bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of a Pseudomonad 2-Nitrobenzoate Nitroreductase and Its Catabolic Pathway-Associated 2-Hydroxylaminobenzoate Mutase and a Chemoreceptor Involved in 2-Nitrobenzoate Chemotaxis
Authors: Iwaki Hiroaki , Muraki Takamichi , Ishihara Shun , Hasegawa Yoshie , Rankin Kathryn N. , Sulea Traian , Boyd Jason , Lau Peter C. K.
Published: February 2007

Pseudomonas fluorescens
strain KU-7 is a prototype microorganism that metabolizes 2-nitrobenzoate (2-NBA) via the formation of 3-hydroxyanthranilate (3-HAA), a known antioxidant and reductant. The initial two steps leading to the sequential formation of 2-hydroxy/aminobenzoate and 3-HAA are catalyzed by a NADPH-dependent 2-NBA nitroreductase (NbaA) and 2-hydroxylaminobenzoate mutase (NbaB), respectively. The 216-amino-acid protein NbaA is 78% identical to a plasmid-encoded hypothetical conserved protein of
Polaromonas
strain JS666; structurally, it belongs to the homodimeric NADH:flavin mononucleotide (FMN) oxidoreductase-like fold family. Structural modeling of complexes with the flavin, coenzyme, and substrate suggested specific residues contributing to the NbaA catalytic activity, assuming a ping-pong reaction mechanism. Mutational analysis supports the roles of Asn40, Asp76, and Glu113, which are predicted to form the binding site for a divalent metal ion implicated in FMN binding, and a role in NADPH binding for the 10-residue insertion in the β5-α2 loop. The 181-amino-acid sequence of NbaB is 35% identical to the 4-hydroxylaminobenzoate lyases (PnbBs) of various 4-nitrobenzoate-assimilating bacteria, e.g.,
Pseudomonas putida
strain TW3. Coexpression of
nbaB
with
nbaA
in
Escherichia coli
produced a small amount of 3-HAA from 2-NBA, supporting the functionality of the
nbaB
gene. We also showed by gene knockout and chemotaxis assays that
nbaY
, a chemoreceptor NahY homolog located downstream of the
nbaA
gene, is responsible for strain KU-7 being attracted to 2-NBA. NbaY is the first chemoreceptor in nitroaromatic metabolism to be identified, and this study completes the gene elucidation of 2-NBA metabolism that is localized within a 24-kb chromosomal locus of strain KU-7.
pilQ
Missense Mutations Have Diverse Effects on PilQ Multimer Formation, Piliation, and Pilus Function in
Neisseria gonorrhoeae
Authors: Helm R. Allen , Barnhart Michelle M. , Seifert H. Steven
Published: February 2007

Type IV pili are required for virulence in
Neisseria gonorrhoeae
, as they are involved in adherence to host epithelium, twitching motility, and DNA transformation. The outer membrane secretin PilQ forms a homododecameric ring through which the pilus is proposed to be secreted.
pilQ
null mutants are nonpiliated, and thus, all pilus-dependent functions are eliminated. Mutagenesis was performed on the middle one-third of
pilQ
, and mutants with colony morphologies consistent with the colony morphology of nonpiliated or underpiliated bacteria were selected. Nineteen mutants, each with a single amino acid substitution, were isolated and displayed diverse phenotypes in terms of PilQ multimer stability, pilus expression, transformation efficiency, and host cell adherence. The 19 mutants were grouped into five phenotypic classes based on functionality. Four of the five mutant classes fit the current model of pilus functionality, which proposes that a functional pilus assembly apparatus, not necessarily full-length pili, is required for transformation, while high levels of displayed pili are required for adherence. One class, despite having an underpiliated colony morphology, expressed high levels of pili yet adhered poorly, demonstrating that pilus expression is necessary but not sufficient for adherence and indicating that PilQ may be directly involved in host cell adherence. The collection of phenotypes expressed by these mutants suggests that PilQ has an active role in pilus expression and function.
Role of L-type calcium channels and PKC in active tone development in rabbit coronary artery
Authors: Cobine Caroline A. , Callaghan Brid P. , Keef Kathleen D.
Published: February 2007

The present study investigated active tone development in isolated ring segments of rabbit epicardial coronary artery. Endothelium-denuded (E−) or endothelium-intact (E+) vessels treated with the NO synthase inhibitor Nω-nitro-l-arginine (100 μM) developed active tone, which was enhanced by stretch and reversed by the NO donor sodium nitroprusside (SNP; IC50= 9 nM). Nifedipine abolished active tone and the contractile response to phorbol dibutyrate (PDBu; 10 nM) with the same potency (IC50= 8 nM), whereas 300 nM PDBu responses were only partially blocked by nifedipine. The classical and novel PKC inhibitors GF-109203X (IC50= 1–2 μM) and chelerythrine (IC50= 4–5 μM) and the classical PKC inhibitor Gö-6976 (IC50= 0.3–0.4 μM) blocked both active tone and 10 nM PDBu responses with similar potency. Active tone development was associated with depolarization of membrane potential ( Em) and a shift to the left of the Em-vs.-contraction relationship determined by varying extracellular potassium. The depolarization and leftward shift were reversed by either chelerythrine (10 μM) or SNP (30 nM). PDBu (100–300 nM) increased peak L-type calcium channel (Cav) currents in isolated coronary myocytes, and this effect was reversed by chelerythrine (1 μM) or Gö-6976 (200 nM). SNP (500 nM) reduced Cavcurrents only in the presence of the PKA blocker 8-bromo-2′- O-monobutyryl-cAMPS, Rp isomer (10 μM). In conclusion, active tone development in coronary artery is suppressed by basal NO release and is dependent on both enhanced Cavactivity and classical PKC activity. Both Em-dependent and -independent processes contribute to contraction. Our results suggest that one Em-independent process is direct enhancement of Cavcurrent by PKC.
Macromolecular transport in heart valves. I. Studies of rat valves with horseradish peroxidase
Authors: Zeng Zhongqing , Yin Yongyi , Huang An-Li , Jan Kung-Ming , Rumschitzki David S.
Published: February 2007

The present study aims to experimentally elucidate subtle structural features of the rat valve leaflet and the related nature of macromolecular transport across its endothelium and in its subendothelial space, information necessary to construct a rational theoretical model that can explain observation. After intravenous injection of horseradish peroxidase (HRP), we perfusion-fixed the aortic valve of normal Sprague-Dawley rats and found under light microscopy that HRP leaked through the leaflet's endothelium at very few localized brown spots, rather than uniformly. These spots grew nearly as rapidly with HRP circulation time before euthanasia as aortic spots, particularly when the time axis only included the time the valve was closed. These results suggest that macromolecular transport in heart valves depends not only on the direction normal to, but also parallel to, the endothelial surface and that convection, as well as molecular diffusion, plays an important role in macromolecular transport in heart valves. Transmission electron microscopy of traverse leaflet sections after 4-min HRP circulation showed a very thin (∼150 nm), sparse layer immediately beneath the endothelium where the HRP concentration was much higher than that in the matrix below it. Nievelstein-Post et al.'s (Nievelstein-Post P, Mottino G, Fogelman A, Frank J. Arterioscler Thromb 14: 1151–1161, 1994) ultrarapid freezing/rotary shadow etching of the normal rabbit valve's subendothelial space supports the existence of this very thin, very sparse “valvular subendothelial intima,” in analogy to the vascular subendothelial intima.
Periodic Solutions of Evolutionm-Laplacian Equations with a Nonlinear Convection Term
Authors: Liu Wenjun
Published: February 2007

We show the existence and gradient estimates of periodic solutions in the case of0≤α<m+1to the evolutionm-Laplacian equations of formut−div{|∇u|m∇u}+b(u)⋅∇u=f(t)uα+h(x,t),inΩ×ℝ1with the Dirichlet boundary value condition.
On a Class of Integrals Involving a Bessel Function Times Gegenbauer Polynomials
Authors: Koumandos Stamatis
Published: February 2007

We provide information and explicit formulae for a class of integrals involving Bessel functions and Gegenbauer polynomials. We present a simple proof of an old formula of Gegenbauer. Some interesting special cases and applications of this result are obtained. In particular, we give a short proof of a recent result of A. A. R. Neves et al. regarding the analytical evaluation of an integral of a Bessel function times associated Legendre functions. These integrals arise in problems of vector diffraction in electromagnetic theory.
Random Three-Step Iteration Scheme and Common Random Fixed Point of Three Operators
Authors: Plubtieng Somyot , Kumam Poom , Wangkeeree Rabian
Published: February 2007

We construct random iterative processes with errors for three asymptotically nonexpansive random operators and study necessary conditions for the convergence of these processes. The results presented in this paper extend and improve the recent ones announced by I. Beg and M. Abbas (2006), and many others.
Metabolic Functions of Peroxisome Proliferator-Activated Receptorβ/δin Skeletal Muscle
Authors: Gaudel Céline , Grimaldi Paul A.
Published: February 2007

Peroxisome proliferator-activated receptors (PPARs) are transcription factors that act as lipid sensors and adapt the metabolic rates of various tissues to the concentration of dietary lipids. PPARs are pharmacological targets for the treatment of metabolic disorders. PPARαand PPARγare activated by hypolipidemic and insulin-sensitizer compounds, such as fibrates and thiazolidinediones. The roles of PPARβ/δin metabolic regulations remained unclear until recently. Treatment of obese monkeys and rodents by specific PPARβ/δagonists promoted normalization of metabolic parameters and reduction of adiposity. Recent evidences strongly suggested that some of these beneficial actions are related to activation of fatty acid catabolism in skeletal muscle and also that PPARβ/δis involved in the adaptive responses of skeletal muscle to environmental changes, such as long-term fasting or physical exercise, by controlling the number of oxidative myofibers. These observations indicated that PPARβ/δagonists might have therapeutic usefulness in metabolic syndrome by increasing fatty acid consumption in skeletal muscle and reducing obesity.
PPARs, Obesity, and Inflammation
Authors: Stienstra Rinke , Duval Caroline , Müller Michael , Kersten Sander
Published: February 2007

The worldwide prevalence of obesity and related metabolic disorders is rising rapidly, increasing the burden on our healthcare system. Obesity is often accompanied by excess fat storage in tissues other than adipose tissue, including liver and skeletal muscle, which may lead to local insulin resistance and may stimulate inflammation, as in steatohepatitis. In addition, obesity changes the morphology and composition of adipose tissue, leading to changes in protein production and secretion. Some of these secreted proteins, including several proinflammatory mediators, may be produced by macrophages resident in the adipose tissue. The changes in inflammatory status of adipose tissue and liver with obesity feed a growing recognition that obesity represents a state of chronic low-level inflammation. Various molecular mechanisms have been implicated in obesity-induced inflammation, some of which are modulated by the peroxisome proliferator-activated receptors (PPARs). PPARs are ligand-activated transcription factors involved in the regulation of numerous biological processes, including lipid and glucose metabolism, and overall energy homeostasis. Importantly, PPARs also modulate the inflammatory response, which makes them an interesting therapeutic target to mitigate obesity-induced inflammation and its consequences. This review will address the role of PPARs in obesity-induced inflammation specifically in adipose tissue, liver, and the vascular wall.
Knowledge and Awareness of Congenital Cytomegalovirus Among Women
Authors: Jeon Jiyeon , Victor Marcia , Adler Stuart P. , Arwady Abigail , Demmler Gail , Fowler Karen , Goldfarb Johanna , Keyserling Harry , Massoudi Mehran , Richards Kristin , Staras Stephanie A. S. , Cannon Michael J.
Published: February 2007

Background. Congenital cytomegalovirus (CMV) infection is a leading cause of disabilities in children, yet the general public appears to have little awareness of CMV. Methods. Women were surveyed about newborn infections at 7
different geographic locations. Results. Of the 643 women surveyed, 142 (22%) had heard of congenital CMV. Awareness increased with increasing levels of education (P<.0001). Women who had worked as a healthcare professional had a higher
prevalence of awareness of CMV than had other women (56% versus 16%, P <.0001). Women who were aware of CMV were most likely to have heard about it from a healthcare
provider (54%), but most could not correctly identify modes of
CMV transmission or prevention. Among common causes of birth defects and
childhood illnesses, women′s awareness of CMV ranked last. Conclusion. Despite its large public health
burden, few women had heard of congenital CMV, and even fewer were aware of
prevention strategies.
Photo‐induced reduction of Noble metal ions to metal nanoparticles on tubular J‐aggregates
Authors: Kirstein Stefan , von Berlepsch Hans , Böttcher Christoph
Published: February 2007

Palladium and silver nanoparticles are formed on the surface of tubular J‐aggregates of an amphiphilic tetrachlorobenzimidacarbocyanine dye by reduction of the respective metal cations in aqueous solution. Upon addition of the palladium complex Na2PdCl4to the aggregate solution, the absorption spectrum shows significant changes which is explained by partial destruction of the aggregates. Cryogenic transmission electron microscopy (cryo‐TEM) images show that the tubular J‐aggregates are randomly covered by well‐separated Pd nanoparticles of approximately 1–3 nm size. Larger particles and higher particle density along the aggregates are obtained when an auxiliary reducing agent is added to the solution. The presence of the metallic particles leads to efficient fluorescence quenching giving clear evidence for super quenching. In similar experiments using AgNO3, silver nanoparticles are grown which are larger in size but less dense distributed along the aggregates. At least in the case of the silver particles, the spontaneous formation of metal nanoparticles is assumed to be initiated by a photo‐induced electron transfer process (PET).
Epoxy‐Carbazole Polymeric Network Nanolayers for Organic Light‐Emitting Devices
Authors: Park Jin-Woo , Lee Eung-Kyoo , Choi Myon Cheon , Kim Hwajeong , Keum Jihwan , Ha Chang-Sik , Kim Youngkyoo
Published: February 2007

We report the study of epoxy‐carbazole polymeric network (ECzPN) nanolayers as a
hole injection/transport layer in organic light‐emitting devices. The ECzPN nanolayers
were prepared by the thermal curing reaction of epoxidized cresol novolak and 3,6‐
diaminocarbazole in the presence of catalytic amount of triphenyl phosphine. The
curing reaction was examined with Fourier transformed infrared spectroscopy, whilst the
thermal stability was studied with thermogravimetric analysis. Optical absorption and
emission spectroscopy were employed to investigate the optical properties of ECzPN
nanolayers, whilst atomic force microscopy was used to examine the surface
nanomorphology of ECzPN nanolayers. The result showed that the device performance
was greatly influenced by the weight ratio of monomers, because the highest occupied
molecular orbital level of ECzPN was significantly changed with the ratio. This is
attributed to the ground‐state complexes induced by the specific interaction (hydrogen
bonding) between the lone pair electrons in amines of carbazole moieties and the
hydroxyl group of ring‐opened epoxide moieties.
Effect of Pressure on Synthesis of Pr‐Doped Zirconia Powders Produced by Microwave‐Driven Hydrothermal Reaction
Authors: Opalinska A. , Leonelli C. , Lojkowski W. , Pielaszek R. , Grzanka E. , Chudoba T. , Matysiak H. , Wejrzanowski T. , Kurzydlowski K. J.
Published: February 2007

A high‐pressure microwave reactor was used to study the hydrothermal synthesis of
zirconia powders doped with 1 mol % Pr. The synthesis was performed in the pressure range
from 2 to 8 MPa corresponding to a temperature range from 215C∘ to 305C∘. This technology
permits a synthesis of nanopowders in short time not limited by thermal inertia of the vessel.
Microwave heating permits to avoid contact of the reactants with heating elements, and is thus
particularly well suited for synthesis of doped nanopowders in high purity conditions.
A mixture of ZrO2 particles with tetragonal and monoclinic crystalline phases, about 15 nm in size, was obtained.
The p/T threshold of about 5‐6 MPa/265–280C∘ was necessary to obtain good quality of
zirconia powder. A new method for quantitative description of grain‐size distribution was applied, which is
based on analysis of the fine structure of the X‐ray diffraction line profiles. It permitted to
follow separately the effect of synthesis conditions on the grain‐size distribution of the
monoclinic and tetragonal phases.
Hematopoietic Prostaglandin D Synthase Suppresses Intestinal Adenomas inApcMin/+ Mice
Authors: Park Jae Man , Kanaoka Yoshihide , Eguchi Naomi , Aritake Kosuke , Grujic Sava , Materi Alicia M. , Buslon Virgilio S. , Tippin Brigette L. , Kwong Alan M. , Salido Eduardo , French Samuel W. , Urade Yoshihiro , Lin Henry J.
Published: February 2007

Aspirin and other nonsteroidal anti-inflammatory drugs prevent some cases of colon cancer by inhibiting prostaglandin (PG) synthesis. PGE2 promotes colon neoplasia, as shown by knockout mouse studies on enzymes and receptors in the PG cascade. A few experiments 20 to 30 years ago suggested that PGD2 may suppress tumors, but a role for biosynthetic enzymes for PGD2 in tumor development has not been studied. We report here that disruption of the gene for hematopoietic PGD synthase in ApcMin/+ mice led to ∼50% more intestinal adenomas compared with controls. Tumor size was not affected. By immunohistochemistry, we detected hematopoietic PGD synthase mainly in macrophages and monocytes of the gut mucosa. The mean number of tumors did not increase with knockout of the gene for the lipocalin type of the enzyme, which is not produced in the intestine. On the other hand, ApcMin/+ mice with transgenic human hematopoietic PGD synthase tended to have 80% fewer intestinal adenomas. The transgene produced high mRNA levels (375-fold over endogenous). There was a suggestion of higher urinary excretion of 11β-PGF2α and a lower excretion of a PGE2 metabolite in transgenic mice, but differences (30–40%) were not statistically significant. The results support an interpretation that hematopoietic PGD synthase controls an inhibitory effect on intestinal tumors. Further studies will be needed to prove possible mechanisms, such as routing of PG production away from protumorigenic PGE2 or inhibition of the nuclear factor-κB cascade by PGD2 metabolites. [Cancer Res 2007;67(3):881–9]
Distinct Genetic Signatures among Pilocytic Astrocytomas Relate to Their Brain Region Origin
Authors: Sharma Mukesh K. , Mansur David B. , Reifenberger Guido , Perry Arie , Leonard Jeffrey R. , Aldape Kenneth D. , Albin Meredith G. , Emnett Ryan J. , Loeser Simon , Watson Mark A. , Nagarajan Rakesh , Gutmann David H.
Published: February 2007

Pilocytic astrocytomas (PAs) are the most common glioma in children. Whereas many PAs are slow-growing or clinically indolent, others exhibit more aggressive features with tumor recurrence and death. To identify genetic signatures that might predict PA clinical behavior, we did gene expression profiling on 41 primary PAs arising sporadically and in patients with neurofibromatosis type 1 (NF1). Whereas no expression signature was found that could discriminate clinically aggressive or recurrent tumors from more indolent cases, PAs arising in patients with NF1 did exhibit a unique gene expression pattern. In addition, we identified a gene expression signature that stratified PAs by location (supratentorial versus infratentorial). Lastly, we also identified a gene expression pattern common to PAs and normal mouse astrocytes and neural stem cells from these distinct brain regions as well as a gene expression pattern shared between PAs and another human glial tumor (ependymoma) arising supratentorially compared with those originating in the posterior fossa. These results suggest that glial tumors share an intrinsic, lineage-specific molecular signature that reflects the brain region in which their nonmalignant predecessors originated. [Cancer Res 2007;67(3):890–900]
In vivo Real-time Tracking of Single Quantum Dots Conjugated with Monoclonal Anti-HER2 Antibody in Tumors of Mice
Authors: Tada Hiroshi , Higuchi Hideo , Wanatabe Tomonobu M. , Ohuchi Noriaki
Published: February 2007

Studies with tracking of single nanoparticles are providing new insights into the interactions and processes involved in the transport of drug carriers in living mice. Here, we report the tracking of a single particle quantum dot (Qdot) conjugated with tumor-targeting antibody in tumors of living mice using a dorsal skinfold chamber and a high-speed confocal microscope with a high-sensitivity camera. Qdot labeled with the monoclonal anti-HER2 antibody was injected into mice with HER2-overexpressing breast cancer to analyze the molecular processes of its mechanistic delivery to the tumor. Movement of single complexes of the Qdot-antibody could be clearly observed at 30 frames/s inside the tumor through a dorsal skinfold chamber. We successfully identified six processes of delivery: initially in the circulation within a blood vessel, during extravasation, in the extracelullar region, binding to HER2 on the cell membrane, moving from the cell membrane to the perinuclear region, and in the perinuclear region. The six processes were quantitatively analyzed to understand the rate-limiting constraints on Qdot-antibody delivery. The movement of the complexes at each stage was “stop-and-go.” The image analysis of the delivery processes of single particles in vivo provides valuable information on antibody-conjugated therapeutic nanoparticles, which will be useful in increasing therapeutic efficacy. [Cancer Res 2007;67(3):1138–44]
Impaired Dihydrotestosterone Catabolism in Human Prostate Cancer: Critical Role of AKR1C2 as a Pre-Receptor Regulator of Androgen Receptor Signaling
Authors: Ji Qing , Chang Lilly , Stanczyk Frank Z. , Ookhtens Murad , Sherrod Andy , Stolz Andrew
Published: February 2007

We previously reported the selective loss of AKR1C2 and AKR1C1 in prostate cancers compared with their expression in paired benign tissues. We now report that dihydrotestosterone (DHT) levels are significantly greater in prostate cancer tumors compared with their paired benign tissues. Decreased catabolism seems to account for the increased DHT levels as expression of AKR1C2 and SRD5A2 was reduced in these tumors compared with their paired benign tissues. After 4 h of incubation with benign tissue samples, 3H-DHT was predominately catabolized to the 5α-androstane-3α,17β-diol metabolite. Reduced capacity to metabolize DHT was observed in tumor samples from four of five freshly isolated pairs of tissue samples, which paralleled loss of AKR1C2 and AKR1C1 expression. LAPC-4 cells transiently transfected with AKR1C1 and AKR1C2, but not AKR1C3, were able to significantly inhibit a dose-dependent, DHT-stimulated proliferation, which was associated with a significant reduction in the concentration of DHT remaining in the media. R1881-stimulated proliferation was equivalent in all transfected cells, showing that metabolism of DHT was responsible for the inhibition of proliferation. PC-3 cells overexpressing AKR1C2 and, to a lesser extent, AKR1C1 were able to significantly inhibit DHT-dependent androgen receptor reporter activity, which was abrogated by increasing DHT levels. We speculate that selective loss of AKR1C2 in prostate cancer promotes clonal expansion of tumor cells by enhancement of androgen-dependent cellular proliferation by reducing DHT metabolism. [Cancer Res 2007;67(3):1361–9]
Insulin-like Growth Factor-I Receptor Signaling Blockade Combined with Radiation
Authors: Allen Gregory W. , Saba Corey , Armstrong Eric A. , Huang Shyh-Min , Benavente Sergio , Ludwig Dale L. , Hicklin Daniel J. , Harari Paul M.
Published: February 2007

Signaling through the insulin-like growth factor-I receptor (IGF-IR) is implicated in cellular proliferation, apoptosis, carcinogenesis, metastasis, and resistance to cytotoxic cancer therapies. Targeted disruption of IGF-IR signaling combined with cytotoxic therapy may therefore yield improved anticancer efficacy over conventional treatments alone. In this study, a fully human anti–IGF-IR monoclonal antibody A12 (ImClone Systems, Inc., New York, NY) is examined as an adjunct to radiation therapy. IGF-IR expression is shown for a diverse cohort of cell lines, whereas targeted IGF-IR blockade by A12 inhibits IGF-IR phosphorylation and activation of the downstream effectors Akt and mitogen-activated protein kinase. Anchorage-dependent proliferation and xenograft growth is inhibited by A12 in a dose-dependent manner, particularly for non–small cell lung cancer lines. Clonogenic radiation survival of H226 and H460 cells grown under anchorage-dependent conditions is impaired by A12, demonstrating a radiation dose-enhancing effect for IGF-IR blockade. Postradiation anchorage-independent colony formation is inhibited by A12 in A549 and H460 cells. In the H460 xenograft model, combining A12 and radiation significantly enhances antitumor efficacy compared with either modality alone. These effects may be mediated by promotion of radiation-induced, double-stranded DNA damage and apoptosis as observed in cell culture. In summary, these results validate IGF-IR signal transduction blockade as a promising strategy to improve radiation therapy efficacy in human tumors, forming a basis for future clinical trials. [Cancer Res 2007;67(3):1155–62]
Development of Rituximab-Resistant Lymphoma Clones with Altered Cell Signaling and Cross-Resistance to Chemotherapy
Authors: Jazirehi Ali R. , Vega Mario I. , Bonavida Benjamin
Published: February 2007

Immunotherapy with rituximab (chimeric anti-CD20 monoclonal antibody, Rituxan), alone or in conjunction with chemotherapy, has significantly improved the treatment outcome of lymphoma patients. Via an elusive mechanism, a subpopulation of patients becomes unresponsive and/or relapses. To recapitulate various aspects of acquired resistance, rituximab-resistant (RR) clones were established from lymphoma lines and compared with parental cells. Surface CD20 expression was diminished in the clones. The clones neither responded to rituximab-mediated growth reduction or complement-dependent cytotoxicity nor underwent apoptosis in response to cross-linked rituximab. Rituximab failed to chemosensitize the RR clones, which exhibited constitutive hyperactivation of the nuclear factor-κB and extracellular signal-regulated kinase 1/2 pathways, leading to overexpression of B-cell lymphoma protein 2 (Bcl-2), Bcl-2–related gene (long alternatively spliced variant of Bcl-x gene), and myeloid cell differentiation 1 and higher drug resistance. Unlike parental cells, rituximab neither inhibited the activity of these pathways nor diminished the expression of resistant factors. Pharmacologic inhibitors of the survival pathways or Bcl-2 family members reduced the activity of these pathways, diminished antiapoptotic protein expression, and chemosensitized the RR clones. These novel in vitro results denote that continuous long-term rituximab exposure culminates in RR clones that do not respond to rituximab-mediated effects, have altered cellular signaling dynamics, and exhibit different genetic and phenotypic properties compared with parental cells. The data also reveal that although RR clones exhibit higher resistance to rituximab and cytotoxic drugs, these clones can be chemosensitized following treatment with pharmacologic inhibitors (e.g., dehydroxymethylepoxyquinomicin, bortezomib, PD098059) that target survival/antiapoptotic pathways. The findings also identify intracellular targets for potential molecular therapeutic intervention to increase treatment efficacy. The significance and potential clinical relevance of the findings are discussed. [Cancer Res 2007;67(3):1270–81]
A Single-Chain Fv Diabody against Human Leukocyte Antigen-A Molecules Specifically Induces Myeloma Cell Death in the Bone Marrow Environment
Authors: Sekimoto Etsuko , Ozaki Shuji , Ohshima Takashi , Shibata Hironobu , Hashimoto Toshihiro , Abe Masahiro , Kimura Naoki , Hattori Kunihiro , Kawai Shigeto , Kinoshita Yasuko , Yamada-Okabe Hisafumi , Tsuchiya Masayuki , Matsumoto Toshio
Published: February 2007

Cross-linked human leukocyte antigen (HLA) class I molecules have been shown to mediate cell death in neoplastic lymphoid cells. However, clinical application of an anti-HLA class I antibody is limited by possible side effects due to widespread expression of HLA class I molecules in normal tissues. To reduce the unwanted Fc-mediated functions of the therapeutic antibody, we have developed a recombinant single-chain Fv diabody (2D7-DB) specific to the α2 domain of HLA-A. Here, we show that 2D7-DB specifically induces multiple myeloma cell death in the bone marrow environment. Both multiple myeloma cell lines and primary multiple myeloma cells expressed HLA-A at higher levels than normal myeloid cells, lymphocytes, or hematopoietic stem cells. 2D7-DB rapidly induced Rho activation and robust actin aggregation that led to caspase-independent death in multiple myeloma cells. This cell death was completely blocked by Rho GTPase inhibitors, suggesting that Rho-induced actin aggregation is crucial for mediating multiple myeloma cell death. Conversely, 2D7-DB neither triggered Rho-mediated actin aggregation nor induced cell death in normal bone marrow cells despite the expression of HLA-A. Treatment with IFNs, melphalan, or bortezomib enhanced multiple myeloma cell death induced by 2D7-DB. Furthermore, administration of 2D7-DB resulted in significant tumor regression in a xenograft model of human multiple myeloma. These results indicate that 2D7-DB acts on multiple myeloma cells differently from other bone marrow cells and thus provide the basis for a novel HLA class I–targeting therapy against multiple myeloma. [Cancer Res 2007;67(3):1184–92]
Evidence that Tumor Necrosis Factor–Related Apoptosis-Inducing Ligand Induction by 5-Aza-2′-Deoxycytidine Sensitizes Human Breast Cancer Cells to Adriamycin
Authors: Xu Jing , Zhou Jun-Ying , Tainsky Michael A. , Wu Gen Sheng
Published: February 2007

The DNA methyltransferase inhibitor 5-aza-2′-deoxycytidine (5-aza-CdR) inhibits DNA methyltransferase activity and sensitizes cancer cells to chemotherapy, but the mechanisms of its sensitization are not fully understood. Here, we show that 5-aza-CdR induces tumor necrosis factor–related apoptosis-inducing ligand (TRAIL) in the human breast cancer MDA-231 cells. Induction of TRAIL by 5-aza-CdR correlated with inactivation of Akt. Furthermore, we show that overexpression of the active form of Akt by adenovirus infection or inhibition of the Akt downstream target glycogen synthase kinase 3 by its pharmacologic inhibitors abolishes TRAIL induction by 5-aza-CdR. Importantly, we show that the combined treatment of breast cancer cells with 5-aza-CdR and Adriamycin significantly increases apoptotic cell death compared with the treatment with either agent alone. Moreover, the combined treatment activated both death receptor and mitochondrial apoptotic pathways, whereas Adriamycin alone activated only the mitochondrial pathway while 5-aza-CdR failed to activate either. More importantly, down-regulation of TRAIL by small interference RNA silencing decreased 5-aza-CdR–mediated Adriamycin-induced caspase activation and apoptosis, thus conferring Adriamycin resistance. Taken together, our results suggest that induction of TRAIL by 5-aza-CdR is critical for enhancing chemosensitivity of breast cancer cells to Adriamycin. [Cancer Res 2007;67(3):1203–11]
A Small-Molecule Enhancer of Signal Transducer and Activator of Transcription 1 Transcriptional Activity Accentuates the Antiproliferative Effects of IFN-γ in Human Cancer Cells
Authors: Lynch Rebecca A. , Etchin Julia , Battle Traci E. , Frank David A.
Published: February 2007

The transcription factor signal transducer and activator of transcription (STAT) 1 can mediate antiproliferative and proapoptotic effects in cancer cells, and a number of mechanisms have been found whereby STAT1 signaling is attenuated in tumors thereby increasing their malignant behavior. Thus, enhancing gene transcription mediated by STAT1 may be an effective approach to cancer therapy. A high-throughput screen was developed to identify molecules that could enhance STAT1-dependent gene expression. Through this approach, it was found that 2-(1,8-naphthyridin-2-yl)phenol (2-NP) caused a 2-fold increase in STAT1-dependent reporter gene expression compared with that seen with maximally effective concentrations of IFN-γ alone. This effect was specific to STAT1 because 2-NP had no effect on unrelated transcription factors such as nuclear factor (NF) κB or the highly homologous transcription factor STAT3. STAT1-dependent gene activation was enhanced by this compound in a variety of human and murine cell lines and was independent of the stimulus used. Furthermore, 2-NP enhanced the expression of the bona fide endogenous STAT1 target gene interferon regulatory factor 1. 2-NP increased the duration of STAT1 tyrosine phosphorylation in response to IFN-γ, and this may underlie its enhancement of STAT1-dependent transcription. Reflecting the fact that STAT1 can exert tumor-suppressive effects, 2-NP enhanced the ability of IFN-γ to inhibit the proliferation of human breast cancer and fibrosarcoma cells. Tumor cells lacking STAT1 were unaffected by either IFN-γ or 2-NP. These findings indicate that enhancement of STAT1 transcriptional activity may have utility in anticancer therapies, and that cell-based screens for modulators of transcription factor function can be a useful approach for drug discovery. [Cancer Res 2007;67(3):1254–61]
Interleukin-1β–Driven Inflammation Promotes the Development and Invasiveness of Chemical Carcinogen–Induced Tumors
Authors: Krelin Yakov , Voronov Elena , Dotan Shahar , Elkabets Moshe , Reich Eli , Fogel Mina , Huszar Monika , Iwakura Yoichiro , Segal Shraga , Dinarello Charles A. , Apte Ron N.
Published: February 2007

The role of microenvironment interleukin 1 (IL-1) on 3-methylcholanthrene (3-MCA)–induced carcinogenesis was assessed in IL-1–deficient mice, i.e., IL-1β−/−, IL-1α−/−, IL-1α/β−/− (double knockout), and mice deficient in the naturally occurring inhibitor of IL-1, the IL-1 receptor antagonist (IL-1Ra). Tumors developed in all wild-type (WT) mice, whereas in IL-1β–deficient mice, tumors developed slower and only in some of the mice. In IL-1Ra–deficient mice, tumor development was the most rapid. Tumor incidence was similar in WT and IL-1α–deficient mice. Histologic analyses revealed fibrotic structures forming a capsule surrounding droplets of the carcinogen in olive oil, resembling foreign body–like granulomas, which appeared 10 days after injection of 3-MCA and persisted until the development of local tumors. A sparse leukocyte infiltrate was found at the site of carcinogen injection in IL-1β–deficient mice, whereas in IL-1Ra–deficient mice, a dense neutrophilic infiltrate was observed. Treatment of IL-1Ra–deficient mice with recombinant IL-1Ra but not with an inhibitor of tumor necrosis factor abrogated the early leukocytic infiltrate. The late leukocyte infiltrate (day 70), which was dominated by macrophages, was also apparent in WT and IL-1α–deficient mice, but was nearly absent in IL-1β–deficient mice. Fibrosarcoma cell lines, established from 3-MCA–induced tumors from IL-1Ra–deficient mice, were more aggressive and metastatic than lines from WT mice; cell lines from IL-1–deficient mice were the least invasive. These observations show the crucial role of microenvironment-derived IL-1β, rather than IL-1α, in chemical carcinogenesis and in determining the invasive potential of malignant cells. [Cancer Res 2007;67(3):1062–71]
Protein Phosphatase-1α Regulates Centrosome Splitting through Nek2
Authors: Mi Jun , Guo Changyue , Brautigan David L. , Larner James M.
Published: February 2007

ATM is a central mediator of the cellular response to the DNA damage produced by ionizing radiation. We recently showed that protein phosphatase 1 (PP1) is activated by ATM. Because Nek2 is activated by autophosphorylation, and because its dephosphorylation is catalyzed by PP1, we asked if the radiation damage signal to Nek2 was mediated by PP1. Overexpression of Nek2 induces premature centrosome splitting probably by phosphorylating centrosome cohesion proteins C-Nap1 and Rootletin. In this study, we show isoform specificity of PP1 binding and regulation of Nek2. Although both PP1α and PP1γ coimmunoprecipitated with Nek2, only PP1α regulated Nek2 function. Ionizing radiation inhibited Nek2 activity, and this response was dependent on ATM and on PP1 binding to Nek2 and coincident with Thr320 dephosphorylation of PP1. Radiation-induced inhibition of centrosome splitting was abrogated in cells expressing Nek2 mutated in the PP1-binding motif outside the kinase domain. Conversely, cells depleted of PP1α by small interfering RNA showed enhanced centrosome splitting and loss of radiation-induced inhibition of centrosome splitting. The identification of a PP1-specific isoform mediating a checkpoint response opens up the possibility of selectively targeting phosphatases as novel radiation sensitizers. [Cancer Res 2007;67(3):1082–9]
Sensitizing Effect of Galectin-7 in Urothelial Cancer to Cisplatin through the Accumulation of Intracellular Reactive Oxygen Species
Authors: Matsui Yoshiyuki , Ueda Shugo , Watanabe Jun , Kuwabara Ichiro , Ogawa Osamu , Nishiyama Hiroyuki
Published: February 2007

To improve chemotherapeutic efficacy in urothelial cancer, it is important to identify predictive markers for chemosensitivity as well as possible molecules accelerating cell killing mechanisms. In this study, we assessed the possibility of galectin-7 to accelerate cis-diamminedichloroplatinum (CDDP)–induced cell killing in vitro and also to predict chemosensitivity against CDDP in urothelial cancer patients. The expression of galectin-7 was analyzed in five bladder cancer cell lines with different p53 status after treatment with CDDP. The roles of galectin-7 in chemosensitivity against CDDP were analyzed by transfection of the galectin-7 gene into several of these cell lines. Furthermore, the relationship between the expression of galectin-7 and the response to neoadjuvant chemotherapy was analyzed in 17 human bladder cancer specimens. Exposure to CDDP induced galectin-7 in cell lines with wild-type p53 but not in those with mutated p53. When the galectin-7 gene was transfected into cell lines with mutated p53, the sensitivity to CDDP increased compared with control transfectants. In addition, galectin-7–transfected cells exhibited more accumulation of intracellular reactive oxygen species and activation of c-Jun NH2-terminal kinase (JNK) and Bax than control transfectants. SP600125, an inhibitor of JNK, or antioxidant N-acetyl-l-cysteine inhibited the enhancement of chemosensitivity against CDDP by galectin-7 transfection. In clinical samples, the expression levels of galectin-7 were significantly lower in urothelial carcinomas compared with normal urothelium. When chemosensitivity was tested, its expression levels were higher in the chemosensitive group than in the chemoresistant group. Galectin-7 is a candidate for a predictive marker of chemosensitivity against CDDP, and the targeted expression of galectin-7 might overcome the chemoresistance of urothelial cancer. [Cancer Res 2007;67(3):1212–20]
Protein Kinase Cε, which Sensitizes Skin to Sun's UV Radiation–Induced Cutaneous Damage and Development of Squamous Cell Carcinomas, Associates with Stat3
Authors: Aziz Moammir H. , Manoharan Herbert T. , Verma Ajit K.
Published: February 2007

Chronic exposure to UV radiation (UVR) is the major etiologic factor in the development of human skin cancers including squamous cell carcinoma (SCC). We have shown that protein kinase Cε (PKCε), a Ca2+-independent, phospholipid-dependent serine/threonine kinase, is an endogenous photosensitizer. PKCε is among the six isoforms (α, δ, ε, η, μ, and ζ) expressed in both mouse and human skin. PKCε transgenic mice, which overexpress PKCε in the basal epidermal cells and cells of the hair follicle, are highly sensitive to UVR-induced cutaneous damage and development of SCC. We now present that PKCε-overexpressing, but not PKCδ-overexpressing, transgenic mice, when exposed to a single (4 kJ/m2) or repeated (four doses, 2 kJ/m2/dose, thrice weekly) UVR, emitted by Kodacel-filtered FS-40 sun lamps, elicit constitutive phosphorylation of signal transducers and activators of transcription 3 (Stat3) at both Tyr705 and Ser727 residues. UVR-induced phosphorylation of Stat3 accompanied increased expression of Stat3-regulated genes (c-myc, cyclin D1, cdc25A, and COX-2). In reciprocal immunoprecipitation/blotting experiments, phosphorylated Stat3 coimmunoprecipitated with PKCε. As observed in vivo using PKCε knockout mice and in vitro in an immunocomplex kinase assay, PKCε phosphorylated Stat3 at Ser727 residue. These results indicate for the first time that (a) PKCε is a Stat3Ser727 kinase; (b) PKCε-mediated phosphorylation of StatSer727 may be essential for transcriptional activity of Stat3; and (c) UVR-induced phosphorylation of Ser727 may be a key component of the mechanism by which PKCε imparts sensitivity to UVR-induced development of SCC. [Cancer Res 2007;67(3):1385–94]
Erlotinib Directly Inhibits HER2 Kinase Activation and Downstream Signaling Events in Intact Cells Lacking Epidermal Growth Factor Receptor Expression
Authors: Schaefer Gabriele , Shao Lihua , Totpal Klara , Akita Robert W.
Published: February 2007

Erlotinib (Tarceva), is an orally available, reversible inhibitor of epidermal growth factor receptor (EGFR; HER1) that exhibits inhibitory activity on purified HER2 kinase at much higher concentrations. Despite the minimal activity on purified protein in vitro, in vivo studies show that erlotinib inhibits the growth of HER2-driven systems effectively. Several hypotheses have been put forward to explain this discrepancy. In particular, it has been suggested that erlotinib might indirectly suppress the activity of HER2 by blocking the ability of EGFR to transactivate it when the two receptors are part of a heterodimer complex. However, an alternative possibility that has not been adequately addressed is whether the direct inhibitory action of erlotinib on the HER2 kinase might account for the observed biological responses. To distinguish between a direct effect of erlotinib on HER2 kinase in intact cells or an indirect effect of erlotinib on HER2 activity that is mediated through EGFR, we generated cell lines that express either EGFR-H2 chimeric receptor or HER2 and HER3 receptors in an EGFR-negative background. We show that dose-dependent inhibition of HER2 was achieved at the receptor level, on downstream signaling molecules, and more importantly was also translated into inhibition of cell growth. Our findings imply that the inhibitory effect of erlotinib in HER2-expressing cells may in part be mediated through direct interaction with HER2 rather than indirectly through a process that requires the presence of EGFR. [Cancer Res 2007;67(3):1228–38]
The Tumor Suppressor Activity of the Lysyl Oxidase Propeptide Reverses the Invasive Phenotype of Her-2/neu–Driven Breast Cancer
Authors: Min Chengyin , Kirsch Kathrin H. , Zhao Yingshe , Jeay Sébastien , Palamakumbura Amitha H. , Trackman Philip C. , Sonenshein Gail E.
Published: February 2007

Expression of the lysyl oxidase gene (LOX) was found to inhibit the transforming activity of the ras oncogene in NIH 3T3 fibroblasts and was hence named the ras recision gene (rrg). Lysyl oxidase (LOX) is synthesized and secreted as a 50-kDa inactive proenzyme (Pro-LOX), which is processed by proteolytic cleavage to a functional 32-kDa enzyme and an 18-kDa propeptide (LOX-PP). Recently, the ras recision activity of the LOX gene in NIH 3T3 cells was mapped to its propeptide region. Here, we show for the first time that LOX-PP inhibits transformation of breast cancer cells driven by Her-2/neu, an upstream activator of Ras. LOX-PP expression in Her-2/neu–driven breast cancer cells in culture suppressed Akt, extracellular signal–regulated kinase, and nuclear factor-κB activation. Her-2/neu–induced epithelial to mesenchymal transition was reverted by LOX-PP, as judged by reduced levels of Snail and vimentin; up-regulation of E-cadherin, γ-catenin, and estrogen receptor α; and decreased ability to migrate or to form branching colonies in Matrigel. Furthermore, LOX-PP inhibited Her-2/neu tumor formation in a nude mouse xenograft model. Thus, LOX-PP inhibits signaling cascades induced by Her-2/neu that promote a more invasive phenotype and may provide a novel avenue for treatment of Her-2/neu–driven breast carcinomas. [Cancer Res 2007;67(3):1105–12]
Intermittent Hypoxia Furthers the Rationale for Hypoxia-Inducible Factor-1 Targeting
Authors: Dewhirst Mark W.
Published: February 2007

Hypoxia-inducible factor-1 (HIF-1) stabilization is a pivotal event in the response to hypoxic stress. A study in the December 15, 2006 issue of Cancer Research shows that HIF-1 stabilization occurs more robustly as a result of intermittent hypoxia compared with chronic hypoxia. The findings of this study suggest that intermittent hypoxia might influence the efficacy of radiotherapy by more strongly affecting the growth and survival of vascular endothelial cells. This finding offers additional encouragement to efforts to target HIF-1 for cancer therapy. [Cancer Res 2007;67(3):854–5]
Experience and Cultural Models Matter: Placing Firm Limits on Childhood Anthropocentrism
Authors: Waxman Sandra , Medin Douglas
Published: February 2007

This paper builds on Hatano and Inagaki’s pioneering work on the role of experience and cultural models in children’s biological reasoning. We use a category-based induction task to consider how experience and cultural models shape rural and urban children’s patterns of biological reasoning. We discuss the implications of these findings for developmental theory and educational practice.
Conceptual Growth in Children and in the Learning Sciences: Giyoo Hatano’s Contributions
Authors: Greeno James G. , Saxe Geoffrey B.
Published: February 2007

In Giyoo Hatano’s passing, we have lost an esteemed colleague and a treasured friend. Among his many contributions to our field, our work, and our lives, we honor and build on his and his colleagues’ work on conceptual growth. We liken the view developed by Hatano and his colleagues to Toulmin’s evolutionary scheme for understanding conceptual growth in intellectual disciplines, and we briefly sketch recent studies and discussions in which our methods and conceptualizations depend upon and draw meaning from Hatano’s work, findings, and insights. He contributed strongly to the evolutionary growth of conceptual understanding in our scientific field, and he was an important hub of communication and interaction among scientists around the world. We are grateful to have had the chance to interact and learn with Giyoo Hatano, and we will gratefully continue our interaction with his ideas, his contributions, and the relations that he forged across national boundaries.