Raphanus sativus and their biological evaluation on anti-inflammatory and antitumor activities

ETHNOPHARMACOLOGICAL RELEVANCE Raphanus sativus seeds (Brassicaceae) known as Raphani Semen have long been used as anti-cancer and/or anti-inflammatory agents in Korean traditional medicine. This study was designed to isolate the bioactive constituents from the seed extracts of Raphanus sativus and evaluate their anti-inflammatory and antitumor activities. MATERIAL AND METHODS Bioassay-guided fractionation and chemical investigation of a methanolic extract of the seeds of Raphanus sativus led to the isolation and identification of seven 4-methylthio-butanyl derivatives. Structural elucidation of the isolated compounds was carried out using 1D and 2D nuclear magnetic resonance (NMR) spectroscopy techniques ((1)H, (13)C, COSY, HMQC and HMBC experiments) and mass spectrometry. RESULTS The isolated compounds were characterized as in the following: three new 4-methylthio-butanyl derivatives, sinapoyl desulfoglucoraphenin (1), (E)-5-(methylsulfinyl)pent-4-enoxylimidic acid methyl ester (2), and (S)-5-((methylsulfinyl)methyl)pyrrolidine-2-thione (3), together with four known compounds, 5-(methylsulfinyl)-4-pentenenitrile (4), 5-(methylsulfinyl)-pentanenitrile (5), sulforaphene (6), and sulforaphane (7). Full NMR data assignments of the three known compounds 4-6 were also reported for the first time. We evaluated the anti-neuroinflammatory effect of 1-7 in lipopolysaccharide-stimulated murine microglia BV2 cells. Compound 1 significantly inhibited nitrite oxide production with IC50 values of 45.36 μM. Moreover, it also reduced the protein expression of inducible nitric oxide synthase. All isolates were also evaluated for their antiproliferative activities against four human tumor cell lines (A549, SK-OV-3, SK-MEL-2, and HCT-15), and all of them showed antiproliferative activity against the HCT-15 cell, with IC50 values of 8.49-23.97 μM. CONCLUSIONS 4-Methylthio-butanyl derivatives were one of the main compositions of Raphanus sativus seeds, and activities demonstrated by the isolated compounds support the ethnopharmacological use of Raphanus sativus seeds (Brassicaceae) as anti-cancer and/or anti-inflammatory agents.

Anti-neuroinflammatory diarylheptanoids from the rhizomes of Dioscorea nipponica

In a continuing search for bioactive constituents from Dioscoreaceae medicinal plants, two new cyclic diarylheptanoids, diosniponol A (1) and B (2), together with 10 known compounds (3-12) were isolated from the rhizomes of Dioscorea nipponica. The structures of these new compounds were determined by spectroscopic analyses, including extensive two-dimensional nuclear magnetic resonance, high-resolution mass spectrometry, and optical rotation. All isolated compounds 1-12 were evaluated for their effects on nitric oxide (NO) production in murine microglia cell line BV-2. Compounds 8 and 11 showed potent inhibitory activities on NO production (IC50 13.36 and 14.36 μM, respectively) without cell toxicity in lipopolysaccharide-activated BV-2 cells.

Matrix Metalloproteinase-8 is a Novel Pathogenetic Factor in Focal Cerebral Ischemia

The neutrophil collagenase matrix metalloproteinase-8 (MMP8) is a recently identified member of MMPs that have important roles in various inflammation-related disorders. Previously, we identified MMP8 as a new neuroinflammatory mediator in activated microglia by regulating TNF-α productivity. Here, we present evidence that MMP8 is a critical factor for brain damage in transient focal cerebral ischemia by modulating neuroinflammation likely microglial activation and TNF-α production. Biochemical analyses showed upregulation of MMP8 expression at mRNA and protein levels in transient middle cerebral artery occlusion/reperfusion (M/R)-challenged brains. Furthermore, double immunolabeling showed that MMP8 expression was upregulated in the activated microglia of M/R-challenged brains. Assessment of infarct volume, neurological score, and survival/death of neural cells revealed that administration of an MMP8 inhibitor (M8I) immediately after reperfusion reduced brain damage. Histological analyses showed that microglial activation and TNF-α expression in ischemic conditions was abrogated by exposure to M8I, as demonstrated in our previous study using cultured microglia. These outcomes from a pharmacological approach were reaffirmed by a genetic approach using a lentiviral system. Intracerebroventricular microinjection of MMP8-specific shRNA lentivirus reduced the extent of ischemia-induced brain damage, as assessed by infarct volume, neurological score, microglial activation, and TNF-α expression. These results suggest a novel pathogenetic role of MMP8 and implicate modulation of its activity as a tractable strategy for therapies against cerebral ischemia.

Dihydrolipoyl dehydrogenase as a potential UVB target in skin epidermis; using an integrated approach of label-free quantitative proteomics and targeted metabolite analysis

UNLABELLED Photodamage is extrinsically induced by overexposure to ultraviolet (UV) radiation, and it increases the risk of various skin disorders. Therefore, discovery of novel biomarkers of photodamage is important. In this study, using LC-MS/MS analysis of epidermis from UVB-irradiated hairless mice, we identified 57 proteins whose levels changed after UVB exposure, and selected 7 proteins related to the tricarboxylic acid (TCA) cycle through pathway analysis. Dihydrolipoyl dehydrogenase (DLD) was the only TCA cycle-associated protein that showed a decreased expression after the UVB exposure. We also performed targeted analysis to detect intermediates and products of the TCA cycle using GC-TOF-MS. Interestingly, malic acid and fumaric acid levels significantly decreased in the UVB-treated group. Our results demonstrate that DLD and its associated metabolites, malic acid and fumaric acid, may be candidate biomarkers of UVB-induced skin photoaging. Additionally, we showed that Aloe vera, a natural skin moisturizer, regulated DLD, malic acid and fumaric acid levels in UVB-exposed epidermis. Our strategy to integrate the proteome and targeted metabolite to detect novel UVB targets will lead to a better understanding of skin photoaging and photodamage. Our study also supports that A. vera exerts significant anti-photodamage activity via regulation of DLD, a novel UVB target, in the epidermis. BIOLOGICAL SIGNIFICANCE This study is the first example of an integration of proteomic and metabolite analysis techniques to find new biomarker candidates for the regulation of the UVB-induced skin photoaging. DLD, malic acid, and fumaric acid can be used for development of cosmeceuticals and nutraceuticals regulating the change of skin metabolism induced by the UVB overexposure. Moreover, this is also the first attempt to investigate the role of the TCA cycle in photodamaged epidermis. Our integration of the proteomic and targeted metabolite analyses will lead to a better understanding of the unidentified photobiological results from UVB-irradiated models and can elicit new diagnostic and treatment strategies based on altered metabolism.

Exogenous S1P Exposure Potentiates Ischemic Stroke Damage That Is Reduced Possibly by Inhibiting S1P Receptor Signaling

Initial and recurrent stroke produces central nervous system (CNS) damage, involving neuroinflammation. Receptor-mediated S1P signaling can influence neuroinflammation and has been implicated in cerebral ischemia through effects on the immune system. However, S1P-mediated events also occur within the brain itself where its roles during stroke have been less well studied. Here we investigated the involvement of S1P signaling in initial and recurrent stroke by using a transient middle cerebral artery occlusion/reperfusion (M/R) model combined with analyses of S1P signaling. Gene expression for S1P receptors and involved enzymes was altered during M/R, supporting changes in S1P signaling. Direct S1P microinjection into the normal CNS induced neuroglial activation, implicating S1P-initiated neuroinflammatory responses that resembled CNS changes seen during initial M/R challenge. Moreover, S1P microinjection combined with M/R potentiated brain damage, approximating a model for recurrent stroke dependent on S1P and suggesting that reduction in S1P signaling could ameliorate stroke damage. Delivery of FTY720 that removes S1P signaling with chronic exposure reduced damage in both initial and S1P-potentiated M/R-challenged brain, while reducing stroke markers like TNF-α. These results implicate direct S1P CNS signaling in the etiology of initial and recurrent stroke that can be therapeutically accessed by S1P modulators acting within the brain.

Dioscorea Extract (DA-9801) Modulates Markers of Peripheral Neuropathy in Type 2 Diabetic db/db Mice

The purpose of this study was to investigate the therapeutic effects of DA-9801, an optimized extract of Dioscorea species, on diabetic peripheral neuropathy in a type 2 diabetic animal model. In this study, db/db mice were treated with DA-9801 (30 and 100 mg/kg, daily, p.o.) for 12 weeks. DA-9801 reduced the blood glucose levels and increased the withdrawal latencies in hot plate tests. Moreover, it prevented nerve damage based on increased nerve conduction velocity and ultrastructural changes. Decrease of nerve growth factor (NGF) may have a detrimental effect on diabetic neuropathy. We previously reported NGF regulatory properties of the Dioscorea genus. In this study, DA-9801 induced NGF production in rat primary astrocytes. In addition, it increased NGF levels in the sciatic nerve and the plasma of type 2 diabetic animals. DA-9801 also increased neurite outgrowth and mRNA expression of Tieg1/Klf10, an NGF target gene, in PC12 cells. These results demonstrated the attenuation of diabetic peripheral neuropathy by oral treatment with DA-9801 via NGF regulation. DA-9801 is currently being evaluated in a phase II clinical study.

Alkaloids from Acorus gramineus Rhizomes and their Biological Activity

As part of our ongoing search for bioactive constituents from natural Korean medicinal resources, a bioassay-guided fractionation and a chemical investigation of the methanolic extract from the rhizomes of Acorus gramineus resulted in the isolation and identification of two alkaloids, including a new aporphine-type alkaloid, named gramichunosin, and a known pyrrole alkaloid. Their structures were determined by a combination of 1D and 2D nuclear magnetic resonance (NMR) spectroscopic analysis and high resolution mass spectrometry (HRMS). This is the first report of alkaloids from A. gramineus. The compounds isolated showed antiproliferative activities against A549, SK-OV-3, SK-MEL-2 and HCT-15 cell lines with EC50 values in the range of 7.46-45.23 μM. Moreover, the anti-neuroinflammatory activities of these compounds were determined by measuring the nitric oxide (NO) levels in the medium using murine microglia BV-2 cells. Compound named gramichunosin inhibited NO production in lipopolysaccharide-stimulated BV-2 with IC50 values of 7.83 μM.

Sarsasapogenin Increases Melanin Synthesis via Induction of Tyrosinase and Microphthalmia-Associated Transcription Factor Expression in Melan-a Cells

Sarsasapogenin (SAR) is a steroidal sapogenin that is used as starting material for the industrial synthesis of steroids. It has various pharmacological benefits, such as antitumor and antidepressant activities. Since its effect on melanin biosynthesis has not been reported, we used murine melanocyte melan-a cells to investigate whether SAR influences melanogenesis. In this study, SAR significantly increased the melanin content of the melan-a cells from 1 to 10 μM. Based on an enzymatic activity assay using melan-a cell lysate, SAR had no effect on tyrosinase and DOPAchrome tautomerase activities. It also did not affect the protein expression of tyrosinase-related protein 1 and DOPAchrome tautomerase. However, protein levels of tyrosinase and microphthalmia-associated transcription factor were strongly stimulated by treatment with SAR. Therefore, our reports suggest that SAR treatment may induce melanogenesis through the stimulation of tyrosinase and microphthalmia-associated transcription factor expression in melan-a cells.

Bioactive sesquiterpenes from the essential oil of Thuja orientalis.

A phytochemical investigation on the essential oil of Thuja orientalis resulted in the isolation and identification of three new sesquiterpenes, 3α-methoxy-4α-epoxythujopsane (1), Δ³,¹⁵-4β-epoxythujopsene (2), and Δ³,⁴-thujopsen-2,15-diol (3), together with eight known sesquiterpenoids (4-11). The structures of these new compounds were elucidated based on spectroscopic data analyses including extensive 2D-NMR data and HR-ESIMS. The full assignments of ¹H and ¹³C NMR chemical shifts for thujopsadiene (4) were obtained by 2D-NMR for the first time. All compounds (1-11) showed antiproliferative activities against the SK-OV-3 and SK-MEL-2 cell lines with IC₅₀ values of 5.85-28.64 µM. In addition, compounds 1, 3, 4, 7, 8, and 9 significantly inhibited nitric oxide production in lipopolysaccharide-activated BV-2 cells with IC₅₀ values of 3.93-17.85 µM without cell toxicity.