Sungsoo Kang

A new class of COX-2 inhibitor, rutaecarpine from Evodia rutaecarpa.

Abstract.Objective and Design: We investigated the effect of a new class of COX-2 inhibitor, rutaecarpine, on the production of PGD2 in bone marrow derived mast cells (BMMC) and PGE2 in COX-2 transfected HEK293 cells. Inflammation was induced by γ-carrageenan in male Splague-Dawley (SD) rats.¶Material: Rutaecarpine (8,13-Dihydroindolo[2,3:3,4]pyridol[2,1-b]quinazolin-5(7H)-one) was isolated from the fruits of Evodia rutaecarpa. BMMC were cultured with WEHI-3 conditioned medium. c-Kit ligand and IL-10 were obtained by their expression in baculovirus.¶Methods: The generation of PGD2 and PGE2 were determined by their assay kit. COX-1 and COX-2 protein and mRNA expression was determined by BMMC in the presence of KL, LPS and IL-10.¶Treatment: Rutaecarpine and indomethacin dissolved in 0.1% carboxymethyl cellulose was administered intraperitoneally and, 1h later, γ-carrageenan solution was injected to right hind paw of rats. Paw volumes were measured using plethysmometer 5h after γ-carrageenan injection.¶Results: Rutaecarpine inhibited COX-2 and COX-1 dependent phases of PGD2 generation in BMMC in a concentration-dependent manner with an IC50 of 0.28 μM and 8.7 μM, respectively. It inhibited COX-2-dependent conversion of exogenous arachidonic acid to PGE2 in a dose-dependent manner by the COX-2-transfected HEK293 cells. However, rutaecarpine inhibited neither PLA2 and COX-1 activity nor COX-2 protein and mRNA expression up to the concentration of 30 μM in BMMC, indicating that rutaecarpine directly inhibited COX-2 activity. Furthermore, rutaecarpine showed in vivo anti-inflammatory activity on rat γ-carrageenan induced paw edema by intraperitoneal administration.¶Conclusion: Anti-inflammatory activity of Evodia rutaecarpa could be attributed at least in part by inhibition of COS-2.

A genome-wide microarray analysis reveals anti-inflammatory target genes of paeonol in macrophages

Abstract.Objective:Paeony root has long been used for its anti-inflammatory effects. In this study, the effects of albiflorin, paeoniflorin, and paeonol, compounds from paeony root, on gene expression profiles were examined in macrophages challenged with the inflammation inducer lipopolysaccharide (LPS).Methods:The RAW264.7 macrophages were treated with LPS in the presence or absence of albiflorin, paeoniflorin, or paeonol. Global mRNA expression levels were detected by using an oligonucleotide microarray platform covering the mouse whole genome.Results:Treatment with LPS caused expression level changes in 1,270 genes by 2 folds or more. Paeonol attenuated the induction level of 355 LPS-responsive genes. Classification of the genes targeted by paeonol according to the Panther group analysis revealed 20 biological processes, 24 molecular functions, and 22 signaling pathways. The Panther signaling pathways highly affected by paeonol included the ‘inflammation mediated by chemokine and cytokine signaling’, ‘interleukin signaling’, and ‘Toll receptor signaling’.Conclusion:Our results demonstrate that paeonol has extensive inhibitory effects on the regulation of inflammation associated gene expression by LPS in macrophages. In addition, the predominant effect of paeonol among the tested compounds suggests that paeonol may be a major ingredient for the anti-inflammatory effect of paeony root.

Antiinflammatory activity of Lonicera japonica

As part of our investigations into new antiinflammatory agents based on plant extracts, the n‐butanol (BuOH) fraction of Lonicera japonica was prepared and its antiinflammatory activity was evaluated using several experimental animal models of inflammation. At oral doses of 100–400u2005mg/kg, the BuOH fraction showed antiinflammatory activity against acute, granulomatic and chronic inflammation models in mice and rats. Although the activity was not potent compared with prednisolone, the results support the traditional use and suggest that this fraction of L. japonica may yield a safe and mild antiinflammatory agent for treating various inflammatory disorders.

Real-time analysis of amyloid fibril formation of α-synuclein using a fibrillation-state-specific fluorescent probe of JC-1

alpha-Synuclein is a pathological component of PD (Parkinsons disease) by participating in Lewy body formation. JC-1 (5,5,6,6-tetrachloro-1,1,3,3-tetraethylbenzimidazolyl carbocyanine iodide) has been shown to interact with alpha-synuclein at the acidic C-terminal region with a K(d) of 2.6 microM. JC-1 can discriminated between the fibrillation states of alpha-synuclein (monomeric, oligomeric intermediate and fibrillar forms) by emitting the enhanced binding fluorescence of different colours at 590, 560 and 538 nm respectively with the common excitation at 490 nm. The fibrillation-state-specific interaction of JC-1 allowed us to perform real-time analyses of the alpha-synuclein fibrillation in the presence of iron as a fibrillation inducer, rifampicin as a fibrillation inhibitor, baicalein as a defibrillation agent and dequalinium as a protofibril inducer. In addition, various alpha-synuclein fibrils with different morphologies prepared with specific ligands such as metal ions, glutathione, eosin and lipids were monitored with their characteristic JC-1-binding fluorescence spectra. FRET (fluorescence resonance energy transfer) between thioflavin-T and JC-1 was also employed to specifically identify the amyloid fibrils of alpha-synuclein. Taken together, we have introduced JC-1 as a powerful and versatile probe to explore the molecular mechanism of the fibrillation process of alpha-synuclein in vitro. It could be also useful in high-throughput drug screening. The specific alpha-synuclein interaction of JC-1 would therefore contribute to our complete understanding of the molecular aetiology of PD and eventual development of diagnostic/therapeutic strategies for various alpha-synucleinopathies.

Papyriflavonol A, a new prenylated flavonol from Broussonetia papyrifera

A new prenylated flavonol, papyriflavonol A, was isolated from the root barks of Broussonetia papyrifera. The structure of this compound was elucidated as 5,7,3,4-tetrahydroxy-6,5-di-(gamma,gamma-dimethylallyl)-flavonol (1) by spectroscopic analysis.

Nanoporous protein matrix made of amyloid fibrils of β2-microglobulin.

Amyloid fibrils are considered as novel nanomaterials because of their nanoscale width, a regular constituting structure of cross β‐sheet conformation, and considerable mechanical strength. By using an amyloidogenic protein of β2‐microglobulin (β2M) related to dialysis‐related amyloidosis, nanoporous protein matrix has been prepared. The β2M granules made of around 15 monomers showed an average size of 23.1 nm. They formed worm‐like fibrils at pH 7.4 in 20 mM sodium phosphate containing 0.15 M NaCl following vigorous nondirectional shaking incubation, in which they became laterally associated and interwound to generate the porous amyloid fibrillar matrix with an average pore size of 30–50 nm. This nanoporous protein matrix was demonstrated to be selectively disintegrated by reducing agents, such as tris‐(2‐carboxyethyl) phosphine. High surface area with nanopores on the surface has been suggested to make the matrix of β2M amyloid fibrils particularly suitable for applications in the area of nanobiotechnology including drug delivery and tissue engineering.

Fibrillar superstructure formation of hemoglobin A and its conductive, photodynamic and photovoltaic effects

The fabrication of biomaterials which serve as functional scaffolds exhibiting diversified effects has been valued. We report here a unique strategy to fibrillate hemoglobin A (HbA), which exhibits multiple photoelectrochemical properties, and a subsequent specific defibrillation procedure. A subtle structural rearrangement of the α/β-subunits within the quaternary structure of HbA is responsible for the HbA fibril formation in the presence of 0.5% CHCl₃. The narrow pH dependence of the suprastructure formation around pH 7.4 illustrates the highly sensitive nature of the structural alteration. The CHCl₃-induced fibrils become disintegrated by ascorbic acid, indicating that the oxidation-reduction process of the iron within the heme moiety could be involved in stabilization of the fibrillar structures. The electron-transferring property of the iron allows the fibrils to exhibit not only their conductive behavior but also a photodynamic effect generating hydroxyl radicals in the presence of H(2)O(2) with light illumination. A photovoltaic effect is also demonstrated with the HbA fibrils, which generate an electric current on the fibril-coated microelectrode upon irradiation at 405nm. Taken together, the multiple effects of HbA fibrils and the selective fibrillation/defibrillation procedures could qualify the fibrils to be employed for various future applications in biotechnology, including bio-machine interfaces.

Removal of intact β2-microglobulin at neutral ph by using seed-conjugated polymer beads prepared with β2-microglobulin-derived peptide (58–67)

Removal of β2‐microglobulin (β2M) from the blood of patients suffering from kidney dysfunction is crucial to protect those individuals from getting the diseased state of dialysis‐related amyloidosis. By harnessing the nucleation‐dependent fibrillation process of amyloidogenesis, a β2M removal strategy has been proposed by preparing seed‐conjugated polymer beads and assimilating soluble β2M to the fibrils on the surface at neutral pH. A novel peptide segment of β2M ranging from residue 58 to residue 67 (Lys‐Asp‐Trp‐Ser‐Phe‐Tyr‐Leu‐Leu‐Tyr‐Tyr), which was capable of being fibrillated at neutral pH was isolated. Charge interaction between the positive N‐terminal lysine and the negative C‐terminal α‐carboxylic group was demonstrated to be critical for the molecular self‐assembly leading to the peptide fibril formation by favoring β‐sheet conformation. Because the peptide fibrils were successful to seed intact β2M at neutral pH, the fibrils were immobilized on polymer beads of HiCore resins, and the resulting seed‐conjugated beads were used to accrete intact β2M in the form of fibrils elongated on the bead surface. Its efficiency of the β2M removal was improved by placing the seed‐immobilized beads in the middle of a continuous flow of the β2M‐containing solution as practiced in the blood circulation during the hemodialysis. Therefore, this β2M removal system is suggested to exhibit high specificity, high binding capacity, and cost‐effectiveness appropriate for eventual clinical application to remove β2M from the blood of renal failure patients.

A new diterpenoid alkaloid from Aconitum jaluense

A new diterpenoid alkaloid, jaluenine (1), has been isolated from the roots of Aconitum jaluense. The structure of jaluenine was determined by spectroscopic methods including two dimensional NMR (1H-1H COSY, HMQC, HMBC, NOESY).