bluuebunny/crossref_metadata_2025 huggingface dataset:
Sources of cortical rhythms in adults during physiological aging: A multicentric EEG study
Authors: Babiloni Claudio , Binetti Giuliano , Cassarino Andrea , Dal Forno Gloria , Del Percio Claudio , Ferreri Florinda , Ferri Raffaele , Frisoni Giovanni , Galderisi Silvana , Hirata Koichi , Lanuzza Bartolo , Miniussi Carlo , Mucci Armida , Nobili Flavio , Rodriguez Guido , Luca Romani Gian , Rossini Paolo M.
Published: August 2005

This electroencephalographic (EEG) study tested whether cortical EEG rhythms (especially delta and alpha) show a progressive increasing or decreasing trend across physiological aging. To this aim, we analyzed the type of correlation (linear and nonlinear) between cortical EEG rhythms and age. Resting eyes‐closed EEG data were recorded in 108 young (Nyoung; age range: 18–50 years, mean age 27.3 ± 7.3 SD) and 107 elderly (Nold; age range: 51–85 years, mean age 67.3 ± 9.2 SD) subjects. The EEG rhythms of interest were delta (2–4 Hz), theta (4–8 Hz), alpha 1 (8–10.5 Hz), alpha 2 (10.5–13 Hz), beta 1 (13–20 Hz), and beta 2 (20–30 Hz). EEG cortical sources were estimated by low‐resolution brain electromagnetic tomography (LORETA). Statistical results showed that delta sources in the occipital area had significantly less magnitude in Nold compared to Nyoung subjects. Similarly, alpha 1 and alpha 2 sources in the parietal, occipital, temporal, and limbic areas had significantly less magnitude in Nold compared to Nyoung subjects. These nine EEG sources were given as input for evaluating the type (linear, exponential, logarithmic, and power) of correlation with age. When subjects were considered as a single group there was a significant linear correlation of age with the magnitude of delta sources in the occipital area and of alpha 1 sources in occipital and limbic areas. The same was true for alpha 2 sources in the parietal, occipital, temporal, and limbic areas. In general, the EEG sources showing significant linear correlation with age also supported a nonlinear correlation with age. These results suggest that the occipital delta and posterior cortical alpha rhythms decrease in magnitude during physiological aging with both linear and nonlinear trends. In conclusion, this new methodological approach holds promise for the prediction of dementia in mild cognitive impairment by regional source rather than surface EEG data and by both linear and nonlinear predictors. Hum Brain Mapp, 2005. © 2005 Wiley‐Liss, Inc.
Motion standstill leads to activation of inferior parietal lobe
Authors: Federspiel Andrea , Volpe Umberto , Horn Helge , Dierks Thomas , Franck Anders , Vannini Patrizia , Wahlund Lars‐Olof , Galderisi Silvana , Maj Mario
Published: August 2005

Previous studies on motion perception revealed motion‐processing brain areas sensitive to changes in luminance and texture (low‐level) and changes in salience (high‐level). The present functional magnetic resonance imaging (fMRI) study focused on motion standstill. This phenomenon, occurring at fast presentation frequencies of visual moving objects that are perceived as static, has not been previously explored by neuroimaging techniques. Thirteen subjects were investigated while perceiving apparent motion at 4 Hz, at 30 Hz (motion standstill), isoluminant static and flickering stimuli, fixation cross, and blank screen, presented randomly and balanced for rapid event‐related fMRI design. Blood oxygenation level‐dependent (BOLD) signal in the occipito‐temporal brain region MT/V5 increased during apparent motion perception. Here we could demonstrate that brain areas like the posterior part of the right inferior parietal lobule (IPL) demonstrated higher BOLD‐signal during motion standstill. These findings suggest that the activation of higher‐order motion areas is elicited by apparent motion at high presentation rates (motion standstill). We interpret this observation as a manifestation of an orienting reaction in IPL towards stimulus motion that might be detected but not resolved by other motion‐processing areas (i.e., MT/V5). Hum Brain Mapp, 2005. © 2005 Wiley‐Liss, Inc.
Tissue Transglutaminase (TG2) Acting as G Protein Protects Hepatocytes Against Fas-Mediated Cell Death in Mice *
Authors: Sarang Zsolt , Molnár Péter , Németh Tamás , Gomba Szabolcs , Kardon Tamás , Melino Gerry , Cotecchia Susanna , Fésüs László , Szondy Zsuzsa
Published: August 2005

Tissue transglutaminase (TG2) is a protein cross-linking enzyme known to be expressed by hepatocytes and to be induced during the

in vivo

hepatic apoptosis program. TG2 is also a G protein that mediates intracellular signaling by the alpha-1b-adrenergic receptor (AR) in liver cells. Fas/Fas ligand interaction plays a crucial role in various liver diseases, and administration of agonistic anti-Fas antibodies to mice causes both disseminated endothelial cell apoptosis and fulminant hepatic failure. Here we report that an intraperitoneal dose of anti-Fas antibodies, which is sublethal for wild-type mice, kills all the TG2 knock-out mice within 20 hours. Although TG2−/− thymocytes exposed to anti-Fas antibodies die at the same rate as wild-type mice, TG2−/− hepatocytes show increased sensitivity toward anti-Fas treatment both

in vivo

and

in vitro,

with no change in their cell surface expression of Fas, levels of FLIPL (FLICE-inhibitory protein), or the rate of I-κBα degradation, but a decrease in the Bcl-xL expression. We provide evidence that this is the consequence of the impaired AR signaling that normally regulates the levels of Bcl-xL in the liver.
In conclusion
, our data suggest the involvement of adrenergic signaling pathways in the hepatic regeneration program, in which Fas ligand-induced hepatocyte proliferation with a simultaneous inhibition of the Fas-death pathway plays a determinant role. (Hepatology 2005.)
The Third Gas: H2S Regulates Perfusion Pressure in Both the Isolated and Perfused Normal Rat Liver and in Cirrhosis *
Authors: Fiorucci Stefano , Antonelli Elisabetta , Mencarelli Andrea , Orlandi Stefano , Renga Barbara , Rizzo Giovanni , Distrutti Eleonora , Shah Vijay , Morelli Antonio
Published: August 2005

The regulation of sinusoidal resistance is dependent on the contraction of hepatic stellate cells (HSC) around sinusoidal endothelial cell (SEC) through paracrine cross-talk of vasoconstrictor and vasodilator agents. Hydrogen sulfide (H2S), a recently discovered gas neurotransmitter, is a putative vasodilator whose role in hepatic vascular regulation and portal hypertension is unexplored. Four-week bile duct–ligated (BDL) rats with cirrhosis and control rats were treated daily with NaHS (56 μmol/kg) for 5 days. Isolated livers were perfused first with NaHS for 20 minutes and then with norepinephrine (NE) and the intrahepatic resistance studied. In normal rats and animals with cirrhosis, administration of NE resulted in a dose-dependent increase of portal pressure. This effect was attenuated by H2S treatment (

P

< .05). The H2S-induced relaxation of hepatic microcirculation was attenuated by glibenclamide, an adenosine triphosphate (ATP)-sensitive K+ channel inhibitor. l-Cysteine, a substrate of cystathionine-gamma-lyase (CSE), decreased vasoconstriction in normal rat livers (

P

< .05) but failed to do so in livers with cirrhosis. BDL resulted in a downregulation of CSE mRNA/protein levels and activity (

P

< .05). Our

in vitro

data demonstrate that CSE is expressed in hepatocytes, HSCs, but not in sinusoidal endothelial cells (SEC). HSC activation downregulates CSE mRNA expression, resulting in a defective production of H2S and abrogation of relaxation induced by l-cysteine.
In conclusion
, CSE-derived H2S is involved in the maintenance of portal venous pressure. The reduction of CSE expression in the liver with cirrhosis contributes to the development of increased intrahepatic resistance and portal hypertension. (Hepatology 2005.)
Role of Adiponectin in the Protective Action of Dietary Saturated Fat Against Alcoholic Fatty Liver in Mice * #
Authors: You Min , Considine Robert V. , Leone Teresa C. , Kelly Daniel P. , Crabb David W.
Published: August 2005

The protective effect of dietary saturated fatty acids against the development of alcoholic liver disease has long been known, but the underlying mechanism is not completely understood. We examined the involvement of the adipocyte hormone adiponectin. Circulating adiponectin levels were significantly elevated by chronic ethanol administration to mice consuming a diet high in saturated fat. The increase in circulating adiponectin was associated with the activation a set of hepatic signaling pathways mediated through AMP-activated protein kinase, PPAR-α, and PPAR-γ coactivator α, which in turn led to markedly increased rates of fatty acid oxidation, prevention of hepatic steatosis, and alleviation of liver enzyme changes. Furthermore, treatment of rat 3T3-L1 adipocytes with saturated fatty acids (palmitic or stearic acids) in the presence of ethanol increased secretion of adiponectin and enhanced activity of a mouse adiponectin promoter.
In conclusion
, the protective action of saturated fat against the development of alcoholic fatty liver in mice is partially mediated through induction of adiponectin. The present findings suggest a novel paradigm for dietary fatty acids in the pathogenesis of alcoholic liver disease and provide a promising therapeutic strategy—nutritional modulation of adiponectin—in treating human alcoholic fatty liver disease. (Hepatology 2005.)
Significance and Therapeutic Potential of Prostaglandin E2 Receptor in Hepatic Ischemia/Reperfusion Injury in Mice *
Authors: Kuzumoto Yukiyasu , Sho Masayuki , Ikeda Naoya , Hamada Kaoru , Mizuno Takashi , Akashi Satoru , Tsurui Yoshikazu , Kashizuka Hisanori , Nomi Takeo , Kubo Atsushi , Kanehiro Hiromichi , Nakajima Yoshiyuki
Published: August 2005

Prostaglandin E2 (PGE2) mediates a variety of innate and adaptive immunity through four distinct receptors: EP1-EP4. It has been suggested that each EP plays a unique and pivotal role in various disease conditions. We investigated the pathophysiological role of EP receptors in hepatic ischemia/reperfusion (I/R) injury. In this study, a 70% hepatic ischemic model was used in male C57BL/6 mice. Selective EP agonists were used to clarify the function of each PGE2 receptor in I/R injury. Although all four receptors were expressed in the naïve liver, EP4 expression was significantly upregulated after hepatic I/R. Although EP1, 2, or 3 agonists did not show any protective effect on liver function, the EP4 agonist significantly inhibited hepatic I/R injury as determined by serological and histological analyses. Furthermore, the EP4 agonist downregulated the local expressions of several proinflammatory cytokines, chemokines, and adhesion molecules in the early phase of reperfusion. In contrast, it augmented the local expression of an anti-inflammatory cytokine, interleukin 10. Additionally, the neutrophil accumulation was also inhibited by EP4 agonist treatment. Finally, to confirm the therapeutic efficacy of the EP4 agonist in hepatic I/R injury, the nonischemic shunt liver was removed after 120 minutes of ischemia, resulting in the death of 86% of control mice within 48 hours. In sharp contrast, 80% of mice treated with the EP4 agonist survived.
In conclusion
, the PGE2-EP4 signaling pathway has an inhibitory role in hepatic I/R injury. An EP4 agonist effectively protects against ischemic injury. (Hepatology 2005.)
A Randomized Unblinded Pilot Study Comparing Albumin Versus Hydroxyethyl Starch in Spontaneous Bacterial Peritonitis * #
Authors: Fernández Javier , Monteagudo Joan , Bargallo Xavier , Jiménez Wladimiro , Bosch Jaume , Arroyo Vicente , Navasa Miguel
Published: August 2005

The administration of albumin improves circulatory function, prevents hepatorenal syndrome, and reduces hospital mortality in patients with cirrhosis and spontaneous bacterial peritonitis. This randomized unblinded pilot study compared the effect of albumin (10 patients) and the synthetic plasma expander hydroxyethyl starch 200/0.5 (10 patients) on the systemic hemodynamics of patients with spontaneous bacterial peritonitis. Baseline measurements were performed within 12 hours after diagnosis of infection. Patients then received 2 doses of the volume expander (1.5 g/kg body weight after baseline measurements and 1 g/kg body weight on day 3). Measurements were repeated after infection resolution. Treatment with albumin was associated with a significant increase in arterial pressure and a suppression of plasma renin activity, indicating an improvement in circulatory function. This occurred in the setting of a significant expansion of central blood volume (increase in cardiopulmonary pressures and atrial natriuretic factor) and an increase in systolic volume and systemic vascular resistance. In contrast, no significant changes were observed in these parameters in patients treated with hydroxyethyl starch. Von Willebrand–related antigen plasma levels significantly decreased in patients treated with albumin but not in those treated with hydroxyethyl starch. Serum nitrates and nitrites increased in patients treated with hydroxyethyl starch but not in those treated with albumin. These data suggest an effect of albumin on endothelial function.
In conclusion
, albumin but not hydroxyethyl starch improves systemic hemodynamics in patients with spontaneous bacterial peritonitis. This effect is due not only to volume expansion but also to an action on the peripheral arterial circulation. (Hepatology 2005.)
Quality of Life in Refractory Ascites: Transjugular Intrahepatic Portal-Systemic Shunting Versus Medical Therapy *
Authors: Campbell Mical S. , Brensinger Colleen M. , Sanyal Arun J. , Gennings Chris , Wong Florence , Kowdley Kris V. , McCashland Timothy , Reddy Rajender K.
Published: August 2005

Uncontrolled studies suggest that transjugular intrahepatic portal-systemic shunting (TIPS) may improve quality of life in patients with refractory ascites. We hypothesized that any improvement of quality of life in patients with TIPS would be matched in controls due to the competing effects of improved ascites and worsened hepatic encephalopathy. Thus, an analysis of quality of life was performed using original data from the North American Study for the Treatment of Refractory Ascites, a multicenter trial of 109 patients randomized to TIPS or repeated large volume paracentesis (LVP) for refractory ascites. Short form 36 (SF-36) surveys were completed at baseline and at 6- and 12-month follow-up. Variables analyzed were: randomization group, number of LVP performed, cumulative volume from LVP, shortness of breath, abdominal distention, abdominal pain, diuretic usage, confusion, hospitalizations, and emergency room visits. Outcomes were changes in physical component scale (PCS) and mental component scale (MCS) of SF-36 results. We constructed multivariable, mixed effects models, including randomization group and baseline MCS and PCS. Changes in PCS and MCS from baseline were similar between the two randomization groups. In multivariate analysis, PCS improvement was associated with lack of confusion, improved ascites, and lack of hospitalizations, but not directly with randomization group. Improvement in MCS was associated with randomization to TIPS and lack of confusion.
In conclusion
, patients with refractory ascites randomized to TIPS or repeated LVP had similar changes in quality of life. Competing effects of hepatic encephalopathy, requirement for repeated LVP, and need for hospitalizations explain similar changes in quality of life between the two groups. (Hepatology 2005.)
Analysis of ISG Expression in Chronic Hepatitis C Identifies Viperin as a Potential Antiviral Effector *
Authors: Helbig Karla J. , Lau Daryl T.-Y. , Semendric Ljiljana , Harley Hugh A. J. , Beard Michael R.
Published: August 2005

Interferon (IFN) α inhibits hepatitis C virus (HCV) replication both clinically and

in vitro

; however, the complete spectrum of interferon-stimulated genes (ISGs) expressed in the HCV-infected liver or the genes responsible for control of HCV replication have not been defined. To better define ISG expression in the chronically infected HCV liver, DNA microarray analysis was performed on 9 individuals with chronic hepatitis C (CHC). A total of 232 messenger RNAs were differentially regulated in CHC compared with nondiseased liver controls. A significant proportion of these were potential ISGs that were transcriptionally elevated, suggesting an ongoing response to endogenous IFN and/or double-stranded RNA. One ISG significantly elevated in all patients was viperin, an evolutionary conserved ISG that has antiviral activity against human cytomegalovirus. Stimulation of Huh-7 and HepG2 cells with IFN-α or -γ revealed viperin is predominantly a type I ISG. Furthermore, viperin expression could also be induced following transfection of Huh-7 cells with either poly(I:C) or HCV RNA. Transient expression of viperin in cells harboring the HCV genomic replicon resulted in a significant decrease in HCV replication, suggesting that viperin has anti-HCV activity.
In conclusion
, even in the face of a persistent HCV infection, there is an active ISG antiviral cellular response, highlighting the complexity of the host viral relationship. Furthermore, ISG viperin has anti-HCV activity

in vitro

; we postulate that viperin, along with other ISGs, acts to limit HCV replication. (Hepatology 2005.)
Theta oscillation‐coupled dendritic spiking integrates inputs on a long time scale
Authors: Huhn Zsófia , Orbán Gergő , Érdi Péter , Lengyel Máté
Published: August 2005

Persistent neural activity lasting for seconds after transient stimulation has been observed in several brain areas. This activity has been taken to be indicative of the integration of inputs on long time scales. Passive membrane properties render neural time constants to be on the order of milliseconds. Intense synaptic bombardment, characteristic of in vivo states, was previously shown to further reduce the time scale of effective integration. We explored how long‐term integration in single cells could be supported by dendritic spikes coupled with the theta oscillation, a prominent brain rhythm often observed during working memory tasks. We used a two‐compartmental conductance‐based model of a hippocampal pyramidal cell to study the interplay of intrinsic dynamics with periodic inputs in the theta frequency band. We show that periodic dendritic spiking integrates inputs by shifting the phase relative to an external oscillation, since spiking frequency is quasi‐linearly modulated by current injection. The time‐constant of this integration process is practically infinite for input intensities above a threshold (the integration threshold) and can be still several hundred milliseconds long below the integration threshold. The somatic compartment received theta frequency stimulation in antiphase with the dendritic oscillation. Consequently, dendritic spikes could only elicit somatic action potentials when they were sufficiently phase‐shifted and thus coincided with somatic depolarization. Somatic depolarization modulated the frequency but not the phase of firing, endowing the cell with the capability to code for two different variables at the same time. Inputs to the dendrite shifted the phase of dendritic spiking, while somatic input was modulating its firing rate. This mechanism resulted in firing patterns that closely matched experimental data from hippocampal place cells of freely behaving rats. We discuss the plausibility of our proposed mechanism and its potential to account for the firing pattern of cells outside the hippocampus during working memory tasks. © 2005 Wiley‐Liss, Inc.
Small interfering RNA targeting survivin sensitizes lung cancer cell with mutant p53 to adriamycin
Authors: Yonesaka Kimio , Tamura Kenji , Kurata Takayasu , Satoh Taroh , Ikeda Masato , Fukuoka Masahiro , Nakagawa Kazuhiko
Published: August 2005

Survivin is a member of the inhibitor of apoptosis protein (IAP) family that is specifically overexpressed in cancer tissues. p53 is one of the tumor suppressor genes; its induction in response to DNA damage causes apoptosis and correlates with drug sensitivity. To investigate the possible regulation of survivin by p53, we examined the level of survivin expression in lung cancer cell lines in response to adriamycin. Levels of survivin mRNA and protein in cell lines with wild‐type p53 decreased dramatically after p53 induction, but no such reduction of survivin was observed in cell lines with mutated or null p53. Inhibition of wild‐type p53 in A549 cells by small interfering (si) RNA significantly upregulated the expression of survivin. Survivin inhibition by siRNA in PC9 cells with mutated p53 significantly depressed cell proliferation. To investigate the sensitivity of cancer cells to adriamycin after inhibition of survivin, we depressed survivin expression using siRNA, and then added adriamycin at an IC50 dose. After a further 48 hr incubation with adriamycin, proliferation was significantly depressed in the cells treated with siRNA targeting survivin, in comparison with siRNA targeting scramble. Furthermore, both TUNEL and pro‐caspase3 expression assay showed a significant increase in apoptosis after combined treatment with adriamycin and siRNA targeting survivin. Our results demonstrate that survivin is downregulated by p53, and that siRNA targeting of survivin increases cell sensitivity to adriamycin and promotes apoptosis. siRNA targeting of survivin could be potentially useful for increasing sensitivity to anticancer drugs, especially in drug‐resistant cells with mutated p53. © 2005 Wiley‐Liss, Inc.
Role of interleukin‐18 and its receptor in hepatocellular carcinoma associated with hepatitis C virus infection
Authors: Asakawa Masami , Kono Hiroshi , Amemiya Hidetake , Matsuda Masanori , Suzuki Tetsuya , Maki Akira , Fujii Hideki
Published: August 2005

Interleukin (IL)‐18 is a proinflammatory cytokine that is up‐regulated in patients with hepatitis C virus (HCV) infection, which is the most common underlying disease in hepatocellular carcinoma (HCC). The purpose of our study was to investigate the role of IL‐18 in HCC associated with HCV infection. Sixty‐five patients with HCC and HCV infections who received curative surgical resections were examined in our study. The expression of the IL‐18 receptor was investigated in HCC tissues obtained from these patients and in 2 HCC cell lines. Nuclear factor (NF)‐κB activity and the expression of Bcl‐xL and xIAP mRNA were tested in the cell lines using recombinant human (rh) IL‐18. The IL‐18 receptor was expressed in both the HCC tissues and the cell lines. NF‐κB activation and the expression of Bcl‐xL and xIAP mRNA were increased by rhIL‐18. Moreover, rhIL‐18 suppressed the apoptosis of HCC cells which was induced by etoposide in vitro. The overall survival rate (55.4%) was significantly worse in the IL‐18 receptor‐positive patients than in the IL‐18 receptor‐negative patients (p = 0.015). In a Cox multivariate analysis, the expression of the IL‐18 receptor was found to be a significant predictor of a poor outcome in HCC patients. The expression of the IL‐18 receptor and an antiapoptotic mechanism involving NF‐κB activation in HCC cells may be implicated in a poor patient outcome. © 2005 Wiley‐Liss, Inc.
Lung cancer risks in relation to vegetable and fruit consumption and smoking
Authors: Rylander Ragnar , Axelsson Gösta
Published: August 2005

In a case‐control study on lung cancer, risk was analysed in relation to smoking habits and frequency of vegetable and fruit consumption. Lung cancer cases in West Sweden and population controls were interviewed using a questionnaire where the frequency of consumption of dietary items and smoking habits were assessed. The material presented comprises 177 female and 359 male confirmed cases of lung cancer and 916 population controls. There was a dose‐response relationship in regard to the number of cigarettes smoked and the number of years smoked, the latter factor being more important. After adjustment for number of cigarettes smoked/day and number of years smoked, the risk for those who seldom consumed vegetables was about twice of that among those who consumed vegetables frequently, both among nonsmokers, smokers and former smokers. This risk increase in relation to vegetable consumption also was present for different smoking categories. A similar tendency, although less pronounced, was found for fruit consumption. The results demonstrate that dietary factors are related to the risk for lung cancer, although smoking is the dominant risk factor. © 2005 Wiley‐Liss, Inc.
Occupational factors and risk of adult bone sarcomas: A multicentric case‐control study in Europe
Authors: Merletti Franco , Richiardi Lorenzo , Bertoni Franco , Ahrens Wolfgang , Buemi Antoine , Costa‐Santos Cristina , Eriksson Mikael , Guénel Pascal , Kaerlev Linda , Jöckel Karl‐Heinz , Llopis‐Gonzalez Agustin , Merler Enzo , Miranda Ana , Morales‐Suárez‐Varela Maria M. , Olsson Håkan , Fletcher Tony , Olsen Jorn
Published: August 2005

We investigated the association between occupational factors and risk of bone sarcoma, a rare tumor with a largely unknown aetiology. A multicentric case‐control study was conducted in 7 European countries in 1995–97. Ninety‐six cases aged 35–69 years with a centrally reviewed diagnosis of bone sarcoma (68 chondrosarcomas and 28 osteosarcomas) were compared to 2,632 population (68%) or colon cancer (32%) controls. Subjects were interviewed to obtain information on occupational, medical and reproductive history, smoking and alcohol consumption and selected exposures including use of pesticides. Response proportions were 90% among cases and 66% among controls. Odds ratios (OR) and 95% confidence intervals (CI) were estimated for selected categories of job titles and branches of industry and for use of pesticides. We found an increased OR for bone sarcoma among blacksmiths, toolmakers, machine‐tool operators (OR = 2.14, 95% CI 1.08–4.26), woodworkers (OR = 2.68, 95% CI 1.36–5.29) and construction workers (OR = 1.62, 95% CI 0.92–2.87). Ever users of pesticide had an OR of 2.33 (95% CI 1.31–4.13), with similar risks for exposure to insecticides and exposure to herbicides. Neither duration of employment in any of the analyzed occupational categories nor duration of use of pesticides showed an increasing trend in the risk of bone sarcoma. ORs of bone sarcoma were 1.03 (95% CI 0.23–4.57), 3.13 (95% CI 1.26–7.76) and 1.44 (95% CI 0.43–4.85) for the first, second and third tertile of days of use of pesticides. Our study suggests that novel and previously reported (woodworking) occupational factors play a role in the aetiology of bone sarcomas. © 2005 Wiley‐Liss, Inc.
A tumor vaccine containing anti‐CD3 and anti‐CD28 bispecific antibodies triggers strong and durable antitumor activity in human lymphocytes
Authors: Haas Claudia , Lulei Maria , Fournier Philippe , Arnold Annette , Schirrmacher Volker
Published: August 2005

We recently reported on newly designed virus‐targeted bispecific CD3‐ and CD28‐binding molecules for human T‐cell activation. When bound via one arm to a human virus‐modified tumor cell vaccine, these reagents caused a polyclonal T‐cell response and overcame the potential various T‐cell evasion mechanisms of tumor cells. In our current study, we demonstrated the induction of strong antitumor activity in human lymphocytes upon coincubation with a virus‐modified tumor vaccine containing anti‐CD3 and anti‐CD28 bispecific antibodies. Blood mononuclear cells or purified T cells that were coincubated with such a tumor vaccine for 3 days were able to destroy monolayers of human breast carcinoma and other carcinoma cells. Serial transfer to new tumor cell monolayers revealed antitumor cytotoxic activity in such effector cells that lasted for about 10 days. Nontumor target cells appeared to be much less sensitive to the activated effector cells. Although the bispecific molecules alone did not activate effector cells, their binding to virus‐infected tumor cells was important and more effective than their binding to free virus. Antitumor activity of the activated effector cells was mediated through soluble factors as well as through direct cell contact of effector cells with the nontargeted bystander tumor cells. Since the virus‐modified tumor vaccine is well tolerated and already exhibits a certain effectiveness in cancer patients, the combination with new bispecific molecules has the potential to introduce additional antitumor effects. The reagents can also be combined with Newcastle Disease Virus (NDV)‐based oncolytic virotherapy. © 2005 Wiley‐Liss, Inc.
Distinct regulation and impact of type 1 T‐cell immunity against HPV16 L1, E2 and E6 antigens during HPV16‐induced cervical infection and neoplasia
Authors: van Poelgeest Mariëtte I.E. , Nijhuis Esther R. , Kwappenberg Kitty M.C. , Hamming Ineke E. , Wouter Drijfhout Jan , Fleuren Gert Jan , van der Zee Ate G.J. , Melief Cornelis J.M. , Kenter Gemma G. , Nijman Hans W. , Offringa Rienk , van der Burg Sjoerd H.
Published: August 2005

Cervical cancer is the possible outcome of a genital infection with high‐risk human papillomavirus type 16 (HPV16) and is preceded by a phase of persistent HPV infection during which the host immune system fails to eliminate the virus. Our previous work showed that failure is reflected by the absence of type 1 T‐cell immunity against HPV16 early antigens E2 and E6 in patients with HPV16+ cervical lesions. We now show that a majority of both patients with cervical lesions and healthy subjects display HPV16 L1 peptide‐specific type 1 T‐cell responses with similar magnitude. The T‐cell response in patients was directed at a broad range of peptides within L1, suggesting that during persistent or repeated exposure to HPV16 L1, the immune system maximizes its efforts to counter the viral challenge. Unlike the type 1 T‐cell responses against HPV16 early antigens E2 and E6, type 1 T‐cell immunity against L1 does not correlate with health or disease. This argues that T‐cell responses against early and late HPV16 antigens essentially differ in the manner in which they are induced and regulated, as well as in their impact on the subsequent stages of HPV16‐induced cervical disease. © 2005 Wiley‐Liss, Inc.
Proliferation and Cdk4 expression in microsatellite unstable colon cancers with TGFBR2 mutations
Authors: Grady William M. , Willis Joseph E. , Trobridge Patty , Romero‐Gallo Judith , Munoz Nina , Olechnowicz Joseph , Ferguson Kelly , Gautam Shiva , Markowitz Sanford D.
Published: August 2005

Approximately 15% of human colon cancers have microsatellite instability (MSI) and carry frameshift mutations in a polyadenine tract (BAT‐RII) in the type II transforming growth factor β (TGF‐β) receptor (TGFBR2), a required component of the TGF‐β receptor. The BAT‐RII mutations in MSI colon cancers make the tumors resistant to the effects of TGF‐β. In cultured epithelial cells, TGF‐β can inhibit cell proliferation and induce apoptosis, and in vitro it can regulate the expression of a variety of cyclins, cyclin‐dependent kinases (cdks) and cdk inhibitors. These effects are context‐ and tissue type‐dependent, raising questions about which of these in vitro effects of TGF‐β signaling inactivation contribute to the formation of primary colon cancer. Thus, this study sought to determine the pathogenetically relevant effects of TGFBR2 inactivation in primary MSI colon cancers with mutant BAT‐RII. Colon cancers with mutant BAT‐RII were found to have increased proliferation compared to cancers with wild‐type BAT‐RII. Assessment of cdk4, cyclin D1 and p27kip1 expression revealed that only cdk4 expression was increased in the cancers with mutant BAT‐RII. In order to determine if TGFBR2 inactivation was the cause of these changes, TGFBR2 was reconstituted in an MSI colon cancer cell line, resulting in decreased proliferation and decreased cdk4 expression and kinase activity. These results suggest that TGFBR2 mutations in primary colon cancers may be responsible for the increased proliferation and cdk4 expression in these tumors and provide evidence that deregulation of cdk4 is a pathogenic in vivo consequence of TGFBR2 inactivation in primary colon cancer. © 2005 Wiley‐Liss, Inc.
Wnt pathway is involved in pleomorphic adenomas induced by overexpression of PLAG1 in transgenic mice
Authors: Zhao Xudong , Ren Weihua , Yang Wenjun , Wang Yi , Kong Hui , Wang Long , Yan Lanzhen , Xu Guojiang , Fei Jian , Fu Jiliang , Zhang Chenping , Wang Zhugang
Published: August 2005

Pleomorphic adenoma gene 1 (PLAG1) was found frequently rearranged and activated in human salivary gland pleomorphic adenomas. It encodes a developmentally regulated transcription factor. Ectopic overexpression of PLAG1 has been proposed to play a crucial role in tumorigenesis of salivary gland pleomorphic adenomas. It was reported that PLAG1 can activate the transcription of insulin‐like growth factor 2 (IGF2), functioning as a protooncogene. In this report, we show that the salivary gland tumors developed in PLAG1 transgenic mice share major histopathologic features with human pleomorphic adenomas. It was found that β‐catenin, the key component of Wnt signaling pathway, was upregulated at transcriptional level in tumors developed in 3 independent transgenic mouse lines. Immunohistochemical staining revealed that expression of β‐catenin as well as c‐myc, downstream of β‐catenin in Wnt signaling pathway, was highly upregulated with overexpression of PLAG1 transgene in tumor and normal transgenic salivary gland tissues. Moreover, we found that PLAG1 can activate the transcription of mouse but not human β‐catenin in the 3T3 cells cotransfected with reporter constructs. Sequence analysis shows there are 4 PLAG1 consensus binding sites in mouse β‐catenin promoter region but not in human. Our findings provide the first in vivo evidence for the oncogenic activity of PLAG1 in pleomorphic adenoma tumorigenesis, reveal a valued animal model for human salivary gland tumors and suggest that Wnt signaling pathway may also contribute to the development of pleomorphic adenomas in transgenic mice. © 2005 Wiley‐Liss, Inc.
Inefficient proteasomal‐degradation pathway stabilizes AP‐2α and activates HER‐2/neu gene in breast cancer
Authors: Li Min , Wang Yingqun , Hung Mien‐Chie , Kannan Perry
Published: August 2005

HER‐2/neu proto‐oncogene is overexpressed in about one fourth of human breast cancers. AP‐2 transcription factors bind to the HER‐2/neu gene promoter and activate its expression. In a striking concurrence, anomalous abundance of AP‐2α protein or its homolog AP‐2γ is also detected with HER‐2/neu protein in mammary tumor‐derived cell lines. This suggests that the deregulation of AP‐2 is the preceding pathogenic event and probably the pivotal one in this type of mammary carcinogenesis. We examined the process of AP‐2α gene expression in mammary carcinoma cell lines to identify where the aberration had occurred. We found no amplification of the AP‐2α gene. Its promoter was marginally upregulated; however, it did not significantly increase the mRNA levels. When the AP‐2α protein was examined, a remarkable stability was seen in breast cancer cell lines MDA‐MB‐453 and SK‐BR‐3, with a half‐life of over 30 hr. This is sharply higher than the approximate 1 hr observed in mammary epithelial cell line MCF‐10A and murine cell line NIH 3T3. Treatment of MCF‐10A and NIH 3T3 cells with the proteasome inhibitor MG‐132 showed that AP‐2α was ubiquitinated and its level significantly increased. Moreover, this increase was accompanied by elevated levels HER‐2/neu protein. In contrast, weaker ubiquitination of AP‐2α was seen in MDA‐MB‐453 and SK‐BR‐3 cancer cells, and MG‐132 treatment did not raise the AP‐2α level any further. These results uncover that unusual stability is the main mechanism that raises the levels of AP‐2 proteins, and in addition, provide the first clue that defective ubiquitin‐dependent proteasomal‐degradation pathway is possibly the prime cause that affects the HER‐2/neu gene and culminates in breast cancer. © 2005 Wiley‐Liss, Inc.
Cytotoxic effects induced by a combination of cyclopamine and gefitinib, the selective hedgehog and epidermal growth factor receptor signaling inhibitors, in prostate cancer cells
Authors: Mimeault Murielle , Moore Erik , Moniaux Nicolas , Hénichart Jean‐Pierre , Depreux Patrick , Lin Ming‐Fong , Batra Surinder K.
Published: August 2005

Although the blockade of the hedgehog cascade by using cyclopamine has been reported to inhibit the growth of some cancer cell types, few studies on the mechanism by which this drug alone or in combination with other cytotoxic agents induces its cytotoxic effect have been reported. In our study, we evaluate, for the first time, the antiproliferative and cytotoxic effects induced by a combination of selective SMO inhibitor, cyclopamine and epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, gefitinib on metastatic prostate cancer (PC) cells. The results revealed that cyclopamine, alone or at a lower concentration in combination with gefitinib, inhibited the growth of sonic hedgehog‐ (SHH), epidermal growth factor‐ (EGF) and serum‐stimulated androgen‐sensitive LNCaP‐C33 and LNCaP‐LN3 and androgen‐independent LNCaP‐C81, DU145 and PC3 cells. The antiproliferative effect of cyclopamine and gefitinib, alone or in combination, was mediated via a blockade of the PC3 cells in the G1 phase of the cell cycle. Importantly, the combined cyclopamine and gefitinib also caused a higher rate of apoptotic death of PC cells compared to single agents. The cytotoxic effect induced by these drugs in PC3 cells appears to be mediated at least, in part, via the mitochondrial pathway through the depolarization of the mitochondrial membrane and the release of cytochrome c and reactive oxygen species into the cytosol. This was also accompanied by the activation of caspase cascades, PARP cleavage and DNA fragmentation. Additionally, the combined cyclopamine and gefitinib were more effective at suppressing the invasiveness of PC3 cells through matrigel in vitro as the drugs alone. These findings indicate that the simultaneous blockade of SHH–GLI‐1 and EGF–EGFR signaling, which results in the growth arrest and massive rate of apoptotic cell death, represents a promising strategy for a more effective treatment of metastatic PC forms. © 2005 Wiley‐Liss, Inc.
Expression of dentin sialophosphoprotein in human prostate cancer and its correlation with tumor aggressiveness
Authors: Chaplet Michaël , Waltregny David , Detry Cédric , Fisher Larry W. , Castronovo Vincent , Bellahcène Akeila
Published: August 2005

Recent studies have demonstrated that two SIBLING family members, bone sialoprotein (BSP) and osteopontin (OPN), are overexpressed in human prostate cancer. The expression of these proteins is associated with the acquisition of a metastatic phenotype by cancer cells and a poor prognosis for the patient. Dentin sialophosphoprotein (DSPP) shares several structural and genetic features with OPN and BSP. The presence of DSPP has been recently established in salivary glands, indicating that its expression is not restricted to mineralized tissues. However, its potential expression in human tumors has not been addressed yet. In this study, we sought to evaluate the expression of DSPP in human prostate cancer. Immunohistochemistry was performed on 69 prostate cancer specimens using LFMb‐21 anti‐DSPP monoclonal antibody. All of the prostate cancer lesions examined expressed detectable levels of DSPP, as compared with no or low level of expression in adjacent normal glands (p < 0.0001). High grade prostatic intraepithelial neoplasia (HGPIN) glands generally displayed DSPP expression levels that were similar to those found in neighboring cancer glands. DSPP expression was significantly associated with the pathological stage (p = 0.0087) and the Gleason score (p = 0.0176) of the tumors. Western Blot was performed on 5 representative prostate tumor extracts and 3 prostatic tumor cell lines (PC3, LNCaP and DU145). All tumor extracts and cell lines analyzed have been found to express DSPP. In addition, in situ hybridization was used to assess the presence of DSPP mRNA. DSPP was detected at the RNA level in both HGPIN and tumoral glands. This study shows for the first time that DSPP is ectopically expressed in human prostate cancer. The expression of this SIBLING protein strongly correlates with conventional histopathological prognostic indicators of prostate cancer progression. © 2005 Wiley‐Liss, Inc.
Aberrant promoter methylation of multiple genes during multistep pathogenesis of colorectal cancers
Authors: Takahashi Takao , Shigematsu Hisayuki , Shivapurkar Narayan , Reddy Jyotsna , Zheng Yingye , Feng Ziding , Suzuki Makoto , Nomura Masaharu , Augustus Meena , Yin Jing , Meltzer Stephen J. , Gazdar Adi F.
Published: August 2005

Aberrant methylation of 5′gene promoter regions associated with gene silencing is an epigenetic phenomenon responsible for silencing of tumor suppressor genes in many cancer types. The aims of our study were to study the role of methylation of a large panel of genes during multistage pathogenesis and to correlate our findings with patient age and other clinico‐pathological features. We investigated the aberrant promoter methylation profile of 19 genes in 92 colorectal cancers (CRCs) and corresponding nonmalignant epithelia (NME) (n = 57), and selected 15 genes for studying 26 colorectal adenomas (CAs). On the Basis of our results, the genes could be divided into 3 groups. Group 1 consisted of 13 genes whose methylation was tumor‐specific. For 8 of these genes, the methylation frequencies in CAs were similar to those of CRCs, but significantly different from the frequencies in NME. Group 2, consisting of 2 genes demonstrating little or no methylation, were present in any sample type. In Group 3, consisting of 4 genes, relatively frequent methylation was present in both CRCs and NME, and the differences between these specimen types were not significant. Methylation of Group 1 genes were tightly correlated with each other, and these genes demonstrated increased methylation frequencies in CRCs with increasing age. Methylation was not correlated with other clinico‐pathological features. In general, methylation frequencies of CAs were intermediate between CRCs and NME. Our study constitutes the most comprehensive methylation profile of CRCs, demonstrates that methylation commences early during CRC pathogenesis and is an age‐related phenomenon. © 2005 Wiley‐Liss, Inc.
Human fibroblast reactions to standard and electropolished titanium and Ti–6Al–7Nb, and electropolished stainless steel
Authors: Meredith D. Osian , Eschbach Lukas , Wood Mairead A. , Riehle Mathis O. , Curtis Adam S.G. , Richards R. Geoffrey
Published: August 2005

Stainless steel (SS), titanium (cpTi), and Ti–6Al–7Nb (TAN) are frequently used metals in orthopedic internal fracture fixation. Although reactivity to SS and cpTi are noted in reference, the soft tissue compatibility of TAN has not been comprehensively studied. This study focuses on the in vitro soft tissue compatibility of TAN in comparison to SS and cpTi using a human fibroblast model. The industrial standard surface finishes of these three materials vary considerably in view of their use in similar applications. To distinguish between material parameters of topography and chemistry, we have included electropolished (e.p) counterparts of the standard preparations of cpTi and TAN in the study (standard SS is e.p). All materials were characterized using atomic force microscopy, profilometry, and scanning electron microscopy. Our findings demonstrate that cell morphology and growth rate was similar for SS, and e.p. cpTi and TAN, with cells well spread and forming a confluent monolayer by 10 days. Cell growth on standard cpTi was similar to the electropolished samples; however, they showed a less spread morphology with more filopodia and surface ruffling present. Cell morphology on standard TAN was rounded or elongated and proliferation was inhibited at all time points, with possible cell necrosis by day 10. We found evidence of endocytosis of β‐phase particles originating from the standard TAN surface. We believe that the particle uptake coupled with the characteristic surface topography contribute to the noncytocompatibility of fibroblasts on standard TAN. © 2005 Wiley Periodicals, Inc. J Biomed Mater Res, 2005
Characterization of polymeric poly(ϵ‐caprolactone) injectable implant delivery system for the controlled delivery of contraceptive steroids
Authors: Dasaratha Dhanaraju Magharla , Gopinath Damodaran , Rafiuddin Ahmed Mohamed , Jayakumar Rajadas , Vamsadhara Chandrasekar
Published: August 2005

Contraceptive steroids levonorgestrel (LNG) and ethinyl estradiol (EE) have been encapsulated with poly(ϵ‐caprolactone) (PCL) microspheres using a w/o/w double emulsion method. The microspheres prepared were smooth and spherical, with a mean size from 8–25 μm. In vitro release profiles of microspheres showed a trend of increasing initially at the first week, and thereafter the release was sustained. At the end of the seventh week LNG/EE from 1:5 and 1:10 PCL microspheres were 60 and 48%, 52 and 46%, respectively. An in vitro degradation study shows that at the 20th week the microspheres maintained the surface integrity. The PCL microspheres showed a triphasic in vivo release profile with an initial burst effect due to the release of the steroid adsorbed on the microsphere surface, a second sustained release phase due to the steroid diffusion through the pores or channels formed in the polymer matrix, and third phase due to polymer bioerodible. Histological examination of PCL microspheres injected intramuscularly into thigh muscle of a rat showed a minimal inflammatory reaction demonstrating that contraceptive steroid‐loaded microspheres were biocompatible. The level of inflammatory cytokines determined by immunostaining for IL‐1α, the tissue response to formulations at the first week was considered mild, whereas at the end of the 20th week the inflammatory response ceased. Thus, this study helped us to evaluate the feasibility of using these microspheres as a long‐acting biodegradable drug delivery system for contraceptive steroids. © 2005 Wiley Periodicals, Inc. J Biomed Mater Res, 2006
A GPIIb/IIIa bioreactor for specific treatment of immune thrombocytopenic purpura, an autoimmune disease. Preparation, in vitro characterization, and preliminary proof‐of‐concept animal studies
Authors: Emregul Emel , David Allan , Balthasar Joseph P. , Yang Victor C.
Published: August 2005

Immune thrombocytopenic purpura (ITP) is an autoimmune disease that affects thousands of Americans each year. The resulting thrombocytopenia, which develops from destruction of platelets (PLT) by anti‐PLT autoantibodies (APAb), is often associated with hemorrhagic complications. Existing therapies are not effective and are associated with significant morbidity. Recently, a new treatment modality using plasmapheresis with a Protein‐A column has shown some clinical promise. Yet, although this method would remove the pathogenic APAb, it would also deplete protective antibodies, thereby weakening the body's self‐defense system. Because about 80% of patients with ITP develop APAb against the GPIIb/IIIa antigens on PLT, a novel approach of attaching a GPIIb/IIIa‐linked bioreactor with an extracorporeal circuit is suggested herein to achieve highly effective/specific APAb removal and overcome shortcomings of plasmapheresis in treating ITP. A hollow fiber‐based bioreactor device was fabricated, and GPIIb/IIIa antigens were immobilized onto the inner lumens of the hollow fibers by using the epichlorohydrin activation method. An optimized bioreactor containing a loading of 1.63 mg GPIIb/IIIa/g fibers and adsorption capacity of 1.9 mg 7E3/g fibers was developed. Preliminary proof‐of‐concept investigation using a 7E3‐induced thrombocytopenic rat model (which mimicked clinical ITP) was carried out. A complete (100%) return of PLT counts to their initial levels was observed in rats within 6 h after the GPIIb/IIIa bioreactor treatment. In addition, a rapid restoration of WBC counts in the treated rats was also found. These preliminary findings shed light of promise of using the GPIIb/IIIa bioreactor approach in achieving highly improved ITP therapy. © 2005 Wiley Periodicals, Inc. J Biomed Mater Res, 2005