c-Myc binds to 5' flanking sequence motifs of the dihydrofolate reductase gene in cellular extracts: role in proliferation.

The dihydrofolate reductase is a key enzyme of the folate metabolism which supplies the cell with dTTPs for DNA synthesis. Using cellular extracts, we demonstrate the formation of c-Myc/Max heterodimers at the dihydrofolate reductase (DHFR) 5' flanking CANNTG (E-box) motifs. The presence of these complexes correlates with c-Myc levels and active cellular proliferation.     

体外冲击波碎石术对兔肝酶活性的影响

采用超微组织化学方法,观察了体外冲击波碎石术(ESWL)对家免肝脏酶活性的影响。实施 ESWL 后,肝细胞的琥珀酸脱氢酶(SDH)和毛细胆管壁上的碱性磷酸酶(ALP)、焦磷酸硫胺素酶(TPPase)反应活性减弱或消失。TPPase 从损伤的肝细胞高尔基体分泌面扁囊、溶酶体样小泡和毛细胆管内溢出,并伴有肝细胞面的质膜上出现了 TPPase 反应产物和形成膜包内凹小泡。结果提示 ESWL 可对肝细胞及毛细胆管的功能和结构有损伤作用。     

“DNA signaturing” database construction for Tetradesmus species identification and phylogenetic relationships of Scenedesmus-like green microalgae (Scenedesmaceae,Chlorophyta)

Species identification of Scenedesmus-like microalgae, comprising Desmodesmus, Tetradesmus, and Scenedesmus, has been challenging due to their high morphological and genetic similarity. After developing a DNA signaturing tool for Desmodesmus identification, we built a DNA signaturing database for Tetradesmus. The DNA signaturing tool contained species-specific nucleotide sequences of Tetradesmus species or strain groups with high similarity in ITS2 sequences. To construct DNA signaturing, we collected data on ITS2 sequences, aligned the sequences, organized the data by ITS2 sequence homology, and determined signature sequences according to hemi-compensatory base changes (hCBC)/CBC data from previous studies. Four Tetradesmus species and 11 strain groups had DNA signatures. The signature sequence of the genus Tetradesmus, TTA GAG GCT TAA GCA AGG ACCC, recognized 86% (157/183) of the collected Tetradesmus strains. Phylogenetic analysis of Scenedesmus-like species revealed that the Tetradesmus species were monophyletic and closely related to each other based on branch lengths. Desmodesmus was suggested to split into two subgenera due to their genetic and morphological distinction. Scenedesmus must be analyzed along with other genera of the Scenedesmaceae family to determine their genetic relationships. Importantly, DNA signaturing was integrated into a database for identifying Scenedesmus-like species through BLAST.     

Chemical Profile and Biological Activities of Fungal Strains Isolated from Piper nigrum Roots: Experimental and Computational Approaches

The current report describes the chemical investigation and biological activity of extracts produced by three fungal strains Fusarium oxysporum, Penicillium simplicissimum, and Fusarium proliferatum isolated from the roots of Piper nigrum L. growing in Vietnam. These fungi were namely determined by morphological and DNA analyses. GC/MS identification revealed that the EtOAc extracts of these fungi were associated with the presence of saturated and unsaturated fatty acids. These EtOAc extracts showed cytotoxicity towards cancer cell lines HepG2, inhibited various microbacterial organisms, especially fungus Aspergillus niger and yeast Candida albicans (the MIC values of 50–100 μg/mL). In α-glucosidase inhibitory assay, they induced the IC₅₀ values of 1.00-2.53 μg/mL were better than positive control acarbose (169.80 μg/mL). The EtOAc extract of F. oxysporum also showed strong anti-inflammatory activity against NO production and PGE-2 level. Four major compounds linoleic acid (37.346 %), oleic acid (27.520 %), palmitic acid (25.547 %), and stearic acid (7.030 %) from the EtOAc extract of F. oxysporum were selective in molecular docking study, by which linoleic and oleic acids showed higher binding affinity towards α-glucosidase than palmitic and stearic acids. In subsequent docking assay with inducible nitric oxide synthase (iNOS), palmitic acid, oleic acid and linoleic acid could be moderate inhibitors.     

Novel (E)-3-(3-Oxo-4-substituted-3,4-dihydro-2H-benzo[b][1,4]oxazin-6-yl)-N-hydroxypropenamides as Histone Deacetylase Inhibitors: Design,Synthesis and Bioevaluation

Herein, we report the design, synthesis and evaluation of novel (E)-3-(3-oxo-4-substituted-3,4-dihydro-2H-benzo[b][1,4]oxazin-6-yl)-N-hydroxypropenamides ( 4 a – i , 7 a – g ) targeting histone deacetylases. Three human cancer cell lines were used to test the cytotoxicity of the synthesized compounds (SW620, colon; PC-3, prostate; NCI−H23, lung cancer); inhibitory activity towards HDAC; anticancer activity; as well as their impact on the cell cycle and apoptosis. As a result, compounds 4 a – i bearing the alkyl substituents seemed to be less potent than the benzyl-containing compounds 7 a – g in all biological assays. Compounds 7 e – f were found to be the most active HDAC inhibitors with IC₅₀ of 1.498±0.020 μM and 1.794±0.159 μM, respectively. In terms of cytotoxicity and anticancer assay, 7 e and 7 f also showed good activity with IC₅₀ values in the micromolar range. In addition, the cell cycle and apoptosis of SW620 were affected by compound 7 f in almost a similar manner to that of reference compound SAHA. Docking assays were carried out for analysis the binding mode and selectivity of this compound toward 8 HDAC isoforms. Overall, our data confirmed that the inhibition of HDAC plays a pivotal role in their anticancer activity.     

新生大鼠辣椒素处理后脊髓内阿片受体的变化

本实验用受体放射自显影方法,研究了新生大鼠辣椒素皮下注射,成年后脊髓内３Ｈ－埃托菲（３Ｈ－ｅｔｏｒｐｈｉｎｅ）结合位点数的变化。发现新生鼠辣椒素处理可以引起脊髓后角浅层及中央管周围’Ｈ－ｅｔｏｒｐｈｉｎｅ结合位点明显减,其中Ⅰ、Ⅱ层自显影银粒数减少３６％左右。Ⅲ层减少２０％,中央管周围减少１１％。而前角及背外测束３Ｈ－ｅｔｏｒｐｈｉｎｅ结合位点数无显著性差异。结果表明脊髓内初级感觉传入细纤维上有阿片受体分布。本研究为探明阿片类物质在脊髓水平的痛觉调控机制提供一定的形态学依据。     

Sequences and Evolution of Human and Squirrel Monkey Blue Opsin Genes

The sequences of the entire blue opsin gene in the squirrel monkey (Saimiri boliviensis) and the five introns of the human blue opsin gene were obtained. Intron 3 of these genes contains an Alu sequence and intron 4 contains a partial mer13 sequence. A comparison of the squirrel monkey opsin sequence with published mammalian opsin sequences shows that features believed to be functionally critical are all conserved. However, the blue opsin has evolved twice as fast as rhodopsin and is only as conservative as the β globin, which has evolved at the average rate of mammalian proteins. Interestingly, the interhelical loops are, on average, actually more conservative than the transmembrane α helical regions. The introns of the blue opsin gene have evolved at the average rate of introns in primate genes. Received: 5 August 1996 / Accepted: 2 October 1996     

Ribosomal DNA and ITS-2 sequence comparisons as a tool for predicting genetic relatedness

The determination of the secondary structure of the internal transcribed spacer (ITS) regions separating nuclear ribosomal RNA genes of Chlorophytes has improved the fidelity of alignment of nuclear ribosomal ITS sequences from related organisms. Application of this information to sequences from green algae and plants suggested that a subset of the ITS-2 positions is relatively conserved. Organisms that can mate are identical at all of these 116 positions, or differ by at most, one nucleotide change. Here we sequenced and compared the ITS-1 and ITS-2 of 40 green flagellates in search of the nearest relative to Chlamydomonas reinhardtii. The analysis clearly revealed one unique candidate, C. incerta. Several ancillary benefits of the analysis included the identification of mislabelled cultures, the resolution of confusion concerning C. smithii, the discovery of misidentified sequences in GenBank derived from a green algal contaminant, and an overview of evolutionary relationships among the Volvocales, which is congruent with that derived from rDNA gene sequence comparisons but improves upon its resolution. The study further delineates the taxonomic level at which ITS sequences, in comparison to ribosomal gene sequences, are most useful in systematic and other studies. Received: 14 February 1997 / Accepted: 28 March 1997     

Cell attachment and spreading on extracellular matrix-coated beads

Parietal yolk sac cells M1536-B3 grown on cytodex 2 beads deposited an extracellular matrix on the surface of the beads. Cell-free matrix-coated beads were isolated by treatment of the cell monolayer with cytochalasin B (CB) at a concentration of 10 μg/ml of phosphate-buffered saline (PBS). The matrix when analysed by electrophoresis on polyacrylamide gels (PAGE) revealed that the major components were laminin and entactin. The matrix-coated beads were used to study the attachment, spreading, and growth of African Green monkey BSC-40, human mammary MCF-7, mouse fibroblast L929, rat liver clone 9, and rat hepatoma H-4-II-E cells in defined serum-free growth medium. The different cell lines exhibited varying responses to matrix-coated vs uncoated beads with respect to rate of attachment, spreading, and growth. One of the most consistent responses observed was the enhancement of cell spreading on matrix-coated beads. The results suggested that the matrix-coated beads will provide a readily available and valuable tool for studies on cell surface-extracellular matrix interactions and the physiological consequences of those interactions.