Synthesis and biological activity of tubercidin analogues of ppp5'A2'p(5'A2'p)n5'A

A series of tubercidin (7-deazaadenosine) analogues of 2-5A of the general formula p5'(c7A)2'p[5'(c7A)-2'p]n5'(c7A) (n = 0-5) were prepared by lead ion catalyzed polymerization of the 5'-phosphoroimidazolidate of tubercidin. Through the corresponding imidazolidates, these oligonucleotide 5'-monophosphates were converted to the 5'-triphosphates. All reported structures were corroborated by enzyme digestion and 1H or 31P nuclear magnetic resonance. When evaluated for its ability to bind to the 2-5 A-dependent endonuclease of mouse L cells, the tubercidin analogue of trimeric 2-5A, namely, ppp5'(c7A)2'p5'(c7A)2'p5'(c7A), and the corresponding tetramer were bound as effectively as 2-5A itself; nonetheless, it and the corresponding tetramer, ppp5'-(c7A)2'p5'(c7A)2'p5'(c7A)2'p5'(c7A), failed to stimulate the 2-5A-dependent endonuclease as judged by its inability to inhibit translation in extracts of mouse L cells programmed with encephalomyocarditis virus RNA and to give rise to ribosomal RNA cleavage in the same cell system under conditions where 2-5A showed activity at 10(-9) M. The trimer, ppp5'(c7A)2'p5'(c7A)2'p5'(c7A), was an antagonist of 2-5A action in the L cell extract. In the lysed rabbit reticulocyte system, both the trimeric and tetrameric tubercidin 2-5A analogues were bound to the 2-5A-dependent endonuclease as well as 2-5A, but in this case, the tetramer triphosphate, ppp5'(c7A)2'p5'(c7A)2'p5'(c7A)2'p5'(c7A), was just as potent an inhibitor of translation as 2-5A tetramer triphosphate. Moreover, this inhibition was prevented by the established 2-5A antagonist p5'A2'p5'A2'p5'A.(ABSTRACT TRUNCATED AT 250 WORDS)     

Metal-binding properties of phytochelatin-related peptides

Phytochelatins (PCs, (gamma Glu-Cys)(n)-Gly, n=2-11) are produced by higher plants, algae and some fungi in order to detoxify Cd(2+) by sequestration to form Cd-PCs complexes. In order to investigate what chemical structures of PCs are responsible for their metal-binding ability, various cysteine-rich peptides ((X-Cys)(7)-Gly, X=Glu, Asp, Lys, Gly, Ser and Gln) were chemically synthesized. Water-solubility, metal-binding property, and detoxification effect toward Cd(2+) were analyzed and compared with those of (gamma EC)(7)G. (SC)(7)G and (QC)(7)G were insoluble at pH below 10, and (GC)(7)G was not soluble at any pH between 1 and 12, indicating that charged side chains were at least required for the molecules to be solubilized in aqueous solution. By spectroscopic analyses using DTNB method and UV method, we found that (EC)(7)G and (DC)(7)G had almost equivalent abilities of Cd(2+)-binding as PC ((gamma EC)(7)G), indicating that the distance between each thiol group was not a major factor for the binding to Cd(2+). (beta DC)(7)G and (KC)(7)G interacted to Cd(2+) with fourth coordination as in the case of other soluble PC-related peptides. However, compared to (gamma EC)(7)G, (beta DC)(7)G displayed a slightly weaker binding to Cd(2+), and (KC)(7)G showed a drastic decrease in binding ability. The affinities of PC-related peptides toward Cd(2+) were evaluated as below; (gamma EC)(7)G=(EC)(7)G=(DC)(7)G>(beta DC)(7)G>(KC)(7)G=weak binding. The results of Cd(2+)-detoxification assays were consistent with the affinity between Cd(2+) and the peptides. We concluded that the structure consisting of thiol and carboxyl groups were essential for the formation of a tight Cd-peptides complex such as Cd-PCs.     

Anti-inflammatory effects and changes in prostaglandin patterns induced by 7beta-hydroxy-epiandrosterone in rats with colitis

High dose levels of dehydroepiandrosterone and its 7-hydroxylated derivatives have been shown to reduce oxidative stress and inflammatory responses in dextran sodium sulfate (DSS)-induced colitis in rats. Another endogenous steroid, 7beta-hydroxy-epiandrosterone (7beta-hydroxy-EpiA) has been shown to exert neuroprotective effects at much smaller doses. Our aims were to evaluate whether 7beta-hydroxy-EpiA pre-treatment prevents DSS-induced colitis and to determine whether the effects involve changes in anti-inflammatory prostaglandin (PG) D(2) and 15-deoxy-Delta(12,14)-PGJ(2) (15d-PGJ(2)) levels. Rats were administered 0.01, 0.1 and 1mg/kg 7beta-hydroxy-EpiA i.p. once a day for 7 days. Thereafter, colitis was induced by administration of 5% DSS in drinking water for 7 days. Levels of the PGs and the expression of cyclooxygenase (COX-2) and PG synthases were assessed during the course of the experiment. Administration of 7beta-hydroxy-EpiA caused a transient increase in COX-2 and PGE synthase expression within 6-15h and augmented colonic tissue levels of 15d-PGJ(2) levels starting at day 2. Treatment with DSS resulted in shortened colon length, depleted mucus in goblet cells and induced oxidative stress. COX-2 and mPGES-1 synthase expression were enhanced and accompanied by increased PGE(2), D(2) and 15d-PGJ(2) production. Although all dose levels of 7beta-hydroxy-EpiA reduced PGE(2) production, only the lowest dose (0.01mg/kg) of the steroid completely prevented colitis damage and tissue inflammation. 7beta-Hydroxy-EpiA pre-treatment prevents the occurrence of DSS-induced colitis through a shift from PGE(2) to PGD(2) production, associated with an early but transient increase in COX-2 expression and a sustained increase in the production of the anti-inflammatory prostaglandin 15d-PGJ(2).     

Measurement of 7 methyl and 7 2 hydroxyethyl guanine DNA adducts in white blood cells of smokers and non smokers

The goal of the present study was to measure the levels of 7-methylguanine and 7-(2- hydroxyethyl)guanine DNA adducts in human white blood cells in relation to smoking. DNA was isolated from samples of 11 smokers and eight non-smokers. The 32P-postlabelled 7-methylguanine and 7-(2-hydroxyethyl)guanine adducts were analysed by thin-layer chromatography (TLC) combined with a high pressure liquid chromatography (HPLC) assay. In smokers the mean 7-methylguanine and 7-(2-hydroxyethyl)guanine levels were 32.3 +/- 7.1 and 6.6 +/- 2.3 adducts per 108 nucleotides respectively. The corresponding values in non-smokers were 25.0 +/- 7.0 and 3.7 +/- 2.4 adducts per 108 nucleotides. There were significantly higher levels of 7-methylguanine and 7-(2-hydroxyethyl)guanine adducts in WBC in smokers than in non-smokers ( p = 0.041; p = 0.018), respectively. A positive correlation between 7-methylguanine and 7-(2-hydroxyethyl)guanine levels was observed.     

Acylated flavonoids and phenol glycosides from Veronica thymoides subsp. pseudocinerea

A new acylated flavone glucoside, 3'-hydroxyscutellarein 7-O-(6'-O-protocatechuoyl)-beta-glucopyranoside (1), and a new phenol glucoside, 3,5-dihydroxyphenethyl alcohol 3-O-beta-glucopyranoside (6) were isolated from Veronica thymoides subsp. pseudocinerea together with seven known flavone, phenol and lignan glycosides; 3'-hydroxyscutellarein 7-O-(6'-O-trans-feruloyl)-beta-glucopyranoside (2), 3'-hydroxy, 6-O-methylscutellarein 7-O-beta-glucopyranoside (3), luteolin 7-O-beta-glucopyranoside (4), isoscutellarein 7-O-(6'-O-acetyl)-beta-allopyranosyl (1' --> 2')-beta-glucopyranoside (5), 3,4-dihydroxyphenethyl alcohol 8-O-beta-glucopyranoside (7), benzyl alcohol 7-O-beta-xylopyranosyl (1" --> 2')-beta-glucopyranoside (8), and (+)-syringaresinol 4'-O-beta-glucopyranoside (9). Compounds 2, 3 and 7-9 were reported for the first time in the genus Veronica. The structures of the isolates were determined by means of spectroscopic (UV, IR, 1D and 2D NMR, HR ESI-MS) methods. Isolated compounds (1-7) exhibited potent radical scavenging activity against the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical.     

Measurement of 7 methyl and 7 2 hydroxyethyl guanine DNA adducts in white blood cells of smokers and non smokers

The goal of the present study was to measure the levels of 7-methylguanine and 7-(2- hydroxyethyl)guanine DNA adducts in human white blood cells in relation to smoking. DNA was isolated from samples of 11 smokers and eight non-smokers. The 32P-postlabelled 7-methylguanine and 7-(2-hydroxyethyl)guanine adducts were analysed by thin-layer chromatography (TLC) combined with a high pressure liquid chromatography (HPLC) assay. In smokers the mean 7-methylguanine and 7-(2-hydroxyethyl)guanine levels were 32.3 +/- 7.1 and 6.6 +/- 2.3 adducts per 108 nucleotides respectively. The corresponding values in non-smokers were 25.0 +/- 7.0 and 3.7 +/- 2.4 adducts per 108 nucleotides. There were significantly higher levels of 7-methylguanine and 7-(2-hydroxyethyl)guanine adducts in WBC in smokers than in non-smokers (p = 0.041; p = 0.018), respectively. A positive correlation between 7-methylguanine and 7-(2-hydroxyethyl)guanine levels was observed.     

Elevation of plasma 7B2 (a novel pituitary protein) in cord blood at obstetrical delivery and the possible correlation with GH

Plasma 7B2-immunoreactivity (7B2-IR) concentrations in umbilical artery (UA), umbilical vein (UV) and maternal vein (MV) were measured by RIA at the time of obstetrical delivery at term. Plasma 7B2-IR concentrations (Mean +/- SEM) in UA (N = 12), UV (N = 16) and MV (N = 16) were 725 +/- 69, 699 +/- 64 and 116 +/- 4.5 pg/ml, respectively. Plasma 7B2-IR concentrations in UA and UV were much higher than those in MV. There was no arterio-venous gradient between UA and UV. A trace amount of 7B2-IR (Mean +/- SEM, 226 +/- 16.8 pg/g tissue) was detected in the placental extracts. A statistically significant positive correlation (r = 0.7595, p less than 0.005) was found between plasma 7B2-IR and GH in the UV. Significant negative correlations between body weight of the neonates and plasma levels of GH (r = -0.6836, p less than 0.01) and 7B2-IR (r = -0.4939, p less than 0.05) were also apparent. When analyzing cord blood plasma using gel permeation chromatography and SDS-polyacrylamide gel electrophoresis, a major peak with an apparent molecular weight of 20,000 was observed. These findings suggest that 7B2-IR in UA and UV originates from the fetus and that 7B2-IR does not permeate through the placenta. The possibility of involvement of 7B2 in fetal growth warrants attention.     

Absence of the P2X7 receptor alters leukocyte function and attenuates an inflammatory response

When challenged with extracellular ATP, leukocytes respond and activate processes attributed to the P2X(7) receptor (P2X(7)R), an unusual ligand-gated ion channel. To prove P2X(7)R involvement, blood samples from P2X(7)R-deficient mice were characterized. Monocytes and lymphocytes associated with wild-type blood responded to ATP and underwent volume/shape changes and shed L-selectin. In contrast, leukocytes from P2X(7)R-deficient animals demonstrated no change in physical properties or L-selectin expression following ATP challenge. Blood stimulated with LPS or ATP individually generated minimal quantities of the leaderless polypeptide IL-1 beta, but sequential treatment of wild-type, but not P2X(7)R-deficient, blood with LPS and ATP yielded large amounts of cell-free cytokine. Based on these differences, wild-type and P2X(7)R-deficient animals were compared following induction of monoclonal anti-collagen-induced arthritis. Ab-treated wild-type animals subsequently challenged with LPS developed inflamed, swollen paws; their joint cartilage demonstrated lesions, loss of proteoglycan content, and the presence of collagen degradation products. P2X(7)R-deficient animals subjected to the same challenge were markedly less affected; both the incidence and severity of disease were reduced. These data indicate that ATP does act via the P2X(7)R to affect leukocyte function and that the P2X(7)R can serve as an important component of an in vivo inflammatory response.     

Characterization of noncovalent complexes of recombinant human monoclonal antibody and antigen using cation exchange, size exclusion chromatography, and BIAcore.

The binding of fully human monoclonal antibodies (MAbs) D2E7 and 2SD4 to their antigen, human tumor necrosis factor-alpha (TNFalpha), was investigated by BIAcore, cation exchange (CIEX), and size exclusion liquid chromatography (SEC) using ultraviolet and laser light scattering detectors. D2E7 has a higher affinity for TNFalpha than 2SD4 and the two antibodies (Abs) differ by 12 amino acids in the antigen (Ag) binding regions. A BIAcore biosensor instrument was used to determine the association, k(on) and dissociation, k(off), rate constants for the binding of TNFalpha to D2E7 and 2SD4. The HPLC methods were used to resolve and to study D2E7, 2SD4, and TNFalpha molecules and the noncovalent complexes of D2E7 and 2SD4 with TNFalpha. The CIEX method demonstrated that all D2E7 charged-variants bound TNFalpha equally well. There was no preferential binding for any one of D2E7 charged-variants to TNFalpha. D2E7 and 2SD4 Abs were resolved by the CIEX method. When a mixture of D2E7 and 2SD4 was mixed with excess TNFalpha, D2E7. TNFalpha complexes were formed before any 2SD4. TNFalpha complexes. Thus, the CIEX method was able to rank the affinities of the MAbs. D2E7 and TNFalpha formed complexes of 600-5000 kDa. The molecular weights of various D2E7. TNFalpha complexes were determined by a SEC method with light scattering (LS) and refractive index (RI) detectors. Upon overnight incubation, a 598-kDa complex emerged as the most stable and the only D2E7. TNFalpha complex. The molar ratio of D2E7 to TNFalpha in this complex was approximately 1:1. Based on molecular weights and the molar ratio, an immune complex, consisting of alternating three D2E7 and three TNFalpha molecules, is proposed as the most stable complex.     

Memory Th2 effector cells can develop in the absence of B7-1/B7-2, CD28 interactions,and effector Th cells after priming with an intestinal nematode parasite

B7-1/B7-2 interactions are required for many Th2-cell mediated primary immune responses including the response that follows infection with the intestinal nematode parasite, Heligmosomoides polygyrus. However, few studies have examined the role of B7-1/B7-2/CD28 interactions in the development of a Th2 memory immune response. We examined the development of the memory Th2 response to H. polygyrus in BALB/c mice deficient in both B7-1 and B7-2 (B7-1/B7-2(-/-)) and in BALB/c mice deficient in CD28 (CD28(-/-)). Following primary inoculation with H. polygyrus, adult worms in the gut were cleared with an anti-helminthic drug and mice were subsequently challenge-inoculated with H. polygyrus larvae. The memory Th2 response is readily distinguished by its inhibitory effect on adult worm maturation, resulting in marked reductions in adult worm egg production that are not observed during the primary immune response. Following H. polygyrus challenge inoculation, comparable decreases in egg production and similar increases in mesenteric lymph node cell IL-4 production were observed in B7-1/B7-2(-/-) and B7-1/B7-2(+/+) mice. However, elevations in total serum IgG1 and IgE were reduced, while increases in serum Ag-specific IgG1 and IgE and germinal center formation were blocked in H. polygyrus-challenged B7-1/B7-2(-/-) mice. In contrast, in H. polygyrus-challenged CD28(-/-) mice, marked elevations in Ag-specific IgG1 and IgE and increased germinal center formation were observed. The results of these studies demonstrate that effector Th2 memory cells that produce IL-4 and mediate host defense can develop when B7-1/B7-2 interactions, and associated effector Th2 cell development, are blocked during priming. However, humoral immunity is impaired and differentially affected in B7-1/B7-2(-/-) mice and CD28(-/-) mice following H. polygyrus challenge.     

Modification of DNA with octadiynyl side chains: synthesis, base pairing, and formation of fluorescent coumarin dye conjugates of four nucleobases by the alkyne--azide "click" reaction

Oligonucleotides incorporating 5-(octa-1,7-diynyl)-2'-deoxycytidine 1a, 5-(octa-1,7-diynyl)-2'-deoxyuridine 2a and 7-deaza-7-(octa-1,7-diynyl)-2'-deoxyguanosine 3a, 7-deaza-7-(octa-1,7-diynyl)-2'-deoxyadenosine 4a were prepared. For this, the phosphoramidites 7, 10, and 13 were synthesized and employed in solid-phase oligonucleotide synthesis. The octa-1,7-diynyl nucleosides 1a- 4a were obtained from their corresponding iodo derivatives using the palladium-assisted Sonogashira cross-coupling reaction. The Tm values demonstrated that DNA duplexes containing octa-1,7-diynyl nucleosides show a positive influence on the DNA duplex stability when they are introduced at the 5-position of pyrimidines or at the 7-position of 7-deazapurines. The terminal alkyne residue of oligonucleotides were selectively conjugated to the azide residue of the nonfluorescent 3-azido-7-hydroxycoumarin ( 38) using the protocol of copper(I)-catalyzed [3 + 2] Huisgen--Sharpless--Meldal cycloaddition "click chemistry" resulting in the formation of strongly fluorescent 1,2,3-triazole conjugates. The fluorescence properties of oligonucleotides with covalently linked coumarin--nucleobase assemblies were investigated. Among the four modified bases, the 7-deazapurines show stronger fluorescence quenching than that of pyrimidines.     

Iridoid glucosides and flavones from the aerial parts of Avicennia marina

Two new iridoid glucosides, namely, 2'-O-[(2E,4E)-5-phenylpenta-2,4-dienoyl]mussaenosidic acid (1; mussaenosidic acid = [1S-(1alpha,4aalpha,7alpha,7aalpha)]-1-(beta-D-glucopyranosyloxy)-1,4a,5,6,7,7a-hexahydro-7-hydroxy-7-methylcyclopenta[c]pyran-4-carboxylic acid) and 2'-O-(4-methoxycinnamoyl)mussaenosidic acid (2), were isolated from the aerial parts of the mangrove plant Avicennia marina. Beside that, one known iridoid glucoside, 2'-O-coumaroylmussaenosidic acid (3) and four known flavones (flavone = 2-phenyl-4H-1-benzopyran-4-one) including 4',5-dihydroxy-3',7-dimethoxyflavone (4), 4',5-dihydroxy-3',5',7-trimethoxyflavone (5), 4',5,7-trihydroxyflavone (6), and 3',4',5-trihydroxy-7-methoxyflavone (7) were also isolated and identified. The structures of these compounds were elucidated by NMR spectroscopy and by low- and high-resolution mass spectrometry. The chemotaxonomic significance of these findings was discussed. In addition, each isolated compound was evaluated for the ability of alpha,alpha-diphenyl-beta-picrylhydrazyl (DPPH) radical-scavenging activity.     

Persistence of Escherichia coli O157:H7 in barley silage: effect of a bacterial inoculant

AIMS: The effect of a lactic acid producing bacterial (LAB) inoculant on the elimination of Escherichia coli O157:H7 from barley forage was assessed. METHODS AND RESULTS: Triplicate mini-silos were prepared for four treatments and six sampling times (1, 3, 7, 15, 30 and 42 d post-ensiling). The treatments were (i) 10(5) cfu g(-1) Pediococcus pentosaceus and Propionibacterium jenzenii (P2); (ii) 10(5) cfu g(-1) E. coli O157:H7 strain 3081 and 10(5) cfu g(-1) E. coli Biotype 1 strains 719IE10, 719IE14 and 614ME49 (EC); (iii) P2 + EC; and (iv) the control (sterile distilled water). Triplicate mini-silos were opened at each sampling time for pH, volatile fatty acid (VFA) and lactate determinations and E. coli, E. coli O157:H7 and LAB were enumerated. On d 3 and 7, numbers of E. coli O157:H7 in P2 + EC were significantly lower than in EC (P < 0;05). Escherichia coli O157:H7 was not detected in P2 + EC and EC at 7 and 15 d post-ensiling, respectively. On d 15 through 42, E. coli Biotype 1 was not detected in P2 + EC or EC. Populations of LAB were higher in P2 and P2 + EC than in the control and EC on d 3 and 7 (P < 0.05). After 3 d of ensiling, lactate levels were higher (P < 0.05) and pH was lower (P < 0.05) in P2 and P2 + EC as compared to the control and EC. Bacteriocins of P2 were not found to be inhibitory to E. coli O157:H7 using the agar-spot procedure. Escherichia coli O157:H7 inoculated into the control silage at a level of 10(3) cfu g(-1) and exposed to aerobic conditions at 22 degrees C was not detected after 1 d and remained undetectable for the 28 d exposure period. CONCLUSIONS: Silage inoculant P2 increased lactate levels and decreased pH more rapidly during ensiling, which appeared to hasten the elimination of E. coli O157:H7 from the silage. SIGNIFICANCE AND IMPACT OF THE STUDY: Results emphasize the importance of adequate ensiling since E. coli O157:H7 may be maintained and spread among cattle through feed.     

Chinese Strains (Type 7) of JC Virus Are Afro-Asiatic in Origin But Are Phylogenetically Distinct from the Mongolian and Indian Strains (Type 2D) and the Korean and Japanese Strains (Type 2A)

We have further characterized the Asian genotypes (Types 2 and 7) and subtypes of JC virus (JCV). Urine samples from 224 individuals with Han and Mongolian populations were collected in five regions in eastern China: Kunming, Chengdu, Shenyang, Chifeng, and Manzhouli. Also, 99 urine samples were collected from coastal and hill groups in Kerala, southern India, and 23 urine samples from Seoul, Korea. PCR products of four typing fragments were sequenced, including two in the VP1 gene, as well as one each in the VT intergenic region and regulatory region. It was possible to clone and sequence a total of 42 JCV whole genomes (~5120 bp). Five genotypes of JCV (Types 7A, 7B, 7C, 2D, and 4) were found in China, four genotypes (Types 2D, 7C, 4, and 1B) in southern India, and three genotypes (Types 7B, 2A, and 1A) in Korea. Type 7A was most prevalent in South China (59–64%) and Type 7B was predominant in northeast China and Inner Mongolia (67–77%). Type 7C strains were spread throughout North and South China (3–14%), while Type 2D strains were found only in the two Mongolian groups (9–10%). In southern India, Type 2D was predominant in the coastal group (95%), and two major types, Type 7C (50%) and Type 2D (35%), were prevalent in the tribal hill groups. In Korea two major genotypes were found: Type 7B (50%) and Type 2A (43%). Phylogenetic reconstruction places the Chinese genotypes in the Afro-Asiatic supercluster, but distinct from the Mongolian and Indian strains (Type 2D), as well as the Korean and Japanese genotype (Type 2A) that predominates in the Americas.     

Immunoglobulin fold characteristics of B7-1 (CD80) and B7-2 (CD86).

B7-1 and B7-2 are expressed on antigen-presenting cells and bind to the CD28 and CTLA-4 receptors on T cells. These interactions trigger a costimulatory pathway that is essential for T-cell activation. B7-1 and B7-2 are members of the immunoglobulin superfamily (IgSF) and, despite sharing common function, have only limited sequence similarity. The B7-1 extracellular region was previously subdivided into 2 IgSF domains, an N-terminal V(ariable)-like domain, followed by a C(onstant)-like domain. We recently reported that the V-like domains of B7-1 and B7-2 share some significant sequence similarities with 3 major histocompatibility complex (MHC)-encoded members of the IgSF. We have now applied inverse folding methodology to assess the compatibility of the B7-1 and B7-2 extracellular region sequences with currently available 3-dimensional structures. In these calculations, the sequences of the N-terminal (V-like) domains in B7-1 and B7-2 were not compatible with known structures, including the IgSF V-set. In contrast, the sequences of the C-like domains were compatible with IgSF C-set structures and were best recognized by the beta 2-microglobulin (beta 2m) domain of MHC Class I. A sequence comparison of the C-like domains in the B7 molecules showed that 11 of 17 rigorously conserved residues in B7-1 and B7-2 are not IgSF C-1 set consensus residues. When mapped onto the corresponding positions of the beta 2m structure, the conserved residues in B7 cluster on the surface, where they may interact with the B7 V-like domain or other molecules.     

岗松化学成分的研究

从岗松的石油醚和氯仿提取组分中分离得到7个化合物,根据NMR、HSQC、HMBC、IR和MS的数据分析及与文献对照,确定它们的结构为白桦脂酸(betulinic acid,1),齐墩果酸(oleanolic acid,2),没食子酸乙酯(ethyl sabre,3),5-羟基-6-甲基-7-甲氧基-二氢黄酮(5-hydroxy-6-methyl-7-methoxy-flavanone,4),5-羟基-7-甲氧基-8-甲基二氢黄酮(5-hydroxy-7-methoxy-8-methylflavanone,5),5-羟基-7-甲氧基-2-异丙基色原酮(5-hydroxy-7-methoxy-2-isopropylchromone,6)和β-谷甾醇(β-sitosterol,7).其中,化合物1～5和7为首次从该植物中分离得到.     

Inhibition of eukaryotic translation by nucleoside 5'-monophosphate analogues of mRNA 5'-cap: changes in N7 substituent affect analogue activity

Nucleotide cap analogues of 7-methylguanosine 5'-monophosphate (m7GMP) were synthesized in which the 7-methyl moiety was replaced with 7-ethyl (e7), 7-propyl (p7), 7-isopropyl (ip7), 7-butyl (b7), 7-isobutyl (ib7), 7-cyclopentyl (cp7), 7-(carboxymethyl) (cm7), 7-benzyl (bn7), 7-(2-phenylethyl) [7-(2-PhEt)], and 7-(1-phenylethyl) [7-(1-PhEt)]. These derivatives were assayed as competitive inhibitors of capped mRNA translation in reticulocyte lysate. We observed that N7 alkyl and alicyclic substituents larger than ethyl significantly decreased the inhibitory activity of these cap analogues presumably by decreasing their affinity for cap binding proteins, which participate in the initiation of translation. This result defined a maximum size for this class of N7 substituents in the nucleotide binding domain of cap binding proteins. Like m7GMP, the N7-substituted GMP derivatives synthesized in this study were found to be predominantly in the anti conformation as determined by proton NMR analyses. However, bn7GMP and 7-(2-PhEt)GMP, which have aromatic N7 substituents, were more effective than m7GMP as competitive inhibitors of translation. The increased affinity of bn7GMP for cap binding proteins was further examined by synthesis of beta-globin mRNA containing 5'-bn7G, 5'-m7G, or 5'-e7G cap structures. These modified mRNAs were tested as translation templates. Messenger RNA capped with bn7G was observed to increase the translation activity of the template 1.8-fold relative to that of its m7G-capped mRNA counterpart. By contrast, e7G-capped mRNA was 25% less active than m7G-capped mRNA.2+V photo-cross-linking of m7G-capped mRNA to cap binding proteins     

Attenuating effect of angiotensin-(1-7) on angiotensin II-mediated NAD(P)H oxidase activation in type 2 diabetic nephropathy of KK-A(y)/Ta mice

ANG-(1-7) is associated with vasodilation and nitric oxide synthase stimulation. However, the role of ANG-(1-7) in type 2 diabetes mellitus is unknown. In this study, we examined the hypothesis that ANG-(1-7) attenuates ANG II-induced reactive oxygen species stress (ROS)-mediated injury in type 2 diabetic nephropathy of KK-A(y)/Ta mice. KK-A(y)/Ta mice were divided into four groups: 1) a control group; 2) ANG II infusion group; 3) ANG II+ANG-(1-7) coinfusion group; and 4) ANG II+ANG-(1-7)+d-Ala(7)-ANG-(1-7) (A779) coinfusion group. In addition, primary mesangial cells were cultured and then stimulated with 25 mM glucose with or without ANG II, ANG-(1-7), and A779. The ANG II+ANG-(1-7) coinfusion group showed a lower urinary albumin/creatinine ratio increase than the ANG II group. ANG-(1-7) attenuated ANG II-mediated NAD(P)H oxidase activation and ROS production in diabetic glomeruli and mesangial cells. ANG II-induced NF-κB and MAPK signaling activation was also attenuated by ANG-(1-7) in the mesangial cells. These findings were related to improved mesangial expansion and to fibronectin and transforming growth factor-β1 production in response to ANG II and suggest that ANG-(1-7) may attenuate ANG II-stimulated ROS-mediated injury in type 2 diabetic nephropathy. The ACE2-ANG-(1-7)-Mas receptor axis should be investigated as a novel target for treatment of type 2 diabetic nephropathy.