Dan Hu

Fusarins G1/G2 (/) and G3/G4 (/), two sets of interesting examples of diastereoisomers with substantially identical NMR data, were discovered from natural products. The reason was discussed and the generally applicable determinant conditions were proposed. The minimum interval for stereoclusters to be entirely segregated was also discussed.

Fusicoccane diterpenoids usually possess a fused 5-8-5 tricyclic ring system, which are biogenetically generated from geranylgeranyl diphosphate (GGDP). In our report, three novel diterpenoid alkaloids with fusicoccane skeleton, pericolactines A-C (1-3), were isolated from Periconia sp.. Their structures with absolute configurations were determined by spectroscopic analyses and quantum chemical ECD calculation. Pericolactines A-C (1-3) are a new class of diterpenoid alkaloids with an unusual fused 5-5-8-5 tetracyclic ring system, which derive from a geranylgeranyl diphosphate (GGDP) and serine conjugated biosynthesis. They belong to the atypical diterpenoid alkaloids.

Bacterial pathogens can exploit metabolic pathways to facilitate their successful infection cycles, but little is known about roles of d-galactosamine (GalN)/N-acetyl-d-galactosamine (GalNAc) catabolism pathway in bacterial pathogenesis. Here, we report the genomic reconstruction of GalN/GalNAc utilization pathway in Streptococci and the diversified aga regulons. We delineated two new paralogous AgaR regulators for the GalN/GalNAc catabolism pathway. The electrophoretic mobility shift assays experiment demonstrated that AgaR2 (AgaR1) binds the predicted palindromes, and the combined in vivo data from reverse transcription quantitative polymerase chain reaction and RNA-seq suggested that AgaR2 (not AgaR1) can effectively repress the transcription of the target genes. Removal of agaR2 (not agaR1) from Streptococcus suis 05ZYH33 augments significantly the abilities of both adherence to Hep-2 cells and anti-phagocytosis against RAW264.7 macrophage. As anticipated, the dysfunction in Aga...

To explore the feasibility of bone marrow mesenchymal stem cells (MSCs) transdifferentiating into corneal epithelial cells in a limbal stem cell deficiency (LSCD) model in rats. Rat MSCs were isolated and purified using a gradient isolation procedure. The cells were induced by rat corneal stromal cells (CSCs) in a transwell co-culture system. The induced MSCs were identified by immunofluorescence staining, flow cytometry, and scanning electron microscopy (SEM). A corneal LSCD model was produced in the right eyes of 48 rats by alkali injury. The eyes of 12 rats without any transplant served as controls (Group 1). Amniotic membranes (AM; Group 2), uninduced MSCs (Group 3), or MSCs induced by CSCs (Group 4), were transplanted onto the cornea of the model (n=12 each). The therapeutic effects of the four groups were evaluated by slit lamp observation, hematoxylin and eosin staining, immunohistochemistry staining, and confocal laser corneal microscopy. Cultivated MSCs were positive for CD29, CD44, and CD90, but negative for CD34, CD45, CD133, and CK12, with typical MSCs characteristics revealed by SEM. After co-culture with CSCs, the induced MSCs expressed positive staining for CK12 with corneal epithelial cell characteristics confirmed by SEM; the induced MSCs were unchanged on the amnion. Compared with the other three groups, the corneal opacity, fluorescence staining, and neovascularization grades were significantly decreased in the induced MSCs group, both on postoperative week four and ten. MSCs induced by CSCs can transdifferentiate into corneal epithelial cells in vitro. The induced MSCs on an amniotic membrane have remarkable effects on the treatment of corneal alkali burn and the reconstruction of the corneal surface of rats.

Sporormiellin A (1), the first tetrahydrofuran-fused furochromone with an unprecedented tetracyclic skeleton, has been obtained from the fungal strain Sporormiella minima. The structure with absolute configuration was elucidated by NMR data, X-ray crystallography, and quantum chemical ECD calculations.

The fungus Nodulisporium sp. (No. 65-12-7-1), which can produce a rare class of steroids (4-methyl-progesteroids) in rice solid medium, was subjected to a one strain-many compounds (OSMAC) approach. It was found to produce ten new 4-methyl-progesteroid derivatives named nodulisporisteroids C-L (1-10) in potato-dextrose-broth (PDB) medium, which showed that the fungus was a prolific producer of 4-methyl-progesteroids. The structures of 1-10 were elucidated by spectroscopic and X-ray crystallographic analysis. Their cytotoxic activities against five human cancer cell lines had been evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method.

In the post genomic era, glycomics--the systematic study of all glycan structures of a given cell or organism--has emerged as an indispensable technology in various fields of biology and medicine. Lectins are regarded as "decipherers of glycans", being useful reagents for their structural analysis, and have been widely used in glycomic studies. However, the inconsistent activity and availability associated with the plant-derived lectins that comprise most of the commercially available lectins, and the limit in the range of glycan structures covered, have necessitated the development of innovative tools via engineering of lectins on existing scaffolds. This review will summarize the current state of the art of lectin engineering and highlight recent technological advances in this field. The key issues associated with the strategy of lectin engineering including selection of template lectin, construction of a mutagenesis library, and high-throughput screening methods are dis...

Three new cyclohexenones (1-3, named sarcosones A-C) and two new isocoumarins (4 and 5), together with five known isocoumarins (6-10), were isolated from the solid cultures of an endophytic fungus Sarcosomataceae sp. NO.49-14-2-1. Their chemical structures were elucidated by analyses of HR-ESI-TOF-MS, (1)H, (13)C NMR, (1)H-(1)H COSY, HSQC, and HMBC spectra. Their absolute configurations were determined via modified Mosher's method and circular dichroism spectra method.

Streptococcal histidine triad proteins HTPs are widely distributed within the Streptococcus genus. Based on the phylogenetic relationship and domain composition, HTPs are classified into type I and type II subfamilies. Previous studies revealed that several pathogenic streptococci contain more than one htp gene. We found that the highly virulent strain of Streptococcus suis 2 (S. suis 2), 05ZYH33 encodes three HTPs, designated HtpsA (previously described as HtpS), HtpsB, and HtpsC. Among them, HtpsC is the only member that contains leucine-rich repeat (LRR) domains at the C-terminal. In this study, we demonstrated that the recombinant HtpsC could bind to two different components of human ECM complex laminin and fibronectin in vitro, suggesting that it is a novel adhesin of S. suis 2. Having constructed an htpsC mutant, we evaluated its role in the pathogenesis of the highly virulent S. suis 2 strain 05ZYH33. Our data showed that inactivation of htpsC significantly affected adherence...

Stachybisbins A (1) and B (2), two new meroterpenoids with unprecedented seco-bisabosqual skeleton, together with three biogenetically related metabolites (3-5), were isolated from a wetland fungal strain of Stachybotrys bisbyi. The structures of the new compounds were determined by spectroscopic analyses, modified Mosher's method, and quantum chemical CD method. The cytotoxic activities of all compounds were tested against HL-60, SMMC-7721, A-549, MCF-7, and SW480 human cancer cell lines.

Pericoterpenoid A (1), a new cadinane-type sesquiterpene, was isolated from an endolichenic fungal strain Periconia sp. (No. 19-4-2-1). Its structure was characterized by analyzing the spectroscopic data (IR, MS, 1D- and 2D-NMR). The antimicrobial activity against Escherichia coli, Staphylococcus aureus, Aspergillus niger, and Candida albicans was evaluated. Pericoterpenoid A showed moderate antimicrobial activity against A. niger and weak activity against C. albicans. This is the first report of the presence of cadinane-type sesquiterpene in Periconia sp.

BmTXKβ, a scorpion toxin isolated from the Chinese scorpion Buthus martensii Karsch (BmK), was expressed as a GST fusion protein in BL21 (DE3) strain. The recombinant GST-BmTXKβ protein was purified by affinity chromatography. When treated with enterokinase, the GST-BmTXKβ fusion protein released an approximate 6.5kDa protein which was the expected size for correctly processed. About 2mg purified recombinant BmTXKβ protein

Streptococcus suis, an emerging infectious pathogen, is the cause of two large-scale outbreaks of human streptococcal toxic shock syndrome in China, and has attracted much attention from the scientific community. The genetic basis of its pathogenesis remains enigmatic, and no effective prevention measures have been established. To better understand the virulence differentiation of S. suis and develop a promising vaccine, we isolated and sequenced a native avirulent S. suis strain (05HAS68). Animal experiments revealed that 05HAS68 is an avirulent strain and could protect piglets from the attack of virulent strains. Comparative genomics analyses demonstrated the genetic basis for the lack of virulence in 05HAS68, which is characterized by the absence of some important virulence-associated factors and the intact 89K pathogenicity island. Lack of virulence was also illustrated by reduced survival of 05HAS68 compared to a virulent strain in pig whole blood. Further investigations reveal...

Among sulphated glycans, little is known about 3'-sulphation because of the lack of useful probes. In the course of molecular engineering of a fungal galectin from Agrocybe cylindracea, we found that a single substitution of Glu86 with alanine resulted in acquisition of specific binding for the 3'-sulpho-Galβ1-4GlcNAc structure. Extensive glyco-technological analysis revealed that this property was obtained in a 'loss-of-function' manner. Though this mutant (E86A) had low total affinity, it showed substantial binding to a naturally occurring N-glycan, of which the terminal galactose is 3-sulphated. Moreover, E86A specifically bound to HeLa cells, in which galactose-3-O-sulfotransferases (Gal3ST2 or Gal3ST3) were over-expressed.

To observe the effect of mitomycin C (MMC) on cultured human nasal mucosa fibroblasts. Cultured human nasal mucosa fibroblasts were exposed to MMC (0.1-0.4 mg/ml) for 1 to 5 minutes. The viability of the fibroblasts was determined by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay; DNA fragmentation (apoptosis) by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL); apoptotic percentage by flow cytometry; and morphology by light microscopy. A portion of the fibroblasts survived the mitomycin treatment and showed evidence of regrowth within 2 to 3 days. These cells reached confluence in 5 to 7 days. The inhibition rates by MTT assay of 0.4 mg/ml MMC for 5-minute exposures was 31.3%. Dose-response effect was noted with the lower concentrations and shorter exposure times where the inhibition rates were lower (but not significantly so). DNA fragmentation was observed in fibroblasts 24 hours after MMC exposure (0.4 mg/ml) for 5 minutes ...

We generated DNA expression vectors encoding the full-length neu cDNA (designated pNeuN), the neu extracellular domain (pNeuE), or the neu extracellular and transmembrane domains (pNeuTM). The 293 cells transfected with pNeuN or pNeuTM expressed the neu extracellular domain on the surface membrane, whereas 293 cells transfected with pNeuE secreted the extracellular domain of neu into the culture supernatant. We examined whether i.m. injection of either of these plasmids could induce protective immunity in FVB/N mice against the adoptive transfer of Tgl-1 cells, a neu-expressing tumor cell line generated from a mouse mammary tumor that spontaneously arose in a FVB/N neu-transgenic mouse. The i.m. injection of pNeuTM or pNeuE, and to a lesser extent pNeuN, induced protective immunity against a subsequent challenge with Tgl-1 cells in FVB/N mice. In addition, the coinjection of a plasmid encoding interleukin-2 (designated pIL-2) augmented the efficacy of each of the pNeu plasmids for i...

Streptococcal pathogens have evolved to express exoglycosidases, one of which is BgaC β-galactosidase, to deglycosidate host surface glycolconjucates with exposure of the polysaccharide receptor for bacterial adherence. The paradigm BgaC protein is the bgaC product of Streptococcus, a bacterial surface-exposed β-galactosidase. Here we report the functional definition of the BgaC homologue from an epidemic Chinese strain 05ZYH33 of the zoonotic pathogen Streptococcus suis. Bioinformatics analyses revealed that S. suis BgaC shared the conserved active sites (W240, W243 and Y454). The recombinant BgaC protein of S. suis was purified to homogeneity. Enzymatic assays confirmed its activity of β-galactosidase. Also, the hydrolysis activity was found to be region-specific and sugar-specific for the Gal β-1,3-GlcNAc moiety of oligosaccharides. Flow cytometry analyses combined with immune electron microscopy demonstrated that S. suis BgaC is an atypical surface-anchored protein in that it la...

Streptococcus suis (S. suis) is a family of pathogenic gram-positive bacterial strains that represents a primary health problem in the swine industry worldwide. S. suis is also an emerging zoonotic pathogen that causes severe human infections clinically featuring with varied diseases/syndromes (such as meningitis, septicemia, and arthritis). Over the past few decades, continued efforts have made significant progress toward better understanding this zoonotic infectious entity, contributing in part to the elucidation of the molecular mechanism underlying its high pathogenicity. This review is aimed at presenting an updated overview of this pathogen from the perspective of molecular epidemiology, clinical diagnosis and typing, virulence mechanism, and protective antigens contributing to its zoonosis.

Talaflavuterpenoid A (1), a new nardosinane-type sesquiterpene, was isolated from the wetland soil-derived fungus Talaromyces flavus BYD07-13, and its structure was elucidated on the basis of HR-MS, NMR, and X-ray diffraction analysis. The absolute configuration of 1 was established by comparing the experimental electronic circular dichroism (ECD) spectrum with the calculated ECD spectra. Its cytotoxic effects on five human tumor cell lines (HL-60, SMMC-7721, A-549, MCF-7, and SW480), and antimicrobial activity against Escherichia coli, Staphylococcus aureus, Candida albicans, and Aspergillus niger were evaluated. This is the first report of the presence of nardosinane-type sesquiterpene in Talaromyces sp.

Major depression is diagnosed in 18% of patients following myocardial infarction (MI), and the antidepressant fluoxetine is shown to effectively decrease depressive symptoms and improve coronary heart disease prognosis. We observed the effect of fluoxetine on cardiac electrophysiology in vivo in a rat model of post-MI depression and the potential mechanism. Eighty adult male Sprague Dawley rats (200-250 g) were randomly assigned to five groups: normal control (control group), MI (MI group), depression (depression group), post-MI depression (model group), and post-MI depression treated with intragastric administration of 10 mg/kg fluoxetine (fluoxetine group). MI was induced by left anterior descending coronary artery ligation. Depression was developed by 4-week chronic mild stress (CMS). Behavior measurement was done before and during the experiment. Electrophysiology study in vivo and Western blot analysis were carried on after 4 weeks of CMS. After 4 weeks of CMS, depression-like ...

Depression is an independent risk factor for cardiovascular events and mortality in patients with myocardial infarction (MI). Excessive sympathetic activation and serious myocardial remodeling may contribute to this association. The aim of this study was to discuss the effect of depression on sympathetic activity and myocardial remodeling after MI. Wild-type (WT) rats were divided into a sham group (Sham), a myocardial infarction group (MI), a depression group (D), and a myocardial infarction plus depression group (MI+D). Compared with controls, the MI+D animals displayed depression-like behaviors and attenuated body weight gain. The evaluation of sympathetic activity showed an increased level in plasma concentrations of epinephrine and norepinephrine and higher expression of myocardial tyrosine hydroxylase in the MI+D group than the control groups (p<0.05 for all). Cardiac function and morphologic analyses revealed a decreased fractional shortening accompanied by increased left ...

The present study investigated the functional involvement of the rat hippocampal formation in easy and difficult visual discrimination learning by measuring regional brain cytochrome oxidase (C.O.) activity, an index of neuronal metabolic activity. The results showed that learning was related to C.O. activity in the CA field and the dentate gyrus of the hippocampal formation of rats in the difficult discrimination, whereas learning was related to C.O. activity only in the dentate gyrus of rats in the easy discrimination. The results suggest that difficult visual discrimination learning required greater involvement of the hippocampal formation than easy visual discrimination learning.

The present study investigated whether more vicarious trial-and-error (VTE) behavior, defined by head movement from one stimulus to another at a choice point during simultaneous discriminations, led to better visual discrimination learning in a Y-maze, and whether VTE behavior was a function of the hippocampus by measuring regional brain cytochrome oxidase (C.O.) activity, an index of neuronal metabolic activity. The results showed that the more VTEs a rat made, the better the rat learned the visual discrimination. Furthermore, both learning and VTE behavior during learning were correlated to C.O. activity in the hippocampus, suggesting that the hippocampus plays a role in VTE behavior during discrimination learning.

ABSTRACT Six new polyesters, talapolyesters A-F (1-4, 14, and 16), together with 11 known ones 15G256 nu (5), 15G256 nu-me (6), 15G256 pi (7), 15G256 beta-2 (8), 15G256 alpha-2 (9), 15G256 alpha-2-me (10), 15G256 iota. (11), 15G256 beta (12), 15G256 alpha (13), 15G256 alpha-1 (15), and 15G256 omega (17), were isolated from the wetland soil-derived fungus Talaromyces flavus BYD07-13, and their structures were determined by NMR and MS spectroscopic data. Among them, 1-4 and 16 were assembled in a different manner from that of the known 256 polyesters. All the polyesters are composed of (R)-2,4-dihydroxy-6-(2-hydroxypropyl)benzoic acid and (R)-3-hydroxybutyric acid/(S)-3,4-dihydroxybutyric acid residues. The absolute configurations of the residues were determined by alkaline hydrolysis. The cytotoxicity against five tumor cell lines of these compounds was examined, and a tight structure-activity relationship is proposed.

Urban residential buildings are formed, maintained and reformed by different external material and energy flows, and their behaviors of input, accumulation and output are characterized by their architectural factors and modes of use that usually determine the consumption of material and energy of a building at its overall life cycle.In this research, we took Beijing city, a rapid developing city

To investigate the feasibility of radiolabelled annexin V imaging for early diagnosis of Parkinson's disease. 99mTc-HYNIC-annexin V was prepared and its binding to apoptotic cell models of Parkinson's disease was studied in vitro. Cellular models of Parkinson's disease were produced by administering different concentrations of 1-methyl-4-phenylpyridinium to PC12 and SH-SY5Y cell lines. Cell apoptosis rates were analysed by flow cytometry. Annexin V was labelled with 99mTc by hydrazinonicotinamide (HYNIC). Cell binding studies were carried out using cellular models of Parkinson's disease. Cell uptake studies were also performed after different levels of MPP treatment, and the correlation between the degree of apoptosis and Tc-HYNIC-annexin V uptake was analysed. The specific activity of 99mTc-HYNIC-annexin V was 3.7-74 x 10 Bq.mg protein. In-vitro binding of 99mTc-HYNIC-annexin V to model cells was specific, saturable and time dependent. Scatchard analysis gave a Kd of 7.16+/-1.78 nmol.l, Bmax values of 179+/-33 fmol per 10(6) cells (PC12) and 220+/-26 fmol per 10(6) cells (SHSY5Y). MPP at different concentrations can induce cell apoptosis in a dose-dependent manner; cellular uptake of 99mTc-HYNIC-annexin V as indicated by membrane-bound radiolabelled annexin V activity was linearly correlated with total fluorescence, as observed by FITC-annexin V flow cytometry (PC12: r=0.924; SH-SY5Y: r=0.937, P<0.01). 99mTc-HYNIC-annexin V retains its receptor-binding activity and has a high affinity to cellular models of Parkinson's disease. The uptake of radioactivity correlated well with cell apoptosis rates; thus, 99mTc-annexin V is a potential imaging agent with which to detect early neuron damage in Parkinson's disease.

Long-term clinical observations and ongoing studies have shown antitumor effects of external Qi of Yan Xin Qigong (YXQG-EQ) that originated from traditional Chinese medicine (TCM). In order to understand the molecular mechanisms underlying the antitumor effects of YXQG-EQ, we investigate the effects of YXQG-EQ on growth and apoptosis in androgen-independent prostate cancer PC3 cells. We found that exposure to YXQG-EQ led to G2/M arrest associated with reduced cyclin B1 expression and apoptosis in PC3 cells. YXQG-EQ treatment inhibited constitutive and epidermal growth factor-induced Akt phosphorylation, basal and TNF-alpha-induced NF-kappaB activation, and downregulated anti-apoptotic Bcl-2 and Bcl-xL expression. In contrast, exposure to YXQG-EQ increased phosphorylation of Akt and Erk1/2 in human umbilical vein endothelial cells (HUVEC), and had no cytotoxic effect on either HUVEC or peripheral blood mononuclear cells (PBMC). These results indicate that YXQG-EQ has profound effects on growth and apoptosis of prostate cancer cells by targeting survival pathways including the Akt and NF-kappa B pathways.

... Jiwei Zhai a , Corresponding Author Contact Information , E-mail The Corresponding Author ,Dan Hu a , Xi Yao a , Zhengkui Xu b and Haydn Chen b. ... 2. Experimental. The barium acetate [Ba(CH 3 COO) 2 ], [Zr(OC 3 H 7 ) 4 ], and [Ti(OC 3 H 7 ) 4 ] were used as starting materials. ...

Malectin is an endoplasmic reticulum-resident lectin, which recognizes di-glucosylated Glc(2)Man(9)GlcNAc(2) (G2M9) N-glycans on newly synthesized glycoproteins. We previously demonstrated that malectin preferentially associates with misfolded glycoproteins and inhibits their secretion (Chen, Y., Hu, D., Yabe, R., Tateno, H., Qin, S. Y., Matsumoto, N., Hirabayashi, J., and Yamamoto, K. (2011) Mol. Biol. Cell 22, 3559-3570). The sugar binding activity of malectin is required for this process. However, because G2M9 N-glycans are generated at the very early stage of processing and are typically found on both misfolded glycoproteins and glycoproteins undergoing folding, other mechanisms must underlie the preference of malectin for misfolded glycoproteins. Here, we searched for proteins that were co-immunoprecipitated with malectin, and we found that malectin formed a stable complex with an endoplasmic reticulum-resident transmembrane protein, ribophorin I. Co-expression of malectin and ribophorin I significantly enhanced the association between malectin and a folding-defective α1-antitrypsin variant (null Hong Kong) and reduced its secretion; however, secretion of wild-type α1-antitrypsin was not affected. The enhanced association and reduced secretion were counteracted by siRNA-mediated down-regulation of ribophorin I. Also, a reporter assay revealed that ribophorin I preferentially interacted with misfolded ribonuclease A but not with the native form, suggesting that ribophorin I may function as a chaperone that recognizes misfolded proteins inside cells. These results provide the first evidence of the mechanism by which malectin preferentially associates with misfolded glycoproteins.