bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison between Disk Diffusion and Microdilution Methods for Determining Susceptibility of Clinical Fungal Isolates to Caspofungin
Authors: Milici Maria Eleonora , Maida Carmelo Massimo , Spreghini Elisabetta , Ravazzolo Barbara , Oliveri Salvatore , Scalise Giorgio , Barchiesi Francesco
Published: August 2007

We compared the caspofungin (CAS) susceptibility testing results generated by the disk diffusion (DD) assay with the results of the Clinical and Laboratory Standards Institute (CLSI) broth microdilution (BD) reference method for 106 yeast isolates. The isolates represented 11 different fungal species, including
Candida albicans
(
n
= 50),
C. parapsilosis
(
n
= 10),
C. glabrata
(
n
= 10),
C. tropicalis
(
n
= 10),
C. guillermondii
(
n
= 6),
C. rugosa
(
n
= 5),
C. krusei
(
n
= 5),
C. kefyr
(
n
= 2),
C. pelliculosa
(
n
= 2),
Saccharomyces cerevisiae
(
n
= 3), and
Geotrichum candidum
(
n
= 3). The DD assay was performed in supplemented Mueller-Hinton agar with CAS, which was tested at concentrations of 2, 10, and 25 μg per disk. MICs and inhibition zone diameters were evaluated at 24 and 48 h. In general, the results obtained by the DD assay correlated well with those obtained by the BD method. In particular, a significant correlation between methods was observed when CAS was used at concentration of 2 μg/disk at a reading time of either 24 or 48 h.
Use of a DNA Microarray for Simultaneous Detection of Antibiotic Resistance Genes among Staphylococcal Clinical Isolates
Authors: Zhu Ling-Xiang , Zhang Zhi-Wei , Wang Can , Yang Hua-Wei , Jiang Di , Zhang Qiong , Mitchelson Keith , Cheng Jing
Published: August 2007

We developed a multiplex asymmetric PCR (MAPCR)-based DNA microarray assay for characterization of the clinically relevant antibiotic resistance genes leading to penicillin, methicillin, aminoglycoside, macrolide, lincosamide, and streptogramin B (MLS
B
) resistance in staphylococci. The DNA-based assay involves detection of specific conserved regions of the
mecA
,
blaZ
(methicillin and penicillin resistance),
aac
(6′)-Ie-
aph
(2‴) (aminoglycoside resistance),
ermA
and
ermC
genes (MLS
B
resistance), and the
msrA
gene (macrolide and streptogramin B resistance). The microarray uses a variable sequence region of the 16S rRNA gene to broadly differentiate between
Staphylococcus aureus
and other coagulase-negative staphylococci (CoNS). The performance of the microarray was validated with a total of 178 clinically important
S. aureus
and 237 CoNS isolates, with correlations of 100% for
S. aureus
to CoNS discrimination and more than 90% for antibiotic resistance between the genotypic analysis determined by the microarray and the phenotype determined by standard methods of species identification and susceptibility testing. The major discrepant results were 17
mecA
-positive CoNS and 60
aac
(6′)-Ie-
aph
(2‴)-positive CoNS isolates measured by microarray that were susceptible to the corresponding antibiotics based on disk diffusion assay. Overall, this microarray-based assay offers a simultaneous, fast (≤5 h), and accurate identification of antibiotic resistance genes from a single colony, as well as species classification. Our extensive validation of the microarray suggests that it may be a useful tool to complement phenotypic susceptibility testing in clinical laboratories and to survey the spread of antibiotic resistance determinants in epidemiological studies.
Predominant Mode of Human Immunodeficiency Virus Transfer between T Cells Is Mediated by Sustained Env-Dependent Neutralization-Resistant Virological Synapses
Authors: Chen Ping , Hübner Wolfgang , Spinelli Matthew A. , Chen Benjamin K.
Published: August 2007

Cell-free human immunodeficiency virus type 1 (HIV-1) can initiate infections, but contact between infected and uninfected T cells can enhance viral spread through intercellular structures called virological synapses (VS). The relative contribution of VS to cell-free viral transfer has not been carefully measured. Using an ultrasensitive, fluorescent virus transfer assay, we estimate that when VS between HIV-expressing Jurkat T cells and primary CD4
+
T cells are formed, cell-associated transfer of virus is 18,000-fold more efficient than uptake of cell-free virus. Furthermore, in contrast to cell-free virus uptake, the VS deposits virus rapidly into focal, trypsin-resistant compartments in target T cells. This massive virus internalization requires Env-CD4 receptor interactions but is resistant to inhibition by patient-derived neutralizing antisera that inhibit homologous cell-free virus. Deleting the Env cytoplasmic tail does not abrogate VS-mediated transfer, but it renders the VS sensitive to neutralizing antibodies, suggesting that the tail limits exposure of VS-neutralizing epitopes on the surface of infected cells. Dynamic live imaging of the VS reveals that HIV-expressing cells are polarized and make sustained, Env-dependent contacts with target cells through uropod-like structures. The polarized T-cell morphology, Env-CD4 coordinated adhesion, and viral transfer from HIV-infected to uninfected cells suggest that VS allows HIV-1 to evade antibody neutralization and to disseminate efficiently. Future studies will discern to what extent this massive viral transfer contributes to productive infection or viral dissemination through the migration of virus-carrying T cells.
Sequence of Human Immunodeficiency Virus Type 1 (HIV-1) Gag Localization and Oligomerization Monitored with Live Confocal Imaging of a Replication-Competent, Fluorescently Tagged HIV-1
Authors: Hübner Wolfgang , Chen Ping , Portillo Armando Del , Liu Yuxin , Gordon Ronald E. , Chen Benjamin K.
Published: August 2007

The assembly of infectious human immunodeficiency virus (HIV) requires that Gag transport and oligomerization be coordinated with its association with other viral proteins, viral RNAs, and cellular membranes. We have developed a replication-competent HIV type 1 molecular clone that carries a Gag-internal or interdomain green fluorescent protein (iGFP) fusion to reveal a physiologically accurate temporal sequence of Gag localization and oligomerization during the formation of infectious HIV. This recombinant HIV is as infectious as native HIV in single-round infectivity assays, validating its use for trafficking studies. It replicates robustly in permissive MT4 cells and is infectious, yet it spreads poorly in other T-cell lines. Immunofluorescence of Gag-iGFP showed a pattern very similar to that of native Gag. However, the intense plasma membrane Gag-iGFP fluorescence contrasts markedly with its immunofluorescence at this site, indicating that many Gag epitopes can be masked by oligomerization. Consistent with this, fluorescence resonance energy transfer studies visualized intense Gag oligomerization at the plasma membrane and weaker oligomerization at cytoplasmic sites. Four-dimensional, time-lapse confocal imaging reveals a temporal progression of Gag distribution over hours in which Gag is initially diffusely localized within the cytoplasm. Plasma membrane signals then accumulate as Gag levels increase and vesicular association appears late, only after plasma membrane site signals have reached high intensity. Lastly, the cell rounds up and HIV protease activation induces diffuse fluorescence throughout the cell. These distinct phases reveal a natural progression of Gag trafficking during the viral gene expression program. HIV Gag-iGFP is a useful tool for dissecting mechanisms of viral assembly and transmission.
Identification of Acetylated, Tetrahalogenated Benzimidazole
d
-Ribonucleosides with Enhanced Activity against Human Cytomegalovirus
Authors: Hwang Jae-Seon , Kregler Oliver , Schilf Rita , Bannert Norbert , Drach John C. , Townsend Leroy B. , Bogner Elke
Published: August 2007

DNA packaging is the key step in viral maturation and involves binding and cleavage of viral DNA containing specific DNA-packaging motifs. This process is mediated by a group of specific enzymes called terminases. We previously demonstrated that the human cytomegalovirus (HCMV) terminase is composed of the large subunit pUL56 and the small subunit pUL89. While the large subunit mediates sequence-specific DNA binding and ATP hydrolysis, pUL89 is required only for duplex nicking. An excellent inhibitor targeting HCMV terminase is 2-bromo-5,6-dichloro-1-(β-
d
-ribofuranosyl)benzimidazole (BDCRB), but it was not developed as an antiviral drug due to its metabolic cleavage in experimental animals. We now have tested several new benzimidazole
d
-ribonucleosides in order to determine whether these compounds represent new, potent inhibitors. Analysis by bioluminometric ATPase activity assays identified two of the new compounds with a high inhibitory effect, 2-bromo-4,5,6-trichloro-1-(2,3,5-tri-
O
-acetyl-β-
d
-ribofuranosyl) benzimidazole (BTCRB) and 2,4,5,6-tetrachloro-1-(2,3,5-tri-
O
-acetyl-β-
d
-ribofuranosyl benzimidazole (Cl
4
RB). By using viral plaque formation, viral yield, and viral growth kinetics, we demonstrated that the two compounds BTCRB and Cl
4
RB had antiviral activities similar to that of BDCRB. Interestingly, BTCRB retained its inhibitory activity after preincubation with HFF cells. By use of electron microscopy, we observed an increase of B capsids and a lack of cytoplasmic capsids in the presence of the compounds that correlated with the virus yield. Furthermore, cleavage of concatenated DNA was inhibited by both compounds, and inhibition by BTCRB was shown to be dose dependent. These results demonstrate that the new compounds are highly active against HCMV and act by mechanisms similar but not identical to those of BDCRB.
Characterization of Low-Pathogenicity H5N1 Avian Influenza Viruses from North America
Authors: Spackman Erica , Swayne David E. , Suarez David L. , Senne Dennis A. , Pedersen Janice C. , Killian Mary Lea , Pasick John , Handel Katherine , Pillai Smitha P. Somanathan , Lee Chang-Won , Stallknecht David , Slemons Richard , Ip Hon S. , Deliberto Tom
Published: August 2007

Wild-bird surveillance in North America for avian influenza (AI) viruses with a goal of early identification of the Asian H5N1 highly pathogenic AI virus has identified at least six low-pathogenicity H5N1 AI viruses between 2004 and 2006. The hemagglutinin (HA) and neuraminidase (NA) genes from all 6 H5N1 viruses and an additional 38 North American wild-bird-origin H5 subtype and 28 N1 subtype viruses were sequenced and compared with sequences available in GenBank by phylogenetic analysis. Both HA and NA were phylogenetically distinct from those for viruses from outside of North America and from those for viruses recovered from mammals. Four of the H5N1 AI viruses were characterized as low pathogenicity by standard in vivo pathotyping tests. One of the H5N1 viruses, A/MuteSwan/MI/451072-2/06, was shown to replicate to low titers in chickens, turkeys, and ducks. However, transmission of A/MuteSwan/MI/451072-2/06 was more efficient among ducks than among chickens or turkeys based on virus shed. The 50% chicken infectious dose for A/MuteSwan/MI/451072-2/06 and three other wild-waterfowl-origin H5 viruses were also determined and were between 10
5.3
and 10
7.5
50% egg infective doses. Finally, seven H5 viruses representing different phylogenetic clades were evaluated for their antigenic relatedness by hemagglutination inhibition assay, showing that the antigenic relatedness was largely associated with geographic origin. Overall, the data support the conclusion that North American H5 wild-bird-origin AI viruses are low-pathogenicity wild-bird-adapted viruses and are antigenically and genetically distinct from the highly pathogenic Asian H5N1 virus lineage.
Escape and Compensation from Early HLA-B57-Mediated Cytotoxic T-Lymphocyte Pressure on Human Immunodeficiency Virus Type 1 Gag Alter Capsid Interactions with Cyclophilin A
Authors: Brockman Mark A. , Schneidewind Arne , Lahaie Matthew , Schmidt Aaron , Miura Toshiyuki , DeSouza Ivna , Ryvkin Faina , Derdeyn Cynthia A. , Allen Susan , Hunter Eric , Mulenga Joseph , Goepfert Paul A. , Walker Bruce D. , Allen Todd M.
Published: August 2007

Certain histocompatibility leukocyte antigen (HLA) alleles are associated with improved clinical outcomes for individuals infected with human immunodeficiency virus type 1 (HIV-1), but the mechanisms for their effects remain undefined. An early CD8+T-cell escape mutation in the dominant HLA-B57-restricted Gag epitope TW10 (TSTLQEQIGW) has been shown to impair HIV-1 replication capacity in vitro. We demonstrate here that this T242N substitution in the capsid protein is associated with upstream mutations at residues H219, I223, and M228in the cyclophilin A (CypA)-binding loop in B57+individuals with progressive disease. In an independent cohort of epidemiologically linked transmission pairs, the presence of these substitutions in viruses encoding T242N was associated with significantly higher plasma viremia in donors, further suggesting that these secondary mutations compensated for the replication defect of T242N. Using NL4-3 constructs, we illustrate the ability of these CypA loop changes to partially restore replication of the T242N variant in vitro. Notably, these mutations also enhanced viral resistance to the drug cyclosporine A, indicating a reduced dependence of the compensated virus on CypA that is normally essential for optimal infectivity. Therefore, mutations in TW10 allow HIV-1 to evade a dominant early CD8+T-cell response, but the benefits of escape are offset by a defect in capsid function. These data suggest that TW10 escape variants undergo a postentry block that is partially overcome by changes in the CypA-binding loop and identify a mechanism for an HIV-1 fitness defect that may contribute to the slower disease progression associated with HLA-B57.
An Active TRIM5 Protein in Rabbits Indicates a Common Antiviral Ancestor for Mammalian TRIM5 Proteins
Authors: Schaller Torsten , Hué Stéphane , Towers Greg J.
Published: August 2007

The recent identification of antiretroviral tripartite motif-bearing restriction factors that protect against retroviral infection has revealed a novel branch of innate immunity. The factors target the retroviral capsid and inhibit infectivity soon after the capsid has entered the cytoplasm by an incompletely characterized mechanism. Restriction is species specific. For example, TRIM5α from Old World monkeys, but not humans, restricts human immunodeficiency virus type 1 infection. Here, we identify an antiviral TRIM5 molecule in rabbits that is closely related to antiviral TRIM5 of both primates and cattle. We demonstrate that the rabbit TRIM5 protein is active against divergent retroviruses and leads to a strong block to viral DNA synthesis and infectivity. Furthermore, we show that antiviral activity is directed against the viral capsid and that human TRIM5 proteins are dominant negative to restriction in rabbit cells. We propose that the sequence and restriction characteristics conserved between restriction factors from primates, cattle, and rabbits indicate that these factors have evolved from a common ancestor with antiretroviral properties.
Myxoma Virus Expressing Human Interleukin-12 Does Not Induce Myxomatosis in European Rabbits
Authors: Stanford Marianne M. , Barrett John W. , Gilbert Philippe-Alexandre , Bankert Richard , McFadden Grant
Published: August 2007

Myxoma virus (MV) is a candidate for oncolytic virotherapy due to its ability to selectively infect and kill tumor cells, yet MV is a species-specific pathogen that causes disease only in European rabbits. To assess the ability of MV to deliver cytokines to tumors, we created an MV (vMyxIL-12) that expresses human interleukin-12 (IL-12). vMyxIL-12 replicates similarly to wild-type MV, and virus-infected cells secrete bioactive IL-12. Yet, vMyxIL-12 does not cause myxomatosis, despite expressing the complete repertoire of MV proteins. Thus, vMyxIL-12 exhibits promise as an oncolytic candidate and is safe in all known vertebrate hosts, including lagomorphs.
Reliability of a new shoulder laxometer to assess inferior glenohumeral joint translation
Authors: Sein M L , Appleyard R C , Walton J R , Bradley T , Murrell G A
Published: August 2007

Objective:

This study tested the reliability of the ORI laxometer, a newly developed non-invasive device for testing inferior translation of the humeral head in humans.

Design:

The instrument was designed to measure displacement between the top of the acromion and the head of the humerus when loaded in a similar fashion to the sulcus sign.

Participants and interventions:

Sixteen healthy subjects (32 shoulders) were measured for inferior glenohumeral joint laxity. One observer used the laxometer in 16 subjects on three separate occasions for the intra-observer trial. Three observers measured the inferior shoulder laxity of six subjects on one occasion in the inter-observer trial. Asymptomatic and unstable shoulders were also compared in 12 subjects with shoulder instability.

Main outcome measurements:

Translation in the glenohumeral joint

Results:

The range of inferior translation of glenohumeral joint in these subjects was between 0.01 mm and 6.5 mm with a mean of 1.5 mm. The intraclass correlation coefficient (ICC) for inter-observer reliability was 0.74. For intra-observer reliability the ICC was 0.76. These results are considered to be good to excellent. There was no advantage gained by using data from all five cycles of testing compared with three cycles. In patients with shoulder instability, laxometer measurements were significantly greater in their unstable shoulders than in their normal shoulders.

Conclusions:

The laxometer is easy to use, painless and gives objective measures for inferior glenohumeral laxity, with good intra- and inter-observer reliabilities. The ORI laxometer may be useful for assessing and monitoring global glenohumeral joint laxity which usually involves inferior laxity.
The effect of chromosome 17 polysomy on HER-2/neu status in breast cancer
Authors: Hyun C L , Lee H E , Kim K S , Kim S-W , Kim J H , Choe G , Park S Y
Published: August 2007

Aim:

To investigate the effect of polysomy 17 on HER-2 status as evaluated by immunohistochemistry (IHC), dual-colour fluorescence in situ hybridisation (FISH) and chromogenic in situ hybridisation (CISH).

Methods:

Dual-probe FISH and single-probe CISH were performed to detect HER-2 gene amplification, and IHC to detect HER-2 expression, on 309 invasive breast cancers.

Results:

Polysomy 17 was detected in 32.0% of the total number of breast cancers; it was detected in 12.3% of the IHC 0 or 1+ cases, 42.8% of the IHC 2+ cases and 66.0% of the IHC 3+ cases (p<0.001). In addition, there was a substantially higher rate of polysomy 17 in the IHC 2+ or 3+/FISH-negative cases than in the IHC 0 or 1+ cases (40.8% vs 12.3%; p<0.001). The FISH and CISH results were concordant in 299 cases (96.8%). Of the 10 discordant cases, FISH suggested amplification in five with disomy 17 and one with monosomy 17, whereas CISH pointed to borderline copy numbers in each of these six cases. The remaining four cases had high polysomy 17; CISH, but not FISH, indicated amplification.

Conclusions:

Results suggest that an increase of HER-2 gene copy number secondary to polysomy 17 leads to HER-2 overexpression in some IHC 2+/3+ breast cancers, without gene amplification. The high level of concordance between FISH and CISH suggests that CISH is a valid alternative to FISH for assessing HER-2 gene amplification. However, cases in which CISH indicates the presence of borderline copy numbers or low levels of amplification may need FISH to rule out polysomy 17 and to determine HER-2 gene amplification status accurately.
A dynamic model of ethical reasoning in speech pathology
Authors: Kenny Belinda , Lincoln Michelle , Balandin Susan
Published: August 2007

Ten new graduate speech pathologists recounted their experiences in managing workplace ethical dilemmas in semi-structured interviews. Their stories were analysed for elements that described the nature and management of the ethical dilemmas. Ethical reasoning themes were generated to reflect the participants’ approaches to managing these dilemmas. Finally, a conceptual model, the Dynamic Model of Ethical Reasoning, was developed. This model incorporates the elements of awareness, independent problem solving, supported problem solving, and decision and outcome evaluation. Features of the model demonstrate the complexity of ethical reasoning and the challenges that new graduates encounter when managing ethical dilemmas. The results have implications for preparing new graduates to manage ethical dilemmas in the workplace.
Ethics, justification and the prevention of spina bifida
Authors: Gagen Wendy Jane , Bishop Jeffrey P
Published: August 2007

During the 1970s, prenatal screening technologies were in their infancy, but were being swiftly harnessed to uncover and prevent spina bifida. The historical rise of this screening process and prevention programme is analysed in this paper, and the role of ethical debates in key studies, editorials and letters reported in theLancet, and other related texts and governmental documents between 1972 and 1983, is considered. The silence that surrounded rigorous ethical debate served to highlight where discussion lay—namely, within the justifications offered for the prevention of spina bifida, and the efficacy and benefits of screening. In other words, the ethical justification for screening and prevention of spina bifida, when the authors are not explicitly interested in ethics, is considered. These justifications held certain notions of disability as costly to society, with an imperative to screen and prevent spina bifida for the good of the society.
Proxy consent: moral authority misconceived
Authors: Wrigley A
Published: August 2007

The Mental Capacity Act 2005 has provided unified scope in the British medical system for proxy consent with regard to medical decisions, in the form of a lasting power of attorney. While the intentions are to increase the autonomous decision making powers of those unable to consent, the author of this paper argues that the whole notion of proxy consent collapses into a paternalistic judgement regarding the other person’s best interests and that the new legislation introduces only an advisor, not a proxy with the moral authority to make treatment decisions on behalf of another. The criticism is threefold. First, there is good empirical evidence that people are poor proxy decision makers as regards accurately representing other people’s desires and wishes, and this is therefore a pragmatically inadequate method of gaining consent. Second, philosophical theory explaining how we represent other people’s thought processes indicates that we are unlikely ever to achieve accurate simulations of others’ wishes in making a proxy decision. Third, even if we could accurately simulate other people’s beliefs and wishes, the current construction of proxy consent in the Mental Capacity Act means that it has no significant ethical authority to match that of autonomous decision making. Instead, it is governed by a professional, paternalistic, best-interests judgement that undermines the intended role of a proxy decision maker. The author argues in favour of clearly adopting the paternalistic best-interests option and viewing the proxy as solely an advisor to the professional medical team in helping make best-interests judgements.
Rapid cold water formation and recrystallization of relict bryophyte tufa at the Fall Creek cold springs, Alberta, Canada
Authors: Rainey Dustin K , Jones Brian
Published: August 2007

The Fall Creek Tufa is a relict, mid- to late-Holocene freshwater carbonate spring deposit that is formed almost entirely of calcite-encased Cratoneuron commutatum gametophytes. The spring is presently active, emitting 13 °C meteorically derived waters, but it is not precipitating calcite. Wood embedded in the youngest part of the tufa yielded a radiocarbon (14C) date of 310 ± 50 years BP. The oldest parts of the deposit could not be dated by radiocarbon methods because of a lack of embedded organics. The Fall Creek Tufa, however, probably began forming ~5000 years BP in response to the same wet climatic conditions that initiated calcite precipitation at Miette Hot Springs in Jasper National Park and the Cave and Basin Hot Springs in Banff National Park. The relict deposit is composed largely of calcite cements that were precipitated around the bryophytes, with lesser amounts of internal sediments. The tufa formed through a repeated four-stage developmental process that involved (I) encrustation, (II) encapsulation, (III) cavity occlusion, and (IV) diagenetic alteration. These stages were temporally and spatially variable. Although a young deposit, much of it has undergone extensive aggrading recrystallization, whereby crystal size has increased as adjacent calcite crystals were sutured together. Epifluorescence micro s copy shows that primary depositional fabrics were masked, but not destroyed, during recrystallization. Narrow ranges of δ18O (VPDB, Vienna Pee-Dee Belemnite standard) values (–18.4 to –17.7) and δ13C (VPDB) values (–1.2 to 0.5) indicate that the calcite was in isotopic equilibrium with the water that it precipitated from, and that diagenesis did not reset the stable oxygen and carbon isotopic compositions.
Glycosides from the North Atlantic sea cucumber Cucumaria frondosa V - Structures of five new minor trisulfated triterpene oligoglycosides, frondosides A7-1, A7-2, A7-3, A7-4, and isofrondoside C
Authors: Silchenko Alexandra S , Avilov Sergey A , Kalinovsky Anatoly I , Dmitrenok Pavel S , Kalinin Vladimir I , Morre Jeffrey , Deinzer Max L , Woodward Carl , Collin Peter D
Published: August 2007

Seven highly polar trisulfated triterpene glycosides belonging to the frondoside A7 group have been isolated from the North Atlantic sea cucumber Cucumaria frondosa. The structures of five new glycosides, frondosides A7-1 (1), A7-2 (2), A7-3 (3), A7-4 (4), and isofrondoside C (5) were elucidated, three of which contained lanostane aglycons without a lactone-ring. Three pairs of the glycosides proved to be isomers by virture of the positions of double bonds in the aglycons nuclei; two pairs of the glycosides differed from each other by the character of the oxygen functionality at C-22. The results from these studies provided the basis for postulating the biosynthetic pathways of norlanostane glycosides in the sea cucumber.Key words: Cucumaria frondosa, sea cucumbers, frondosides, triterpene glycosides.
KAUFFMAN STATE SUMS AND BRACKET DEFORMATION
Authors: APOSTOLAKIS NIKOS , KAISER UWE
Published: August 2007

We derive a formula expanding the bracket with respect to a natural deformation parameter. The expansion is in terms of a two-variable polynomial algebra of diagram resolutions generated by basic operations involving the Goldman bracket. A functorial characterization of this algebra is given. Differentiability properties of the star product underlying the Kauffman bracket are discussed.
Localization of AMP kinase is regulated by stress, cell density, and signaling through the MEK→ERK1/2 pathway
Authors: Kodiha Mohamed , Rassi James G. , Brown Claire M. , Stochaj Ursula
Published: August 2007

5′-AMP-activated protein kinase (AMPK) serves as an energy sensor and is at the center of control for a large number of metabolic reactions, thereby playing a crucial role in Type 2 diabetes and other human diseases. AMPK is present in the nucleus and cytoplasm; however, the mechanisms that regulate the intracellular localization of AMPK are poorly understood. We have now identified several factors that control the distribution of AMPK. Environmental stress regulates the intracellular localization of AMPK, and upon recovery from heat shock or oxidant exposure AMPK accumulates in the nuclei. We show that under normal growth conditions AMPK shuttles between the nucleus and the cytoplasm, a process that depends on the nuclear exporter Crm1. However, nucleocytoplasmic shuttling does not take place in high-density cell cultures, for which AMPK is confined to the cytoplasm. Furthermore, we demonstrate that signaling through the mitogen-activated protein kinase kinase (MEK)→extracellular signal-regulated kinase 1/2 (ERK1/2) cascade plays a crucial role in controlling the proper localization of AMPK. As such, pharmacological inhibitors that interfere with this pathway alter AMPK distribution under nonstress conditions. Taken together, our studies identify novel links between the physiological state of the cell, the activation of MEK→ERK1/2 signaling, and the nucleocytoplasmic distribution of AMPK. This sets the stage to develop new strategies to regulate the intracellular localization of AMPK and thereby the modification of targets that are relevant to human disease.
Allogeneic bone marrow transplantation for hepatocellular carcinoma: hepatocyte growth factor suppresses graft-vs.-host disease
Authors: Yoshida Yasuhiko , Hirano Tadamichi , Son Gakuhei , Iimuro Yuji , Imado Takehito , Iwasaki Tsuyoshi , Fujimoto Jiro
Published: August 2007

Allogeneic bone-marrow transplantation (BMT) can induce a powerful graft-vs.-tumor (GVT) effect not only on hematological malignancies but also on solid tumors. However, graft-vs.-host disease (GVHD) is a major complication of allogeneic BMT. We assessed GVT effect on hepatocellular carcinoma (HCC) and the effects of hepatocyte growth factor (HGF) gene transduction on GVHD in HCC transplanted mice. (C57BL/6 × C3H/HeJ)F1(B6C3F1, H-2bxk) mice were used as recipients and C3H/HeJ(H-2k) mice were used as donors. Hepa1-a (a C57L mouse-derived hepatoma cell, H-2b) was subcutaneously injected into the recipient mice. Tumor bearing mice were treated in the following ways: group 1, no treatment; group 2, total body irradiation (TBI); group 3, TBI and BMT; group 4, TBI and BMT with empty vector; group 5, TBI and BMT with HGF gene transduction; group 6, TBI and BMT with administration of FK506, a representative immunosuppressive agent. Acute GVHD was assessed by histological examination of the liver, small intestines, and large intestines. Tumor growth was markedly suppressed in mice that received an allogeneic BMT. Donor-derived CD8+T cells had infiltrated into the tumor, and cytotoxic CD8+T cells against HCC were present. However, among the four groups that received a BMT, this suppressive effect was weaker in group 6 compared with the other three groups ( groups 3, 4, and 5). HGF gene transduction improved GVHD while preserving the GVT effects. Allogeneic BMT markedly suppresses the growth of HCC. Simultaneous HGF gene transfer can suppress GVHD while preserving the GVT effect.
Blockade of purinergic 2 receptors attenuates the mechanoreceptor component of the exercise pressor reflex
Authors: Kindig Angela E. , Hayes Shawn G. , Kaufman Marc P.
Published: August 2007

The finding that pyridoxalphosphate-6-azophenyl-2,4-disulfonic acid (PPADS), a P2 antagonist, attenuated the pressor response to calcaneal tendon stretch, a purely mechanical stimulus, raises the possibility that P2 receptors sensitize mechanoreceptors to static contraction of the triceps surae muscles. The mechanical component of the exercise pressor reflex, which is evoked by static contraction, can be assessed by measuring renal sympathetic nerve activity during the first 2–5 s of this maneuver. During this period of time, group III mechanoreceptors often discharge explosively in response to the sudden tension developed at the onset of contraction. In decerebrated cats, we, therefore, examined the effect of PPADS (10 mg/kg) injected into the popliteal artery on the renal sympathetic and pressor responses to contraction and stretch. We found that PPADS significantly attenuated the renal sympathetic response to contraction, with the effect starting 2 s after its onset and continuing throughout its 60-s period. PPADS also significantly attenuated the renal sympathetic nerve response to stretch, but did so after a latency of 10 s. Our findings lead us to conclude that P2 receptors sensitize group III muscle afferents to contraction. The difference in the onset latency between the PPADS-induced attenuation of the renal sympathetic response to contraction and the renal sympathetic response to stretch is probably due to the sensitivities of different populations of group III afferents to ATP released during contraction and stretch.
Mechanisms of impaired calcium handling underlying subclinical diastolic dysfunction in diabetes
Authors: Lacombe Véronique A. , Viatchenko-Karpinski Serge , Terentyev Dmitry , Sridhar Arun , Emani Sitaramesh , Bonagura John D. , Feldman David S. , Györke Sandor , Carnes Cynthia A.
Published: August 2007

Isolated diastolic dysfunction is found in almost half of asymptomatic patients with well-controlled diabetes and may precede diastolic heart failure. However, mechanisms that underlie diastolic dysfunction during diabetes are not well understood. We tested the hypothesis that isolated diastolic dysfunction is associated with impaired myocardial Ca2+ handling during type 1 diabetes. Streptozotocin-induced diabetic rats were compared with age-matched placebo-treated rats. Global left ventricular myocardial performance and systolic function were preserved in diabetic animals. Diabetes-induced diastolic dysfunction was evident on Doppler flow imaging, based on the altered patterns of mitral inflow and pulmonary venous flows. In isolated ventricular myocytes, diabetes resulted in significant prolongation of action potential duration compared with controls, with afterdepolarizations occurring in diabetic myocytes ( P < 0.05). Sustained outward K+ current and peak outward component of the inward rectifier were reduced in diabetic myocytes, while transient outward current was increased. There was no significant change in L-type Ca2+ current; however, Ca2+ transient amplitude was reduced and transient decay was prolonged by 38% in diabetic compared with control myocytes ( P < 0.05). Sarcoplasmic reticulum Ca2+ load (estimated by measuring the integral of caffeine-evoked Na+-Ca2+ exchanger current and Ca2+ transient amplitudes) was reduced by ∼50% in diabetic myocytes ( P < 0.05). In permeabilized myocytes, Ca2+ spark amplitude and frequency were reduced by 34 and 20%, respectively, in diabetic compared with control myocytes ( P < 0.05). Sarco(endo)plasmic reticulum Ca2+-ATPase-2a protein levels were decreased during diabetes. These data suggest that in vitro impairment of Ca2+ reuptake during myocyte relaxation contributes to in vivo diastolic dysfunction, with preserved global systolic function, during diabetes.
Liver injury after an aggressive encounter in male mice
Authors: Sánchez Olga , Viladrich Meritxell , Ramírez Ignasi , Soley Maria
Published: August 2007

Acute and intense psychological stressors induce cell damage in several organs, including the heart and the liver. Much less is known about social stress. In male mice, aggressive behavior is the most common social stressor. It is remarkable that upon fighting, submandibular salivary glands release a number of peptides into the bloodstream including epidermal growth factor (EGF). We showed previously that released EGF protects the heart from cell damage in this particular stressful situation. Here, we studied the effect of an aggressive encounter on the liver and whether EGF has a similar effect on this organ. An aggressive encounter in male mice caused inflammatory response and a transient increase in plasma alanine and aspartate transaminase activities. At 3 h, focal infiltration of neutrophils was observed in liver parenchyma. These cells accumulate on eosinophilic hepatocytes, which may correspond to dying cells. A few hours later, evidence of necrotic lesion was observed. Surgical excision of submandibular glands, sialoadenectomy, did not prevent the rise in plasma EGF concentration and did not affect the increase in plasma transaminase activities. Neither did the administration of tyrphostin AG-1478 (inhibitor of EGF receptor kinase) alter the increase in plasma alanine transaminase activity. However, it did enhance the rise in both aspartate transaminase and creatine kinase activity, suggesting heart damage. We conclude that an aggressive encounter causes mild liver damage and that released EGF does not protect this organ, in contrast to its effect on the heart.
Intestinal carbonic anhydrase, bicarbonate, and proton carriers play a role in the acclimation of rainbow trout to seawater
Authors: Grosell Martin , Gilmour Katie M. , Perry Steven F.
Published: August 2007

Abrupt transfer of rainbow trout from freshwater to 65% seawater caused transient disturbances in extracellular fluid ionic composition, but homeostasis was reestablished 48 h posttransfer. Intestinal fluid chemistry revealed early onset of drinking and slightly delayed intestinal water absorption that coincided with initiation of NaCl absorption and HCO3−secretion. Suggestive of involvement in osmoregulation, relative mRNA levels for vacuolar H+-ATPase (V-ATPase), Na+-K+-ATPase, Na+/H+exchanger 3 (NHE3), Na+-HCO3−cotransporter 1, and two carbonic anhydrase (CA) isoforms [a general cytosolic isoform trout cytoplasmic CA (tCAc) and an extracellular isoform trout membrane-bound CA type IV (tCAIV)], were increased transiently in the intestine following exposure to 65% seawater. Both tCAc and tCAIV proteins were localized to apical regions of the intestinal epithelium and exhibited elevated enzymatic activity after acclimation to 65% seawater. The V-ATPase was localized to both basolateral and apical regions and exhibited a 10-fold increase in enzymatic activity in fish acclimated to 65% seawater, suggesting a role in marine osmoregulation. The intestinal epithelium of rainbow trout acclimated to 65% seawater appears to be capable of both basolateral and apical H+extrusion, likely depending on osmoregulatory status and intestinal fluid chemistry.
Mechanisms responsible for the enhanced pumping capacity of the in situ winter flounder heart (Pseudopleuronectes americanus)
Authors: Mendonça Paula C. , Genge A. Gaylene , Deitch Eric J. , Gamperl A. Kurt
Published: August 2007

In situ Starling and power output curves and in vitro pressure-volume curves were determined for winter flounder hearts, as well as the hearts of two other teleosts (Atlantic salmon and cod). In situ maximum cardiac output was not different between the three species (∼62 ml·min−1·kg−1). However, because of the small size of the flounder heart, maximum stroke volume per milliliter per gram ventricle was significantly greater (2.3) compared with cod (1.7) and salmon (1.4) and is the highest reported for teleosts. The maximum power output of the flounder heart (7.6 mW/g) was significantly lower than that measured in the salmon (9.7) and similar to the cod (7.8) but was achieved at a much lower output pressure (4.9 vs. 8.0 and 6.2 kPa, respectively). Although the flounder heart could not perform resting levels of cardiac function at subambient pressures, it was much more sensitive to filling pressure, a finding supported by pressure-volume curves, which showed that the flounder's heart chambers were more compliant. Finally, we report that the flounder's bulbus:ventricle mass ratio (0.59) was significantly higher than in the cod (0.37) and salmon (0.22). These data, which support previous studies suggesting that the flatfish cardiovascular system is a high-volume, low-pressure design, show that vis-à-fronte filling is not important in flatfish, and that some fish can achieve high levels of cardiac output by vis-à-tergo filling alone; and suggest that a large compliant bulbus assists the flounder heart in delivering extremely large stroke volumes at pressures that do not become limiting.
Neonatal dietary supplementation of arachidonic acid increases prostaglandin levels in adipose tissue but does not promote fat mass development in guinea pigs
Authors: Aprikian Olivier , Reynaud Denis , Pace-Asciak Cecil , Leone Patricia , Blancher Florence , Monnard Irina , Darimont Christian , Macé Katherine
Published: August 2007

The role of arachidonic acid (AA) on the development of adipose tissue is still controversial since its metabolites, i.e., prostaglandins, can either stimulate or inhibit preadipocyte differentiation in vitro. In the present study, we evaluated the effects of early postnatal supplementation of AA on body weight and adipose tissue development in guinea pigs. Male newborn guinea pigs were fed for 21 days ( day 21) with diets (milk and pellet) supplemented (+AA) or not (−AA) with 1.2% (total fatty acids) AA. From day 21 to day 105 both groups were fed a chow diet. The 21-days-old +AA pups showed a twofold higher AA accretion in phospholipids associated with a two- to sixfold increase in several prostaglandins, such as 6-keto PGF1α(the stable hydrolysis product of PGI2), PGF2α, PGE2, and PGD2in adipose tissue, compared with the −AA group. No difference in fat pad and body weight, aP2, and leptin gene expression in adipose tissue, fasting plasma glucose, free-fatty acids, and triglyceride concentration was observed between groups at day 21 or day 105. These results show that dietary supplementation of AA during the suckling/weaning period increases prostaglandin levels in adipose tissue but does not influence early fat mass development in the guinea pig.
A novel mechanism of renal blood flow autoregulation and the autoregulatory role of A1adenosine receptors in mice
Authors: Just Armin , Arendshorst William J.
Published: August 2007

Autoregulation of renal blood flow (RBF) is mediated by a fast myogenic response (MR; ∼5 s), a slower tubuloglomerular feedback (TGF; ∼25 s), and potentially additional mechanisms. A1adenosine receptors (A1AR) mediate TGF in superficial nephrons and contribute to overall autoregulation, but the impact on the other autoregulatory mechanisms is unknown. We studied dynamic autoregulatory responses of RBF to rapid step increases of renal artery pressure in mice. MR was estimated from autoregulation within the first 5 s, TGF from that at 5–25 s, and a third mechanism from 25–100 s. Genetic deficiency of A1AR (A1AR−/−) reduced autoregulation at 5–25 s by 50%, indicating a residual fourth mechanism resembling TGF kinetics but independent of A1AR. MR and third mechanism were unaltered in A1AR−/−. Autoregulation in A1AR−/− was faster at 5–25 than at 25–100 s suggesting two separate mechanisms. Furosemide in wild-type mice (WT) eliminated the third mechanism and enhanced MR, indicating TGF-MR interaction. In A1AR−/−, furosemide did not further impair autoregulation at 5–25 s, but eliminated the third mechanism and enhanced MR. The resulting time course was the same as during furosemide in WT, indicating that A1AR do not affect autoregulation during furosemide inhibition of TGF. We conclude that at least one novel mechanism complements MR and TGF in RBF autoregulation, that is slower than MR and TGF and sensitive to furosemide, but not mediated by A1AR. A fourth mechanism with kinetics similar to TGF but independent of A1AR and furosemide might also contribute. A1AR mediate classical TGF but not TGF-MR interaction.
Integration of jugular venous return and circle of Willis in a theoretical human model of selective brain cooling
Authors: Neimark Matthew A. , Konstas Angelos-Aristeidis , Laine Andrew F. , Pile-Spellman John
Published: August 2007

A three-dimensional mathematical model was developed to examine the induction of selective brain cooling (SBC) in the human brain by intracarotid cold (2.8°C) saline infusion (ICSI) at 30 ml/min. The Pennes bioheat equation was used to propagate brain temperature. The effect of cooled jugular venous return was investigated, along with the effect of the circle of Willis (CoW) on the intracerebral temperature distribution. The complete CoW, missing A1 variant (mA1), and fetal P1 variant (fP1) were simulated. ICSI induced moderate hypothermia (defined as 32–34°C) in the internal carotid artery (ICA) territory within 5 min. Incorporation of the complete CoW resulted in a similar level of hypothermia in the ICA territory. In addition, the anterior communicating artery and ipsilateral posterior communicating artery distributed cool blood to the contralateral anterior and ipsilateral posterior territories, respectively, imparting mild hypothermia (35 and 35.5°C respectively). The mA1 and fP1 variants allowed for sufficient cooling of the middle cerebral territory (30–32°C). The simulations suggest that ICSI is feasible and may be the fastest method of inducing hypothermia. Moreover, the effect of convective heat transfer via the complete CoW and its variants underlies the important role of CoW anatomy in intracerebral temperature distributions during SBC.
Visual-Shift Adaptation Is Composed of Separable Sensory and Task-Dependent Effects
Authors: Simani M. C. , McGuire L. M. M. , Sabes P. N.
Published: August 2007

Visuomotor coordination requires both the accurate alignment of spatial information from different sensory streams and the ability to convert these sensory signals into accurate motor commands. Both of these processes are highly plastic, as illustrated by the rapid adaptation of goal-directed movements following exposure to shifted visual feedback. Although visual-shift adaptation is a widely used model of sensorimotor learning, the multifaceted adaptive response is typically poorly quantified. We present an approach to quantitatively characterizing both sensory and task-dependent components of adaptation. Sensory aftereffects are quantified with “alignment tests” that provide a localized, two-dimensional measure of sensory recalibration. These sensory effects obey a precise form of “additivity,” in which the shift in sensory alignment between vision and the right hand is equal to the vector sum of the shifts between vision and the left hand and between the right and left hands. This additivity holds at the exposure location and at a second generalization location. These results support a component transformation model of sensory coordination, in which eye–hand and hand–hand alignment relies on a sequence of shared sensory transformations. We also ask how these sensory effects compare with the aftereffects measured in target reaching and tracking tasks. We find that the aftereffect depends on both the task performed during feedback-shift exposure and on the testing task. The results suggest the presence of both a general sensory recalibration and task-dependent sensorimotor effect. The task-dependent effect is observed in highly stereotyped reaching movements, but not in the more variable tracking task.
Corticofugal Modulation of the Tactile Response Coherence of Projecting Neurons in the Gracilis Nucleus
Authors: Castellanos Nazareth P. , Malmierca Eduardo , Nuñez Angel , Makarov Valeri A.
Published: August 2007

Precise and reproducible spike timing is one of the alternatives of the sensory stimulus encoding. We test coherence (repeatability) of the response patterns elicited in projecting gracile neurons by tactile stimulation and its modulation provoked by electrical stimulation of the corticofugal feedback from the somatosensory (SI) cortex. To gain the temporal structure we adopt the wavelet-based approach for quantification of the functional stimulus–neural response coupling. We show that the spontaneous firing patterns (when they exist) are essentially random. Tactile stimulation of the neuron receptive field strongly increases the spectral power in the stimulus and 5- to 15-Hz frequency bands. However, the functional coupling (coherence) between the sensory stimulus and the neural response exhibits ultraslow oscillation (0.07 Hz). During this oscillation the stimulus coherence can temporarily fall below the statistically significant level, i.e., the functional stimulus–response coupling may be temporarily lost for a single neuron. We further demonstrate that electrical stimulation of the SI cortex increases the stimulus coherence for about 60% of cells. We find no significant correlation between the increment of the firing rate and the stimulus coherence, but we show that there is a positive correlation with the amplitude of the peristimulus time histogram. The latter argues that the observed facilitation of the neural response by the corticofugal pathway, at least in part, may be mediated through an appropriate ordering of the stimulus-evoked firing pattern, and the coherence enhancement is more relevant in gracilis nucleus than an increase of the number of spikes elicited by the tactile stimulus.