bluuebunny/crossref_metadata_2025 huggingface dataset:
Endothelial nitric oxide synthase is dynamically expressed during bone marrow stem cell differentiation into endothelial cells
Authors: Liu Zhenguo , Jiang Yuehua , Hao Hong , Gupta Kalpna , Xu Jian , Chu Ling , McFalls Edward , Zweier Jay , Verfaillie Catherine , Bache Robert J.
Published: June 2007

This study was designed to investigate the developmental expression of endothelial nitric oxide synthase (eNOS) during stem cell differentiation into endothelial cells and to examine the functional status of the newly differentiated endothelial cells. Mouse adult multipotent progenitor cells (MAPCs) were used as the source of stem cells and were induced to differentiate into endothelial cells with vascular endothelial growth factor (VEGF) in serum-free medium. Expression of eNOS in the cells during differentiation was evaluated with real-time PCR, nitric oxide synthase (NOS) activity, and Western blot analysis. It was found that eNOS, but no other NOS, was present in undifferentiated MAPCs. eNOS expression disappeared in the cells immediately after induction of differentiation. However, eNOS expression reoccurred at day 7 during differentiation. Increasing eNOS mRNA, protein content, and activity were observed in the cells at days 14 and 21 during differentiation. The differentiated endothelial cells formed dense capillary networks on growth factor-reduced Matrigel. VEGF-stimulated phosphorylation of extracellular signal-regulated kinase (ERK)-1 and ERK-2 occurred in these cells, which was inhibited by NOS inhibitor NG-nitro-l-arginine methyl ester. In conclusion, these data demonstrate that eNOS is present in MAPCs and is dynamically expressed during the differentiation of MAPCs into endothelial cells in vitro.
Nicotine alters lung branching morphogenesis through the α7 nicotinic acetylcholine receptor
Authors: Wongtrakool Cherry , Roser-Page Susanne , Rivera Hilda N. , Roman Jesse
Published: June 2007

There is abundant epidemiological data linking prenatal environmental tobacco smoke with childhood asthma and wheezing, but the underlying molecular and physiological mechanisms that occur in utero to explain this link remain unelucidated. Several studies suggest that nicotine, which traverses the placenta, is a causative agent. Therefore, we studied the effects of nicotine on lung branching morphogenesis using embryonic murine lung explants. We found that the expression of α7 nicotinic acetylcholine receptors, which mediate many of the biological effects of nicotine, is highest in pseudoglandular stage lungs compared with lungs at later stages. We then studied the effects of nicotine in the explant model and found that nicotine stimulated lung branching in a dose-dependent fashion. α-Bungarotoxin, an antagonist of α7 nicotinic acetylcholine receptors, blocked the stimulatory effect of nicotine, whereas GTS-21, a specific agonist, stimulated branching, thereby mimicking the effects of nicotine. Explants deficient in α7 nicotinic acetylcholine receptors did not respond to nicotine. Nicotine also stimulated the growth of the explant. Altogether, these studies suggest that nicotine stimulates lung branching morphogenesis through α7 nicotinic acetylcholine receptors and may contribute to dysanaptic lung growth, which in turn may predispose the host to airway disease in the postnatal period.
Interleukin-17A modulates human airway epithelial responses to human rhinovirus infection
Authors: Wiehler Shahina , Proud David
Published: June 2007

Human rhinovirus (HRV) infections are associated with exacerbations of asthma and chronic obstructive pulmonary disease that are characterized by a selective neutrophil infiltration. IL-17A, a cytokine derived primarily from activated T cells, has been linked to neutrophilic inflammation of the airways. We hypothesized that IL-17A alters the response of HRV-infected epithelial cells to modulate airway inflammatory cell populations. IL-17A synergistically enhanced HRV-16-induced epithelial production of the neutrophil chemoattractant, IL-8, as well as human β-defensin-2 (HBD-2), a chemoattractant for immature dendritic cells and memory T cells, but suppressed viral production of the eosinophil chemoattractant, RANTES. These effects were not due to alterations of viral uptake or replication by IL-17A. The synergy between HRV-16 and IL-17A for IL-8 protein production was both dose- and time-dependent. IL-8 induction by IL-17A or HRV-16, alone and in combination, was reduced by inhibitors of the p38 and p44/42 MAPK pathways. By contrast, induction of HBD-2 depended on the activation of the p38 and JNK pathways. The ability of IL-17A to synergistically enhance HRV-induced IL-8 is mediated posttranscriptionally, since IL-8 promoter activation by the combination of the two stimuli was merely additive, whereas the combination of IL-17A and HRV-16 led to stabilization of IL-8 mRNA. Similarly, stimulation of HBD-2 promoter constructs by the combination of IL-17A and HRV-16 was no more than the sum of the individual responses. Further studies are needed to examine HBD-2 mRNA stability. Taken together, these data represent the first demonstration that IL-17A can modify epithelial responses to HRV in a manner that would be expected to favor the recruitment of neutrophils, immature dendritic cells, and memory T cells to the airways.
Effects of prior very-heavy intensity exercise on indices of aerobic function and high-intensity exercise tolerance
Authors: Ferguson C. , Whipp B. J. , Cathcart A. J. , Rossiter H. B. , Turner A. P. , Ward S. A.
Published: June 2007

A recent bout of high-intensity exercise can alter the balance of aerobic and anaerobic energy provision during subsequent exercise above the lactate threshold (θL). However, it remains uncertain whether such “priming” influences the tolerable duration of subsequent exercise through changes in the parameters of aerobic function [e.g., θL, maximum oxygen uptake (V̇o2max)] and/or the hyperbolic power-duration (P-t) relationship [critical power (CP) and the curvature constant (W′)]. We therefore studied six men performing cycle ergometry to the limit of tolerance; gas exchange was measured breath-by-breath and arterialized capillary blood [lactate] was measured at designated intervals. On different days, each subject completed 1) an incremental test (15 W/min) for estimation of θL and measurement of the functional gain (ΔV̇o2/ΔWR) and V̇o2peak and 2) four constant-load tests at different work rates (WR) for estimation of CP, W′, and V̇o2max. All tests were subsequently repeated with a preceding 6-min supra-CP priming bout and an intervening 2-min 20-W recovery. The hyperbolicity of the P-t relationship was retained postpriming, with no significant difference in CP (241 ± 39 vs. 242 ± 36 W, post- vs. prepriming), V̇o2max (3.97 ± 0.34 vs. 3.93 ± 0.38 l/min), ΔV̇o2/ΔWR (10.7 ± 0.3 vs. 11.1 ± 0.4 ml·min−1·W−1), or the fundamental V̇o2 time constant (25.6 ± 3.5 vs. 28.3 ± 5.4 s). W′ (10.61 ± 2.07 vs. 16.13 ± 2.33 kJ) and the tolerable duration of supra-CP exercise (−33 ± 11%) were each significantly reduced, despite a less-prominent V̇o2 slow component. These results suggest that, following supra-CP priming, there is either a reduced depletable energy resource or a residual fatigue-metabolite level that leads to the tolerable limit before this resource is fully depleted.
Optimization of a dried residue specimen preparation method for quantifying analytes in plutonium metal using WDXRF
Authors: Worley Christopher G. , Colletti Lisa P.
Published: June 2007

A novel method for preparing thin films was investigated for quantifying gallium and iron in plutonium solutions using WDXRF. This technique was developed to eliminate the potential for radioactive liquid to leak into the spectrometer, decrease specimen preparation time, and minimize waste. Samples were cast in μL quantities onto Kapton, and a surfactant was added to disperse the solution uniformly across the Kapton. After drying the specimens, they were sealed in a cell for analysis. Results to date indicate the method can provide a relative precision of ∼0.5% for gallium and ∼2% for iron, which is more than sufficient for routine sample analyses.
Nondestructive analysis of a painting, a national treasure in Japan
Authors: Hayakawa Yasuhiro , Shirono Seiji , Miura Sadatoshi , Matsushima Tomohide , Uchida Tokugo
Published: June 2007

The painting materials and drawing techniques of a pair of two-panel folding screens entitled Red and White Plum Blossoms by Ogata Korin, a national treasure in Japan, were investigated directly and nondestructively by XRF, X-ray radiography, and high resolution digital imaging. Several assumptions were made about the materials used in the red and white plum trees drawn on the right and left screens, respectively, and the river drawn at the center. By the present investigation, the materials used for the paintings were revealed, and some of them contradicted what have been previously believed.
Human Tumor Xenografts Recurring after Radiotherapy Are More Sensitive to Anti–Vascular Endothelial Growth Factor Receptor-2 Treatment than Treatment-Naive Tumors
Authors: Kozin Sergey V. , Winkler Frank , Garkavtsev Igor , Hicklin Daniel J. , Jain Rakesh K. , Boucher Yves
Published: June 2007

The effects of antiangiogenic therapy on tumors relapsing after irradiation are not known. To this end, we irradiated human tumors growing s.c. in nude mice with a single dose of 20 or 30 Gy. Compared with primary (treatment-naive) xenografts, the growth rate of recurrent tumors was 1.6-fold slower, which is consistent with the known “tumor bed effect.” For similar size tumors, recurrences had fewer functional vessels, a reduced vessel coverage by perivascular cells, and were more necrotic. Placenta growth factor concentration was significantly lower in relapses, whereas vascular endothelial growth factor (VEGF) levels were similar between primary and recurrent tumors. On the other hand, fibrillar collagen deposition was significantly increased in recurrent tumors. This radiation-induced fibrosis was partially responsible for the slower growth of recurrences; the i.t. injection of collagenase increased the growth rate of tumor relapses without affecting primary tumor growth. The mouse-specific VEGF receptor 2–blocking antibody DC101 induced a 2.2-fold longer growth delay in recurrent tumors compared with treatment-naive tumors. DC101 significantly decreased the interstitial fluid pressure and did not change the functional vessel density and perivascular cell coverage in both tumor variants. Interestingly, DC101 induced a rapid (2 days after treatment initiation) and significant decrease in tumor cell proliferation in recurrent but not in primary tumors. Thus, our results show that the stromal compartment and the response to antiangionenic therapy of primary and in-field recurrent tumors are significantly different. Our findings suggest that antiangiogenic agents could be effective in the treatment of patients with relapses after radiotherapy. [Cancer Res 2007;67(11):5076–82]
Mapping Genes that Contribute to Daunorubicin-Induced Cytotoxicity
Authors: Duan Shiwei , Bleibel Wasim K. , Huang Rong Stephanie , Shukla Sunita J. , Wu Xiaolin , Badner Judith A. , Dolan M. Eileen
Published: June 2007

Daunorubicin is an anthracycline antibiotic agent used in the treatment of hematopoietic malignancies. Toxicities associated with this agent include myelosuppression and cardiotoxicity; however, the genes or genetic determinants that contribute to these toxicities are unknown. We present an unbiased genome-wide approach that incorporates heritability, whole-genome linkage analysis, and linkage-directed association to uncover genetic variants contributing to the sensitivity to daunorubicin-induced cytotoxicity. Cell growth inhibition in 324 Centre d' Etude du Polymorphisme Humain lymphoblastoid cell lines (24 pedigrees) was evaluated following treatment with daunorubicin for 72 h. Heritability analysis showed a significant genetic component contributing to the cytotoxic phenotypes (h2 = 0.18–0.63 at 0.0125, 0.025, 0.05, 0.1, 0.2, and 1.0 μmol/L daunorubicin and at the IC50, the dose required to inhibit 50% cell growth). Whole-genome linkage scans at all drug concentrations and IC50 uncovered 11 regions with moderate peak LOD scores (>1.5), including 4q28.2 to 4q32.3 with a maximum LOD score of 3.18. The quantitative transmission disequilibrium tests were done using 31,312 high-frequency single-nucleotide polymorphisms (SNP) located in the 1 LOD confidence interval of these 11 regions. Thirty genes were identified as significantly associated with daunorubicin-induced cytotoxicity (P ≤ 2.0 × 10−4, false discovery rate ≤ 0.1). Pathway and functional gene ontology analysis showed that these genes were overrepresented in the phosphatidylinositol signaling system, axon guidance pathway, and GPI-anchored proteins family. Our findings suggest that a proportion of susceptibility to daunorubicin-induced cytotoxicity may be controlled by genetic determinants and that analysis using linkage-directed association studies with dense SNP markers can be used to identify the genetic variants contributing to cytotoxicity. [Cancer Res 2007;67(11):5425–33]
Reversible Smad-Dependent Signaling between Tumor Suppression and Oncogenesis
Authors: Sekimoto Go , Matsuzaki Koichi , Yoshida Katsunori , Mori Shigeo , Murata Miki , Seki Toshihito , Matsui Hirofumi , Fujisawa Jun-ichi , Okazaki Kazuichi
Published: June 2007

Cancer cells often gain advantage by reducing the tumor-suppressive activity of transforming growth factor-β (TGF-β) together with stimulation of its oncogenic activity as in Ras-transformed cells; however, molecular mechanisms remain largely unknown. TGF-β activates both its type I receptor (TβRI) and c-Jun NH2-terminal kinase (JNK), which phosphorylate Smad2 and Smad3 at the COOH-terminal (pSmad2/3C) and linker regions (pSmad2/3L). Here, we report that Ras transformation suppresses TβRI-mediated pSmad3C signaling, which involves growth inhibition by down-regulating c-Myc. Instead, hyperactive Ras constitutively stimulates JNK-mediated pSmad2/3L signaling, which fosters tumor invasion by up-regulating plasminogen activator inhibitor-1 and matrix metalloproteinase-1 (MMP-1), MMP-2, and MMP-9. Conversely, selective blockade of linker phosphorylation by a mutant Smad3 lacking JNK-dependent phosphorylation sites results in preserved tumor-suppressive function via pSmad3C in Ras-transformed cells while eliminating pSmad2/3L–mediated invasive capacity. Thus, specific inhibition of the JNK/pSmad2/3L pathway should suppress cancer progression by shifting Smad-dependent signaling from oncogenesis to tumor suppression. [Cancer Res 2007;67(11):5090–6]
Retroviral Insertional Mutagenesis Identifies Genes that Collaborate with NUP98-HOXD13 during Leukemic Transformation
Authors: Slape Christopher , Hartung Helge , Lin Ying-Wei , Bies Juraj , Wolff Linda , Aplan Peter D.
Published: June 2007

The t(2;11)(q31;p15) chromosomal translocation results in a fusion between the NUP98 and HOXD13 genes and has been observed in patients with myelodysplastic syndrome (MDS) or acute myelogenous leukemia. We previously showed that expression of the NUP98-HOXD13 (NHD13) fusion gene in transgenic mice results in an invariably fatal MDS; approximately one third of mice die due to complications of severe pancytopenia, and about two thirds progress to a fatal acute leukemia. In the present study, we used retroviral insertional mutagenesis to identify genes that might collaborate with NHD13 as the MDS transformed to an acute leukemia. Newborn NHD13 transgenic mice and littermate controls were infected with the MOL4070LTR retrovirus. The onset of leukemia was accelerated, suggesting a synergistic effect between the NHD13 transgene and the genes neighboring retroviral insertion events. We identified numerous common insertion sites located near protein-coding genes and confirmed dysregulation of a subset of these by expression analyses. Among these genes were Meis1, a known collaborator of HOX and NUP98-HOX fusion genes, and Mn1, a transcriptional coactivator involved in human leukemia through fusion with the TEL gene. Other putative collaborators included Gata2, Erg, and Epor. Of note, we identified a common insertion site that was >100 kb from the nearest coding gene, but within 20 kb of the miR29a/miR29b1 microRNA locus. Both of these miRNA were up-regulated, demonstrating that retroviral insertional mutagenesis can target miRNA loci as well as protein-coding loci. Our data provide new insights into NHD13-mediated leukemogenesis as well as retroviral insertional mutagenesis mechanisms. [Cancer Res 2007;67(11):5148–55]
Somatic Mutations in the Tyrosine Kinase Domain of Epidermal Growth Factor Receptor (EGFR) Abrogate EGFR-Mediated Radioprotection in Non–Small Cell Lung Carcinoma
Authors: Das Amit K. , Chen Benjamin P. , Story Michael D. , Sato Mitsuo , Minna John D. , Chen David J. , Nirodi Chaitanya S.
Published: June 2007

The epidermal growth factor receptor (EGFR) is an important determinant of radioresponse, whose elevated expression and activity frequently correlates with radioresistance in several cancers, including non–small cell lung carcinoma (NSCLC). We reported recently that NSCLC cell lines harboring somatic, activating mutations in the tyrosine kinase domain (TKD) of the EGFR exhibit significant delays in the repair of DNA double-strand breaks (DSB) and poor clonogenic survival in response to radiation. Here, we explore the mechanisms underlying mutant EGFR-associated radiosensitivity. In three representative NSCLC cell lines, we show that, unlike wild-type (WT) EGFR, receptors with common oncogenic TKD mutations, L858R or ΔE746-E750, are defective in radiation-induced translocation to the nucleus and fail to bind the catalytic and regulatory subunits of the DNA-dependent protein kinase (DNA-PK), a key enzyme in the nonhomologous end-joining repair pathway. Moreover, despite the presence of WT EGFR, stable exogenous expression of either the L858R or the ΔE746-E750 mutant forms of EGFR in human bronchial epithelial cells significantly delays repair of ionizing radiation (IR)–induced DSBs, blocks the resolution of frank or microhomologous DNA ends, and abrogates IR-induced nuclear EGFR translocation or binding to DNA-PK catalytic subunit. Our study has identified a subset of naturally occurring EGFR mutations that lack a critical radioprotective function of EGFR, providing valuable insights on how the EGFR mediates cell survival in response to radiation in NSCLC cell lines. [Cancer Res 2007;67(11):5267–74]
Metastatic Potential of 21T Human Breast Cancer Cells Depends on Akt/Protein Kinase B Activation
Authors: Qiao Meng , Iglehart J. Dirk , Pardee Arthur B.
Published: June 2007

Most cancer lethality is caused by metastasis. To gain insight into the molecular basis of tumor progression to metastasis, we used the 21T series of human mammary epithelial cells obtained by successive biopsies from one breast cancer patient. The c-erbB2 gene is amplified and overexpressed in each of three 21T tumor lines. The erbB receptor tyrosine kinase–activated phosphatidylinositol 3-kinase/Akt signaling cascade is crucial for the development and maintenance of epithelial cells, and dysregulation of this pathway is frequently associated with cellular transformation and cancer. For Akt to be fully activated, Ser473 on its COOH terminus needs to be phosphorylated. We detected more Ser473 Akt phosphorylation in MT cells, derived from a pleural effusion, compared with cells from the primary tumor. This phosphorylation has recently been shown to be catalyzed by mammalian target of rapamycin (mTOR)/rictor kinase. By using genetic and pharmacologic activators and inhibitors, we showed that Ser473 Akt phosphorylation is more sensitive to mTOR/rictor inhibition in metastatic tumor cells than normal mammary epithelial and primary tumor cells. The mTOR/rictor kinase activity was indispensable for both Ser473 Akt phosphorylation and migration of metastatic MT2 cells. In addition, a large decrease of protein phosphatase PH domain leucine-rich repeat protein phosphatase (PHLPP) was found, which could be responsible for the overexpression of Ser473 Akt in MT cells. Our data indicate that these breast cancer cells acquire new vulnerabilities, rictor and PHLPP, which might provide an Achilles' heel for therapeutic intervention of breast cancer metastasis. [Cancer Res 2007;67(11):5293–9]
Identification and Characterization of Novel Spliced Variants of Neuregulin 4 in Prostate Cancer
Authors: Hayes Nandini V.L. , Blackburn Edith , Smart Laura V. , Boyle Mary M. , Russell Graham A. , Frost Teresa M. , Morgan Byron J.T. , Baines Anthony J. , Gullick William J.
Published: June 2007

Purpose: The neuregulin (NRG) 1, 2, and 3 genes undergo extensive alternative mRNA splicing, which results in variants that show structural and functional diversity. The aims of this study were to establish whether the fourth member of this family, NRG4, is expressed in prostate cancer, if it is alternatively spliced and whether any functional differences between the variants could be observed.

Experimental Design: The expression of NRG4 was determined using immunohistochemical staining of 40 cases of primary prostate cancer. Bioinformatic analysis and reverse transcription-PCR (RT-PCR) using NRG4 isotype-specific primers on a panel of normal and prostate cancer cell lines were used to identify alternatively spliced NRG4 variants. Expression of these variants was determined using isotype-specific antibodies. Transfection into Cos-7 cells of two of these green fluorescent protein-tagged variants allowed analysis of their subcellular location. Four of the variants were chemically synthesized and tested for their ability to activate the ErbB4 receptor.

Results: NRG4 was variably expressed in the cytoplasm in the majority of prostate cancer cases, and in a subset of cases in the membrane, high levels were associated with advanced disease stage. Four novel NRG4 splice variants (NRGA2, NRG4 B1-3) were characterized, where each seemed to have a different subcellular location and were also expressed in the cytoplasm of the prostate tumors. NRG4 B3 was also present in endothelial cells. In transfected cells, the A type variant (NRG4 A1) was localized to the membrane, whereas the B type variant (NRG4 B1), which lacks the predicted transmembrane region, had an intracellular localization. Only the variants with an intact epidermal growth factor–like domain activated ErbB4 signaling.

Conclusion: NRG4 overexpression is associated with advanced-stage prostate cancer. The alternative splice variants may have different roles in cell signaling, some acting as classic receptor ligands and some with as-yet unknown functions.
Inhibition of p53-Murine Double Minute 2 Interaction by Nutlin-3A Stabilizes p53 and Induces Cell Cycle Arrest and Apoptosis in Hodgkin Lymphoma
Authors: Drakos Elias , Thomaides Athanasios , Medeiros L. Jeffrey , Li Jiang , Leventaki Vasiliki , Konopleva Marina , Andreeff Michael , Rassidakis George Z.
Published: June 2007

Purpose: p53 is frequently expressed but rarely mutated in Hodgkin and Reed-Sternberg (HRS) cells of Hodgkin's lymphoma (HL). p53 protein levels are regulated by murine double minute 2 (MDM2) through a well-established autoregulatory feedback loop. In this study, we investigated the effects of nutlin-3A, a recently developed small molecule that antagonizes MDM2 and disrupts the p53-MDM2 interaction, on p53-dependent cell cycle arrest and apoptosis in cultured HRS cells.

Experimental Design: HL cell lines carrying wild-type (wt) or mutated p53 gene were treated with the potent MDM2 inhibitor nutlin-3A or a 150-fold less active enantiomer, nutlin-3B.

Results: We show that nutlin-3A, but not nutlin-3B, stabilizes p53 in cultured HRS cells carrying wt p53 gene resulting in p53-dependent cell cycle arrest and apoptosis. Cell cycle arrest was associated with up-regulation of the cyclin-dependent kinase inhibitor p21. Nutlin-3A–induced apoptotic cell death was accompanied by Bax and Puma up-regulation and caspase-3 cleavage and was abrogated, in part, by inhibition of caspase-9 and caspase-3 activity. By contrast, no effects on cell cycle or apoptosis were found in HL cell lines harboring mutated p53 gene. Furthermore, combined treatment with nutlin-3A and doxorubicin revealed enhanced cytotoxicity in HRS cells with wt p53 gene. Blocking of nuclear export by leptomycin B, or inhibition of proteasome by MG132, stabilized p53 at a level comparable with that of nutlin-3A treatment in HRS cells with wt p53.

Conclusions: These data suggest that nutlin-3A stabilized p53 by preventing MDM2-mediated p53 degradation in HRS cells. wt p53 stabilization and activation by nutlin-3A may be a novel therapeutic approach for patients with HL.
Insulin-Like Growth Factor-I Produced by Seminal Vesicles: Relationship to Intraepithelial Basal Cell Hyperplasia in the Prostate
Authors: Garcia Fernando U. , Urbanska Katarzyna , Koltowski Lauren , Reiss Krzysztof , Sell Christian
Published: June 2007

Purpose: This study examined the seminal vesicle fluid (SVF) as a potential local source of insulin-like growth factor-I (IGF-I) in the peripheral zone of the prostate.

Experimental Design: IGF-I levels in seminal fluid were measured. The levels of the IGF-I receptor (IGF-IR) in its active, phosphorylated form as well as direct downstream targets were examined in the peripheral zone of the prostate.

Results: In situ, we find that the IGF-IR is activated in the peripheral zone in areas of atrophy, prostatic intraepithelial hyperplasia, and cancer. In addition, immunostaining reveals preferential activation of the IGF-IR in p63-positive cells in areas of intermediate basal cell hyperplasia in the peripheral zone, indicating that prostate progenitor cells are highly sensitive to increases in local IGF-I levels. These areas of basal cell hyperplasia occur at high incidence in the peripheral zone of the prostate. Relatively high levels of IGF-I were identified in SVF. In addition, we find that SVF can stimulate the proliferation of both normal and cancer-derived prostate cells.

Conclusions: These results suggest that SVF is a local source of IGF-I that provides chronic stimulation of prostate cells. This chronic stimulation could contribute to the development of prostate cancer in older men.
Changes of Biochemical Markers of Bone Turnover and YKL-40 Following Hormonal Treatment for Metastatic Prostate Cancer Are Related to Survival
Authors: Johansen Julia S. , Brasso Klaus , Iversen Peter , Teisner Børge , Garnero Patrick , Price Paul A. , Christensen Ib Jarle
Published: June 2007

Purpose: Elevated serum levels of biochemical markers of bone turnover and YKL-40 in patients with metastatic prostate cancer (PC) at the time of diagnosis are associated to poor prognosis. In this study, we evaluated the value of these biomarkers in monitoring the patients during hormonal treatment.

Experimental Design: Serum procollagen type I N-terminal propeptide (PINP), bone-specific alkaline phosphatase (BAP), CTX-I, and YKL-40 were determined by ELISA in a longitudinal study of 106 patients with metastatic PC during treatment with total androgen ablation or parenteral estrogen. Serum samples were collected with 3 months interval. Median observation time was 4.9 years (range, 3.6-6.2). A total of 78 patients died (64 within 7 months following the last blood sampling).

Results: After 6 months treatment, serum PINP, BAP, and YKL-40 decreased (P < 0.0001), but not serum CTX-I compared with baseline values. Univariate Cox analysis showed that serum PINP at 6 months [log transformed and treated as a continuous variable; hazard ratio (HR), 2.2; P < 0.0001], serum BAP (HR, 1.8; P < 0.0001), and serum CTX-I (HR, 2.4; P < 0.0001), but not serum YKL-40 (HR, 1.4; P = 0.16) were associated with survival. Multivariate Cox analysis including the biomarkers 6 months after the start of treatment showed that Soloway score (HR, 3.9; P = 0.013), WHO tumor grade (HR, 3.9; P = 0.004), and serum PINP (HR, 2.2; P < 0.0001) were independent prognostic variables of survival. Scoring the biomarkers during treatment as time-dependent covariates in univariate Cox regression analysis showed that increases in serum PINP (HR, 2.0; P < 0.0001), BAP (HR, 2.1; P < 0.0001), and YKL-40 (HR, 2.1; P < 0.0001) were predictors of early death.

Conclusions: Serial monitoring of serum PINP, BAP, CTX-I, and YKL-40 in metastatic PC patients during hormonal treatment provided information of prognosis.
Nuclear Factor-κB–Related Serum Factors as Longitudinal Biomarkers of Response and Survival in Advanced Oropharyngeal Carcinoma
Authors: Allen Clint , Duffy Sonia , Teknos Theodoros , Islam Mozaffarul , Chen Zhong , Albert Paul S. , Wolf Gregory , Van Waes Carter
Published: June 2007

Purpose: Cytokines and growth factors modulated by transcription factor nuclear factor-κB and secreted by tumor and stromal cells are detectable in serum of patients with advanced cancers, including head and neck squamous cell carcinomas (SCC). Longitudinal changes in these serum factors could be early biomarkers of treatment response and survival.

Experimental Design: Interleukin (IL)-6, IL-8, growth-related oncogene-1 (GRO-1), vascular endothelial growth factor (VEGF), and hepatocyte growth factor (HGF) concentrations were determined by Luminex multiplex assay using serum obtained at baseline and every 3 months in a prospective study of 30 patients with locally advanced (stage III/IV) oropharyngeal SCC receiving chemoradiation therapy. The relationship between baseline and direction of change in individual and multiple cytokines with cause-specific and disease-free survival was determined by Cox proportional hazards models and Kaplan-Meier survival analysis. Statistical analyses included adjustment for smoking status and response to chemoradiation.

Results: Three-year cause-specific and disease-free survival was 74.4% and 68.9%. Nonsmoking history (P = 0.05) and higher baseline VEGF (P = 0.003) correlated with increased survival. Longitudinal increases in levels of individual factors predicted decreased cause-specific survival when adjusted for smoking history [IL-6: relative risk (RR), 3.8; 95% confidence interval (95% CI), 2.0-7.4; P = 0.004; IL-8: RR, 1.6; 95% CI, 1.2-2.2; P = 0.05; VEGF: RR, 3.0; 95% CI, 1.6-5.6; P = 0.01; HGF: RR, 2.9; 95% CI, 1.9-4.4; P = 0.02; and GRO-1: RR, 1.2; 95% CI, 1.1-1.3; P = 0.02]. For a given individual, large increases in the upper quartile for any three or more factors predicted poorer cause-specific survival compared with patients with two or fewer large increases in factor levels (P = 0.004).

Conclusions: Pretreatment VEGF levels and longitudinal change in IL-6, IL-8, VEGF, HGF, and GRO-1 may be useful as biomarkers for response and survival in patients with locally advanced oropharyngeal and head and neck SCC treated with chemoradiation.
Distinction of Hereditary Nonpolyposis Colorectal Cancer and Sporadic Microsatellite-Unstable Colorectal Cancer through Quantification of MLH1 Methylation by Real-time PCR
Authors: Bettstetter Marcus , Dechant Stephan , Ruemmele Petra , Grabowski Monika , Keller Gisela , Holinski-Feder Elke , Hartmann Arndt , Hofstaedter Ferdinand , Dietmaier Wolfgang
Published: June 2007

Purpose: Promoter hypermethylation occurs frequently in tumors and leads to silencing of tumor-relevant genes like tumor suppressor genes. In a subset of sporadic colorectal cancers (CRC), inactivation of the mismatch repair gene MLH1 due to promoter methylation causes high level of microsatellite instability (MSI-H). MSI-H is also a hallmark of hereditary nonpolyposis colorectal cancer (HNPCC) in which mismatch repair inactivation results from germ-line mutations. For differentiation of sporadic and hereditary MSI-H tumor patients, MLH1 promoter methylation analysis is a promising tool but is not yet used in daily diagnostics because only qualitative techniques without standardization are available. The aim of this study is to establish a reliable and quantitative MLH1 methylation analysis technique and to define valid MLH1 methylation cutoff values for HNPCC diagnostics.

Experimental Design: We developed a new real-time PCR–based technique to detect and quantify methylation of both proximal and distal hMLH1 promoter regions. We established and validated this technique in a cohort of 108 CRCs [94 MSI-H and 16 microsatellite stable (MSS) cases] comprising a reference (n = 58) and a tester tumor group (n = 50).

Results: The reference tumor group contained 28 HNPCC with proven germ-line mutations or positive Amsterdam I criteria (median age, 37 years) and loss of MLH1 expression, 14 sporadic MSI-H CRC tumors with loss of MLH1 expression and BRAF V600E mutation (median age, 80.5 years), and 16 sporadic MSS CRC (median age, 76.5 years). No MLH1 promoter methylation could be found in any MSS tumors. HNPCC patients showed no or low level of MLH1 promoter methylation. A cutoff value of 18% methylation extent could be determined in this study to define MLH1 hypermethylation specific for sporadic MSI-H cases. Methylation could also be verified qualitatively by melting point analysis. BRAF V600E mutations were not detected in any HNPCC patients (n = 22 informative cases).

Conclusion: According to the present data, quantitative MLH1 methylation analysis in MSI-H CRC is a valuable molecular tool to distinguish between HNPCC and sporadic MSI-H CRC. The detection of a BRAF V600E mutation further supports the exclusion of HNPCC.
A Polymorphism in HDM2 (SNP309) Associates with Early Onset in Superficial Tumors, TP53 Mutations, and Poor Outcome in Invasive Bladder Cancer
Authors: Sanchez-Carbayo Marta , Socci Nicholas D. , Kirchoff Thomas , Erill Nadina , Offit Keneth , Bochner Bernard H. , Cordon-Cardo Carlos
Published: June 2007

Purpose: The HDM2 gene represents one of the central nodes in the p53 pathway. A recent study reported the association of a single nucleotide polymorphism (SNP309) in the HDM2 promoter region with accelerated tumor formation in both hereditary and sporadic cancers. In this study, we aim to evaluate the SNP309 in bladder cancer and to link it to TP53 status.

Experimental Design: SNP309 genotyping and TP53 mutation status were done on 141 bladder tumors and 8 bladder cancer cell lines using a RFLP strategy and TP53 genotyping arrays, respectively. Transcript profiling of a subset of cases (n = 41) was done using oligonucleotide arrays to identify genes differentially expressed regarding their SNP309 status.

Results: Of 141 bladder tumors analyzed, 36.9% displayed the SNP309 wild-type (WT; T/T) genotype, whereas 11.3% were homozygous (G/G) and 51.8% were heterozygous (T/G) cases. Patients with superficial disease and the G/G genotype had an earlier age on onset than those with the T/G or T/T genotypes (P = 0.029). Tumors with SNP309 WT genotype significantly displayed TP53 mutations when compared with tumors harboring G/G or T/G genotypes (P < 0.05). SNP309 WT cases had a poorer overall survival than cases with G/G and T/G genotypes (P < 0.05). TP53 mutation status provided enhanced prognostic value (P < 0.001). Transcript profiling identified TP53 targets among those differentially expressed between tumors displaying G/G or T/G SNP309 versus WT cases.

Conclusions: SNP309 is a frequent event in bladder cancer, related to earlier onset of superficial disease and TP53 mutation status. SNP309 genotypes were found to be associated with clinical outcome.
KIF14 Messenger RNA Expression Is Independently Prognostic for Outcome in Lung Cancer
Authors: Corson Timothy W. , Zhu Chang Qi , Lau Suzanne K. , Shepherd Frances A. , Tsao Ming-Sound , Gallie Brenda L.
Published: June 2007

Purpose: The mitotic kinesin KIF14 is overexpressed in multiple cancers including lung cancer. Therefore, we investigated KIF14 expression in association with clinical variables and the effect of KIF14 on in vitro colony formation in non–small-cell lung carcinoma.

Experimental Design: RNA was extracted from 129 untreated, resected tumors and KIF14 expression was quantified by real-time reverse transcription-PCR. Associations with clinical variables were determined by standard statistical methods. KIF14 expression was knocked down by small interfering RNA in H1299 and HeLa cells; proliferation and growth in soft agar were assayed.

Results: Squamous cell carcinoma had the highest KIF14 level, followed by large-cell undifferentiated carcinoma, then adenocarcinoma (P = 0.002). KIF14 level decreased with differentiation (P = 0.01) but was not associated with pathologic stage, T or N stage, or sex. When dichotomized about the median, KIF14 overexpression significantly decreased disease-free survival (Kaplan-Meier log-rank, P = 0.01) and trended toward decreasing overall survival (P = 0.08). In a univariate Cox proportional hazard regression, increasing KIF14 expression decreased disease-free survival [P = 0.01; hazard ratio, 1.44 (95% confidence interval, 1.09-1.91)]. In a multivariate Cox regression, including stage, differentiation, histology, and tumor purity as covariates, KIF14 overexpression remained an independent prognostic factor for disease-free survival [P = 0.01; hazard ratio, 1.45 (95% confidence interval, 1.09-1.94)]. Knockdown of KIF14 in non–small-cell lung carcinoma and cervical carcinoma cell lines decreased proliferation and colony formation in soft agar.

Conclusions: KIF14 expression is independently prognostic for disease-free survival in lung cancer and knockdown decreases tumorigenicity in vitro, showing that it is a clinically relevant oncogene and an exciting therapeutic target for further study.
Chromosome 22q11.2 Deletion Presenting with Immune-Mediated Cytopenias, Macrothrombocytopenia and Platelet Dysfunction
Authors: Akar N.A. , Adekile A.D.
Published: June 2007

Objective: To report a case of chromosome 22q11.2 deletion presenting with large platelets, platelet dysfunction, immune-mediated thrombocytopenia and neutropenia, in addition to other features of the disease. Case Presentation and Intervention: The patient presented in the neonatal period with tetralogy of Fallot, subtle dysmorphic features and thrombocytopenia. Fluorescent in situ hybridization analysis confirmed the diagnosis of chromosome 22q11.2 deletion. Further investigations showed immune thrombocytopenia and neutropenia in addition to reduced expression of platelet GPIb and abnormal platelet aggregation studies. CD4:CD8 ratio was reversed. His cardiac abnormality was successfully corrected surgically. He had mild recurrent bacterial infections. Recurrent epistaxis was becoming increasingly more severe, and he had cognitive developmental and speech delay. His serum calcium, phosphorus and parathormone have remained normal. Conclusions: Immune thrombocytopenia can coexist with macrothrombocytopenia and platelet dysfunction in chromosome 22q11.2 deletion and may present with significant bleeding episodes.
Three Successive Cases of Enteric Anisakiasis
Authors: Ishida Masaharu , Harada Akihiko , Egawa Shinichi , Watabe Shuichi , Ebina Nobuo , Unno Michiaki
Published: June 2007

Enteric anisakiasis is relatively rare, and the preoperative diagnosis is difficult. We report 3 cases of enteric anisakiasis: 1 was confirmed by operation, and the other 2 cases were suspected by the patient history. The 1st patient was a 48-year-old female presenting with abdominal pain. An abdominal computed tomography scan showed a dilated small intestine and accumulation of ascites. We performed partial resection of the small intestine, and an Anisakis nematode was found on the wall of the resected bowel. After surgery, the detailed history revealed that the patient consumed a raw sardine with vinegar and miso, fermented soybean paste. Three days after her operation, 2 men presenting with abdominal pain visited the hospital, and computed tomography scans of the patients showed dilated small intestines and accumulation of ascites. By taking patient histories, we found that both of them consumed sardines with vinegar and miso, and both were given conservative treatment with fasting and transfusion. By the experience of the 1st case, we could diagnose the following 2 cases as having enteric anisakiasis. In conclusion, the possibility of anisakiasis should be considered in patients with abdominal pain after ingesting raw fish, especially when intestinal obstruction is suspected.
Vascular Remodeling
Authors: Korshunov Vyacheslav A. , Schwartz Stephen M. , Berk Bradford C.
Published: June 2007

An important concept for vascular remodeling, termed Glagov’s phenomenon, is that arteries remodel to maintain constant flow despite increases in atherosclerotic lesion mass. Although Glagov’s phenomenon was originally described only for the case of arterial remodeling in response to growth of atherosclerotic plaques, experimental and clinical observations indicate that blood flow properties influence remodeling after angioplasty, hypertension, and flow diversion as well as atherosclerotic plaque progression. This review attempts to define Glagov’s observation in terms of the physical parameters of blood in conduit arteries that must determine the remodeling response. Next we review experiments that have begun to identify specific molecules that influence vascular remodeling and therefore may serve as mediators for the phenomena. More comprehensive analyses of the specific molecular pathways in the vessels that determine constant flow may provide new therapeutic approaches to regulate vascular remodeling.
Nox1 Mediates Basic Fibroblast Growth Factor-Induced Migration of Vascular Smooth Muscle Cells
Authors: Schröder Katrin , Helmcke Ina , Palfi Katalin , Krause Karl-Heinz , Busse Rudi , Brandes Ralf P.
Published: June 2007

Objectives—

Basic fibroblast growth factor (bFGF) stimulates vascular smooth muscle cell (SMC) migration. We determined whether bFGF increases SMC reactive oxygen-species (ROS) and studied the role of ROS for SMC migration.

Methods and Results—

bFGF rapidly increased rat SMC ROS formation and migration through pathways sensitive to inhibition of NADPH oxidases, PI3-kinase, protein kinase C, and Rac-1. SiRNA directed against the NADPH oxidase Nox4 impaired basal but not bFGF-induced ROS formation and did not affect migration. In contrast, siRNA against Nox1 blocked the agonist-induced ROS generation as well as the bFGF-induced migration. Agonist-induced migration was also attenuated in SMC derived from Nox1 y/− mice and transduction of Nox1 restored normal migration. Likewise, SMC outgrowth in response to bFGF was attenuated in aortic segments from Nox1 y/− mice as compared with Nox1 y/+ mice. bFGF activated JNK but not Src in a Nox1-dependent manner. Consequently, phosphorylation of the adaptor protein paxillin, which is central for migration and secretion of matrix-metalloproteinases, were dependent on Nox1 as well as JNK but not Src.

Conclusions—

These data demonstrate that bFGF activates the Nox1-containing NADPH oxidase and that bFGF through a pathway involving ROS and JNK stimulates SMC migration.
Peripheral Circadian Clocks in the Vasculature
Authors: Reilly Dermot F. , Westgate Elizabeth J. , FitzGerald Garret A.
Published: June 2007

Living organisms have adapted to the daily rotation of the earth and regular changes in the light environment. Life forms anticipate environmental transitions, adapt their own physiology, and perform activities at behaviorally advantageous times during the day. This is achieved by means of endogenous circadian clocks that can be synchronized to the daily changes in external cues, most notably light and temperature. For many years it was thought that neurons of the suprachiasmatic nucleus (SCN) uniquely controlled circadian rhythmicity of peripheral tissues via neural and humoral signals. The cloning and characterization of mammalian clock genes revealed that they are expressed in a circadian manner throughout the body. It is now accepted that peripheral cells, including those of the cardiovascular system, contain a circadian clock similar to that in the SCN. Many aspects of cardiovascular physiology are subject to diurnal variation, and serious adverse cardiovascular events including myocardial infarction, sudden cardiac death, and stroke occur with a frequency conditioned by time of day. This has raised the possibility that biological responses under the control of the molecular clock might interact with environmental cues to influence the phenotype of human cardiovascular disease.
Relationship Between a Systemic Inflammatory Marker, Plaque Inflammation, and Plaque Characteristics Determined by Intravascular Optical Coherence Tomography
Authors: Raffel O. Christopher , Tearney Guillermo J. , Gauthier Denise DeJoseph , Halpern Elkan F. , Bouma Brett E. , Jang Ik-Kyung
Published: June 2007

Objective—

The purpose of this study was to evaluate the relationships between the peripheral white blood cell (WBC) count, local plaque fibrous cap macrophage density, and the morphological features and presence of thin-cap fibroatheromas (TCFA) identified by optical coherence tomography (OCT).

Methods and Results—

OCT was performed in patients undergoing catheterization. Images were analyzed using validated criteria for plaque characteristics. Baseline WBC count correlated with macrophage density (
r
=0.483,
P
<0.001). Both parameters were associated with lipid-rich plaque and correlated inversely with plaque fibrous cap thickness (
r
=−0.547 for macrophage density and −0.423 for WBC count,
P
<0.015). Plaques classified as TCFA had a higher median macrophage density than non-TCFA plaques (7.4 versus 4.99,
P
<0.001). Patients with TCFA had a higher WBC count compared with those without TCFA (11.0 versus 7.9,
P
=0.007). Receiver operator curves for WBC count, macrophage density, and these combined parameters for prediction of TCFA showed the area under the curves were 0.88, 0.91, and 0.97 (
P
<0.001), respectively.

Conclusion—

This study provides the first in vivo data linking the peripheral WBC count, plaque fibrous cap macrophage density, and the characteristics and presence of TCFA. Macrophage density correlated with the WBC count, and both parameters independently and particularly in combination predict the presence of TCFA.
Mechanical Stretch Simulates Proliferation of Venous Smooth Muscle Cells Through Activation of the Insulin-Like Growth Factor-1 Receptor
Authors: Cheng Jizhong , Du Jie
Published: June 2007

Objective—

Activation and proliferation of vascular smooth muscle cells (VSMCs) occur in the venous neointima of vein grafts. VSMCs in a grafted vein are subjected to mechanical stretch; our goal is to understand the essential mechanical stretch-regulated signals that influence VSMCs during neointimal formation in vein grafts.

Methods and Results—

In cultured vein VSMCs, mechanical stretch induces proliferation and upregulation of both IGF-1 and IGF-1R. Stretch of VSMCs sustained tyrosine phosphorylation of both IGF-1R and its substrate, IRS-1; these responses were related to mechanical stretch-induced activation of Src and autocrine IGF-1 production. Mechanical stretch-activated IGF-1R is functional because there is a prolonged activation of IRS-1-associated phosphatidylinositol-3 kinase (PI3K). When we knocked out IGF-1R, the mechanical stretch-induced increase in VSMC proliferation was blocked. To link mechanical stretch-activated IGF-1R cell signaling to venous VSMC proliferation in vivo, we also studied a vein graft model. Tamoxifen-inducible null deletion of IGF-1R in mice reduced the formation of neointima in the vein graft.

Conclusions—

Our results demonstrate for the first time that mechanical stretch activates IGF-1/IGF-1R signals in venous VSMCs, and we have uncovered a signaling pathway that leads to neointima formation in vivo.