bluuebunny/crossref_metadata_2025 huggingface dataset:
Vacuolar H+-ATPase in human breast cancer cells with distinct metastatic potential: distribution and functional activity
Authors: Sennoune Souad R. , Bakunts Karina , Martínez Gloria M. , Chua-Tuan Jenny L. , Kebir Yamina , Attaya Mohamed N. , Martínez-Zaguilán Raul
Published: February 2004

Tumor cells thrive in a hypoxic microenvironment with an acidic extracellular pH. To survive in this harsh environment, tumor cells must exhibit a dynamic cytosolic pH regulatory system. We hypothesize that vacuolar H+-ATPases (V-ATPases) that normally reside in acidic organelles are also located at the cell surface, thus regulating cytosolic pH and exacerbating the migratory ability of metastatic cells. Immunocytochemical data revealed for the first time that V-ATPase is located at the plasma membrane of human breast cancer cells: prominent in the highly metastatic and inconspicuous in the lowly metastatic cells. The V-ATPase activities in isolated plasma membranes were greater in highly than in lowly metastatic cells. The proton fluxes via V-ATPase evaluated by fluorescence spectroscopy in living cells were greater in highly than in lowly metastatic cells. Interestingly, lowly metastatic cells preferentially used the ubiquitous Na+/H+ exchanger and HCO3−-based H+-transporting mechanisms, whereas highly metastatic cells used plasma membrane V-ATPases. The highly metastatic cells were more invasive and migratory than the lowly metastatic cells. V-ATPase inhibitors decreased the invasion and migration in the highly metastatic cells. Altogether, these data indicate that V-ATPases located at the plasma membrane are involved in the acquisition of a more metastatic phenotype.
Urea transport in MDCK cells that are stably transfected with UT-A1
Authors: Fröhlich Otto , Klein Janet D. , Smith Pauline M. , Sands Jeff M. , Gunn Robert B.
Published: February 2004

Progress in understanding the cell biology of urea transporter proteins has been hampered by the lack of an appropriate cell culture system. The goal of this study was to create a polarized epithelial cell line that stably expresses the largest of the rat renal urea transporter UT-A isoforms, UT-A1. The gene for UT-A1 was cloned into pcDNA5/FRT and transfected into Madin-Darby canine kidney (MDCK) cells with an integrated Flp recombination target site. The cells from a single clone were grown to confluence on collagen-coated membranes until the resistance was >1,500 Ω·cm2. Transepithelial [14C]urea fluxes were measured at 37°C in a HCO3−/CO2 buffer, pH 7.4, with 5 mM urea. The baseline fluxes were not different between unstimulated UT-A1-transfected MDCK cells and nontransfected or sham-transfected MDCK cells. However, only in the UT-A1-transfected cells was UT-A1 protein expressed (as measured by Western blot analysis) and urea transport stimulated by forskolin or arginine vasopressin. Forskolin and arginine vasopressin also increased the phosphorylation of UT-A1. Thionicotinamide, dimethylurea, and phloretin inhibited the forskolin-stimulated [14C]urea fluxes in the UT-A1-transfected MDCK cells. These characteristics mimic those seen in rat terminal inner medullary collecting ducts. This new polarized epithelial cell line stably expresses UT-A1 and reproduces several of the physiological responses observed in rat terminal inner medullary collecting ducts.
Modulation of vascular smooth muscle cell migration by calcium/ calmodulin-dependent protein kinase II-δ2
Authors: Pfleiderer Paul J. , Lu Katherine Kun , Crow Michael T. , Keller Rebecca S. , Singer Harold A.
Published: February 2004

Previous studies demonstrated a requirement for multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMKII) in PDGF-stimulated vascular smooth muscle (VSM) cell migration. In the present study, molecular approaches were used specifically to assess the role of the predominant CaMKII isoform (δ2 or δC) on VSM cell migration. Kinase-negative (K43A) and constitutively active (T287D) mutant forms of CaMKIIδ2 were expressed using recombinant adenoviruses. CaMKII activities were evaluated in vitro by using a peptide substrate and in intact cells by assessing the phosphorylation of overexpressed phospholamban on Thr17, a CaMKII-selective phosphorylation site. Expression of kinase-negative CaMKIIδ2 inhibited substrate phosphorylation both in vitro and in the intact cell, indicating a dominant-negative function with respect to exogenous substrate. However, overexpression of the kinase-negative mutant failed to inhibit endogenous CaMKIIδ2 autophosphorylation on Thr287 after activation of cells with ionomycin, and in fact, these subunits served as a substrate for the endogenous kinase. Constitutively active CaMKIIδ2 phosphorylated substrate in vitro without added Ca2+/calmodulin and in the intact cell without added Ca2+-dependent stimuli, but it inhibited autophosphorylation of endogenous CaMKIIδ2 on Thr287. Basal and PDGF-stimulated cell migration was significantly enhanced in cells expressing kinase-negative CaMKIIδ2, an effect opposite that of KN-93, a chemical inhibitor of CaMKII activation. Expression of the constitutively active CaMKIIδ2 mutant inhibited PDGF-stimulated cell migration. These studies point to a role for the CaMKIIδ2 isoform in regulating VSM cell migration. An inclusive interpretation of results using both pharmacological and molecular approaches raises the hypothesis that CaMKIIδ2 autophosphorylation may play an important role in PDGF-stimulated VSM cell migration.
Extracellular pressure stimulates macrophage phagocytosis by inhibiting a pathway involving FAK and ERK
Authors: Shiratsuchi Hiroe , Basson Marc D.
Published: February 2004

We hypothesized that changes in extracellular pressure during inflammation or infection regulate macrophage phagocytosis through modulating the focal adhesion kinase (FAK)-ERK pathway. Undifferentiated (monocyte-like) or PMA-differentiated (macrophage-like) THP-1 cells were incubated at 37°C with serum-opsonized latex beads under ambient or 20-mmHg increased pressure. Pressure did not affect monocyte phagocytosis but significantly increased macrophage phagocytosis (29.9 ± 1.8 vs. 42.0 ± 1.6%, n = 9, P < 0.001). THP-1 macrophages constitutively expressed activated FAK, ERK, and Src. Exposure of macrophages to pressure decreased ERK and FAK-Y397 phosphorylation (77.6 ± 7.9%, n = 7, P < 0.05) but did not alter FAK-Y576 or Src phosphorylation. FAK small interfering RNA (SiRNA) reduced FAK expression by >75% and the basal amount of phosphorylated FAK by 25% and significantly increased basal macrophage phagocytosis ( P < 0.05). Pressure inhibited FAK-Y397 phosphorylation in mock-transfected or scrambled SiRNA-transfected macrophages, but phosphorylated FAK was not significantly reduced further by pressure in cells transfected with FAK SiRNA. Pressure increased phagocytosis in all three groups. However, FAK-SiRNA-transfected cells exhibited only 40% of the pressure effect on phagocytosis observed in scrambled SiRNA-transfected cells so that phagocytosis inversely paralleled FAK activation. PD-98059 (50 μM), an ERK activation inhibitor, increased basal phagocytosis (26.9 ± 1.8 vs. 31.7 ± 1.1%, n = 15, P < 0.05), but pressure did not further increase phagocytosis in PD-98059-treated cells. Pressure also inhibited ERK activation after mock transfection or transfection with scrambled SiRNA, but transfection of FAK SiRNA abolished ERK inhibition by pressure. Pressure did not increase phagocytosis in MonoMac-1 cells that do not express FAK. Increased extracellular pressure during infection or inflammation enhances macrophage phagocytosis by inhibiting FAK and, consequently, decreasing ERK activation.
Regulation of prostate-specific antigen by activin A in prostate cancer LNCaP cells
Authors: Fujii Yasuhisa , Kawakami Satoru , Okada Yohei , Kageyama Yukio , Kihara Kazunori
Published: February 2004

Activins are multifunctional growth and differentiation factors and stimulate FSH-β gene expression and FSH secretion by the pituitary gonadotropes. Follistatins bind activin, resulting in the neutralization of activin bioactivity. The activin/follistatin system is present in the prostate tissue. Prostate-specific antigen (PSA) plays an important role in male reproductive physiology as well as being very important as a tumor marker for prostate cancer. Thus the regulation of PSA has important clinical implications. Previous studies showed that PSA is primarily regulated by androgens. In the present study, we evaluated the direct effects of activin A on the proliferation and PSA production of prostate cancer LNCaP cells, which express functional activin receptors and androgen receptor and PSA. LNCaP cells were treated with activin A and 5α-dihydrotestosterone (DHT) with or without their antagonists (follistatin or the nonsteroidal anti-androgen bicalutamide). Activin A decreased cell growth of LNCaP cells in a dose-dependent manner, whereas DHT increased it in a biphasic manner. In contrast to their opposing actions on cell growth, both activin A and DHT upregulated PSA gene expression and increased PSA secretion by LNCaP cells. The effects of activin A and DHT to increase PSA production were synergistic or additive. Follistatin or bicalutamide was without effect on cell growth or PSA production. The effects of activin A on LNCaP cells were blocked by follistatin, not by bicalutamide, whereas effects of DHT were prevented by bicalutamide, not by follistatin. Activin A upregulates PSA production, and the effect is through an androgen receptor-independent pathway. The activin/follistatin system can be a physiological modulator of PSA gene transcription and secretion in the prostate tissue, and activins may cooperate with androgen to upregulate PSA in vivo.
Mucosal angiogenesis regulation by CXCR4 and its ligand CXCL12 expressed by human intestinal microvascular endothelial cells
Authors: Heidemann Jan , Ogawa Hitoshi , Rafiee Parvaneh , Lügering Norbert , Maaser Christian , Domschke Wolfram , Binion David G. , Dwinell Michael B.
Published: February 2004

Mice genetically deficient in the chemokine receptor CXCR4 or its ligand stromal cell-derived factor (SDF)-1/CXCL12 die perinatally with marked defects in vascularization of the gastrointestinal tract. The aim of this study was to define the expression and angiogenic functions of microvascular CXCR4 and SDF-1/CXCL12 in the human intestinal tract. Studies of human colonic mucosa in vivo and primary cultures of human intestinal microvascular endothelial cells (HIMEC) in vitro showed that the intestinal microvasculature expresses CXCR4 and its cognate ligand SDF-1/CXCL12. Moreover, SDF-1/CXCL12 stimulation of HIMEC triggers CXCR4-linked G proteins, phosphorylates ERK1/2, and activates proliferative and chemotactic responses. Pharmacological studies indicate SDF-1/CXCL12 evokes HIMEC chemotaxis via activation of ERK1/2 and phosphoinositide 3-kinase signaling pathways. Consistent with chemotaxis and proliferation, endothelial tube formation was inhibited by neutralizing CXCR4 or SDF-1/CXCL12 antibodies, as well as the ERK1/2 inhibitor PD-98059. Taken together, these data demonstrate an important mechanistic role for CXCR4 and SDF-1/CXCL12 in regulating angiogenesis within the human intestinal mucosa.
Mitogen-activated protein kinases regulate COX-2 and mucosal recovery in ischemic-injured porcine ileum
Authors: Shifflett Donnie E. , Jones Samuel L. , Moeser Adam J. , Blikslager Anthony T.
Published: February 2004

Mitogen-activated protein kinase (MAPK) pathways transduce signals from a diverse array of extracellular stimuli. The three primary MAPK-signaling pathways are the extracellular regulated kinases (ERK1/2), p38 MAPK, and c-Jun NH2-terminal kinase (JNK). Previous research in our laboratory has shown that COX-2-elaborated prostanoids participate in recovery of mucosal barrier function in ischemic-injured porcine ileum. Because COX-2 expression is regulated in part by MAPKs, we postulated that MAPK pathways would play an integral role in recovery of injured mucosa. Porcine mucosa was subjected to 45 min of ischemia, after which tissues were mounted in Ussing chambers, and transepithelial electrical resistance (TER) was monitored as an index of recovery of barrier function. Treatment of tissues with the p38 MAPK inhibitor SB-203580 (0.1 mM) or the ERK1/2 inhibitor PD-98059 (0.1 mM) abolished recovery. Western blot analysis revealed that SB-203580 inhibited upregulation of COX-2 that was observed in untreated ischemic-injured mucosa, whereas PD-98059 had no effect on COX-2 expression. Inhibition of TER recovery by SB-203580 or PD-98059 was overcome by administration of exogenous prostaglandin E2(1 μM). The JNK inhibitor SP-600125 (0.1 mM) significantly increased TER and resulted in COX-2 upregulation. COX-2 expression appears to be positively and negatively regulated by the p38 MAPK and the JNK pathways, respectively. Alternatively, ERK1/2 appear to be involved in COX-2-independent reparative events that remain to be defined.
Bacterial activation of β-catenin signaling in human epithelia
Authors: Sun Jun , Hobert Michael E. , Rao Anjali S. , Neish Andrew S. , Madara James L.
Published: February 2004

The mucosal lining of the human intestine is constantly bathed in a milieu of commensal gut flora, the vast majority of these being nonpathogenic microorganisms. Here, we demonstrate that microbial-epithelial cell interactions not only affect proinflammatory pathways but also influence β-catenin signaling, a key component in regulating epithelial cell proliferation. The nonpathogenic Salmonella strain PhoPcactivates the β-catenin signaling pathway of human epithelia via a blockade of β-catenin degradation. Normal β-catenin ubiquitination necessary for constitutive β-catenin degradation is abolished, allowing the accumulation and translocation of β-catenin to the nucleus. Transcriptional activation mediated by the β-catenin/T cell factor complex increases c-myc expression and enhances cell proliferation. We also show that the Salmonella effector protein AvrA is involved in modulating this β-catenin activation. These data suggest that nonvirulent bacterial-epithelial interactions can influence β-catenin signaling and cell growth control in a manner previously unsuspected.
Gestational and early postnatal dietary NaCl levels affect NaCl intake, but not stimulated water intake, by adult rats
Authors: Curtis Kathleen S. , Krause Eric G. , Wong Donna L. , Contreras Robert J.
Published: February 2004

We examined body fluid regulation by weanling (21–25 days) and adult (>60 days) male rats that were offspring of dams fed chow containing either 0.1, 1, or 3% NaCl throughout gestation and lactation. Weanling rats were maintained on the test diets until postnatal day 30 and on standard 1% NaCl chow thereafter. Ad libitum water intake by weanlings was highest in those fed 3% NaCl and lowest in those fed 0.1% NaCl. Adult rats maintained on standard NaCl chow consumed similar amounts of water after overnight water deprivation or intravenous hypertonic NaCl (HS) infusion regardless of early NaCl condition. Moreover, baseline and HS-stimulated plasma Na+ concentrations also were similar for the three groups. Nonetheless, adult rats in the early 3% NaCl group consumed more of 0.5 M NaCl after 10 days of dietary Na+ deprivation than did rats in either the 1% or 0.1% NaCl group. Interestingly, whether NaCl was consumed in a concentrated solution in short-term, two-bottle tests after dietary Na+ deprivation or in chow during ad libitum feeding, adult rats in the 3% NaCl group drank less water for each unit of NaCl consumed, whereas rats in the 0.1% NaCl group drank more water for each unit of NaCl consumed. Thus gestational and early postnatal dietary NaCl levels do not affect stimulated water intake or long-term body fluid regulation. Together with our previous studies, these results suggest that persistent changes in NaCl intake and in water intake associated with NaCl ingestion reflect short-term behavioral effects that may be attributable to differences in NaCl taste processing.
Localized compliance of small airways in excised rat lungs using microfocal X-ray computed tomography
Authors: Sera Toshihiro , Fujioka Hideki , Yokota Hideo , Makinouchi Akitake , Himeno Ryutaro , Schroter Robert C. , Tanishita Kazuo
Published: February 2004

Airway compliance is a key factor in understanding lung mechanics and is used as a clinical diagnostic index. Understanding such mechanics in small airways physiologically and clinically is critical. We have determined the “morphometric change” and “localized compliance” of small airways under “near”-physiological conditions; namely, the airways were embedded in parenchyma without dehydration and fixation. Previously, we developed a two-step method to visualize small airways in detail by staining the lung tissue with a radiopaque solution and then visualizing the tissue with a cone-beam microfocal X-ray computed tomography system (Sera et al. J Biomech 36: 1587-1594, 2003). In this study, we used this technique to analyze changes in diameter and length of the same small airways (∼150 μm ID) and then evaluated the localized compliance as a function of airway generation ( Z). For smaller (<300-μm-diameter) airways, diameter was 36% larger at end-tidal inspiration and 89% larger at total lung capacity; length was 18% larger at end-tidal inspiration and 43% larger at total lung capacity than at functional residual capacity. Diameter, especially at smaller airways, did not behave linearly with V1/3 (where V is volume). With increasing lung pressure, diameter changed dramatically at a particular pressure and length changed approximately linearly during inflation and deflation. Percentage of airway volume for smaller airways did not behave linearly with that of lung volume. Smaller airways were generally more compliant than larger airways with increasing Z and exhibited hysteresis in their diameter behavior. Airways at higher Z deformed at a lower pressure than those at lower Z. These results indicated that smaller airways did not behave homogeneously.
Is the blood flow response to a single contraction determined by work performed?
Authors: Hamann Jason J. , Buckwalter John B. , Clifford Philip S. , Shoemaker J. Kevin
Published: February 2004

Nine healthy volunteers performed a series of single handgrip isometric contractions to test the hypothesis that the blood flow response to a contraction is determined solely by the tension-time index (isometric analog of work). Contractions were performed in duplicate at 15, 30, and 60% of maximal voluntary contraction (MVC) at durations of 0.5, 1, and 2 s. Forearm blood flow (FBF) was measured beat by beat by using Doppler ultrasound. Peak FBF responded in a graded fashion to graded increases in peak tension with contraction time held constant (35, 56, and 90 ml/min for 15, 30, and 60% MVC for 1 s, respectively). When tension was kept constant, peak FBF responded in a graded fashion to graded increases in duration (77, 90, and 97 ml/min for 60% MVC for 0.5, 1, and 2 s). With a constant tension-time index, peak FBF responded in a graded fashion to graded increases in peak tension (48, 56, and 77 ml/min for 15% MVC/2 s, 30% MVC/1 s, and 60% MVC/0.5 s). Similar trends were also observed for total postcontraction hyperemia. Blood flow increased regardless of whether the change in tension-time index was accomplished by an increase in tension or duration of contraction. However, with a constant tension-time index, the change in blood flow was related to the peak tension developed. Our results suggest that the blood flow response to a single muscle contraction is not determined solely by the work performed (tension-time index) but also by the number of muscle fibers recruited.
Mechanical compression-induced pressure sores in rat hindlimb: muscle stiffness, histology, and computational models
Authors: Linder-Ganz E. , Gefen A.
Published: February 2004

Pressure sores affecting muscles are severe injuries associated with ischemia, impaired metabolic activity, excessive tissue deformation, and insufficient lymph drainage caused by prolonged and intensive mechanical loads. We hypothesize that mechanical properties of muscle tissue change as a result of exposure to prolonged and intensive loads. Such changes may affect the distribution of stresses in soft tissues under bony prominences and potentially expose additional uninjured regions of muscle tissue to intensified stresses. In this study, we characterized changes in tangent elastic moduli and strain energy densities of rat gracilis muscles exposed to pressure in vivo (11.5, 35, or 70 kPa for 2, 4, or 6 h) and incorporated the abnormal properties that were measured in finite element models of the head, shoulders, pelvis, and heels of a recumbent patient. Using in vitro uniaxial tension testing, we found that tangent elastic moduli of muscles exposed to 35 and 70 kPa were 1.6-fold those of controls ( P < 0.05, for strains ≤5%) and strain energy densities were 1.4-fold those of controls ( P < 0.05, for strains ≥5%). Histological (phosphotungstic acid hematoxylin) evaluation showed that this stiffening accompanied extensive necrotic damage. Incorporating these effects into the finite element models, we were able to show that the increased muscle stiffness in widening regions results in elevated tissue stresses that exacerbate the potential for tissue necrosis. Interfacial pressures could not predict deep muscle (e.g., longissimus or gluteus) stresses and injuring conditions. We conclude that information on internal muscle stresses is required to establish new criteria for pressure sore prevention.
Ventilatory dynamics and control of blood gases after maximal exercise in the Thoroughbred horse
Authors: Padilla Danielle J. , McDonough Paul , Kindig Casey A. , Erickson Howard H. , Poole David C.
Published: February 2004

Despite enormous rates of minute ventilation (V̇e) in the galloping Thoroughbred (TB) horse, the energetic demands of exercise conspire to raise arterial Pco2(i.e., induce hypercapnia). If locomotory-respiratory coupling (LRC) is an obligatory facilitator of high V̇e in the horse such as those found during galloping (Bramble and Carrier. Science 219: 251–256, 1983), V̇e should drop precipitously when LRC ceases at the galloptrot transition, thus exacerbating the hypercapnia. TB horses ( n = 5) were run to volitional fatigue on a motor-driven treadmill (1 m/s increments; 14–15 m/s) to study the dynamic control of breath-by-breath V̇e, O2uptake, and CO2output at the transition from maximal exercise to active recovery (i.e., trotting at 3 m/s for 800 m). At the transition from the gallop to the trot, V̇e did not drop instantaneously. Rather, V̇e remained at the peak exercising levels (1,391 ± 88 l/min) for ∼13 s via the combination of an increased tidal volume (12.6 ± 1.2 liters at gallop; 13.9 ± 1.6 liters over 13 s of trotting recovery; P < 0.05) and a reduced breathing frequency [113.8 ± 5.2 breaths/min (at gallop); 97.7 ± 5.9 breaths/min over 13 s of trotting recovery ( P < 0.05)]. Subsequently, V̇e declined in a biphasic fashion with a slower mean response time (85.4 ± 9.0 s) than that of the monoexponential decline of CO2output (39.9 ± 4.7 s; P < 0.05), which rapidly reversed the postexercise arterial hypercapnia (arterial Pco2at gallop: 52.8 ± 3.2 Torr; at 2 min of recovery: 25.0 ± 1.4 Torr; P < 0.05). We conclude that LRC is not a prerequisite for achievement of V̇e commensurate with maximal exercise or the pronounced hyperventilation during recovery.
Transfer function analysis of heart rate variability in response to water intake: correlation with gastric myoelectrical activity
Authors: Chen C. L. , Lin H. H. , Orr William C. , Yang Cheryl C. H. , Kuo Terry B. J.
Published: February 2004

We utilized transfer function analysis of heart rate variability (HRV) and respiration to investigate the effect of water intake on gastric myoelectrical activity and its relationship to vagal activity. The electrogastrography (EGG) and HRV were recorded simultaneously before and after drinking 500 ml of water in 10 healthy subjects. We observed good linearity between lung volumes and HRV signals at a ventilatory rate between 0.2 and 0.4 Hz before and after water intake. The EGG power of 3 cycles/min increased remarkably after the water intake. We found that there was a significant increase in the magnitude of the respiration-HRV transfer function after water intake ( P < 0.05). The EGG 3 cycles/min power was positively correlated with the transfer magnitude throughout the study ( r = 0.54, P = 0.01). These results confirm that transfer function analysis of HRV sensitively identifies subtle changes in the respiratory sinus arrhythmia that occurs with water intake. The present findings suggest that transfer function analysis of HRV and respiration after water intake can be used to evaluate vagal nervous activity in the human gut.
Control of cutaneous vascular conductance and sweating during recovery from dynamic exercise in humans
Authors: Journeay W. Shane , Reardon Francis D. , Martin C. Ryan , Kenny Glen P.
Published: February 2004

The purpose of the study was to examine the effect of 1) passive (assisted pedaling), 2) active (loadless pedaling), and 3) inactive (motionless) recovery modes on mean arterial pressure (MAP), skin blood flow (SkBF), and sweating during recovery after 15 min of dynamic exercise. It was hypothesized that an active recovery mode would be most effective in attenuating the fall in MAP, SkBF, and sweating during exercise recovery. Six male subjects performed 15 min of cycle ergometer exercise at 70% of their predetermined peak oxygen consumption followed by 15 min of 1) active, 2) passive, or 3) inactive recovery. Mean skin temperature (T̄sk), esophageal temperature (Tes), SkBF, sweating, cardiac output (CO), stroke volume (SV), heart rate (HR), total peripheral resistance (TPR), and MAP were recorded at baseline, end exercise, and 2, 5, 8, 12, and 15 min postexercise. Cutaneous vascular conductance (CVC) was calculated as the ratio of laser-Doppler blood flow to MAP. In the active and passive recovery modes, CVC, sweat rate, MAP, CO, and SV remained elevated over inactive values ( P < 0.05). The passive mode was equally as effective as the active mode in maintaining CO, SV, MAP, CVC, and sweat rate above inactive recovery. Sweat rate was different among all modes after 8 min of recovery ( P < 0.05). TPR during active recovery remained significantly lower than during recovery in the passive and inactive modes ( P < 0.05). No differences in either Tesor T̄skwere observed among conditions. Given that MAP was higher during passive and active recovery modes than during inactive recovery suggests differences in CVC may be due to differences in baroreceptor unloading and not factors attributed to central command. However, differences in sweat rate may be influenced by factors such as central command and mechanoreceptor stimulation.
Metabolic effects of induced alkalosis during progressive forearm exercise to fatigue
Authors: Raymer Graydon H. , Marsh Greg D. , Kowalchuk John M. , Thompson R. Terry
Published: February 2004

Metabolic alkalosis induced by sodium bicarbonate (NaHCO3) ingestion has been shown to enhance performance during brief high-intensity exercise. The mechanisms associated with this increase in performance may include increased muscle phosphocreatine (PCr) breakdown, muscle glycogen utilization, and plasma lactate (Lac-pl) accumulation. Together, these changes would imply a shift toward a greater contribution of anaerobic energy production, but this statement has been subject to debate. In the present study, subjects ( n = 6) performed a progressive wrist flexion exercise to volitional fatigue (0.5 Hz, 14–21 min) in a control condition (Con) and after an oral dose of NaHCO3 (Alk: 0.3 g/kg; 1.5 h before testing) to evaluate muscle metabolism over a complete range of exercise intensities. Phosphorus-31 magnetic resonance spectroscopy was used to continuously monitor intracellular pH, [PCr], [Pi], and [ATP] (brackets denote concentration). Blood samples drawn from a deep arm vein were analyzed with a blood gas-electrolyte analyzer to measure plasma pH, Pco2, and [Lac-]pl, and plasma [Formula: see text] was calculated from pH and Pco2. NaHCO3 ingestion resulted in an increased ( P < 0.05) plasma pH and [Formula: see text] throughout rest and exercise. Time to fatigue and peak power output were increased ( P < 0.05) by ∼12% in Alk. During exercise, a delayed ( P < 0.05) onset of intracellular acidosis (1.17 ± 0.26 vs. 1.28 ± 0.22 W, Con vs. Alk) and a delayed ( P < 0.05) onset of rapid increases in the [Pi]-to-[PCr] ratio (1.21 ± 0.30 vs. 1.30 ± 0.30 W) were observed in Alk. No differences in total [H+], [Pi], or [Lac-]pl accumulation were detected. In conclusion, NaHCO3 ingestion was shown to increase plasma pH at rest, which resulted in a delayed onset of intracellular acidification during incremental exercise. Conversely, NaHCO3 was not associated with increased [Lac-]pl accumulation or PCr breakdown.
Stent design properties and deployment ratio influence indexes of wall shear stress: a three-dimensional computational fluid dynamics investigation within a normal artery
Authors: LaDisa John F. , Olson Lars E. , Guler Ismail , Hettrick Douglas A. , Audi Said H. , Kersten Judy R. , Warltier David C. , Pagel Paul S.
Published: February 2004

Restenosis limits the effectiveness of stents, but the mechanisms responsible for this phenomenon remain incompletely described. Stent geometry and expansion during deployment produce alterations in vascular anatomy that may adversely affect wall shear stress (WSS) and correlate with neointimal hyperplasia. These considerations have been neglected in previous computational fluid dynamics models of stent hemodynamics. Thus we tested the hypothesis that deployment diameter and stent strut properties (e.g., number, width, and thickness) influence indexes of WSS predicted with three-dimensional computational fluid dynamics. Simulations were based on canine coronary artery diameter measurements. Stent-to-artery ratios of 1.1 or 1.2:1 were modeled, and computational vessels containing four or eight struts of two widths (0.197 or 0.329 mm) and two thicknesses (0.096 or 0.056 mm) subjected to an inlet velocity of 0.105 m/s were examined. WSS and spatial WSS gradients were calculated and expressed as a percentage of the stent and vessel area. Reducing strut thickness caused regions subjected to low WSS (<5 dyn/cm2) to decrease by ∼87%. Increasing the number of struts produced a 2.75-fold increase in exposure to low WSS. Reducing strut width also caused a modest increase in the area of the vessel experiencing low WSS. Use of a 1.2:1 deployment ratio increased exposure to low WSS by 12-fold compared with stents implanted in a 1.1:1 stent-to-vessel ratio. Thinner struts caused a modest reduction in the area of the vessel subjected to elevated WSS gradients, but values were similar for the other simulations. The results suggest that stent designs that reduce strut number and thickness are less likely to subject the vessel to distributions of WSS associated with neointimal hyperplasia.
Monkey Hand Postural Synergies During Reach-to-Grasp in the Absence of Vision of the Hand and Object
Authors: Mason Carolyn R. , Theverapperuma Lalin S. , Hendrix Claudia M. , Ebner Timothy J.
Published: February 2004

Understanding how the CNS controls reach-to-grasp will require behavioral and neurophysiological studies of reach-to-grasp in the monkey, including the evaluation of whole-hand grasp with explicit force requirements. In this study, monkeys performed a reach-to-grasp task in which the size, shape, and orientation of the objects were varied. The monkeys were required to grasp each object at five force levels based on visual feedback. Seventeen positions on the wrist and hand were monitored to quantify kinematics. Hand shaping began with initiation of reach and continued throughout the reach, matching object properties even without vision of the hand or object. Grasp aperture scaled to object size. Singular value decomposition analysis of the marker positions identified two dominant hand postures. The first eigenvector or “eigenposture” consisted of an open hand configuration midway between flexion and extension that explained >93% of the variance. The second eigenposture consisted of hyperextension of all joints that accounted for another 4–5% of the variance. The two eigenpostures were similar across force levels and between monkeys. Reach kinematics consisted of a U-shaped hand path with a bell-shaped velocity profile. Trajectory and speed were independent of grasp force and object properties. In summary, hand shaping during the reach occurred without vision of the hand or object, and hand kinematics were not dependent on grasp force. Furthermore, the reach was independent of grasp force and object properties. These observations imply that the kinematics of reach-to-grasp and grasp force are controlled independently. Similar to humans, monkeys may use a simplifying strategy to reduce the degrees of freedom of the hand during reach-to-grasp.
Mechanisms for the Modulation of Native Glycine Receptor Channels by Ethanol
Authors: Eggers Erika D. , Berger Albert J.
Published: February 2004

Previously, we showed that ethanol increases synaptic glycine currents, an effect that depends on ethanol concentration and developmental age of the preparation. Glycine receptor (GlyR) subunits undergo a shift from α2/β to α1/β from neonate to juvenile ages, with synaptic glycine currents from neonate hypoglossal motoneurons (HMs) being less sensitive to ethanol than those from juvenile HMs. Here we investigate whether these dose and developmental effects are also present in excised membrane patches containing GlyRs and if ethanol changes response kinetics. We excised outside-out patches from rat HM somata and applied glycine using either a picospritzer or piezo stack translator. Ethanol (100 mM) increased the response to glycine (200 μM) of patches from neonate and juvenile HMs. However, 30 mM ethanol increased the response from only juvenile HM patches. Using a lower concentration of glycine (30 μM) to observe single channel openings, we found that 100 mM ethanol increased the number of GlyRs that open in response to glycine and decreased first latency to channel opening. To investigate GlyR kinetic properties, we rapidly applied 1 mM glycine for 1 ms and found that glycine currents were increased by ethanol (100 mM) at both ages. For patches from juvenile HMs, ethanol consistently decreased response rise-time and increased response decay time. Using kinetic modeling, we determined that ethanol's potentiation of the glycine response arises from an increase in the glycine association ( kon) and a decrease in the dissociation ( koff) rate constants, resulting in increased glycine affinity of the GlyR.
A comprehensive nonredundant expressed sequence tag collection for the developing Rattus norvegicus heart
Authors: Laffin Jennifer J.S. , Scheetz Todd E. , de Fatima Bonaldo Maria , Reiter Rebecca S. , Chang Shereen , Eyestone Mari , Abdulkawy Hakeem , Brown Bartley , Roberts Chad , Tack Dylan , Kucaba Tamara , Lin Jim Jung-Ching , Sheffield Val C. , Casavant Thomas L. , Soares M. Bento
Published: February 2004

Congenital heart defects affect ∼1,000,000 people in the United States, with 40,000 new births contributing to that number every year. A large percentage of these defects can be attributed to septal defects. We assembled a nonredundant collection of over 12,000 expressed sequence tags (ESTs) from a total of 30,000 ESTs, with the ultimate goal of identifying spatially and/or temporally regulated genes during heart septation. These ESTs were compiled from nonnormalized, normalized, and serially subtracted cDNA libraries derived from two sets of tissue samples. The first includes microdissected rat hearts from embryonic (E) days E13, E15, and E16.5–E18.5 and adult heart. The second includes hearts from embryonic days E17, E19, and E21 and postnatal (P) days P1, P12, P74, and P200. Over 6,000 novel ESTs were identified in the libraries derived from these two sets of tissues, all of which have been contributed to the NCBI rat UniGene collection. It is anticipated that such EST and cDNA clone resources will prove invaluable to gene expression studies aimed at the understanding of the molecular mechanisms underlying heart septation defects.
Brain Cholesterol: Long Secret Life Behind a Barrier
Authors: Björkhem Ingemar , Meaney Steve
Published: February 2004

Although an immense knowledge has accumulated concerning regulation of cholesterol homeostasis in the body, this does not include the brain, where details are just emerging. Approximately 25% of the total amount of the cholesterol present in humans is localized to this organ, most of it present in myelin. Almost all brain cholesterol is a product of local synthesis, with the blood-brain barrier efficiently protecting it from exchange with lipoprotein cholesterol in the circulation. Thus, there is a highly efficient apolipoprotein-dependent recycling of cholesterol in the brain, with minimal losses to the circulation. Under steady-state conditions, most of the de novo synthesis of cholesterol in the brain appears to be balanced by excretion of the cytochrome P-450-generated oxysterol 24S-hydroxycholesterol. This oxysterol is capable of escaping the recycling mechanism and traversing the blood-brain barrier. Cholesterol levels and cholesterol turnover are affected in neurodegenerating disorders, and the capacity for cholesterol transport and recycling in the brain seems to be of importance for the development of such diseases. The possibility has been discussed that administration of inhibitors of cholesterol synthesis may reduce the prevalence of Alzheimer disease. No firm conclusions can, however, be drawn from the studies presented thus far. In the present review, the most recent advances in our understanding of cholesterol turnover in the brain is discussed.
Embryonic Cell Lines with Endothelial Potential: An In Vitro System for Studying Endothelial Differentiation
Authors: Yin Yijun , Que Jianwen , Teh Ming , Cao Wei Ping , El Oakley Reida Menshawe , Lim Sai-Kiang
Published: February 2004

Objective—Endothelial differentiation is a fundamental process in angiogenesis and vasculogenesis with implications in development, normal physiology, and pathology. To better understand this process, an in vitro cellular system that recapitulates endothelial differentiation and is amenable to experimental manipulations is required.

Methods and Results—Embryonic cell lines that differentiate exclusively into endothelial cells were derived from early mouse embryos using empirical but reproducible culture techniques without viral or chemical transformation. The cells were not pluripotent and expressed reduced levels ofOct 4andRex-1. They were non-tumorigenic with a population doubling time of ≈15 hours. When plated on matrigel, they readily differentiated to form patent tubular structures with diameters of 30 to 150 μm. The differentiated cells endocytosed acetylated low-density lipoprotein (LDL) and began to express endothelial-specific markers such as CD34, CD31, Flk-1, TIE2, P-selectin, Sca-1, and thy-1. They also expressed genes essential for differentiation and maintenance of endothelial lineages, eg, Flk-1, vascular endothelial growth factor (VEGF), and angiopoietin-1. When transplanted into animal models, these cells incorporated into host vasculature.

Conclusions—These cell lines can undergo in vitro and in vivo endothelial differentiation that recapitulated known endothelial differentiation pathways. Therefore, they are ideal for establishing an in vitro cellular system to study endothelial differentiation.
Plant Sterol-Fortified Orange Juice Effectively Lowers Cholesterol Levels in Mildly Hypercholesterolemic Healthy Individuals
Authors: Devaraj Sridevi , Jialal Ishwarlal , Vega-López Sonia
Published: February 2004

Objective—

Hypercholesterolemia is a major risk factor for coronary artery disease. Therapeutic lifestyle changes include dietary modifications such as inclusion of phytosterols, which effectively lowers low-density lipoprotein (LDL) cholesterol in margarines and other fats. Their effectiveness in nonfat moieties is not yet established. The aim of this study was to examine if phytosterols alter the plasma lipoprotein profile when incorporated into nonfat orange juice.

Methods and Results—

After a 2-week run-in phase with orange juice, 72 mildly hypercholesterolemic healthy subjects were randomized to receive either placebo orange juice (placebo OJ) or plant sterol-fortified orange juice (sterol OJ) (2g/d) for 8 weeks. Fasting blood was obtained at baseline, after 2 weeks of OJ, and after 8 weeks of placebo/sterol-OJ supplementation. Sterol OJ supplementation significantly decreased total (7.2%), LDL (12.4%), and non-high-density lipoprotein (HDL) cholesterol (7.8%) compared with baseline and compared with placebo OJ (
P
<0.01). Apolipoprotein B levels were significantly decreased (9.5%) with sterol OJ. There were no significant changes in HDL cholesterol or triglycerides with the sterol OJ. While folate and B12 levels significantly increased, homocysteine levels were unchanged.

Conclusions—

Orange juice fortified with plant sterols are effective in reducing LDL cholesterol and could easily be incorporated into the therapeutic lifestyle changes dietary regimen.
Improvement of Left Ventricular Function After Cardiac Resynchronization Therapy Is Predicted by Tissue Doppler Imaging Echocardiography
Authors: Penicka Martin , Bartunek Jozef , De Bruyne Bernard , Vanderheyden Marc , Goethals Marc , De Zutter Marc , Brugada Pedro , Geelen Peter
Published: February 2004

Background—

Cardiac resynchronization therapy was shown to reverse left ventricular (LV) remodeling in patients with congestive heart failure (CHF). However, the prediction of benefit is controversial. We aimed to investigate predictive factors of LV functional recovery and reversed remodeling after biventricular pacing.

Methods and Results—

Forty-nine consecutive patients with CHF and a wide QRS complex (182±32 ms) were studied by echocardiography before resynchronization. Intraventricular and interventricular asynchrony and their combination were assessed by pulsed-wave tissue Doppler imaging from measurements of regional electromechanical coupling times in basal segments of the right and left ventricle. At 6-month follow-up, responders were defined by a relative increase in LV ejection fraction ≥25% compared with baseline (n=27). Receiver operating curve analysis revealed the degree of intraventricular asynchrony (area under the curve=0.77), interventricular asynchrony (area under the curve=0.69), and their combination (area under the curve=0.84) as the best predictors of functional recovery after resynchronization. In addition, the degree of intraventricular and interventricular asynchrony correlated significantly with the improvement of LV ejection fraction (
r
=0.73,
P
<0.0001), end-diastolic diameter (
r
=−0.59,
P
<0.0001), and end-systolic diameter (
r
=−0.48,
P
<0.001) at follow-up. QRS duration and conventional echo-Doppler indices were not predictive of reversed LV remodeling.

Conclusions—

In patients with CHF, the degree of intraventricular and interventricular asynchrony and their combination are the best predictive factors of LV functional recovery and reversed remodeling after cardiac resynchronization therapy.
Ezetimibe Effectively Reduces Plasma Plant Sterols in Patients With Sitosterolemia
Authors: Salen G. , von Bergmann K. , Lütjohann D. , Kwiterovich P. , Kane J. , Patel S.B. , Musliner T. , Stein P. , Musser B. ,   the Multicenter Sitosterolemia Study Group
Published: February 2004

Background—

Sitosterolemia is a recessively inherited disorder that results from mutations in either ABCG5 or G8 proteins, with hyperabsorption of dietary sterols and decreased hepatic excretion of plant sterols and cholesterol. As a consequence of markedly elevated plasma and tissue sitosterol and campesterol levels, premature atherosclerosis develops.

Methods and Results—

In this multicenter, double-blind, randomized, placebo-controlled study, we examined whether treatment with ezetimibe, an inhibitor of cholesterol absorption, reduces plant sterol levels in patients with sitosterolemia. After a 3-week placebo run-in, 37 patients were randomized to receive placebo (n=7) or ezetimibe 10 mg/d (n=30) for 8 weeks. Sitosterol concentrations decreased by 21% (
P
<0.001) in patients treated with ezetimibe compared with a nonsignificant 4% rise in those on placebo (between-group
P
<0.001). The reduction in sitosterol from baseline was progressive, with further decline observed at each subsequent biweekly visit. Campesterol also progressively declined, with a mean decrease after 8 weeks of 24% with ezetimibe and a mean increase of 3% with placebo treatment (between-group
P
<0.001). Reductions in plant sterol concentrations were similar irrespective of whether patients were undergoing concomitant treatment with resin or statin. Reductions in total sterols and apolipoprotein B were also observed. Ezetimibe was well tolerated, with no serious treatment-related adverse events or discontinuations due to adverse events being reported.

Conclusions—

Ezetimibe produced significant and progressive reductions in plasma plant sterol concentrations in patients with sitosterolemia, consistent with the hypothesis that ezetimibe inhibits the intestinal absorption of plant sterols as well as cholesterol, leading to reductions in plasma concentrations.
Endogenous Adrenomedullin Protects Against Vascular Response to Injury in Mice
Authors: Kawai Junsuke , Ando Katsuyuki , Tojo Akihiro , Shimosawa Tatsuo , Takahashi Katsutoshi , Onozato Maristela Lika , Yamasaki Masao , Ogita Teruhiko , Nakaoka Takashi , Fujita Toshiro
Published: February 2004

Background—

In our previous study, adrenomedullin (AM) overexpression could limit the arterial intimal hyperplasia induced by cuff injury in rats. However, it remains to be elucidated whether endogenous AM plays a role against vascular injury.

Methods and Results—

We used the AM knockout mice to investigate the effect of endogenous AM. Compared with wild-type (AM
+/+
) mice, heterozygous AM knockout (AM
+/−
) mice had the increased intimal thickening of the cuff-injured femoral artery, concomitantly with lesser AM staining. In AM
+/−
mice, cuff placement increased both the production of superoxide anions (O
2
−
) measured by coelentarazine chemiluminescence and the immunostaining of p67
phox
and gp91
phox
, subunits of NAD(P)H oxidase in the adventitia, associated with the increment of CD45-positive leukocytes, suggesting that the stimulated formation of radical oxygen species accompanied chronic adventitial inflammation. Not only the AM gene transfection but also the treatment of NAD(P)H oxidase inhibitor apocynin and membrane-permeable superoxide dismutase mimetic tempol could limit cuff-induced intimal hyperplasia in AM
+/−
mice, associated with the inhibition of O
2
−
formation in cuff-injured artery.

Conclusions—

The overproduction of oxidative stress induced by the increased NAD(P)H oxidase activity might be involved in cuff-injured arterial intimal hyperplasia in AM
+/−
mice. Thus, it is suggested that endogenous AM possesses a protective action against the vascular response to injury, possibly through the inhibition of oxidative stress production.