bluuebunny/crossref_metadata_2025 huggingface dataset:
Genomic and Biological Characterization of Exon 4 KRAS Mutations in Human Cancer
Authors: Janakiraman Manickam , Vakiani Efsevia , Zeng Zhaoshi , Pratilas Christine A. , Taylor Barry S. , Chitale Dhananjay , Halilovic Ensar , Wilson Manda , Huberman Kety , Ricarte Filho Julio Cezar , Persaud Yogindra , Levine Douglas A. , Fagin James A. , Jhanwar Suresh C. , Mariadason John M. , Lash Alex , Ladanyi Marc , Saltz Leonard B. , Heguy Adriana , Paty Philip B. , Solit David B.
Published: June 2010

Mutations in RAS proteins occur widely in human cancer. Prompted by the confirmation of KRAS mutation as a predictive biomarker of response to epidermal growth factor receptor (EGFR)–targeted therapies, limited clinical testing for RAS pathway mutations has recently been adopted. We performed a multiplatform genomic analysis to characterize, in a nonbiased manner, the biological, biochemical, and prognostic significance of Ras pathway alterations in colorectal tumors and other solid tumor malignancies. Mutations in exon 4 of KRAS were found to occur commonly and to predict for a more favorable clinical outcome in patients with colorectal cancer. Exon 4 KRAS mutations, all of which were identified at amino acid residues K117 and A146, were associated with lower levels of GTP-bound RAS in isogenic models. These same mutations were also often accompanied by conversion to homozygosity and increased gene copy number, in human tumors and tumor cell lines. Models harboring exon 4 KRAS mutations exhibited mitogen-activated protein/extracellular signal-regulated kinase kinase dependence and resistance to EGFR-targeted agents. Our findings suggest that RAS mutation is not a binary variable in tumors, and that the diversity in mutant alleles and variability in gene copy number may also contribute to the heterogeneity of clinical outcomes observed in cancer patients. These results also provide a rationale for broader KRAS testing beyond the most common hotspot alleles in exons 2 and 3. Cancer Res; 70(14); 5901–11. ©2010 AACR.
Development of a Lung Cancer Therapeutic Based on the Tumor Suppressor MicroRNA-34
Authors: Wiggins Jason F. , Ruffino Lynnsie , Kelnar Kevin , Omotola Michael , Patrawala Lubna , Brown David , Bader Andreas G.
Published: June 2010

Tumor suppressor microRNAs (miRNA) provide a new opportunity to treat cancer. This approach, “miRNA replacement therapy,” is based on the concept that the reintroduction of miRNAs depleted in cancer cells reactivates cellular pathways that drive a therapeutic response. Here, we describe the development of a therapeutic formulation using chemically synthesized miR-34a and a lipid-based delivery vehicle that blocks tumor growth in mouse models of non–small-cell lung cancer. This formulation is effective when administered locally or systemically. The antioncogenic effects are accompanied by an accumulation of miR-34a in the tumor tissue and downregulation of direct miR-34a targets. Intravenous delivery of formulated miR-34a does not induce an elevation of cytokines or liver and kidney enzymes in serum, suggesting that the formulation is well tolerated and does not induce an immune response. The data provide proof of concept for the systemic delivery of a synthetic tumor suppressor mimic, obviating obstacles associated with viral-based miRNA delivery and facilitating a rapid route for miRNA replacement therapy into the clinic. Cancer Res; 70(14); 5923–30. ©2010 AACR.
Breast Carcinoma–Associated Fibroblasts Rarely Contain p53 Mutations or Chromosomal Aberrations
Authors: Hosein Abdel Nasser , Wu Min , Arcand Suzanna L. , Lavallée Sylvie , Hébert Josée , Tonin Patricia N. , Basik Mark
Published: June 2010

It has become increasingly clear that the cells within the tumor microenvironment play a critical role in cancer growth and metastasis. Studies in experimental models suggest that carcinoma-associated fibroblasts (CAF) differ from normal fibroblasts and are capable of promoting cancer progression through a variety of mechanisms. At present, a definitive view is lacking on whether genomic abnormalities are present and whether they might underlie the observed phenotypic differences. This study reports the molecular analysis of the largest series of breast CAFs reported to date, with an array comparative genomic hybridization–based DNA copy number analysis of cultured CAFs derived from 25 freshly resected human breast cancers. We found DNA copy number changes consisting of the whole arm of chromosomes 6p and 9p plus interstitial 4q loss in only one sample. No abnormalities were observed in non–tumor-associated fibroblast counterparts. Karyotyping of the same CAF revealed further chromosomal abnormalities, which included clonal loss of chromosomes, chromosomal duplications, and less frequent chromosomal rearrangements. These abnormalities were not associated with alterations in the global gene expression profile of this particular CAF, relative to its non–tumor-associated fibroblast counterpart. Moreover, this particular patient's CAF also displayed the only p53 mutation in the cohort, the first time such a mutation has been reported in freshly cultured human CAFs. These findings argue that the procancerous effects of CAFs are unlikely to be due to DNA copy number–type genomic abnormalities in the CAFs themselves. As such, breast CAFs should be mainly regarded as genomically stable cellular constituents that exist within complex cancer microenvironments. Cancer Res; 70(14); 5770–7. ©2010 AACR.
Depletion of Tumor-Associated Macrophages Enhances the Effect of Sorafenib in Metastatic Liver Cancer Models by Antimetastatic and Antiangiogenic Effects
Authors: Zhang Wei , Zhu Xiao-Dong , Sun Hui-Chuan , Xiong Yu-Quan , Zhuang Peng-Yuan , Xu Hua-Xiang , Kong Ling-Qun , Wang Lu , Wu Wei-Zhong , Tang Zhao-You
Published: June 2010

Purpose: To investigate the role of macrophages in tumor progression under sorafenib treatment and to explore whether combination of drugs that deplete macrophages improved the antitumor effect of sorafenib.

Experimental Design: Tumor growth, lung metastasis, and tumor angiogenesis were observed in HCCLM3-R and SMMC7721, two human hepatocellular carcinoma xenograft nude mouse models, when treated with sorafenib (30 mg/kg daily, n = 6 per group) or a vehicle as control. Macrophage infiltration was measured in the peripheral blood and in sorafenib-treated tumor by immunohistochemistry and flow cytometry with F4/80 antibody and CD11b antibody. The effect of macrophage depletion on tumor angiogenesis and metastasis after sorafenib treatment, using two drug target macrophages, zoledronic acid (ZA) and clodrolip, was measured in the two models of hepatocellular carcinoma.

Results: Although sorafenib significantly inhibited tumor growth and lung metastasis, it induced a significant increase in peripheral recruitment and intratumoral infiltration of F4/80- and CD11b-positive cells, which was accompanied with elevation of colony-stimulating factor-1, stromal-derived factor 1α, and vascular endothelial growth factor in the tumor and elevation of plasma colony-stimulating factor-1 and mouse vascular endothelial growth factor in peripheral blood, suggesting the role of macrophages in tumor progression under sorafenib treatment. Depletion of macrophages by clodrolip or ZA in combination with sorafenib significantly inhibited tumor progression, tumor angiogenesis, and lung metastasis compared with mice treated with sorafenib alone. ZA was more effective than clodrolip.

Conclusions: Macrophages may have an important role in tumor progression under sorafenib treatment. ZA is promising when combined with sorafenib to enhance its antitumor effect. Clin Cancer Res; 16(13); 3420–30. ©2010 AACR.
Chemotherapy-Induced Activation of ADAM-17: A Novel Mechanism of Drug Resistance in Colorectal Cancer
Authors: Kyula Joan N. , Van Schaeybroeck Sandra , Doherty Joanne , Fenning Catherine S. , Longley Daniel B. , Johnston Patrick G.
Published: June 2010

Purpose: We have shown previously that exposure to anticancer drugs can trigger the activation of human epidermal receptor survival pathways in colorectal cancer (CRC). In this study, we examined the role of ADAMs (a disintegrin and metalloproteinases) and soluble growth factors in this acute drug resistance mechanism.

Experimental Design: In vitro and in vivo models of CRC were assessed. ADAM-17 activity was measured using a fluorometric assay. Ligand shedding was assessed by ELISA or Western blotting. Apoptosis was assessed by flow cytometry and Western blotting.

Results: Chemotherapy (5-fluorouracil) treatment resulted in acute increases in transforming growth factor-α, amphiregulin, and heregulin ligand shedding in vitro and in vivo that correlated with significantly increased ADAM-17 activity. Small interfering RNA–mediated silencing and pharmacologic inhibition confirmed that ADAM-17 was the principal ADAM involved in this prosurvival response. Furthermore, overexpression of ADAM-17 significantly decreased the effect of chemotherapy on tumor growth and apoptosis. Mechanistically, we found that ADAM-17 not only regulated phosphorylation of human epidermal receptors but also increased the activity of a number of other growth factor receptors, such as insulin-like growth factor-I receptor and vascular endothelial growth factor receptor.

Conclusions: Chemotherapy acutely activates ADAM-17, which results in growth factor shedding, growth factor receptor activation, and drug resistance in CRC tumors. Thus, pharmacologic inhibition of ADAM-17 in conjunction with chemotherapy may have therapeutic potential for the treatment of CRC. Clin Cancer Res; 16(13); 3378–89. ©2010 AACR.
Prognostic Significance of TRAIL Signaling Molecules in Stage II and III Colorectal Cancer
Authors: McLornan Donal P. , Barrett Helen L. , Cummins Robert , McDermott Ultan , McDowell Cliona , Conlon Susie J. , Coyle Victoria M. , Van Schaeybroeck Sandra , Wilson Richard , Kay Elaine W. , Longley Daniel B. , Johnston Patrick G.
Published: June 2010

Purpose: We previously found that cellular FLICE-inhibitory protein (c-FLIP), caspase 8, and tumor necrosis factor–related apoptosis-inducing ligand (TRAIL) receptor 2 (DR5) are major regulators of cell viability and chemotherapy-induced apoptosis in colorectal cancer. In this study, we determined the prognostic significance of c-FLIP, caspase 8, TRAIL and DR5 expression in tissues from patients with stage II and III colorectal cancer.

Experimental Design: Tissue microarrays were constructed from matched normal and tumor tissue derived from patients (n = 253) enrolled in a phase III trial of adjuvant 5-fluorouracil–based chemotherapy versus postoperative observation alone. TRAIL, DR5, caspase 8, and c-FLIP expression levels were determined by immunohistochemistry.

Results: Colorectal tumors displayed significantly higher expression levels of c-FLIP (P < 0.001), caspase 8 (P = 0.01), and DR5 (P < 0.001), but lower levels of TRAIL (P < 0.001) compared with matched normal tissue. In univariate analysis, higher TRAIL expression in the tumor was associated with worse overall survival (P = 0.026), with a trend to decreased relapse-free survival (RFS; P = 0.06), and higher tumor c-FLIP expression was associated with a significantly decreased RFS (P = 0.015). Using multivariate predictive modeling for RFS in all patients and including all biomarkers, age, treatment, and stage, we found that the model was significant when the mean tumor c-FLIP expression score and disease stage were included (P < 0.001). As regards overall survival, the overall model was predictive when both TRAIL expression and disease stage were included (P < 0.001).

Conclusions: High c-FLIP and TRAIL expression may be independent adverse prognostic markers in stage II and III colorectal cancer and might identify patients most at risk of relapse. Clin Cancer Res; 16(13); 3442–51. ©2010 AACR.
Immunohistochemical Detection of EGFR Mutation Using Mutation-Specific Antibodies in Lung Cancer
Authors: Kitamura Atsuko , Hosoda Waki , Sasaki Eiichi , Mitsudomi Tetsuya , Yatabe Yasushi
Published: June 2010

Purpose: Patients with mutations of epidermal growth factor receptor (EGFR) receive more benefit from EGFR-tyrosine kinase inhibitor treatment. However, usually such treatment is used to treat advanced lung cancer and only small biopsy samples are available for mutational analysis. We used immunohistochemistry to examine recently developed antibodies specific to major hotspot mutations of L858R and DEL E746-A750.

Experimental Design: We used five series of lung cancers: 47 non–small cell lung cancers (NSCLC) to evaluate various types of EGFR mutations, a consecutive series of 238 NSCLCs to study the sensitivity and specificity, 11 NSCLCs with both EGFR mutation and amplification to examine the spatial distribution, 32 patients treated with gefitinib to compare clinical responses, and 15 NSCLCs to explore changes associated with acquired T790M mutation.

Results: Each antibody specifically recognized the corresponding mutation but also recognized other types of mutations. Overall specificity and sensitivity were 96% and 47%, respectively. The positive reaction showed heterogeneous distribution that agreed with the expression of the total EGFR molecule, part of which was associated with gene amplification. A clinical response to gefitinib treatment correlated with the reaction, although one of the two patients with a positive reaction responded well despite having the wild-type EGFR. Acquired T790M mutation did not change the reaction to the antibodies.

Conclusions: On some characteristics, the positive reaction to mutation-specific antibodies differs from the molecular EGFR mutation. Therefore, this study revealed that not all patients with EGFR mutations can be selected using these mutation-specific antibodies. Clin Cancer Res; 16(13); 3349–55. ©2010 AACR.
Sorafenib, a Multikinase Inhibitor, Enhances the Response of Melanoma to Regional Chemotherapy
Authors: Augustine Christina K. , Toshimitsu Hiroaki , Jung Sin-Ho , Zipfel Patricia A. , Yoo Jin S. , Yoshimoto Yasunori , Selim M. Angelica , Burchette James , Beasley Georgia M. , McMahon Nicole , Padussis James , Pruitt Scott K. , Ali-Osman Francis , Tyler Douglas S.
Published: June 2010

Melanoma responds poorly to standard chemotherapy due to its intrinsic chemoresistance. Multiple genetic and molecular defects, including an activating mutation in the BRaf kinase gene, are associated with melanoma, and the resulting alterations in signal transduction pathways regulating proliferation and apoptosis are thought to contribute to its chemoresistance. Sorafenib, a multikinase inhibitor that targets BRaf kinase, is Food and Drug Administration approved for use in advanced renal cell and hepatocellular carcinomas. Although sorafenib has shown little promise as a single agent in melanoma patients, recent clinical trials suggest that, when combined with chemotherapy, it may have more benefit. We evaluated the ability of sorafenib to augment the cytotoxic effects of melphalan, a regional chemotherapeutic agent, and temozolomide, used in systemic and regional treatment of melanoma, on a panel of 24 human melanoma-derived cell lines and in an animal model of melanoma. Marked differences in response to 10 μmol/L sorafenib alone were observed in vitro across cell lines. Response to sorafenib significantly correlated with extracellular signal-regulated kinase (ERK) downregulation and loss of Mcl-1 expression (P < 0.05). Experiments with the mitogen-activated protein kinase/ERK kinase inhibitor U0126 suggest a unique role for ERK downregulation in the observed effects. Sorafenib in combination with melphalan or temozolomide led to significantly improved responses in vitro (P < 0.05). In the animal model of melanoma, sorafenib in combination with regional melphalan or regional temozolomide was more effective than either treatment alone in slowing tumor growth. These results show that sorafenib in combination with chemotherapy provides a novel approach to enhance chemotherapeutic efficacy in the regional treatment of in-transit melanoma. Mol Cancer Ther; 9(7); 2090–101. ©2010 AACR.
A Genetically Encoded Multifunctional TRAIL Trimer Facilitates Cell-Specific Targeting and Tumor Cell Killing
Authors: Spitzer Dirk , McDunn Jonathan E. , Plambeck-Suess Stacey , Goedegebuure Peter S. , Hotchkiss Richard S. , Hawkins William G.
Published: June 2010

Tumor necrosis factor–related apoptosis-inducing ligand (TRAIL, Apo2L) has been shown to exhibit potent and specific apoptotic activity against tumor cells. Several TRAIL constructs have been tried in patients, and the molecule remains under active clinical investigation. Native and recombinant TRAIL must form a homotrimer to become biologically active. However, noncovalently associated TRAIL displays a high degree of sensitivity to degradation, which limits its therapeutic potential. To enforce trimerization of the recombinant protein, we developed a covalently linked TRAIL trimer (TR3) by genetic fusion. This molecular drug design conferred improved stability without altering the native killing ability of TRAIL. Target specificity was shown by blocking TR3 activity with soluble death receptor 5 (DR5-Fc). In addition, we have shown that TR3 is amenable to further, genetic modifications. The incorporation of additional functional domains to TR3, such as antibody fragments (scFvs) that allow for a more cell-specific delivery of the agent, is stoichiometrically controlled and inconsequential with regard to the bioactivity of TRAIL. As proof of this concept, TR3 activity was targeted to the mouse RBC membrane. TR3-decorated RBCs were effectively capable of target cell killing in a model of pancreatic cancer. TR3 represents a generally applicable platform tool to study basic mechanisms along the death receptor pathway. More importantly, the ability to target TR3 to a cell surface presents the opportunity to create a cancer-selective drug with fewer off-target toxicities and enhanced killing capacities. Mol Cancer Ther; 9(7); 2142–51. ©2010 AACR.
Combining the FLT3 Inhibitor PKC412 and the Triterpenoid CDDO-Me Synergistically Induces Apoptosis in Acute Myeloid Leukemia with the Internal Tandem Duplication Mutation
Authors: Ahmad Rehan , Liu Suiyang , Weisberg Ellen , Nelson Erik , Galinsky Ilene , Meyer Colin , Kufe Donald , Kharbanda Surender , Stone Richard
Published: June 2010

Mutations of the FLT3 receptor tyrosine kinase consisting of internal tandem duplications (ITD) have been detected in blasts from 20% to 30% of patients with acute myeloid leukemia (AML) and are associated with a poor prognosis. FLT3/ITD results in constitutive autophosphorylation of the receptor and factor-independent survival in leukemia cell lines. The C-28 methyl ester of the oleane triterpenoid (CDDO-Me) is a multifunctional molecule that induces apoptosis of human myeloid leukemia cells. Here, we report that CDDO-Me blocks targeting of NFκB to the nucleus by inhibiting IκB kinase β–mediated phosphorylation of IκBα. Moreover, CDDO-Me blocked constitutive activation of the signal transducer and activator of transcription 3. We report the potent and selective antiproliferative effects of CDDO-Me on FLT3/ITD-positive myeloid leukemia cell lines and primary AML cells. The present studies show that CDDO-Me treatment results in caspase-3–mediated induction of apoptosis of FLT3/ITD-expressing cells and its antiproliferative effects are synergistic with PKC412, a FLT3-tyrosine kinase inhibitor currently in clinical trials. Taken together, our studies indicate that CDDO-Me greatly enhanced the efficacy of the FLT3 inhibitor PKC412, suggesting that combining two separate pathway inhibitors might be a viable therapeutic strategy for AML associated with a FLT3/ITD mutation. Mol Cancer Res; 8(7); 986–93. ©2010 AACR.
Optical Measurement of Rectal Microvasculature as an Adjunct to Flexible Sigmoidosocopy: Gender-Specific Implications
Authors: Roy Hemant K. , Gomes Andrew J. , Ruderman Sarah , Bianchi Laura K. , Goldberg Michael J. , Stoyneva Valentina , Rogers Jeremy D. , Turzhitsky Vladimir , Kim Young , Yen Eugene , Jameel Mohammed , Bogojevic Andrej , Backman Vadim
Published: June 2010

Flexible sigmoidoscopy is a robust, clinically validated, and widely available colorectal cancer screening technique that is currently sanctioned by major guideline organizations. Given that endoscopic visualization is generally limited to the distal third of the colon and women tend to have a proclivity for proximal lesions, the flexible sigmoidoscopy performance is markedly inferior in women than in men. Our group has shown that by using a novel light-scattering approach, we were able to detect an early increase in blood supply (EIBS) in the distal colonic mucosa, which served as a marker of field carcinogenesis and, hence, proximal neoplasia. Therefore, we sought to ascertain whether rectal EIBS would improve flexible sigmoidoscopy, especially in women. A polarization-gated spectroscopy fiber-optic probe was used to assess EIBS in the endoscopically normal rectum (n = 366). When compared with gender-matched neoplasia-free controls, females with advanced proximal neoplasia (n = 10) had a robust (60%; P = 0.002) increase in rectal mucosal oxyhemoglobin content whereas the effect size in males was less marked (33%; P = 0.052). In women, addition of rectal oxyhemoglobin tripled the sensitivity for advanced neoplasia over flexible sigmoidoscopy alone. Indeed, the performance characteristics seemed to be excellent (sensitivity, 100%; specificity, 76.8%; positive predictive value, 32.6%; and negative predictive value, 100%). A variety of nonneoplastic factors were assessed and did not confound the relationship between rectal EIBS and advanced neoplasia. Therefore, using rectal EIBS in combination with flexible sigmoidoscopy mitigated the gender gap and may allow flexible sigmoidoscopy to be considered as a viable colorectal cancer screening test in women. Cancer Prev Res; 3(7); 844–51. ©2010 AACR.
One Year Recurrence of Aberrant Crypt Foci
Authors: Pinsky Paul F. , Fleshman James , Mutch Matt , Rall Christopher , Charabaty Aline , Seligson David , Dry Sarah , Umar Asad , Schoen Robert E.
Published: June 2010

Aberrant crypt foci (ACF) are putative precursors of colorectal adenomas and have been postulated as a potential biomarker for colorectal cancer. Few studies have followed subjects after ACF removal to monitor recurrence. Subjects enrolled in the Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial were recruited for a study of ACF. A standardized protocol using magnified endoscopy and mucosal staining with methylene blue was implemented to detect rectal ACF. After removal of all baseline ACF, subjects returned 1 year later and recurrent ACF were observed and biopsied. A total of 434 of 505 (86%) subjects observed at baseline returned for the year 1 exam. The mean number of ACF at year 1 was strongly correlated with the number at baseline; subjects with 0, 1, 2 to 3, 4 to 6, and 7+ ACF at baseline had a mean of 1.2, 1.4, 1.7, 3.0, and 5.5 ACF, respectively, at year 1. ACF prevalence and mean count at year 1 (61% and 1.93, respectively), were only slightly lower than the corresponding values at year 0 (69% and 2.25, respectively). The locations of ACF at year 1 and baseline were significantly correlated. Of 96 ACF assessed for histology, 70 (73%) were hyperplastic and none were dysplastic. After removal of ACF at baseline, ACF counts 1 year later were only slightly reduced and were significantly correlated with the baseline ACF count. The results of this study do not support a role for ACF in clinical practice. Cancer Prev Res; 3(7); 839–43. ©2010 AACR.
Screening for Phenylketonuria
Authors: Bodamer Olaf A.
Published: June 2010

Newborn screening for phenylketonuria (PKU) started with Robert Guthrie (1916–1995) who developed the bacterial inhibition test for the semiquantitative analysis of phenylalanine, which was the first test suitable for high throughput analysis. In addition, he introduced the ‘Guthrie filter card’ as a transport medium for dried blood which is still used today. Realizing the potential of his approach for early diagnosis and the fact that a low-phenylalanine diet prevented the neurological sequelae of untreated PKU, Robert Guthrie became the first and utmost advocate of newborn screening for PKU. Following the first PKU newborn screening program in Buffalo, N.Y., USA, in the 1960s, additional newborn screening programs were initiated around the world in the 1960s and 1970s. Newborn screening has since been recognized as an important public health measure, and most countries have ongoing newborn screening programs for PKU and other inborn errors of metabolism. Since the first programs, it has been recognized that early diagnosis of PKU and subsequent initiation of a low phenylalanine diet results in normal neurological outcomes – in contrast to the severe mental retardation in untreated PKU. Today’s newborn screening laboratories use photometric assays or tandem mass spectrometry for analysis of phenylalanine rather than the bacterial inhibition test. This has led to an increased number of cases with hyperphenylalaninemia that often do not require dietary treatment. Any elevated phenylalanine level in a neonate needs to be followed by a second specimen for repeat analysis of phenylalanine and tyrosine. Confirmation of elevated phenylalanine levels with low to normal tyrosine levels requires analysis of urine pterines and dihydropterine reductase activity in red cells to rule out an inborn error of biopterin metabolism. In any neonate whose blood phenylalanine levels exceed 6 mg/dl, a tetrahydrobiopterin loading test should be performed and dietary therapy should be initiated. The level at which dietary therapy is started may be different between the USA, UK and continental Europe.
Effects of Cholesterol and Its 24S-OH and 25-OH Oxysterols on Choline Acetyltransferase-Positive Neurons in Brain Slices
Authors: Ullrich Celine , Pirchl Michael , Humpel Christian
Published: June 2010

In Alzheimer’s disease (AD) neurons expressing the enzyme choline acetyltransferase (ChAT) degenerate and a loss of cholinergic activity directly correlates with cognitive decline. Recent studies have suggested that cholesterol plays a role in AD. The aim of the present study was to explore if cholesterol and its oxysterols, 24S-hydroxycholesterol (24S-OH Chol) and 25-hydroxycholesterol (25-OH Chol), affect ChAT-positive neurons in organotypic brain slices of the basal nucleus of Meynert (nBM). We showed that slices expressed approximately 140 ChAT-positive neurons/slice after 2 weeks when incubated with nerve growth factor (NGF). This number markedly decreased when incubated without NGF to approximately 20 neurons/slice. Cholesterol and 24S-OH Chol delayed this decrease in ChAT-positive neurons. In contrast, 25-OH Chol induced a decline in ChAT-positive neurons in 2-week-old slices within 4 days. The effects of cholesterol and its oxysterols were exhibited in a dose- and time-dependent way. Our results show that cholesterol and 24S-OH Chol delays the decrease in ChAT-positive neurons, while 25-OH Chol rapidly decreases ChAT expression, suggesting differential mechanisms on ChAT expression in cholinergic nBM neurons.
Stroke Awareness in Denmark
Authors: Truelsen Thomas , Krarup Lars-Henrik
Published: June 2010

Background: This is the first study to examine the awareness of major stroke symptoms and stroke risk factors among the general population in Denmark. Early recognition of stroke warning signs and means of reducing stroke occurrence could improve the treatment and prevention of stroke. Methods: A total of 3,520 subjects were invited to participate in the study, using a web-based, closed questionnaire. The recruitment was stratified by region, age and gender to represent the composition of the general Danish population aged 40+ years. Enrolment was competitive with a predefined target of 800 responses. The subjects were asked to rank the 4 most important acute stroke symptoms, and the 3 most important risk factors for stroke. Multivariable ordinal logistic regression was used for assessing factors associated with awareness of stroke symptoms and major stroke risk factors. Results: From December 12 to December 17, 2008, a total of 811 subjects (mean age: 58 years; SD: 11 years) were included, of which 405 (50%) were women. The 4 most frequently identified stroke symptoms were sudden onset of: speech disturbances (n = 636; 78.4%), facial palsy (n = 450; 55.5%), hemiparesis (n = 435; 53.6%) and visual disturbances (n = 429; 52.9%). The 3 most frequently identified stroke risk factors were: hypertension (n = 586; 72.3%), history of cerebrovascular disease (n = 401; 49.5%) and high concentration of cholesterol (n = 269; 33.2%). Compared with men, women had better knowledge of major stroke symptoms (OR = 1.33; 95% CI: 1.12–1.50) and stroke risk factors (OR = 1.33; 95% CI: 1.10–1.51) in multivariable analyses. Conclusions: The awareness of major stroke symptoms is insufficient. Whereas hypertension was the most often mentioned stroke risk factor, few subjects recognized tobacco smoking and diabetes mellitus as major risk factors for stroke.
RCAS-1 Serum and Tumor Levels in Head and Neck Squamous Cell Carcinoma
Authors: Brunner M. , Erovic B. , Heiduschka G. , Schneider S. , Pammer J. , Frantal S. , Thurnher D.
Published: June 2010

Background/Aims: RCAS-1 is a transmembrane protein that is involved in the evasion of host immune surveillance by tumor cells. It has been found to be a valuable prognostic and diagnostic marker in a number of different malignancies. The objective of the study was to analyze the potency of RCAS-1 as a biomarker in the serum of patients with head and neck squamous cell carcinoma (HNSCC). Methods: ELISA was performed with prospectively collected serum samples from 60 patients with HNSCC (taken at the time of diagnosis, after 3 and 12 months) and from 31 healthy controls. To correlate serum levels with RCAS-1 expression in the tumor, immunohistochemical staining of RCAS-1 was done using a tissue microarray. Results: Surprisingly, median sRCAS-1 levels were basically identical between tumor patients and controls. Interestingly, patients with low RCAS-1 values at the time of diagnosis had better disease-free survival. 62% of tumor samples expressed RCAS-1 but we could not demonstrate a correlation between protein expression and serum levels. Conclusion: This study was the first to correlate RCAS-1 levels in the serum and in the tumor of the same patients. RCAS-1 seems to have prognostic properties although larger studies will be necessary to fully evaluate its role in HNSCC.
Elevated Plasma Levels of Antimicrobial Polypeptides in Patients with Severe Sepsis
Authors: Berkestedt Ingrid , Herwald Heiko , Ljunggren Lennart , Nelson Axel , Bodelsson Mikael
Published: June 2010

We wanted to investigate if plasma levels of antimicrobial polypeptides (AMPs) are increased in severe sepsis and if they correlate with severity and mortality. Samples were collected from 31 sepsis patients at the intensive care unit. The Sequential Organ Failure Assessment (SOFA) score and 90-day mortality were registered, and inflammatory markers and AMP levels were measured by ELISA. A median SOFA score (13) and cardiovascular SOFA score (3) indicated multiorgan failure with severe circulatory derangement, and elevated cytokine levels indicated inflammatory activation. Levels of bactericidal/permeability-increasing protein, heparin-binding protein, α-defensins and lactoferrin but not LL-37 were elevated in sepsis patients compared with controls. Bactericidal/permeability-increasing protein levels correlated with mortality, with lower levels in survivors. Levels of all AMPs, except LL-37, positively correlated with the cardiovascular SOFA score. In conclusion, levels of several AMPs are increased in sepsis and correlate with circulatory derangement. This probably reflects neutrophil activation as part of an innate immune response.
Epicardial Perivascular Adipose-Derived Leptin Exacerbates Coronary Endothelial Dysfunction in Metabolic Syndrome via a Protein Kinase C-β Pathway
Authors: Payne Gregory A. , Borbouse Léna , Kumar Sajel , Neeb Zachary , Alloosh Mouhamad , Sturek Michael , Tune Johnathan D.
Published: June 2010

Objective—

Factors released by perivascular adipose tissue (PVAT) disrupt coronary endothelial function via phosphorylation of endothelial NO synthase by protein kinase C (PKC)-β. However, our understanding of how PVAT potentially contributes to coronary disease as a complication of obesity/metabolic syndrome (MetS) remains limited. The current study investigated whether PVAT-derived leptin impairs coronary vascular function via PKC-β in MetS.

Methods and Results—

Coronary arteries with and without PVAT were collected from lean or MetS Ossabaw miniature swine for isometric tension studies. Endothelial-dependent vasodilation to bradykinin was significantly reduced in MetS. PVAT did not affect bradykinin-mediated dilation in arteries from lean swine but significantly exacerbated endothelial dysfunction in arteries from MetS swine. PVAT-induced impairment was reversed by inhibition of either PKC-β with ruboxistaurin (Eli Lilly and Company, Indianapolis, Ind) or leptin receptor signaling with a recombinant, pegylated leptin antagonist. Western blot and immunohistochemical analyses demonstrated increased PVAT-derived leptin and coronary leptin receptor density with MetS. Coronary PKC-β activity was increased in both MetS arteries exposed to PVAT and lean arteries exposed to leptin. Finally, leptin-induced endothelial dysfunction was reversed by ruboxistaurin.

Conclusion—

Increases in epicardial PVAT leptin exacerbate coronary endothelial dysfunction in MetS via a PKC-β-dependent pathway. These findings implicate PVAT-derived leptin as a potential contributor to coronary atherogenesis in MetS.
Genetic Determinants of Major Blood Lipids in Pakistanis Compared With Europeans
Authors: Saleheen Danish , Soranzo Nicole , Rasheed Asif , Scharnagl Hubert , Gwilliam Rhian , Alexander Myriam , Inouye Michael , Zaidi Moazzam , Potter Simon , Haycock Philip , Bumpstead Suzanna , Kaptoge Stephen , Di Angelantonio Emanuele , Sarwar Nadeem , Hunt Sarah E. , Sheikh Nasir , Shah Nabi , Samuel Maria , Haider Shajjia Razi , Murtaza Muhammed , Thompson Alexander , Gobin Reeta , Butterworth Adam , Ahmad Usman , Hakeem Abdul , Zaman Khan Shah , Kundi Assadullah , Yaqoob Zia , Cheema Liaquat Ali , Qamar Nadeem , Faruqui Azhar , Mallick Nadeem Hayat , Azhar Muhammad , Samad Abdus , Ishaq Muhammad , Rasheed Syed Zahed , Jooma Rashid , Niazi Jawaid Hassan , Gardezi Ali Raza , Memon Nazir Ahmed , Ghaffar Abdul , Rehman Fazal-ur , Hoffmann Michael Marcus , Renner Wilfried , Kleber Marcus E. , Grammer Tanja B. , Stephens Jonathon , Attwood Anthony , Koch Kerstin , Hussain Mustafa , Kumar Kishore , Saleem Asim , Kumar Kishwar , Daood Muhammad Salman , Gul Aftab Alam , Abbas Shahid , Zafar Junaid , Shahid Faisal , Bhatti Shahzad Majeed , Ali Syed Saadat , Muhammad Fahim , Sagoo Gurdeep , Bray Sarah , McGinnis Ralph , Dudbridge Frank , Winkelmann Bernhard R. , Böehm Bernhard , Thompson Simon , Ouwehand Willem , März Winfried , Frossard Philippe , Danesh John , Deloukas Panos
Published: June 2010

Background—

Evidence is sparse about the genetic determinants of major lipids in Pakistanis.

Methods and Results—

Variants (n=45 000) across 2000 genes were assessed in 3200 Pakistanis and compared with 2450 Germans using the same gene array and similar lipid assays. We also did a meta-analysis of selected lipid-related variants in Europeans. Pakistani genetic architecture was distinct from that of several ethnic groups represented in international reference samples. Forty-one variants at 14 loci were significantly associated with levels of HDL-C, triglyceride, or LDL-C. The most significant lipid-related variants identified among Pakistanis corresponded to genes previously shown to be relevant to Europeans, such as
CETP
associated with HDL-C levels (rs711752;
P
<10
−13
),
APOA5/ZNF259
(rs651821;
P
<10
−13
) and
GCKR
(rs1260326;
P
<10
−13
) with triglyceride levels; and
CELSR2
variants with LDL-C levels (rs646776;
P
<10
−9
). For Pakistanis, these 41 variants explained 6.2%, 7.1%, and 0.9% of the variation in HDL-C, triglyceride, and LDL-C, respectively. Compared with Europeans, the allele frequency of rs662799 in
APOA5
among Pakistanis was higher and its impact on triglyceride concentration was greater (
P
-value for difference <10
−4
).

Conclusions—

Several lipid-related genetic variants are common to Pakistanis and Europeans, though they explain only a modest proportion of population variation in lipid concentration. Allelic frequencies and effect sizes of lipid-related variants can differ between Pakistanis and Europeans.
Circadian Rhythms in Patients With ST-Elevation Myocardial Infarction
Authors: Holmes David R. , Aguirre Frank V. , Aplin Richard , Lennon Ryan J. , Nestler David M. , Bell Malcolm R. , Rihal Charanjit S. , Ting Henry H.
Published: June 2010

Background—

Circadian rhythms with regard to time of symptom onset for patients with acute myocardial infarction have been observed, although their relationship to outcomes has been debated. We evaluated these rhythms in patients with ST-elevation myocardial infarction as a function of the 24-hour circadian cycle.

Methods and Results—

The relationship between onset of symptoms during the 24-hour circadian cycle and prehospital delays from symptom onset to hospital arrival, timeliness of reperfusion, and in-hospital death was assessed in 2143 patients with ST-elevation myocardial infarction presenting from 2004–2008 at 1 of 3 tertiary-care healthcare ST-elevation myocardial infarction systems. There was a significant association between time of onset and the circadian cycle, with the greatest percentage (39%) of patients experiencing onset between 8
am
and 3
pm
(
P
<0.001). Time of onset was associated with prehospital delay and timeliness of reperfusion. Patients with onset from 12
am
to 5:59
am
had median prehospital delays of 121 minutes versus 70 minutes from 12
pm
to 5:59
pm
(
P
<0.001). Patients with onset time from 12
am
to 5:59
am
had median door-to-balloon times of 75 minutes versus 60 minutes from 6
am
to 11:59
am
(
P
<0.001). Using multivariable modeling to control for baseline patient characteristics, prehospital delay, and timeliness of reperfusion, there was no significant association between time of symptom onset with in-hospital death.

Conclusions—

Patients with ST-elevation myocardial infarction exhibit significant circadian patterns in symptom onset, prehospital delay, and timeliness of reperfusion. Patients who develop symptoms from 12
am
to 5:59
am
present with longer prehospital delays and have longer door-to-balloon times. After multivariable adjustment, there was no significant association between circadian patterns of time of onset and in-hospital death.
T-Tubule Remodeling During Transition From Hypertrophy to Heart Failure
Authors: Wei Sheng , Guo Ang , Chen Biyi , Kutschke William , Xie Yu-Ping , Zimmerman Kathy , Weiss Robert M. , Anderson Mark E. , Cheng Heping , Song Long-Sheng
Published: June 2010

Rationale:

The transverse tubule (T-tubule) system is the ultrastructural substrate for excitation–contraction coupling in ventricular myocytes; T-tubule disorganization and loss are linked to decreased contractility in end stage heart failure (HF).

Objective:

We sought to examine (1) whether pathological T-tubule remodeling occurs early in compensated hypertrophy and, if so, how it evolves during the transition from hypertrophy to HF; and (2) the role of junctophilin-2 in T-tubule remodeling.

Methods and Results:

We investigated T-tubule remodeling in relation to ventricular function during HF progression using state-of-the-art confocal imaging of T-tubules in intact hearts, using a thoracic aortic banding rat HF model. We developed a quantitative T-tubule power (TT
power
) index to represent the integrity of T-tubule structure. We found that discrete local loss and global reorganization of the T-tubule system (leftward shift of TT
power
histogram) started early in compensated hypertrophy in left ventricular (LV) myocytes, before LV dysfunction, as detected by echocardiography. With progression from compensated hypertrophy to early and late HF, T-tubule remodeling spread from the LV to the right ventricle, and TT
power
histograms of both ventricles gradually shifted leftward. The mean LV TT
power
showed a strong correlation with ejection fraction and heart weight to body weight ratio. Over the progression to HF, we observed a gradual reduction in the expression of a junctophilin protein (JP-2) implicated in the formation of T-tubule/sarcoplasmic reticulum junctions. Furthermore, we found that JP-2 knockdown by gene silencing reduced T-tubule structure integrity in cultured adult ventricular myocytes.

Conclusions:

T-tubule remodeling in response to thoracic aortic banding stress begins before echocardiographically detectable LV dysfunction and progresses over the development of overt structural heart disease. LV T-tubule remodeling is closely associated with the severity of cardiac hypertrophy and predicts LV function. Thus, T-tubule remodeling may constitute a key mechanism underlying the transition from compensated hypertrophy to HF.
The (Pro)renin Receptor/ATP6AP2 is Essential for Vacuolar H
+
-ATPase Assembly in Murine Cardiomyocytes
Authors: Kinouchi Kenichiro , Ichihara Atsuhiro , Sano Motoaki , Sun-Wada Ge-Hong , Wada Yoh , Kurauchi-Mito Asako , Bokuda Kanako , Narita Tatsuya , Oshima Yoichi , Sakoda Mariyo , Tamai Yoshitaka , Sato Hiromu , Fukuda Keiichi , Itoh Hiroshi
Published: June 2010

Rationale
:

The (pro)renin receptor [(P)RR], encoded in
ATP6AP2
, plays a key role in the activation of local renin-angiotensin system (RAS). A truncated form of (P)RR, termed M8.9, was also found to be associated with the vacuolar H
+
-ATPase (V-ATPase), implicating a non–RAS-related function of ATP6AP2.

Objective
:

We investigated the role of (P)RR/ATP6AP2 in murine cardiomyocytes.

Methods and Results
:

Cardiomyocyte-specific ablation of
Atp6ap2
resulted in lethal heart failure; the cardiomyocytes contained RAB7- and lysosomal-associated membrane protein 2 (LAMP2)-positive multivesicular vacuoles, especially in the perinuclear regions. The myofibrils and mitochondria remained at the cell periphery. Cardiomyocyte death was accompanied by numerous autophagic vacuoles that contained undigested cellular constituents, as a result of impaired autophagic degradation. Notably, ablation of
Atp6ap2
selectively suppressed expression of the V
O
subunits of V-ATPase, resulting in deacidification of the intracellular vesicles. Furthermore, the inhibition of intracellular acidification by treatment with bafilomycin A1 or chloroquine reproduced the phenotype observed for the (P)RR/ATP6AP2-deficient cardiomyocytes.

Conclusions
:

Genetic ablation of
Atp6ap2
created a loss-of-function model for V-ATPase. The gene product of
ATP6AP2
is considered to act as in 2 ways: (1) as (P)RR, exerting a RAS-related function; and (2) as the V-ATPase-associated protein, exerting a non–RAS-related function that is essential for cell survival.
Perception of Weight-Bearing Distribution During Sit-to-Stand Tasks in Hemiparetic and Healthy Individuals
Authors: Brière Anabèle , Lauzière Séléna , Gravel Denis , Nadeau Sylvie
Published: June 2010

Background and Purpose—It is unknown whether hemiparetic individuals are aware of their weight-bearing asymmetry during sit-to-stand tasks. This study compared the error between hemiparetic and healthy individuals’ perception of weight-bearing and their actual weight-bearing distribution during the sit-to-stand task and analyzed the association between the knee extensor muscle strength and the weight-bearing distribution and perception.

Methods—Nineteen unilateral hemiparetic subjects and 15 healthy individuals participated in the study. They performed the sit-to-stand transfer on force platforms under different foot placements (spontaneous and symmetrical) and had to rate their perceived weight-bearing distribution at the lower limbs on a visual analog scale. The strength of the knee extensors was assessed with a Biodex dynamometer.

Results—The hemiparetic individuals presented greater weight-bearing asymmetry and errors of perception than the healthy individuals. Although no significant association was found between strength and weight-bearing perception, moderate associations were found between strength and weight-bearing distribution for both the spontaneous (r=0.75,P<0.01) and symmetrical (r=0.71,P<0.01) foot position conditions.

Conclusions—This study revealed that individuals with hemiparesis after a stroke do not perceive themselves as asymmetrical when executing the sit-to-stand transfer and that the knee extensor strength is a factor linked to their weight-bearing asymmetry, not to their perception.
Individual Differences in Nucleus Accumbens Dopamine Receptors Predict Development of Addiction-Like Behavior: A Computational Approach
Authors: Piray Payam , Keramati Mohammad Mahdi , Dezfouli Amir , Lucas Caro , Mokri Azarakhsh
Published: June 2010

Clinical and experimental observations show individual differences in the development of addiction. Increasing evidence supports the hypothesis that dopamine receptor availability in the nucleus accumbens (NAc) predisposes drug reinforcement. Here, modeling striatal-midbrain dopaminergic circuit, we propose a reinforcement learning model for addiction based on the actor-critic model of striatum. Modeling dopamine receptors in the NAc as modulators of learning rate for appetitive—but not aversive—stimuli in the critic—but not the actor—we define vulnerability to addiction as a relatively lower learning rate for the appetitive stimuli, compared to aversive stimuli, in the critic. We hypothesize that an imbalance in this learning parameter used by appetitive and aversive learning systems can result in addiction. We elucidate that the interaction between the degree of individual vulnerability and the duration of exposure to drug has two progressive consequences: deterioration of the imbalance and establishment of an abnormal habitual response in the actor. Using computational language, the proposed model describes how development of compulsive behavior can be a function of both degree of drug exposure and individual vulnerability. Moreover, the model describes how involvement of the dorsal striatum in addiction can be augmented progressively. The model also interprets other forms of addiction, such as obesity and pathological gambling, in a common mechanism with drug addiction. Finally, the model provides an answer for the question of why behavioral addictions are triggered in Parkinson's disease patients by D2 dopamine agonist treatments.
Sound Retrieval and Ranking Using Sparse Auditory Representations
Authors: Lyon Richard F. , Rehn Martin , Bengio Samy , Walters Thomas C. , Chechik Gal
Published: June 2010

To create systems that understand the sounds that humans are exposed to in everyday life, we need to represent sounds with features that can discriminate among many different sound classes. Here, we use a sound-ranking framework to quantitatively evaluate such representations in a large-scale task. We have adapted a machine-vision method, the passive-aggressive model for image retrieval (PAMIR), which efficiently learns a linear mapping from a very large sparse feature space to a large query-term space. Using this approach, we compare different auditory front ends and different ways of extracting sparse features from high-dimensional auditory images. We tested auditory models that use an adaptive pole–zero filter cascade (PZFC) auditory filter bank and sparse-code feature extraction from stabilized auditory images with multiple vector quantizers. In addition to auditory image models, we compare a family of more conventional mel-frequency cepstral coefficient (MFCC) front ends. The experimental results show a significant advantage for the auditory models over vector-quantized MFCCs. When thousands of sound files with a query vocabulary of thousands of words were ranked, the best precision at top-1 was 73% and the average precision was 35%, reflecting a 18% improvement over the best competing MFCC front end.