bluuebunny/crossref_metadata_2025 huggingface dataset:
K-Ras Mutations and Treatment Outcome in Colorectal Cancer Patients Receiving Exclusive Fluoropyrimidine Therapy
Authors: Etienne-Grimaldi Marie-Christine , Formento Jean-Louis , Francoual Mireille , François Eric , Formento Patricia , Renée Nicole , Laurent-Puig Pierre , Chazal Maurice , Benchimol Daniel , Delpero Jean-Robert , Letoublon Christian , Pezet Denis , Seitz Jean-François , Milano Gérard
Published: August 2008

Purpose: K-Ras mutations predict resistance to anti–epidermal growth factor receptor (EGFR) monoclonal antibodies. Because combinations of anti-EGFR with 5-fluorouracil (5-FU)-based chemotherapy are promising treatments, we analyzed the effect of K-Ras mutations in patients having received exclusive 5-FU therapy.

Experimental Design: This study was conducted on 93 stage IV colorectal cancer patients with unresectable measurable liver metastasis receiving 5-FU-leucovorin (56 men and 37 women; 77 cancer deaths). Liver metastases (n = 93) along with primary tumors (n = 48) were analyzed for K-Ras mutations (codons 12 and 13), p53 mutations (exons 4-9), p53 polymorphism (codon 72), thymidylate synthase (TS) polymorphism (28-bp repeats including G>C mutation), methylenetetrahydrofolate reductase polymorphism (677C>T, 1298A>C), thymidylate synthase (TS) activity, dihydropyrimidine dehydrogenase activity, folylpolyglutamate synthase activity, and p53 protein expression.

Results: Thirty-six of 93 (38.7%) metastases were K-Ras mutated (30 at codon 12 and 6 at codon 13). Mutated primary tumors (16 of 48) matched perfectly with mutated metastases. The additional analyzed tumor markers were not different between K-Ras mutated and wild-type tumors. The objective response rate was 37%: 44.4% in K-Ras mutated versus 32.1% in wild-type K-Ras metastasis (P = 0.27). Low TS activity in metastasis was the only significant predictor of tumor response (P = 0.047). K-Ras status did not influence specific survival.

Conclusions: The present data indicate a perfect concordance of K-Ras mutations between primary and liver metastasis and suggest that any predictive and/or prognostic value of K-Ras mutations in treatments combining anti-EGFR monoclonal antibodies with 5-FU should be exclusively linked to the anti-EGFR agent.
Improved Identification of von Hippel-Lindau Gene Alterations in Clear Cell Renal Tumors
Authors: Nickerson Michael L. , Jaeger Erich , Shi Yangu , Durocher Jeffrey A. , Mahurkar Sunil , Zaridze David , Matveev Vsevolod , Janout Vladimir , Kollarova Hellena , Bencko Vladimir , Navratilova Marie , Szeszenia-Dabrowska Neonilia , Mates Dana , Mukeria Anush , Holcatova Ivana , Schmidt Laura S. , Toro Jorge R. , Karami Sara , Hung Rayjean , Gerard Gary F. , Linehan W. Marston , Merino Maria , Zbar Berton , Boffetta Paolo , Brennan Paul , Rothman Nathaniel , Chow Wong-Ho , Waldman Frederic M. , Moore Lee E.
Published: August 2008

Purpose: To provide a comprehensive, thorough analysis of somatic mutation and promoter hypermethylation of the von Hippel-Lindau (VHL) gene in the cancer genome, unique to clear cell renal cancer (ccRCC). Identify relationships between the prevalence of VHL gene alterations and alteration subtypes with patient and tumor characteristics.

Experimental Design: As part of a large kidney cancer case-control study conducted in Central Europe, we analyzed VHL mutations and promoter methylation in 205 well-characterized, histologically confirmed patient tumor biopsies using a combination of sensitive, high-throughput methods (endonuclease scanning and Sanger sequencing) and analysis of 11 CpG sites in the VHL promoter.

Results: We identified mutations in 82.4% of cases, the highest VHL gene mutation prevalence reported to date. Analysis of 11 VHL promoter CpG sites revealed that 8.3% of tumors were hypermethylated and all were mutation negative. In total, 91% of ccRCCs exhibited alteration of the gene through genetic or epigenetic mechanisms. Analysis of patient and tumor characteristics revealed that certain mutation subtypes were significantly associated with Fuhrman nuclear grade, metastasis, node positivity, and self-reported family history of RCC.

Conclusion: Detection of VHL gene alterations using these accurate, sensitive, and practical methods provides evidence that the vast majority of histologically confirmed ccRCC tumors possess genetic or epigenetic alteration of the VHL gene and support the hypothesis that VHL alteration is an early event in ccRCC carcinogenesis. These findings also indicate that VHL molecular subtypes can provide a sensitive marker of tumor heterogeneity among histologically similar ccRCC cases for etiologic, prognostic, and translational studies.
CHEK2 1100delC Is a Susceptibility Allele for HNPCC-Related Colorectal Cancer
Authors: Wasielewski Marijke , Vasen Hans , Wijnen Juul , Hooning Maartje , Dooijes Dennis , Tops Carli , Klijn Jan G.M. , Meijers-Heijboer Hanne , Schutte Mieke
Published: August 2008

Purpose: The pathogenic CHEK2 1100delC variant is firmly established as a breast cancer susceptibility allele. Dutch CHEK2 1100delC breast cancer families frequently also include colorectal cancer cases, and the variant is particularly prevalent among breast cancer families with hereditary breast and colorectal cancer. Yet, it is still unclear whether CHEK2 1100delC also confers a colorectal cancer risk independent of its breast cancer risk.

Experimental Design:CHEK2 1100delC was genotyped in the index cases of 369 Dutch colorectal cancer families that had been excluded for familial breast cancer. The cohort included 132 cases with familial adenomatous polyposis (FAP) and FAP-related disease, and 237 cases with hereditary nonpolyposis colorectal cancer (HNPCC) and HNPCC-related disease.

Results: None of the FAP/FAP-related cases carried the CHEK2 1100delC variant. In contrast, CHEK2 1100delC was present in 10 of 237 (4.2%) HNPCC/HNPCC-related cases that was significantly more prevalent than the 1.0% Dutch population frequency (odds ratio, 4.3; 95% confidence interval, 1.7-10.7; P = 0.002). Nine of the 10 CHEK2 1100delC colorectal cancer cases met the revised Amsterdam and/or Bethesda criteria. The 10 CHEK2 1100delC colorectal cancer families had a high-risk cancer inheritance pattern, including 35 colorectal cancer cases, 9 cases with polyps, and 21 cases with other tumor types.

Conclusion: Our analysis provides strong evidence that the 1100delC variant of CHEK2 confers a colorectal cancer risk in HNPCC/HNPCC-related families, supporting the hypothesis that CHEK2 is a multiorgan cancer susceptibility gene.
Delineation of TMPRSS2-ERG Splice Variants in Prostate Cancer
Authors: Hu Ying , Dobi Albert , Sreenath Taduru , Cook Christopher , Tadase Atekelt Y. , Ravindranath Lakshmi , Cullen Jennifer , Furusato Bungo , Chen Yongmei , Thangapazham Rajesh L. , Mohamed Ahmed , Sun Chen , Sesterhenn Isabell A. , McLeod David G. , Petrovics Gyorgy , Srivastava Shiv
Published: August 2008

Purpose: The expression of the ETS-related gene (ERG) is low or undetectable in benign prostate epithelial cells. High prevalence of ERG overexpression in prostate cancer cells due to TMPRSS2-ERG fusions suggest for causal roles of ERG protein in the neoplastic process. TMPRSS2-ERG fusion junctions have been extensively studied in prostate cancer. However, virtually nothing is known about the nature of full-length transcripts and encoded proteins. This study focuses on qualitative and quantitative features of full-length TMPRSS2-ERG transcripts in prostate cancer.

Experimental Design: Full-length TMPRSS2-ERG transcripts were cloned and sequenced from a cDNA library generated from pooled RNA of six TMPRSS2-ERG fusion–positive prostate tumors. The encoded ERG proteins were analyzed in HEK293 cells. Copy numbers of TMPRSS2-ERG splice variants were determined by quantitative reverse transcription-PCR in laser capture microdissected prostate cancer cells.

Results: Two types of TMPRSS2-ERG cDNAs were identified: type I, which encodes full-length prototypical ERG protein (ERG1, ERG2, ERG3), and type II, encoding truncated ERG proteins lacking the ETS domain (ERG8 and a new variant, TEPC). In microdissected prostate tumor cells from 122 patients, relative abundance of these variants was in the following order: ERG8 > TEPC > ERG 3 > ERG1/2 with combined overexpression rate of 62.3% in prostate cancer. Increased ratio of type I over type II splice forms showed a trend of correlation with less favorable pathology and outcome.

Conclusions: Qualitative and quantitative features of specific ERG splice variants defined here promise to enhance the utility of ERG as a biomarker and therapeutic target in prostate cancer.
Three-Gene Expression Signature Predicts Survival in Early-Stage Squamous Cell Carcinoma of the Lung
Authors: Skrzypski Marcin , Jassem Ewa , Taron Miquel , Sanchez Jose Javier , Mendez Pedro , Rzyman Witold , Gulida Grazyna , Raz Dan , Jablons David , Provencio Mariano , Massuti Bartomeu , Chaib Imane , Perez-Roca Laia , Jassem Jacek , Rosell Rafael
Published: August 2008

Purpose: Adjuvant treatment may improve survival in early-stage squamous cell carcinoma (SCC) of the lung; however, the absolute gain is modest and mainly limited to stage II-IIIA. Current staging methods are imprecise indications of prognosis, but high-risk patients can be identified by gene expression profiling and considered for adjuvant therapy.

Experimental Design: The expression of 29 genes was assessed by reverse transcriptase quantitative PCR in frozen primary tumor specimens obtained from 66 SCC patients who had undergone surgical resection. Expression values were dichotomized using the median as a cutoff value. We used a risk score to develop a gene expression model for the prediction of survival.

Results: The univariate analysis of gene expression in the training cohort identified 10 genes with significant prognostic value: CSF1, EGFR, CA IX, PH4, KIAA0974, ANLN, VEGFC, NTRK1, FN1, and INR1. In the multivariate Cox model, CSF1 (hazard ratio, 3.5; P = 0.005), EGFR (hazard ratio, 2.7; P = 0.02), CA IX (hazard ratio, 0.2; P < 0.0001), and tumor size >4 cm (hazard ratio, 2.7; P = 0.02) emerged as significant markers for survival. The high prognostic value of a risk score based on the expression of the three genes (CSF1, EGFR, and CA IX) was positively validated in a separate cohort of 26 patients in an independent laboratory (P = 0.05).

Conclusions: The three-gene signature is strongly associated with prognosis in early-stage SCC. Positive independent validation suggests its suitability for selecting SCC patients with an increased risk of death who might benefit from adjuvant treatment.
Alterations in Hepatic Metabolism in
fld
Mice Reveal a Role for Lipin 1 in Regulating VLDL-Triacylglyceride Secretion
Authors: Chen Zhouji , Gropler Matthew C. , Norris Jin , Lawrence John C. , Harris Thurl E. , Finck Brian N.
Published: August 2008

Objective—

Lipin 1 controls fatty acid metabolism in the nucleus as a transcriptional regulator and in the cytosol as an enzyme catalyzing the penultimate step in phosphoglycerol triacylglyceride (TAG) synthesis. We sought to evaluate the effects of lipin 1 on hepatic TAG synthesis and secretion by gain-of-function and loss-of-function approaches.

Methods and Results—

Rates of TAG synthesis were not impaired in hepatocytes isolated from adult lipin 1–deficient (
fld
) mice and were actually increased in 14-day-old
fld
mice. Additionally, compared to littermate controls, VLDL-TAG secretion rates were markedly increased in
fld
mice of both ages. Lipin 1 overexpression did not alter TAG synthesis rates but significantly suppressed VLDL-TAG secretion. The lipin 1-mediated suppression of VLDL-TAG secretion was linked to the peptide motif mediating its transcriptional-regulatory effects. However, the expression of candidate genes required for VLDL assembly and secretion was unaltered by lipin 1 activation or deficiency. Finally, the hepatic expression of lipin 1 was diminished in obese insulin-resistant mice, whereas adenoviral-mediated overexpression of lipin 1 in liver of these mice inhibits VLDL-TAG secretion and improves hepatic insulin signaling.

Conclusions—

Collectively, these studies reveal new and unexpected effects of lipin 1 on hepatic TAG metabolism and obesity-related hepatic insulin resistance.
Direct Comparison of Whole-Heart Navigator-Gated Magnetic Resonance Coronary Angiography and 40- and 64-Slice Multidetector Row Computed Tomography to Detect the Coronary Artery Stenosis in Patients Scheduled for Conventional Coronary Angiography
Authors: Pouleur Anne-Catherine , le Polain de Waroux Jean-Benoît , Kefer Joelle , Pasquet Agnès , Vanoverschelde Jean-Louis , Gerber Bernhard L.
Published: August 2008

Background—

Both whole-heart magnetic resonance coronary angiography (WH-MRCA) and multidetector computed tomography (MDCT) have been proposed for the noninvasive identification of the coronary stenosis. The authors sought to directly compare the diagnostic accuracy of these noninvasive imaging techniques using the invasive quantitative coronary angiography as a reference standard.

Methods and Results—

Seventy-seven consecutive patients (56 men, 61�14 years) prospectively underwent WH-MRCA and 40- or 64-slice MDCT before the quantitative coronary angiography. Diagnostic accuracy of WH-MRCA and MDCT for the visual identification of >50% diameter stenosis in segments of >1.5 mm size was compared using the quantitative coronary angiography as a reference. According to the quantitative coronary angiography, 49 of 992 coronary segments >1.5 mm diameter had >50% diameter stenosis. MDCT had a higher success rate (100% versus 88%,
P
<0.001) and enabled identification of more segments (963 versus 726,
P
<0.001) than did WH-MRCA. On a per-segment basis, WH-MRCA had similar sensitivity (47/49 or 96% versus 48/49 or 98%,
P
=0.9) but significantly lower specificity (644/943 or 68% versus 863/943 or 92%,
P
<0.001) and accuracy (691/992 or 70% versus 911/992 or 92%,
P
<0.001) for the detection of >50% diameter stenosis than did MDCT. On a per-patient basis, the sensitivity was similar (17/17 or 100% versus 16/17 or 94%,
P
=0.9), but specificity (43/60 or 72% versus 53/60 or 88%,
P
=0.024) and diagnostic accuracy (60/77 or 78%, versus 69/77 or 90%,
P
=0.044) of WH-MRCA for the detection of >50% diameter stenosis were significantly lower than of MDCT.

Conclusions—

Because of the higher success rate and higher number of interpretable segments, 40- or 64-slice MDCT performs better than WH-MRCA.
123
I-
m
IBG Scintigraphy to Predict Inducibility of Ventricular Arrhythmias on Cardiac Electrophysiology Testing
Authors: Bax Jeroen J. , Kraft Otakar , Buxton Alfred E. , Fjeld Jan Gunnar , Pařízek Petr , Agostini Denis , Knuuti Juhani , Flotats Albert , Arrighi James , Muxi Africa , Alibelli Marie-Jeanne , Banerjee Gopa , Jacobson Arnold F.
Published: August 2008

Background—

Disturbances of autonomic function after infarction are associated with both total mortality and sudden death. Although many imaging techniques for assessing the cardiac autonomic nervous system have been studied, the clinical usefulness of these techniques remains uncertain. This exploratory pilot study examined the relationship between abnormalities of ventricular sympathetic innervation delineated by scintigraphic imaging with
123
I-
m
IBG and inducible ventricular tachyarrhythmias in patients with left ventricular dysfunction and previous myocardial infarction.

Methods and Results—

Fifty patients underwent electrophysiological (EP) testing and 15-minute and 4-hour planar and single photon emission computed tomography (SPECT) imaging with
123
I-
m
IBG and SPECT imaging with
99m
Tc-tetrofosmin. The primary efficacy variables were the 4-hour heart:mediastinum ratio (H/M) and the
123
I-
m
IBG/
99m
Tc-tetrofosmin SPECT mismatch score. EP studies were categorized as positive (EP
+
) or negative (EP
−
) for inducibility of sustained (>30 seconds) ventricular tachyarrhythmias. Thirty patients were EP
+
, and 20 were EP
−
. There were no significant differences in the 4-hour H/M ratios or
123
I-
m
IBG/
99m
Tc-tetrofosmin SPECT mismatch scores between the two groups. In a multivariable analysis using all
123
I-
m
IBG and
99m
Tc-tetrofosmin SPECT measurements, the only variable that showed a significant difference between EP
+
and EP
−
patients was the 4-hour
123
I-
m
IBG SPECT defect score. A 4-hour
123
I-
m
IBG SPECT defect score of ≥37 yielded a sensitivity of 77% and specificity of 75% for predicting EP results.

Conclusions—

The standard indices of
123
I-
m
IBG imaging (H/M and innervation-perfusion mismatch score) are not predictive of EP test results. The association of
123
I-
m
IBG SPECT defect severity with EP test inducibility in this exploratory study will require confirmation in a larger cohort of patients.
Stretch-Induced Activation of the Transcription Factor Activator Protein-1 Controls Monocyte Chemoattractant Protein-1 Expression During Arteriogenesis
Authors: Demicheva Elena , Hecker Markus , Korff Thomas
Published: August 2008

Cerebral, coronary, and peripheral artery diseases combined represent the most frequent cause of death in developed nations. The underlying progressive occlusion of large conductance arteries can partially be compensated for by transformation of preexisting collateral arterioles to small artery bypasses, a process referred to as arteriogenesis. Because biomechanical forces have been implicated in the initiation of arteriogenesis, we have investigated the mechanosensitive expression of a pivotal proarteriogenic molecule, monocyte chemoattractant protein (MCP)-1, which governs the recruitment of circulating monocytes to the wall of the remodeling collateral arterioles. Using a new ear artery ligation model and the classic hindlimb ischemia model in mice, we noted that MCP-1 expression is significantly increased in collateral arterioles undergoing arteriogenesis already 24 hours after its onset. By mimicking proarteriogenic perfusion conditions in small mouse arteries, we observed that MCP-1 expression is predominantly upregulated in the smooth muscle cells, which solely sense changes in circumferential wall tension or stretch. Subsequent analyses of cultured endothelial and smooth muscle cells confirmed that cyclic stretch but not shear stress upregulates MCP-1 expression in these cells. Blockade of the mechanosensitive transcription factor activator protein-1 by using a specific decoy oligodeoxynucleotide abolished this stretch-induced MCP-1 expression. Likewise, topical administration of the decoy oligodeoxynucleotide to the mouse ear abrogated arteriogenesis through downregulation of MCP-1 expression and monocyte recruitment. Collectively, these findings point toward a stretch-induced activator protein-1–mediated rise in MCP-1 expression in vascular smooth muscle cells as a critical determinant for the initiation of arteriogenesis.
p21-Activated Kinase Signaling Regulates Oxidant-Dependent NF-κB Activation by Flow
Authors: Orr A. Wayne , Hahn Cornelia , Blackman Brett R. , Schwartz Martin Alexander
Published: August 2008

Disturbed blood flow induces inflammatory gene expression in endothelial cells, which promotes atherosclerosis. Flow stimulates the proinflammatory transcription factor nuclear factor (NF)-κB through integrin- and Rac-dependent production of reactive oxygen species (ROS). Previous work demonstrated that NF-κB activation by flow is matrix-specific, occurring in cells on fibronectin but not collagen. Activation of p21-activated kinase (PAK) followed the same matrix-dependent pattern. We now show that inhibiting PAK in cells on fibronectin blocked NF-κB activation by both laminar and oscillatory flow in vitro and at sites of disturbed flow in vivo. Constitutively active PAK rescued flow-induced NF-κB activation in cells on collagen. Surprisingly, PAK was not required for flow-induced ROS production. Instead, PAK modulated the ability of ROS to activate the NF-κB pathway. These data demonstrate that PAK controls NF-κB activation by modulating the sensitivity of cells to ROS.
On Free-Edge Effect and Onset of Delamination in FRPC Laminates Using Mixed Finite Element Model
Authors: Ramtekkar G.S. , Desai Y.M.
Published: August 2008

Free-edge stress analysis and the onset of delamination at the free-edges of FRPC laminates have been studied in the present paper. The stress analysis has been performed using a layer-wise 3-D mixed finite element model developed by the authors. Because the mixed FE formulation appropriately models the kinetics and the kinematics of laminated structures, the high stress gradients at the free-edge region of composite laminates have been appositely mapped. The onset of delamination failure has been computed using the tensor polynomial (quadratic) failure criterion. Results have been compared with the theoretical and experimental results available in the literature. It has been shown that the present results are in good agreement with the experimental observations.
Effect of Surface Quality of Rayon-Base Carbon Fabrics on Wettability of Phenolic Prepreg
Authors: Liu L. , Huang Y.D. , Hu B.S. , Zhang X.Z. , Wu L.N.
Published: August 2008

The wettability between rayon-base carbon fabrics (CF) and phenolic resin is key to prepreg quality and the total quality of the composites. Three types of rayon-base CF with different characteristics were selected. The surface sizing, its content, and the degree of graphitization of CF, and their effects on the wettability of CF by phenolic resin were analyzed. The results showed that the content of surface sizing and the degree of graphitization of CF were the main factors to determine the wettability.
Feeling Like a Nurse
Authors: Spence Deborah , Smythe Elizabeth
Published: August 2008

Purpose: To explore the essential meaning of being a nurse. Design: Nine registered nurses were each asked to write a story of a time when they felt like a nurse. Analysis was informed by Heideggerian hermeneutic philosophy, seeking to reveal the phenomenon of “feeling like a nurse.” Methods: As part of a workshop on phenomenological methodology, participants were invited to reflect on a personal experience of feeling like a nurse. These documents were analyzed according to the method of van Manen. Participants were kept informed throughout each phase. Findings: Feelings announce primordial meaning of feeling like a nurse. Nurses experience the call as mood attuned by an anxiety that creates possibilities for authentic caring. It is a way of being that encompasses watching and acting, doing to and caring for, and taking over and giving back. Moreover, it is after the encounter that the essential meaning is more clearly revealed. We argue that there is value in continuing to question the meaning of “being a nurse.” Amid a complex and increasingly technological world, this calls the profession to remember the human encounter at the heart of all nursing.
Tragedy, Reconciliation and Reconstruction
Authors: Frost Mervyn
Published: August 2008

This article explores the uses of tragedy as a mode of analysis in international relations. In tragic analyses, actors are portrayed as acting ethically, but through their deeds they bring about consequences that are contrary to the values in the name of which the deeds were undertaken. The good deeds bring about ethically obnoxious consequences. The article demonstrates how tragic analyses can be made of the actions of collective actors such as states and nations. Examples from Rhodesia, South Africa and the Balkans are used to demonstrate this. Tragic stories elicit sympathy for the protagonists. Such accounts are compared with rival accounts of the same acts, in terms of `just war theory', for example, which accounts do not generate sympathy but call forth emotions of outrage and condemnation. Finally, a case is made for the use of the tragic form of analysis in international affairs. Such analyses highlight the tensions and contradictions between rival social practices and point the way towards political transformations that will make a repetition of those cases of tragedy less likely in future.
Targeting of antigens to B cells augments antigen-specific T-cell responses and breaks immune tolerance to tumor-associated antigen MUC1
Authors: Ding Chuanlin , Wang Li , Marroquin Jose , Yan Jun
Published: August 2008

B cells are antibody (Ab)–secreting cells as well as potent antigen (Ag)–presenting cells that prime T-cell activation, which evokes great interest in their use for vaccine development. Here, we targeted ovalbumin (OVA) to B cells via CD19 and found that a single low dose of anti–CD19-OVA conjugates, but not isotype mAb-OVA, stimulated augmented CD4 and CD8 T-cell proliferation and expansion. Administration of TLR9 agonist CpG could significantly enhance long-term T-cell survival. Similar results were obtained when the tumor-associated Ag MUC1 was delivered to B cells. MUC1 transgenic (Tg) mice were previously found to lack effective T-cell help and produce low-titer of anti-MUC1 Abs after vaccination. Targeting MUC1 to B cells elicited high titer of anti-MUC1 Abs with different isotypes, predominantly IgG2a and IgG2b, in MUC1 Tg mice. The isotype switching of anti-MUC1 Ab was CD4 dependent. In addition, IFN-γ–producing CD8 T cells and in vivo cytolytic activity were significantly increased in these mice. The mice also showed significant resistance to MUC1+ lymphoma cell challenge both in the prophylactic and therapeutic settings. We conclude that Ags targeting to B cells stimulate CD4 and CD8 T-cell responses as well as Th-dependent humoral immune responses.
Multiparameter flow cytometric remission is the most relevant prognostic factor for multiple myeloma patients who undergo autologous stem cell transplantation
Authors: Paiva Bruno , Vidriales Maria-Belén , Cerveró Jorge , Mateo Gema , Pérez Jose J. , Montalbán Maria A. , Sureda Anna , Montejano Laura , Gutiérrez Norma C. , de Coca Alfonso García , de las Heras Natalia , Mateos Maria V. , López-Berges Maria C. , García-Boyero Raimundo , Galende Josefina , Hernández Jose , Palomera Luis , Carrera Dolores , Martínez Rafael , de la Rubia Javier , Martín Alejandro , Bladé Joan , Lahuerta Juan J. , Orfao Alberto , San Miguel Jesús F.
Published: August 2008

Minimal residual disease (MRD) assessment is standard in many hematologic malignancies but is considered investigational in multiple myeloma (MM). We report a prospective analysis of the prognostic importance of MRD detection by multiparameter flow cytometry (MFC) in 295 newly diagnosed MM patients uniformly treated in the GEM2000 protocol VBMCP/VBAD induction plus autologous stem cell transplantation [ASCT]). MRD status by MFC was determined at day 100 after ASCT. Progression-free survival (PFS; median 71 vs 37 months, P < .001) and overall survival (OS; median not reached vs 89 months, P = .002) were longer in patients who were MRD negative versus MRD positive at day 100 after ASCT. Similar prognostic differentiation was seen in 147 patients who achieved immunofixation-negative complete response after ASCT. Moreover, MRD− immunofixation-negative (IFx−) patients and MRD− IFx+ patients had significantly longer PFS than MRD+ IFx− patients. Multivariate analysis identified MRD status by MFC at day 100 after ASCT as the most important independent prognostic factor for PFS (HR = 3.64, P = .002) and OS (HR = 2.02, P = .02). Our findings demonstrate the clinical importance of MRD evaluation by MFC, and illustrate the need for further refinement of MM re-sponse criteria. This trial is registered at http://clinicaltrials.gov under identifier NCT00560053.
The role of neutrophils in the upper and lower respiratory tract during influenza virus infection of mice
Authors: Tate Michelle D , Brooks Andrew G , Reading Patrick C
Published: August 2008

Background

Neutrophils have been shown to play a role in host defence against highly virulent and mouse-adapted strains of influenza virus, however it is not clear if an effective neutrophil response is an important factor moderating disease severity during infection with other virus strains. In this study, we have examined the role of neutrophils during infection of mice with influenza virus strain HKx31, a virus strain of the H3N2 subtype and of moderate virulence for mice, to determine the role of neutrophils in the early phase of infection and in clearance of influenza virus from the respiratory tract during the later phase of infection.

Methods

The anti-Gr-1 monoclonal antibody (mAb) RB6-8C5 was used to (i) identify neutrophils in the upper (nasal tissues) and lower (lung) respiratory tract of uninfected and influenza virus-infected mice, and (ii) deplete neutrophils prior to and during influenza virus infection of mice.

Results

Neutrophils were rapidly recruited to the upper and lower airways following influenza virus infection. We demonstrated that use of mAb RB6-8C5 to deplete C57BL/6 (B6) mice of neutrophils is complicated by the ability of this mAb to bind directly to virus-specific CD8+T cells. Thus, we investigated the role of neutrophils in both the early and later phases of infection using CD8+T cell-deficient B6.TAP-/-mice. Infection of B6.TAP-/-mice with a low dose of influenza virus did not induce clinical disease in control animals, however RB6-8C5 treatment led to profound weight loss, severe clinical disease and enhanced virus replication throughout the respiratory tract.

Conclusion

Neutrophils play a critical role in limiting influenza virus replication during the early and later phases of infection. Furthermore, a virus strain of moderate virulence can induce severe clinical disease in the absence of an effective neutrophil response.
HAPSIMU: a genetic simulation platform for population-based association studies
Authors: Zhang Feng , Liu Jianfeng , Chen Jie , Deng Hong-Wen
Published: August 2008

Background

Population structure is an important cause leading to inconsistent results in population-based association studies (PBAS) of human diseases. Various statistical methods have been proposed to reduce the negative impact of population structure on PBAS. Due to lack of structural information in real populations, it is difficult to evaluate the impact of population structure on PBAS in real populations.

Results

We developed a genetic simulation platform, HAPSIMU, based on real haplotype data from the HapMap ENCODE project. This platform can simulate heterogeneous populations with various known and controllable structures under the continuous migration model or the discrete model. Moreover, both qualitative and quantitative traits can be simulated using additive genetic model with various genetic parameters designated by users.

Conclusion

HAPSIMU provides a common genetic simulation platform to evaluate the impact of population structure on PBAS, and compare the relative performance of various population structure identification and PBAS methods.
Peripheral arterial occlusive disease: Global gene expression analyses suggest a major role for immune and inflammatory responses
Authors: Fu Shijun , Zhao Haiguang , Shi Jiantao , Abzhanov Arhat , Crawford Keith , Ohno-Machado Lucila , Zhou Jianqin , Du Yanzhi , Kuo Winston Patrick , Zhang Ji , Jiang Mier , Jin Jason Gang
Published: August 2008

Background

Peripheral arterial disease (PAD), a major manifestation of atherosclerosis, is associated with significant cardiovascular morbidity, limb loss and death. However, mechanisms underlying the genesis and progression of the disease are far from clear. Genome-wide gene expression profiling of clinical samples may represent an effective approach to gain relevant information.

Results

After histological classification, a total of 30 femoral artery samples, including 11 intermediate lesions, 14 advanced lesions and 5 normal femoral arteries, were profiled using Affymetrix microarray platform. Following real-time RT-PCR validation, different algorithms of gene selection and clustering were applied to identify differentially expressed genes. Under a stringent cutoff, i.e., a false discovery rate (FDR) <0.5%, we found 366 genes were differentially regulated in intermediate lesions and 447 in advanced lesions. Of these, 116 genes were overlapped between intermediate and advanced lesions, including 68 up-regulated genes and 48 down-regulated ones. In these differentially regulated genes, immune/inflammatory genes were significantly up-regulated in different stages of PAD, (85/230 in intermediate lesions, 37/172 in advanced lesions). Through literature mining and pathway analysis using different databases such as Gene Ontology (GO), and the Kyoto Encyclopedia of Gene and Genomics (KEGG), genes involved in immune/inflammatory responses were significantly enriched in up-regulated genes at different stages of PAD(p < 0.05), revealing a significant correlation between immune/inflammatory responses and disease progression. Moreover, immune-related pathways such as Toll-like receptor signaling and natural killer cell mediated cytotoxicity were particularly enriched in intermediate and advanced lesions (P < 0.05), highlighting their pathogenic significance during disease progression.

Conclusion

Lines of evidence revealed in this study not only support previous hypotheses, primarily based on studies of animal models and other types of arterial disease, that inflammatory responses may influence the development of PAD, but also permit the recognition of a wide spectrum of immune/inflammatory genes that can serve as signatures for disease progression in PAD. Further studies of these signature molecules may eventually allow us to develop more sophisticated protocols for pharmaceutical interventions.
Collagen I but not Matrigel matrices provide an MMP-dependent barrier to ovarian cancer cell penetration
Authors: Sodek Katharine L , Brown Theodore J , Ringuette Maurice J
Published: August 2008

Background

The invasive potential of cancer cells is usually assessedin vitrousing Matrigel as a surrogate basement membrane. Yet cancer cell interaction with collagen I matrices is critical, particularly for the peritoneal metastatic route undertaken by several cancer types including ovarian. Matrix metalloprotease (MMP) activity is important to enable cells to overcome the barrier constraints imposed by basement membranes and stromal matricesin vivo. Our objective was to compare matrices reconstituted from collagen I and Matrigel as representative barriers for ovarian cancer cell invasion.

Methods

The requirement of MMP activity for ovarian cancer cell penetration of Matrigel and collagen matrices was assessed in 2D transwell and 3D spheroid culture systems.

Results

The broad range MMP inhibitor GM6001 completely prevented cell perforation of polymerised collagen I-coated transwell membranes. In contrast, GM6001 decreased ES-2 cell penetration of Matrigel by only ~30% and had no effect on HEY cell Matrigel penetration. In 3D culture, ovarian cancer cells grown as spheroids also migrated into surrounding Matrigel matrices despite MMP blockade. In contrast, MMP activity was required for invasion into 3D matrices of collagen I reconstituted from acid-soluble rat-tail collagen I, but not from pepsin-extracted collagen I (Vitrogen/Purecol), which lacks telopeptide regions.

Conclusion

Matrigel does not form representative barriers to ovarian cancer cells in either 2D or 3D culture systems. Our findings support the use of collagen I rather than Matrigel as a matrix barrier for invasion studies to better approximate critical interactions and events associated with peritoneal metastasis.
Charting calcium-regulated apoptosis pathways using chemical biology: role of calmodulin kinase II
Authors: Olofsson Maria Hägg , Havelka Aleksandra Mandic , Brnjic Slavica , Shoshan Maria C , Linder Stig
Published: August 2008

Background

Intracellular free calcium ([Ca2+]i) is a key element in apoptotic signaling and a number of calcium-dependent apoptosis pathways have been described. We here used a chemical biology strategy to elucidate the relative importance of such different pathways.

Results

A set of 40 agents ("bioprobes") that induce apoptosis was first identified by screening of a chemical library. Using p53, AP-1, NFAT and NF-κB reporter cell lines, these bioprobes were verified to induce different patterns of signaling. Experiments using the calcium chelator BAPTA-AM showed that Ca2+was involved in induction of apoptosis by the majority of the bioprobes and that Ca2+was in general required several hours into the apoptosis process. Further studies showed that the calmodulin pathway was an important mediator of the apoptotic response. Inhibition of calmodulin kinase II (CaMKII) resulted in more effective inhibition of apoptosis compared to inhibition of calpain, calcineurin/PP2B or DAP kinase. We used one of the bioprobes, the plant alkaloid helenalin, to study the role of CaMKII in apoptosis. Helenalin induced CaMKII, ASK1 and Jun-N-terminal kinase (JNK) activity, and inhibition of these kinases inhibited apoptosis.

Conclusion

Our study shows that calcium signaling is generally not an early event during the apoptosis process and suggests that a CaMKII/ASK1 signaling mechanism is important for sustained JNK activation and apoptosis by some types of stimuli.
Alternating evolutionary pressure in a genetic algorithm facilitates protein model selection
Authors: Offman Marc N , Tournier Alexander L , Bates Paul A
Published: August 2008

Background

Automatic protein modelling pipelines are becoming ever more accurate; this has come hand in hand with an increasingly complicated interplay between all components involved. Nevertheless, there are still potential improvements to be made in template selection, refinement and protein model selection.

Results

In the context of an automatic modelling pipeline, we analysed each step separately, revealing several non-intuitive trends and explored a new strategy for protein conformation sampling using Genetic Algorithms (GA). We apply the concept of alternating evolutionary pressure (AEP), i.e. intermediate rounds within the GA runs where unrestrained, linear growth of the model populations is allowed.

Conclusion

This approach improves the overall performance of the GA by allowing models to overcome local energy barriers. AEP enabled the selection of the best models in 40% of all targets; compared to 25% for a normal GA.
An approach to analyse the specific impact of rapamycin on mRNA-ribosome association
Authors: Genolet Raphael , Araud Tanguy , Maillard Laetitia , Jaquier-Gubler Pascale , Curran Joseph
Published: August 2008

Background

Recent work, using both cell culture model systems and tumour derived cell lines, suggests that the differential recruitment into polysomes of mRNA populations may be sufficient to initiate and maintain tumour formation. Consequently, a major effort is underway to use high density microarray profiles to establish molecular fingerprints for cells exposed to defined drug regimes. The aim of these pharmacogenomic approaches is to provide new information on how drugs can impact on the translational read-out within a defined cellular background.

Methods

We describe an approach that permits the analysis of de-novo mRNA-ribosome association in-vivo during short drug exposures. It combines hypertonic shock, polysome fractionation and high-throughput analysis to provide a molecular phenotype of translationally responsive transcripts. Compared to previous translational profiling studies, the procedure offers increased specificity due to the elimination of the drugs secondary effects (e.g. on the transcriptional read-out). For this pilot "proof-of-principle" assay we selected the drug rapamycin because of its extensively studied impact on translation initiation.

Results

High throughput analysis on both the light and heavy polysomal fractions has identified mRNAs whose re-recruitment onto free ribosomes responded to short exposure to the drug rapamycin. The results of the microarray have been confirmed using real-time RT-PCR. The selective down-regulation of TOP transcripts is also consistent with previous translational profiling studies using this drug.

Conclusion

The technical advance outlined in this manuscript offers the possibility of new insights into mRNA features that impact on translation initiation and provides a molecular fingerprint for transcript-ribosome association in any cell type and in the presence of a range of drugs of interest. Such molecular phenotypes defined pre-clinically may ultimately impact on the evaluation of a particular drug in a living cell.
The Alternative Splicing Mutation Database: a hub for investigations of alternative splicing using mutational evidence
Authors: Bechtel Jason M , Rajesh Preeti , Ilikchyan Irina , Deng Ying , Mishra Pankaj K , Wang Qi , Wu Xiaochun , Afonin Kirill A , Grose William E , Wang Ye , Khuder Sadik , Fedorov Alexei
Published: August 2008

Background

Some mutations in the internal regions of exons occur within splicing enhancers and silencers, influencing the pattern of alternative splicing in the corresponding genes. To understand how these sequence changes affect splicing, we created a database of these mutations.

Findings

The Alternative Splicing Mutation Database (ASMD) serves as a repository for all exonic mutations not associated with splicing junctions that measurably change the pattern of alternative splicing. In this initial published release (version 1.2), only human sequences are present, but the ASMD will grow to include other organisms, (see Availability and requirements section for the ASMD web address).

This relational database allows users to investigate connections between mutations and features of the surrounding sequences, including flanking sequences, RNA secondary structures and strengths of splice junctions. Splicing effects of the mutations are quantified by the relative presence of alternative mRNA isoforms with and without a given mutation. This measure is further categorized by the accuracy of the experimental methods employed. The database currently contains 170 mutations in 66 exons, yet these numbers increase regularly.

We developed an algorithm to derive a table of oligonucleotide Splicing Potential (SP) values from the ASMD dataset. We present the SP concept and tools in detail in our corresponding article.

Conclusion

The current data set demonstrates that mutations affecting splicing are located throughout exons and might be enriched within local RNA secondary structures. Exons from the ASMD have below average splicing junction strength scores, but the difference is small and is judged not to be significant.
Intervertebral disc cells as competent phagocytes in vitro: implications for cell death in disc degeneration
Authors: Jones Philip , Gardner Lucy , Menage Janis , Williams Gwyn T , Roberts Sally
Published: August 2008

Introduction

Apoptosis has been reported to occur in the intervertebral disc. Elsewhere in the body, apoptotic cells are cleared from the system via phagocytosis by committed phagocytes such as macrophages, reducing the chance of subsequent inflammation. These cells, however, are not normally present in the disc. We investigated whether disc cells themselves can be induced to become phagocytic and so have the ability to ingest and remove apoptotic disc cells, minimising the damage to their environment.

Method

Bovine nucleus pulposus cells from caudal intervertebral discs were grown in culture and exposed to both latex particles (which are ingested by committed phagocytes) and apoptotic cells. Their response was monitored via microscopy, including both fluorescent and video microscopy, and compared with that seen by cell lines of monocytes/macrophages (THP-1 and J774 cells), considered to be committed phagocytes, in addition to a nonmacrophage cell line (L929 fibroblasts). Immunostaining for the monocyte/macrophage marker, CD68, was also carried out.

Results

Disc cells were able to ingest latex beads at least as efficiently, if not more so, than phagocytic THP-1 and J774 cells. Disc cells ingested a greater number of beads per cell than the committed phagocytes in a similar time scale. In addition, disc cells were able to ingest apoptotic cells when cocultured in monolayer with a UV-treated population of HeLa cells. Apoptotic disc cells, in turn, were able to stimulate phagocytosis by the committed macrophages. CD68 immunostaining was strong for THP-1 cells but negligible for disc cells, even those that had ingested beads.

Conclusion

In this study, we have shown that intervertebral disc cells are capable of behaving as competent phagocytes (that is, ingesting latex beads) and apoptotic cells. In terms of number of particles, they ingest more than the monocyte/macrophage cells, possibly due to their greater size. The fact that disc cells clearly can undergo phagocytosis has implications for the intervertebral disc in vivo. Here, where cell death is reported to be common yet there is normally no easy access to a macrophage population, the endogenous disc cells may be encouraged to undergo phagocytosis (for example, of neighbouring cells within cell clusters).
Vital Statistics, and Observations on the first thousand Female Patients admitted into the Somerset County Lunatic Asylum; the Results compared with an equal number of Male Patients, together with an Analysis of the Causes of Death in both Sexes
Authors: Boyd R.
Published: August 2008

In a previous number (39) of this Journal are recorded the statistics of the first thousand cases of male patients admitted into the Somerset Lunatic Asylum, with an analysis of the causes of death. The same method is followed in the present communication with regard to the first thousand female admissions; a comparison is made between the sexes, and a general review from some authorities on these subjects.
On some of the Causes of Insanity
Authors: Maudsley Henry
Published: August 2008

It is not an easy matter, at least I have not found it so in my experience, when brought face to face with an actual case of in sanity, and asked to state the cause of it, to do so definitely and satisfactorily. The uncertainty springs from the fact that, in the great majority of cases, there has been a concurrence of co-operating conditions, not one single effective cause. Two persons are exposed to a similar heavy mental shock: one of them is driven mad by it, but the other is not. Can we say then that the madness has been produced by a moral cause? Not accurately so; for in the former case there has been some innate vice of nervous constitution, some predisposition of it to disease, whereby insanity has been produced by a cause which has had no such ill effect in the latter case. The entire causes have not, then, been in reality the same. And what we have to bear in mind is, that all the conditions which conspire to the production of an effect, whether visibly active or seemingly passive, are alike causes, alike agents, and that therefore all the conditions, whether they are in the patient himself or in the circum stances of life in which he is placed, which in a given case co-operate in the production of insanity, must properly be viewed as its causes. Mental derangement sometimes appears as the natural issue of all the precedent conditions of life, mental and bodily, the outcome of the individual character as affected by certain circumstances; the germs of the disease have been latent in the foundations of the character, and the final outbreak is but the explosion of a long train of antecedent preparations. In vain, then, is it in many cases to attempt to fix accurately on a single cause, moral or physical; a common mistake on the part of those, who think to do so being to fix upon what is really an early symptom of the disease as the supposed cause of it. Religion, self-abuse, intemperance, have all at times been put down as the causes of mental derangement, when they were really morbid symptoms.
The Medical Treatment of Insanity
Authors: Clouston T. S.
Published: August 2008

Might it not be possible greatly to extend our knowledge of the effects of medicines in insanity by treating similar classes of cases in all asylums in a similar way for a certain time? Might we not in this way arrive at a much greater degree of scientific accuracy in our treatment of certain forms of insanity than we are able to practise at present?