bluuebunny/crossref_metadata_2025 huggingface dataset:
Contribution of Endocytic Motifs in the Cytoplasmic Tail of Herpes Simplex Virus Type 1 Glycoprotein B to Virus Replication and Cell-Cell Fusion
Authors: Beitia Ortiz de Zarate Igor , Cantero-Aguilar Lilia , Longo Magalie , Berlioz-Torrent Clarisse , Rozenberg Flore
Published: October 2007

The use of endocytic pathways by viral glycoproteins is thought to play various functions during viral infection. We previously showed in transfection assays that herpes simplex virus type 1 (HSV-1) glycoprotein B (gB) is transported from the cell surface back to the
trans
-Golgi network (TGN) and that two motifs of gB cytoplasmic tail, YTQV and LL, function distinctly in this process. To investigate the role of each of these gB trafficking signals in HSV-1 infection, we constructed recombinant viruses in which each motif was rendered nonfunctional by alanine mutagenesis. In infected cells, wild-type gB was internalized from the cell surface and concentrated in the TGN. Disruption of YTQV abolished internalization of gB during infection, whereas disruption of LL induced accumulation of internalized gB in early recycling endosomes and impaired its return to the TGN. The growth of both recombinants was moderately diminished. Moreover, the fusion phenotype of cells infected with the gB recombinants differed from that of cells infected with the wild-type virus. Cells infected with the YTQV-mutated virus displayed reduced cell-cell fusion, whereas giant syncytia were observed in cells infected with the LL-mutated virus. Furthermore, blocking gB internalization or impairing gB recycling to the cell surface, using drugs or a transdominant negative form of Rab11, significantly reduced cell-cell fusion. These results favor a role for endocytosis in virus replication and suggest that gB intracellular trafficking is involved in the regulation of cell-cell fusion.
RNA Editing of the Human Herpesvirus 8 Kaposin Transcript Eliminates Its Transforming Activity and Is Induced during Lytic Replication
Authors: Gandy Sharon Z. , Linnstaedt Sarah D. , Muralidhar Sumitra , Cashman Kathleen A. , Rosenthal Leonard J. , Casey John L.
Published: October 2007

Human herpesvirus 8 is the etiologic agent associated with Kaposi's sarcoma and primary effusion lymphoma (PEL). The K12 RNA, which produces as many as three variants of the kaposin protein, as well as a microRNA, is the most abundant transcript expressed in latent Kaposi's sarcoma-associated herpesvirus infection, and yet it is also induced during lytic replication. The portion of the transcript that includes the microRNA and the kaposin A sequence has been shown to have tumorigenic potential. Genome coordinate 117990, which is within this transcript, has been found to be heterogeneous, primarily in RNAs but also among viral DNA sequences. This sequence heterogeneity affects an amino acid in kaposins A and C and the microRNA. The functional effects of this sequence heterogeneity have not been studied, and its origin has not been definitively settled; both RNA editing and heterogeneity at the level of the viral genome have been proposed. Here, we show that transcripts containing A at position 117990 are tumorigenic, while those with G at this position are not. Using a highly sensitive quantitative assay, we observed that, in PEL cells under conditions where more than 60% of cDNAs derived from K12 RNA transcripts have G at coordinate 117990, there is no detectable G in the viral DNA sequence at this position, only A. This result is consistent with RNA editing by one of the host RNA adenosine deaminases (ADARs). Indeed, we observed that purified human ADAR1 efficiently edits K12 RNA in vitro. Remarkably, the amount of editing correlated with the replicative state of the virus; editing levels were nearly 10-fold higher in cells treated to induce lytic viral replication. These results suggest that RNA editing controls the function of one segment of the kaposin transcript, such that it has transforming activity during latent replication and possibly another, as-yet-undetermined, function during lytic replication.
Adhesion Molecule Interactions Facilitate Human Immunodeficiency Virus Type 1-Induced Virological Synapse Formation between T Cells
Authors: Jolly Clare , Mitar Ivonne , Sattentau Quentin J.
Published: October 2007

Human immunodeficiency virus type 1 (HIV-1) can spread between CD4+T cells by using a virological synapse (VS). The VS assembly is a cytoskeleton-driven process dependent on HIV-1 envelope glycoprotein (Env)-receptor engagement and is hypothesized to require adhesion molecule interactions. Here we demonstrate that leukocyte function-associated antigen 1 (LFA-1), intercellular adhesion molecule 1 (ICAM-1), and ICAM-3 are enriched at the VS and that inhibition of these interactions influences conjugate formation and reduces VS assembly. Moreover, CD4+T cells deficient in LFA-1 or with modified LFA-1 function were less able to support VS assembly and cell-cell transfer of HIV-1. Thus, cognate adhesion molecule interactions at the VS are important for HIV-1 spread between T cells.
Tioman Virus, a Paramyxovirus of Bat Origin, Causes Mild Disease in Pigs and Has a Predilection for Lymphoid Tissues
Authors: Yaiw Koon Chu , Bingham John , Crameri Gary , Mungall Bruce , Hyatt Alex , Yu Meng , Eaton Bryan , Shamala Devi , Wang Lin-Fa , Thong Wong Kum
Published: October 2007

Disease manifestation, pathology, and tissue tropism following infection with Tioman virus (TioPV), a newly isolated, bat-derived paramyxovirus, was investigated in subcutaneously (
n
= 12) and oronasally (
n
= 4) inoculated pigs. Pigs were either asymptomatic or developed pyrexia, but all of the animals produced neutralizing antibodies. The virus (viral antigen and/or genome) was detected in lymphocytes of the thymus, tonsils, spleen, lymph nodes and Peyer's patches (ileum), tonsillar epithelium, and thymic epithelioreticular cells. Virus was isolated from oral swabs but not from urine. Our findings suggest that the pig could act as an intermediate or amplifying host for TioPV and that oral secretion is a possible means of viral transmission.
Human Papillomavirus Type 16 E7 Oncoprotein Associates with the Centrosomal Component γ-Tubulin
Authors: Nguyen Christine L. , Eichwald Catherine , Nibert Max L. , Münger Karl
Published: October 2007

Expression of a high-risk human papillomavirus (HPV) E7 oncoprotein is sufficient to induce aberrant centrosome duplication in primary human cells. The resulting centrosome-associated mitotic abnormalities have been linked to the development of aneuploidy. HPV type 16 (HPV16) E7 induces supernumerary centrosomes through a mechanism that is at least in part independent of the inactivation of the retinoblastoma tumor suppressor pRb and is dependent on cyclin-dependent kinase 2 activity. Here, we show that HPV16 E7 can concentrate around mitotic spindle poles and that a small pool of HPV16 E7 is associated with centrosome fractions isolated by sucrose density gradient centrifugation. The targeting of HPV16 E7 to the centrosome, however, was not sufficient for centrosome overduplication. Nonetheless, we found that HPV16 E7 can associate with the centrosomal regulator γ-tubulin and that the recruitment of γ-tubulin to the centrosome is altered in HPV16 E7-expressing cells. Since the association of HPV16 E7 with γ-tubulin is independent of pRb, p107, and p130, our results suggest that the association with γ-tubulin contributes to the pRb/p107/p130-independent ability of HPV16 E7 to subvert centrosome homeostasis.
Allogeneic Differences in the Dependence on CD4+T-Cell Help for Virus-Specific CD8+T-Cell Differentiation
Authors: Kemball Christopher C. , Szomolanyi-Tsuda Eva , Lukacher Aron E.
Published: October 2007

CD4+T-cell help enables antiviral CD8+T cells to differentiate into fully competent memory cells and sustains CD8+T-cell-mediated immunity during persistent virus infection. We recently reported that mice of C57BL/6 and C3H strains differ in their dependence on CD28 and CD40L costimulation for long-term control of infection by polyoma virus, a persistent mouse pathogen. In this study, we asked whether mice of these inbred strains also vary in their requirement for CD4+T-cell help for generating and maintaining polyoma virus-specific CD8+T cells. CD4+T-cell-depleted C57BL/6 mice mounted a robust antiviral CD8+T-cell response during acute infection, whereas unhelped CD8+T-cell effectors in C3H mice were functionally impaired during acute infection and failed to expand upon antigenic challenge during persistent infection. Using (C57BL/6 × C3H)F1mice, we found that the dispensability for CD4+T-cell help for the H-2b-restricted polyoma virus-specific CD8+T-cell response during acute infection extends to the H-2k-restricted antiviral CD8+T cells. Our findings demonstrate that dependence on CD4+T-cell help for antiviral CD8+T-cell effector differentiation can vary among allogeneic strains of inbred mice.
Targeted Deletion of Regions Rich in Immune-Evasive Genes from the Cytomegalovirus Genome as a Novel Vaccine Strategy
Authors: Čičin-Šain Luka , Bubić Ivan , Schnee Margit , Ruzsics Zsolt , Mohr Christian , Jonjić Stipan , Koszinowski Ulrich H.
Published: October 2007

Human cytomegalovirus (CMV), a ubiquitous human pathogen, is a leading cause of congenital infections and represents a serious health risk for the immunosuppressed patient. A vaccine against CMV is currently not available. CMV is characterized by its large genome and by multiple genes modulating the immunity of the host, which cluster predominantly at genome termini. Here, we tested whether the deletion of gene blocks rich in immunomodulatory genes could be used as a novel concept in the generation of immunogenic but avirulent, herpesvirus vaccines. To generate an experimental CMV vaccine, we selectively deleted 32 genes from the mouse cytomegalovirus (MCMV) genome. The resulting mutant grew to titers similar to that of wild-type MCMV in vitro. In vivo, the mutant was 10,000-fold attenuated and well tolerated, even by highly susceptible mice deficient for B, T, and NK cells or for the interferon type I receptor. Equally relevant for safety concerns, immune suppression did not lead to the mutant's reactivation from latency. Immunization with the replication-competent mutant, but not with inactivated virus, resulted in protective immunity, which increased over time. Vaccination induced MCMV-specific antibodies and a strong T-cell response. We propose that a targeted and rational approach can improve future herpesvirus vaccines and vaccine vectors.
EVALUATION OF RELATIVISTIC TRANSITION MATRIX ELEMENTS OF HYDROGENIC ATOMS USING PLANE WAVE EXPANSION
Authors: SOLDATOV A. V. , SEKE J. , ADAM G.
Published: October 2007

For the first time to our knowledge, a general, explicit formula for exact transition matrix elements in relativistic hydrogenic atoms is derived, by using the plane-wave expansion for the electromagnetic-field vector potential. By applying the obtained formula, discrete-discrete and discrete-continuous matrix elements are evaluated analytically and numerically.
SUBALGEBRAS OF THE LIE ALGEBRA R6AND THEIR APPLICATIONS
Authors: ZHANG YU-FENG , FAN EN-GUI
Published: October 2007

As we all know, the Hamiltonian systems are the same describing forms as Newton mechanics and Lagrange mechanics. Therefore, researching for a new Hamiltonian structure of the soliton equations has important significance. In the paper, firstly, with the help of the Lie algebra R6, a few types of subalgebras are constructed, from which the corresponding equivalent tensor systems are given. For their applications, two integrable couplings hierarchies along with the multi-potential component functions generated from the soliton theory and the Virasoro symmetric algebra are obtained. Secondly, the Hamiltonian structures of the above integrable couplings are worked out, which may become another describing expression for the Newton and Lagrange mechanics. In particular, one of the integrable couplings presented above reduces to the famous AKNS hierarchy of soliton equations.
TWO-DIMENSIONAL ANISOTROPIC HEISENBERG FERROMAGNET IN COEXISTING TRANSVERSE AND LONGITUDINAL MAGNETIC FIELDS
Authors: HU AI-YUAN , CHEN YUAN
Published: October 2007

The two-dimensional spin-1/2 anisotropic Heisenberg ferromagnet is investigated in coexisting transverse and longitudinal magnetic fields. Using the Green function treatment, the magnetization and susceptibility are studied as a function of temperature, anisotropy and magnetic fields. The effects of exchange anisotropy and transverse magnetic field on the magnetic properties of the system are discussed.
Impact of genetic background and aging on mesenteric collateral growth capacity in Fischer 344, Brown Norway, and Fischer 344 × Brown Norway hybrid rats
Authors: Sheridan Kevin M. , Ferguson Michael J. , Distasi Matthew R. , Witzmann Frank A. , Dalsing Michael C. , Miller Steven J. , Unthank Joseph L.
Published: September 2007

Available studies indicate that both genetic background and aging influence collateral growth capacity, but it is not known how their combination affects collateral growth. We evaluated collateral growth induced by ileal artery ligation in Fischer 344 (F344), Brown Norway (BN), and the first generation hybrid of F344 × BN (F1) rats available for aging research from the National Institute on Aging. Collateral growth was determined by paired diameter measurements in anesthetized rats immediately and 7 days postligation. In 3-mo-old rats, significant collateral growth occurred only in BN (35% ± 11%, P < 0.001). The endothelial cell number in arterial cross sections was also determined, since this precedes shear-mediated luminal expansion. When compared with the same animal controls, the intimal cell number was increased only in BN rats (92% ± 21%, P < 0.001). The increase in intimal cell number and the degree of collateral luminal expansion in BN rats was not affected by age from 3 to 24 mo. Immunohistochemical studies demonstrated that intimal cell proliferation was much greater in the collaterals of BN than of F1 rats. The remarkable difference between these three strains of rats used in aging research and the lack of an age-related impairment in the BN rats are novel observations. These rat strains mimic clinical observations of interindividual variation in collateral growth capacity and the impact of age on arteriogenesis and should be useful models to investigate the molecular mechanisms responsible for such differences.
Na+/H+ exchanger inhibitor cariporide attenuates the mitochondrial Ca2+ overload and PTP opening
Authors: Toda Takako , Kadono Toshie , Hoshiai Minako , Eguchi Yu , Nakazawa Shinpei , Nakazawa Hiroe , Higashijima Naoko , Ishida Hideyuki
Published: September 2007

The Na+/H+ exchanger (NHE) inhibitor cariporide has a cardioprotective effect in various animal models of myocardial ischemia-reperfusion. Recent studies have suggested that cariporide interacts with mitochondrial Ca2+ overload and the mitochondrial permeability transition (MPT); however, the precise mechanisms remain unclear. Therefore, we examined whether cariporide affects mitochondrial Ca2+ overload and MPT. Isolated adult rat ventricular myocytes were used to study the effects of cariporide on hypercontracture induced by ouabain or phenylarsine oxide (PAO). Mitochondrial Ca2+ concentration ([Ca2+]m) and the mitochondrial membrane potential (ΔΨm) were measured by loading myocytes with rhod-2 and JC-1, respectively. We also examined the effect of cariporide on the MPT using tetramethylrhodamine methyl ester (TMRM) and oxidative stress generated by laser illumination. Cariporide (1 μM) prevented ouabain-induced hypercontracture (from 40 ± 2 to 24 ± 2%, P < 0.05) and significantly attenuated ouabain-induced [Ca2+]m overload (from 149 ± 6 to 121 ± 5% of the baseline value, P < 0.05) but did not affect ΔΨm. These results indicate that cariporide attenuates the [Ca2+]m overload without the accompanying depolarization of ΔΨm. Moreover, cariporide increased the time taken to induce the MPT (from 79 ± 11 to 137 ± 20 s, P < 0.05) and also attenuated PAO-induced hypercontracture (from 59 ± 3 to 50 ± 4%, P < 0.05). Our data indicate that cariporide attenuates [Ca2+]m overload and MPT. Thus these effects might potentially contribute to the mechanisms of cardioprotection afforded by NHE inhibitors.
Insulin-induced myocardial protection in isolated ischemic rat hearts requires p38 MAPK phosphorylation of Hsp27
Authors: Li Gefeng , Ali Imtiaz S. , Currie R. William
Published: September 2007

Six hours after insulin treatment, hearts express heat shock protein 70 (Hsp70) and have improved contractile function after ischemia-reperfusion injury. In this study we examined hearts 1 h after insulin treatment for contractile function and for expression of Hsp70 and Hsp27. Adult, male Sprague-Dawley rats were assigned to groups: 1) sham, 2) control, 3) insulin injected (200 μU/g body wt), 4) heat shock treated (core body temperature, 42°C for 15 min), and 5) heat shock and insulin treated. At 1 h after these treatments, hearts were isolated, equilibrated to Langendorff perfusion for 30 min, and then subjected for 30 min no-flow global ischemia (37°C) followed by 2 h of reperfusion. Insulin-treated hearts had significantly increased contractile function compared with control hearts. At 1 h after insulin treatment, a minimal change in Hsp70 and Hsp27 content were detected. By 3 h after insulin treatment, a significant increase in Hsp70, but not Hsp27, was detected by Western blot analysis. By immunofluorescence, minimal Hsp70 was detected in insulin-treated hearts, whereas Hsp27 was detected in all hearts, indicative of its constitutive expression. Phosphospecific isoforms of Hsp27 were detected in insulin-, heat shock-, and heat shock and insulin-treated hearts. After ischemia and reperfusion, the insulin-treated hearts had significantly elevated levels of phosphorylated Hsp27. Inhibition of p38 MAPK with SB-203580 blocked the insulin-induced phosphorylation of Hsp27 and the improved functional recovery. In conclusion, insulin induces an apparent rapid phosphorylation of Hsp27 that is associated with improved functional recovery after ischemia-reperfusion injury.
Enhanced mitochondrial superoxide in hyperglycemic endothelial cells: direct measurements and formation of hydrogen peroxide and peroxynitrite
Authors: Quijano Celia , Castro Laura , Peluffo Gonzalo , Valez Valeria , Radi Rafael
Published: September 2007

Hyperglycemic challenge to bovine aortic endothelial cells (BAECs) increases oxidant formation and cell damage that are abolished by MnSOD overexpression, implying mitochondrial superoxide (O2•−) as a central mediator. However, mitochondrial O2•−and its steady-state concentrations have not been measured directly yet. Therefore, we aimed to detect and quantify O2•−through different techniques, along with the oxidants derived from it. Mitochondrial aconitase, a sensitive target of O2•−, was inactivated 60% in BAECs incubated in 30 mM glucose (hyperglycemic condition) with respect to cells incubated in 5 mM glucose (normoglycemic condition). Under hyperglycemic conditions, increased oxidation of the mitochondrially targeted hydroethidine derivative (MitoSOX) to hydroxyethidium, the product of the reaction with O2•−, could be specifically detected. An 8.8-fold increase in mitochondrial O2•−steady-state concentration (to 250 pM) and formation rate (to 6 μM/s) was estimated. Superoxide formation increased the intracellular concentration of both hydrogen peroxide, measured as 3-amino-2,4,5-triazole-mediated inactivation of catalase, and nitric oxide-derived oxidants (i.e., peroxynitrite), evidenced by immunochemical detection of 3-nitrotyrosine. Oxidant formation was further evaluated by chloromethyl dichlorodihydrofluorescein (CM-H2DCF) oxidation. Exposure to hyperglycemic conditions triggered the oxidation of CM-H2DCF and was significantly reduced by pharmacological agents that lower the mitochondrial membrane potential, inhibit electron transport (i.e., myxothiazol), and scavenge mitochondrial oxidants (i.e., MitoQ). In BAECs devoid of mitochondria (rho0cells), hyperglycemic conditions did not increase CM-H2DCF oxidation. Mitochondrial O2•−formation in hyperglycemic conditions was associated with increased glucose metabolization in the Krebs cycle and hyperpolarization of the mitochondrial membrane.
Atherogenic vascular and lipid phenotypes in young patients with Type 1 diabetes are associated with diabetes high-risk HLA genotype
Authors: Odermarsky Michal , Nilsson Anita , Lernmark Åke , Sjöblad Sture , Liuba Petru
Published: September 2007

Expression of human leukocyte antigen (HLA) class II molecules on islet endothelial cells is a central vascular event in the pathogenesis of Type 1 diabetes. Previous studies demonstrated the ability of other vascular endothelial cells to express HLA and thereby to process islet autoantigens on their surface. We investigated whether the HLA-DQ2/8 genotype, which confers the highest risk for Type 1 diabetes, is associated with early atherosclerosis in youths with this disease. Brachial artery endothelium-dependent, flow-mediated dilation (BA-FMD) and carotid artery intima-media thickness (CA-IMT), as well as markers of systemic inflammation [C-reactive protein (CRP), fibrinogen, and orosomucoid], HbA1C, LDL, HDL, and total cholesterol, were assessed in 86 children and adolescents with Type 1 diabetes (mean age and diabetes duration, 15 and 7 yr, respectively) between 2004 and 2006. HLA genotypes were determined in dried blood spots by an oligoblot hybridization method. As a result, HLA-DQ2/8 was detected in 34 patients (DQ2/8). When this group was compared with the remaining patients (non-DQ2/8, n = 52), there were no differences in age, diabetes duration, HbA1C, body mass index, inflammatory markers, and IMT ( P ≥ 0.4). In the DQ2/8 group, LDL-to-HDL ratio was elevated compared with that in the non-DQ2/8 group (1.8 vs. 1.3, respectively; P = 0.001), whereas FMD did not significantly differ between the groups (5.3% vs. 6.7%, respectively; P = 0.08). When patients were further categorized in relation to CRP (cut-off value, 1 mg/l), BA-FMD was significantly lower (3%, P < 0.01), whereas LDL-to-HDL ratio increased further (2.2, P < 0.001) in the subgroup of DQ2/8 and CRP ≥ 1 patients compared with the remaining three subgroups. These associations remained significant after adjustment for age, diabetes duration, and HbA1Cby analysis of covariance. The brachial artery responses to nitroglycerine were similar in all subgroups. In conclusion, the diabetes-predisposing HLA-DQ2/8 genotype in children and adolescents with Type 1 diabetes interferes with endothelial and lipid-related mechanisms of early atherosclerosis, possibly in part through inflammatory pathways.
Effects of voluntary wheel running on cardiac function and myosin heavy chain in chemically gonadectomized rats
Authors: Hydock David S. , Lien Chia-Ying , Schneider Carole M. , Hayward Reid
Published: September 2007

Reducing testosterone and estrogen levels with a luteinizing hormone-releasing hormone agonist such as Zoladex (i.e., chemical gonadectomy) is a common treatment for many prostate and breast cancer patients, respectively. There are reports of surgical gonadectomy inducing cardiac dysfunction, and exercise has been shown to be cardioprotective under these circumstances. Minimal research has been done investigating the effects of chemical gonadectomy and increased physical activity on cardiac function. The purpose of this investigation was to examine the effects of chemical gonadectomy and physical activity on cardiac function. Male (M) and female (F) Sprague-Dawley rats received either Zoladex treatment (Zol) that suppressed gonadal function for 8 wk or control implants (Con) and either were allowed unlimited access to voluntary running wheels (WR) or remained sedentary (Sed) throughout the treatment period. In vivo and ex vivo left ventricle (LV) function were then assessed, and myosin heavy chain (MHC) expression was analyzed to help explain LV functional differences. Hearts from M Sed+Zol exhibited significantly lower aortic blood flow velocity, developed pressure, and maximal rate of pressure development and higher β-MHC expression than M Sed+Con. Hearts from F Sed+Zol exhibited significantly lower LV wall thicknesses, fractional shortening, and developed pressure and higher β-MHC expression than F Sed+Con. This cardiac dysfunction was not evident in hearts from M or F WR+Zol, and this was associated with a preservation of the MHC isoform distribution. Thus an 8-wk chemical gonadectomy with Zoladex promoted cardiac dysfunction in male and female rats, and voluntary wheel running protected against this cardiac dysfunction.
p38 and ERK1/2 MAPKs mediate the interplay of TNF-α and IL-10 in regulating oxidative stress and cardiac myocyte apoptosis
Authors: Dhingra Sanjiv , Sharma Anita K. , Singla Dinender K. , Singal Pawan K.
Published: September 2007

It is known that TNF-α increases the production of ROS and decreases antioxidant enzymes, resulting in an increase in oxidative stress. IL-10 appears to modulate these effects. The present study investigated the role of p38 and ERK1/2 MAPKs in mediating the interplay of TNF-α and IL-10 in regulating oxidative stress and cardiac myocyte apoptosis in Sprague-Dawley male rats. Isolated adult cardiac myocytes were exposed to TNF-α (10 ng/ml), IL-10 (10 ng/ml), and IL-10 + TNF-α ( ratio 1) for 4 h. H2O2(100 μM) as a positive control and the antioxidant Trolox (20 μmol/l) were used to confirm the involvement of oxidative stress. H2O2treatment increased oxidative stress and apoptosis; TNF-α mimicked these effects. Exposure to TNF-α significantly increased ROS production, caused cell injury, and increased the number of apoptotic cells and Bax-to-Bcl-xl ratio. This change was associated with an increase in the phospho-p38 MAPK-to-total p38 MAPK ratio and a decrease in the phospho-ERK1/2-to-total ERK1/2 ratio. IL-10 treatment by itself had no effect on these parameters, but it prevented the above-listed changes caused by TNF-α. The antioxidant Trolox modulated TNF-α-induced changes in Bax/Bcl-xl, cell injury, and MAPKs. Preexposure of cells to the p38 MAPK inhibitor SB-203580 prevented TNF-α-induced changes. Inhibition of the ERK pathway with PD-98059 attenuated the protective role of IL-10 against TNF-α-induced apoptosis. This study provides evidence in support of the essential role of p38 and ERK1/2 MAPKs in the interactive role of TNF-α and IL-10 in cardiac myocyte apoptosis.
Anabolic steroids induce cardiac renin-angiotensin system and impair the beneficial effects of aerobic training in rats
Authors: Rocha F. L. , Carmo E. C. , Roque F. R. , Hashimoto N. Y. , Rossoni L. V. , Frimm C. , Anéas I. , Negrão C. E. , Krieger J. E. , Oliveira E. M.
Published: September 2007

We evaluated the effects of swimming and anabolic steroids (AS) on ventricular function, collagen synthesis, and the local renin-angiotensin system in rats. Male Wistar rats were randomized into control (C), steroid (S; nandrolone decanoate; 5 mg/kg sc, 2×/wk), steroid + losartan (SL; 20 mg·kg−1·day−1), trained (T), trained + steroid (T+S), and trained + steroid + losartan (T+SL; n = 14/group) groups. Swimming was performed 5 times/wk for 10 wk. Serum testosterone increased in S and T+S. Resting heart rate was lower in T and T+S. Percent change in left ventricular (LV) weight-to-body weight ratio increased in S, T, and T+S. LV systolic pressure declined in S and T+S. LV contractility increased in T ( P < 0.05). LV relaxation increased in T ( P < 0.05). It was significantly lower in T+S compared with C. Collagen volumetric fraction (CVF) and hydroxyproline were higher in S and T+S than in C and T ( P < 0.05), and the CVF and LV hypertrophy were prevented by losartan treatment. LV-ANG I-converting enzyme activity increased (28%) in the S group (33%), and type III collagen synthesis increased (56%) in T+S but not in T group. A positive correlation existed between LV-ANG I-converting enzyme activity and collagen type III expression ( r2 = 0.88; P < 0.05, for all groups). The ANG II and angiotensin type 1a receptor expression increased in the S and T+S groups but not in T group. Supraphysiological doses of AS exacerbated the cardiac hypertrophy in exercise-trained rats. Exercise training associated with AS induces maladaptive remodeling and further deterioration in cardiac performance. Exercise training associated with AS causes loss of the beneficial effects in LV function induced by exercising. These results suggest that aerobic exercise plus AS increases cardiac collagen content associated with activation of the local renin-angiotensin system.
Surfactant protein A regulates IgG-mediated phagocytosis in inflammatory neutrophils
Authors: Wofford Jessica A. , Wright Jo Rae
Published: September 2007

Surfactant proteins (SP)-A and SP-D have been shown to affect the functions of a variety of innate immune cells and to interact with various immune proteins such as complement and immunoglobulins. The goal of the current study is to test the hypothesis that SP-A regulates IgG-mediated phagocytosis by neutrophils, which are major effector cells of the innate immune response that remove invading pathogens by phagocytosis and by extracellular killing mediated by reactive oxygen and nitrogen. We have previously shown that SP-A stimulates chemotaxis by inflammatory, but not peripheral, neutrophils. To evaluate the ability of SP-A to modulate IgG-mediated phagocytosis, polystyrene beads were coated with BSA and treated with anti-BSA IgG. SP-A significantly and specifically enhanced IgG-mediated phagocytosis by inflammatory neutrophils, but it had no effect on beads not treated with IgG. SP-A bound to IgG-coated beads and enhanced their uptake via direct interactions with the beads as well as direct interactions with the neutrophils. SP-A did not affect reactive oxygen production or binding of IgG to neutrophils and had modest effects on polymerization of actin. These data suggest that SP-A plays an important role in mediating the phagocytic response of neutrophils to IgG-opsonized particles.
ENaC α-subunit variants are expressed in lung epithelial cells and are suppressed by oxidative stress
Authors: Xu Haishan , Chu Shijian
Published: September 2007

Amiloride-sensitive epithelial sodium channel (ENaC) is a major sodium channel in the lung facilitating fluid absorption. ENaC is composed of α-, β-, and γ-subunits, and the α-subunit is indispensable for ENaC function in the lung. In human lungs, the α-subunit is expressed as various splice variants. Among them, α1- and α2-subunits are two major variants with different upstream regulatory sequences that possess similar channel characteristics when tested in Xenopus oocytes. Despite the importance of α-ENaC, little was known about the relative abundance of its variants in lung epithelial cells. Furthermore, lung infection and inflammation are often accompanied by reduced α-ENaC expression, oxidative stress, and pulmonary edema. However, it was not clear how oxidative stress affects expression of α-ENaC variants. In this study, we examined relative expression levels of α-subunit variants in four human lung epithelial cell lines. We also tested the hypothesis that oxidative stress inhibits α-ENaC expression. Our results show that both α1- and α2-ENaC variants are expressed in the cells we tested, but relative abundance varies. In the two monolayer-forming cell lines, H441 and Calu-3, α2-ENaC is the predominant variant. We also show that H2O2 specifically suppresses α1- and α2-ENaC variant expression in H441 and Calu-3 cells in a dose-dependent fashion. This suppression is achieved by inhibition of their promoters and is attenuated by dexamethasone. These data demonstrate the importance of the α2-subunit variant and suggest that glucocorticoids and antioxidants may be useful in correcting infection/inflammation-induced lung fluid imbalance.
Association of VASP with TRPC4 in PKG-mediated inhibition of the store-operated calcium response in mesangial cells
Authors: Wang Xiaoxia , Pluznick Jennifer L. , Settles Deann C. , Sansom Steven C.
Published: October 2007

We tested the hypotheses that the NO-cGMP-PKG pathway mediates inhibition of the store-operated cation channel (SOC) in human glomerular mesangial cells (HMC) and that TRPC4, a molecular component of SOC in HMC, is associated with PKG-phosphorylated vasodilator-stimulated phosphoprotein (VASP). Using fura 2 ratiometry, we measured intracellular Ca2+concentration [Ca2+]ito determine whether sodium nitroprusside (SNP), an NO donor, and 8-Br-cGMP affected SOC-TRPC4 via PKG. We found that the SOC response in HMC was attenuated in the presence of 100 μM SNP, an NO donor, or 100 μM 8-Br-cGMP. Addition of DT-3 (2.5 μM), a specific PKG-1α inhibitor, reversed the effects of 8-Br-cGMP on the SOC response. Application of 100 μM cAMP did not significantly inhibit the SOC response. RT-PCR and Western blotting revealed PKG-1α transcript and protein in HMC. Immunocytochemical analysis localized PKG-1α to the cytoplasm and plasma membrane of HMC. Previous studies have shown that PKG-mediated phosphorylation of VASP attenuates cellular Ca2+entry, resulting in altered growth and proliferation. Therefore, we used Western blotting and immunocytochemistry to determine whether PKG-phosphorylated VASP associates with TRPC4. Western blot analysis revealed that 8-Br-cGMP enhanced the phosphorylation of VASP at serine 239 (Ser239), a known PKG phosphorylation site, in HMC within 5 min. Coimmunoprecipitation and coimmunostaining showed that P-Ser239-VASP associated with TRPC4. However, VASP that was unphosphorylated at Ser239 was not associated with TRPC4. These results indicate that VASP has a role in the NO/PKG-1α-mediated inhibition of the TRPC4-SOC response in HMC.
Synapsin Regulates Basal Synaptic Strength, Synaptic Depression, and Serotonin-Induced Facilitation of Sensorimotor Synapses in Aplysia
Authors: Fioravante Diasinou , Liu Rong-Yu , Netek Anne K. , Cleary Leonard J. , Byrne John H.
Published: October 2007

Synapsin is a synaptic vesicle-associated protein implicated in the regulation of vesicle trafficking and transmitter release, but its role in heterosynaptic plasticity remains elusive. Moreover, contradictory results have obscured the contribution of synapsin to homosynaptic plasticity. We previously reported that the neuromodulator serotonin (5-HT) led to the phosphorylation and redistribution of Aplysia synapsin, suggesting that synapsin may be a good candidate for the regulation of vesicle mobilization underlying the short-term synaptic plasticity induced by 5-HT. This study examined the role of synapsin in homosynaptic and heterosynaptic plasticity. Overexpression of synapsin reduced basal transmission and enhanced homosynaptic depression. Although synapsin did not affect spontaneous recovery from depression, it potentiated 5-HT–induced dedepression. Computational analysis showed that the effects of synapsin on plasticity could be adequately simulated by altering the rate of Ca2+-dependent vesicle mobilization, supporting the involvement of synapsin not only in homosynaptic but also in heterosynaptic forms of plasticity by regulating vesicle mobilization.
Peptide Hormone Modulation of a Neuronally Modulated Motor Circuit
Authors: Kirby Matthew S. , Nusbaum Michael P.
Published: October 2007

Rhythmically active motor circuits are influenced by neuronally released and circulating hormone modulators, but there are few systems in which the influence of a peptide hormone modulator on a neuronally modulated motor circuit has been determined. We performed such an analysis in the isolated crab stomatogastric nervous system by assessing the influence of the hormone crustacean cardioactive peptide (CCAP) on the gastric mill (chewing) rhythm elicited by identified modulatory projection neurons. The gastric mill circuit is located in the stomatogastric ganglion. In situ, this ganglion is located within the ophthalmic artery and thus is in the path of circulating hormones such as CCAP. Focally-applied CCAP directly excited some gastric mill neurons, including the gastric mill central pattern generator neurons LG and Int1, but it did not elicit a sustained gastric mill rhythm. At concentrations as low as 10−10M, however, CCAP did influence gastric mill rhythms elicited by coactivating the projection neurons MCN1 and CPN2 and by selectively stimulating MCN1. In both cases, CCAP slowed this rhythm by selectively prolonging the protraction phase, although its influence on the MCN1-elicited rhythm was limited to those with relatively brief cycle periods. Interestingly, CCAP also reduced the threshold MCN1 firing frequency for activating the gastric mill rhythm. Last, the gastric mill neurons that exhibited altered activity during these CCAP-influenced rhythms did not correspond completely to the set of CCAP-responsive neurons. These results highlight the ability of hormonal modulation to enhance the flexibility provided by the neuronal modulation of rhythmically active motor circuits.
Control of Movement Variability and the Regulation of Limb Impedance
Authors: Lametti Daniel R. , Houle Guillaume , Ostry David J.
Published: October 2007

Humans routinely make movements to targets that have different accuracy requirements in different directions. Examples extend from everyday occurrences such as grasping the handle of a coffee cup to the more refined instance of a surgeon positioning a scalpel. The attainment of accuracy in situations such as these might be related to the nervous system's capacity to regulate the limb's resistance to displacement, or impedance. To test this idea, subjects made movements from random starting locations to targets that had shape-dependent accuracy requirements. We used a robotic device to assess both limb impedance and patterns of movement variability just as the subject reached the target. We show that impedance increases in directions where required accuracy is high. Independent of target shape, patterns of limb stiffness are seen to predict spatial patterns of movement variability. The nervous system is thus seen to modulate limb impedance in entirely predictable environments to aid in the attainment of reaching accuracy.
Spatial and Functional Architecture of the Mammalian Brain Stem Respiratory Network: A Hierarchy of Three Oscillatory Mechanisms
Authors: Smith J. C. , Abdala A. P. L. , Koizumi H. , Rybak I. A. , Paton J. F. R.
Published: October 2007

Mammalian central pattern generators (CPGs) producing rhythmic movements exhibit extremely robust and flexible behavior. Network architectures that enable these features are not well understood. Here we studied organization of the brain stem respiratory CPG. By sequential rostral to caudal transections through the pontine-medullary respiratory network within an in situ perfused rat brain stem–spinal cord preparation, we showed that network dynamics reorganized and new rhythmogenic mechanisms emerged. The normal three-phase respiratory rhythm transformed to a two-phase and then to a one-phase rhythm as the network was reduced. Expression of the three-phase rhythm required the presence of the pons, generation of the two-phase rhythm depended on the integrity of Bötzinger and pre-Bötzinger complexes and interactions between them, and the one-phase rhythm was generated within the pre-Bötzinger complex. Transformation from the three-phase to a two-phase pattern also occurred in intact preparations when chloride-mediated synaptic inhibition was reduced. In contrast to the three-phase and two-phase rhythms, the one-phase rhythm was abolished by blockade of persistent sodium current ( INaP). A model of the respiratory network was developed to reproduce and explain these observations. The model incorporated interacting populations of respiratory neurons within spatially organized brain stem compartments. Our simulations reproduced the respiratory patterns recorded from intact and sequentially reduced preparations. Our results suggest that the three-phase and two-phase rhythms involve inhibitory network interactions, whereas the one-phase rhythm depends on INaP. We conclude that the respiratory network has rhythmogenic capabilities at multiple levels of network organization, allowing expression of motor patterns specific for various physiological and pathophysiological respiratory behaviors.
Expression of epidermal growth factor (EGF)/transforming growth factor-α by human lung cancer cells determines their response to EGF receptor tyrosine kinase inhibition in the lungs of mice
Authors: Wu Wenjuan , O'Reilly Michael S. , Langley Robert R. , Tsan Rachel Z. , Baker Cheryl H. , Bekele Neby , Tang Xi Ming , Onn Amir , Fidler Isaiah J. , Herbst Roy S.
Published: October 2007

Epidermal growth factor receptor (EGFR) has been extensively targeted in the treatment of non–small cell lung cancer, producing responses in a small number of patients. To study the role of ligand expression in mediating response to EGFR antagonism, we injected NCI-H441 [EGFR and EGF/transforming growth factor-α (TGF-α) positive] or PC14-PE6 (EGFR positive and EGF/TGF-α negative) human lung adenocarcinoma cells into the lungs of nude mice. We randomized the mice to receive treatment with the EGFR tyrosine kinase inhibitors gefitinib or AEE788 or vehicle. Treatment of mice bearing NCI-H441 but not PC14-PE6 lung tumors resulted in a significant reduction in primary tumor growth, pleural effusion, and lymph node metastasis. Immunohistochemical analyses revealed that NCI-H441 and PC14-PE6 cells expressed EGFR but that the expression of EGF/TGF-α was high in NCI-H441 cells and very low in PC14-PE6 cells. Consequently, EGFR was activated in both tumor and tumor-associated endothelial cells in the NCI-H441 tumors but not in the PC14-PE6 tumors. Antagonism of EGFR signaling by treatment of mice with AEE788 decreased proliferation and increased apoptosis of both tumor cells and tumor-associated endothelial cells in NCI-H441 tumors but not in PC14-PE6 tumors. However, after transfection of PC14-PE6 cells with TGF-α, lung tumors derived from the transfected cells expressed and activated EGFR in both tumor and tumor-associated endothelial cells and tumors responded to treatment with AEE788. Collectively, these results strongly suggest that the response of human lung cancers growing orthotopically in mice to the inhibition of EGFR signaling is determined by ligand (EGF/TGF-α) expression by tumor cells. Our findings provide an additional explanation for the susceptibility of lung cancers to treatment with EGFR tyrosine kinase inhibitors. [Mol Cancer Ther 2007;6(10):2652–63]
Synergistic cytotoxicity between tamoxifen and the plant toxin persin in human breast cancer cells is dependent on Bim expression and mediated by modulation of ceramide metabolism
Authors: Roberts Caroline G. , Gurisik Ebru , Biden Trevor J. , Sutherland Robert L. , Butt Alison J.
Published: October 2007

Phytochemicals have provided an abundant source of novel therapeutics for the treatment of human cancers. We have previously described a novel plant toxin, persin, derived from avocado leaves, which has unique in vivo actions in the mammary epithelium and Bim-dependent, cytotoxic effects in human breast cancer cells in vitro. Compounds structurally similar to persin, such as the polyunsaturated fatty acid, conjugated linoleic acid, can attenuate steroid hormone receptor signaling and modulate the response of breast cancer cells to antiestrogens. Here, we provide evidence that persin may have similar effects by showing its potent proapoptotic synergy with the antiestrogen 4-hydroxytamoxifen. However, although persin transcriptionally down-regulates estrogen receptor (ER) expression, unlike conjugated linoleic acid, it also shows efficacy in ER-negative breast cancer cells, both alone and in combination with 4-hydroxytamoxifen, whereas normal breast epithelial cells are unaffected, suggesting it may act via a distinct, ER-independent mechanism. These proapoptotic synergistic interactions are associated with increased de novo ceramide synthesis and are dependent on expression of the proapoptotic protein Bim. These data show that persin should be further investigated as a potential novel cancer therapeutic agent because it significantly enhances the sensitivity of breast cancer cells to the cytotoxic effects of tamoxifen, regardless of their ER status, while displaying apparent specificity for the malignant phenotype. [Mol Cancer Ther 2007;6(10):2777–85]
Subungual Penetration of Dibutyl Phthalate in Human Fingernails
Authors: Jackson E.M.
Published: October 2007

Dibutyl phthalate (DBP) has a wide variety of manufacturing applications and is used in both commercial and consumer products. Results of animal reproductive toxicity and teratogenicity animal studies have not been consistent in identifying DBP as a reproductive toxicant. Expert reviews for its use in consumer products have consistently concluded that it is not a reproductive risk to consumers. Results from a subungual penetration study of 100% fluid DBP applied to human fingernails showed levels of penetration at the limits of chemical detection. Even if DBP penetrated the human fingernail, its rapid metabolism by the human body would prevent its having any toxic reproductive effects. Furthermore, DBP functions as a plasticizer in consumer products such as cosmetic nail products (nail polish, basecoats, topcoats, nail hardeners), resulting in its becoming unavailable for subungual penetration seconds after application of the cosmetic nail product since it is then trapped in the rapidly forming coating.
Dry Powder Inhalers: Which Factors Determine the Frequency of Handling Errors?
Authors: Wieshammer Siegfried , Dreyhaupt Jens
Published: October 2007

Background: Dry powder inhalers are often used ineffectively, resulting in a poor level of disease control. Objectives: To determine how often essential mistakes are made in the use of Aerolizer®, Discus®, HandiHaler® and Turbuhaler® and to study the effects of age, severity of airflow obstruction and previous training in inhalational technique by medical personnel on the error rate. Methods: Two hundred and twenty-four newly referred outpatients (age 55.1 ± 20 years) were asked how they had been acquainted with the inhaler and to demonstrate their inhalational technique. Results: The inhaler-specific error rates were as follows: Aerolizer 9.1%, Discus 26.7%, HandiHaler 53.1% and Turbuhaler 34.9%. Compared to Aerolizer, the odds ratio of an ineffective inhalation was higher for HandiHaler (9.82, p < 0.01) and Turbuhaler (4.84, p < 0.05). The error rate increased with age and with the severity of airway obstruction (p < 0.01). When training had been given as opposed to no training, the odds ratio of ineffective inhalation was 0.22 (p < 0.01). If Turbuhaler is used, the estimated risks range from 9.8% in an 18-year-old patient with normal lung function and previous training to 83.2% in an 80-year-old patient with moderate or severe obstruction who had not received any training. Conclusions: Dry powder inhalers are useful in the management of younger patients with normal lung function or mild airway obstruction. In older patients with advanced chronic obstructive pulmonary disease, the risk of ineffective inhalation remains high despite training in inhalational technique. A metered-dose inhaler with a spacer might be a valuable treatment alternative in a substantial proportion of these patients.