Sunghyouk Park

Glycyrrhizic acid affords robust neuroprotection in the postischemic brain via anti-inflammatory effect by inhibiting HMGB1 phosphorylation and secretion

High mobility group box 1 (HMGB1) is an endogenous danger signal molecule. In a previous report, we showed that HMGB1 is massively released during NMDA-induced acute damaging process in the postischemic brain and triggers inflammatory processes, like microglial activation. siRNA-mediated HMGB1 knockdown markedly reduced infarct volumes, confirming the crucial role played by HMGB1 in the postischemic brain. In the present study, we showed neuroprotective effects of glycyrrhizin (GL) in the postischemic rat brain after middle cerebral artery occlusion (MCAO). GL, a triterpene present in the roots and rhizomes of licorice, Glycyrrhiza glabra, has been shown to have anti-inflammatory and anti-viral effects. It has been reported that GL binds directly to HMGB1, and inhibits its chemoattractant and mitogenic activities. The administration of GL (10mg/kg) intravenously at 3 or 6h after MCAO reduced infarct volumes to 12.9±4.2% and 46.2±9.9%, respectively, of untreated control. This neuroprotective effect was accompanied by improvements in motor impairment and neurological deficits and suppressions of microglia activation and proinflammatory cytokine induction. Interestingly, GL almost completely blocked HMGB1 secretion in the postischemic brain and in lipopolysaccharide (LPS)-treated microglia cells. Furthermore, HMGB1 phosphorylation, which is the initial step for HMGB1 secretion, and the interaction between HMGB1 and protein kinase C (PKC) or calcium/calmodulin-dependent protein kinase IV (CaMKIV) were suppressed dose-dependently by GL. Here, we hypothesized that the blockage for the putative phosphorylation sites in HMGB1 by GL might be attributed to this suppression. In addition to the anti-inflammatory effects, we found that GL has anti-excitotoxic and anti-oxidative effects in neurons. Together these results indicate that GL has neuroprotective efficacy in the postischemic brain via its anti-inflammatory, anti-excitotoxic, and anti-oxidative effects and in particular, it exerts anti-inflammatory effect, at least in part, by inhibiting HMGB1 secretion.

A new NMR-based metabolomics approach for the diagnosis of biliary tract cancer.

BACKGROUND & AIMS Biliary tract cancer is highly lethal at presentation, with increasing mortality worldwide. Current diagnostic measures employing multiple criteria such as imaging, cytology, and serum tumor markers are not satisfactory, and a new diagnostic tool is needed. Because bile is a cognate metabolite-rich bio-fluid in the biliary ductal system, we tested a new metabolomic approach to develop an effective diagnostic tool. METHODS Biles were collected prospectively from patients with cancer (n=17) or benign biliary tract diseases (n=21) with percutaneous or endoscopic methods. Nuclear magnetic resonance spectra (NMR) of these biles were analyzed using orthogonal partial least square discriminant analysis (OPLS-DA). RESULTS The metabolomic 2-D score plot showed good separation between cancer and benign groups. The contributing NMR signals were analyzed using a statistical TOCSY approach. The diagnostic performance assessed by leave-one-out analysis exhibited 88% sensitivity and 81% specificity, better than the conventional markers (CEA, CA19-9, and bile cytology). CONCLUSION The NMR-based metabolomics approach provides good performance in discriminating cancer and benign biliary duct diseases. The excellent predictability of the method suggests that it can, at least, augment the currently available diagnostic approaches.

Application of a 1H nuclear magnetic resonance (NMR) metabolomics approach combined with orthogonal projections to latent structure-discriminant analysis as an efficient tool for discriminating between Korean and Chinese herbal medicines

Correct identification of the origins of herbal medical products is becoming increasingly important in tandem with the growing interest in alternative medicine. However, visual inspection of raw material is still the most widely used method, and newer scientific approaches are needed. To develop a more objective and efficient tool for discriminating herbal origins, particularly Korean and Chinese, we employed a nuclear magnetic resonance (NMR)-based metabolomics approach combined with an orthogonal projections to latent structure-discriminant analysis (OPLS-DA) multivariate analysis. We first analyzed the constituent metabolites of Scutellaria baicalensis through NMR studies. Subsequent holistic data analysis with OPLS-DA yielded a statistical model that could cleanly discriminate between the sample groups even in the presence of large structured noise. An analysis of the statistical total correlation spectroscopy (STOCSY) spectrum identified citric acid and arginine as the key discriminating metabolites for Korean and Chinese samples. As a validation of the discrimination model, we performed blind prediction tests of sample origins using an external test set. Our model correctly predicted the origins of all of the 11 test samples, demonstrating its robustness. We tested the wider applicability of the developed method with three additional herbal medicines from Korea and China and obtained very high prediction accuracy. The solid discriminatory power and statistical validity of our method suggest its general applicability for determining the origins of herbal medicines.

NMR-Based Metabolomics Approach for the Differentiation of Ginseng (Panax ginseng) Roots from Different Origins

Agro-herbal materials vary in prices and qualities depending on the origin and age and the differentiation is both scientific and public health issue. Here, we describe a metabolomics approach used to discriminate ginseng roots from different sources. Six different types of ginseng roots from China and Korea were analyzed by NMR-based metabolomics. Chinese Dangsam showed prominent differences and was easily differentiated. The difference was mainly due to the large signals in the sugar region. We further analyzed the metabolomics results in subgroups. Jeonra (Korean), Choongcheong (Korean), and Chinese ginseng in subgroup 1 could be easily differentiated by the first two principal components. The loading plot for PC1 showed that the Jeonra and Chinese ginseng roots were mainly separated by sugar signals and methyl signals but that they were reverse-correlated. A diffusion-ordered spectroscopy (DOSY) analysis showed that the methyl signals are from high molecular weight compounds and that the sugar signals are either from oligosaccharides or ginsenosides. In subgroup 2, composed of Korean Choongcheong ginseng at different ages, we were able to see age-dependent transitions in the score plot. We believe our approach can be applied to detecting the adulteration of ginseng root powders and other herbal products from different origins.

Dynamic interaction of CD2 with the GYF and the SH3 domain of compartmentalized effector molecules

Intracellular protein interaction domains are essential for eukaryotic signaling. In T cells, the CD2BP2 adaptor binds two membrane‐proximal proline‐rich motifs in the CD2 cytoplasmic tail via its GYF domain, thereby regulating interleukin‐2 production. Here we present the structure of the GYF domain in complex with a CD2 tail peptide. Unlike SH3 domains, which use two surface pockets to accommodate proline residues of ligands, the GYF domain employs phylogenetically conserved hydrophobic residues to create a single interaction surface. NMR analysis shows that the Fyn but not the Lck tyrosine kinase SH3 domain competes with CD2BP2 GYF‐domain binding to the same CD2 proline‐rich sequence in vitro. To test the in vivo significance of this competition, we used co‐immunoprecipitation experiments and found that CD2BP2 is the ligand of the membrane‐proximal proline‐rich tandem repeat of CD2 in detergent‐ soluble membrane compartments, but is replaced by Fyn SH3 after CD2 is translocated into lipid rafts upon CD2 ectodomain clustering. This unveils the mechanism of a switch of CD2 function due to an extracellular mitogenic signal.

An HR-MAS MR Metabolomics Study on Breast Tissues Obtained with Core Needle Biopsy

Background Much research has been devoted to the development of new breast cancer diagnostic measures, including those involving high-resolution magic angle spinning (HR-MAS) magnetic resonance (MR) spectroscopic techniques. Previous HR-MAS MR results have been obtained from post-surgery samples, which limits their direct clinical applicability. Methodology/Principal Findings In the present study, we performed HR-MAS MR spectroscopic studies on 31 breast tissue samples (13 cancer and 18 non-cancer) obtained by percutaneous core needle biopsy. We showed that cancer and non-cancer samples can be discriminated very well with Orthogonal Projections to Latent Structure-Discriminant Analysis (OPLS-DA) multivariate model on the MR spectra. A subsequent blind test showed 69% sensitivity and 94% specificity in the prediction of the cancer status. A spectral analysis showed that in cancer cells, taurine- and choline-containing compounds are elevated. Our approach, additionally, could predict the progesterone receptor statuses of the cancer patients. Conclusions/Significance HR-MAS MR metabolomics on intact breast tissues obtained by core needle biopsy may have a potential to be used as a complement to the current diagnostic and prognostic measures for breast cancers.

Gallbladder carcinoma and chronic cholecystitis:differentiation with two-phase spiral CT

The objective of the present study was to determine whether an analysis of two-phase spiral computed tomographic (CT) features provides a sound basis for the differential diagnosis between gallbladder carcinoma and chronic cholecystitis. Eighty-two patients, 35 with gallbladder carcinoma and 47 with chronic cholecystitis, underwent two-phase spiral CT. We reviewed the two-phase spiral CT features of thickness and enhancement pattern of the gallbladder wall seen during the arterial and venous phases. Mean wall thicknesses were 12.6 mm in the gallbladder carcinoma group and 6.9 mm in the chronic cholecystitis group. The common enhancement patterns seen in gallbladder carcinoma were (a) a highly enhanced thick inner wall layer during the arterial phase that showed isoattenuation with the adjacent hepatic parenchyma during the venous phase (16 of 35, 45.7%) and (b) a highly enhanced thick inner wall layer during both phases (eight of 35, 22.9%). The most common enhancement pattern of chronic cholecystitis was isoattenuation of the thin inner wall layer during both phases (42 of 47, 89.4%). In conclusion, awareness of the wall thickening and enhancement patterns in gallbladder carcinoma and chronic cholecystitis on two-phase spiral CT appears to be valuable in differentiating these two different disease entities.

Sungsanpin, a Lasso Peptide from a Deep-Sea Streptomycete

Sungsanpin (1), a new 15-amino-acid peptide, was discovered from a Streptomyces species isolated from deep-sea sediment collected off Jeju Island, Korea. The planar structure of 1 was determined by 1D and 2D NMR spectroscopy, mass spectrometry, and UV spectroscopy. The absolute configurations of the stereocenters in this compound were assigned by derivatizations of the hydrolysate of 1 with Marfeys reagents and 2,3,4,6-tetra-O-acetyl-β-d-glucopyranosyl isothiocyanate, followed by LC-MS analysis. Careful analysis of the ROESY NMR spectrum and three-dimensional structure calculations revealed that sungsanpin possesses the features of a lasso peptide: eight amino acids (-Gly(1)-Phe-Gly-Ser-Lys-Pro-Ile-Asp(8)-) that form a cyclic peptide and seven amino acids (-Ser(9)-Phe-Gly-Leu-Ser-Trp-Leu(15)) that form a tail that loops through the ring. Sungsanpin is thus the first example of a lasso peptide isolated from a marine-derived microorganism. Sungsanpin displayed inhibitory activity in a cell invasion assay with the human lung cancer cell line A549.

Replicon size and excision repair as factors in the inhibition and recovery of DNA synthesis from ultraviolet damage.

Initiation of DNA replication and chain growth, analyzed by alkaline sucrose gradient sedimentation, was interrupted to different extents in different cell types by irradiation with ultraviolet light. Within the first hour of irradiation DNA replication was reduced in a manner that depended on the average number of lesions per replicating unit (replicon). At low numbers of lesions per replicon, inhibition of replicon initiation was the predominant response; at higher numbers of lesions per replicon, blockage of chain growth was also observed. After irradiation with a dose that initially blocked chain growth, the rate at which cells recovered their ability to synthesize increasingly more and larger size DNA was a function both of replicon size and of excision repair capacity. Cells with small replicons recovered more rapidly than cells with large replicons, and excision repair-deficient cells recovered less rapidly than excision-competent cells. These observations indicate that excision repair capacity and replicon size play major roles in the response of DNA replication to ultraviolet damage.

Predicting idiopathic toxicity of cisplatin by a pharmacometabonomic approach

Cisplatin has been one of the most widely used anticancer agents, but its nephrotoxicity remains a dose-limiting complication. Here, we evaluated the idiopathic nature and the predose prediction of cisplatin-induced nephrotoxicity using a nuclear magnetic resonance (NMR)-based pharmacometabonomic approach. Cisplatin produced serious toxic responses in some animals (toxic group), but had little effect in others (nontoxic group), as judged by hematological and histological results. The individual metabolic profiles, assessed by urine NMR spectra, showed large differences between the post-administration profiles of the two groups, indicating the relevance of the NMR approach. Importantly, multivariate analysis of the NMR data showed that the toxic and nontoxic groups can be differentiated based on the pretreatment metabolite profiles. Leave-one-out analysis, performed to evaluate the practical performance of our approach, gave a 66% accuracy rate in predicting toxic responses based on the pretreatment metabolite profiles. Hence, we provide a working model that can explain the idiopathic toxicity mechanism based on marker metabolites found by NMR analysis consistent with tissue NADH measurements. Thus, a pharmacometabonomic approach using pretreatment metabolite profiles may help expedite personalized chemotherapy of anticancer drugs.