bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression of Antimicrobial Peptides in Cecal Tonsils of Chickens Treated with Probiotics and Infected with
Salmonella enterica
Serovar Typhimurium
Authors: Akbari Mohammad Reza , Haghighi Hamid Reza , Chambers James R. , Brisbin Jennifer , Read Leah R. , Sharif Shayan
Published: October 2008

Several strategies currently exist for control of
Salmonella enterica
serovar Typhimurium colonization in the chicken intestine, among which the use of probiotics is of note. Little is known about the underlying mechanisms of probiotic-mediated reduction of
Salmonella
colonization. In this study, we asked whether the effect of probiotics is mediated by antimicrobial peptides, including avian beta-defensins (also called gallinacins) and cathelicidins. Four treatment groups were included in this study: a negative-control group, a probiotic-treated group, a
Salmonella
-infected group, and a probiotic-treated and
Salmonella
-infected group. On days 1, 3, and 5 postinfection (p.i.), the cecal tonsils were removed, and RNA was extracted and used for measurement of avian beta-defensin 1 (AvBD1), AvBD2, AvBD4, AvBD6, and cathelicidin gene expression by real-time PCR. The expressions of all avian beta-defensins and cathelicidin were detectable in all groups, irrespective of treatment and time point. Probiotic treatment and
Salmonella
infection did not affect the expression of any of the investigated genes on day 1 p.i. Furthermore, probiotic treatment had no significant effect on the expression of the genes at either 3 or 5 days p.i. However, the expression levels of all five genes were significantly increased (
P
< 0.05) in response to
Salmonella
infection at 3 and 5 days p.i. However, administration of probiotics eliminated the effect of
Salmonella
infection on the expression of antimicrobial genes. These findings indicate that the expression of antimicrobial peptides may be repressed by probiotics in combination with
Salmonella
infection or, alternatively, point to the possibility that, due to a reduction in
Salmonella
load in the intestine, these genes may not be induced.
Enhanced Gamma Interferon Responses of Mouse Spleen Cells following Immunotherapy for Tuberculosis Relapse
Authors: Gil Olga , Vilaplana Cristina , Guirado Evelyn , Díaz Jorge , Cáceres Neus , Singh Mahavir , Cardona Pere-Joan
Published: October 2008

Gamma interferon responses of spleen cells in mice were examined during postchemotherapy relapse of intraperitoneally induced latent tuberculous infection. The mycobacterial extract RUTI, which prevented the relapse, significantly enhanced the immune responses to secreted and structural recombinant mycobacterial antigens, suggesting that RUTI-mediated protection was mediated by activated T cells.
Validation of an Enzyme-Linked Immunosorbent Assay for Diagnosis of Human Trichinellosis
Authors: Gómez-Morales Maria Angeles , Ludovisi Alessandra , Amati Marco , Cherchi Simona , Pezzotti Patrizio , Pozio Edoardo
Published: October 2008

Trichinellosis is a zoonotic disease caused by the consumption of raw or semiraw meat from different animals harboring
Trichinella
larvae in their muscles. Since there are no pathognomonic signs, diagnosis can be difficult; for this reason, serology is important. The objective of this study was to validate an enzyme-linked immunosorbent assay (ELISA) using excretory/secretory antigens to detect anti-
Trichinella
immunoglobulin G antibodies in human sera. A total of 3,505 human serum samples were tested. A receiver-operator characteristic (ROC) curve analysis was performed. The accuracy of the test was determined by calculating the area under the curve, which was equal to 0.999, indicating high accuracy. The coefficient of variation calculated for data from four serum samples in eight working sessions was no higher than 5% for the positive sera or 14% for the negative sera. Moreover, the analysis of the differences in optical density between duplicates indicated a high repeatability for the ELISA. At the ROC optimized cutoff, the sensitivity and specificity of the test were, respectively, 99.2% and 90.6% (specificity of 95.6% when excluding the samples from multiparasitized persons from Tanzania). The validated ELISA showed good performance in terms of sensitivity, repeatability, and reproducibility, whereas the specificity was limited. These results suggest that this test is suitable for detecting anti-
Trichinella
antibodies in human sera for diagnostic purposes, whereas its use in epidemiological surveys could be questionable.
Assessment of Three Commercially Available Serologic Assays for Detection of Antibodies to
Mycobacterium tuberculosis
and Identification of Active Tuberculosis
Authors: Anderson Brian L. , Welch Ryan J. , Litwin Christine M.
Published: October 2008

Tuberculosis (TB) caused by
Mycobacterium tuberculosis
remains a major world disease, with approximately 9 million new cases each year. Identification and treatment of active disease are essential for TB control. Serology may offer increased detection of active disease in patients with a positive tuberculin skin test (TST) or QuantiFERON-TB (QFT-G). The InBios Active Tb
Detect
immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA), IBL
M. tuberculosis
IgG ELISA, and Anda Biologics TB ELISAs were evaluated for the ability to detect
M. tuberculosis
antibodies in patients with active disease. Agreement, sensitivity, and specificity for each ELISA were determined and compared to those for culture or amplified direct detection and
M. tuberculosis
low-risk control patients. The InBios Active Tb
Detect
ELISA had an agreement of 96.2%, a sensitivity of 83.3%, and a specificity of 98.9%. The IBL
M. tuberculosis
ELISA had an agreement of 84.0%, a sensitivity of 5.6%, and a specificity of 100.0%. The agreement, sensitivity, and specificity of the Anda Biologics TB ELISA were 74.2%, 83.3%, and 72.0%, respectively. The sensitivity for detecting
M. tuberculosis
antibodies in human immunodeficiency virus-associated TB was 50% for both the InBios Active Tb
Detect
ELISA and the Anda Biologics TB ELISA and 0% for the IBL
M. tuberculosis
ELISA. The positivity rates for InBios Active Tb
Detect
ELISA, IBL
M. tuberculosis
ELISA, and Anda Biologics TB ELISA in latently infected individuals positive by TST and/or QFT-G were 5.1%, 0.0%, and 30.8%, respectively. It can be concluded that the InBios Active Tb
Detect
IgG ELISA is superior to the other ELISAs in accurately detecting active TB.
Seroprevalence of Antibodies against Serogroup C Meningococci in England in the Postvaccination Era
Authors: Trotter Caroline L. , Borrow Ray , Findlow Jamie , Holland Ann , Frankland Sarah , Andrews Nick J. , Miller Elizabeth
Published: October 2008

The United Kingdom introduced meningococcal serogroup C conjugate (MCC) vaccines in 1999, resulting in substantial declines in serogroup C disease and carriage. Here, we measured the age-specific prevalence of serum bactericidal antibodies (SBA) to
Neisseria meningitidis
serogroup C and immunoglobulin G (IgG) concentrations to serogroups A, C, W-135, and Y in 2,673 serum samples collected in England between 2000 and 2004. We compared the seroprevalence of SBA titers of ≥8 in the postvaccination era with results from an earlier prevaccination study conducted using the same methods. We found that the percentages of individuals with protective SBA titers were higher in 2000 to 2004 in all of the age groups targeted for MCC vaccination. In the postvaccine era, the prevalence of protective titers was high (75%) in children who had recently been offered routine immunization, but this fell to 36% more than 18 months after scheduled immunization. In the cohorts targeted in the catch-up campaign, the percentage achieving SBA titers of ≥8 was higher in children offered the vaccine at ages 5 to 17 years than in children offered the vaccine at ages 1 to 4 years. The geometric mean concentration (GMC) IgG for serogroup C followed a similar pattern, corresponding to the age at and time since scheduled MCC vaccination. Serogroup-specific IgG GMCs for W-135 and Y were low and showed little variation by age. Serogroup A IgG GMCs were higher, possibly reflecting exposure to cross-reacting antigens. Although the incidence of serogroup C disease remains low due to persisting herd effects, population antibody levels to serogroup C meningococci should be monitored so that potentially susceptible age groups can be identified should herd immunity wane.
Orally Administered
Mycobacterium vaccae
Modulates Expression of Immunoregulatory Molecules in BALB/c Mice with Pulmonary Tuberculosis
Authors: Hernández-Pando Rogelio , Aguilar Diana , Orozco Hector , Cortez Yuriria , Brunet Laura Rosa , Rook Graham A.
Published: October 2008

The environmental saprophyte
Mycobacterium vaccae
induces a Th1 response and cytotoxic T cells that recognize
M. tuberculosis
, and by subcutaneous injection, it is therapeutic for pulmonary tuberculosis (TB) induced by high-dose challenge in BALB/c mice. However,
M. vaccae
also drives regulatory T cells that inhibit Th2 responses, and this is seen in allergy models, not only following subcutaneous injection but also after oral administration. An oral immunotherapeutic for TB would be clinically useful, so we investigated
M. vaccae
given orally by gavage at 28-day intervals in the TB model. We used two different protocols: starting the oral
M. vaccae
either 1 day before or 32 days after infection with
M. tuberculosis
. Throughout the infection (until 120 days), we monitored outcome (CFU), molecules involved in the development of immunoregulation (Foxp3, hemoxygenase 1, idoleamine 2,3-dioxygenase, and transforming growth factor β [TGF-β]), and indicators of cytokine balance (tumor necrosis factor, inducible nitric oxide synthase, interleukin-4 [IL-4], and IL-4δ2; an inhibitory splice variant of IL-4 associated with improved outcome in human TB). Oral
M. vaccae
had a significant effect on CFU and led to increased expression of Th1 markers and of IL-4δ2, while suppressing IL-4, Foxp3, and TGF-β. When administered 1 day before infection, oral
M. vaccae
induced a striking peak of expression of hemoxygenase 1. In conclusion, we show novel information about the expression in TB of murine IL-4δ2 and molecules involved in immunoregulation and show that these can be modulated by oral administration of a saprophytic mycobacterium. A clinical trial of oral
M. vaccae
in extensively drug-resistant TB might be justified.
Cigarette fires and burns in a population of New Zealand smokers
Authors: Smith J , Bullen C , Laugesen M , Glover M
Published: October 2008

Objectives:

To identify the proportion of adult cigarette smokers who have experienced cigarette-caused fires and burns and to describe smoker characteristics associated with increased risk of cigarette-caused fires and burns.

Methods:

Data on cigarette-caused fires and burns were collected in the baseline questionnaire of a randomised trial of a smoking cessation intervention conducted in New Zealand between March 2006 and May 2007. Participants were adult callers to a national smoking cessation counselling service. Lifetime prevalence estimates of cigarette-caused fires and burns were obtained and associations between smoker characteristics and risk of fires and burns examined using logistic regression.

Results:

Of 1097 participants in the trial at baseline, 75 (6.8%) reported past experience of ⩾1 fires caused by cigarettes (96 fires reported in total) and 658 (60.0%) described at least 1 cigarette-caused burn. In all, 57 participants (5.2%) reported burns that required medical attention. Male sex and Māori ethnicity (indigenous New Zealanders, who comprise 15% of the national population and among whom 42% of adults are smokers) were associated with increased risk of cigarette-caused fires. Male sex, younger age, younger age of smoking initiation, being unmarried, having a partner who smoked, having a higher education level and an annual income of $20 000 or more were associated with increased risk of cigarette burn injuries.

Conclusions:

The results indicate that cigarette-caused fires and burns are common among New Zealand smokers, are a source of inequality and therefore deserve greater attention from health advocates and policymakers.
How does increasingly plainer cigarette packaging influence adult smokers’ perceptions about brand image? An experimental study
Authors: Wakefield M A , Germain D , Durkin S J
Published: October 2008

Background:

Cigarette packaging is a key marketing strategy for promoting brand image. Plain packaging has been proposed to limit brand image, but tobacco companies would resist removal of branding design elements.

Method:

A 3 (brand types) × 4 (degree of plain packaging) between-subject experimental design was used, using an internet online method, to expose 813 adult Australian smokers to one randomly selected cigarette pack, after which respondents completed ratings of the pack.

Results:

Compared with current cigarette packs with full branding, cigarette packs that displayed progressively fewer branding design elements were perceived increasingly unfavourably in terms of smokers’ appraisals of the packs, the smokers who might smoke such packs, and the inferred experience of smoking a cigarette from these packs. For example, cardboard brown packs with the number of enclosed cigarettes displayed on the front of the pack and featuring only the brand name in small standard font at the bottom of the pack face were rated as significantly less attractive and popular than original branded packs. Smokers of these plain packs were rated as significantly less trendy/stylish, less sociable/outgoing and less mature than smokers of the original pack. Compared with original packs, smokers inferred that cigarettes from these plain packs would be less rich in tobacco, less satisfying and of lower quality tobacco.

Conclusion:

Plain packaging policies that remove most brand design elements are likely to be most successful in removing cigarette brand image associations.
Welcome to Serbia: feel free to smoke
Authors: O’Rourke M , Djukic J
Published: October 2008

This paper reviews tobacco use and control policies in Serbia. Tobacco contributes significantly to the burden of disease in Serbia, and it has one of the highest age standardised lung cancer incidence rates in Europe. Until recently there were few restrictions on smoking or tangible tobacco control policies. Serbia has, however, now embarked on a legislative and regulatory path towards greater tobacco control, but implementation and enforcement are still lax. Increased government interventions are needed to reduce the prevalence of smoking in the population.
A STUDY OF THE TRAP-DOOR PROBLEM IN A GRANULAR MASS
Authors: Ladanyi B. , Hoyaux B.
Published: October 2008

By using Schneebeli's rod material for simulating an ideal granular mass in plane strain, a series of model trap-door tests has been performed with the main objective of checking the validity of the classic bin theory when applied to this class of problems. In the tests, the pressure on the trap-door was measured, and the rod displacement trajectories were photographically recorded. A comparison of measured and calculated pressures acting on the trap-door both during its downward and upward movement, shows that the pressures can reasonably well be predicted by assuming the existence of two limiting vertical failure planes as in the classical bin theory, provided proper considerations are made concerning the ratio and the value of normal stresses acting along these planes.
EARTH PRESSURES ON OTTAWA•OUTFALL SEWER TUNNEL
Authors: Eden W. J. , Bozozuk M.
Published: October 2008

Measurements of surface settlement, changes in piezometric levels and earth pressures were undertaken on a 10-ft diameter tunnel that passed through extremely sensitive and moderately over-consolidated Leda clay in Ottawa. The instrumentation is described, and the results are given for a period of 5 years, from 1 year before construction until 4 years after completion. Results of the measurements are compared with those obtained from a tunnel of similar size in Detroit clay. Because of the method of construction there was opportunity to conduct measurements on both the rather flexible temporary steel lining and the more rigid permanent reinforced concrete lining.Very little surface settlement was measured owing to overconsolidation of the clay and the method of construction. The tunnel had considerable influence on the piezometric regime in the clay, acting as a drainage sink. Earth pressures were in accordance with previous measurements on tunnels, with vertical pressures approaching the full overburden pressure.
SULFATE ACTION ON CONCRETE IN WINNIPEG SOILS
Authors: Whitehead M. B. , Hurwitz L. E. , Kavanagh R. J. , Kuluk A. , Malus J. G. , McKnight R. W. , Nizalik R. G. , Priestley A.
Published: October 2008

The evolution of recommendations for the design of sulfate-resistant concrete is described. The need for such concrete in sulfate-bearing soils such as occur in the Winnipeg area is brought out by descriptions of the nature of the attack and of case histories.
EFFECTS OF ENVIRONMENT ON THE PERFORMANCE OF SHALLOW FOUNDATIONS
Authors: Hamilton J. J.
Published: October 2008

In addition to its structural role, the foundation usually separates the man-made indoor climate from the actual environment of the ground. The implications of this secondary role are sometimes not fully appreciated by the designer, with serious consequences. The structural success of shallow foundations on volume-changing clays may be governed by their net effects on the subsoil environment. The significance of changes from natural ground conditions by construction must be appreciated at least qualitatively by the foundation designer. Natural and artificial changes in the thermal, the moisture, and the stress environments of subsoils are discussed in this paper, with particular emphasis on the clay soils and the climatic conditions of Western Canada.
CONSTRUCTION METHOD AND STABILITY OF EMBANKMENTS ON MUSKEG
Authors: Raymond Gerald P
Published: October 2008

A method for the design and construction of relatively thick highway embankments across muskeg deposits is described. Using theoretical soil-mechanics concepts it is predicted that the long-term stability of the muskeg necessitates the use of berms despite the fact that an undrained strength analysis based on in situ vane tests may predict a reasonable factor of safety. Consideration of both theoretical soil-mechanics concepts and practical considerations suggested that the interaction between the soil and the berm dictates that berm construction should be from the outer limits towards the center and should be placed prior to the central fill.Several case records are presented to support the conclusions drawn and furthermore suggest that the width of the berms should be [Formula: see text] to 2 times the thickness of the muskeg (peat plus lake marl).
Placental HIFs as markers of cerebral hypoxic distress in fetal mice
Authors: Trollmann Regina , Strasser Katja , Keller Stephan , Antoniou Xenia , Grenacher Beat , Ogunshola Omolara O. , Dötsch Jörg , Rascher Wolfgang , Gassmann Max
Published: October 2008

Reduced oxygen supply during the pre- and perinatal period often leads to acquired neonatal brain damage. So far, there are no reliable markers available to assess the hypoxic cerebral damage and the resulting prognosis during the immediate postnatal period. Thus we aimed to determine whether the hypoxia-inducible transcription factors (HIF-1 and HIF-2) and/or their target genes in the placenta represent reliable indicators of hypoxic distress of the developing brain during systemic hypoxia at the end of gestation. To this end, pregnant mice were exposed to systemic hypoxia (inspired O2 fraction: 6%, 6 h) at gestational day 20. This hypoxic exposure significantly increased HIF-1α and HIF-2α protein levels in brain and placental tissue. Compared with normoxic controls, an increase of HIF-1α-immunoreactive neurons and HIF-2α-positive glial cells and vascular endothelial cells was observed in hypoxic cerebral cortex and hippocampus. In placenta, HIF-1α and HIF-2α were expressed in labyrinthine layer with increased staining intensity during hypoxia compared with normoxia. Significant upregulation of VEGF mRNA and protein in brain and placenta, as well as erythropoietin protein in placenta, indicated activity of the HIF system upon fetal hypoxia. Notably, hypoxia did not affect expression of the HIF target genes inducible nitric oxide synthase and GLUT-1. Taken together, at gestational day 20, systemic hypoxia led to upregulation of HIF-α in mouse brain that was temporally paralleled in placenta, implying that α-subunits of both HIF-1 and HIF-2 are indeed early markers of hypoxic distress in vivo. If our data reflect the situation in humans, analysis of the placenta will allow early identification of the hypoxic brain distress occurring near birth.
Microarray analysis of the transcriptome of the subfornical organ in the rat: regulation by fluid and food deprivation
Authors: Hindmarch Charles , Fry Mark , Yao Song T. , Smith Pauline M. , Murphy David , Ferguson Alastair V.
Published: October 2008

We have employed microarray technology using Affymetrix 230 2.0 genome chips to initially catalog the transcriptome of the subfornical organ (SFO) under control conditions and to also evaluate the changes (common and differential) in gene expression induced by the challenges of fluid and food deprivation. We have identified a total of 17,293 genes tagged as present in one of our three experimental conditions, transcripts, which were then used as the basis for further filtering and statistical analysis. In total, the expression of 46 genes was changed in the SFO following dehydration compared with control animals (22 upregulated and 24 downregulated), with the largest change being the greater than fivefold increase in brain-derived neurotrophic factor (BDNF) expression, while significant changes in the expression of the calcium-sensing (upregulated) and apelin (downregulated) receptors were also reported. In contrast, food deprivation caused greater than twofold changes in a total of 687 transcripts (222 upregulated and 465 downregulated), including significant reductions in vasopressin, oxytocin, promelanin concentrating hormone, cocaine amphetamine-related transcript (CART), and the endothelin type B receptor, as well as increases in the expression of the GABAB receptor. Of these regulated transcripts, we identified 37 that are commonly regulated by fasting and dehydration, nine that were uniquely regulated by dehydration, and 650 that are uniquely regulated by fasting. We also found five transcripts that were differentially regulated by fasting and dehydration including BDNF and CART. In these studies we have for the first time described the transcriptome of the rat SFO and have in addition identified genes, the expression of which is significantly modified by either water or food deprivation.
Selective estrogen receptor-α and estrogen receptor-β agonists rapidly decrease pulmonary artery vasoconstriction by a nitric oxide-dependent mechanism
Authors: Lahm Tim , Crisostomo Paul R. , Markel Troy A. , Wang Meijing , Wang Yue , Tan Jiangning , Meldrum Daniel R.
Published: October 2008

Both endogenous and exogenous estrogen decrease pulmonary artery (PA) vasoconstriction. Whether these effects are mediated via estrogen receptor (ER)-α or ER-β, and whether the contribution of ERs is stimulus-dependent, remains unknown. We hypothesized that administration of the selective ER-α agonist propylpyrazole triol (PPT) and/or the selective ER-β agonist diarylpropiolnitrile (DPN) rapidly decreases PA vasoconstriction induced by pharmacologic and hypoxic stimuli via a nitric oxide (NO)-dependent mechanism. PA rings ( n = 3–10/group) from adult male Sprague-Dawley rats were suspended in physiologic organ baths. Force displacement was measured. Vasoconstrictor responses to phenylephrine (10−8M − 10−5M) and hypoxia (Po2 35–45 mmHg) were determined. Endothelium-dependent and -independent vasorelaxation were measured by generating dose-response curves to acetylcholine (10−8M − 10−4M) and sodium nitroprusside (10−9M − 10−5M). PPT or DPN (10−9M − 5 × 10−5M) were added to the organ bath in the presence and absence of the NO-synthase inhibitor Nω-nitro-l-arginine methyl ester (l-NAME) (10−4M). Selective ER-α activation (PPT, 5 × 10−5M) rapidly (<20 min) decreased phenylephrine-induced vasoconstriction. This effect, as well as PPT's effects on endothelium-dependent vasorelaxation, were neutralized by l-NAME. In contrast, selective ER-β activation (DPN, 5 × 10−5M) rapidly decreased phase II of hypoxic pulmonary vasoconstriction (HPV). l-NAME eliminated this phenomenon. Lower PPT or DPN concentrations were less effective. We conclude that both ER-α and ER-β decrease PA vasoconstriction. The immediate onset of effect suggests a nongenomic mechanism. The contribution of specific ERs appears to be stimulus specific, with ER-α primarily modulating phenylephrine-induced vasoconstriction, and ER-β inhibiting HPV. NO inhibition eliminates these effects, suggesting a central role for NO in mediating the pulmonary vascular effects of both ER-α and ER-β.
Aquaporin 2 expression increased by glucagon in normal rat inner medullary collecting ducts
Authors: Yano Yuristella , Cesar Katia R. , Araujo Magali , Rodrigues Adilson C. , Andrade Lucia C. , Magaldi Antonio J.
Published: October 2008

It is well known that Glucagon (Gl) is released after a high protein diet and participates in water excretion by the kidney, principally after a protein meal. To study this effect in in vitro perfused inner medullary collecting ducts (IMCD), the osmotic water permeability (Pf; μm/s) at 37°C and pH 7.4 in normal rat IMCDs ( n = 36) perfused with Ringer/HCO3 was determined. Gl (10−7 M) in absence of Vasopressin (AVP) enhanced the Pf from 4.38 ± 1.40 to 11.16 ± 1.44 μm/s ( P < 0.01). Adding 10−8, 10−7, and 10−6 M Gl, the Pf responded in a dose-dependent manner. The protein kinase A inhibitor H8 blocked the Gl effect. The specific Gl inhibitor, des-His1-[Glu9] glucagon (10−7 M), blocked the Gl-stimulated Pf but not the AVP-stimulated Pf. There occurred a partial additional effect between Gl and AVP. The cAMP level was enhanced from the control 1.24 ± 0.39 to 59.70 ± 15.18 fm/mg prot after Gl 10−7 M in an IMCD cell suspension. The immunoblotting studies indicated an increase in AQP2 protein abundance of 27% (cont 100.0 ± 3.9 vs. Gl 127.53; P = 0.0035) in membrane fractions extracted from IMCD tubule suspension, incubated with 10−6 M Gl. Our data showed that 1) Gl increased water absorption in a dose-dependent manner; 2) the anti-Gl blocked the action of Gl but not the action of AVP; 3) Gl stimulated the cAMP generation; 4) Gl increased the AQP2 water channel protein expression, leading us to conclude that Gl controls water absorption by utilizing a Gl receptor, rather than a AVP receptor, increasing the AQP2 protein expression.
Pyramidal Neurons Switch From Integrators In Vitro to Resonators Under In Vivo-Like Conditions
Authors: Prescott Steven A. , Ratté Stéphanie , De Koninck Yves , Sejnowski Terrence J.
Published: October 2008

During wakefulness, pyramidal neurons in the intact brain are bombarded by synaptic input that causes tonic depolarization, increased membrane conductance (i.e., shunting), and noisy fluctuations in membrane potential; by comparison, pyramidal neurons in acute slices typically experience little background input. Such differences in operating conditions can compromise extrapolation of in vitro data to explain neuronal operation in vivo. For instance, pyramidal neurons have been identified as integrators (i.e., class 1 neurons according to Hodgkin's classification of intrinsic excitability) based on in vitro experiments but that classification is inconsistent with the ability of hippocampal pyramidal neurons to oscillate/resonate at theta frequency since intrinsic oscillatory behavior is limited to class 2 neurons. Using long depolarizing stimuli and dynamic clamp to reproduce in vivo-like conditions in slice experiments, we show that CA1 hippocampal pyramidal cells switch from integrators to resonators, i.e., from class 1 to class 2 excitability. The switch is explained by increased outward current contributed by the M-type potassium current IM, which shifts the balance of inward and outward currents active at perithreshold potentials and thereby converts the spike-initiating mechanism as predicted by dynamical analysis of our computational model. Perithreshold activation of IM is enhanced by the depolarizing shift in spike threshold caused by shunting and/or sodium channel inactivation secondary to tonic depolarization. Our conclusions were validated by multiple comparisons between simulation and experimental data. Thus even so-called “intrinsic” properties may differ qualitatively between in vitro and in vivo conditions.
Network analysis of temporal effects of intermittent and sustained hypoxia on rat lungs
Authors: Wu Wei , Dave Nilesh B. , Yu Guoying , Strollo Patrick J. , Kovkarova-Naumovski Elizabeta , Ryter Stefan W. , Reeves Stephen R. , Dayyat Ehab , Wang Yang , Choi Augustine M. K. , Gozal David , Kaminski Naftali
Published: October 2008

The molecular networks underlying the lung response to hypoxia are not fully understood. We employed systems biology approaches to study temporal effects of intermittent or sustained hypoxia on gene expression in rat lungs. We obtained gene expression profiles from rats exposed to intermittent or sustained hypoxia lasting 0–30 days and identified differentially expressed genes, their patterns, biological processes, and regulatory networks critical for lung response to intermittent or sustained hypoxia. We validated selected genes with quantitative real-time PCR. Intermittent and sustained hypoxia induced two distinct sets of genes in rat lungs that displayed different temporal expression patterns. Intermittent hypoxia induced genes mostly involved in ion transport and homeostasis, neurological processes, and steroid hormone receptor activity, while sustained hypoxia induced genes principally participating in immune responses. The intermittent hypoxia-activated network suggested a role for cross talk between estrogen receptor 1 (ESR1) and other key proteins in hypoxic responses. The sustained hypoxia-activated network was indicative of vascular remodeling and pulmonary hypertension. We confirmed the temporal expression changes of 12 genes (including the Esr1 gene and 4 ESR1 target genes) in intermittent hypoxia and 8 genes in sustained hypoxia with quantitative real-time PCR. Conclusions: intermittent and sustained hypoxia induced distinct gene expression patterns in rat lungs. The functional characteristics of genes activated by these two distinct perturbations suggest their roles in the downstream physiological effects of intermittent and sustained hypoxia. Our results demonstrate the discovery potential of applying systems biology approaches to the understanding of mechanisms underlying hypoxic lung response.
Regulation of Estrogenic Effects by Beclin 1 in Breast Cancer Cells
Authors: John Shali , Nayvelt Irina , Hsu Hui-Chen , Yang PingAr , Liu Wensheng , Das Gokul M. , Thomas Thresia , Thomas T.J.
Published: October 2008

Beclin 1 is an essential mediator of autophagy and a regulator of cell growth and cell death. We examined the effect of Beclin 1 overexpression on the action of estradiol (E2) and two antiestrogens, raloxifene and 4-hydroxytamoxifen, in estrogen receptor α (ERα)-positive MCF-7 breast cancer cells. [3H]-thymidine incorporation studies showed that Beclin 1–overexpressing cells (MCF-7.beclin) had a lower proliferative response to E2 compared with cells transfected with vector control (MCF-7.control). There was only a 35% increase in [3H]-thymidine incorporation, after 24 hours of E2 treatment of MCF-7.beclin cells compared with untreated cells, whereas this increase was 2-fold for MCF-7.control cells. E2-induced changes in the expression of early-response genes were examined by real-time quantitiative PCR. There were significant differences in the pattern of expression of E2-induced genes c-myc, c-fos, Erg-1, and Nur77 between MCF-7.beclin and MCF-7.control cells two hours after treatment. Although E2-induced growth of MCF-7.control cells was completely inhibited by 500 nmol/L raloxifene or 500 nmol/L 4-hydroxytamoxifen, these concentrations of antiestrogens had no significant effect on the growth of MCF-7.beclin cells. Confocal microscopic and coimmunoprecipitation studies showed evidence for colocalization and association of Beclin 1 and ERα. In addition, E2 caused a decrease in Akt phosphorylation in MCF-7.beclin cells, compared with a 3-fold increase in MCF-7 cells, five minutes after treatment. These results indicate that Beclin 1 can down-regulate estrogenic signaling and growth response, and contribute to the development of antiestrogen resistance. This observation might be useful to define and overcome antiestrogen resistance of breast cancer. [Cancer Res 2008;68(19):7855–63]
The Polycomb Gene Product BMI1 Contributes to the Maintenance of Tumor-Initiating Side Population Cells in Hepatocellular Carcinoma
Authors: Chiba Tetsuhiro , Miyagi Satoru , Saraya Atsunori , Aoki Ryutaro , Seki Atsuyoshi , Morita Yohei , Yonemitsu Yutaka , Yokosuka Osamu , Taniguchi Hideki , Nakauchi Hiromitsu , Iwama Atsushi
Published: October 2008

Side population (SP) cell analysis and sorting have been successfully applied to hepatocellular carcinoma (HCC) cell lines to identify a minor cell population with cancer stem cell properties. However, the molecular mechanisms operating in SP cells remain unclear. The polycomb gene product BMI1 plays a central role in the self-renewal of somatic stem cells in a variety of tissues and organs and seems to be implicated in tumor development. In this study, we determined the critical role of BMI1 in the maintenance of cancer stem cells with the SP phenotype in HCC cell lines. BMI1 was preferentially expressed in SP cells in Huh7 and PLC/PRF/5 HCC cells compared with the corresponding non-SP cells. Lentiviral knockdown of BMI1 considerably decreased the number of SP cells in both Huh7 and PLC/PRF/5 cells. Long-term culture of purified SP cells resulted in a drastic reduction in the SP subpopulation upon the BMI1 knockdown, indicating that BMI1 is required for the self-renewal of SP cells in culture. More importantly, the BMI1 knockdown abolished the tumor-initiating ability of SP cells in nonobese diabetic/severe combined immunodeficiency mice. Derepression of the INK4A and ARF genes that are major targets for BMI1 was not necessarily associated with impaired self-renewal of SP cells caused by BMI1 knockdown. In conclusion, our findings define an important role for BMI1 in the maintenance of tumor-initiating SP cells in HCC. BMI1 might be a novel therapeutic target for the eradication of cancer stem cells in HCC. [Cancer Res 2008;68(19):7742–9]
A Bicistronic CYCLIN D1-TROP2 mRNA Chimera Demonstrates a Novel Oncogenic Mechanism in Human Cancer
Authors: Guerra Emanuela , Trerotola Marco , Dell' Arciprete Roberta , Bonasera Veronica , Palombo Barbara , El-Sewedy Tarek , Ciccimarra Tommaso , Crescenzi Carlo , Lorenzini Franco , Rossi Cosmo , Vacca Giovanna , Lattanzio Rossano , Piantelli Mauro , Alberti Saverio
Published: October 2008

A chimeric CYCLIN D1-TROP2 mRNA was isolated from human ovarian and mammary cancer cells. The CYCLIN D1-TROP2 mRNA was shown to be a potent oncogene as it transforms naïve, primary cells in vitro and induces aggressive tumor growth in vivo in cooperation with activated RAS. Silencing of the chimeric mRNA inhibits the growth of breast cancer cells. The CYCLIN D1-TROP2 mRNA was expressed by a large fraction of the human gastrointestinal, ovarian, and endometrial tumors analyzed. It is most frequently detected in intestinal cell aneuploid cancers and it is coexpressed with activated RAS oncogenes, consistent with a cooperative transforming activity in human cancers. The chimeric mRNA is a bicistronic transcript of post transcriptional origin that independently translates the Cyclin D1 and Trop-2 proteins. This is a novel mechanism of CYCLIN D1 activation that achieves the truncation of the CYCLIN D1 mRNA in the absence of chromosomal rearrangements. This leads to a higher CYCLIN D1 mRNA stability, with inappropriate expression during the cell cycle. The stabilized CYCLIN D1 mRNA cooperates with TROP2 in stimulating the growth of the expressing cells. These findings show a novel epigenetic, oncogenic mechanism, which seems to be widespread in human cancers. [Cancer Res 2008;68(19):8113–21]
Senescence-Associated Exosome Release from Human Prostate Cancer Cells
Authors: Lehmann Brian D. , Paine Matthew S. , Brooks Adam M. , McCubrey James A. , Renegar Randall H. , Wang Rong , Terrian David M.
Published: October 2008

Males of advanced age represent a rapidly growing population at risk for prostate cancer. In the contemporary setting of earlier detection, a majority of prostate carcinomas are still clinically localized and often treated using radiation therapy. Our recent studies have shown that premature cellular senescence, rather than apoptosis, accounts for most of the clonogenic death induced by clinically relevant doses of irradiation in prostate cancer cells. We show here that this treatment-induced senescence was associated with a significantly increased release of exosome-like microvesicles. In premature senescence, this novel secretory phenotype was dependent on the activation of p53. In addition, the release of exosome-like microvesicles also increased during proliferative senescence in normal human diploid fibroblasts. These data support the hypothesis that senescence, initiated either by telomere attrition (e.g., aging) or DNA damage (e.g., radiotherapy), may induce a p53-dependent increase in the biogenesis of exosome-like vesicles. Ultrastructural analysis and RNA interference–mediated knockdown of Tsg101 provided significant evidence that the additional exosomes released by prematurely senescent prostate cancer cells were principally derived from multivesicular endosomes. Moreover, these exosomes were enriched in B7-H3 protein, a recently identified diagnostic marker for prostate cancer, and an abundance of what has recently been termed “exosomal shuttle RNA.” Our findings are consistent with the proposal that exosomes can transfer cargos, with both immunoregulatory potential and genetic information, between cells through a novel mechanism that may be recruited to increase exosome release during accelerated and replicative cellular senescence. [Cancer Res 2008;68(19):7864–71]
SCL/TAL1 Interrupting Locus Derepresses GLI1 from the Negative Control of Suppressor-of-Fused in Pancreatic Cancer Cell
Authors: Kasai Kenji , Inaguma Shingo , Yoneyama Akiko , Yoshikawa Kazuhiro , Ikeda Hiroshi
Published: October 2008

As a physically binding protein of GLI1 transcription factor, Suppressor-of-Fused (SUFU) has been placed in the center of negative regulation of Hedgehog (Hh) signaling. SUFU tethers GLI1 in cytoplasm, and in some circumstances, it moves into the nucleus in association with GLI1, leading to the suppression of GLI1 target gene expression by recruiting a corepressor complex. The activated transcriptional function of GLI1 is important for cellular proliferation in a variety of human cancers. However, it has not been revealed how GLI1 is derepressed from SUFU-mediated suppression. Here, we show SCL/TAL1 interrupting locus (SIL) product, a cytoplasmic protein overexpressed in pancreatic ductal adenocarcinoma (PDA), is responsible for the derepression of GLI1. We found SIL associated with the carboxyl terminus of SUFU, one of two distinct GLI1-binding domains, and this association was responsible for cytoplasmic tethering of SUFU. Overexpressed SIL attenuated SUFU-mediated cytoplasmic tethering and target gene suppression of GLI1. Knockdown of SIL in PDA cells conversely induced the nuclear accumulation of SUFU in association with GLI1 and the transcriptional suppression of GLI1 target genes. Importantly, we also showed that oncogenic K-RAS, and not Sonic hedgehog, enhanced the SIL association with the amino-terminus of SUFU, the other GLI1-binding domain that led to further increase of nuclear translocation of GLI1. These results uncover the role of SIL in derepressing GLI1 from the negative control of SUFU, which is a crucial step for activating Hh signaling in cancer cells. [Cancer Res 2008;68(19):7723–9]
Prostaglandin E2 Regulates Tumor Angiogenesis in Prostate Cancer
Authors: Jain Shalini , Chakraborty Goutam , Raja Remya , Kale Smita , Kundu Gopal C.
Published: October 2008

In cancer management, the cyclooxygenase (COX)–targeted approach has shown great promise in anticancer therapeutics. However, the use of COX-2 inhibitors has side effects and health hazards; thus, targeting its major metabolite prostaglandin E2 (PGE2)–mediated signaling pathway might be a rational approach for the next generation of cancer management. Recent studies on several in vitro and in vivo models have revealed that elevated expression of COX-2 correlates with prostate tumor growth and angiogenesis. In this study, we have shown the in-depth molecular mechanism and the PGE2 activation of the epidermal growth factor receptor and β3 integrin through E prostanoid 2 (EP2)–mediated and EP4-mediated pathways, which lead to activator protein-1 (AP-1) activation. Moreover, PGE2 also induces activating transcription factor-4 (ATF-4) activation and stimulates cross-talk between ATF-4 and AP-1, which is unidirectional toward AP-1, which leads to the increased expressions of urokinase-type plasminogen activator and vascular endothelial growth factor and, eventually, regulates prostate tumor cell motility. In vivo Matrigel angiogenesis assay data revealed that PGE2 induces angiogenesis through EP2 and EP4. Human prostate cancer specimen analysis also supported our in vitro and in vivo studies. Our data suggest that targeting PGE2 signaling pathway (i.e., blocking EP2 and EP4 receptors) might be a rational therapeutic approach for overcoming the side effects of COX-2 inhibitors and that this might be a novel strategy for the next generation of prostate cancer management. [Cancer Res 2008;68(19):7750–9]
Glucagon-like Peptide-2 Does Not Modify the Growth or Survival of Murine or Human Intestinal Tumor Cells
Authors: Koehler Jacqueline A. , Harper Will , Barnard Maja , Yusta Bernardo , Drucker Daniel J.
Published: October 2008

Glucagon-like peptide-2 (GLP-2) secreted from enteroendocrine cells exerts proabsorptive, regenerative, and cytoprotective actions in the normal and injured gut epithelium. Hence, sustained GLP-2 receptor (GLP-2R) activation represents a strategy under investigation for the prevention and treatment of chemotherapy-induced mucositis. Nevertheless, the consequences of increased GLP-2R signaling for the growth and survival of intestinal tumor cells remain poorly understood. We studied the proliferative and cytoprotective actions of GLP-2 in human colon cancer cells stably transfected with the GLP-2R and in nude mice harboring GLP-2R+ human colon cancer cells. The importance of the GLP-2R for tumor growth was also examined in ApcMin/+ mice chronically treated with exogenous GLP-2 and in ApcMin/+:Glp2r−/− mice. GLP-2 increased cyclic AMP accumulation and produced cell-specific activation of growth and survival pathways in DLD-1, SW480, and HT29 cells. However, GLP-2 did not stimulate cell growth or attenuate cycloheximide-, LY294002-, indomethacin-, or chemotherapy-induced cytotoxicity in vitro. Moreover, chronic GLP-2 administration had no effect on the growth of human colon cancer cell xenografts in nude mice in vivo. Daily GLP-2 treatment for 7 weeks increased growth of normal gut mucosa but did not increase the number or size of polyps in ApcMin/+ mice, and genetic disruption of the Glp2r gene in ApcMin/+ mice did not modify polyp size or number. Taken together, although GLP-2R activation engages signaling pathways promoting cell proliferation and cytoprotection in the normal gut epithelium, sustained direct or indirect modulation of GLP-2R signaling does not modify intestinal tumor cell growth or survival. [Cancer Res 2008;68(19):7897–904]
Development of Cellular Immune Responses against PAX5, a Novel Target for Cancer Immunotherapy
Authors: Yan Mengyong , Himoudi Nourredine , Pule Martin , Sebire Neil , Poon Edmund , Blair Allison , Williams Owen , Anderson John
Published: October 2008

PAX5 is a member of the PAX family of developmental transcription factors with an important role in B-cell development. Its expression in normal adult tissue is limited to the hemopoietic system, but it is aberrantly expressed in a number of solid cancers and leukemias where it functions as an oncogene. We therefore hypothesized that anti-PAX5 immune responses could be used to target a number of malignancies without significant toxicity. We screened PAX5 peptides for the ability to bind HLA-A2 and identified a novel sequence, TLPGYPPHV (referred to as TLP). CTL lines against TLP were generated from peripheral blood of five normal HLA-A2–positive blood donors and showed specific HLA-A2–restricted killing against PAX5-expressing target cells. We generated high-avidity CTL clones from these lines capable of killing cells pulsed with <1 nmol/L of TLP and killing a range of PAX5-expressing malignant cell lines. I.v. injection of an anti-PAX5 CTL clone into immunodeficient mice bearing s.c. human tumors resulted in specific growth inhibition of PAX5-expressing tumors. This knowledge can be used for the therapeutic generation of CTL lines or the cloning of high-avidity T-cell receptor genes for use in adoptive immunotherapy. [Cancer Res 2008;68(19):8058–65]
Cancer Immunotherapy Targeting the High Molecular Weight Melanoma-Associated Antigen Protein Results in a Broad Antitumor Response and Reduction of Pericytes in the Tumor Vasculature
Authors: Maciag Paulo Cesar , Seavey Matthew M. , Pan Zhen-Kun , Ferrone Soldano , Paterson Yvonne
Published: October 2008

The high molecular weight melanoma-associated antigen (HMW-MAA), also known as melanoma chondroitin sulfate proteoglycan, has been used as a target for the immunotherapy of melanoma. This antigen is expressed on the cell surface and has a restricted distribution in normal tissues. Besides its expression in a broad range of transformed cells, this antigen is also found in pericytes, which are important for tumor angiogenesis. We generated a recombinant Listeria monocytogenes (Lm-LLO-HMW-MAA-C) that expresses and secretes a fragment of HMW-MAA (residues 2,160–2,258) fused to the first 441 residues of the listeriolysin O (LLO) protein. Immunization with Lm-LLO-HMW-MAA-C was able to impede the tumor growth of early established B16F10-HMW-MAA tumors in mice and both CD4+ and CD8+ T cells were required for therapeutic efficacy. Immune responses to a known HLA-A2 epitope present in the HMW-MAA2160-2258 fragment was detected in the HLA-A2/Kb transgenic mice immunized with Lm-LLO-HMW-MAA-C. Surprisingly, this vaccine also significantly impaired the in vivo growth of other tumorigenic cell lines, such as melanoma, renal carcinoma, and breast tumors, which were not engineered to express HMW-MAA. One hypothesis is that the vaccine could be targeting pericytes, which are important for tumor angiogenesis. In a breast tumor model, immunization with Lm-LLO-HMW-MAA-C caused CD8+ T-cell infiltration in the tumor stroma and a significant decrease in the number of pericytes in the tumor blood vessels. In conclusion, a Lm-based vaccine against HMW-MAA can trigger cell-mediated immune responses to this antigen that can target not only tumor cells but also pericytes in the tumor vasculature. [Cancer Res 2008;68(19):8066–75]