bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of Novel Genes in Intestinal Tissue That Are Regulated after Infection with an Intestinal Nematode Parasite
Authors: Datta R. , deSchoolmeester M. L. , Hedeler C. , Paton N. W. , Brass A. M. , Else K. J.
Published: June 2005

Infection of resistant or susceptible mice withTrichuris murisprovokes mesenteric lymph node responses which are polarized towards Th2 or Th1, respectively. These responses are well documented in the literature. In contrast, little is known about the local responses occurring within the infected intestine. Through microarray analyses, we demonstrate that the gene expression profile of infected gut tissue differs according to whether the parasite is expelled or not. Genes differentially regulated postinfection in resistant BALB/c mice include several antimicrobial genes, in particular, intelectin (Itln). In contrast, analyses in AKR mice which ultimately progress to chronic infection provide evidence for a Th1-dominated mucosa with up-regulated expression of genes regulated by gamma interferon. Increases in the expression of genes associated with tryptophan metabolism were also apparent with the coinduction of tryptophanyl tRNA synthetase (Wars) and indoleamine-2,3-dioxygenase (Indo). With the emerging literature on the role of these gene products in the suppression of T-cell responses in vitro and in vivo, their up-regulated expression here may suggest a role for tryptophan metabolism in the parasite survival strategy.
Protection againstLeptospira interrogansSensu Lato Challenge by DNA Immunization with the Gene Encoding Hemolysin-Associated Protein 1
Authors: Branger C. , Chatrenet B. , Gauvrit A. , Aviat F. , Aubert A. , Bach J. M. , André-Fontaine G.
Published: June 2005

The use of DNA constructs encoding leptospiral proteins is a promising new approach for vaccination against leptospirosis. In previous work we determined that immunization with hemolysis-associated protein 1 (Hap1) (LipL32) expressed by adenovirus induced significant protection against a virulentLeptospirachallenge in gerbils. To avoid the use of the adenovirus vector, we checked for clinical protection against lethal challenge by DNA vaccination. A DNA vaccine expressing Hap1 was designed to enhance the direct gene transfer of this protein into gerbils. A challenge was performed 3 weeks after the last immunization with a virulent strain of serovar canicola. Our results show that the cross-protective effect with pathogenic strains ofLeptospira, shared by Hap1, could be mediated by the DNA plasmid vector. This finding should facilitate the design and development of a new generation of vaccines against bacteria, particularlyLeptospira interroganssensu lato.
Multiple Insertional Events, Restricted by the Genetic Background, Have Led to Acquisition of Pathogenicity Island II
J96
-Like Domains among
Escherichia coli
Strains of Different Clinical Origins
Authors: Bidet Philippe , Bonacorsi Stéphane , Clermont Olivier , De Montille Caroline , Brahimi Naima , Bingen Edouard
Published: June 2005

We investigated the dissemination of pathogenicity island (PAI) II
J96
-like elements (
hra
,
hly
,
cnf1
, and
pap
) among 455
Escherichia coli
isolates from children and adults with urinary tract infection (UTI), neonates with meningitis or colonized healthy neonates, and 74 reference strains by means of PCR phylogenetic grouping, ribotyping, and PCR analysis of virulence genes. Colocalization of these genes was documented by pulsed-field gel electrophoresis followed by Southern hybridization and long-range PCR (LRPCR) between the
hra
and the
papG
alleles. Site-specific insertion of the PAI was determined by LRPCR between
hra
and tRNA flanking sequences.
hra
,
hly
, and
cnf1
were found in 113 isolates and consistently colocalized, constituting the backbone of PAI II
J96
-like domains. The prevalence of PAI II
J96
-like domains was significantly higher among UTI isolates than among neonatal meningitis and commensal isolates. These domains were restricted to a few ribotypes of group B2. In contrast to the consistent colocalization of
hra
,
hly
, and
cnf1
, the
pap
operon was varied: 12% of strains exhibited an allelic exchange of the
papG
class III allele (
papGIII
) for the
papG
class II allele (
papGII
) (only UTI isolates), and the
pap
operon was deleted in 23% of strains. No strains harbored
papGIII
outside the PAI, which appears to be the only source of this allele. PAI II
J96
-like domains were inserted in the vicinities of three different tRNAs—
pheU
(54%),
leuX
(29%), and
pheV
(15%)—depending on the genetic backgrounds and origins of the isolates. Multiple insertional events restricted by the genetic background have thus led to PAI II
J96
acquisition. Specific genetic backgrounds and insertion sites may have played a role in additional recombination processes for
E. coli
adaptation to different ecological niches.
Effects of Pregnancy and Intensity of
Plasmodium falciparum
Transmission on Immunoglobulin G Subclass Responses to Variant Surface Antigens
Authors: Megnekou Rosette , Staalsoe Trine , Taylor Diane W , Leke Rose , Hviid Lars
Published: June 2005

Placenta-sequestering
Plasmodium falciparum
involved in the pathogenesis of pregnancy-associated malaria (PAM) in otherwise clinically immune women expresses particular variant surface antigens (VSA
PAM
) on the surface of infected erythrocytes that differ from VSA found in parasitized nonpregnant individuals (non-PAM type VSA). We studied levels of immunoglobulin G (IgG) and IgG subclasses with specificity for VSA
PAM
and for non-PAM type VSA in pregnant and nonpregnant women from two sites with different endemicities in Cameroon. We found that VSA
PAM
-specific responses depended on the pregnancy status, parity, gestational age, and parasite transmission intensity, whereas only the parasite transmission intensity influenced the levels of IgG specific for non-PAM type VSA. For both types of VSA, the responses were dominated by the cytophilic subclass IgG1, followed by IgG3. In pregnant women, the levels of VSA
PAM
-specific antibodies either were very low or negative or were very high, whereas the levels of the antibodies specific for non-PAM type VSA were uniformly high. Interestingly, the levels of VSA
PAM
-specific IgG1 increased with increasing gestational age, while the levels of the corresponding IgG3 tended to decrease with increasing gestational age. The IgG subclass responses with specificity for non-PAM type VSA did not vary significantly with gestational age. Taken together, our data indicate that IgG1 and to a lesser extent IgG3 are the main subclasses involved in acquired VSA
PAM
-specific immunity to pregnancy-associated malaria.
Microarray Analysis of Quorum-Sensing-Regulated Genes inPorphyromonas gingivalis
Authors: Yuan Lihui , Hillman Jeffrey D. , Progulske-Fox Ann
Published: June 2005

Quorum sensing is a phenomenon defined as gene regulation in response to cell density that regulates various functions in bacteria. The periodontopathogenPorphyromonas gingivalispossesses aluxSgene homologue that may encode a quorum-sensing system. In order to identify genes ofP. gingivalisthat are regulated byluxS, gene expression analysis was done using microarrays and RNA samples from the W83 wild-type strain and an isogenicluxSmutant, LY2001. The results indicated that 17 open reading frames (ORFs) in LY2001 are upregulated and two are downregulated. Real-time PCR was done to confirm the microarray results. Among the upregulated ORFs is a group of stress-related genes, includinghtrA,clpB,groEL,dnaK, and the F subunit of alkyl hydroperoxide reductase. This suggested thatluxSis involved in stress gene regulation inP. gingivalis. Stress response experiments, including high-temperature survival, resistance to hydrogen peroxide (H2O2), and survival during exposure to low and high pH, were performed on theP. gingivaliswild-type and LY2001 strains. LY2001 had a significantly higher survival rate than did W83 when stressed at 50°C. No difference was found at pH 5, but LY2001 had increased survival compared to W83 at pH 9. LY2001 also survived better than W83 when stressed with 0.35 mM H2O2. These results suggest thatluxSmight be involved in promoting survival ofP. gingivalisin the host by regulating its response to host-induced stresses such as temperature, H2O2, and pH.
Superoxide Production in
Galleria mellonella
Hemocytes: Identification of Proteins Homologous to the NADPH Oxidase Complex of Human Neutrophils
Authors: Bergin David , Reeves Emer P. , Renwick Julie , Wientjes Frans B. , Kavanagh Kevin
Published: June 2005

The insect immune response has a number of structural and functional similarities to the innate immune response of mammals. The objective of the work presented here was to establish the mechanism by which insect hemocytes produce superoxide and to ascertain whether the proteins involved in superoxide production are similar to those involved in the NADPH oxidase-induced superoxide production in human neutrophils. Hemocytes of the greater wax moth (
Galleria mellonella
) were shown to be capable of phagocytosing bacterial and fungal cells. The kinetics of phagocytosis and microbial killing were similar in the insect hemocytes and human neutrophils. Superoxide production and microbial killing by both cell types were inhibited in the presence of the NADPH oxidase inhibitor diphenyleneiodonium chloride. Immunoblotting of
G. mellonella
hemocytes with antibodies raised against human neutrophil
phox
proteins revealed the presence of proteins homologous to gp91
phox
, p67
phox
, p47
phox
, and the GTP-binding protein rac 2. A protein equivalent to p40
phox
was not detected in insect hemocytes. Immunofluorescence analysis localized insect 47-kDa and 67-kDa proteins throughout the cytosol and in the perinuclear region. Hemocyte 67-kDa and 47-kDa proteins were immunoprecipitated and analyzed by matrix-assisted laser desorption ionization—time of flight analysis. The results revealed that the hemocyte 67-kDa and 47-kDa proteins contained peptides matching those of p67
phox
and p47
phox
of human neutrophils. The results presented here indicate that insect hemocytes phagocytose and kill bacterial and fungal cells by a mechanism similar to the mechanism used by human neutrophils via the production of superoxide. We identified proteins homologous to a number of proteins essential for superoxide production in human neutrophils and demonstrated that significant regions of the 67-kDa and 47-kDa insect proteins are identical to regions of the p67
phox
and p47
phox
proteins of neutrophils.
Neisseria gonorrhoeaeKills Carcinoembryonic Antigen-Related Cellular Adhesion Molecule 1 (CD66a)-Expressing Human B Cells and Inhibits Antibody Production
Authors: Pantelic Milica , Kim Young-June , Bolland Silvia , Chen Ines , Shively John , Chen Tie
Published: June 2005

Neisseria gonorrhoeaecells (gonococci [GC]), the etiological agents for gonorrhea, can cause repeated infections. During and after gonococcal infection, local and systemic antigonococcal antibody levels are low. These clinical data indicate the possibility that GC may suppress immune responses during infection. Carcinoembryonic antigen-related cellular adhesion molecule 1 (CEACAM1 or CD66a), a receptor for GC opacity (Opa) proteins, was shown to mediate inhibitory signals. In the present study, human B cells were activated by interleukin-2 to express CEACAM1 and then stimulated to secrete antibodies and simultaneously coincubated with Opa−and OpaI GC of strain MS11. Our results show that this OpaI GC has the ability to inhibit antibody production. The interaction of GC and CEACAM1 with human peripheral B cells also results in induction of cell death. The same findings were observed in DT40 B cells. This CEACAM1-promoted cell death pathway does not involve the inhibitory signals or the tyrosine phosphatases SHP-1 and SHP-2 but depends on Bruton's tyrosine kinase in DT40 cells. Our results suggest thatNeisseria gonorrhoeaepossesses the ability to suppress antibody production by killing CEACAM1-expressing B cells.
The Gonococcal Fur-Regulated
tbpA
and
tbpB
Genes Are Expressed during Natural Mucosal Gonococcal Infection
Authors: Agarwal Sarika , King Carol A. , Klein Ellen K. , Soper David E. , Rice Peter A. , Wetzler Lee M. , Genco Caroline A.
Published: June 2005

Iron is limiting in the human host, and bacterial pathogens respond to this environment by regulating gene expression through the ferric uptake regulator protein (Fur). In vitro studies have demonstrated that
Neisseria gonorrhoeae
controls the expression of several critical genes through an iron- and Fur-mediated mechanism. While most in vitro experiments are designed to determine the response of
N. gonorrhoeae
to an exogenous iron concentration of zero, these organisms are unlikely to be exposed to such severe limitations of iron in vivo. To determine if
N. gonorrhoeae
expresses iron- and Fur-regulated genes in vivo during uncomplicated gonococcal infection, we examined gene expression profiles of specimens obtained from male subjects with urethral infections. RNA was isolated from urethral swab specimens and used as a template to amplify, by reverse transcriptase PCR (RT-PCR), gonococcal genes known to be regulated by iron and Fur (
tbpA
,
tbpB
, and
fur
). The constitutively expressed gonococcal
rmp
gene was used as a positive control. RT-PCR analysis indicated that gonorrhea-positive specimens where
rmp
expression was seen were also 93% (51/55)
fbpA
positive, 87% (48/55)
tbpA
positive, and 86% (14 of 16 tested)
tbpB
positive. In addition, we detected a
fur
transcript in 79% (37 of 47 tested) of positive specimens. We also measured increases in levels of immunoglobulin G antibody against TbpA (91%) and TbpB (73%) antigens in sera from infected male subjects compared to those in uninfected controls. A positive trend between
tbpA
gene expression and TbpA antibody levels in sera indicated a relationship between levels of gene expression and immune response in male subjects infected with gonorrhea for the first time. These results indicate that gonococcal iron- and Fur-regulated
tbpA
and
tbpB
genes are expressed in gonococcal infection and that male subjects with mucosal gonococcal infections exhibit antibodies to these proteins.
Inhibition of Chondroitin-4-Sulfate-Specific Adhesion of
Plasmodium falciparum
-Infected Erythrocytes by Sulfated Polysaccharides
Authors: Andrews Katherine T. , Klatt Nicole , Adams Yvonne , Mischnick Petra , Schwartz-Albiez Reinhard
Published: June 2005

Adhesion of
Plasmodium falciparum
-infected erythrocytes to placental chondroitin 4-sulfate (CSA) has been linked to the severe disease outcome of pregnancy-associated malaria. Soluble polysaccharides that release mature-stage parasitized erythrocytes into the peripheral circulation may help elucidate these interactions and have the potential to aid in developing therapeutic strategies. We have screened a panel of 11 sulfated polysaccharides for their capacities to inhibit adhesion of infected erythrocytes to CSA expressed on CHO-K1 cells and ex vivo human placental tissue. Two carrageenans and a cellulose sulfate (CS10) were able to inhibit adhesion to CSA and to cause already bound infected erythrocytes to de-adhere in a dose-dependent manner. CS10, like CSA and in contrast to all other compounds tested, remained bound to infected erythrocytes after washing and continued to inhibit binding. Both carrageenans and CS10 inhibited adhesion to placental tissue. Although highly sulfated dextran sulfate can inhibit CSA-mediated adhesion to CHO cells, this polysaccharide amplified adhesion to placental tissue severalfold, demonstrating the importance of evaluating inhibitory compounds in systems as close to in vivo as possible. Interestingly, and in contrast to all other compounds tested, which had a random distribution of sulfate groups, CS10 exhibited a clustered sulfate pattern along the polymer chain, similar to that of the undersulfated placental CSA preferred by placental-tissue-binding infected erythrocytes. Therefore, the specific antiadhesive capacity observed here seems to depend not only on the degree of charge and sulfation but also on a particular pattern of sulfation.
Effects of a Probiotic Strain ofEnterococcus faeciumon the Rate of Natural Chlamydia Infection in Swine
Authors: Pollmann M. , Nordhoff M. , Pospischil A. , Tedin K. , Wieler L. H.
Published: June 2005

Chlamydiae are obligately intracellular pathogens which cause infections associated with a broad range of diseases in both livestock and humans. In addition, a large proportion of animals may become persistently infected asymptomatic carriers and serve as reservoirs for other animals which also shed these potential zoonotic pathogens. Reducing the chlamydial load of animals is therefore of major importance, and since large-scale antibiotic treatment is neither desired nor feasible, alternative means of prevention are needed. Here we performed a study comparing the efficacy of a probiotic strain ofEnterococcus faeciumon the reduction of both the rate of natural infection and the shedding of chlamydiae in swine. The presence ofChlamydiaceaewas detected by species-specific PCR of fecal samples of sows taken at three times prior to the birth of piglets. Piglets delivered from chlamydia-positive sows in either the control or the probiotic group were also examined for the frequency of chlamydiae at various ages. Eighty-five percent of the piglets from the control group were found to be chlamydia positive, whereas chlamydiae were found in only 60% of piglets from the probiotic group, results confirmed by fluorescence in situ hybridization and immunohistology, which showed higher rates of infection in the control group. In addition to the reduced frequency of chlamydia-positive piglets in the probiotic group, the time of appearance of positive samples was delayed. To our knowledge, these data show for the first time that a probiotic strain ofE. faeciumcan reduce the rate of carryover infections of piglets by obligate intracellular pathogens.
SlyA Regulates Function of
Salmonella
Pathogenicity Island 2 (SPI-2) and Expression of SPI-2-Associated Genes
Authors: Linehan Sheena A. , Rytkönen Anne , Yu Xiu-Jun , Liu Mei , Holden David W.
Published: June 2005

During the systemic phase of murine infection with
Salmonella enterica
serovar Typhimurium, bacterial virulence is correlated with the ability to grow and survive within host macrophages.
Salmonella
pathogenicity island 2 (SPI-2), encoding a type three secretion system, has emerged as an important contributor to
Salmonella
intracellular growth. SPI-2 mutants have been proposed to be more accessible than wild-type
Salmonella
to oxyradicals generated by the NADPH phagocyte oxidase. We performed mixed infections of mice to investigate the relationship between SPI-2 and SlyA, a transcriptional regulator that confers resistance to oxyradicals. In mixed-infection experiments, the SPI-2 null mutant was severely attenuated in virulence, whereas
slyA
mutants were only mildly attenuated. Surprisingly, further experiments indicated that the function of SPI-2 was partially dependent on
slyA
. The intracellular behavior of a
slyA
mutant in infected cells was consistent with inefficient SPI-2 expression, as formation of
Salmonella
-induced filaments and the intracellular F-actin meshwork, features that depend on SPI-2, were present at abnormally low frequencies. Furthermore, the translocated levels of the SPI-2 effector SseJ were severely reduced in a strain carrying a mutation in
slyA
. We used flow cytometry to investigate the role of SlyA in expression of green fluorescent protein (GFP) from transcriptional fusions with promoters of either of two other SPI-2 effector genes,
sifB
and
sifA
. The
slyA
mutant exhibited reduced GFP expression from both promoters. Combining mutations in
slyA
and other regulators of SPI-2 indicated that SlyA acts through the SsrAB two-component regulatory system. SlyA exhibits partial functional redundancy with OmpR-EnvZ and contributes to the transcriptional response to low osmolarity and the absence of calcium, two environmental stimuli that promote SPI-2 gene expression.
LidA, a Translocated Substrate of the
Legionella pneumophila
Type IV Secretion System, Interferes with the Early Secretory Pathway
Authors: Derré Isabelle , Isberg Ralph R.
Published: June 2005

Legionella pneumophila
uses a type IV secretion system to deliver effector molecules into the host cell and establish its replication vacuole. In this study, we investigated the role of LidA, a translocated substrate associated with the surface of the
L. pneumophila
-containing vacuole. LidA is secreted into the host cell throughout the replication cycle of the bacteria and associates with compartments of the early secretory pathway. When overexpressed in mammalian cells or yeast, LidA interferes with the early secretory pathway, probably via a domain predicted to be rich in coiled-coil structure. Finally, during intracellular replication, the replication vacuoles are in close contact with the endoplasmic reticulum-Golgi intermediate compartment and the Golgi apparatus, suggesting a positive correlation between intracellular growth and association of the vacuole with compartments of the early secretory pathway. We propose that LidA is involved in the recruitment of early secretory vesicles to the
L. pneumophila
-containing vacuole and that the vacuole associates with the secretory pathway to facilitate this process.
Role of the Distal
sarA
Promoters in SarA Expression in
Staphylococcus aureus
Authors: Cheung Ambrose L. , Manna Adhar C.
Published: June 2005

The global regulatory locus
sarA
comprises a 375-bp open reading frame that is driven by three promoters, the proximal P1 and distal P3 and P2 promoters. We mutated the weaker P3 and P2 promoters to ascertain the effect of the change on SarA protein and target gene expression. Our results indicated that the solely active P1 promoter led to a lower SarA protein level, which has an effect on
agr
transcription and subsequently had corresponding effects on
hla
,
sspA
, and
spa
transcription, probably in both
agr
-independent and
agr
-dependent manners.
Role of Motility and Flagellin Glycosylation in the Pathogenesis of
Pseudomonas aeruginosa
Burn Wound Infections
Authors: Arora Shiwani K. , Neely Alice N. , Blair Barbara , Lory Stephen , Ramphal Reuben
Published: June 2005

In this study, we tested the contribution of flagellar motility, flagellin structure, and its glycosylation in
Pseudomonas aeruginosa
using genetically defined flagellar mutants. All mutants and their parent strains were tested in a burned-mouse model of infection. Motility and glycosylation of the flagellum appear to be important determinants of flagellar-mediated virulence in this model. This is the first report where genetically defined flagellar variants of
P. aeruginosa
were tested in the burned-mouse model of infection.
Structural Diversity of Streptokinase and Activation of Human Plasminogen
Authors: Lizano Sergio , Johnston Kenneth H.
Published: June 2005

The β domain of streptokinase is required for plasminogen activation and contains a region of sequence diversity associated with infection and disease in group A streptococci. We report that mutagenesis of this polymorphic region does not alter plasminogen activation, which suggests an alternative function for this molecular motif in streptococcal disease.
Certain Site-Directed, Nonenzymatically Active Mutants of the
Haemophilus influenzae
P4 Lipoprotein Are Able To Elicit Bactericidal Antibodies
Authors: Green Bruce A. , Baranyi Elizabeth , Reilly Thomas J. , Smith Arnold L. , Zlotnick Gary W.
Published: June 2005

The
Haemophilus influenzae
P4 lipoprotein (
hel
) is a potential component of a nontypeable
H. influenzae
otitis media vaccine. Since P4 is known to be an enzyme, nonenzymatically active forms of recombinant P4 are required. After site-directed mutagenesis of the
hel
gene, three of the mutated proteins were shown to be vaccine candidates.
SET MAPPING REFLECTION
Authors: MOORE JUSTIN TATCH
Published: June 2005

In this note we will discuss a new reflection principle which follows from the Proper Forcing Axiom. The immediate purpose will be to prove that the bounded form of the Proper Forcing Axiom implies both that 2ω = ω2 and that [Formula: see text] satisfies the Axiom of Choice. It will also be demonstrated that this reflection principle implies that □(κ) fails for all regular κ > ω1.
HOMOLOGOUS NON-ISOTOPIC SYMPLECTIC TORI IN A K3 SURFACE
Authors: ETGÜ TOLGA , PARK B. DOUG
Published: June 2005

For each member of an infinite family of homology classes in the K3 surface E(2), we construct infinitely many non-isotopic symplectic tori representing this homology class. This family has an infinite subset of primitive classes. We also explain how these tori can be non-isotopically embedded as homologous symplectic submanifolds in many other symplectic 4-manifolds including the elliptic surfaces E(n) for n > 2.
ON UNIQUENESS AND DIFFERENTIABILITY IN THE SPACE OF YAMABE METRICS
Authors: ANDERSON MICHAEL T.
Published: June 2005

It is shown that there is a unique Yamabe representative for a generic set of conformal classes in the space of metrics on any manifold. At such classes, the scalar curvature functional is shown to be differentiable on the space of Yamabe metrics. In addition, some sufficient conditions are given which imply that a Yamabe metric of locally maximal scalar curvature is necessarily Einstein.
CONNECTIONS FOR GENERAL GROUP ACTIONS
Authors: LEWIS DEBRA , NIGAM NILIMA , OLVER PETER J.
Published: June 2005

We introduce the notion of a partial connection, which extends to manifolds with nonfree group actions most of the key features of connections on principal bundles. We investigate several properties of curvature in this setting and show that (partial) moving frames determine (partial) connection forms.
DIFFERENTIAL EQUATIONS AND INTERTWINING OPERATORS
Authors: HUANG YI-ZHI
Published: June 2005

We show that if every module W for a vertex operator algebra V = ∐n∈ℤV(n) satisfies the condition dim W/C1(W)<∞, where C1(W) is the subspace of W spanned by elements of the form u-1w for u ∈ V+ = ∐n>0 V(n) and w ∈ W, then matrix elements of products and iterates of intertwining operators satisfy certain systems of differential equations. Moreover, for prescribed singular points, there exist such systems of differential equations such that the prescribed singular points are regular. The finiteness of the fusion rules is an immediate consequence of a result used to establish the existence of such systems. Using these systems of differential equations and some additional reductivity conditions, we prove that products of intertwining operators for V satisfy the convergence and extension property needed in the tensor product theory for V-modules. Consequently, when a vertex operator algebra V satisfies all the conditions mentioned above, we obtain a natural structure of vertex tensor category (consequently braided tensor category) on the category of V-modules and a natural structure of intertwining operator algebra on the direct sum of all (inequivalent) irreducible V-modules.
Reciprocal changes in endogenous ghrelin and growth hormone during fasting in healthy women
Authors: Koutkia Polyxeni , Schurgin Sunita , Berry Jacqueline , Breu Jeff , Lee B. S. Hang , Klibanski Anne , Grinspoon Steven
Published: June 2005

Ghrelin stimulates growth hormone (GH) secretion, but it is unknown whether there is a feedback of GH on ghrelin secretion. In this study, we characterized the relatedness of GH and ghrelin in a model of acute caloric deprivation in 10 healthy women (age 26.7 ± 1.6 yr) during a 4-day fast in the early follicular phase. GH, ghrelin, and cortisol were assessed every hour over 24 h during an isocaloric diet and after a 4-day complete fast. Sampling during a normal diet at baseline demonstrated that ghrelin decreased 17.9% within 1 h after meals ( P < 0.0001), but there was no meal effect on GH. BMI (22.3 ± 0.4 vs. 21.5 ± 0.4 kg/m2, P < 0.0001) and IGF-I (312 ± 28 vs.124 ± 22 ng/ml, P < 0.0001) decreased during fasting. Mean 24-h GH increased (2.6 ± 0.5 vs. 5.6 ± 0.5 ng/ml, P < 0.001), but ghrelin decreased (441.3 ± 59.7 vs. 359.8 ± 54.2 pg/ml, P = 0.012). The peak ghrelin level decreased from 483.5 to 375.6 pg/ml ( P < 0.0001), and the time of the peak ghrelin changed from 0415 to 1715. In contrast, the diurnal pattern of GH was maintained, with increases in the nadir (1.1 to 3.4 ng/ml) and peak GH concentrations (4.1 to 7.9 ng/ml) from the fed to fasted state ( P < 0.0001). The change in morning GH concentrations was inversely related to the change in ghrelin ( r = −0.79, P = 0.012). During complete short-term caloric deprivation in healthy women, ghrelin decreases, even as GH rises, and these processes appear to be reciprocal, suggesting that GH exhibits feedback inhibition on ghrelin. Our data provide new evidence of the physiological relationship of GH and ghrelin in response to changes in protein-energy metabolism.
Measurement of selective effect of insulin on glucose disposal from labeled glucose oral test minimal model
Authors: Man Chiara Dalla , Caumo Andrea , Basu Rita , Rizza Robert , Toffolo Gianna , Cobelli Claudio
Published: June 2005

The oral glucose minimal model (OMM) measures insulin sensitivity (SI) and the glucose rate of appearance (Ra) of ingested glucose in the presence of physiological changes of insulin and glucose concentrations. However, SI of OMM measures the overall effect of insulin on glucose utilization and glucose production. In this study we show that, by adding a tracer to the oral dose, e.g., of a meal, and by using the labeled version of OMM, OMM* to interpret the data, one can measure the selective effect of insulin on glucose disposal, [Formula: see text]. Eighty-eight individuals underwent both a triple-tracer meal with the tracer-to-tracee clamp technique, providing a model-independent reference of the Ra of ingested glucose ([Formula: see text]) and an insulin-modified labeled intravenous glucose tolerance test (IVGTT*). We show that OMM* provides not only a reliable means of tracing the Ra of ingested glucose (Ra meal) but also accurately measures [Formula: see text]. We do so by comparing OMM* Ra meal with the model-independent [Formula: see text] provided by the tracer-to-tracee clamp technique, while OMM* [Formula: see text] is compared with both [Formula: see text], obtained by using as known input [Formula: see text], and with [Formula: see text] measured during IVGTT*.
Stimulation of Na+transport by AVP is independent of PKA phosphorylation of the Na-K-ATPase in collecting duct principal cells
Authors: Mordasini David , Bustamante Mauro , Rousselot Martine , Martin Pierre-Yves , Hasler Udo , Féraille Eric
Published: June 2005

Arginine-vasopressin (AVP) stimulates Na+transport and Na-K-ATPase activity via cAMP-dependent PKA activation in the renal cortical collecting duct (CCD). We investigated the role of the Na-K-ATPase in the AVP-induced stimulation of transepithelial Na+transport using the mpkCCDc14cell model of mammalian collecting duct principal cells. AVP (10−9M) stimulated both the amiloride-sensitive transepithelial Na+transport measured in intact cells and the maximal Na pump current measured by the ouabain-sensitive short-circuit current in apically permeabilized cells. These effects were associated with increased Na-K-ATPase cell surface expression, measured by Western blotting after streptavidin precipitation of biotinylated cell surface proteins. The effects of AVP on Na pump current and Na-K-ATPase cell surface expression were dependent on PKA activity but independent of increased apical Na+entry. Time course experiments revealed that in response to AVP, the cell surface expression of both endogenous Na-K-ATPase and hybrid Na pumps containing a c- myc-tagged wild-type human α1-subunit increased transiently. Na-K-ATPase cell surface expression was maximal after 30 min and then declined toward baseline after 60 min. Immunoprecipitation experiments showed that PKA activation did not alter total phosphorylation levels of the endogenous Na-K-ATPase α-subunit. In addition, mutation of the PKA phosphorylation site (S943A or S943D) did not alter the time course of increased cell surface expression of c- myc-tagged Na-K-ATPase in response to AVP or to dibutyryl-cAMP. Therefore, stimulation of Na-K-ATPase cell surface expression by AVP is dependent on PKA but does not rely on α1-subunit phosphorylation on serine 943 in the collecting duct principal cells.
Mechanoregulation of intracellular Ca2+concentration is attenuated in collecting duct of monocilium-impairedorpkmice
Authors: Liu Wen , Murcia Noel S. , Duan Yi , Weinbaum Sheldon , Yoder Bradley K. , Schwiebert Erik , Satlin Lisa M.
Published: June 2005

Autosomal recessive polycystic kidney disease (ARPKD) is characterized by the progressive dilatation of collecting ducts, the nephron segments responsible for the final renal regulation of sodium, potassium, acid-base, and water balance. Murine models of ARPKD possess mutations in genes encoding cilia-associated proteins, including Tg737 in orpk mice. New findings implicate defects in structure/function of primary cilia as central to the development of polycystic kidney disease. Our group (Liu W, Xu S, Woda C, Kim P, Weinbaum S, and Satlin LM, Am J Physiol Renal Physiol 285: F998–F1012, 2003) recently reported that increases in luminal flow rate in rabbit collecting ducts increase intracellular Ca2+concentration ([Ca2+]i) in cells therein. We thus hypothesized that fluid shear acting on the apical membrane or hydrodynamic bending moments acting on the cilium increase renal epithelial [Ca2+]i. To further explore this, we tested whether flow-induced [Ca2+]itransients in collecting ducts from mutant orpk mice, which possess structurally abnormal cilia, differ from those in controls. Isolated segments from 1- and 2-wk-old mice were microperfused in vitro and loaded with fura 2; [Ca2+]iwas measured by digital ratio fluorometry before and after the rate of luminal flow was increased. All collecting ducts responded to an increase in flow with an increase in [Ca2+]i, a response that appeared to be dependent on luminal Ca2+entry. However, the magnitude of the increase in [Ca2+]iin 2- but not 1-wk-old mutant orpk animals was blunted. We speculate that this defect in mechano-induced Ca2+signaling in orpk mice leads to aberrant structure and function of the collecting duct in ARPKD.
Exercise-induced increase in muscle insulin sensitivity
Authors: Holloszy John O.
Published: June 2005

Exercise/muscle contraction activates glucose transport. The increase in muscle glucose transport induced by exercise is independent of insulin. As the acute effect of exercise on glucose transport wears off, it is replaced by an increase in insulin sensitivity. An increase in insulin sensitivity results in a shift in the insulin dose-response curve to the left, with a decrease in the concentration of insulin needed to induce 50% of the maximal response. This phenomenon, which plays a major role in rapid muscle glycogen accumulation after exercise, is not mediated by amplification of the insulin signal. Development of the increase in insulin sensitivity after contractions does not require protein synthesis or activation of p38 MAPK. It does require the presence of a serum protein during the period of contractile activity. The effect of exercise on muscle insulin sensitivity is mimicked by hypoxia and by treatment of muscles with 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside to activate AMP-activated protein kinase. The postexercise increase in sensitivity of muscle glucose transport to activation is not specific for insulin but also involves an increased susceptibility to activation by a submaximal contraction/hypoxia stimulus. The increase in insulin sensitivity is mediated by translocation of more GLUT4 glucose transporters to the cell surface in response to a submaximal insulin stimulus. Although the postexercise increase in muscle insulin sensitivity has been characterized in considerable detail, the basic mechanisms underlying this phenomenon remain a mystery.
Contraction signaling to glucose transport in skeletal muscle
Authors: Jessen Niels , Goodyear Laurie J.
Published: June 2005

Contracting skeletal muscles acutely increases glucose transport in both healthy individuals and in people with Type 2 diabetes, and regular physical exercise is a cornerstone in the treatment of the disease. Glucose transport in skeletal muscle is dependent on the translocation of GLUT4 glucose transporters to the cell surface. It has long been believed that there are two major signaling mechanisms leading to GLUT4 translocation. One mechanism is insulin-activated signaling through insulin receptor substrate-1 and phosphatidylinositol 3-kinase. The other is an insulin-independent signaling mechanism that is activated by contractions, but the mediators of this signal are still unknown. Accumulating evidence suggests that the energy-sensing enzyme AMP-activated protein kinase plays an important role in contraction-stimulated glucose transport. However, more recent studies in transgenic and knockout animals show that AMP-activated protein kinase is not the sole mediator of the signal to GLUT4 translocation and suggest that there may be redundant signaling pathways leading to contraction-stimulated glucose transport. The search for other possible signal intermediates is ongoing, and calcium, nitric oxide, bradykinin, and the Akt substrate AS160 have been suggested as possible candidates. Further research is needed because full elucidation of an insulin-independent signal leading to glucose transport would be a promising pharmacological target for the treatment of Type 2 diabetes.
Expressional reprogramming of survival pathways in rat cardiocytes by neuregulin-1β
Authors: Giraud Marie-Noëlle , Flück Martin , Zuppinger Christian , Suter Thomas M.
Published: June 2005

Neuregulin/ErbB2-induced kinase signaling provides essential survival and protection clues for functional integrity of the adult heart and skeletal muscle. To define the regulatory pathways involved in neuregulin-dependent muscle cell survival, we set out to map the largely unknown transcript targets of this growth/differentiation factor in cardiocytes. Freshly isolated adult primary rat cardiocytes were treated for 24 h with recombinant human neuregulin-1β (NRG-1β, 30 ng/ml). Transcript level alterations in NRG-1β-treated and control cardiocytes ( n = 6) were identified with Atlas Rat Toxicology 1.2 cDNA arrays (BD Clontech) and established permutation L1 regression analysis. Selected transcriptional adjustments were confirmed by RT-PCR and Western blotting. Involvement of MAPK pathways was verified with the inhibitor PD-98059. Application of the single dose of NRG-1β to quiescent cardiocytes induced expressional reprogramming of distinct cellular processes. This response included a prominent 50–100% increase in transcripts of multiple redox systems. It also involved a comparable mRNA augmentation of protein synthetic and folding factors together with augmented message for the trigger of cardiac hypertrophy, cyclin D1 (CCND1). First evidence for a role of neuregulin in promotion of mitochondrial turnover, voltage-gated ion channel expression, and the suppression of fatty acid transporter mRNAs was revealed. Subsequent analysis confirmed a corresponding upregulation of redox factor proteins thioredoxin and the thioredoxin reductase 1, GSTP-1, and CCND1 and demonstrated downregulation of the related transcripts by PD-98059 in neuregulin-stimulated cultures. These MAPK-dependent expressional adjustments point to novel oxidative defense and hypertrophy pathways being involved in the longer lasting protective function of neuregulin in the heart.
Chronic hypoxia modulates relations among calcium, myosin light chain phosphorylation, and force differently in fetal and adult ovine basilar arteries
Authors: Nauli Surya M. , Williams James M. , Gerthoffer William T. , Pearce William J.
Published: June 2005

The present study tests the hypothesis that age-related differences in contractility of cerebral arteries from hypoxic animals involve changes in myofilament Ca2+sensitivity. Basilar arteries from term fetal and nonpregnant adult sheep maintained 110 days at 3,820 m were used in measurements of cytosolic calcium concentration ([Ca2+]i), myosin light chain phosphorylation, and contractile tensions induced by graded concentrations of K+or serotonin (5-HT). Slopes relating [Ca2+]ito tension were similar in fetal (0.83 ± 0.07) and adult (1.02 ± 0.08) arteries for K+contractions but were significantly greater for fetal (3.77 ± 0.64) than adult (2.00 ± 0.13) arteries for 5-HT contractions. For both K+and 5-HT contractions, these relations were left shifted in fetal compared with adult arteries, indicating greater Ca2+sensitivity in fetal arteries. In contrast, slopes relating [Ca2+]iand %myosin phosphorylation for K+contractions were less in fetal (0.37 ± 0.08) than adult (0.81 ± 0.07) arteries, and the fetal curves were right shifted. For 5-HT contractions, the slope of the Ca2+-phosphorylation relation was similar in fetal (0.33 ± 0.09) and adult (0.33 ± 0.23) arteries, indicating that 5-HT depressed Ca2+-induced myosin phosphorylation in adult arteries. For slopes relating %myosin phosphorylation and tension, fetal values (K+: 1.52 ± 0.22, 5-HT: 7.66 ± 1.70) were less than adult values (K+: 2.13 ± 0.30, 5-HT: 8.29 ± 2.40) for both K+- and 5-HT-induced contractions, although again fetal curves were left shifted relative to the adult. Thus, in hypoxia-acclimatized basilar arteries, a downregulated ability of Ca2+to promote myosin phosphorylation is offset by an upregulated ability of phosphorylated myosin to produce force yielding an increased fetal myofilament Ca2+sensitivity. Postnatal maturation reprioritizes the mechanisms regulating hypoxic contractility through changes in the source of activator Ca2+, the pathways governing myosin light chain phosphorylation, and its interaction with actin.
Firing rates of motor units in human vastus lateralis muscle during fatiguing isometric contractions
Authors: Adam Alexander , De Luca Carlo J.
Published: June 2005

We investigated the firing rate of motor units in the vastus lateralis muscle in five healthy young men (mean = 21.4 yr, SD = 0.9) during a sequence of isometric constant-torque contractions repeated to exhaustion. The contractions were sustained at 20% of the maximal voluntary level, measured at the beginning of the test sequence. Electromyographic (EMG) signals were recorded via quadrifilar fine-wire electrodes and subsequently decomposed into their constituent motor unit action potentials to obtain the motor unit firing times. In addition, we measured the whole muscle mechanical properties during the fatigue task using electrical stimulation. The firing rate of motor units first decreased within the first 10–20% of the endurance time of the contractions and then increased. The firing rate increase was accompanied by recruitment of additional motor units as the force output remained constant. The elicited twitch and tetanic torque responses first increased and then decreased. The two processes modulated in a complementary fashion at the same time. Our data suggest that, when the vastus lateralis muscle is activated to maintain a constant torque output, its motoneuron pool receives a net excitatory drive that first decreases to compensate for the short-lived potentiation of the muscle force twitch and then increases to compensate for the diminution of the force twitch. The underlying inverse relationship between the firing rate and the recruitment threshold that has been reported for nonfatigued contractions is maintained. We, therefore, conclude that the central nervous system control of vastus lateralis motor units remains invariant during fatigue in submaximal isometric isotonic contractions.
Target-Specific Short-Term Dynamics Are Important for the Function of Synapses in an Oscillatory Neural Network
Authors: Mamiya Akira , Nadim Farzan
Published: June 2005

Short-term dynamics such as facilitation and depression are present in most synapses and are often target-specific even for synapses from the same type of neuron. We examine the dynamics and possible functions of two synapses from the same presynaptic neuron in the rhythmically active pyloric network of the spiny lobster. Using simultaneous recordings, we show that the synapses from the lateral pyloric (LP) neuron to the pyloric dilator (PD; a member of the pyloric pacemaker ensemble) and the pyloric constrictor (PY) neurons both show short-term depression. However, the postsynaptic potentials produced by the LP-to-PD synapse are larger in amplitude, depress less, and recover faster than those produced by the LP-to-PY synapse. The main function of the LP-to-PD synapse is to slow down the pyloric rhythm. However, in some cases, it slows down the rhythm only when it is fast and has no effect or to speeds up when it is slow. In contrast, the LP-to-PY synapse functions to delay the activity of the PY neuron; this delay increases as the cycle period becomes longer. Using a computational model, we show that the short-term dynamics of synaptic depression observed for each of these synapses are tailored to their individual functions and that replacing the dynamics of either synapse with the other would disrupt these functions. Together, the experimental and modeling results suggest that the target-specific features of short-term synaptic depression are functionally important for synapses efferent from the same presynaptic neuron.
Long-Term Outcome of Ph-Negative Acute Lymphoblastic Leukaemia in Adults: A Single Centre Experience
Authors: Castagnola Carlo , Lunghi Monia , Caberlon Sabrina , Bonfichi Maurizio , Pascutto Cristiana , Lazzarino Mario
Published: June 2005

Background and Objectives: In adult acute lymphoblastic leukaemia (ALL), unlike in childhood ALL, the percentage of long-term remitters and survivors has not improved significantly over the last decades. In the present analysis, we describe a series of adult ALL patients consecutively treated with the same regimen in order to analyse prognostic factors and treatment outcome as well as to define new risk-oriented strategies. Design and Methods: From 1990 to 1998, 102 newly diagnosed ALL patients were referred to our division, 83 of them were eligible for the present study. Median age was 31 years (range 13–76); 77.1% had B-lineage ALL and 22.9% T-lineage ALL; 36.1% showed associated myeloid markers. All patients received an induction phase treatment, consisting of a 4-week cycle with vincristine, daunorubicin, L-asparaginase and desametasone; the consolidation phase included cyclophosphamide, cytarabine, 6-mercaptopurine and central nervous system (CNS) prophylaxis, followed by three of months maintenance (methotrexate + 6-mercaptopurine), re-induction (4-week cycle with vincristine, adriamicin, desametasone), and 2-year maintenance with methotrexate + 6-mercaptopurine. Results: Complete remission (CR) was achieved in 66 patients (79.5%); 20.5% of patients were resistant. The relapse rate was 60.2%. There were 10 CNS relapses (accounting for 12% of all patients, 15% of all CRs and 20% of all relapses). One patient had an ovarian and 2 had a breast relapse. Eleven patients remained in first continuous CR after chemotherapy. Median overall survival (OS) and disease-free survival (DFS) were 1.8 and 1.0 years, respectively, with a median follow-up of 5.6 years (range 0.3–12.1). Initial white blood cell count ≤30 × 109/l, age <35 years, and time to complete remission ≤40 days were the most significant prognostic factors for OS (p < 0.05). These three characteristics defined a group of patients with a better prognosis (5-year OS: 58%). They represented, however, only 18% in this series of adult patients. Interpretation and Conclusions: In our cohort, we were able to define a small subgroup of adult ALL patients with a better outcome. However, the majority of patients is at a high risk of failure when treated with standard protocols. There is a need for new, more active regimens for these patients.