bluuebunny/crossref_metadata_2025 huggingface dataset:
Thrombospondin‐1‐induced vascular smooth muscle cell chemotaxis: The role of the type 3 repeat and carboxyl terminal domains
Authors: Lee Taeseung , Nesselroth Susan M. , Olson Eric T. , Esemuede Nowokere , Lawler Jack , Sumpio Bauer E. , Gahtan Vivian
Published: May 2003

Thrombospondin‐1 (TSP‐1), an acute phase reactant implicated in vascular disease, is a matricellular glycoprotein with six domains that confer different functions. The authors have shown TSP‐1 induces vascular smooth muscle cell (VSMC) chemotaxis via extracellular signal‐regulated kinases‐1 and ‐2 (ERK) and p38 kinase (p38) and that a fusion protein of the carboxyl terminal (COOH) and type 3 repeat (T3) domains independently induce VSMC chemotaxis. The purpose of this study was to determine whether COOH‐, T3‐induced VSMC chemotaxis, or both, is dependent upon ERK or p38 activation. To determine if the T3, COOH, or type 2 repeat domain (T2, control domain not associated with chemotaxis) activate ERK, p38, or both, VSMCs were exposed to each fusion protein (20 μg/ml for 15, 30, 60, or 120 min), serum‐free media (SFM, negative control), or TSP‐1 (20 μg/ml for 30 min, positive control). Western immunoblotting was performed for activation studies. Using a microchemotaxis chamber, VSMCs pre‐incubated in SFM, DMSO (vehicle control), PD98059 (10 μM), or SB202190 (10 μM) were exposed to each domain, TSP‐1, or SFM. After 4 h (37°C), migrated VSMCs were recorded as cells/five fields (400 ×) and analyzed by paired t‐test. ERK was activated by T2, T3, and COOH. However, p38 was activated by T3 and COOH, but not T2. T3 and COOH‐induced VSMC chemotaxis were inhibited by PD98059 or SB202190, but more completely by SB202190. The T2 domain had no effect on VSMC chemotaxis. These results suggest activation of the p38 pathway may be more specific than ERK for COOH‐ and T3‐induced VSMC chemotaxis. © 2003 Wiley‐Liss, Inc.
ATP modulates load‐inducible IL‐1β, COX 2, and MMP‐3 gene expression in human tendon cells
Authors: Tsuzaki M. , Bynum D. , Almekinders L. , Yang X. , Faber J. , Banes A.J.
Published: May 2003

Tendon cells receive mechanical signals from the load bearing matrices. The response to mechanical stimulation is crucial for tendon function. However, overloading tendon cells may deteriorate extracellular matrix integrity by activating intrinsic factors such as matrix metalloproteinases (MMPs) that trigger matrix destruction. We hypothesized that mechanical loading might induce interleukin‐1beta (IL‐1β) in tendon cells, which can induce MMPs, and that extracellular ATP might inhibit the load‐inducible gene expression. Human tendon cells isolated from flexor digitorum profundus tendons (FDPs) of four patients were made quiescent and treated with ATP (10 or 100 μM) for 5 min, then stretched equibiaxially (1 Hz, 3.5% elongation) for 2 h followed by an 18‐h‐rest period. Stretching induced IL‐1β, cyclooxygenase 2 (COX 2), and MMP‐3 genes but not MMP‐1. ATP reduced the load‐inducible gene expression but had no effect alone. A medium change caused tendon cells to secrete ATP into the medium, as did exogenous UTP. The data demonstrate that mechanical loading induces ATP release in tendon cells and stimulates expression of IL‐1β, COX 2, and MMP‐3. Load‐induced endogenous IL‐1β may trigger matrix remodeling or a more destructive pathway(s) involving IL‐1β, COX 2, and MMP‐3. Concomitant autocrine and paracrine release of ATP may serve as a negative feedback mechanism to limit activation of such an injurious pathway. Attenuation or failure of this negative feedback mechanism may result in the progression to tendinosis. © 2003 Wiley‐Liss, Inc.
Struktur der Randschicht nichtrostender Stähle nach Tieftemperatur‐Nitrierung – eine Mößbauerstudie
Authors: Pietzsch C. , Brusky U. , Spies H.‐J.
Published: May 2003

Durch die Nitrierung nichtrostender Stähle bei Temperaturen zwischen 300 °C und 400 °C können Randschichten hoher Härte mit Stickstoffgehalten von 8 wt.% bis 12 wt. % erzeugt werden.

In der vorliegenden Arbeit werden die Phasenbildungen nach Gas – und Plasmanitrierung beschrieben.

Zur Untersuchung der Phasenbildung und Phasenzusammensetzung wurden die Mößbauerspektroskopie in der Varianten CEMS und XCMS und die Röntgenbeugung eingesetzt. Die chemische Zusammensetzung wurde mittels GDOS, die Härte als Martenshärte unter Prüfkraft bestimmt.

Trotz unterschiedlicher Wirkprinzipien konnten mit den im Rahmen der Untersuchungen eingesetzten Prüfmethoden keine signifikanten Unterschiede zwischen der Struktur und den Eigenschaften von gas‐ und plasmanitrierten Proben gefunden werden.

Deutliche Unterschiede bestehen aber zwischen dem Nitrierverhalten unterschiedlicher Stähle.
Untersuchungen zum Boriereinfluss auf das Kavitationsverhalten von Fe‐Werkstoffen
Authors: Demirci A.H.
Published: May 2003

Ziel der im Folgenden beschriebenen Untersuchungen war es, am Beispiel eines Werkzeugstahles (X165CrMoV12) einen Überblick über den Einfluss einer Borierbehandlung auf das Kavitationsverhalten von Fe‐Werkstoffen zu erhalten.

Das Kavitationsverhalten wurde qualitativ im Rasterelektronenmikroskop und quantitativ durch Gewichtsvergleich beurteilt, wobei vorgesehen war, die Beanspruchungszeit unter sonst konstant gehaltenen, definierten Beanspruchungsbedingungen zu variieren.
Role of surgery in children with rhabdomyosarcoma
Authors: Schalow Erica L. , Broecker Bruce H.
Published: May 2003

Background

Rhabdomyosarcoma (RMS) is a common soft tissue sarcoma of childhood. Historically, surgery has played a central role in the management of children with this tumor, though with surgery alone survival rates were poor. With current multimodal (surgery, radiotherapy and chemotherapy) treatment of these patients, survival has dramatically improved and, with this improvement, there has been an evolution of the role of surgery in the management of this condition.

Material and Method

The contemporary published literature (English) regarding surgical aspects of pediatric rhabdomyosarcoma was reviewed and evaluated.

Results

Multimodal therapy has improved the survival of children with RMS from 25% in 1970 to greater than 70% today. Surgical procedures for childhood RMS today are less apt to be exenterative or multilating than those employed thirty years ago.

Conclusions

Surgery plays a vital role in the diagnosis and treatment of children with RMS. This role has evolved in the context of multimodal therapy and improved survival to an emphasis on less radical procedures with decreased morbidity. Med Pediatr Oncol 2003;41:1–6. © 2003 Wiley‐Liss, Inc.
European training programme in paediatric haematology and oncology
Authors: Mann Jillian R. ,   on behalf of SIOP Europe and ESPHI
Published: May 2003

The European Board of Paediatrics has recommended for all paediatricians a three‐year Common Trunk training in basic paediatrics, followed for tertiary care specialists, by three years of training in the relevant specialty. Paediatric Haematology and Oncology is a complex specialty with a broad range of activities, ranging from the care of children with leukaemia and solid tumours including those of the central nervous system, to all types of non‐malignant haematologic disorders and laboratory haematology pursuits. A modular training programme has been recommended to provide a broad standard of training across the whole specialty which should last at least 18 months, and a further period of 18 months that may be used flexibly to prepare trainees for their anticipated future careers. Med Pediatr Oncol 2003;40:65–66. © 2003 Wiley‐Liss, Inc.
Prolylhydroxylase and procollagen type III in long‐term survivors of acute lymphoblastic leukemia (ALL): A biochemical approach to HCV‐related liver disease
Authors: Leonardi Salvatore , La Spina Milena , La Rosa Mario , Schilirò Gino
Published: May 2003

Background

We examined two proteins, prolylhydroxylase (hPH) and procollagen type III (PIIIP), as possible non‐invasive HCV‐related markers of liver disease. The purpose of this study was to assess whether the measurement of these proteins could serve to monitor HCV related liver damage in acute lymphoblastic leukaemia (ALL) patients.

Procedure

A total of 34 ALL patients, 24 HCV‐seropositive and 10 HCV‐seronegative, who had had increased transaminase values (ALT) for almost 6 months were studied. Serum hPH concentrations were determined by an immuno‐enzymatic assay kit. PIIIP was assayed by the radioimmunoassay method.

Results

Both hPH and PIIIP were increased in ALL patients with chronic hepatitis C. Serum hPH levels were significantly elevated in those with chronic hepatitis C with either normal or high transaminases when compared to those who never were HCV seropositive. The sensitivity and specificity of these protein measurements to evaluate hepatic fibrosis were not supported by histologic confirmation because only 6 out of 12 patients with chronic hepatitis had a liver biopsy.

Conclusions

Our study suggests that PIIIP and hPH values are significantly higher in ALL patients with chronic HCV with either normal or high transaminases. This might suggest that the liver damage is more marked in patients with chronic hepatitis and that the liver damage is related to the HCV rather than chemotherapy. Future studies correlating histologic findings with the serum biochemical markers are required to establish the sensitivity and specificity of hPH and PIIIP in predicting hepatic fibrosis and to confirm this association. Med Pediatr Oncol 2003;41:17–20. © 2003 Wiley‐Liss, Inc.
Iterationsverfahren zur exakten Restmode‐Korrektur bei Frequenzganganalysen
Authors: Dietrich G.K.P.
Published: May 2003

In this paper we formulate frequency response methods which determine iteratively the exact steady state response of linear mechanical vibration systems, i.e., no forces are neglected by these algorithms. We presuppose that only a small number of eigenfrequencies and ‐modes of the systems is known. The result of methods of approximation may be iteratively improved toward the exact result.
inhibits antigen‐mediated Syk, but not Lyn tyrosine kinase activation in mast cells
Authors: Vallé Alain , Kinet Jean-Pierre
Published: May 2003

High affinity IgE receptors (αβγ
2) mediate the activation of the non‐receptor tyrosine kinases Lyn and Syk. Here we show that the antioxidant drug (NAC) inhibits antigen‐mediated Syk activation whereas Lyn activation and phosphorylation of β and γ is maintained. Furthermore, NAC inhibits antigen‐mediated calcium mobilization and exocytosis in a dose‐dependent manner, but does not inhibit ionomycin‐induced exocytosis. These data support a model in which the activation of Lyn is responsible for receptor phosphorylation and precedes the activation of Syk. The inhibition of Syk activation by NAC may be relevant to B and T cell antigen receptors, which are also linked to Syk/ZAP70 tyrosine kinases.
Fluorescence spectra of bacteriorhodopsin and the intermediates O and Q at room temperature
Authors: Ohtani Hiroyuki , Tsukamoto Yasuhisa , Sakoda Yuusaku , Hamaguchi Hiro-o
Published: May 2003

An unequivocal answer is given to the question of why the reported fluorescence spectra of bacteriorhodopsin (bR568) have been different from one another. The inconsistency is shown to arise from the accumulation of the fluorescent intermediates O and Q (K
N) by cw excitation light. Their fractions in the photo‐stationary states depend on the excitation power and the suspension pH. We report the intermediate‐free fluorescence spectrum of bR568 obtained with a weak excitation source (632.8 nm, 5.3 × 1015 − 1.9 × 1016 photons cm−2 · s−1) and a near‐IR sensitive intensified photodiode array system. The fluorescence maxima of the spectra, F(λ) and , are located at 755 ± 10 nm and 12700 ± 200 cm−1, respectively. The spectrum of O is identical to that of the deionized purple membrane bR605(F
max = 750 ± 5 nm, f
max = 13,000 ± 100 cm−1). Q (K
N) exhibits a blue‐shifted spectrum more than that of bR568, (F
max < 720 nm, f
max < 13,400 cm−1).
Effect of the C‐terminal proline repeats on ordered packing of squid rhodopsin and its mobility in membranes
Authors: Venien-Bryan Catherine , Davies Anthony , Langmack Keith , Baverstock Jenny , Watts Anthony , Marsh Derek , Saibil Helen
Published: May 2003

Negative stain electron microscopy and saturation transfer electron spin resonance spectroscopy have been used to compare the lattice ordering and in‐plane membrane mobility of full‐length and C‐terminally cleaved squid rhodopsin. The C‐terminus of squid rhodopsin contains a negatively charged region followed by 9–10 repeats of a proline‐rich sequence, not found in rhodopsins other than those of cephalopod invertebrates, but similar proline repeats are found in other, unrelated membrane proteins. We find that the proline repeats cluster the rhodopsins into small groups, interfering with two‐dimensional crystallization and maintaining their mobility in the membrane.
Self‐aggregation of purified and membrane‐bound erythrocyte CD38 induces extensive decrease of its ADP‐ribosyl cyclase activity
Authors: Zocchi Elena , Franco Luisa , Guida Lucrezia , Calder Lesley , De Flora Antonio
Published: May 2003

The transmembrane glycoprotein CD38 is a bifunctional enzyme that catalyzes at its ectocellular domain both the synthesis and the hydrolysis of cyclic ADP‐ribose (cADPR). The complete reaction, converting NAD+ to nicotinamide and ADP‐ribose, reproduces an NAD+glycohydrolase (NADase) reaction. CD38 purified from human erythrocyte membranes has been recently shown to undergo stable oligomerization induced by either NAD+ or β‐mercaptoethanol. We demonstrate that oligomerization is also triggered by reduced glutathione (GSH) and that the GSH‐induced self‐aggregation of purified CD38 is accompanied by extensive and comparable decrease of its ADP‐ribosyl cyclase and NADase activities. GSH‐induced oligomerization of CD38 and strong enzyme inactivation take place also in situ on erythrocyte membranes.
Ferrodoxin‐dependent CO2 fixation in bean sprouts
Authors: Hirasawa Masakazu , Koguchi-Kamioka Osamu , Li Xin , Knaff David B.
Published: May 2003

Extracts of bean sprouts are capable of reducing ferrodoxin and of catalyzing the incorporation of bicarbonate and acetyl coenzyme A into an organic compound that is likely to be pyruvate, in a reaction that requires reduced ferredoxin. The rate of the reaction, the first known for which ferredoxin appears to serve as the direct reductant for CO2 fixation in a higher plant, depends on the concentrations of both ferredoxin and bicarbonate, with half‐maximal rates being observed at ferredoxin and bicarbonate concentrations of 0.8 μM and 200 μM, respectively.
Isolation and identification of a μ‐calpain‐protein kinase Cα complex in skeletal muscle
Authors: Savart Michel , Verret Catherine , Dutaud Dominique , Touyarot Katia , Elamrani Najat , Ducastaing André
Published: May 2003

A μ‐calpain‐PKC complex was isolated from rabbit skeletal muscle by ultracentrifugation and by anion‐exchange chromatography. The PKC associated to μ‐calpain was stimulated by calcium, phosphatidylserine and diacylglycerol, and corresponds to a conventional PKC (cPKC). This complex presents an apparent molecular mass close to 190 kDa and is composed of one μ‐calpain molecule and of one cPKC molecule. Using monoclonal antibodies specific for the different cPKC isoforms, the isoenzyme associated to μ‐calpain was identified as cPKCα. Immunofluorescence staining reveals a co‐localization of μ‐calpain and cPKCα on the muscle fibre plasma membranes.
Stress signal, mediated by a Hogl‐like MAP kinase, controls sexual development in fission yeast
Authors: Kato Tomohisa , Okazaki Koei , Murakami Hiroshi , Stettler Sophie , Fantes Peter A. , Okayama Hiroto
Published: May 2003

We identified the phhl
+ gene that encodes a MAP kinase as the effector of Wis1 MAP kinase kinase in fission yeast, which is highly homologous with HOG1 of S. cerevisiae. Heterothalic phh1 dsiruptant is phenotypically indistinguishable from wis1 deletion mutant, both displaying the same extent of partial sterility and enhanced sensitivity to a variety of stress. In phh1 disruptant, nitrogen starvation‐induced expression of ste11
+, a key controller of sexual differentiation, is markedly diminished. Ectopic expression of ste11
+ effectively restores fertility, but not stress resistance, to the phh1 disruptant. These data show that stress signal, mediated by a MAP kinase, is required for efficient start of sexual differentiation.
The yeast multidrug transporter Pdr5 of the plasma membrane is ubiquitinated prior to endocytosis and degradation in the vacuole
Authors: Egner Ralf , Kuchler Karl
Published: May 2003

We have recently demonstrated that the Pdr5 ATP binding cassette multidrug transporter is a short‐lived protein, whose biogenesis involves cell surface targeting followed by endocytosis and delivery to the vacuole for proteolytic turnover [Egner, R., Mahé, Y., Pandjaitan, R., and Kuchler, K. (1995) Mol. Cell. Biol. 15, 5879–5887]. Using c‐myc epitope‐tagged ubiquitin, we now have shown that Pdr5 is a ubiquitinated plasma membrane protein in vivo. Ubiquitination of Pdr5 was detected in both wild type and conditional end4 mutants defective in endocytic vesicle formation. Likewise, the Ste6 a‐factor pheromone transporter, which represents another short‐lived ABC transporter whose turnover requires vacuolar proteolysis, was also found to be ubiquitinated, and ubiquitin‐modified Ste6 massively accumulated in end4 mutants at the restrictive temperature. By contrast, the plasma membrane ATPase Pma1, a long‐lived and metabolically very stable protein, was found not to be ubiquitinated. Our results imply a novel function for ubiquitin in protein trafficking and suggest that ubiquitination of certain short‐lived plasma membrane proteins may trigger their endocytic delivery to the vacuole for proteolytic turnover.
PRK1 phosphorylates MARCKS at the PKC sites: serine 152, serine 156 and serine 163
Authors: Palmer Ruth H. , Schönwaßer Dorothee C. , Rahman Dinah , Pappin Darryl J.C. , Herget Thomas , Parker Peter J.
Published: May 2003

The 80kDa yristolated lanine‐ich ‐inase ubstrate (MARCKS) in a major in vivo substrate of protein kinase C (PKC). Here we report that MARCKS is a major substrate for the lipid‐activated PKC‐related kinase (PRK1) in cell extracts. Furthermore, PRK1 is shown to phosphorylate MARCKS on the same sites as PKC in vitro. Thus, control of MARCKS phosphorylation on these previously identified ‘PKC’ sites may be regulated under certain circumstances by PRK as well as PKC mediated signalling pathways. The implications for MARCKS as a marker of PKC activation and as a point of signal convergence are discussed.
Identification of differentially expressed genes induced in pancreatic islet neogenesis
Authors: Rafaeloff Ronit , Qin Xiao F. , Barlow Scott W. , Rosenberg Lawrence , Vinik Aaron I.
Published: May 2003

Cellophane wrapping of the hamster pancreas induces islet neogenesis. We have used the mRNA differential display technique to select for genes expressed during islet neogenesis but not in control pancreata. Ten candidate clones have been identified. Upon sequencing, 6 clones showed a high degree of homology to known genes, 1 showed some, and 3 showed no homology to genes of known sequence. Thus, mRNA differential display is a useful technique to identify genes induced during islet neogenesis, and in combination with screening hamster pancreatic cDNA libraries for full length clones, will enhance the likelihood of capturing the participants in this process.
The high‐affinity binding of Clostridium botulinum type B neurotoxin to synaptotagmin II associated with gangliosides GT1b/GD1a
Authors: Nishiki Tei-ichi , Tokuyama Yoshimi , Kamata Yoichi , Nemoto Yasuo , Yoshida Akira , Sato Kazuki , Sekiguchi Mariko , Takahashi Masami , Kozaki Shunji
Published: May 2003

125I‐labeled botulinum type B neurotoxin was shown to bind specifically to recombinant rat synaptotagmins I and II. Binding required reconstitution of the recombinant proteins with gangliosides GT1b/GD1a. Scatchard plot analyses revealed a single class of binding site with dissociation constants of 0.23 and 2.3 nM for synaptotagmin II and synaptotagmin I, respectively, values very similar to those of the high‐ (0.4 nM) and low‐affinity (4.1 nM) binding sites in synaptosomes. The high‐affinity binding of neurotoxin to synaptosomes was specifically inhibited by a monoclonal antibody recognizing with the amino‐terminal region of synaptotagmin II. These results suggest that this region of synaptotagmin II participates in the formation of the high‐affinity toxin binding site by associating with specific gangliosides.
Evidence for an association of ndh B, ndh J gene products and ferredoxin‐NADP‐reductase as components of a chloroplastic NAD(P)H dehydrogenase complex
Authors: Guedeney Geneviève , Corneille Sylvie , Cuiné Stéphan , Peltier Gilles
Published: May 2003

Using non‐denaturing gel electrophoresis and staining with nitro‐blue tetrazolium, we reveal the presence of two NAD(P)H oxidoreductase activity bands within thylakoids membranes of Solanum tuberosum L. Second dimension SDS‐PAGE and Western analysis show that one of the activity bands contains several polypeptides, two of them being recognized by antibodies directed against peptides corresponding to conserved domains of chloroplastic genes products NDH B and NDH J (at 32 and 18 kDa, respectively). Both activity bands also contain a polypeptide (around 36 kDa) recognized by an antibody directed against ferredoxin‐NADP+‐reductase (FNR). We conclude from these results that both chloroplastic ndh B and ndh J gene products are components of a thylakoid NAD(P)H dehydrogenase complex. The association with FNR is suggested to allow the complex to use NADPH instead of NADH as a preferential substrate.
The human G protein β4 subunit: gene structure, expression, Gγ and effector interaction
Authors: Rosskopf Dieter , Nikula Christiane , Manthey Iris , Joisten Markus , Frey Ulrich , Kohnen Stefanie , Siffert Winfried
Published: May 2003

The aim of this study was the characterization of the human Gβ4 subunit of heterotrimeric G proteins. Human Gβ4 is widely expressed. Its gene is located on chromosome 3 with a genomic structure indistinguishable from that of the genes of Gβ1 to Gβ3, but entirely different from Gβ5. In vitro translation co‐precipitation analyses revealed that Gβ4 can form stable dimers with Gγ1, Gγ2, Gγ3, Gγ4, Gγ5, Gγ7, Gγ10, Gγ11, Gγ12, and Gγ13, dimers which were also able to stimulate phospholipase β2.
The C‐terminal peptide of thrombospondin‐1 stimulates distinct signaling pathways but induces an activation‐independent agglutination of platelets and other cells
Authors: Voit Simone , Udelhoven Michael , Lill Gereon , Aktas Barsom , Nieswandt Bernhard , Schrör Karsten , Weber Artur-Aron
Published: May 2003

A peptide from the C‐terminal domain of thrombospondin‐1 (4N1‐1) has been proposed to stimulate platelet aggregation by a novel mechanism involving both an activation‐independent agglutination and an activation‐dependent, glycoprotein (GP) IIb/IIIa‐mediated aggregation which involves GPVI signaling but does not involve CD47. The present study demonstrates that 4N1‐1 stimulated a different pattern of signal transduction pathways than the GPVI agonist convulxin. Furthermore, 4N1‐1‐induced platelet aggregation was activation‐independent and not dependent on GPVI or GPIIb/IIIa. Interestingly, 4N1‐1 also stimulated activation‐independent agglutination of different megakaryocytic and non‐megakaryocytic cells. 4N1‐1‐induced cell agglutination but not platelet signaling was inhibited by anti‐CD47 antibodies.
Snorkeling of lysine side chains in transmembrane helices: how easy can it get?
Authors: Strandberg Erik , Killian J.Antoinette
Published: May 2003

Transmembrane segments of proteins are often flanked by lysine residues. The side chains of these residues may snorkel, i.e. they may bury themselves with their aliphatic part in the hydrophobic region of the lipid bilayer, while positioning the charged amino group in the more polar interface. Here we estimate the free energy cost of snorkeling from thermodynamical calculations based on studies with synthetic transmembrane peptides [Strandberg et al. (2002) Biochemistry 41, 7190–7198]. The value is estimated to be between 0.07 and 0.7 kcal mol−1 for a lysine side chain. This very low value indicates that snorkeling may be a common process, which should be taken into consideration both in experimental and in theoretical studies on protein–lipid interactions.
Growth phase‐dependent changes in the subcellular localization of pre‐B‐cell colony‐enhancing factor1
Authors: Kitani Takako , Okuno Sachiko , Fujisawa Hitoshi
Published: May 2003

A cDNA encoding the homolog of the human pre‐B‐cell colony‐enhancing factor (PBEF), a cytokine‐like secreted protein, was isolated from a rat cDNA library. This protein existed in both the cytoplasm and nucleus of the cells, and the amount was higher in the cytoplasm than in the nucleus of proliferating PC‐12 and Swiss 3T3 cells but higher in the nucleus than in the cytoplasm of the PC‐12 cells treated with nerve growth factor and the 3T3 cells grown to a confluent state. Thus, the so‐called PBEF is not a cytokine‐like secreted protein but an intracellular protein associated with the cell cycle.
Transient increase of phosphatidylcholine in plant cells in response to phosphate deprivation
Authors: Jouhet Juliette , Maréchal Eric , Bligny Richard , Joyard Jacques , Block Maryse A
Published: May 2003

In plants, phosphate deprivation is normally known to decrease the phospholipid content consistent with a mobilization of the phosphate reserve, and conversely to increase non‐phosphorous membrane lipids such as digalactosyldiacylglycerol. We report here that unexpectedly, at an early stage of phosphate starvation, phosphatidylcholine (PC) increases transiently. We also show that a significant pool of diacylglycerol (DAG) with the same fatty acid composition as that of PC is present and moreover increases in response to phosphate deprivation. The evolution of the molecular profile of the newly synthesized galactolipids is compatible with a utilization of DAG accumulating from PC hydrolysis, achieved after selection of their acyl molecular species by the galactolipid synthesizing enzymes.
Protein kinase D: a family affair
Authors: Rykx An , De Kimpe Line , Mikhalap Svetlana , Vantus Tibor , Seufferlein Thomas , Vandenheede Jackie R. , Van Lint Johan
Published: May 2003

The protein kinase D family of enzymes consists of three isoforms: PKD1/PKCμ, PKD2 and PKD3/PKCν. They all share a similar architecture with regulatory sub‐domains that play specific roles in the activation, translocation and function of the enzymes. The PKD enzymes have recently been implicated in very diverse cellular functions, including Golgi organization and plasma membrane directed transport, metastasis, immune responses, apoptosis and cell proliferation.
The metabolic role of human ADH3 functioning as ethanol dehydrogenase
Authors: Lee Shou-Lun , Wang Ming-Fang , Lee An-I , Yin Shih-Jiun
Published: May 2003

Human class III alcohol dehydrogenase (ADH3), also known as glutathione‐dependent formaldehyde dehydrogenase, exhibited non‐hyperbolic kinetics with ethanol at a near physiological pH 7.5. The S
0.5 and k
cat were determined to be 3.4±0.3 M and 33±3 min−1, and the Hill coefficient (h) 2.21±0.09, indicating positive cooperativity. Strikingly, the S
0.5 for ethanol was found to be 5.4×106‐fold higher than the K
m for S‐(hydroxymethyl)glutathione, a classic substrate for the enzyme, whereas the k
cat for the former was 41% lower than that for the latter. Isotope effects on enzyme activity suggest that hydride transfer may be rate‐limiting in the oxidation of ethanol. Kinetic simulations using the experimentally determined Hill constant suggest that gastric ADH3 may highly effectively contribute to the first‐pass metabolism at 0.5–3 M ethanol, an attainable range in the gastric lumen during alcohol consumption. The positive cooperativity mainly accounts for this metabolic role of ADH3.
Defined subcomplexes of the A1 ATPase from the archaeon Methanosarcina mazei Gö1: biochemical properties and redox regulation
Authors: Lemker Thorsten , Grüber Gerhard , Schmid Roland , Müller Volker
Published: May 2003

The potential A1 ATPase genes ahaA, ahaB, ahaC, ahaD, ahaE, ahaF, and ahaG from the anaerobic archaeon Methanosarcina mazei Gö1 were overexpressed in Escherichia coli DK8 (pTL2). An A1 complex was purified to apparent homogeneity and shown by Western blot and N‐terminal sequence analyses to contain subunits A, B, C, D, and F but to be devoid of subunits E and G. Further removal of subunit C was without effect on ATPase activity. The enzyme was most active at pH 5.2 and required bisulfite and acetate for maximal activity. Kinetic studies confirmed three new inhibitors for A1 ATPases (diethylstilbestrol and its derivatives hexestrol and dienestrol) and identified redox modulation as a new type of regulation of archaeal A1 ATPases.
Substitutions in the Escherichia coli RNA polymerase σ70 factor that affect recognition of extended −10 elements at promoters
Authors: Sanderson Andrew , Mitchell Jennie E , Minchin Stephen D , Busby Stephen J.W
Published: May 2003

Previous work has shown that the base sequence of the DNA segment immediately upstream of the −10 hexamer at bacterial promoters (the extended −10 element) can make a significant contribution to promoter strength. Guided by recently published structural information, we used alanine scanning and suppression mutagenesis of Region 2.4 and Region 3.0 of the Escherichia coli RNA polymerase σ70 subunit to identify amino acid sidechains that play a role in recognition of this element. Our study shows that changes in these regions of the σ70 subunit can affect the recognition of different extended −10 element sequences.
Palmitate modulates the early steps of insulin signalling pathway in pancreatic islets
Authors: Haber Esther P , Hirabara Sandro M , Gomes André D , Curi Rui , Carpinelli Angelo R , Carvalho Carla R.O
Published: May 2003

Insulin stimulates its own secretion and synthesis by pancreatic β‐cells. Although the exact molecular mechanism involved is unknown, changes in β‐cell insulin signalling have been recognized as a potential link between insulin resistance and its impaired release, as observed in non‐insulin‐dependent diabetes. However, insulin resistance is also associated with elevated plasma levels of free fatty acids (FFA) that are well known modulators of insulin secretion by pancreatic islets. This information led us to investigate the effect of FFA on insulin receptor signalling in pancreatic islets. Exposure of pancreatic islets to palmitate caused up‐regulation of several insulin‐induced activities including tyrosine phosphorylation of insulin receptor and pp185. This is the first evidence that short exposure of these cells to 100 μM palmitate activates the early steps of insulin receptor signalling. 2‐Bromopalmitate, a carnitine palmitoyl‐CoA transferase‐1 inhibitor, did not affect the effect of the fatty acid. Cerulenin, an acylation inhibitor, abolished the palmitate effect on protein levels and phosphorylation of insulin receptor. This result supports the proposition that protein acylation may be an important mechanism by which palmitate exerts its modulating effect on the intracellular insulin signalling pathway in rat pancreatic islets.
OmpF enhances the ability of BtuB to protect susceptible Escherichia coli cells from colicin E9 cytotoxicity
Authors: Law Christopher J , Penfold Christopher N , Walker Daniel C , Moore Geoffrey R , James Richard , Kleanthous Colin
Published: May 2003

The outer membrane (OM) vitamin B12 receptor, BtuB, is the primary receptor for E group colicin adsorption to Escherichia coli. Cell death by this family of toxins requires the OM porin OmpF but its role remains elusive. We show that OmpF enhances the ability of purified BtuB to protect bacteria against the endonuclease colicin E9, demonstrating either that the two OM proteins form the functional receptor or that OmpF is recruited for subsequent translocation of the bacteriocin. While stable binary colicin E9–BtuB complexes could be readily shown in vitro, OmpF‐containing complexes could not be detected, implying that OmpF association with the BtuB–colicin complex, while necessary, must be weak and/or transient in nature.
Activity of coliphage HK022 excisionase (Xis) in the absence of DNA binding
Authors: Gottfried Pnina , Silberstein Nava , Yagil Ezra , Kolot Mikhail
Published: May 2003

A mutated excisionase (Xis) protein of coliphage HK022 whose single Cys residue was replaced by Ser does not bind to its two tandem binding sites (X1, X2) on the P arm of attR. Despite its DNA‐binding inability the protein showed 30% excision activity of the wild type Xis both in vitro and in vivo. This partial activity is attributed to the interaction of Xis with integrase that is retained in the mutant protein. This protein–protein interaction occurs in the absence of DNA binding.
Motilin–bicelle interactions: membrane position and translational diffusion
Authors: Andersson August , Mäler Lena
Published: May 2003

The interaction between the peptide hormone motilin and bicelles has been investigated by pulsed field gradient‐nuclear magnetic resonance methods and by the use of paramagnetic probes. Diffusion coefficients were measured for motilin, the phospholipids with and without motilin, and for tetramethylsilane. The results show that around 90% of motilin is bound to acidic bicelles and 84% of motilin is bound to neutral bicelles. It is found that the apparent bicelle size is reduced by the presence of motilin. This cannot be explained by changes in 1,2‐dihexanoyl‐sn‐glycero‐3‐phosphatidylcholine solubility. The use of paramagnetic agents to investigate the position of motilin shows that the turn in the N‐terminus of motilin is inserted into the bicelle, while the helix most likely resides within the head‐group layer.
Spectral imaging and its applications in live cell microscopy
Authors: Zimmermann Timo , Rietdorf Jens , Pepperkok Rainer
Published: May 2003

In biological microscopy, the ever expanding range of applications requires quantitative approaches that analyze several distinct fluorescent molecules at the same time in the same sample. However, the spectral properties of the fluorescent proteins and dyes presently available set an upper limit to the number of molecules that can be detected simultaneously with common microscopy methods. Spectral imaging and linear unmixing extends the possibilities to discriminate distinct fluorophores with highly overlapping emission spectra and thus the possibilities of multicolor imaging. This method also offers advantages for fast multicolor time‐lapse microscopy and fluorescence resonance energy transfer measurements in living samples. Here we discuss recent progress on the technical implementation of the method, its limitations and applications to the imaging of biological samples.
Modulating viral gene expression by aptamers to RNA structures
Authors: Toulmé Jean‐Jacques , Darfeuille Fabien , Kolb Gaëlle , Chabas Sandrine , Staedel Cathy
Published: May 2003

Oligonucleotides exhibiting a strong affinity and a high specificity for RNA hairpins were obtained by in vitro selection. Such oligomers give rise to loop—loop complexes with the target hairpins: the trans‐activation responsive (TAR) element of the Human Immunodeficiency virus‐1 (HIV‐1) or subdomains of the Hepatitis C virus (HCV) mRNA. Chemically modified derivatives of an antiTAR aptamer were shown to compete out the binding of the viral protein Tat and to selectively inhibit the in vitro TAR‐dependent transcription of a reporter gene. In addition, antisense oligomers derived from sequences selected against the domain IIId of the HCV internal ribosome entry site were shown to specifically block translation both in a cell‐free assay and in cultured cells.
rhBMP‐2 injected in a calcium phosphate paste (α‐BSM) accelerates healing in the rabbit ulnar osteotomy model
Authors: Li R. H. , Bouxsein M. L. , Blake C. A. , D'Augusta D. , Kim H. , Li X. J. , Wozney J. M. , Seeherman H. J.
Published: May 2003

This study evaluated the ability of recombinant human bone morphogenetic protein‐2 (rhBMP‐2) delivered in an injectable calcium phosphate carrier (α‐BSM) to accelerate healing in a rabbit ulna osteotomy model compared to untreated surgical controls. Healing was assessed by radiography, histology and biomechanics. Bilateral mid‐ulnar osteotomies were created in 16 skeletally mature rabbits. One limb in each animal was injected with either 0.1 mg rhBMP‐2/α‐BSM (BMP) (N = 8) or buffer/α‐BSM (BSM) (N = 8). Contralateral osteotomies served as untreated surgical controls (SXCT). Gamma scintigraphy showed 75%, 45% and 5% of the initial 125I‐rhBMP‐2 dose was retained at the osteotomy site at 3 h, 1 week and 3 weeks. The biological activity of rhBMP‐2 (alkaline phosphatase activity from bioassay) extracted from α‐BSM incubated in vitro up to 30 days at 37°C was unchanged. Radiographs demonstrated complete bridging of the BMP limbs at 4 weeks whereas none of the BSM or SXCT limbs were bridged. Post‐mortem peripheral quantitative computed tomography determined mineralized callus area was 62% greater in BMP limbs compared to SXCT limbs. Torsional stiffness and strength were 63% and 103% greater in BMP limbs compared to SXCT limbs. There was no difference in torsional properties between BSM and SXCT limbs. Failure occurred outside the osteotomy in four out of seven of the BMP limbs. All BSM and SXCT limbs failed through the osteotomy. Histology showed bony bridging of the osteotomy and no residual carrier in the BMP limbs. BSM and SXCT groups showed less mature calluses composed of primarily fibrocartilaginous tissue and immature bone in the osteotomy gap. These data indicate rhBMP‐2 delivered in α‐BSM accelerated healing in a rabbit ulna osteotomy model compared to BSM and SXCT groups. © 2003 Orthopaedic Research Society. Published by Elsevier Science Ltd. All rights reserved.
Institutions and services of psychiatric care in Saxony-Anhalt: assessment with the European Services Mapping Schedule
Authors: Brieger Peter , Wetzig Fanny , Böcker Felix M..
Published: May 2003

This article describes and analyses the availability of outpatient, inpatient and community-based psychiatric care in Saxony-Anhalt, one of the federal states in the eastern part of Germany. The European Services Mapping Schedule was used to classify 365 institutions. Outpatient care was provided by an average of four private practice psychiatrists per 100 000 inhabitants, which is low when compared to the German average. Ten secure beds (fo-rensic), 48 acute beds, 13 elective beds and 13 day hospital places per 100 000 inhabitants were available for inpatient care. Non-acute non-hospital residential services with indefinite stay and with 24 h support amounted to 240 places per 100 000, with regional differences ranging from less than 100 to more than 1000. Other facilities offering paid work or work-related activities were scarce and some services providing structured activity or social contact were available only in urban agglomerations. Overall, psychiatric care in Saxony-Anhalt is fragmented as regards providers and funding.
Fallen heroes: explaining the failure of the Gdansk shipyard, and the successful early reform strategies in Szczecin and Gdynia
Authors: Keat Preston
Published: May 2003

In this paper, I examine the causes of failure in the Gdansk shipyard, and alternatively, the determinants of success in Szczecin and Gdynia. I give particular attention to the role of the initial owner of these companies—the Polish government. Why were these three firms treated differently by the government, and how did this affect their approaches to restructuringŒ I find that in the cases of Szczecin and Gdynia, initial government aid conditionality, combined with pro-active employee support of reforms, produced environments where restructuring had a real chance. In Gdansk, the government and company employees failed to reach agreement on a strategy for initiating reforms. Both feared the potential consequences (economic for the workers and political for the government) of embarking on reforms that were in no way guaranteed to succeed. What emerged was an ongoing stalemate, with company management rejecting a series of reform projects and the government continuing to offer soft credits to keep the unprofitable firm alive. This stalemate, or sub-optimal equilibrium, was linked to the shipyard’s uniquely credible threats of intransigence.
Immunocytochemical study of the glucocorticoid receptor in rat liver nuclei after hyperthermic stress
Authors: Čvoro Aleksandra , Korać Aleksandra , Matić Gordana
Published: May 2003

The presence of glucocorticoid receptors (GR) in rat liver nuclei over a 24 h time period following hyperthermic stress at 41 °C was immunocytologically studied using unfixed nuclear smears. Liver nuclei in unstressed animals were found to be immunonegative for GR. However, intense GR immunopositivity followed by a subsequent gradual decrease in receptor levels was observed in the nuclei of test animals during the first 2 h after stress. This stress‐related increase in the receptor nuclear level was greater than the increase seen after dexamethasone administration. These results suggest that hyperthermic stress could potentiate the hormonal stimulation of receptor nuclear translocation.
Suppression of anoikis by v‐Src but not by activated c‐H‐Ras in human gallbladder epithelial cells
Authors: Hisano Chizuko , Tanaka Risa , Fujishima Hiromitsu , Ariyama Hiroshi , Tsuchiya Tanji , Tatsumoto Takashi , Mitsugi Kenji , Nakamura Minoru , Nakano Shuji
Published: May 2003

Detachment of anchorage‐dependent normal epithelial cells from their substratum causes the type of apoptosis known as anoikis, whereas malignant cells can proliferate independently of anchorage. Because src and ras oncogenes are activated in many human cancers, we investigated their role and downstream signaling pathways in anoikis resistance, using HAG‐1 human epithelial cells transfected with v‐src or activated H‐ras. Consequently, anchorage‐dependent mock‐ or ras‐transfected cells underwent anoikis. In contrast, anchorage‐independent v‐Src‐transformed cells did not exhibit such apoptotic features. Focal adhesion kinase (FAK), a transducer of integrin, was only activated in v‐Src‐transformed cells. Herbimycin A, an Src kinase inhibitor, reduced tyrosyl phosphorylation of FAK and reversed resistance to anoikis. However, both protein kinase C (PKC) and phophatidylinositol‐3 (PI‐3) kinase inhibitors failed to induce anoikis. These data suggest that the ability of activated Src to prevent anoikis may be mediated by Src to a downstream signaling pathway involving FAK, but not Ras, PI‐3 kinase, or PKC.
Short chain fatty acids stimulate feline colonic smooth muscle contraction
Authors: Rondeau Mark P , Meltzer Karen , Michel Kathryn E , McManus Catherine M , Washabau Robert J
Published: May 2003

The effect of short chain fatty acids (SCFA) on feline colonic smooth muscle contraction was evaluated in vitro. Colonic tissue was obtained from seven healthy male and female adult cats and seven healthy male and female kittens. Longitudinal and circular colonic smooth muscle strips from proximal and distal colon were incubated with SCFA (acetate, butyrate and propionate; 1–100 mM). SCFA-induced contractions were compared to responses obtained using maximal concentrations (10−4 M) of acetylcholine (ACh). The calcium dependence of the SCFA response was investigated by incubating with nifedipine (1 μM) or verapamil (1 μM).

Acetate, butyrate and propionate elicited isometric stress responses (0.25–1.98×104 N/m2) in longitudinal, but not circular, smooth muscle from both the proximal and distal colon of adult cats. Maximal responses were attained at 50 and 100 mM SCFA. Maximal butyrate and propionate responses were 29 and 19% of the maximal ACh response (10−4 M), respectively. Acetate was least effective in stimulating contractile responses. Nifedipine and verapamil abolished all responses. Contractile responses in kittens were similar to those observed in adult cats, but were smaller in amplitude.

Results of these studies have shown that SCFA stimulate longitudinal colonic smooth muscle contractions in kittens and adult cats in vitro. These SCFA-induced contractions involve activation of calcium influx. These in vitro findings may account for some of the effects of dietary fiber on feline colonic motility in vivo.
Kidney transplants for children under 1 year of age – a single‐center experience
Authors: Khwaja Khalid , Humar Abhinav , Najarian John S.
Published: May 2003

Abstract: From September 20, 1970 to October 24, 2001, we performed 46 kidney transplants in infants under 1 yr old at the University of Minnesota. This article reviews the preoperative care, surgical technique, and immunosuppression. Recipients included 16 females and 30 males; the youngest recipient was 6 wk old. The mean pretransplant height was 62.8 cm, which increased to 77 cm at 1 yr post‐transplant and to 104 cm at 5 yr. We used 40 living donors (all but 1 were related to the recipient) and 6 cadaver donors. The overall actuarial graft survival was 85% at 1 yr and 70% at 5 yr. In the cyclosporine era, graft survival improved to 91% at 1 yr and 80% at 5 yr. Death with function was the most common cause of graft loss (n = 5), followed by biopsy‐proven chronic rejection (n = 4), biopsy‐proven recurrent disease (n = 3), and graft thrombosis (n = 2). Patient survival was 91% at 1 yr and 86% at 5 yr. In the cyclosporine era, patient survival was 100% at 5 yr and 85% at 10 yr. We concluded that an early transplant is the best treatment option for infants under 1 yr old with chronic renal failure. Whenever possible, adult living kidney donors should be used.
Pilot study describing the use of pravastatin in pediatric renal transplant recipients
Authors: Butani Lavjay , Pai Monica V. , Makker Sudesh P.
Published: May 2003

Abstract: Renal transplant (Tx) recipients frequently develop hypercholesterolemia. Pravastatin (P) has been shown to be effective in adult renal Tx recipients, not only in reducing serum cholesterol, but possibly also in decreasing graft rejection. However, there are no data on the use of P in children following renal transplantation. We conducted a retrospective case–control study evaluating the safety and efficacy of P (10–20 mg/day) in reducing hypercholesterolemia, when used pre‐emptively in the post‐Tx period in seven children, compared with an historical control (C) group of nine children who had not received P. The two groups were comparable with respect to their demographics and in their pretransplant serum cholesterol. Compared with the C group, the mean serum cholesterol in the P group was lower at 3 months (159 mg/dL vs. 225 mg/dL), 6 months (134 mg/dL vs. 200 mg/dL), 9 months (134 mg/dL vs. 209 mg/dL), and 12 months (125 mg/dL vs. 195 mg/dL) (p < 0.005 for all, Student's two‐tailed t‐test). At 1 month only 43% of the P group had hypercholesterolemia compared with 67% of the controls; by 12 months this difference was even more significant (0% in the P group vs. 45% in the C group). None of the treated patients developed any adverse reactions. This study demonstrates that the pre‐emptive use of P in pediatric renal Tx recipients appears to be effective in significantly reducing serum cholesterol. Whether this effect will translate into improved allograft and patient survival in the long term cannot be predicted at present and will require additional studies to evaluate.