bluuebunny/crossref_metadata_2025 huggingface dataset:
Echocardiographic Assessment of Left Ventricular Function in Type 1 Gaucher's Disease
Authors: Koželj Mirta , Zver Samo , Zadnik Vesna
Published: July 2010

There is predominate opinion among physicians managing type 1 Gauchers' disease (GD) that cardiac involvement is not an issue in these patients. In order to follow this hypothesis, we prospectively investigated 15 adult imiglucerase-treated type 1 GD patients by echocardiography, Doppler, and tissue Doppler echocardiography. This was a case-controlled study with 18 matched healthy volunteers. The obtained data was correlated with the levels of NT-proBNP (brain natriuretic peptide). None of the GD patients had clinical signs of heart disease. In 3 of the 15 patients, we observed echocardiographic signs of aortic and mitral valve calcification. The left ventricular systolic function was within normal limits. Compared to the control group, there was no statistically significant difference observed in the most sensitive indices of left ventricular diastolic function, parameterEm(P=.095), andE/Emratio (P=.097), as demonstrated by tissue Doppler echocardiography. However, there was a positive correlation between theE/Emratio and NT-proBNP plasma levels (P=.009). In conclusion, the prospective echocardiographic study of type 1 GD patients did not validate any left ventricular dysfunction. But, theE/Emratio showed a strong statistical correlation with the most sensitive indicators of heart failure, NT-proBNP. Research on larger groups of patients and the usage of even more sensitive methods as strain-rate imaging will be necessary to confirm eventual myocardial involvement in GD patients.
Continuum Plate Theory and Atomistic Modeling to Find the Flexural Rigidity of a Graphene Sheet Interacting with a Substrate
Authors: Roberts M. W. , Clemons C. B. , Wilber J. P. , Young G. W. , Buldum A. , Quinn D. D.
Published: July 2010

Using a combination of continuum modeling, atomistic simulations, and numerical optimization, we estimate the flexural rigidity of a graphene sheet. We consider a rectangular sheet that is initially parallel to a rigid substrate. The sheet interacts with the substrate by van der Waals forces and deflects in response to loading on a pair of opposite edges. To estimate the flexural rigidity, we model the graphene sheet as a continuum and numerically solve an appropriate differential equation for the transverse deflection. This solution depends on the flexural rigidity. We then use an optimization procedure to find the value of the flexural rigidity that minimizes the difference between the numerical solutions and the deflections predicted by atomistic simulations. This procedure predicts a flexural rigidity of 0.26 nNnm=1.62 eV.
Tumor Cells Secrete Galectin-1 to Enhance Endothelial Cell Activity
Authors: Thijssen Victor L. , Barkan Batya , Shoji Hiroki , Aries Ingrid M. , Mathieu Véronique , Deltour Louise , Hackeng Tilman M. , Kiss Robert , Kloog Yoel , Poirier Françoise , Griffioen Arjan W.
Published: July 2010

Tumor angiogenesis is a key event in cancer progression. Here, we report that tumors can stimulate tumor angiogenesis by secretion of galectin-1. Tumor growth and tumor angiogenesis of different tumor models are hampered in galectin-1–null (gal-1−/−) mice. However, tumor angiogenesis is less affected when tumor cells express and secrete high levels of galectin-1. Furthermore, tumor endothelial cells in gal-1−/− mice take up galectin-1 that is secreted by tumor cells. Uptake of galectin-1 by cultured endothelial cells specifically promotes H-Ras signaling to the Raf/mitogen-activated protein kinase/extracellular signal-regulated kinase (Erk) kinase (Mek)/Erk cascade and stimulates endothelial cell proliferation and migration. Moreover, the activation can be blocked by galectin-1 inhibition as evidenced by hampered membrane translocation of H-Ras.GTP and impaired Raf/Mek/Erk phosphorylation after treatment with the galectin-1–targeting angiogenesis inhibitor anginex. Altogether, these data identify galectin-1 as a proangiogenic factor. These findings have direct implications for current efforts on galectin-1–targeted cancer therapies. Cancer Res; 70(15); 6216–24. ©2010 AACR.
Akt3-Mediated Resistance to Apoptosis in B-RAF–Targeted Melanoma Cells
Authors: Shao Yongping , Aplin Andrew E.
Published: July 2010

Melanoma cells are highly resistant to anoikis, a form of apoptosis induced in nonadherent/inappropriate adhesion conditions. Depleting B-RAF or the prosurvival Bcl-2 family protein Mcl-1 renders mutant B-RAF melanoma cells susceptible to anoikis. In this study, we examined the effect of targeting B-RAF on the survival of primary stage melanoma cells cultured in three-dimensional type I collagen gels, which partially mimics the dermal microenvironment. Depletion/inhibition of B-RAF with small interfering RNA or the mutant B-RAF inhibitor, PLX4720, induced apoptosis of mutant B-RAF melanoma cells in three-dimensional collagen. Apoptosis was dependent on two upregulated BH3-only proteins, Bim-EL and Bmf, and was inhibited by ectopic Mcl-1 expression. Akt3 activation has been associated with the survival of melanoma cells. Mutant B-RAF melanoma cells ectopically expressing a constitutively activated form of Akt3 or endogenously expressing mutant Akt3 were protected from apoptosis induced by B-RAF knockdown or PLX4720 treatment. Furthermore, intrinsically resistant metastatic melanoma cells displayed elevated Akt phosphorylation in three-dimensional collagen and were rendered susceptible to PLX4720 by Akt3 knockdown. Importantly, myristylated Akt3 prevented B-RAF targeting–induced upregulation of Bim-EL and Bmf in three-dimensional collagen and partially protected Mcl-1–depleted cells from apoptosis. These findings delineate how mutant B-RAF protects melanoma cells from apoptosis and provide insight into possible resistance mechanisms to B-RAF inhibitors. Cancer Res; 70(16); 6670–81. ©2010 AACR.
Fructose Induces Transketolase Flux to Promote Pancreatic Cancer Growth
Authors: Liu Haibo , Huang Danshan , McArthur David L. , Boros Laszlo G. , Nissen Nicholas , Heaney Anthony P.
Published: July 2010

Carbohydrate metabolism via glycolysis and the tricarboxylic acid cycle is pivotal for cancer growth, and increased refined carbohydrate consumption adversely affects cancer survival. Traditionally, glucose and fructose have been considered as interchangeable monosaccharide substrates that are similarly metabolized, and little attention has been given to sugars other than glucose. However, fructose intake has increased dramatically in recent decades and cellular uptake of glucose and fructose uses distinct transporters. Here, we report that fructose provides an alternative substrate to induce pancreatic cancer cell proliferation. Importantly, fructose and glucose metabolism are quite different; in comparison with glucose, fructose induces thiamine-dependent transketolase flux and is preferentially metabolized via the nonoxidative pentose phosphate pathway to synthesize nucleic acids and increase uric acid production. These findings show that cancer cells can readily metabolize fructose to increase proliferation. They have major significance for cancer patients given dietary refined fructose consumption, and indicate that efforts to reduce refined fructose intake or inhibit fructose-mediated actions may disrupt cancer growth. Cancer Res; 70(15); 6368–76. ©2010 AACR.
Combinatorial Chemoprevention Reveals a Novel Smoothened-Independent Role of GLI1 in Esophageal Carcinogenesis
Authors: Ilyas Sumera I. , DeMars Cathrine J. , Comba Andrea , Gainullin Vladimir G. , Rizvi Zaheer , Almada Luciana L. , Wang Kenneth , Lomberk Gwen , Fernández-Zapico Martin E. , Buttar Navtej S.
Published: July 2010

Reflux-induced injury promotes esophageal adenocarcinoma, one of the most rapidly increasing, highly lethal cancers in Western countries. Here, we investigate the efficacy of a combinatorial chemoprevention strategy for esophageal adenocarcinoma and characterize the underlying molecular mechanisms. Specifically, our approach involves the use of ursodeoxycholic acid (Urso) due to its ability to decrease injury-inducing bile salts in combination with Aspirin to mitigate the consequences of injury. We find that Urso-Aspirin combination reduces the risk of adenocarcinoma in vivo in animals with reflux, decreases the proliferation of esophageal adenocarcinoma cells, and downregulates a key cell cycle regulator, CDK2. Mechanistically, using cell growth, luciferase reporter, expression, and chromatin immunoprecipitation assays, we identify GLI1, a Hedgehog-regulated transcription factor, as a novel target of Urso-Aspirin combination. We show that GLI1 is upregulated during esophageal carcinogenesis, and GLI1 can bind to the CDK2 promoter and activate its expression. Although the Urso-Aspirin combination downregulates GLI1, the GLI1 overexpression not only abrogates the effect of this combination on proliferation but it also restores CDK-2 expression. These findings support that the chemopreventive effect of the Urso-Aspirin combination occurs, at least in part, through a novel GLI1-CDK2–dependent mechanism. To further understand the regulation of CDK2 by GLI1, both pharmacologic and RNAi-mediated approaches show that GLI1 is a transcriptional activator of CDK2, and this regulation occurs independent of Smoothened, the central transducer of the Hedgehog canonical pathway. Collectively, these results identify a novel GLI1-to-CDK2 pathway in esophageal carcinogenesis, which is a bona fide target for effective combinatorial chemoprevention with Urso and Aspirin. Cancer Res; 70(17); 6787–96. ©2010 AACR.
A KRAS-Variant in Ovarian Cancer Acts as a Genetic Marker of Cancer Risk
Authors: Ratner Elena , Lu Lingeng , Boeke Marta , Barnett Rachel , Nallur Sunitha , Chin Lena J. , Pelletier Cory , Blitzblau Rachel , Tassi Renata , Paranjape Trupti , Hui Pei , Godwin Andrew K. , Yu Herbert , Risch Harvey , Rutherford Thomas , Schwartz Peter , Santin Alessandro , Matloff Ellen , Zelterman Daniel , Slack Frank J. , Weidhaas Joanne B.
Published: July 2010

Ovarian cancer (OC) is the single most deadly form of women's cancer, typically presenting as an advanced disease at diagnosis in part due to a lack of known risk factors or genetic markers of risk. The KRAS oncogene and altered levels of the microRNA (miRNA) let-7 are associated with an increased risk of developing solid tumors. In this study, we investigated a hypothesized association between an increased risk of OC and a variant allele of KRAS at rs61764370, referred to as the KRAS-variant, which disrupts a let-7 miRNA binding site in this oncogene. Specimens obtained were tested for the presence of the KRAS-variant from nonselected OC patients in three independent cohorts, two independent ovarian case-control studies, and OC patients with hereditary breast and ovarian cancer syndrome (HBOC) as well as their family members. Our results indicate that the KRAS-variant is associated with more than 25% of nonselected OC cases. Further, we found that it is a marker for a significant increased risk of developing OC, as confirmed by two independent case-control analyses. Lastly, we determined that the KRAS-variant was present in 61% of HBOC patients without BRCA1 or BRCA2 mutations, previously considered uninformative, as well as in their family members with cancer. Our findings strongly support the hypothesis that the KRAS-variant is a genetic marker for increased risk of developing OC, and they suggest that the KRAS-variant may be a new genetic marker of cancer risk for HBOC families without other known genetic abnormalities. Cancer Res; 70(16); 6509–15. ©2010 AACR.
The Combination of Multiple Receptor Tyrosine Kinase Inhibitor and Mammalian Target of Rapamycin Inhibitor Overcomes Erlotinib Resistance in Lung Cancer Cell Lines through c-Met Inhibition
Authors: Nakachi Ichiro , Naoki Katsuhiko , Soejima Kenzo , Kawada Ichiro , Watanabe Hideo , Yasuda Hiroyuki , Nakayama Sohei , Yoda Satoshi , Satomi Ryosuke , Ikemura Shinnosuke , Terai Hideki , Sato Takashi , Ishizaka Akitoshi
Published: July 2010

Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKI) show antitumor activity in a subset of non–small cell lung cancer (NSCLC) patients. However, the initial tumor response is followed by recurrence. Several studies have suggested the importance of other receptor tyrosine kinases (RTK) and downstream kinases as potential targets in the treatment of NSCLC. We used the multiple-RTK inhibitor AEE788, which inhibits EGFR, vascular endothelial growth factor receptor, and human epidermal growth factor receptor 2, with and without the downstream kinase inhibitor RAD001 (an inhibitor of mammalian target of rapamycin). AEE788 inhibited cell growth more effectively than did erlotinib in three NSCLC cell lines examined (A549, H1650, and H1975). However, in the EGFR-TKI–resistant cell line H1975 harboring T790M resistance mutation, cell growth inhibition by AEE788 was only mild, and the phosphorylation of its leading targets such as EGFR and vascular endothelial growth factor receptor 2 was not inhibited. In H1975, AEE788 induced significantly greater cell growth inhibition when combined with RAD001 than when used alone. This cooperative effect was not seen with the combination of erlotinib and RAD001. We found that c-Met was highly phosphorylated in this cell line, and the phosphorylated c-Met was inhibited effectively by AEE788. Using a phospho-RTK array, the phosphorylation of c-Met and insulin-like growth factor-I receptor was inhibited by AEE788. These results suggest that upstream RTK inhibitor overcomes the acquired resistance to EGFR-TKI when combined with downstream kinase inhibitor. Thus, the combined inhibition of upstream and downstream RTKs is a promising strategy for the treatment of NSCLC. Mol Cancer Res; 8(8); 1142–51. ©2010 AACR.
BAD, a Proapoptotic Member of the BCL2 Family, Is a Potential Therapeutic Target in Hepatocellular Carcinoma
Authors: Galmiche Antoine , Ezzoukhry Zakaria , François Catherine , Louandre Christophe , Sabbagh Charles , Nguyen-Khac Eric , Descamps Véronique , Trouillet Nathalie , Godin Corinne , Regimbeau Jean-Marc , Joly Jean-Paul , Barbare Jean-Claude , Duverlie Gilles , Mazière Jean-Claude , Chatelain Denis
Published: July 2010

Proteins of the BCL2 family are key regulators of apoptosis. Their expression levels are frequently altered in cancers, enabling tumor cells to survive. To gain insight into the pathogenesis of hepatocellular carcinoma (HCC), we performed a comprehensive survey of the expression of the members of the BCL2 family in samples obtained from surgically resected HCCs. Here, we report the occurrence of a new molecular anomaly, consisting of a strong reduction in the expression of the proapoptotic protein BAD in HCC compared with surrounding nontumoral tissue. We investigate the function of BAD in a panel of HCC cell lines. Using gene overexpression and RNA interference, we show that BAD is involved in the cytotoxic effects of sorafenib, a multikinase blocker, which is currently the sole therapeutic drug effective for the treatment of HCC. Finally, we report that ABT-737, a compound that interacts with proteins of the BCL2 family and exhibits a BAD-like reactivity, sensitizes HCC cells toward sorafenib-induced apoptosis. Collectively, our findings indicate that BAD is a key regulator of apoptosis in HCC and an important determinant of HCC cell response to sorafenib. Mol Cancer Res; 8(8); 1116–25. ©2010 AACR.
Factors Affecting the Decision of Breast Cancer Patients to Undergo Contralateral Prophylactic Mastectomy
Authors: Yi Min , Hunt Kelly K. , Arun Banu K. , Bedrosian Isabelle , Barrera Angelica Gutierrez , Do Kim-Anh , Kuerer Henry M. , Babiera Gildy V. , Mittendorf Elizabeth A. , Ready Kaylene , Litton Jennifer , Meric-Bernstam Funda
Published: July 2010

Increasing numbers of women with breast cancer are electing for contralateral prophylactic mastectomy (CPM) to reduce the risk of developing contralateral breast cancer. The objective of this study was to identify factors that may affect a patient's decision to undergo CPM. We identified 2,504 women with stage 0 to III unilateral primary breast cancer who underwent breast surgery at our institution from January 2000 to August 2006 from a prospectively maintained database. We did logistic regression analyses to determine which factors were associated with undergoing CPM. Of 2,504 breast cancer patients, 1,223 (48.8%) underwent total mastectomy. Of the 1,223 patients who underwent mastectomy, 284 (23.2%) underwent immediate or delayed CPM. There were 33 patients (1.3%) who had genetic testing before the surgery, with the use of testing increasing in the latter years of the study (0.1% in 2000-2002 versus 2.0% in 2003-2006; P < 0.0001). Multivariable analysis revealed several factors that were associated with a patient undergoing CPM: age younger than 50 years, white ethnicity, family history of breast cancer, BRCA1/2 mutation testing, invasive lobular histology, clinical stage, and use of reconstruction. We identified specific patient and tumor characteristics associated with the use of CPM. Although genetic testing is increasing, most women undergoing CPM did not have a known genetic predisposition to breast cancer. Evidence-driven models are needed to better inform women of their absolute risk of contralateral breast cancer as well as their competing risk of recurrence from the primary breast cancer to empower them in their active decision making. Cancer Prev Res; 3(8); 1026–34. ©2010 AACR.
Oral Selenium Supplementation Has No Effect on Prostate-Specific Antigen Velocity in Men Undergoing Active Surveillance for Localized Prostate Cancer
Authors: Stratton M. Suzanne , Algotar Amit M. , Ranger-Moore James , Stratton Steven P. , Slate Elizabeth H. , Hsu Chiu-Hsieh , Thompson Patricia A. , Clark Larry C. , Ahmann Frederick R.
Published: July 2010

The Nutritional Prevention of Cancer trial showed a 52% lower incidence of prostate cancer in men supplemented with selenium. As a result, our study was designed to assess whether selenium supplementation attenuates the progression of prostate cancer. A phase 2 randomized, double-blind, placebo-controlled clinical trial was conducted in men with localized nonmetastatic prostate cancer who had elected to forgo active treatment and be followed by active surveillance. A total of 140 men were randomized to placebo (n = 46), 200 μg/d (n = 47), or 800 μg/d (n = 47) selenium p.o. (as selenized yeast) and followed every 3 months for up to 5 years. Prostate-specific antigen (PSA) velocity was used as a marker of prostate cancer progression and was estimated using mixed-effects regression. Adjusting for age, body mass index, baseline selenium, smoking, baseline PSA, race, PSA method, and Gleason score, PSA velocities for the 200 μg/d and 800 μg/d treatment groups were not statistically significantly different from placebo (P = 0.32 and P = 0.61, respectively). In the highest quartile of baseline selenium, men supplemented with 800 μg selenium showed statistically significantly higher PSA velocity as compared with placebo (P = 0.018). Selenium supplementation did not show a protective effect on PSA velocity in subjects with localized prostate cancer. On the contrary, supplementation with high-dose selenium was observed to be a risk factor for increased PSA velocity in men with high baseline plasma selenium concentrations. Cancer Prev Res; 3(8); 1035–43. ©2010 AACR.
Maternal Thyroid Function at 11–13 Weeks of Gestation in Women with Hypothyroidism Treated by Thyroxine
Authors: Ashoor Ghalia , Rotas Michael , Maiz Nerea , Kametas Nikos A. , Nicolaides Kypros H.
Published: July 2010

Objective: The aim of this study in pregnant women with hypothyroidism treated by levothyroxine is to examine the interrelations between thyroid-stimulating hormone (TSH), free thyroxine (FT4) and free tri-iodothyronine (FT3), and examine whether in such patients the treatment is adequate. Methods: This was a retrospective cross-sectional study. Maternal serum concentrations of FT3, FT4 and TSH were measured at 11–13 weeks in 164 singleton pregnancies from women with hypothyroidism before pregnancy receiving treatment with thyroxine. The values were compared to the results in 4,318 normal singleton pregnancies. Results: In the hypothyroid group, compared to the normal group, there was an increase in median TSH (1.990 vs. 1.007 MoM) and FT4 (1.052 vs. 0.992 MoM) and decrease in FT3 (0.901 vs. 0.991 MoM). Serum FT4 was at or above the 2.5th centile in 158 (96.3%) cases but TSH was above the 97.5th centile in 48 (29.3%) and FT3 was below the 2.5th centile in 49 (29.9%) cases. In both the hypothyroid and unaffected groups there were significant associations between TSH and FT4, TSH and FT3 and between FT3 and FT4. Conclusions: In a high proportion of pregnant women with hypothyroidism treated with levothyroxine there is evidence of persistent hypothyroidism because the treatment is inadequate in correcting the levels of FT3.
Sera of Elderly Bullous Pemphigoid Patients with Associated Neurological Diseases Recognize Bullous Pemphigoid Antigens in the Human Brain
Authors: Chen Jinbo , Li Li , Chen Jiang , Zeng Yueping , Xu Haoxiang , Song Yang , Wang Baoxi
Published: July 2010

Background: Bullous pemphigoid (BP) is an acquired autoimmune dermatosis that is often associated with various neurological diseases (ND) in the elderly. Previously, we reported that BP antigen 1 (BPAG1) of the mouse brain can be recognized by serum samples of BP patients with ND (BP+ND), indicating the possibility that BPAG1 could act as a shared autoantigen of the skin and brain. However, it is not known whether the serum of BP+ND patients could recognize BPAG1 in the human brain. Objectives: The aim of this study is to determine whether sera of BP+ND patients could recognize BP antigens in human brain tissue. Methods: Serum samples of BP+ND patients and those of age- and sex-matched BP patients without ND, ND patients without BP, and normal controls were obtained. Serum samples were examined by immunoblotting against protein extracts of human skin and brain. Results: A 230-kDa protein of human epidermal extract was recognized by 16 of 22 (72.7%) serum samples of elderly BP+ND patients, whereas it was recognized by 50, 0, and 0% of serum samples of BP, ND, and controls, respectively. A 230-kDa protein of human brain extract was recognized by 12 of 22 (54.5%) serum samples of BP+ND patients, whereas it was recognized by 9.1, 9.1, and 4.5% of serum samples of BP, ND, and controls, respectively. A 180- and a 165-kDa protein could also be recognized by a proportion of serum samples of BP and BP+ND patients. Conclusion: This study demonstrated that both human skin and brain contain immunogenic BPAG1 in BP+ND patients. We speculate that alterations of the central nervous system in the course of pathological changes of elderly ND patients could expose the neural isoforms of BPAG1 or other BP antigens. Autoantibodies against these antigens in the brain may cross-react with antigen in the skin, which may underlie the pathological development of BP.
Effects of Blood Pressure Changes on Alzheimer’s Disease
Authors: Feldstein Carlos A.
Published: July 2010

Alzheimer’s disease is the most frequent cause of dementia. Whereas other major causes of death have been decreasing, the number of deaths due to Alzheimer’s disease is rising. As there is no cure for this type of dementia at present, preventive measures have assumed great importance. By analyzing data from available longitudinal studies, the current review presents evidence supporting a link between Alzheimer’s disease and blood pressure changes.
Nitric Oxide Is Involved in the Response of the Isolated Intact and Epithelium-Denuded Rat Trachea to the β2 Adrenergic Receptor Agonist Salbutamol
Authors: Montalvo F. , Cantres-Fonseca O. , Santos G. , Vega M. , Torres I. , Carmona J. , Dexter D. , Santacana G.
Published: July 2010

Background: The involvement of the airway smooth muscle mediator nitric oxide (NO) in the actions of the β2 agonist salbutamol (Sal), a well- known bronchodilator, is very poorly understood. Objectives: To determine if endogenous NO release is a major factor in the Sal-induced relaxation of the carbachol- and electrical field-stimulated rat trachea and determine the role of the tracheal epithelium as the possible source of NO involved in these effects. Methods: Isolated carbachol- or electric field-stimulated pre-contracted in vitro male Sprague Dawley rat tracheas (with epithelium intact or denuded) were relaxed with incremental or discrete concentrations of Sal in the presence and absence of the NO synthesis inhibitor L-NAME. Results: Epithelium-denuded tracheas showed a reduced relaxation response to Sal. L-NAME (1 mM) similarly decreased the sensitivity of the rat tracheas to Sal in both epithelium-intact and -denuded conditions. In the presence of L-NAME, high concentrations of Sal induced an unexpectedly large relaxation response in carbachol-stimulated rat tracheas with both intact and denuded epithelium. Sal relaxation was also affected by L-NAME in electrical field-stimulated epithelium-intact and -denuded tracheas. Conclusion: The results suggest that NO derived from sources other than the epithelium is an important mediator of the Sal-induced relaxation in rat tracheas.
TRA-418, a thromboxane A2 receptor antagonist and prostacyclin receptor agonist, inhibits platelet-leukocyte interaction in human whole blood
Authors: Ohno Michihiro , Yamada Naohiro , Ohtake Atsushi , Matsushita Teruo , Miyamoto Mitsuko
Published: July 2010

TRA-418, a compound with both thromboxane A2 receptor (TP receptor) antagonistic and prostacyclin receptor (IP receptor) agonistic activities, was synthesised in our laboratory as a new antithrombotic agent. In this study, we examined the effects of TRA-418 on platelet-leukocyte interactions in human whole blood. Platelet-leukocyte interactions were induced by U-46619 in the presence of epinephrine (U-46619 + epinephrine) or with thrombin receptor agonist peptide 1–6 (TRAP). Platelet-leukocyte interactions were assessed by flow cytometry, with examination of both platelet-neutrophil and platelet-monocyte complexes. In a control experiment, the TP receptor antagonist SQ-29548 significantly inhibited the induction of platelet-leukocyte complexes by the combination of U-46619 and epinephrine, but not TRAP-induced formation of platelet-leukocyte complexes. Conversely, the IP receptor agonist beraprost sodium inhibited platelet-leukocyte complex formation induced by both methods, although the IC50 values of beraprost sodium for U-46619 + epinephrine were at least 10-fold greater than for TRAP. Under such conditions, TRA-418 inhibited both U-46619 + epinephrine-induced and TRAP-induced platelet-leukocyte complex formation in a concentration-dependent manner, in a similar range. These results suggest that TRA-418 exerts its inhibitory effects on platelet-leukocyte interactions by acting as a TP receptor antagonist as well as an IP receptor agonist in an additive or synergistic manner. These inhibitory effects of TRA-418 on formation of platelet-leukocyte complexes suggest the compound is beneficial effects as an antithrombotic agent.
Association study between polymorphims in GAS6-TAM genes and carotid atherosclerosis
Authors: Hurtado Begoña , Abasolo Nerea , Muñoz Xavier , García Nadia , Benavente Yolanda , Rubio Francisco , de Frutos Pablo García , Krupinski Jerzy , Sala Núria
Published: July 2010

Carotid atherosclerosis (CA) is one of the most common causes of stroke, and recent studies suggest that pathways initiated by the interaction of the plasma vitamin K-dependent protein GAS6 with the tyrosine kinase receptors TYRO3, AXL and MERTK (TAM) may have a relevant role in atherogenesis. Furthermore, our previous studies indicated an association between GAS6 and stroke. The aim of this study was to analyse the genetic association between SNPs and haplotypes in GAS6-TAM genes and CA. We performed a case-control study with 233 CA patients confirmed by nuclear magnetic resonance angiography and 202 patients who suffered from cardioembolic (non atherogenic) stroke. For all included subjects information on established risk factors was available. Genotyping of 16 selected tagSNPs was performed by real-time PCR, using either FRET or TaqMan probes. Adjusted logistic regression (LR) analyses indicated that rs2289743 in TYRO3 and rs869016 in MERTK were associated to CA, decreasing its risk (OR [95%CI]=0.39 [0.16–0.94] and OR [95%CI]=0.31 [0.14–0.69], respectively). Linkage disequilibrium results were consistent with the haplotype blocks described in HapMap and adjusted LR analyses revealed that the haplotype ACAA in MERTK, containing the minor allele of the associated SNP, was also associated to CA. No association was observed with GAS6 and AXL variants, which suggests that CA is not the mechanism underlying the reported association between GAS6 and stroke. The association between TYRO3 and MERTK variants and carotid atherosclerosis found in this study reinforces a physiological role of the GAS6-TAM pathway in atherogenesis.
Differential sensitivity and kinetics of response of different ex vivo tests monitoring functional variability of platelet response to clopidogrel
Authors: Sollier Claire Bal dit , Berge Natacha , Boval Bernadette , Dubar Michel , Drouet Ludovic
Published: July 2010

We previously showed that variability of response to clopidogrel is linked to occupancy of the P2Y12 receptor by clopidogrel active-metabolite, and that maximal platelet aggregation intensity (MAI) measured by light transmission aggregometry (LTA) correlates with occupancy. The present study compared a range of ex vivo platelet tests at various levels of P2Y12 occupancy. After screening with clopidogrel 75 mg/day for seven days, subjects were selected to obtain ‘low’, ‘average’ and ‘high’ responders and randomised to clopidogrel (75 mg/day days 1–10; 300 mg day 11), or placebo. Assays were LTA in platelet-rich plasma using 2, 5 and 20 μM ADP, VerifyNow® P2Y12, flow cytometric analysis of platelet activation markers and vasodilator-stimulated phospho-protein (VASP) index, and a thromboelastographic test that is sensitive to clopidogrel. The reference test was P2Y12 receptor occupancy, measured by binding of 33P-2MeS-ADP to platelets. MAI showed the best correlation with P2Y12 occupancy. Similar results were seen with different ADP concentrations and when LTA data were expressed as inhibition of platelet aggregation. A plot of free receptors/cell versus VASP index was biphasic, with poor correlation for low-level P2Y12 occupancy. Sensitivity of the VerifyNow P2Y12 assay decreased at higher clopidogrel responses. Thromboelastography and P-selectin expression had poor correlation with receptor occupancy. In conclusion, LTA data correlate best with P2Y12 occupancy, the gold standard for detecting clopidogrel’s effect at the receptor level. Our results highlight a limitation of the VASP index, which appears unable to distinguish low, average and high responders early after clopidogrel initiation when P2Y12 occupancy is still low.
Factor XIII-A is involved in the regulation of gene expression in alternatively activated human macrophages
Authors: Töröcsik Dániel , Széles Lajos , Paragh György , Rákosy Zsuzsa , Bárdos Helga , Nagy László , Balázs Margit , Inbal Aida , Ádány Róza
Published: July 2010

Factor XIII subunit A (FXIII-A) is one of the most overrepresented genes that is expressed during the alternative activation of macrophages. Based on its substrate profile and its cellular localisation, FXIII-A is thought to function as an intracellular/intranuclear transglutaminase. Our aim was to find role for the intracellular FXIII-A by comparing the microarray profiles of alternatively activated monocyte-derived macrophages. Microarray analyses of FXIII-A-deficient patients and healthy controls were evaluated, followed by functional clustering of the differentially expressed genes. After a 48-hour differentiation in the presence of interleukin 4 (IL4), 1,017 probes out of the 24,398 expressed in macrophages from FXIII-A- deficient samples were IL4 sensitive, while only 596 probes were IL4 sensitive in wild-type samples. Of these genes, 307 were induced in both the deficient and the wild-type macrophages. Our results revealed that FXIII-A has important role(s) in mediating gene expression changes in macrophages during alternative activation. Functional clustering of the target genes carried out using Cytoscape/BiNGO and Ingenuity Pathways Analysis programs showed that, in the absence of FXIII-A, the most prominent differences are related to immune functions and to wound response. Our findings suggest that functional impairment of macrophages at the level of gene expression regulation plays a role in the wound healing defects of FXIII-A-deficient patients.
Phosphate and vascular calcification: Emerging role of the sodium-dependent phosphate co-transporter PiT-1
Authors: Lau Wei Ling , Festing Maria , Giachelli Cecilia
Published: July 2010

Elevated serum phosphate is a risk factor for vascular calcification and cardiovascular events in kidney disease as well as in the general population. Elevated phosphate levels drive vascular calcification, in part, by regulating vascular smooth muscle cell (VSMC) gene expression, function, and fate. The type III sodium-dependent phosphate co-transporter, PiT-1, is necessary for phosphate-induced VSMC osteochondrogenic phenotype change and calcification, and has recently been shown to have unexpected functions in cell proliferation and embryonic development.
Differential effects of TAK-442, a novel orally active direct factor Xa inhibitor, and ximelagatran, a thrombin inhibitor, on factor V-mediated feedback on coagulation cascade and bleeding
Authors: Konishi Noriko , Hiroe Katsuhiko , Kawamura Masaki
Published: July 2010

Thrombin amplifies the blood coagulation via factor V (FV)-mediated positive feedback loop. We hypothesised that factor Xa (FXa) inhibitors would interfere more gradually with this feedback activation loop than thrombin inhibitors, thereby achieving a better balance between haemostasis and prevention of thrombosis. In this study, we compared the effects of TAK-442, a novel FXa inhibitor, versus ximelagatran, a thrombin inhibitor, on FV-mediated positive feedback, venous thrombosis and bleeding. In normal plasma, TAK-442 delayed the onset of tissue factor-induced thrombin generation and prolonged prothrombin time (PT) with more gradual concentration-response curve than melagatran, the active form of ximelagatran. The effect of melagatran on the onset of thrombin generation decreased in an FVa-concentration-dependent manner in FV-deficient plasma supplemented with FVa. Furthermore, in FV-deficient plasma, the PT-prolonging potency of melagatran was markedly increased with a change in its concentration-response curve from steep to gradual. In the rat venous thrombosis model, TAK-442 (10 mg/kg, p.o.) prevented thrombus formation by 55% with 1.2 times prolongation of PT; a similar effect was observed in ximelagatran-treated (3 mg/kg, p.o.) rats. TAK-442 at 100 mg/kg prolonged PT by only 2.1 times with no change in bleeding time (BT), whereas ximelagatran at 10 mg/kg prolonged PT by 3.9 times and significantly increased BT. These results suggest that the differential effects of the two agents on FV-mediated amplification of thrombin generation may underlie the observation of a wider therapeutic window for TAK-442 than for ximelagatran.
Shear stress regulates inflammatory and thrombogenic gene transcripts in cultured human endothelial progenitor cells
Authors: Lund Trine , Hermansen Stig , Andreasen Thomas , Olsen Jan , Østerud Bjarne , Myrmel Truls , Ytrehus Kirsti
Published: July 2010

Shear stress has an established effect on mature endothelial cells, but less is known about how shear stress regulates endothelial progenitor cells (EPCs). In vitro expanded EPCs isolated from adult human blood represent a novel tool in regenerative vessel therapy. However, in vitro culturing may generate cells with unfavourable properties. The aim of the present study was therefore to assess whether shear stress may influence the inflammatory and thrombotic phenotype of in vitro expanded EPCs. In late outgrowth EPCs, 6 hours of shear stress (6.0 dynes/ cm2) significantly reduced the mRNA levels of IL-8, COX2, and tissue factor (TF) compared to static controls. This was associated with a reduced TF activity. In contrast, mRNA expression of NOS3 was significantly increased following 6 and 24 hours of shear stress. In accordance with this, NOS3 protein expression was increased following 24 hours of shear stress. Overall stimulation with the proinflammatory mediator, TNFα, for the final 2 hours increased the mRNA expression of IL-6, IL-8, MCP-1, ICAM1, and TF. However exposure to 6 hours of shear stress significantly suppressed the inductory potential of TNFα to increase the mRNA levels of IL-6, IL-8, COX2, and TF. Additionally, TNFα increased TF activity approximately 10 times, an effect that was also significantly reduced by exposure to 6 and 24 hours of shear stress. The effect of shear on the gene levels of TF and NOS3 were not blocked by the NOS inhibitor L-NAME. These observations suggest that EPCs are capable of functionally responding to shear stress.
A splice variant of ADAMTS13 is expressed in human hepatic stellate cells and cancerous tissues
Authors: Hunt Ryan , Hershko Klilah , Pommier Elie , Geetha S. , Blaisdell Adam , Marple Andrew , Roma Isabella , Newell Jordan , Allen Courtni , Friedman Scott , Shomron Noam , Hamasaki-Katagiri Nobuko , Kimchi-Sarfaty Chava
Published: July 2010

Although ADAMTS13, the von Willebrand factor (VWF)-cleaving protease, is expressed in a range of tissues, the physiological significance of tissue-specific ADAMTS13 alternative splicing isoforms have yet to be clarified. Screening a panel of human tissues and cell lines revealed a spliced ADAMTS13 transcript in hepatic stellate cells and a hepatoma cell line that retains the 25th intron. A nonsense codon within the intron truncates the protease, which gains 64 novel amino acids in lieu of both CUB domains. This isoform, while retaining VWF-cleaving capability, accumulates intracellularly and its biological inaccessibility may prevent its participation in regulating haemostasis and other physiologic functions.
The effect of ethanol and its metabolism on fibrinolysis
Authors: Vorster Hester , Jerling Johann , Venter Christine , Kotze Retha , Bornman Elsabe , Malfliet Joyce , Rijken Dingeman , Pieters Marlien
Published: July 2010

The role of ethanol metabolism in possible haemostatic cardioprotective effects has not yet been determined. To this end, we investigated the effect of a moderate dose of ethanol (35 g) and its metabolism, on haemostatic variables over 14 hours (h). Eighteen Caucasian males participated in a placebo-controlled, randomised, cross-over study. Blood was collected prior to alcohol consumption, and at 10 time points for 14 h. Blood ethanol peaked at 1 h and was cleared after 8 h following ethanol consumption, significantly increasing plasma acetate (p=0.0028). Ethanol did not influence the coagulation factors significantly. PAI-1act increased (p<0.0001) and tPAact (p=0.047) decreased following alcohol consumption, reaching maximum (0.69 to 22.2 IU/ml) and minimum (0.88 to 0.33 IU/ml) levels at 5 h, respectively. Significantly increased plasma clot lysis times (46.8 to 67.6 minutes) and reduced global fibrinolytic capacity of whole blood, measured as D-dimer production during incubation of blood clots (2.26 to 0.29 μg/ml), were found at 5 h. Except for PAI-1act (borderline significance; p=0.05), there was no significant difference in the fibrinolytic markers between the two groups the following morning. Moderate ethanol consumption resulted in a significant temporary fibrinolysis inhibition. Any protective effects of moderate ethanol consumption on cardiovascular disease do not appear to be due to improvement in fibrinolytic potential within the first 14 h following consumption. The use of global fibrinolytic assays is recommended for determining the true effect of ethanol on fibrinolysis.
CYP1A1, smoking and venous thromboembolism
Authors: Kong Fan-Cui , Zhang Wei-Juan , Zhu Jin , Zheng Wen-Jie , Wang He-Yao , Wang Chen
Published: July 2010

Since CYP1A1 enzyme is involved in metabolism of tobacco carcinogens, the CYP1A1 gene may be of relevance to smoking-induced differences in the risks of venous thromboembolism (VTE). We conducted a case-control study to investigate genetic polymorphisms in CYP1A1 that might modify the risk of developing VTE. A total of 425 Chinese patients with VTE and 527 VTE-free control individuals, matched by age and gender, were included in this analysis. The MspI and Ile462Val polymorphisms in CYP1A1 gene were analysed using the Amplification Refractory Mutation System (ARMS) method. The Ile462Val AG variant and combined AG and GG variant was significantly associated with VTE, adjusted for age, gender, weight and contraceptives (OR=1.362, 95%CI 1.026, 1.809, p=0.033; OR=1.420, 95% CI 1.081, 1.865, p=0.012, respectively); The AG and GG combined variant was still significantly associated with VTE when adjusting further for smoking (OR=1.344, 95%CI 1.019,1.772, p=0.036) A more than two-fold increase for VTE was associated with the Ile462Val combined variant of AG+GG (OR of 2.805, 95% CI 1.250, 6.293, p=0.012) in the smokers. Genetic variations of CYP1A1 Ile462Val contribute to susceptibility to smoking-induced VTE in the Chinese populations. A two-fold increase in the risk in the smokers in the patients who carry CYP1A1 Ile462Val variant alleles has demonstrated the importance of gene-environment interactions in the development of this disease.
Clinical decision rule and D-dimer have lower clinical utility to exclude pulmonary embolism in cancer patients
Authors: van Sluis Geerte , Kamphuisen Pieter , Söhne Maaike , Leebeek Frank , Bossuyt Patrick , Büller Harry , Douma Renée
Published: July 2010

Patients with malignancy frequently present with clinically suspected pulmonary embolism (PE). However, the safe and efficient combination of a clinical decision rule (CDR) and D-dimer test to rule out PE performs less well in patients with malignancy. We examined potential explanations and analysed whether elevating the D-dimer cut-off could improve the clinical utility. We used data on consecutive patients with suspected PE included in a multicenter management study. The performance of the Wells CDR and the D-dimer test was compared between patients with and without malignancy and multivariable analysis was used to compare the weights of the CDR variables. Furthermore, we combined the CDR (cut-off ≤4) with different D-dimer cut-off levels for the exclusion of PE. Of 3,306 patients with suspected PE, 475 (14%) had cancer. The Wells rule variables were less diagnostic in cancer patients. Increasing the D-dimer cut-off level to 700 μg/l for all ages or using an age-dependent cut-off resulted in an increase in the proportion of patients in whom PE could be excluded from 8.4% to 13% and 12%, respectively. The corresponding false-negative rates were 1.6% (95% confidence interval 0.3–8.7%) and 0.0% (0.0–6.3%). The Wells CDR and D-dimer perform less well in patients with suspected PE if they have cancer. Individual variables in the Wells rule are less diagnostic in cancer patients than in non-cancer patients with suspected PE. A CDR combined with an age-dependent D-dimer cut-off shows a modest improvement of the strategy in cancer patients.
Impact of race and gender on antithrombotic therapy
Authors: Capodanno Davide , Angiolillo Dominick
Published: July 2010

Large randomised trials support the use of a variety of antithrombotic drugs for treatment of atherothrombotic disease processes leading to cardiovascular disease. The heterogeneous case-mix of patients enrolled in these trials, however, hamper the attempt to generalise their findings to subgroups which are not sufficiently represented in the study population, such as women and ethnic minorities. Sex- and race-specific disparities in the clinical presentation, management, and outcomes of coronary artery disease may relate to underlying differences in thrombotic profiles and response to antithrombotic therapies. The present manuscript provides an overview of the currently available data on the epidemiology of coronary artery disease based on gender and race as well as the biological considerations for their differences in thrombosis and haemostasis and effects of antithrombotic therapy.
Relationship between dietary vitamin K intake and the stability of anticoagulation effect in patients taking long-term warfarin
Authors: Kim Kyun Hee , Choi Won Suk , Lee Jang Hoon , Lee Hyejin , Yang Dong Heon , Chae Shung Chull
Published: July 2010

Little study has been performed on the effect of vitamin K intake on the variability of warfarin’s anticoagulant effects over long period of time. We estimated average vitamin K intake in the patients taking warfarin and evaluated its relation with the stability of anticoagulation effect. We estimated average daily vitamin K intake based on a three-day food diary in 66 patients taking warfarin regularly for ≥one year and divided them into three groups of equal number according to vitamin K intake. Stability of anticoagulant effect was compared in these groups using the coefficient of variation (CV) of the prothrombin time expressed in international normalised ratio (INR) and the CV of warfarin doses. Median daily vitamin K intake was 161.3 μg/day (31.3 μg/day – 616.6 μg/day). CVs of both INR and warfarin doses were negatively and independently correlated with dietary vitamin K intake (r=-0.293, p=0.017 and r= –0.350, p=0.004, respectively). CV of INR was significantly different among three groups of vitamin K intake (p<0.05 in ANOVA). High vitamin K intake (>195.7 μg/day) group had lower CV of INR than the low intake (<126.5 μg/day) group (19.2 ± 8.96 % vs. 25.5 ± 8.61 %, p<0.05). CV of warfarin doses was also significantly different among the groups (p<0.05 in Jonckheere-Terpstra test). However, the significance of difference between high and low vitamin intake groups was marginal (p=0.046 in Mann-Whitney test). In conclusion, long-term anticoagulation effect of warfarin is more stable in the patients who take greater than a certain amount of dietary vitamin K.

Footnote: The content of this paper was submitted as partial fulfillment of the requirement for the degree of Master of Medical Science (Kim KH) to the Kyungpook National University.