Effects of heptapeptide selank on genetically-based and situation-provoked symptoms of depression in behavior in WAG/Rij and Wistar rats, and in BALB/c mice

A synthetic derivative of the endogenous peptide tuftsin heptapeptide selank (Thr-Lys-Pro-Arg-Pro-Gly-Pro) possesses an anxiolytic and psychostimulant effect, and represents a working element of a new peptide drug having completed the third phase of the clinical testing as a selective anxiolytic. The neurobiochemical spectrum of selank action combines mechanisms which are characteristics of antidepressants and psychostimulants: activation of the brain monoaminergic systems, dopamine synthesis and turnover, and modulation of the tyrosine hydroxylase activity. The aim of this study was to investigate the effect of selank in a new model of inherited (genetically-based) symptoms of depression in behavior of inbred WAG/Rij rats in comparison with its effect on situation-provoked symptoms of depression in behavior of BALB/c mice. Outbred Wistar rats constituted control group. Selank in high doses (1000-2000 microg/kg), after repeated injection counteracted symptoms of depression in behavior of WAG/Rij rats (increased immobilization in the forced swimming test and decreased sucrose intake or preference (anhedonia)). Selank in low doses (100 and 300 microg/kg) after single injection reduced the duration of immobility of BALB/c mice in the forced swimming test, but did not exert significant effect after repeated injection or after injection in high doses (600 and 900 microg/kg). Selank did not affect the level of general locomotor activity and anxiety in WAG/Rij rats, and did not exert substantial effect on the behavior of control Wistar rats. The results demonstrate the presence of antidepressant component in the spectrum of neuropsychotrophyc activity of selank and indicate the higher reliability of a new experimental model of depression (the WAG/Rij rats) as compared to the standard forced swimming test for the determination of antidepressant activity of a pharmacological drug.     

Effect of selank on the main carboxypeptidases in the rat nervous tissue

Effect of single selank administration on activity of carboxypeptidase H and of phenylmethylsulfonylfluoride-inhibited carboxypeptidase - enzymes detaching arginine and lysine from C-terminus of molecules-precursors of biologically active peptides was studied. The preparation has been shown to cause long, preserved for 24 h changes of activities of these carboxypeptidases. It is suggested that the change in activity of the studied enzymes can be one of mechanisms of regulation of level of neuropeptides at action of selank.     

Increase in terminal restriction fragments of Bacteroidetes-derived 16S rRNA genes after administration of short-chain fructooligosaccharides

It is well known that short chain fructooligosaccharides (scFOS) modify intestinal microbiota in animals as well as in humans. Since most murine intestinal bacteria are still uncultured, it is difficult for a culturing method to detect changes in intestinal microbiota after scFOS administration in a mouse model. In this study, we sought markers of positive change in murine intestinal microbiota after scFOS administration using terminal restriction fragment length polymorphism (T-RFLP) analysis, which is a culture-independent method. The T-RFLP profiles showed that six terminal restriction fragments (T-RFs) were significantly increased after scFOS administration. Phylogenetic analysis of the 16S rRNA partial gene sequences of murine fecal bacteria suggested that four of six T-RFs that increased after scFOS administration were derived from the 16S rRNA genes of the class Bacteroidetes. Preliminary quantification of Bacteroidetes by real-time PCR suggests that the 16S rRNA genes derived from Bacteroidetes were increased by scFOS administration. Therefore, the T-RFs derived from Bacteroidetes are good markers of change of murine intestinal microbiota after scFOS administration.     

Genetic and physical map of the structural genes (nifH,D,K) coding for the nitrogenase complex of Rhodopseudomonas capsulata.

Functional genes coding for the structural components of the nitrogenase complex (nifH,D,K) have been cloned on an 11.8-kilobase-pair HindIII fragment of DNA from the photosynthetic bacterium Rhodopseudomonas capsulata. The genes were physically mapped by hybridization of individual cloned nif genes from Klebsiella pneumoniae and Anabaena sp. strain 7120 to Southern blots of HindIII digests of the cloned R. capsulata fragment, after introduction of HindIII sites into the latter at specified locations by insertion of Tn5. Plasmids with the 11.8-kilobase-pair HindIII fragment containing the Tn5 insertions were also used for complementation tests with chromosomal Nif- mutations and for the generation of subfragments to locate those mutations by marker rescue. The R. capsulata nifH,D,K genes comprise a single unit of expression, with the same organization and polarity as found in K. pneumoniae. However, the R. capsulata nifH,D,K fragment did not complement Nif- point mutations in the corresponding Klebsiella genes, and the Klebsiella nif genes did not function in R. capsulata.     

Big form of gamma-melanotropin-like immunoreactivity in normal human plasma

Using a radioimmunoassay for gamma 3-melanotropin (gamma 3-MSH), gamma-melanotropin-like immunoreactivity (gamma-MSH-LI) was detected in plasma extracts of normal subjects before and after metyrapone administration. Plasma gamma-MSH-LI from four normal men rose significantly after a single oral administration of metyrapone (30 mg/kg of body weight). Gel chromatographic study of plasma extract after metyrapone administration showed a single peak of gamma-MSH-LI near the elution position of mouse 16K fragment, however smaller forms of gamma-MSH-LI were not detected.     

Quantitation of DNase I sensitivity in Xenopus chromatin containing active and inactive globin, albumin and vitellogenin genes.

The disappearance of defined restriction fragments of the beta 1-globin, an albumin and the A1 vitellogenin gene was quantitated after DNase I digestion and expressed by a sensitivity factor defined by a mathematical model. Analysis of naked DNA showed that the gene fragments have similar but not identical sensitivity factors. DNase I digestion of chromatin revealed for the same gene fragments sensitivity factors differing over a much wilder range. This is correlated to the activity of the genes analyzed: the beta 1-globin gene fragment is more sensitive to DNase I in chromatin of erythrocytes compared to hepatocytes whereas the albumin gene fragment is more sensitive to DNase I in chromatin of hepatocytes. The A1 vitellogenin gene has the same DNase I sensitivity in both cell types. Comparing the DNase I sensitivity of the three genes in their inactive state we suggest that different chromatin conformations may exist for inactive genes.     

Mapping polypeptide hormone genes in the mouse: somatostatin, glucagon, calcitonin, and parathyroid hormone

Mouse-Chinese hamster hybrids segregating mouse chromosomes were analyzed by Southern hybridization techniques to map the genes for somatostatin (Smst), glucagon (Gcg), calcitonin (Calc), and parathyroid hormone (Pth). The mouse gene for somatostatin, detected on a 20-kb EcoRI fragment, is located on mouse chromosome 16. Glucagon cDNA hybridized to a 14-kb EcoRI fragment residing on chromosome 2. Calcitonin and parathyroid hormone genes, detected on 7.8-kb HindIII and 6.0-kb BamHI fragments, respectively, were on mouse chromosome 7. The calcitonin and parathyroid hormone genes appear to be part of a larger linkage group which has been conserved in mouse and man.     

Expression of c-jun and Egr-1 genes in rat liver in response to partial hepatectomy, and injection of turpentine.

In rat liver stimulated by partial hepatectomy a significant expression of Egr-1 and to less extent c-jun genes was observed within 15-30 min after the surgery. The induction is transient and vanishes within one hour. Egr-1 and c-jun were also induced in liver after subcutaneous injection of turpentine, a strong inducer of acute phase response. A maximum activation of these genes was observed within 4-9 hours after administration of turpentine. Both Egr-1 and c-jun extend the list of "immediate early" genes involved in proliferative response and constitute a group of genes inducible at the beginning of acute phase response.     

青岛文昌鱼LIM类基因Bblim同源框区的cDNA克隆、序列分析及其在胚胎发育中的表达(英文)

不同卵裂球发育命运的特化、亦即胚胎细胞的分化是动物胚胎发育的重要特征。多数胚胎细胞尽管形态特征完全一致,却具有完全不同的发育命运。预示着：在这些细胞中存在有决定发育命运的因素———决定子。本工作克隆了青岛文昌鱼ＬＩＭ类同源框基因的同源框片段。目的在于揭示决定子的分子本质。青岛近海采集性成熟的成年青岛文昌鱼,收集未受精卵、受精卵以及各个不同时期的胚胎,液氮冻存备用。分别制备总ＲＮＡ。根据其它动物ＬＩＭ类同源框基因的序列设计引物（Ｔａｂ．１）,连续进行ＲＴＰＣＲ和ＰＣＲ两次扩增。其中,原肠胚来源的第二次ＰＣＲ产物经电泳鉴定（Ｆｉｇ．１）后,酶切、克隆入质粒、测序、将该片段所在的基因命名为Ｂｂｌｉｍ基因,该片段称为Ｂｂｌｉｍ同源框。根据Ｂｂｌｉｍ基因同源框的核苷酸序列推导出其相应的氨基酸序列（Ｆｉｇ．２）,与其它ＬＩＭ类同源框基因进行比较（Ｆｉｇ．３）后,认为：Ｂｂｌｉｍ基因可归入ｌｉｍ３类基因。比较胚胎发育各个不同时期第二次ＰＣＲ产物的含量———即Ｂｂｌｉｍ基因的转录（Ｆｉｇ．４）,提示：该基因可能在受·精·后·和·原·肠·形·成·期·前·后·两个发育阶段起作用。此外,Ｂｂｌｉｍ基因的同源域与海鞘Ｈｒｌｉｍ的     

Cloning of the DNA repair genes mtcA, mtcB, uvsC, uvsD, uvsE and the leuB gene from Deinococcus radiodurans

A gene library from Deinococcus radiodurans has been constructed in the cosmid pJBFH. A 51.5-kb hybrid cosmid, pUE40, that transduced Escherichia coli HB101 from leucine dependence to independence was selected, and a 6.9-kb fragment which carried the leuB gene from D. radiodurans was subcloned into the EcoRI site of pAT153. The DNA repair genes mtcA, mtcB, uvsC, uvsD and uvsE, which code for two D. radiodurans UV endonucleases were identified by transforming appropriate repair-deficient mutants of D. radiodurans to repair proficiency with DNA derived from the gene library. Hybrid cosmid pUE50 (37.9 kb) containing an insert carrying both the mtcA and mtcB genes was selected and 5.6- and 2.7-kb DNA fragments carrying mtcA and mtcB, respectively, i.e., the genes that code for UV endonuclease alpha, were subcloned into the EcoRI site of pAT153. The three genes uvsC, uvsD and uvsE, that code for UV endonuclease beta, were all present in the 46.0-kb hybrid cosmid pUE60. The uvsE gene in a 12.2-kb fragment was subcloned into the HindIII site of pAT153 and the size of the insert reduced to 6.1 kb by deletion of a 6.7-kb fragment from the hybrid plasmid pUE62. None of the uvs genes introduced into the UV-sensitive E. coli CSR603 (uvrA-) was able to complement its repair defect. The mtcA, uvsC, uvsD and uvsE genes were found in the 52.5-kb hybrid cosmid pUE70. It is concluded that the DNA repair genes mtcA, mtcB, uvsC, uvsD and uvsE are located within an 83.0-kb fragment of the D. radiodurans genome.     

Analysis of cloned structural and regulatory genes for carbohydrate utilization in Pseudomonas aeruginosa PAO.

Five of the genes required for phosphorylative catabolism of glucose in Pseudomonas aeruginosa were ordered on two different chromosomal fragments. Analysis of a previously isolated 6.0-kb EcoRI fragment containing three structural genes showed that the genes were present on a 4.6-kb fragment in the order glucose-binding protein (gltB)-glucokinase (glk)-6-phosphogluconate dehydratase (edd). Two genes, glucose-6-phosphate dehydrogenase (zwf) and 2-keto-3-deoxy-6-phosphogluconate aldolase (eda), shown by transductional analysis to be linked to gltB and edd, were cloned on a separate 11-kb BamHI chromosomal DNA fragment and then subcloned and ordered on a 7-kb fragment. The 6.0-kb EcoRI fragment had been shown to complement a regulatory mutation, hexR, which caused noninducibility of four glucose catabolic enzymes. In this study, hexR was mapped coincident with edd. A second regulatory function, hexC, was cloned within a 0.6-kb fragment contiguous to the edd gene but containing none of the structural genes. The phenotypic effect of the hexC locus, when present on a multicopy plasmid, was elevated expression of glucokinase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydratase, and 2-keto-3-deoxy-6-phosphogluconate aldolase activities in the absence of inducer.     

Cloning, sequencing, and expression of Bacillus subtilis genes involved in ATP-dependent nuclease synthesis.

The genes encoding the subunits of the Bacillus subtilis ATP-dependent nuclease (add genes) have been cloned. The genes were located on an 8.8-kb SalI-SmaI chromosomal DNA fragment. Transformants of a recBCD deletion mutant of Escherichia coli with plasmid pGV1 carrying this DNA fragment showed ATP-dependent nuclease activity. Three open reading frames were identified on the 8.8-kb SalI-SmaI fragment, which could encode three proteins with molecular masses of 135 (AddB protein), 141 (AddA protein), and 28 kDa. Only the AddB and AddA proteins are required for ATP-dependent exonuclease activity. Both the AddB and AddA proteins contained a conserved amino acid sequence for ATP binding. In the AddA protein, a number of small regions were present showing a high degree of sequence similarity with regions in the E. coli RecB protein. The AddA protein contained six conserved motifs which were also present in the E. coli helicase II (UvrD protein) and the Rep helicase, suggesting that these motifs are involved in the DNA unwinding activity of the enzyme. When linked to the T7 promoter, a high level of expression was obtained in E. coli.     

An integrated study of acute effects of valproic acid in the liver using metabonomics, proteomics, and transcriptomics platforms

An integrated omics approach was undertaken in order to elucidate a systems biology level understanding of the acute hepatotoxcity of valproic acid (VPA). Metabonomics, proteomics and gene expression microarray platforms were employed in this systems biology study. CD-1 female pregnant mice were injected subcutaneously with 600 mg/kg VPA or vehicle control. Urine, serum, and liver tissue were collected at 6, 12, and 24 h after dosing. Principal component analysis (PCA) of the metabonomics data showed clustering of the dosed groups away from the controls for the urine samples. Looser clustering was seen in the other sample sets investigated. However, VPA administration resulted in altered glucose concentrations in urine samples at 12 and 24 h and in aqueous liver tissue extracts at 12 h after VPA administration. Proteomics studies identified two proteins, glycogen phosphorylase and amylo-1,6-glucosidase, which were increased in dosed animals relative to control. Both of these proteins are involved in converting glycogen to glucose. Examination of the expression of 20,000 liver genes did not reveal significantly altered expression at 6, 12, or 24 h after VPA exposure. The combined studies indicated a perturbation in the glycogenolysis pathway following administration of VPA.     

hydγ, a gene from Desulfovibrio vulgaris (Hildenborough) encodes a polypeptide homologous to the periplasmic hydrogenase

Downstream of the genes for the structural alpha and beta subunits of the periplasmic Desulfovibrio vulgaris (Hildenborough) hydrogenase a DNA fragment was detected with sequence homology to these genes. This fragment was cloned in Escherichia coli and the nucleotide sequence was determined. A gene was detected on the fragment with coding capacity for a 65.8 kDa polypeptide, hyd gamma. The central part of hyd gamma has an unusually high degree of homology with the alpha subunit and the C-terminal part has similarity with the beta subunit. These results strongly suggest that the three genes for hyd gamma and the alpha and beta subunits derive from one common ancestor gene. We succeeded in the identification of the translational product of this gene in E. coli, but were unable to determine the function of hyd gamma after expression in E. coli.     

Comparative stabilities in vitro and in vivo of a recombinant mouse antibody FvCys fragment and a bisFvCys conjugate.

A murine antibody FvCys fragment with a single additional cysteine residue at the C terminus of the VH domain was expressed in Escherichia coli from a modified expression plasmid containing the structural genes for the VH and VL domains derived from the anti-lysozyme hybridoma D1.3. Chemical cross-linking between the introduced sulfhydryl groups of two FvCys fragments by means of bis-maleimidohexane was used to generate a bisFvCys conjugate. The stability of the bisFvCys conjugate and an FvCys analogue that had been reacted with N-ethyl-maleimide to block the free sulfhydryl group, FvCys(BL), were compared after 125I-labeling. The bisFvCys conjugate was completely stable to incubation in solution at 37 degrees C for 24 h whereas only 60% of the FvCys(BL) fragment remained soluble. After i.v. administration to normal Wistar rats, both Fv proteins were rapidly cleared from the circulation with biphasic kinetics that were best fitted to a two-compartment open pharmacokinetic model. The alpha-phase half-life of the bisFvCys conjugate, 0.32 h, was significantly longer than that of the FvCys(BL) fragment, 0.15 h (p less than 0.001) whereas there was no significant difference between the beta-phase half-lives, 1.4 to 1.6h. No chain cleavage or covalent attachment to serum protein was detected by SDS-PAGE analysis of serum samples. However, gel permeation HPLC revealed that both Fv proteins associated with serum proteins in vivo and in vitro.