bluuebunny/crossref_metadata_2025 huggingface dataset:
Independent Ancient Polyploidy Events in the Sister Families Brassicaceae and Cleomaceae
Authors: Schranz M. Eric , Mitchell-Olds Thomas
Published: April 2006

Recent studies have elucidated the ancient polyploid history of the Arabidopsis thaliana (Brassicaceae) genome. The studies concur that there was at least one polyploidy event occurring some 14.5 to 86 million years ago (Mya), possibly near the divergence of the Brassicaceae from its sister family, Cleomaceae. Using a comparative genomics approach, we asked whether this polyploidy event was unique to members of the Brassicaceae, shared with the Cleomaceae, or an independent polyploidy event in each lineage. We isolated and sequenced three genomic regions from diploid Cleome spinosa (Cleomaceae) that are each homoeologous to a duplicated region shared between At3 and At5, centered on the paralogs of SEPALLATA (SEP) and CONSTANS (CO). Phylogenetic reconstructions and analysis of synonymous substitution rates support the hypothesis that a genomic triplication in Cleome occurred independently of and more recently than the duplication event in the Brassicaceae. There is a strong correlation in the copy number (single versus duplicate) of individual genes, suggesting functionally consistent influences operating on gene copy number in these two independently evolving lineages. However, the amount of gene loss in Cleome is greater than in Arabidopsis. The genome of C. spinosa is only 1.9 times the size of A. thaliana, enabling comparative genome analysis of separate but related polyploidy events.
Factors Associated with High Milk Test Day Somatic Cell Counts in Large Dairy Herds in Brandenburg. II. Milking Practices
Authors: Köster G. , Tenhagen B.‐A. , Scheibe N. , Heuwieser W.
Published: April 2006

The objective of this study was to examine the influences of different milking practices on cow udder health in 80 large dairy herds (range 100–1100 cows) in Brandenburg, Germany. Milking practices were evaluated during one complete milking using a standardized data capture form. The somatic cell count (SCC) of all lactating cows on each farm was determined monthly by the local milk recording association ‘Landeskontrollverband Brandenburg’. Factor analysis was used to investigate the relationship between the different aspects of the milking practices. The components extracted by the factor analysis were examined for their influence on the SCC of the current month (CMSCC) and the year before the visit (YASCC) using univariate analysis of variance. Three components were extracted from the milking practices. ‘Reasonable use of water’ was significantly related to CMSCC (P = 0.019) and YASCC (P = 0.003). It included information on the use of a hose to clean udders before milking, cleaning of the floor between groups and use of water to clean teats. ‘Attention of the milkers’ was also significantly associated with CMSCC (P = 0.012) and YASCC (P = 0.014). It included information on the accuracy of mastitis detection by foremilk screening and the regular use of post‐milking teat and cluster disinfection. The component ‘preparation routines’ (method of udder cleaning and forestripping) did not significantly influence CMSCC and YASCC. These results indicate that excessive use of water in the parlour during milking time is harmful to udder health and that the consistency of procedures in the milking parlour presents significant room for improvement in large dairy herds in Brandenburg.
Developmental signalling pathways in lung cancer
Authors: DANIEL Vincent C. , PEACOCK Craig D. , WATKINS D. Neil
Published: April 2006

Abstract:  Hedgehog, Notch and Wnt signalling are all essential for axial patterning and progenitor cell fates in signalling pathways conserved from flies to humans. Aberrant activation of these pathways is observed in a wide variety of cancers, suggesting that these embryonic signalling pathways contribute in a fundamental way to the evolution and maintenance of a malignant phenotype. Because all three of these pathways participate in lung development, recent studies have begun to explore the connection between lung development, airway epithelial repair and lung cancer. Development, repair and malignant transformation of the neuroendocrine lineage are all accompanied by aberrant Hedgehog pathway activation, whereas Notch and Wnt signalling may be important in other airway cell types. Small molecule targeting of these pathways may provide therapeutic opportunities in lung cancer. The plant‐derived alkaloid cyclopamine is a naturally occurring Hedgehog pathway inhibitor that shows therapeutic promise in small cell lung cancer, a highly aggressive neuroendocrine tumour. A more detailed understanding of how embryonic signalling pathways participate in airway epithelial repair and tumourigenesis may reveal more novel therapeutic vulnerabilities in lung cancer.
CROSS-COUNTRY HIERARCHICAL STRUCTURE AND CURRENCY CRISES
Authors: ORTEGA GUILLERMO J. , MATESANZ DAVID
Published: April 2006

Using data from a sample of 28 representatives countries, we propose a classification of currency crises consequences based on the ultrametric analysis of the real exchange rate movements time series, without any further assumption. By using the matrix of synchronous linear correlation coefficients and the appropriate metric distance between pairs of countries, we were able to construct a hierarchical tree of countries. This economic taxonomy provides relevant information regarding liaisons between countries and a meaningful insight about the contagion phenomenon.
CONSTRAINTS ON MASS MATRICES DUE TO MEASURED PROPERTY OF THE MIXING MATRIX
Authors: CHATURVEDI S. , GUPTA VIRENDRA
Published: April 2006

It is shown that two specific properties of the unitary matrix V can be expressed directly in terms of the matrix elements and eigenvalues of the hermitian matrix M which is diagonalized by V. These are the asymmetry Δ(V) = |V12|2-|V21|2, of V with respect to the main diagonal and the Jarlskog invariant [Formula: see text]. These expressions for Δ(V) and J(V) provide constraints on possible mass matrices from the available data on V.
SIMULATION OF ELECTRON SWARM PARAMETERS IN AIR
Authors: SETTAOUTI A. , SETTAOUTI L.
Published: April 2006

The electron transport of air in a uniform electric field is investigated by a Monte–Carlo simulation. The simulation results obtained are compared with the available data in the literature. The result of Monte–Carlo simulation shows that electron molecule collision cross sections adopted in the simulation result in good agreement with the experimental values over the range of E/N investigated (E is the electric field and N is the gas number density of background gas molecules).
INFLUENCE OF THE GUPTA POTENTIAL ON THE MELTING BEHAVIORS OF CLUSTERS
Authors: REN SHI-WEI
Published: April 2006

By using the standard microcanonical molecular-dynamics simulations, the melting behaviors of the clusters bound by Gupta potentials are investigated. The calculations indicate that these clusters can show different melting behaviors. The origins of these processes are studied. The effects of the attractive range and the repulsive part of the interatomic interaction on the melting behaviors of the clusters are compared and analyzed.
Cardiac fibroblasts: friend or foe?
Authors: Baudino Troy A. , Carver Wayne , Giles Wayne , Borg Thomas K.
Published: April 2006

Cardiac function is determined by the dynamic interaction of various cell types and the extracellular matrix that composes the heart. This interaction varies with the stage of development and the degree and duration of mechanical, chemical, and electrical signals between the various cell types and the ECM. Understanding how these complex signals interact at the molecular, cellular, and organ levels is critical to understanding the function of the heart under a variety of physiological and pathophysiological conditions. Quantitative approaches, both in vivo and in vitro, are essential to understand the dynamic interaction of mechanical, chemical, and electrical stimuli that govern cardiac function. The fibroblast can thus be a friend in normal function or a foe in pathophysiological conditions.
Sodium current function in adult and aged canine atrial cells
Authors: Baba Shigeo , Dun Wen , Hirose Masanori , Boyden Penelope A.
Published: April 2006

The incidence of atrial fibrillation increases with age, but it is unknown whether there are changes in the intrinsic function of Na+ currents in cells of the aged atria. Thus, we studied right (RA) and left (LA) atrial cells from two groups of dogs, adult and aged (>8 yr), to determine the change in Na+ currents with age. In this study all dogs were in normal sinus rhythm. Whole cell voltage clamp techniques were used to compare the Na+ currents in the two cell groups. Immunocytochemical studies were completed for the Na+ channel protein Nav1.5 to determine whether there was structural remodeling of this protein with age. In cells from aged animals, we found that Na+ currents are similar to those we measured in adult atria. However, Na+ current ( INa) density of the aged atria differed depending on the atrial chamber with LA cell currents being larger than RA cell currents. Thus with age, the difference in INa density between atrial chambers remains. INa kinetic differences between aged and adult cells included a significant acceleration into the inactivated state and an enhanced use-dependent decrease in peak current in aged RA cells. Finally, there is no structural remodeling of the cardiac Na+ channel protein Nav1.5 in the aged atrial cell. In conclusion, with age there is no change in INa density, but there are subtle kinetic differences contributing to slight enhancement of use dependence. There is no structural remodeling of the fast Na+ current protein with age.
A polymerized bovine hemoglobin oxygen carrier preserves regional myocardial function and reduces infarct size after acute myocardial ischemia
Authors: George Isaac , Yi Geng-Hua , Schulman Allison R. , Morrow Brad T. , Cheng Yanping , Gu Anguo , Zhang Geping , Oz Mehmet C. , Burkhoff Daniel , Wang Jie
Published: April 2006

The purpose of this study was to test if HBOC-201, a hemoglobin-based oxygen-carrying solution, can decrease infarct size (or Inf) during acute, severe myocardial ischemia and reperfusion. To test the impact of HBOC-201 on infarct size, ischemia was produced in 18 dogs by coronary stenosis to achieve 80–95% flow reduction for 195 min along with pacing 10% above the spontaneous heart rate, followed by 180 min of reperfusion. Animals were randomized to intravenous infusion of HBOC-201 (1 g/kg) ( n = 6), normal saline (NS) ( n = 6), or phenylephrine (Phe) ( n = 6, as a control for the increased blood pressure seen with HBOC-201), given 15 min after the start of ischemia. Amount of infarct was quantified as the ratio between area at risk (AAR) and Inf after Evans blue and 2,3,5-triphenyltetrazolium chloride staining. Hearts were divided into five layers from base ( layer A) to apex ( layer E) and photographed for digital image analysis of AAR and Inf. Regional myocardial function (RMF) was also measured after 60 min of ischemia and 15 min of reperfusion. Inf/AAR was significantly reduced after HBOC-201 therapy (4.4 ± 2.2%) vs. NS (26.0 ± 3.6%) and Phe (25.7 ± 4.1%) (both, P < 0.05). RMF after reperfusion was restored to 92% of baseline with HBOC-201 compared with 11% of baseline after NS ( P < 0.05) and 49% after Phe ( P = not significant). HBOC-201 administration after induction of severe myocardial ischemia by acute coronary stenosis reduces infarct size and improves myocardial viability.
Novel role for K+-dependent Na+/Ca2+exchangers in regulation of cytoplasmic free Ca2+and contractility in arterial smooth muscle
Authors: Dong Hui , Jiang Yanfen , Triggle Chris R. , Li Xiaofang , Lytton Jonathan
Published: April 2006

Cytoplasmic free Ca2+([Ca2+]cyt) is essential for the contraction and relaxation of blood vessels. The role of plasma membrane Na+/Ca2+exchange (NCX) activity in the regulation of vascular Ca2+homeostasis was previously ascribed to the NCX1 protein. However, recent studies suggest that a relatively newly discovered K+-dependent Na+/Ca2+exchanger, NCKX (gene family SLC24), is also present in vascular smooth muscle. The purpose of the present study was to identify the expression and function of NCKX in arteries. mRNA encoding NCKX3 and NCKX4 was demonstrated by RT-PCR and Northern blot in both rat mesenteric and aortic smooth muscle. NCXK3 and NCKX4 proteins were also demonstrated by immunoblot and immunofluorescence. After voltage-gated Ca2+channels, store-operated Ca2+channels, and Na+pump were pharmacologically blocked, when the extracellular Na+was replaced with Li+(0 Na+) to induce reverse mode (Ca2+entry) activity of Na+/Ca2+exchangers, a large increase in [Ca2+]cytsignal was observed in primary cultured aortic smooth muscle cells. About one-half of this [Ca2+]cytsignal depended on the extracellular K+. In addition, after the activity of NCX was inhibited by KB-R7943, Na+replacement-induced Ca2+entry was absolutely dependent on extracellular K+. In arterial rings denuded of endothelium, a significant fraction of the phenylephrine-induced and nifedipine-resistant aortic or mesenteric contraction could be prevented by removal of extracellular K+. Taken together, these data provide strong evidence for the expression of NCKX proteins in the vascular smooth muscle and their novel role in mediating agonist-stimulated [Ca2+]cytand thereby vascular tone.
Pulsatile blood flow in the entire coronary arterial tree: theory and experiment
Authors: Huo Yunlong , Kassab Ghassan S.
Published: April 2006

The pulsatility of coronary circulation can be accurately simulated on the basis of the measured branching pattern, vascular geometry, and material properties of the coronary vasculature. A Womersley-type mathematical model is developed to analyze pulsatile blood flow in diastole in the absence of vessel tone in the entire coronary arterial tree on the basis of previously measured morphometric data. The model incorporates a constitutive equation of pressure and cross-section area relation based on our previous experimental data. The formulation enables the prediction of the impedance, the pressure distribution, and the pulsatile flow distribution throughout the entire coronary arterial tree. The model is validated by experimental measurements in six diastolic arrested, vasodilated porcine hearts. The agreement between theory and experiment is excellent. Furthermore, the present pulse wave results at low frequency agree very well with previously published steady-state model. Finally, the phase angle of flow is seen to decrease along the trunk of the major coronary artery and primary branches toward the capillary vessels. This study represents the first, most extensive validated analysis of Womersley-type pulse wave transmission in the entire coronary arterial tree down to the first segment of capillaries. The present model will serve to quantitatively test various hypotheses in the coronary circulation under pulsatile flow conditions.
Inward rectifying potassium channels facilitate cell-to-cell communication in hamster retractor muscle feed arteries
Authors: Jantzi Micaela C. , Brett Suzanne E. , Jackson William F. , Corteling Randolph , Vigmond Edward J. , Welsh Donald G.
Published: April 2006

This study examined whether inward rectifying K+(KIR) channels facilitate cell-to-cell communication along skeletal muscle resistance arteries. With the use of feed arteries from the hamster retractor muscle, experiments examined whether KIRchannels were functionally expressed and whether channel blockade attenuated the conduction of acetylcholine-induced vasodilation, an index of cell-to-cell communication. Consistent with KIRchannel expression, this study observed the following: 1) a sustained Ba2+-sensitive, K+-induced dilation in preconstricted arteries; 2) a Ba2+-sensitive inwardly rectifying K+current in arterial smooth muscle cells; and 3) KIR2.1 and KIR2.2 expression in the smooth muscle layer of these arteries. It was subsequently shown that the discrete application of acetylcholine elicits a vasodilation that conducts with limited decay along the feed artery wall. In the presence of 100 μM Ba2+, the local and conducted response to acetylcholine was attenuated, a finding consistent with a role for KIRin facilitating cell-to-cell communication. A computational model of vascular communication accurately predicted these observations. Control experiments revealed that in contrast to Ba2+, ATP-sensitive- and large-conductance Ca2+activated-K+channel inhibitors had no effect on the local or conducted vasodilatory response to acetylcholine. We conclude that smooth muscle KIRchannels play a key role in facilitating cell-to-cell communication along skeletal muscle resistance arteries. We attribute this facilitation to the intrinsic property of negative slope conductance, a biophysical feature common to KIR2.1- and 2.2-containing channels, which enables them to increase their activity as a cell hyperpolarizes.
Does Hyperglycemia Reduce Proliferation or Increase Apoptosis?
Authors: Davidson Sean M. , Yellon Derek M.
Published: April 2006

Diabetes mellitus is a major risk factor for the development of vascular complications. We hypothesized that hyperglycemia decreases endothelial cell (EC) proliferation and survival via phosphatidylinositol 3-kinase (PI3k) and Akt signaling pathways. We cultured human umbilical vein ECs (HUVEC) in 5, 20, or 40 mM d-glucose. Cells grown in 5, 20, and 40 mM mannitol served as a control for osmotic effects. We measured EC proliferation for up to 15 days. We assessed apoptosis by annexin V and propidium iodide staining and flow cytometry, analyzed cell lysates obtained on culture day 8 for total and phosphorylated PI3k and Akt by Western blot analysis, and measured Akt kinase activity using a GSK fusion protein. HUVEC proliferation was also tested in the presence of pharmacological inhibitors of PI3k-Akt (wortmannin and LY294002) and after transfection with a constitutively active Akt mutant. ECs in media containing 5 mM d-glucose (control) exhibited log-phase growth on days 7–10. d-Glucose at 20 and 40 mM significantly decreased proliferation versus control ( P < 0.05 for both), whereas mannitol did not impair EC proliferation. Apoptosis increased significantly in HUVEC exposed to 40 mM d-glucose. d-Glucose at 40 mM significantly decreased tyrosine-phosphorylated PI3k, threonine 308-phosphorylated-Akt, and Akt activity relative to control 5 mM d-glucose. Pharmacological inhibition of PI3k-Akt resulted in a dose-dependent decrease in EC proliferation. Transfection with a constitutively active Akt mutant protected ECs by enhancing proliferation when grown in 20 and 40 mM d-glucose. We conclude that d-glucose regulates Akt signaling through threonine phosphorylation of Akt and that hyperglycemia-impaired PI3k-Akt signaling may promote EC proliferative dysfunction in diabetes.
Neutrophil activation is modulated by sex hormones after trauma-hemorrhagic shock and burn injuries
Authors: Deitch Edwin A. , Ananthakrishnan Preya , Cohen David B. , Xu Da Zhong , Feketeova Eleonora , Hauser Carl J.
Published: April 2006

Recent literature indicates that females are more resistant to shock-, trauma-, and sepsis-induced immune dysfunction and organ injury than are males. Consequently, using trauma-hemorrhagic shock (T/HS) and burn models, we tested whether the neutrophil response to trauma occurred in a sexually dimorphic fashion and, if so, the role of sex hormones. Neutrophil activation, as reflected by CD11b expression and respiratory burst activity, was increased to a greater extent in male rats than in female rats after T/HS or burn injury. Testosterone appeared to potentiate neutrophil activation, because castration reduced neutrophil activation, whereas ovariectomy had little effect. Mechanistically, this sexually dimorphic neutrophil response appeared to be due to both cellular and humoral factors. Evidence for a cellular difference between male and female neutrophils is based on the observation that naive female neutrophils were more resistant to activation by burn or T/HS plasma and lymph than naive male neutrophils and that this resistance varied over the estrus cycle. Additionally, the humoral environment was more neutrophil activating in male rats, because burn and T/HS plasma and lymph from male rats activated naive male neutrophils to a greater extent than comparable samples from females. Last, on the basis of in vitro experiments examining the effects of estrogen on calcium signaling, it appears that estrogen limits trauma-induced neutrophil activation, at least in part, by limiting the entry of calcium into the cell via store-operated calcium entry mechanisms. In conclusion, there is a striking sexual dimorphism in neutrophil responses after trauma, and these changes reflect both cellular resistance to activation as well as a less activating humoral environment.
Chronic systemic delivery of angiopoietin-2 reveals a possible independent angiogenic effect
Authors: Bureau W. , Van Slyke P. , Jones J. , Han R. N. N. , Ward Nicole L. , Stewart D. J. , Dumont D. J.
Published: April 2006

Angiopoietin-2 has been implicated in the angiogenic response; however, this response has been tied to the expression of VEGF, and an independent angiogenic role has yet to be described. In this report, we detail the generation of transgenic mice that conditionally express angiopoietin-2 in the liver, resulting in sustained increases in circulating levels. These animals survive gestation and present with several vascular abnormalities, including an increase in the diameter of myocardial coronary vessels and a reduction in the density of endocardial vessels. In the lung, prominent increases in vessel diameter were observed. These vascular remodeling changes occurred in the absence of any apparent increase in VEGF expression. Our results illustrate that chronic systemic delivery of angiopoietin-2 induces angiogenesis in the absence of increased VEGF expression and that angiopoietin-2 promotes myocardial coronary vessel remodeling.
Exercise training improves aging-induced downregulation of VEGF angiogenic signaling cascade in hearts
Authors: Iemitsu Motoyuki , Maeda Seiji , Jesmin Subrina , Otsuki Takeshi , Miyauchi Takashi
Published: April 2006

Exercise training improves aging-induced deterioration of angiogenesis in the heart. However, the mechanisms underlying exercise-induced improvement of capillary density in the aged heart are unclear. Vascular endothelial growth factor (VEGF) is implicated in angiogenesis, which activated angiogenic signaling cascade through Akt and endothelial nitric oxide synthase (eNOS)-related pathway. We hypothesized that VEGF angiogenic signaling cascade in the heart contributes to a molecular mechanism of exercise training-induced improvement of capillary density in old age. With the use of hearts of sedentary young rats (4 mo old), sedentary aged rats (23 mo old), and exercise-trained aged rats (23 mo old, swim training for 8 wk), the present study investigated whether VEGF and VEGF-related angiogenic molecular expression in the aged heart is affected by exercise training. Total capillary density in the heart was significantly lower in the sedentary aged rats compared with the sedentary young rats, whereas that in the exercise-trained rat was significantly higher than the sedentary aged rats. The mRNA and protein expressions of VEGF and of fms-like tyrosine kinase-1 (Flt-1) and fetal liver kinase-1 (Flk-1), which are main VEGF receptors, in the heart were significantly lower in the sedentary aged rats compared with the sedentary young rats, whereas those in the exercise-trained rats were significantly higher than those in the sedentary aged rats. The phosphorylation of Akt protein and eNOS protein in the heart corresponded to the changes in the VEGF protein levels. These findings suggest that exercise training improves aging-induced downregulation of cardiac VEGF angiogenic signaling cascade, thereby contributing to the exercise training-induced improvement of angiogenesis in old age.
Disruption of COX-2 modulates gene expression and the cardiac injury response to doxorubicin
Authors: Neilan Tomas G. , Doherty Glen A. , Chen Gang , Deflandre Catherine , McAllister Hester , Butler Ryan K. , McClelland Sarah E. , Kay Elaine , Ballou Leslie R. , Fitzgerald Desmond J.
Published: April 2006

To determine the role of cyclooxygenase (COX)-2 in anthracycline-induced cardiac toxicity, we administered doxorubicin (Dox) to mice with genetic disruption of COX-2 (COX-2−/−). After treatment with Dox, COX-2−/−mice had increased cardiac dysfunction and cardiac cell apoptosis compared with Dox-treated wild-type mice. The expression of the death-associated protein kinase-related apoptosis-inducing protein kinase-2 was also increased in Dox-treated COX-2−/−animals. The altered gene expression, cardiac injury, and dysfunction after Dox treatment in COX-2−/−mice was attenuated by a stable prostacyclin analog, iloprost. Wild-type mice treated with Dox developed cardiac fibrosis that was absent in COX-2−/−mice and unaffected by iloprost. These results suggest that genetic disruption of COX-2 increases the cardiac dysfunction after treatment with Dox by an increase in cardiac cell apoptosis. This Dox-induced cardiotoxicity in COX-2−/−mice was attenuated by a prostacyclin analog, suggesting a protective role for prostaglandins in this setting.
Thyroid hormone downregulates the expression and function of sarcoplasmic reticulum-associated CaM kinase II in the rabbit heart
Authors: Jiang Mao , Xu Ande , Narayanan Njanoor
Published: April 2006

Phosphorylation of sarcoplasmic reticulum (SR) Ca2+-cycling proteins by a membrane-associated Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) is a well-documented physiological mechanism for regulation of transmembrane Ca2+fluxes and the cardiomyocyte contraction-relaxation cycle. The present study investigated the effects of l-thyroxine-induced hyperthyroidism on protein expression of SR CaM kinase II and its substrates, endogenous CaM kinase II-mediated SR protein phosphorylation, and SR Ca2+pump function in the rabbit heart. Membrane vesicles enriched in junctional SR (JSR) or longitudinal SR (LSR) isolated from euthyroid and hyperthyroid rabbit hearts were utilized. Endogenous CaM kinase II-mediated phosphorylation of ryanodine receptor-Ca2+release channel (RyR-CRC), Ca2+-ATPase, and phospholamban (PLN) was significantly lower (30–70%) in JSR and LSR vesicles from hyperthyroid than from euthyroid rabbit heart. Western immunoblotting analysis revealed significantly higher (∼40%) levels of sarco(endo)plasmic reticulum Ca2+-ATPase isoform 2 (SERCA2) in JSR, but not in LSR, from hyperthyroid than from euthyroid rabbit heart. Maximal velocity of Ca2+uptake was significantly increased in JSR (130%) and LSR (50%) from hyperthyroid compared with euthyroid rabbit hearts. Apparent affinity of the Ca2+-ATPase for Ca2+did not differ between the two groups. Protein levels of PLN and CaM kinase II were significantly lower (30–40%) in JSR, LSR, and ventricular tissue homogenates from hyperthyroid rabbit heart. These findings demonstrate selective downregulation of expression and function of CaM kinase II in hyperthyroid rabbit heart in the face of upregulated expression and function of SERCA2 predominantly in the JSR compartment.
Protective effects of epoxyeicosatrienoic acids on human endothelial cells from the pulmonary and coronary vasculature
Authors: Dhanasekaran Anuradha , Al-Saghir Rula , Lopez Bernardo , Zhu Daling , Gutterman David D. , Jacobs Elizabeth R. , Medhora Meetha
Published: April 2006

Epoxyeicosatrienoic acids (EETs) are cytochrome P-450 (CYP) metabolites synthesized from the essential fatty acid arachidonic acid to generate four regioisomers, 14,15-, 11,12-, 8,9-, and 5,6-EET. Cultured human coronary artery endothelial cells (HCAECs) contain endogenous EETs that are increased by stimulation with physiological agonists such as bradykinin. Because EETs are known to modulate a number of vascular functions, including angiogenesis, we tested each of the four regioisomers to characterize their effects on survival and apoptosis of HCAECs and cultured human lung microvascular endothelial cells (HLMVECs). A single application of physiologically relevant concentration of 14,15-, 11,12-, and 8,9-EET but not 5,6-EET (0.75–300 nM) promoted concentration-dependent increase in cell survival of HLMVECs and HCAECs after removal of serum. The lipids also protected the same cells from death via the intrinsic, as well as extrinsic, pathways of apoptosis. EETs did not increase intracellular calcium concentration ([Ca2+]i) or phosphorylate mitogen-activated protein kinase p44/42 when applied to these cells, and their protective action was attenuated by the phosphotidylinositol-3 kinase inhibitor wortmannin (10 μM) but not the cyclooxygenase inhibitor indomethacin (20 μM). Our results demonstrate for the first time the capacity of EETs to enhance human endothelial cell survival by inhibiting both the intrinsic, as well as extrinsic, pathways of apoptosis, an important underlying mechanism that may promote angiogenesis and endothelial survival during atherosclerosis and related cardiovascular ailments.
Cardioprotective mechanisms ofPrunus cerasus(sour cherry) seed extract against ischemia-reperfusion-induced damage in isolated rat hearts
Authors: Bak Istvan , Lekli Istvan , Juhasz Bela , Nagy Norbert , Varga Edit , Varadi Judit , Gesztelyi Rudolf , Szabo Gergo , Szendrei Levente , Bacskay Ildiko , Vecsernyes Miklos , Antal Miklos , Fesus Laszlo , Boucher Francois , de Leiris Joel , Tosaki Arpad
Published: April 2006

The effects of kernel extract obtained from sour cherry ( Prunus cerasus) seed on the postischemic cardiac recovery were studied in isolated working rat hearts. Rats were treated with various daily doses of the extract for 14 days, and hearts were then isolated and subjected to 30 min of global ischemia followed by 120 min of reperfusion. The incidence of ventricular fibrillation (VF) and tachycardia (VT) fell from their control values of 92% and 100% to 50% (not significant) and 58% (not significant), 17% ( P < 0.05), and 25% ( P < 0.05) with the doses of 10 mg/kg and 30 mg/kg of the extract, respectively. Lower concentrations of the extract (1 and 5 mg/kg) failed to significantly reduce the incidence of VF and VT during reperfusion. Sour cherry seed kernel extract (10 and 30 mg/kg) significantly improved the postischemic recovery of cardiac function (coronary flow, aortic flow, and left ventricular developed pressure) during reperfusion. We have also demonstrated that the extract-induced protection in cardiac function significantly reflected in a reduction of infarct size. Immunohistochemistry indicates that a reduction in caspase-3 activity and apoptotic cells by the extract, beside other potential action mechanisms of proanthocyanidin, trans-resveratrol, and flavonoid components of the extract, could be responsible for the cardioprotection in ischemic-reperfused myocardium.
Redox regulation of endogenous substrate oxidation by cardiac mitochondria
Authors: Korge Paavo , Weiss James N.
Published: April 2006

Reactive oxygen species (ROS) play important roles in regulating mitochondrial function, as well as in ischemia-reperfusion injury and cardioprotection. Here we show that, in the absence of exogenous substrates, cardiac mitochondria have a surprisingly large capacity to phosphorylate ADP by oxidizing endogenous substrates, provided that H2O2is removed from the extramitochondrial environment and a reduced environment is maintained in the matrix. In isolated mitochondria without exogenous substrates, addition of catalase and the membrane-permeant reducing agent N-acetylcysteine (Nac) or the ROS scavenger mercaptopropionyl glycine significantly increased the ability to phosphorylate added ADP, as demonstrated by 1) full recovery of membrane potential (Δψ) and matrix volume from ADP-induced dissipation and shrinkage, 2) ADP-dependent increase in O2consumption, and 3) enhanced rate of ATP synthesis. Removal of extramitochondrial H2O2by catalase was required to stimulate endogenous substrate oxidation, as shown by the increase in O2consumption and Δψ. This effect was greatly enhanced by addition of Nac or mercaptopropionyl glycine to suppress oxidation-induced ROS increases in the matrix. Theoretical considerations, as well as reversible inhibition of O2consumption with 3-mercaptopropionic acid and pyruvate in state 3, indicate that these substrates are fatty acids. Under in vivo conditions in which powerful antioxidant conditions are maintained, this mechanism may be important in stimulation of β-oxidation and ATP production at low levels of extramitochondrial fatty acids. Incapacitation of this mechanism may potentially contribute to mitochondrial dysfunction during oxidative stress.
Sp3 inhibits Sp1-mediated activation of the cardiac troponin T promoter and is downregulated during pathological cardiac hypertrophy in vivo
Authors: Azakie Anthony , Fineman Jeffrey R. , He Youping
Published: April 2006

Combinatorial interactions between cis elements and trans-acting factors are required for regulation of cardiac gene expression during normal cardiac development and pathological cardiac hypertrophy. Sp factors bind GC boxes and are implicated in recruitment and assembly of the basal transcriptional complex. In this study, we show that the cardiac troponin T (cTnT) promoter contains a GC box that is necessary for basal and cAMP-mediated activity of cTnT promoter constructs transfected in embryonic cardiomyocytes. Cardiac nuclear proteins bind the cTnT GC box in a sequence-specific fashion and consist of Sp1, Sp2, and Sp3 protein factors. By chromatin immunoprecipitation, Sp1 binds the cTnT promoter “in vivo.” Cotransfected Sp1 trans-activates the cTnT promoter in cardiomyocytes in culture. Sp3 represses Sp1-mediated transcriptional activation of the cTnT gene in embryonic cardiomyocytes. Sp3 repression of Sp1-mediated cTnT promoter activation is dose dependent, inferring a mechanism of competitive binding/inhibition. To evaluate the role of Sp factors in cardiac gene expression in vivo, we have established a clinically relevant animal model of pathological cardiac hypertrophy where the fetal cardiac program is activated. In this animal model, cardiac hypertrophy results from increased left-right shunting, volume loading of the left ventricle, and pressure loading of the right ventricle. Sp1 expression is increased in all four hypertrophied cardiac chambers, whereas Sp3 expression is diminished. This observation is consistent with the in vitro activating function of Sp1 and inhibitory effects of Sp3 on activity of cTnT promoter constructs. Sp factor levels are modulated during the hypertrophic cardiac program in vivo.
Restoring depressed HERG K+ channel function as a mechanism for insulin treatment of abnormal QT prolongation and associated arrhythmias in diabetic rabbits
Authors: Zhang Yiqiang , Xiao Jiening , Wang Huizhen , Luo Xiaobin , Wang Jingxiong , Villeneuve Louis R. , Zhang Haiqing , Bai Yunlong , Yang Baofeng , Wang Zhiguo
Published: April 2006

Abnormal QT prolongation (QT-P) in diabetic patients has become a nonnegligible clinical problem and has attracted increasing attention from basic scientists, because it increases the risk of lethal ventricular arrhythmias. Correction of QT-P may be an important measure in minimizing sudden cardiac death in diabetic patients. Here we report the efficacy of insulin in preventing QT-P and the associated arrhythmias and the mechanisms underlying the effects in a rabbit model of type 1 insulin-dependent diabetes mellitus (IDDM). The heart rate-corrected QT (QTc) interval and action potential duration were considerably prolonged, with frequent ventricular tachycardias. The rapid delayed rectifier K+ current ( IKr) was markedly reduced in IDDM hearts, and hyperglycemia depressed the function of the human ether-a-go-go-related gene (HERG), which conducts IKr. The impairment was primarily ascribed to the enhanced oxidative damage to the myocardium, as indicated by the increased intracellular level of reactive oxygen species and simultaneously decreased endogenous antioxidant reserve and by the increased lipid peroxidation and protein oxidation. Moreover, IDDM or hyperglycemia resulted in downregulation of HERG protein level. Insulin restored the depressed IKr/HERG and prevented QTc/action potential duration prolongation and the associated arrhythmias, and the beneficial actions of insulin are partially due to its antioxidant ability. Our study represents the first documentation of oxidative stress as the major metabolic mechanism for HERG K+ dysfunction, which causes diabetic QT-P, and suggests IKr/HERG as a potential therapeutic target for treatment of the disorder.
Activation of endoplasmic reticulum stress response during the development of ischemic heart disease
Authors: Azfer Asim , Niu Jianli , Rogers Linda M. , Adamski Frances M. , Kolattukudy Pappachan E.
Published: April 2006

Endoplasmic reticulum (ER) stress has been found to be associated with neurodegenerative diseases and diabetes mellitus. Whether ER stress is involved in the development of heart disease is not known. Cardiac-specific expression of monocyte chemoattractant protein-1 (MCP-1) in mice causes the development of ischemic heart disease. Here we report that microarray analysis of gene expression changes in the heart of these transgenic mice revealed that a cluster of ER stress-related genes was transcriptionally activated in the heart during the development of ischemic heart disease. The gene array results were verified by quantitative real-time PCR that showed highly elevated transcript levels of genes involved in unfolded protein response such as ER and cytoplasmic chaperones, oxidoreductases, protein disulfide isomerase (PDI) family, and ER-associated degradation system such as ubiquitin. Immunoblot analysis confirmed the expression of chaperones, PDI, and ubiquitin. Immunohistochemical analyses showed that ER stress proteins were associated mainly with the degenerating cardiomyocytes. A novel ubiquitin fold modifier (Ufm1) that has not been previously associated with ER stress and not found to be induced under any condition was also found to be upregulated in the hearts of MCP mice (transgenic mice that express MCP-1 specifically in the heart). The present results strongly suggest that activation of ER stress response is involved in the development of ischemic heart disease in this murine model.
Detection of allergen-induced airway hyperresponsiveness in isolated mouse lungs
Authors: Witzenrath Martin , Ahrens Birgit , Kube Stefanie M. , Braun Armin , Hoymann Heinz G. , Hocke Andreas C. , Rosseau Simone , Suttorp Norbert , Hamelmann Eckard , Schütte Hartwig
Published: April 2006

Airway hyperresponsiveness (AHR) is a hallmark of bronchial asthma. Important features of this exaggerated response to bronchoconstrictive stimuli have mostly been investigated in vivo in intact animals or in vitro in isolated tracheal or bronchial tissues. Both approaches have important advantages but also certain limitations. Therefore, the aim of our study was to develop an ex vivo model of isolated lungs from sensitized mice for the investigation of airway responsiveness (AR). BALB/c mice were sensitized by intraperitoneal ovalbumin (Ova) and subsequently challenged by Ova inhalation. In vivo AR was measured in unrestrained animals by whole body plethysmography after stimulation with aerosolized methacholine (MCh) with determination of enhanced pause ( Penh). Twenty-four hours after each Penh measurement, airway resistance was continuously registered in isolated, perfused, and ventilated lungs on stimulation with inhaled or intravascular MCh or nebulized Ova. In a subset of experiments, in vivo AR was additionally measured in orotracheally intubated, spontaneously breathing mice 24 h after Penh measurement, and lungs were isolated further 24 h later. Isolated lungs of allergen-sensitized and -challenged mice showed increased AR after MCh inhalation or infusion as well as after specific provocation with aerosolized allergen. AR was increased on days 2 and 5 after Ova challenge and had returned to baseline on day 9. AHR in isolated lungs after aerosolized or intravascular MCh strongly correlated with in vivo AR. Pretreatment of isolated lungs with the β2-agonist fenoterol diminished AR. In conclusion, this model provides new opportunities to investigate mechanisms of AHR as well as pharmacological interventions on an intact organ level.
Role of caveolin-1 in p42/p44 MAP kinase activation and proliferation of human airway smooth muscle
Authors: Gosens Reinoud , Stelmack Gerald L. , Dueck Gordon , McNeill Karol D. , Yamasaki Akira , Gerthoffer William T. , Unruh Helmut , Gounni Abdelilah Soussi , Zaagsma Johan , Halayko Andrew J
Published: April 2006

Chronic airways diseases, including asthma, are associated with an increased airway smooth muscle (ASM) mass, which may contribute to chronic airway hyperresponsiveness. Increased muscle mass is due, in part, to increased ASM proliferation, although the precise molecular mechanisms for this response are not completely clear. Caveolae, which are abundant in smooth muscle cells, are membrane microdomains where receptors and signaling effectors can be sequestered. We hypothesized that caveolae and caveolin-1 play an important regulatory role in ASM proliferation. Therefore, we investigated their role in p42/p44 MAPK signaling and proliferation using human ASM cell lines. Disruption of caveolae using methyl-β-cyclodextrin and small interfering (si)RNA-knockdown of caveolin-1 caused spontaneous p42/p44 MAPK activation; additionally, caveolin-1 siRNA induced ASM proliferation in mitogen deficient conditions, suggesting a key role for caveolae and caveolin-1 in maintaining quiescence. Moreover, caveolin-1 accumulates twofold in myocytes induced to a contractile phenotype compared with proliferating ASM cells. Caveolin-1 siRNA failed to increase PDGF-induced p42/p44 MAPK activation and cell proliferation, however, indicating that PDGF stimulation actively reversed the antimitogenic control by caveolin-1. Notably, the PDGF induced loss of antimitogenic control by caveolin-1 coincided with a marked increase in caveolin-1 phosphorylation. Furthermore, the strong association of PDGF receptor-β with caveolin-1 that exists in quiescent cells was rapidly and markedly reduced with agonist addition. This suggests a dynamic relationship in which mitogen stimulation actively reverses caveolin-1 suppression of p42/p44 MAPK signal transduction. As such, caveolae and caveolin-1 coordinate PDGF receptor signaling, leading to myocyte proliferation, and inhibit constitutive activity of p42/p44 MAPK to sustain cell quiescence.
Apoptosis of pulmonary microvascular endothelial cells stimulates vascular smooth muscle cell growth
Authors: Sakao Seiichiro , Taraseviciene-Stewart Laimute , Wood Kathy , Cool Carlyne D. , Voelkel Norbert F.
Published: April 2006

We have previously hypothesized that the development of severe angioproliferative pulmonary hypertension is associated with not only initial endothelial cell (EC) apoptosis followed by the emergence of apoptosis-resistant proliferating EC but also with proliferation of vascular smooth muscle cells (VSMC). We have demonstrated that EC death results in the selection of an apoptosis-resistant, proliferating, and phenotypically altered EC phenotype. We postulate here that the initial apoptosis of EC induces the release of mediators that cause VSMC proliferation. We cultured EC in an artificial capillary CellMax system designed to simulate the highly efficient functions of the human capillary system. We induced apoptosis of microvascular EC using shear stress and the combined VEGF receptor (VEGFR-1 and -2) inhibitor SU-5416. Flow cytometry for the proliferation marker bromodeoxyuridine showed that serum-free medium conditioned by apoptosed EC induced proliferation of VSMC, whereas serum-free medium conditioned by nonapoptosed EC did not. We also show that medium conditioned by apoptosed EC is characterized by increased concentrations of transforming growth factor (TGF)-β1 and VEGF compared with medium conditioned by nonapoptosed EC and that TGF-β1 blockade prevented the proliferation of cultured VSMC. In conclusion, EC death induced by high shear stress and VEGFR blockade leads to the production of factors, in particular TGF-β1, that activate VSMC proliferation.
GRO-α regulation in airway smooth muscle by IL-1β and TNF-α: role of NF-κB and MAP kinases
Authors: Issa Razao , Xie Shaoping , Lee Kang-Yun , Stanbridge Rex D. , Bhavsar Pankaj , Sukkar Maria B. , Chung Kian Fan
Published: April 2006

Airway smooth muscle cells (ASMC) are a source of inflammatory chemokines that may propagate airway inflammatory responses. We investigated the production of the CXC chemokine growth-related oncogene protein-α (GRO-α) from ASMC induced by cytokines and the role of MAPK and NF-κB pathways. ASMC were cultured from human airways, grown to confluence, and exposed to cytokines IL-1β and TNF-α after growth arrest. GRO-α release, measured by ELISA, was increased by >50-fold after IL-1β (0.1 ng/ml) or 5-fold after TNF-α (1 ng/ml) in a dose- and time-dependent manner. GRO-α release was not affected by the T helper type 2 cytokines IL-4, IL-10, and IL-13. IL-1β and TNF-α also induced GRO-α mRNA expression. Supernatants from IL-1β-stimulated ASMC were chemotactic for neutrophils; this effect was inhibited by anti-GRO-α blocking antibody. AS-602868, an inhibitor of IKK-2, and PD-98059, an inhibitor of ERK, inhibited GRO-α release and mRNA expression, whereas SP-600125, an inhibitor of JNK, reduced GRO-α release without effect on mRNA expression. SB-203580, an inhibitor of p38 MAPK, had no effect. AS-602868 but not PD-98059 or SP-600125 inhibited p65 DNA-binding induced by IL-1β and TNF-α. By chromatin immunoprecipitation assay, IL-1β and TNF-α enhanced p65 binding to the GRO-α promoter, which was inhibited by AS-602868. IL-1β- and TNF-α-stimulated expression of GRO-α from ASMC is regulated by independent pathways involving NF-κB activation and ERK and JNK pathways. GRO-α released from ASMC participates in neutrophil chemotaxis.
SERPINB1 upregulation is associated with in vivo complex formation with neutrophil elastase and cathepsin G in a baboon model of bronchopulmonary dysplasia
Authors: Yasumatsu Ryuji , Altiok Ozden , Benarafa Charaf , Yasumatsu Chie , Bingol-Karakoc Gulbin , Remold-O'Donnell Eileen , Cataltepe Sule
Published: April 2006

Bronchopulmonary dysplasia (BPD) continues to be a major cause of morbidity in premature infants. An imbalance between neutrophil elastase and its inhibitors has been implicated in BPD. Serine protease inhibitor (SERPIN)B1 is an inhibitor of neutrophil proteases, including neutrophil elastase (NE) and cathepsin G (cat G). Recent studies suggest that SERPINB1 could provide protection in the airways by regulating excess protease activity associated with inflammatory lung disorders. In this study, we determined the distribution and ontogeny of SERPINB1 in the baboon lung and characterized the expression of SERPINB1 in baboon models of BPD. SERPINB1 expression was detected in the conducting airway and glandular epithelial cells in addition to neutrophils, macrophages, and mast cells. SERPINB1 mRNA and protein expression increased with advancing gestational age and in the new BPD model. In contrast, SERPINB1 expression levels were decreased in the old BPD model. Furthermore, SERPINB1 was detected as a high-molecular-mass (HMM) complex in lung tissue and bronchoalveolar lavage fluid samples from the BPD group. Analysis of the HMM complex by coimmunoprecipitation showed that these complexes were formed between SERPINB1 and NE or cat G. High-performance liquid chromatography (HPLC) ion trap mass spectrometry verified the presence of SERPINB1 in HMM complexes. Finally, NE activity level was compared between new and old baboon models of BPD and was found to be significantly lower in new BPD. Thus SERPINB1 upregulation in new BPD may be protective by contributing to the regulation of neutrophil proteases NE and cat G.