bluuebunny/crossref_metadata_2025 huggingface dataset:
In vitro Anthelmintic Activity of Aqueous and Ethanolic Extract of Senna italica (Caesalpiniaceae) on Three-stages of Haemonchus contortus
Authors: Yongwa Gilbert , Ngnoda Belga François Ngnodandi François , Ndjonka Dieudonné , Saotoing Pierre
Published: April 2020

A phytochemical screening and in vitro anthelminthic activity of aqueous and ethanolic extract of Senna italica on Haemonchus contortus were conducted. Polyphenol, tannin and flavonoid contents were determined by using gallic acid and rutin. egg hatching inhibition test was carried out on fresh eggs; larval mortality test was conducted on infective larvae (L3) and adult worm mortality test was conducted. Eggs, larvae and adults worms were incubated in aqueous and ethanolic extract of S. italica at different concentrations (0.1; 0.3; 0.5; 0.7 and 1 mg/mL). All extracts of plants showed an effect on all stages of H. contortus with high efficiency variations depending on the dose used. The inhibition of eggs hatching rate increased from 8.67±1.53% to 65.67±1.15% and from 24.67±1.53% to 80±1.73% respectively for aqueous and ethanolic extract of S. italic. The larval mortality rate increased from 12.22±2.34% to 56.67±4.9% after 24 h and from 45.28±4.11% to 91.25±3.73% after 48 h for aqueous extract and from 30.07±2.84% to 70.83±6.31% after 24 h and from 48.79±3.73% to 96.25±4.79% after 48 h for ethanolic extract. After 24 h of exposure to aqueous and ethanolic extract of S. italica, the adult mortality rate varies from 55.56±9.62% to 83.33±9.62% for aqueous extract and from 61.11±9.62% to 88.89±9.62%.These in vitro results confirm the use of S. italica in traditional medicine.
Changes in nuclear content of protein conjugate histone H2A—ubiquitin during rooster spermatogenesis
Authors: Agell Neus , Chiva Manel , Mezquita Cristóbal
Published: July 2002

Electrophoretic analysis of acid‐soluble chromosomal proteins isolated from rooster testis cell nuclei at different stages of spermatogenesis, revealed that the nuclear content of a protein identified by its solubility, electrophoretic mobility and amino acid analysis as the protein conjugate histone H2A—ubiquitin (uH2A, A24) changed markedly from meiotic cells to late spermatids. The protein was not detectable in tetraploid primary spermatocytes; it was present in 1.7% of the total amount of nucleosomal core histones in early spermatids and reached its maximum level (3.5% and 11%) at the end of spermiogenesis, when histones are replaced by the protamine galline.
Specific binding of chartreusin, an antitumor antibiotic, to DNA
Authors: Uramoto Masakazu , Kusano Tomonobu , Nishio Tomoko , Isono Kiyoshi , Shishido Kazuo , Ando Tadahiko
Published: July 2002

It was evidenced that the binding of the agent is not at random and correlates to the sequence on DNA stretch.
Phosphorylethanolamine ‐ the major constituent of the phosphomonoester peak observed by 31P‐NMR on developing dog brain
Authors: Gyulai Laszlo , Bolinger Lizann , Leigh John S. , Barlow Clyde , Chance Britton
Published: July 2002

31P‐NMR spectra of newborn dog brains exhibit a prominent phosphomonoester (PME) peak (6.78 ± SD. 0.05 ppm from phosphocreatine peak), similar to those of human neonates. Studies were undertaken to identify the chemical constituents of this peak. Brains of puppies were funnel frozen for methanol‐HCl‐perchloric acid extraction after in vivo 31P‐NMR spectra were taken. The pK of the major component of the PME region in the NMR spectrum of extract was 5.4, corresponding to that of phosphorylethanolamine (PEt). Addition of PEt increased the major peak on the PME region over a wide range of pH, while addition of phosphorylcholine or ribose 5‐phosphate yielded distinct peaks. We suggest that the major constituent of phosphomonoester peak of 31P‐NMR spectra of newborn dog brain is phosphorylethanolamine. Biochemical mechanisms relevant to changes of phosphorylethanolamine during brain development are discussed.
Nicotine‐evoked disassembly of cortical actin filaments in adrenal chromaffin cells
Authors: Cheek Timothy R. , Burgoyne Robert D.
Published: July 2002

Using rhodamine‐phalloidin staining it was found that actin filaments are concentrated in the cortex of resting chromaffin cells. Cortical actin filaments were disassembled 15 s after stimulation by nicotine and had reassembled 30 s later. Actin filament disassembly following nicotinic stimulation was also detected using the DNase I inhibition assay. Disassembly was independent of external calcium, insensitive to trifluoperazine and was not elicited by high K+, muscarinic agonists or phorbol ester. Disassembly of cortical actin filaments may allow access of secretory granules to exocytotic sites and act in conjunction with a rise in intracellular free calcium to bring about the full secretory response due to nicotinic agonists.
Nucleotide sequence of the PR‐1 gene of Nicotiana tabacum
Authors: Ohshima Masahiro , Matsuoka Makoto , Yamamoto Naoki , Tanaka Yoshiyuki , Kano-Murakami Yuriko , Ozeki Yoshihiro , Kato Akira , Harada Naoki , Ohashi Yuko
Published: July 2002

A gene encoding one of the pathogenesis‐related proteins, PR1a, and two related pseudogenes were isolated from Nicotiana tabacum. The cloned PR1a gene (pPR‐γ) and one of the pseudogenes (pPR‐α) were sequenced and found to have similar structures. The sequence of pPR‐γ was quite similar to that of the cDNA clone of PR1a. The plasmid pPR‐γ did not contain an intron and had a typical promoter sequence in the 5′‐flanking region.
Electron microscopy and image analysis of the multicatalytic proteinase
Authors: Baumeister W. , Dahlmann B. , Hegerl R. , Kopp F. , Kuehn L. , Pfeifer G.
Published: July 2002

On electron micrographs, negatively stained multicatalytic proteinase molecules are viewed end‐on (ring shaped) or side‐on (rectangular shaped). For aurothioglucose, ammonium molybdate‐ and phosphotungstate‐stained molecules, the dimensions measured are consistent. In contrast, uranyl acetate‐staining reveals ring‐shaped particles which vary in diameter between 12 and 16 nm. This is due to a partial collapse and substantial flattening of the structure. Digital image analysis of side‐on views of the particles reveals a tripartite, reel‐shaped structure. Within the ring‐like, end‐on projections of ammonium molybdate‐stained molecules six local centres of mass can be discerned; their position appears to depart, however, from a true six‐fold symmetry.
Spontaneous polymerization of the antibiotic peptide magainin 2
Authors: Urrutia Raul , Cruciani Ricardo A. , Barker Jeffery L. , Kachar Bechara
Published: July 2002

We describe here the ability of the magainin 2 peptide to assemble spontaneously into characteristic 13‐nm diameter filaments having a 30 nm periodic helical substructure. Optimal conditions for extensive polymerization into filaments of several hundred microns required low pH and high ionic strength. Polymerization of the magainin 2 peptide may be involved in its recently described in vitro membrane‐disrupting and antibiotic activities.
Differential activities of two distinct endothelin family peptides on ileum and coronary artery
Authors: Ishida Norio , Tsujioka Keiji , Tomoi Masaaki , Saida Kaname , Mitsui Youji
Published: July 2002

A synthesized mouse vasoactive intestinal contractor peptide, which belongs to a novel member of the endothelin family, induced a prolonged contraction in mouse ileum as well as porcine coronary artery in vitro. Studies comparing the effects of vasoactive intestinal contractor and endothelin on different tissues revealed that the maximum ileum contraction of vasoactive intestinal contractor was much higher than that of endothelin in both guinea pig and mouse systems, but that the vasoconstriction activity of vasoactive intestinal contractor was weaker than that of endothelin in porcine artery. These resuts show that vasoactive intestinal contractor might be a novel gastrointestinal hormone.
A cysteine proteinase cDNA from Trypanosoma brucei predicts an enzyme with an unusual C‐terminal extension
Authors: Mottram Jeremy C. , North Michael J. , Barry J.David , Coombs Graham H.
Published: July 2002

A cDNA for a Trypanosoma brucei cysteine proteinase has been cloned and sequenced. The deduced protein can be divided into four domains, based on homologies with other cysteine proteinases: the pre‐, pro‐ and central regions show considerable homology to the cathepsin L class of mammalian enzymes, whilst the long C‐terminal extension distinguishes the trypanosome enzyme from all mammalian cysteine proteinases reported. This 108 amino acid extension, which includes 9 contiguous prolines near the junction with the central domain, appears likely to be processed in part to produce the mature enzyme, and may be involved in targetting the protein within the cell. The trypanosome genome contains more than 20 copies of the cysteine proteinase gene arranged in a long tandem array.
Calcyclin is a calcium and zinc binding protein
Authors: Filipek Anna , Heizmann Claus W. , Kúznicki Jacek
Published: July 2002

Calcyclin, a cell cycle regulated protein, was recently purified from Ehrlich ascites tumour (EAT) cells and shown to be a calcium binding protein. Here we show that calcyclin monomer and dimer also bind zinc ions. Zinc binding sites seem to be different from calcium binding sites since: preincubation with Ca2+ lacks effect on the binding of Zn2+, and Ca2+ (but not Zn2+) increases tyrosine fluorescence intensity. Binding of Zn2+ reduces the extent of the conformational changes induced by Ca2+, and seems to affect Ca2+‐binding. The data suggest that Ca2+‐ and Zn2+‐ might trigger the biological activity of calcyclin.
Endothelin stimulates hypertrophy and contractility of neonatal rat cardiac myocytes in a serum‐free medium
Authors: Suzuki Takahiko , Hoshi Hiroyoshi , Mitsui Youji
Published: July 2002

The effect of endothelin (ET) on rat cardiac myocytes cultured in a serum‐free, defined medium was determined. ET stimulated cardiac myocyte hypertrophy in a dose‐dependent manner as determined by the protein synthesis and cell surface area. Since the myocyte hypertrophy was abolished by H‐7, a protein kinase C inhibitor, ET‐receptor mediated protein kinase C activation may be involved in cardiac myocyte hypertrophy. At the same time, ET also stimulated myocyte contractility in this medium, and this stimulatory effect was inhibited by nicardipine. This result indicates that the influx of extracellular calcium ion is necessary for the stimulation of contractility induced by ET.
Cerulenin, an inhibitor of lipid synthesis, blocks vesicular stomatitis virus RNA replication
Authors: Pérez Luis , Carrasco Luis
Published: July 2002

The replication of genomex of animal viruses in the cytoplasm of susceptible cells is usually coupled to specialized membrane structures. The inhibitor of lipid synthesis cerulenin blocks the formation of vesicular stomatitis virus polypeptides when added to cells soon after virus entry, but has much less effect on viral translation, or the acylation of the glycoprotein G, when cerulenin is added late during infection. By contrast, cerulenin powerfully blocks viral RNA synthesis or the incorporation of glycerol into lipids when present at any time after VSV‐infection. These findings suggest that the systhesis of VSV RNA is dependent on continuous synthesis of lipids.
How does protein synthesis give rise to the 3D‐structure?
Authors: Ptitsyn O.B.
Published: July 2002

The recent experimental data on stages and kinetic intermediates in protein folding are reviewed. It is emphasized that these data are consistent with the ‘framework model’ proposed by the author in 1973. The model implies that protein folds by stage mechanism (secondary structure — molten globule state — native state) in such a way that the results of previous stages are not reconsidered in subsequent ones. Arguments are presented that both these hypotheses and available experimental data do not contradict the assumption that native structures of at least small proteins are nevertheless under thermodynamic rather than kinetic control, i.e. correspond to global minima of free energy.
Identification of hydrophobic fragments of α1‐antitrypsin and Cl protease inhibitor in human bile, plasma and spleen
Authors: Johansson Jan , Gröndal Staffan , Sjövall Jan , Jörnvall Hans , Curstedt Tore
Published: July 2002

Hydrophobic peptides were isolated from the phospholipid fraction of human bile, plasma and spleen by exclusion chromatography in organic solvents. From plasma, the activation peptide or Cl protease inhibitor was recovered, from spleen the activation peptide of α1‐antitrypsin, and from bile, both these peptides, as well as a fragment generated by proteolytic cleavage of α1‐antitrypsin six residues N‐terminal of the Pl–Pl ' peptide bond. Cleavages in this region inactivate antiproteases but have previously not been reported to occur in vivo. These peptides in human bile may reflect physiological actions in regulation of antiproteolytic activity or bile secretion processes, and/or be of importance for the physicochemical state of cholesterol, phospholipids and bile acids in bile.
Selective binding of anchorin CII (annexin V) to type II and X collagen and to chondrocalcin (C‐propeptide of type II collagen) Implications for anchoring function between matrix vesicles and matrix proteins
Authors: Kirsch Thorsten , Pfäffle Michael
Published: July 2002

Anchorin CII is a collagen binding protein of the annexin family associated with plasma membranes of chondrocytes, osteoblasts, and many other cells. As a major, constituent of cartilage‐derived matrix vesicles it has been shown to bind to native type II and X collagen. In accordance with this observation, here we show the localization of anchorin CII in the extracellular matrix of calcifying cartilage in the fetal human growth plate, and that it was restricted to the chondrocyte surface in proliferating and resting cartilage. Furthermore, we present evidence, using a slot blot assay, that anchorin CII not only binds to native type II and X collagen, but also to chondrocalcin, the carboxy‐terminal extension of type II procollagen in a calcuim‐independent manner, Pepsin digestion of type II collagen results in loss of anchorin CII binding, confirming our previous notion that the telopeptide region of type II collagen carries anchorin CII binding sites.
Singlet oxygen induces collagenase expression in human skin fibroblasts
Authors: Scharffetter-Kochanek Karin , Wlaschek Meinhard , Briviba Karlis , Sies Helmut
Published: July 2002

Singlet oxygen generated in a dark reaction by thermodissociation of an endoperoxide (NDPO2) elicits an increase in mRNA of interstitial collagenase (MMP‐1) in cultured human fibroblasts. The effect is enhanced in deuterium oxide‐based medium and is abolished in the presence of non‐toxic doses of sodium azide. In contrast, the mRNA level of the tissue inhibitor of metalloproteinases (TIMP‐1) remains unaltered under these experimental conditions. These observations support the suggestion that an unbalanced synthesis of collagenase and TIMP reported to occur following UV‐A irradiation or during inflammatory conditions may be mediated by singlet oxygen.
Cloning and functional expression of a tetrabenazine sensitive vesicular monoamine transporter from bovine chromaffin granules
Authors: Howell Mike , Shirvan Anat , Stern-Bach Yael , Steiner-Mordoch Sonia , Strasser Jane E. , Dean Gary E. , Schuldiner Shimon
Published: July 2002

Using oligonucleotide primers derived from the vesicular monoamine transporters sequences, a cDNA predicted to encode the bovine chromaffin granule amine transporter has been cloned (b‐VMAT2). Surprisingly, its structure is more similar to the rat brain transporter (VMAT2), than to the rat adrenal counterpart (VMAT1). Unlike rat VMAT1, bovine VMAT2 appears to be expressed both in the adrenal medulla and the brain, as judged by Northern analysis. After modification/deletion of the seven amino acids at the N‐terminus of the protein it was expressed in a functional form. The order of affinity of the bovine VMAT2 transporter to substrates is: serotonin>dopamine = norepinephrine>epinephrine. Also, the recombinant bovine adrenal transporter is highly sensitive to tetrabenazine, in sharp contrast to the rat adrenal transporter. The findings indicate, therefore, a clear species variation in which structure and function of the bovine adrenal transporter resemble the rat brain protein, while its tissue distribution is distinct from both types of rat proteins. In addition, the predicted protein sequence is identical to the experimentally determined N‐terminus sequence of the purified vesicular amine transporter [Stern‐Bach et al. (1992) Proc. Natl. Acad. Sci. USA 89, 9730‐9733].
Does a synthetic peptide containing the leucine‐zipper domain of c‐myb form an α‐helical structure in solution?
Authors: Ebneth Andreas , Adermann Knut , Wolfes Heiner
Published: July 2002

We have examined a synthetic peptide containing the putative leucine zipper domain of the chicken c‐myb proto‐oncogene using circular dichroism (CD) spectroscopy. The peptide adopts an α‐helical structure only at low temperatures and in the presence 2,2,2‐trifluoroethanol.
Vaccinia virus DNA topoisomerase I preferentially removes positive supercoils from DNA
Authors: Fernandez-Beros Maria-Elena , Tse-Dinh Yuk-Ching
Published: July 2002

Type I DNA topoisomerases homologous to Escherichia coli topoisomerase I normally only remove negative supercoils from DNA. Topoisomerases I from various eukaryotes share sequence homology and remove both positive and negative supercoils from DNA. Here we report that vaccinia virus topoisomerase I has significant difference in substrate preference from the other homologous type I topoisomerases. Vaccinia virus topoisomerase I shows a definite preference for removal of positive supercoils. In contrast, topoisomerase I from human, wheat germ and Saccharomyces cerevisiae has little preference between positive and negative supercoils. The vaccinia enzyme may have evolved for functions required for optimal viral growth.
And still they are moving… Dynamic properties of caveolae
Authors: Kurzchalia Teymuras V. , Parton Robert G.
Published: July 2002

Caveolae are structures found on the surface of many mammalian cells. In the last few years the biogenesis and the function of these organelles have been intensively investigated but many challenging questions remain. One of these is whether caveolae are statically attached to the cytoplasmic surface of the plasma membrane or are moving to other intracellular organelles. Also the cycling of the caveolar coat component, VIP21‐caveolin, is a subject of intensive discussion. The solution to these problems could give an insight into the understanding of caveolar function.
P‐glycoprotein is localized in caveolae in resistant cells and in brain capillaries
Authors: Demeule Michel , Jodoin Julie , Gingras Denis , Béliveau Richard
Published: July 2002

A significant proportion of P‐glycoprotein (P‐gp) and caveolin was co‐localized in caveolae isolated from resistant (CHRC5) cells overexpressing P‐gp and from drug‐sensitive Chinese hamster ovary cells (AuxB1). The proportion of P‐gp and caveolin associated with caveolar microdomains was higher in CHRC5 cells grown in the presence of P‐gp substrates (cyclosporin A or colchicine) than in untreated CHRC5 cells. Co‐immunoprecipitation of P‐gp and caveolin from CHRC5 lysates suggests that there is a physical interaction between them. Furthermore, co‐localization of P‐gp and caveolin was found in caveolae from brain capillaries, indicating that this association also takes place in vivo.
Olive oil phenolics are dose‐dependently absorbed in humans
Authors: Visioli Francesco , Galli Claudio , Bornet Francis , Mattei Alissa , Patelli Rossana , Galli Giovanni , Caruso Donatella
Published: July 2002

Olive oil phenolic constituents have been shown, in vitro, to be endowed with potent biological activities including, but not limited to, an antioxidant action. To date, there is no information on the absorption and disposition of such compounds in humans. We report that olive oil phenolics, namely tyrosol and hydroxytyrosol, are dose‐dependently absorbed in humans after ingestion and that they are excreted in the urine as glucuronide conjugates. Furthermore, an increase in the dose of phenolics administered increased the proportion of conjugation with glucuronide.
Interaction of a novel cysteine and histidine‐rich cytoplasmic protein with galectin‐3 in a carbohydrate‐independent manner
Authors: Menon Rajesh P. , Strom Molly , Hughes R.Colin
Published: July 2002

We have used the yeast two‐hybrid system to search for cytoplasmic proteins that might assist in the intracellular trafficking of the soluble β‐galactoside‐binding protein, galectin‐3. We utilised as bait murine full‐length galectin‐3 to screen a murine 3T3 cDNA library. Several interacting clones were found to encode a partial open reading frame and a full‐length clone was obtained by rapid amplification of cDNA ends methodology. In various assays in vitro the novel protein was shown to bind galectin‐3 in a carbohydrate‐independent manner. The novel protein contains an unusually high content of cysteine and histidine residues and shows significant sequence homologies with several metal ion‐binding motifs present in known proteins. Confocal immunofluorescence microscopy of permeabilised 3T3 cells shows a prominent perinuclear, as well as cytoplasmic, localisation of the novel protein.
Amyloids protect the silkmoth oocyte and embryo
Authors: Iconomidou Vassiliki A. , Vriend Gert , Hamodrakas Stavros J.
Published: July 2002

Chorion is the major component of silkmoth eggshell. More than 95% of its dry mass consists of proteins that have remarkable mechanical and chemical properties protecting the oocyte and the developing embryo from a wide range of environmental hazards. We present data from electron microscopy (negative staining and shadowing), X‐ray diffraction and modeling studies of synthetic peptide analogues of silkmoth chorion proteins indicating that chorion is a natural amyloid. The folding and self‐assembly models of chorion peptides strongly support the β‐sheet helix model of amyloid fibrils proposed recently by Blake and Serpell [Structure 4 (1996) 989–998].
Differential skeletal muscle expression of myostatin across teleost species, and the isolation of multiple myostatin isoforms
Authors: Roberts Steven B , Goetz Frederick W
Published: July 2002

Two myostatin (MSTN) isoforms were isolated from brook trout with 92% identity in corresponding regions at the nucleotide level. One isoform was isolated from muscle and brain and the second from ovarian tissue. To our knowledge this is the first time two MSTN isoforms have been isolated from a given vertebrate species. Within the brain, MSTN transcripts were localized to the optic lobes, hindbrain, and hypothalamus. In the trout ovary, MSTN transcripts were upregulated at ovulation in several females. MSTN cDNA fragments were also isolated from several other fish species and differential expression of MSTN among muscle fiber types was observed.
Role of AP2/EREBP transcription factors in gene regulation during abiotic stress
Authors: Kizis Dimosthenis , Lumbreras Victoria , Pagès Montserrat
Published: July 2002

Crop plants are exposed to many types of abiotic stress during their life cycle. Water deficit derived from drought, low temperature or high salt concentration in the soil, is one of the most common environmental stresses that affects growth and development of plants through alterations in metabolism and gene expression. Adaptation to these conditions may involve passive tolerance or active homeostatic mechanisms for maintaining water balance. Active responses occur at different levels in the plant and may represent a concomitant protection against other types of stress such as pathogen attack. Many morphological and physiological adaptations to water stress are under the control of the plant hormone abscisic acid and involve specific activation of target genes that in one way or another protect cells against water deficit or participate in the regulation of the drought response. Here, we discuss recent advances in our understanding of drought adaptation mediated by specific changes in gene expression and the role of AP2/EREBP nuclear factors in these processes.
Domain II of m‐calpain is a Ca2+‐dependent cysteine protease
Authors: Hata Shoji , Sorimachi Hiroyuki , Nakagawa Kazuhiro , Maeda Tatsuya , Abe Keiko , Suzuki Koichi
Published: July 2002

Calpain, a Ca2+‐dependent cytosolic cysteine protease, proteolytically modulates specific substrates involved in Ca2+‐mediated intracellular events, such as signal transduction, cell cycle, differentiation, and apoptosis. The 3D structure of m‐calpain, in the absence of Ca2+, revealed that the two subdomains (domains IIa and IIb) of the protease domain (II) have an ‘open’ conformation, probably due to interactions with other domains. Although the presence of an EF‐hand structure was once predicted in the protease domain, no explicit Ca2+‐binding structure was identified in the 3D structure. Therefore, it is predicted that if the protease domain is excised from the calpain molecule, it will have a Ca2+‐independent protease activity. In this study, we have characterized a truncated human m‐calpain that consists of only the protease domain. Unexpectedly, the proteolytic activity was Ca2+‐dependent, very weak, and not effectively inhibited by calpastatin, a calpain inhibitor. Ca2+‐dependent modification of the protease domain by the cysteine protease inhibitor, E‐64c, was clearly observed as a SDS–PAGE migration change, indicating that the conformational changes of this domain are a result of Ca2+ binding. These results suggest that the Ca2+ binding to domain II, as well as to domains III, IV, and VI, is critical in the process of complete activation of calpain.
Monocyte chemoattractant protein‐2 can exert its effects through the MCP‐1 receptor (CC CKR2B)
Authors: Yamagami Shinsuke , Tanaka Hiroko , Endo Noriaki
Published: July 2002

We studied the activities of the monocyte chemoattractant proteins MCP‐1, MCP‐2 and MCP‐3 on human embryonic kidney 293‐EBNA cells transfected with the MCP‐1 receptor (CC CKR2B). At 4 nM, MCP‐2 induced a Ca2+ influx which was as potent as that with MCP‐1 at 4 nM, although the increase by MCP‐2 became saturated at higher concentrations. In addition, all three MCPs showed dose‐dependent inhibition of adenylyl cyclase activity stimulated by forskolin (IC50 values: 0.3 nM for MCP‐1, 7 nM for MCP‐2, and 1.5 nM for MCP‐3). In conclusion, our data indicate that MCP‐2 can exert its effects through the MCP‐1 receptor, CC CKR2B.
Expression of a lipocalin in Pichia pastoris: secretion, purification and binding activity of a recombinant mouse major urinary protein
Authors: Ferrari E , Lodi T , Sorbi R.T , Tirindelli R , Cavaggioni A , Spisni A
Published: July 2002

The proteins of the mouse major urinary protein complex (MUP), members of the lipocalin family, bind volatile pheromones and interact with the vomeronasal neuroepithelium of the olfactory system. We report the expression of a MUP protein using its native signal sequence for secretion in the methylotrophic yeast, Pichia pastoris. Mature recombinant MUP (rMUP) is secreted at a concentration of 270 mg/l in minimal medium and it is isolated from the culture supernatant by one step ion‐exchange chromatography in a nearly pure form. Binding activity, tested with an odorant molecule which displays high affinity for native MUP, indicates that rMUP has a behavior similar to the native one. This finding suggests that the protein, and in particular its hydrophobic binding pocket, is properly folded.
Regulation of capsular polysialic acid biosynthesis by N‐acetyl‐d‐mannosamine, an intermediate of sialic acid metabolism
Authors: Revilla-Nuin Beatriz , Rodriguez-Aparicio Leandro B , Ferrero Miguel A , Reglero Angel
Published: July 2002

N‐Acetyl‐d‐mannosamine (ManNAc) is a specific substrate for the synthesis of N‐acetylneuraminic acid, the essential precursor of bacterial capsular polysialic acid (PA). When Escherichia coli K92 used ManNAc as a carbon source, we observed a dramatic reduction (up to 90%) in in vivo PA production. Experiments in which the carbon source was changed revealed that the maximal inhibitory effect occurred when this sugar was present in the medium before the logarithmic phase of bacterial growth had started. Enzymatic analysis revealed that high concentrations of ManNAc‐6‐phosphate inhibit NeuAc lyase, the enzyme that synthesizes NeuAc for PA biosynthesis in E. coli. These results indicate that ManNAc‐6‐phosphate is able to regulate NeuAc lyase activity and modulate the PA synthesis.
Identification of a cytoplasmic motif in the erythropoietin receptor required for receptor internalization
Authors: Levin Iris , Cohen Jacob , Supino-Rosin Lia , Yoshimura Akihiko , Watowich Stephanie S , Neumann Drorit
Published: July 2002

Erythropoietin (EPO) promotes the viability, proliferation and differentiation of mammalian erythroid progenitor cells via its specific cell surface receptor. The EPO receptor (EPO‐R) is a member of the cytokine receptor superfamily and is comprised of one identified subunit which homodimerizes upon ligand binding. To study the role of the intracellular domain of the EPO‐R in the endocytosis of EPO, we compared the rate and extent of 125I‐EPO endocytosis by wild type (wt) EPO‐R and five cytoplasmically truncated EPO‐Rs: 1–251 EPO‐R, 1–257 EPO‐R, 1–267 EPO‐R, 1–276 EPO‐R and 1–306 EPO‐R which contain 4, 10, 20, 29 or 59 amino acids of the cytoplasmic domain, respectively. We also studied an EPO‐R mutant (PB) which lacks amino acid residues 281–300 of the cytoplasmic domain. The experiments were conducted in COS 7 cells transfected with the EPO‐R cDNAs and in Ba/F3 cells stably expressing the wt EPO‐R, 1–251 or 1–257 EPO‐R. Cells expressing wt EPO‐R, PB EPO‐R (Δ281–300), 1–276 EPO‐R or 1–306 EPO‐R internalized approximately 50% of 125I‐EPO bound to the cell surface, while cells expressing 1–251, 1–257 or 1–267 EPO‐R internalized only 25% of the bound 125I‐EPO. The steady‐state expression levels of these latter receptors on the cell surface were typically 2–5‐fold higher than wt EPO‐R. Our data indicate that amino acid residues 267–276 (FEGLFTTHK) of the EPO‐R cytoplasmic domain may have a role in receptor internalization. Metabolic labeling experiments suggest that in transiently transfected COS 7 cells most of the wt EPO‐R and 1–257 EPO‐Rs do not exit the ER and may be degraded there. The half‐life of both receptors was essentially similar and was in the range of 1 h. In Ba/F3 cells the mature Golgi processed 1–257 EPO‐R was more stable than the corresponding form of the wt EPO‐R, possibly contributing to its higher cell surface expression.
Hyperleptinaemia in mice induced by administration of the tyrosine hydroxylase inhibitor α‐methyl‐p‐tyrosine
Authors: Rayner D.Vernon , Simón Edurne , Duncan Jacqueline S. , Trayhurn Paul
Published: July 2002

α‐methyl‐p‐tyrosine (αMPT), an inhibitor of tyrosine hydroxylase, was administered to mice to block noradrenaline synthesis. Ten hours after injection of αMPT there was a 6‐fold increase in plasma leptin. The level of ob mRNA in epididymal white adipose tissue was also increased, but UCP1 mRNA in brown fat fell. In contrast to lean mice, ob mRNA in white fat of ob/ob mice was not increased by αMPT. αMPT raised plasma leptin in fasted as well as fed mice. Hyperleptinaemia was attenuated by treatment with a β3‐adrenoceptor agonist. Inhibition of noradrenaline synthesis leads to the rapid induction of hyperleptinaemia; it is suggested that sympathetic tone plays a pivotal role in regulating leptin production.
Increased caveolin‐3 levels in mdx mouse muscles
Authors: Vaghy Pal L. , Fang Jin , Wu Wenrong , P. Vaghy Laszlo
Published: July 2002

The density of skeletal muscle caveolae is increased in Duchenne muscular dystrophy and its genetic homologue, the mdx mouse. This structural change is significant as it may indicate muscle regeneration. We identified in mdx mouse tibialis anterior muscles significantly increased levels of caveolin‐3, the chief protein in muscle caveolae, and reduced levels of neuronal nitric oxide synthase, an enzyme regulated by caveolin‐3. Similar changes occurred in the corresponding mRNA levels. These data suggest that induction of caveolin‐3 occurs and this may at least partly be responsible for increased number of caveolae, altered nNOS‐caveolin cycle, and regeneration of dystrophic muscles.
Genomic organization and promoter function of the mouse uncoupling protein 2 (UCP2) gene
Authors: Yamada Masanobu , Hashida Tetsu , Shibusawa Nobuyuki , Iwasaki Toshiharu , Murakami Masami , Monden Tsuyoshi , Satoh Teturou , Mori Masatomo
Published: July 2002

We cloned and characterized the mouse uncoupling protein 2 (UCP2) gene and its promoter region. The gene spans approximately 6.3 kb and contains eight exons and seven introns. Two short exons are located in the 5′ untranslated region, and each of the remaining exons encodes one of the transmembrane domains. 3′‐RACE analysis showed that a polyadenylation signal 257 bp downstream from the stop codon was functional. Primer extension analysis indicated a single transcriptional start site 369 bp upstream from the translational start site. The promoter region lacks both TATA and CAAT boxes but is GC‐rich. A construct containing 1250 bp of the promoter region showed significant activity in all 6 cell lines examined, and the region between −160 and −678 bp exhibited strong positive regulatory activity. These features of the UCP2 gene are different from those of the UCP1 gene and may contribute to its ubiquitous expression.
New muteins of RNase A with enhanced antitumor action
Authors: Cafaro Valeria , Bracale Aurora , Di Maro Antimo , Sorrentino Salvatore , D'Alessio Giuseppe , Di Donato Alberto
Published: July 2002

Monomeric bovine pancreatic RNase A has been transformed into a dimeric ribonuclease with antitumor activity (Di Donato, A., Cafaro, V. and D'Alessio, G. (1994) J. Biol. Chem. 269, 17394–17396). This was accomplished by replacing the residues located in the RNase chain at positions 19, 28, 31, and 32, with proline, leucine, and two cysteine residues, respectively, i.e. those present at identical positions in the subunit of bovine seminal RNase, a dimeric RNase of the pancreatic‐type superfamily, endowed with a powerful antitumor action. However, as an antitumor agent this mutant dimeric RNase A is not as powerful as seminal RNase. We report here site‐directed mutagenesis experiments which have led to the identification of two other amino acid residues, glycine 38 and 111, whose substitution in the polypeptide chain of the first generation dimeric mutant of RNase A, is capable of conferring to the mutein the full cytotoxic activity characteristic of native seminal RNase.
Human Prop‐1: cloning, mapping, genomic structure
Authors: Duquesnoy Philippe , Roy Anne , Dastot Florence , Ghali Isis , Teinturier Cécile , Netchine Irène , Cacheux Valère , Hafez Mona , Salah Nermine , Chaussain Jean-Louis , Goossens Michel , Bougnères Pierre , Amselem Serge
Published: July 2002

Prop‐1 is a newly isolated pituitary‐specific paired‐like homeodomain transcription factor whose cDNA sequence is well known in mouse. To study its involvement in human combined pituitary hormone deficiency (CPHD), we have isolated the human cDNA ortholog and determined the exon/intron organization and chromosomal localization of the human gene. A Prop‐1 defect was characterized in three CPHD families. One missense mutation (R73C) involves a residue conserved in 95% of the more than 400 homeodomain proteins so far identified; in vitro splicing assays demonstrated the functional importance of the second defect, whereas the remaining mutation is a frameshift. Given the disease phenotype documented in the patients, these data, which will facilitate molecular investigations in other patients, demonstrate the crucial role of Prop‐1 in the proper development of somatotrophs, lactotrophs, thyreotrophs and gonadotrophs.
Wounding and chemicals induce expression of the Arabidopsis thaliana gene Thi2.1, encoding a fungal defense thionin, via the octadecanoid pathway
Authors: Bohlmann Holger , Vignutelli Alberto , Hilpert Bernadette , Miersch Otto , Wasternack Claus , Apel Klaus
Published: July 2002

In seedlings of Arabidopsis thaliana the thionin gene Thi2.1 is inducible by methyl jasmonate, wounding, silver nitrate, coronatine, and sorbitol. We have used a biochemical and genetic approach to test the signal transduction of these different inducers. Both exogenously applied jasmonates and jasmonates produced endogenously upon stress induction, lead to GUS expression in a Thi2.1 promoter‐uidA transgenic line. No GUS expression was observed in a coi1 mutant background which lacks jasmonate perception whereas methyl jasmonate and coronatine but not the other inducers were able to overcome the block in jasmonic acid production in a fad3‐2 fad7‐2 fad8 mutant background. Our results show conclusively that all these inducers regulate Thi2‐1 gene expression via the octadecanoid pathway.
Identification and cloning of a novel cellular protein Naf1, Nef‐associated factor 1, that increases cell surface CD4 expression1
Authors: Fukushi Masaya , Dixon Jill , Kimura Tohru , Tsurutani Naomi , Dixon Michael J , Yamamoto Naoki
Published: July 2002

The nef gene of human and simian immunodeficiency virus is a key factor in acquired immunodeficiency syndrome pathogenesis and virus replication. Several Nef‐induced phenomena, including the down‐regulation of CD4 molecule, have been previously reported. In this study, we have identified and cloned a novel cellular protein Naf1 (Nef‐associated factor 1), which associated with Nef in the yeast two‐hybrid system and pull‐down assay. The Naf1 gene generates two isoforms (Naf1α and β) containing four coiled‐coil structures. The Naf1 mRNA is ubiquitously expressed in human tissues with strong expression in peripheral blood lymphocytes and spleen. Naf1 overexpression increased cell surface CD4 expression. Nef suppressed this Naf1‐induced augmentation of CD4 expression, providing a novel mode of Nef action in CD4 down‐regulation.
STABLE: protein‐DNA fusion system for screening of combinatorial protein libraries in vitro
Authors: Doi Nobuhide , Yanagawa Hiroshi
Published: July 2002

We have developed a new method that permits the complete in vitro construction and selection of peptide or protein libraries. This method relies on an in vitro transcription/translation reaction compartmentalized in water in oil emulsions. In each emulsion compartment, streptavidin (STA)‐fused polypeptides are synthesized and attached to the encoding DNA via its biotin label. The resulting protein‐DNA fusion molecules recovered from the emulsion can be subjected to affinity selection based on the properties of the peptide portion, whose sequence can be determined from that of its DNA‐tag. This method, named ‘STABLE’ (STA‐biotin linkage in emulsions), should be useful for rapid in vitro evolution of proteins and for ligand‐based selection of cDNA libraries.
Cloning of an α‐glucosidase gene from Thermococcus hydrothermalis by functional complementation of a Saccharomyces cerevisiae mal11 mutant strain
Authors: Galichet Arnaud , Belarbi Abdel
Published: July 2002

α‐Glucosidase is found in methanogenic and thermophilic archaea and also in eukaryotes and bacteria. The gene encoding the enzyme was cloned from Thermococcus hydrothermalis by complementation of a Saccharomyces cerevisiae deficiency maltase mutant strain. The gDNA clone isolated encodes an open reading frame corresponding to a protein of 242 amino acids. The protein shows 42% identity to a Pyrococcus horikoshii unknown ORF but no similarities were obtained with polysaccharidase sequences.
Sertaconazole in the treatment of mycoses: from dermatology to gynecology
Authors: Torres J , Márquez M , Camps F
Published: July 2002

Sertaconazole is a pharmaceutical product in the form of a cream, gel, powder and solution for dermatological use and vaginal cream, tablets and ovules for gynecological use. It is marketed in 24 countries and registered in a further 22. The active ingredient is 2% sertaconazole nitrate. Sertaconazole nitrate is an azole antifungal agent, with notable antifungal activity. Its molecule has a highly lipophilic fragment. This is a review of the efficacy and safety of all pharmaceutical forms of sertaconazole in order to provide data on vaginal sertaconazole for marketing purposes.
New considerations for the pathogenesis of endometriosis
Authors: Gazvani R , Templeton A
Published: July 2002

Objectives: To review the available evidence regarding the immunological, epidemiological and other factors involved in the pathogenesis of endometriosis. Methods: Literature review. Results: Endometriosis remains a poorly‐understood disease of unknown etiology and pathogenesis. Conclusions: There is evidence to suggest that alterations in the immune response, whether genetically transmitted or environmentally induced, predispose women to the ectopic implantation of endometrial cells transported into the peritoneal cavity by way of retrograde menstruation. This predisposition may exist because of an impaired peritoneal clearing of endometrial cells and fragments or because of pathological angiogenesis.
Fetal heart rate patterns associated with periventricular leukomalacia
Authors: Okamura M. , Itakura A. , Kurauchi O. , Hayakawa F. , Mizutani S. , Tomoda Y.
Published: July 2002

Objective: Our purpose was to assess the applicability of fetal heart rate (FHR) monitoring to detect fetuses at risk of developing periventricular leukomalacia (PVL). Methods: FHR tracings obtained for babies delivered under 33 weeks' gestation and with a birth weight under 2000 g were assessed for baseline heart rate, variability, deceleration and ‘flip flap’ (an oscillatory tracing pattern). Results: PVL developed in 19 of the 103 infants studied. All of these infants were among the fetuses who exhibited average and increased variability. In addition, PVL was detected in 10 (47.6%) of the 21 flip flap positive fetuses, and in 9 (11.0%) of the 82 flip flap negative fetuses. The incidence of PVL was significantly higher in the flip flap positive fetuses (P < 0.005). Conclusion: The possibility that an unstable intrauterine environment, reflected by a flip flap pattern, is related to the subsequent development of PVL is indicated.
Morphological alterations of hypothalamic nucleidue to intrahypothalamic hyperinsulinism in newborn rats
Authors: Plagemann A. , Harder T. , Rake A. , Janert U. , Melchior K. , Rohde W. , Dörner G.
Published: July 2002

In former studies, a temporary, intrahypothalamically localized hyperinsulinism duringbrain development was shown to result in overweight and metabolic disturbances during later lifein rats. Therefore, we tested the hypothesis whether intrahypothalamic insulin treatment duringearly postnatal life may lead to hypothalamic morphological alterations, i.e., of numerical densityof neurons and area of neuronal nuclei or area of neuronal cytoplasm, in this animal model. Forthis purpose, on the 8th day of age in Wistar rats a long‐acting insulin was bilaterally applicatedstereotactically into the hypothalamus (12 mIU on each side), while in controls the insulin‐freeagar‐vehicle was given only. By computer‐assisted morphometric analysis on the 15th day of life adecrease of the mean area of neuronal nuclei and the mean nucleus‐cytoplasm‐ratio within theVMN of the insulin‐treated animals was observed, as compared to control rats (P < 0.05), while no significant alterations were found in the lateral hypothalamic area (LHA). Analysisof topographically distinct parts of the VMN revealed significant reductions of the mean area ofneuronal nuclei (P < 0.001) and nucleus‐cytoplasm‐ratio (P < 0.05) inthe anterior part of the VMN (VMNpa). Furthermore, in the ventrolateral part (VMNpv) adecreased mean neuronal density was observed in the insulin group (P < 0.01). Incontrast, the dorsomedial part of the VMN (VMNpd) displayed an increased mean neuronaldensity in the insulin‐treated animals (P < 0.05). In the dorsomedial hypothalamicnucleus (DMN) a significant increase of the mean area of neuronal nuclei (P < 0.01)and the area of neuronal cytoplasm were observed (P < 0.001). These alterationswere accompanied by a significantly elevated mean numerical density of astrocytes (positive forglial fibriallary acidic protein ; GFAP+) within the periventricular hypothalamic area (PER) of theinsulin‐treated rats (P < 0.05). These observations speak for a varying vulnerabilityof LHA, DMN and distinct parts of the VMN to hyperinsulinism during early development,possibly leading to a disturbed organization and, consecutively, permanent dysfunction of thesemorphologically connected and functionally interacting hypothalamic nuclei.
Spatiotemporal pattern of the postnatal astrogliogenesis in the rat hippocampal formation
Authors: Setkowicz Zuzanna , Pawliński Rafał , Ziaja Marek , Janeczko Krzysztof
Published: July 2002

Newborn, 2‐, 4‐, 8‐, 16‐ and 30‐day‐old Wistar rats were injected with 3H‐thymidine and sacrificed following 4 h. survival time. Brain sections containing the dorsal part of hippocampal formation were immunostained for S‐100β protein and subjected to autoradiography to visualize proliferating astrocytes. Microscopical observations revealed age‐dependent changes in the number and distribution of proliferating astrocytes. The changes were considered as being related to the neurogenetic gradient characteristic to the hippocampal formation.
Depression and panic: Comorbidity
Authors: Baldwin DS
Published: July 2002

Panic disorder is a common condition. Epidemiological studies throughout the world consistently indicate that the lifetime prevalence of panic disorder (with or without agoraphobia) is between 1.5% and 3.5%. Panic disorder shows substantial comorbidity with other forms of mental illness. Major depressive disorder occurs in 50 to 65% of individuals with panic disorder and there is considerable cross-sectional and longitudinal comorbidity with recurrent brief depression and dysthymia. Phobic anxiety disorders, most notably social phobia and generalised anxiety disorder, commonly occur with panic disorder, especially in individuals with more severe agoraphobia. Approximately 35 to 50% of individuals with panic disorder in community settings also have agoraphobia. Panic disorder also shows significant comorbidity with physical illness. Compared with individuals without or with some other psychiatric diagnosis, patients with panic disorder have an increased risk of suffering from multiple medically unexplained symptoms and are associated with high use of medical services and increased mortality from both cardiovascular and cerebrovascular disease.
Subjective quality of life in schizophrenic patients living in the community. Relationship to clinical and social characteristics
Authors: Bengtsson-Tops A. , Hansson L.
Published: July 2002

The aims of this study were to assess the quality of life among 120 schizophrenic patients who were attending a psychiatric outpatient department and to investigate which socio-demographic and clinical factors influenced their subjective quality of life. Quality of life was assessed by the Lanchashire quality of life profile, social functioning was judged according to the Global Assessment of Functioning (GAF) scale, and psychopathology was rated by means of the Brief Psychiatric Rating Scale (BPRS). Both objective and subjective life conditions indicated an impaired quality of life for the patients. The areas of finance and work had the largest proportion of dissatisfied patients. Socio-demographic indicators showed to have a weak influence on the patient's self-assessed quality of life while clinical factors, such as psychopathology, strongly influenced the patient's life satisfaction. It is concluded that there is a need for further emphasise on the clinical, financial, and social interventions for this group of patients.