bluuebunny/crossref_metadata_2025_split huggingface dataset:
Targeted Degradation of KRAS by an Engineered Ubiquitin Ligase Suppresses Pancreatic Cancer Cell Growth In Vitro and In Vivo
Authors: Ma Yihui , Gu Yumei , Zhang Qiang , Han Yongqing , Yu Shuangni , Lu Zhaohui , Chen Jie 
Published: January 2013

Abstract
KRAS is an attractive pancreatic ductal adenocarcinoma (PDAC) therapeutic target. E3 ligase is thought to be the component of the ubiquitin conjugation system that is directly responsible for substrate recognition. In this study, an engineered E3 ubiquitin ligase (RC-U) was generated to target the KRAS oncoprotein for ubiquitination and degradation. The engineered E3 ubiquitin ligases (RC-U) were constructed (pRC-U and lentivirus-expressing RC-U). After transfecting the pRC-U plasmid into human pancreatic cancer cells, KRAS expression levels were determined. KRAS expression was also evaluated in cells transfected with pRC-U and treated with MG-132 or cycloheximide. Interactions between RC-U and KRAS as well as whether RC-U could ubiquitinate KRAS were investigated. Extracellular signal–regulated kinase 1/2 (ERK1/2) and phosphorylated ERK 1/2 (pERK1/2) levels were examined in pancreatic cancer cells transfected with pRC-U. The effects of RC-U on pancreatic cancer cell growth were assessed. RC-U decreased KRAS protein levels. After pRC-U transfection, KRAS stability was increased in the presence of MG-132. HEK 293T cells were transfected with a mutant KRAS construct together with pRC-U and incubated with cycloheximide to inhibit new protein synthesis. The exogenous mutant KRAS oncoprotein was degraded more quickly. RC-U can bind KRAS and KRAS can be ubiquitinated by RC-U. pERK1/2 protein levels were decreased. RC-U resulted in reduced cell proliferation in vitro and in vivo. KRAS destruction by RC-U occurred through a ubiquitin-dependent, proteasome-mediated degradation pathway. RC-U inhibited pancreatic cancer cell growth in vitro and in vivo. Mol Cancer Ther; 12(3); 286–94. ©2012 AACR.
A Second-Generation 2-Methoxyestradiol Prodrug Is Effective against Barrett's Adenocarcinoma in a Mouse Xenograft Model
Authors: Kambhampati Suman , Rajewski Roger A. , Tanol Mehmet , Haque Inamul , Das Amlan , Banerjee Snigdha , Jha Saheli , Burns Douglas , Borrego-Diaz Emma , Van Veldhuizen Peter J. , Banerjee Sushanta K. 
Published: January 2013

Abstract
2-Methoxyestradiol (2-ME2) is an endogenous metabolite of estradiol. In preclinical models, 2-ME2 is effective against different types of tumors. Unfortunately, only low systemic concentrations of 2-ME2 can be achieved following oral administration, even after very high doses are administered to patients. In an effort to solve this problem, we have now synthesized and tested a new prodrug of 2-ME2 that is water-soluble due to a bioreversible hydrophilic group added at the 3-position and that more effectively resists metabolic inactivation due to an ester moiety added to mask the 17-position alcohol. We are reporting here for the first time that this double prodrug of 2-ME2 is effective as an antiproliferative and anticancer agent for both in vitro and in vivo studies against Barrett esophageal adenocarcinoma (BEAC) and provided greater potency than 2-ME2 in inhibiting the growth of BEAC xenografts. Finally, studies indicate that, like 2-ME2, the 2-ME2-PD1 exhibits anticancer effect through possible disruption of microtubule network. Mol Cancer Ther; 12(3); 255–63. ©2012 AACR.
Dynamin-Related Protein-1 Controls Fusion Pore Dynamics During Platelet Granule Exocytosis
Authors: Koseoglu Secil , Dilks James R. , Peters Christian G. , Fitch-Tewfik Jennifer L. , Fadel Nathalie A. , Jasuja Reema , Italiano Joseph E. , Haynes Christy L. , Flaumenhaft Robert 
Published: January 2013

Objective—
Platelet granule exocytosis serves a central role in hemostasis and thrombosis. Recently, single-cell amperometry has shown that platelet membrane fusion during granule exocytosis results in the formation of a fusion pore that subsequently expands to enable the extrusion of granule contents. However, the molecular mechanisms that control platelet fusion pore expansion and collapse are not known.

Methods and Results—
We identified dynamin-related protein-1 (Drp1) in platelets and found that an inhibitor of Drp1, mdivi-1, blocked exocytosis of both platelet dense and α-granules. We used single-cell amperometry to monitor serotonin release from individual dense granules and, thereby, measured the effect of Drp1 inhibition on fusion pore dynamics. Inhibition of Drp1 increased spike width and decreased prespike foot events, indicating that Drp1 influences fusion pore formation and expansion. Platelet-mediated thrombus formation in vivo after laser-induced injury of mouse cremaster arterioles was impaired after infusion of mdivi-1.

Conclusion—
These results demonstrate that inhibition of Drp1 disrupts platelet fusion pore dynamics and indicate that Drp1 can be targeted to control thrombus formation in vivo.
Kalirin Promotes Neointimal Hyperplasia by Activating Rac in Smooth Muscle Cells
Authors: Wu Jiao-Hui , Fanaroff Alexander C. , Sharma Krishn C. , Smith Liisa S. , Brian Leigh , Eipper Betty A. , Mains Richard E. , Freedman Neil J. , Zhang Lisheng 
Published: January 2013

Objective—

Kalirin is a multifunctional protein that contains 2 guanine nucleotide exchange factor domains for the GTPases Rac1 and RhoA. Variants of
KALRN
have been associated with atherosclerosis in humans, but Kalirin’s activity has been characterized almost exclusively in the central nervous system. We therefore tested the hypothesis that Kalirin functions as a Rho-guanine nucleotide exchange factor in arterial smooth muscle cells (SMCs).

Approach and Results—

Kalirin-9 protein is expressed abundantly in aorta and bone marrow, as well as in cultured SMCs, endothelial cells, and macrophages. Moreover, arterial Kalirin was upregulated during early atherogenesis in apolipoprotein E-deficient mice. In cultured SMCs, signaling was affected similarly in 3 models of Kalirin loss-of-function: heterozygous
Kalrn
deletion, Kalirin RNAi, and treatment with the Kalirin Rho-guanine nucleotide exchange factor -1 inhibitor 1-(3-nitrophenyl)-1H-pyrrole-2,5-dione. With reduced Kalirin function, SMCs showed normal RhoA activation but diminished Rac1 activation, assessed as reduced Rac-GTP levels, p21-activated kinase autophosphorylation, and SMC migration.
Kalrn
–/+
SMCs proliferated 30% less rapidly than wild-type SMCs. Neointimal hyperplasia engendered by carotid endothelial denudation was ≈60% less in
Kalrn
–/+
and SMC-specific
Kalrn
–/+
mice than in control mice.

Conclusion—
Kalirin functions as a guanine nucleotide exchange factor for Rac1 in SMCs, and promotes SMC migration and proliferation both in vitro and in vivo.
Key Role of Estrogens and Endothelial Estrogen Receptor α in Blood Flow–Mediated Remodeling of Resistance Arteries
Authors: Tarhouni K. , Guihot A. L. , Freidja M. L. , Toutain B. , Henrion B. , Baufreton C. , Pinaud F. , Procaccio V. , Grimaud L. , Ayer A. , Loufrani L. , Lenfant F. , Arnal J. F. , Henrion D. 
Published: January 2013

Objective—
Flow- (shear stress–)mediated outward remodeling of resistance arteries is involved in collateral growth during postischemic revascularization. As this remodeling is especially important during pregnancy, we hypothesized that estrogens may be involved. A surgical model eliciting a local increase in blood flow in 1 mesenteric resistance artery was used in 3-month-old ovariectomized female rats either treated with 17-β-estradiol (E2) or left untreated.

Methods and Results—

After 14 days, arterial diameter was greater in high-flow arteries than in normal-flow vessels. An ovariectomy suppressed high-flow remodeling, while E2 restored it. High-flow remodeling was absent in mice lacking the estrogen receptor α but not estrogen receptor β. The kinetics of inflammatory marker expression, macrophage infiltration, oxidative stress, and metaloproteinases expression were not altered by the absence of E2 after 2 and 4 days, that is, during remodeling. Nevertheless, E2 was required for the increase in endothelial nitric oxide synthase expression and activation at day 4 when diameter expansion occurs. Finally, the impact of E2 on the endothelium appeared crucial for high-flow remodeling, as this E2 action was abrogated in mice lacking endothelial NOS, as well as in Tie2-Cre(+) ERα
f/f
mice.

Conclusion—
We demonstrate the essential role of E2 and endothelial estrogen receptor α in flow-mediated remodeling of resistance arteries in vivo.
Histone Deacetylase 9 Promotes Angiogenesis by Targeting the Antiangiogenic MicroRNA-17–92 Cluster in Endothelial Cells
Authors: Kaluza David , Kroll Jens , Gesierich Sabine , Manavski Yosif , Boeckel Jes-Niels , Doebele Carmen , Zelent Arthur , Rössig Lothar , Zeiher Andreas M. , Augustin Hellmut G. , Urbich Carmen , Dimmeler Stefanie 
Published: January 2013

Objective—
Histone deacetylases (HDACs) modulate gene expression by deacetylation of histone and nonhistone proteins. Several HDACs control angiogenesis, but the role of HDAC9 is unclear.

Methods and Results—
Here, we analyzed the function of HDAC9 in angiogenesis and its involvement in regulating microRNAs. In vitro, silencing of HDAC9 reduces endothelial cell tube formation and sprouting. Furthermore, HDAC9 silencing decreases vessel formation in a spheroid-based Matrigel plug assay in mice and disturbs vascular patterning in zebrafish embryos. Genetic deletion of HDAC9 reduces retinal vessel outgrowth and impairs blood flow recovery after hindlimb ischemia. Consistently, overexpression of HDAC9 increases endothelial cell sprouting, whereas mutant constructs lacking the catalytic domain, the nuclear localization sequence, or sumoylation site show no effect. To determine the mechanism underlying the proangiogenic effect of HDAC9, we measured the expression of the microRNA (miR)-17–92 cluster, which is known for its antiangiogenic activity. We demonstrate that silencing of HDAC9 in endothelial cells increases the expression of miR-17–92. Inhibition of miR-17–20a rescues the sprouting defects induced by HDAC9 silencing in vitro and blocking miR-17 expression partially reverses the disturbed vascular patterning of HDAC9 knockdown in zebrafish embryos.

Conclusion—
We found that HDAC9 promotes angiogenesis and transcriptionally represses the miR-17–92 cluster.
Reduced Atherosclerotic Burden in Subjects With Genetically Determined Low Oxidative Stress
Authors: Violi Francesco , Pignatelli Pasquale , Pignata Claudio , Plebani Alessandro , Rossi Paolo , Sanguigni Valerio , Carnevale Roberto , Soresina Annarosa , Finocchi Andrea , Cirillo Emilia , Catasca Elisa , Angelico Francesco , Loffredo Lorenzo 
Published: January 2013

Objective—
NADPH oxidase, one of the most important enzymes producing reactive oxidant species, is suggested to play a role in experimental atherosclerosis, but its role in human atherosclerosis is still unclear. We hypothesized that a reduced activity of NADPH oxidase might be linked to a reduced atherosclerotic burden.

Methods and Results—

Thirty-one women carriers of hereditary deficiency of NOX2, the catalytic subunit of NADPH oxidase, were matched for sex and age with 31 controls and 31 obese women. Flow-mediated dilation and intima-media thickness, 2 surrogate markers of atherosclerosis, serum activity of NOX2, urinary isoprostanes, serum levels of nitrite/nitrate, and platelet production of isoprostanes and nitrite/nitrate were determined. Compared with controls (5.7±3.0% and 0.60±0.11 mm), carriers of NOX2 deficiency had higher flow-mediated dilation (9.2±5.0%;
P
<0.001) and lower intima-media thickness (0.50±0.11 mm;
P
=0.002), whereas obese women had lower flow-mediated dilation (3.2±2.1%;
P
=0.007) and higher intima-media thickness (0.71±0.15 mm;
P
<0.001). Compared with controls, carriers of NOX2 deficiency had lower urinary isoprostanes (132.6±87.3 versus 82.3±46.0 pg/mg creatinine;
P
=0.007) and serum NOX2 activity (24.9±19.3 versus 12.8±11.9 pg/mL;
P
=0.004) and higher serum nitrite/nitrate (23.8±7.6 versus 30.5±6.3 µmol/L;
P
<0.001), whereas obese women had higher urinary isoprostanes (132.6±87.3 versus 182.2±84.6 pg/mg creatinine;
P
=0.008) and serum NOX2 activity (24.9±19.3 versus 36.1±18.6 pg/mL;
P
=0.008) and lower serum nitrite/nitrate (23.8±7.6 versus 12.6±4.2 µmol/L;
P
<0.001). Flow-mediated dilation correlated with intima-media thickness (
r
=–0.433;
P
<0.001), serum NOX2 activity (
r
=–325;
P
<0.001), and urinary isoprostanes (
r
=–0.314;
P
=0.002). Ex vivo study showed that, compared with controls, platelets from carriers of NOX2 deficiency had lower isoprostanes (
P
<0.001) and higher nitrite/nitrate (
P
<0.001), whereas platelets from obese women had higher isoprostanes (
P
<0.001) and lower nitrite/nitrate (
P
=0.013).

Conclusion—
The study shows reduced atherosclerotic burden in carriers of NOX2 deficiency, suggesting that oxidative stress generated by this enzymatic pathway is implicated in human atherosclerosis.
Aberrant Endoplasmic Reticulum Stress in Vascular Smooth Muscle Increases Vascular Contractility and Blood Pressure in Mice Deficient of AMP-Activated Protein Kinase-α2 In Vivo
Authors: Liang Bin , Wang Shuangxi , Wang Qilong , Zhang Wencheng , Viollet Benoit , Zhu Yi , Zou Ming-Hui 
Published: January 2013

Objective—
The endoplasmic reticulum (ER) plays a critical role in ensuring proper folding of newly synthesized proteins. Aberrant ER stress is reported to play a causal role in cardiovascular diseases. However, the effects of ER stress on vascular smooth muscle contractility and blood pressure remain unknown. The aim of this study was to investigate whether aberrant ER stress causes abnormal vasoconstriction and consequent high blood pressure in mice.

Methods and Results—
ER stress markers, vascular smooth muscle contractility, and blood pressure were monitored in mice. Incubation of isolated aortic rings with tunicamycin or MG132, 2 structurally unrelated ER stress inducers, significantly increased both phenylephrine-induced vasoconstriction and the phosphorylation of myosin light chain (Thr18/Ser19), both of which were abrogated by pretreatment with chemical chaperones or 5-Aminoimidazole-4-carboxamide ribonucleotide and metformin, 2 potent activators for the AMP-activated protein kinase. Consistently, administration of tauroursodeoxycholic acid or 4-phenyl butyric acid, 2 structurally unrelated chemical chaperones, in AMP-activated protein kinase-α2 knockout mice lowered blood pressure and abolished abnormal vasoconstrictor response of AMP-activated protein kinase-α2 knockout mice to phenylephrine. Consistently, tunicamycin (0.01 μg/g per day) infusion markedly increased both systolic and diastolic blood pressure, both of which were ablated by coadministration of 4-phenyl butyric acid. Furthermore, 4-phenyl butyric acid or tauroursodeoxycholic acid, which suppressed angiotensin II infusion–induced ER stress markers in vivo, markedly lowered blood pressure in angiotensin II–infused mice in vivo.

Conclusion—
We conclude that ER stress increases vascular smooth muscle contractility resulting in high blood pressure, and AMP-activated protein kinase activation mitigates high blood pressure through the suppression of ER stress in vivo.
Notch-Dependent Regulation of the Ischemic Vasodilatory Response—Brief Report
Authors: Chang Alex C.Y. , Patenaude Alexandre , Lu Katherine , Fuller Megan , Ly Michelle , Kyle Alastair , Golbidi Saeid , Wang Yingjin , Walley Keith , Minchinton Andrew , Laher Ismail , Karsan Aly 
Published: January 2013

Objective—
We have recently described that Notch activates nitric oxide (NO) signaling in the embryonic endocardium. Both Notch signaling and NO signaling have been shown to be important during adult arteriogenesis. Notch has been shown to be required for remodeling of the collateral vessels, whereas NO is required for the initial vasodilatory response to ischemia. Whether Notch also has an impact on the vasodilatory phase of arteriogenesis after ischemia is not known. We tested the hypothesis that endothelial cell-Notch function is required for NO induction and vasodilation, in response to ischemia in the adult vasculature.

Methods and Results—
We observed a significant decrease in NO levels in the dorsal aorta using a mouse model where Notch was inhibited in endothelial cell in a Tet-inducible fashion. In a femoral artery ligation model, inhibition of endothelial cell-Notch reduced reperfusion and NO generation, as quantified by laser Doppler perfusion imaging and by phosphoendothelial NO synthase, nitrotyrosine, and phosphovasodilator-stimulated phosphoprotein staining, respectively.

Conclusion—
Endothelial Notch activation is required for NO production and reactive vasodilation in a femoral artery ligation model.
Leukocyte-Specific CCL3 Deficiency Inhibits Atherosclerotic Lesion Development by Affecting Neutrophil Accumulation
Authors: de Jager Saskia C.A. , Bot Ilze , Kraaijeveld Adriaan O. , Korporaal Suzanne J.A. , Bot Martine , van Santbrink Peter J. , van Berkel Theo J.C. , Kuiper Johan , Biessen Erik A.L. 
Published: January 2013

Objective—
Despite common disbelief that neutrophils are involved in atherosclerosis, evidence is accumulating for a causal role of neutrophils in atherosclerosis. CC chemokine ligand (CCL)3 is an inflammatory chemokine and its expression is significantly increased during atherosclerotic lesion formation in mice. It has recently been shown that under conditions of inflammation neutrophils can migrate along a CCL3 gradient. In this study, we aimed to elucidate the role of leukocyte-derived CCL3 in atherogenesis.

Methods and Results—

Irradiated low density lipoprotein receptor
–/–
mice, reconstituted with CCL3
–/–
or littermate bone marrow showed markedly reduced CCL3 response to lipopolysaccharide treatment, establishing the critical relevance of leukocytes as source of CCL3. Hematopoietic deficiency of CCL3 significantly reduced aortic sinus lesion formation by 31% after 12 weeks of western-type diet. Interestingly, whereas plaque macrophage, collagen, and vascular smooth muscle cell content were unchanged, neutrophil adhesion to and presence in plaques was significantly attenuated in CCL3
–/–
chimeras. These mice had reduced circulating neutrophil numbers, which could be ascribed to an increased neutrophil turnover and CCL3
–/–
neutrophils were shown to be less responsive toward the neutrophil chemoattractant CXC chemokine ligand 1.

Conclusion—
Our data indicate that under conditions of acute inflammation leukocyte-derived CCL3 can induce neutrophil chemotaxis toward the atherosclerotic plaque, thereby accelerating lesion formation.
Pericytes From Infantile Hemangioma Display Proangiogenic Properties and Dysregulated Angiopoietin-1
Authors: Boscolo Elisa , Mulliken John B. , Bischoff Joyce 
Published: January 2013

Objective—
Infantile hemangioma (IH) is a rapidly growing vascular tumor affecting newborns. It is composed of immature endothelial cells and pericytes that proliferate into a disorganized mass of blood vessels. We isolated pericytes from IH (Hem-pericytes) to test our hypothesis that Hem-pericytes are unable to stabilize blood vessels.

Methods and Results—
We injected pericytes in vivo, in combination with endothelial cells, and found that Hem-pericytes formed more microvessels compared with control retinal pericytes. We, thereby, analyzed proangiogenic properties of the Hem-pericytes. They grew fast in vitro, and were unable to stabilize endothelial cell growth and migration, and expressed high levels of vascular endothelial growth factor-A compared with retinal pericytes. Hem-pericytes from proliferating phase IH showed lower contractility in vitro, compared with Hem-pericytes from the involuting phase and retinal pericytes. Consistent with a diminished ability to stabilize endothelium, angiopoietin 1 was reduced in Hem-pericytes compared with retinal pericytes. Normal retinal pericytes in which angiopoietin 1 was silenced produced conditioned medium that stimulated endothelial cell proliferation and migration.

Conclusion—
We report the first successful isolation of patient-derived pericytes from IH tissue. Hem-pericytes exhibited proangiogenic properties and low levels of angiopoietin 1, consistent with a diminished ability to stabilize blood vessels in IH.
Smoking-Thrombolysis Paradox
Authors: Kufner Anna , Nolte Christian H. , Galinovic Ivana , Brunecker Peter , Kufner Gerald M. , Endres Matthias , Fiebach Jochen B. , Ebinger Martin 
Published: January 2013

Background and Purpose—
The so-called smoking-thrombolysis paradox of an improved outcome after thrombolysis was first described in smokers with myocardial infarction. We investigated whether reperfusion rates and clinical outcome differ between smokers and nonsmokers with ischemic stroke after intravenous tissue plasminogen activator.

Methods—
Consecutive acute ischemic stroke patients, who had magnetic resonance imaging before and 1 day after thrombolysis, were included for analysis. All of the patients received intravenous tissue plasminogen activator within 4.5 hours. Reperfusion was defined as a 75% reduction in perfusion deficit (mean transit time >6 s) after thrombolysis compared with baseline. Magnetic resonance angiography was used to evaluate arterial stenosis and occlusion. Functional outcome was assessed 3 months after stroke using the modified Rankin Score.

Results—

Of 148 patients, 21.6% were smokers (n=32). Smokers were younger (median, 61 years [
SD
, 9.4 years] versus 75 years [
SD
, 11.6 years];
P
<0.001), less often women (28% versus 51%;
P
=0.03), had lower baseline glucose levels (median, 6.2 mmol/L [interquartile range, 5.7–6.8 mmol/L] versus 6.7 mmol/L [interquartile range, 6.1–8.2 mmol/L];
P
<0.01) and higher baseline perfusion deficits (median, 53 mL [interquartile range, 13–141 mL] versus 17 mL [interquartile range, 2–66 mL];
P
=0.04). In a backward stepwise regression analysis including age, sex, hypertension, glucose, perfusion deficit, and smoking, smoking had an odds ratio of 4 (95% confidence interval, 1–16;
P
=0.03) for reperfusion and 6 (95% confidence interval, 1–30;
P
=0.05) for recanalization (regression analysis for recanalization also included localization of arterial occlusion). Smokers had a better outcome (modified Rankin Score=0–2) than nonsmokers (77% versus 55%;
P
=0.05).

Conclusions—
Smoking is independently associated with recanalization and reperfusion, indicating that thrombolytic therapy acts more effectively in smokers; because of small numbers, these results should be considered preliminary.

Clinical Trial Registration—

URL:
http://clinicaltrials.gov
. Unique Identifier: NCT00715533.