bluuebunny/crossref_metadata_2025 huggingface dataset:
TRPC1 functions as a store-operated Ca2+channel in intestinal epithelial cells and regulates early mucosal restitution after wounding
Authors: Rao Jaladanki N. , Platoshyn Oleksandr , Golovina Vera A. , Liu Lan , Zou Tongtong , Marasa Bernard S. , Turner Douglas J. , Yuan Jason X.-J. , Wang Jian-Ying
Published: November 2005

An increase in cytosolic free Ca2+concentration ([Ca2+]cyt) results from Ca2+release from intracellular stores and extracellular Ca2+influx through Ca2+-permeable ion channels and is crucial for initiating intestinal epithelial restitution to reseal superficial wounds after mucosal injury. Capacitative Ca2+entry (CCE) induced by Ca2+store depletion represents a major Ca2+influx mechanism, but the exact molecular components constituting this process remain elusive. This study determined whether canonical transient receptor potential (TRPC)1 served as a candidate protein for Ca2+-permeable channels mediating CCE in intestinal epithelial cells and played an important role in early epithelial restitution. Normal intestinal epithelial cells (the IEC-6 cell line) expressed TRPC1 and TPRC5 and displayed typical records of whole cell store-operated Ca2+currents and CCE generated by Ca2+influx after depletion of intracellular stores. Induced TRPC1 expression by stable transfection with the TRPC1 gene increased CCE and enhanced cell migration during restitution. Differentiated IEC-Cdx2L1 cells induced by forced expression of the Cdx2 gene highly expressed endogenous TRPC1 and TRPC5 and exhibited increased CCE and cell migration. Inhibition of TRPC1 expression by small interfering RNA specially targeting TRPC1 not only reduced CCE but also inhibited cell migration after wounding. These findings strongly suggest that TRPC1 functions as store-operated Ca2+channels and plays a critical role in intestinal epithelial restitution by regulating CCE and intracellular [Ca2+]cyt.
Endotoxin enhances microvascular thrombosis in mouse cremaster venules via a TLR4-dependent, neutrophil-independent mechanism
Authors: Rumbaut Rolando E. , Bellera Ricardo V. , Randhawa Jaspreet K. , Shrimpton Corie N. , Dasgupta Swapan K. , Dong Jing-Fei , Burns Alan R.
Published: November 2005

Endotoxemia promotes adhesive interactions between platelets and microvascular endothelium in vivo. We sought to determine whether endotoxin (lipopolysaccharide, LPS) modified platelet thrombus formation in mouse cremaster venules and whether Toll-like receptor 4 (TLR4) and neutrophils were involved in the response. Intravital videomicroscopy was performed in the cremaster microcirculation of pentobarbital-anesthetized mice; venular platelet thrombi were induced with a light/dye endothelial injury model. C57BL/6 mice treated with Escherichia coli endotoxin had enhanced rates of venular platelet thrombus formation: the time to microvessel occlusion was reduced by ∼50% ( P < 0.005) compared with saline-treated animals. Enhanced microvascular thrombosis was evident as early as 2 h after LPS administration. LPS had no effect on thrombosis in either of two mouse strains with altered TLR4 signaling (C57BL/10ScNJ or C3H/HeJ), whereas it enhanced thrombosis in the control strains (C57BL/10J and C3H/HeN). LPS also enhanced platelet adhesion to endothelium in the absence of light/dye injury. Platelet adhesion, but not enhanced thrombosis, was inhibited by depletion of circulating neutrophils. LPS failed to enhance platelet aggregation ex vivo and did not influence platelet P-selectin expression, a marker of platelet activation. These findings support the notion that endotoxemia promotes platelet thrombus formation independent of neutrophils and without enhancement of platelet aggregation, via a TLR4-dependent mechanism.
Matrix metalloproteinase expression in vein grafts: role of inflammatory mediators and extracellular signal-regulated kinases-1 and -2
Authors: Sharony Ram , Pintucci Giuseppe , Saunders Paul C. , Grossi Eugene A. , Baumann F. Gregory , Galloway Aubrey C. , Mignatti Paolo
Published: November 2005

Matrix metalloproteinases (MMPs) play key roles in vascular remodeling. We characterized the role of inflammatory mediators and extracellular signal-regulated kinases (ERKs) in the control of arterialized vein graft expression of MMP-9, MMP-2, and membrane-type 1-MMP (MT1-MMP) and of the tissue inhibitor of metalloproteinases-2 (TIMP-2). For this purpose we used a canine model of jugular vein to carotid artery interposition graft and analyzed the vein grafts at various postoperative times (30 min to 28 days) using the contralateral vein as a control. To study the role of ERK-1/2, veins were incubated with the mitogen-activated protein kinase kinase (MEK-1/2) inhibitor UO126 for 30 min before being grafted. Vein graft extracts were analyzed for MMPs, TIMP-2, tumor necrosis factor-α (TNF-α), polymorphonuclear neutrophil (PMN) infiltration, myeloperoxidase (MPO), and thrombin activity, and for ERK-1/2 activation. Vein graft arterialization resulted in rapid and sustained (8 h to 28 days) upregulation of vein graft-associated MMP-9, MMP-2, MT1-MMP, thrombin activity, and TNF-α levels with concomitant TIMP-2 downregulation. MMP-2 activation preceded MT1-MMP upregulation. PMN infiltration and vein graft-associated MPO activity increased within hours after arterialization, indicating a prompt, local inflammatory response. In cultured smooth muscle cells, both thrombin and TNF-α upregulated MT1-MMP expression; however, only thrombin activated MMP-2. Inhibition of ERK-1/2 activation blocked arterialization-induced upregulation of MMP-2, MMP-9, and MT1-MMP. Thus, thrombin, inflammatory mediators, and activation of the ERK-1/2 pathway control MMP and TIMP-2 expression in arterialized vein grafts.
Nonlinear isochrones in murine left ventricular pressure-volume loops: how well does the time-varying elastance concept hold?
Authors: Claessens T. E. , Georgakopoulos D. , Afanasyeva M. , Vermeersch S. J. , Millar H. D. , Stergiopulos N. , Westerhof N. , Verdonck P. R. , Segers P.
Published: November 2005

tThe linear time-varying elastance theory is frequently used to describe the change in ventricular stiffness during the cardiac cycle. The concept assumes that all isochrones (i.e., curves that connect pressure-volume data occurring at the same time) are linear and have a common volume intercept. Of specific interest is the steepest isochrone, the end-systolic pressure-volume relationship (ESPVR), of which the slope serves as an index for cardiac contractile function. Pressure-volume measurements, achieved with a combined pressure-conductance catheter in the left ventricle of 13 open-chest anesthetized mice, showed a marked curvilinearity of the isochrones. We therefore analyzed the shape of the isochrones by using six regression algorithms (two linear, two quadratic, and two logarithmic, each with a fixed or time-varying intercept) and discussed the consequences for the elastance concept. Our main observations were 1) the volume intercept varies considerably with time; 2) isochrones are equally well described by using quadratic or logarithmic regression; 3) linear regression with a fixed intercept shows poor correlation ( R2 < 0.75) during isovolumic relaxation and early filling; and 4) logarithmic regression is superior in estimating the fixed volume intercept of the ESPVR. In conclusion, the linear time-varying elastance fails to provide a sufficiently robust model to account for changes in pressure and volume during the cardiac cycle in the mouse ventricle. A new framework accounting for the nonlinear shape of the isochrones needs to be developed.
Synchronous progression of calcium transient-dependent beating and sarcomere destruction in apoptotic adult cardiomyocytes
Authors: Maruyama Rumi , Takemura Genzou , Tohse Noritsugu , Ohkusa Tomoko , Ikeda Yasuhiro , Tsuchiya Kunihiko , Minatoguchi Shinya , Matsuzaki Masunori , Fujiwara Takako , Fujiwara Hisayoshi
Published: November 2005

During early apoptosis, adult cardiomyocytes show unusual beating, suggesting possible participation of abnormal Ca2+transients in initiation of apoptotic processes in this cell type. Simultaneously with the beating, these cells show dynamic structural alteration resulting from cytoskeletal disintegration that is quite rapid. Because of the specialized structure and extensive cytoskeleton of cardiomyocytes, we hypothesized that its degradation in so short a time would require a particularly efficient mechanism. To better understand this mechanism, we used serial video microscopy to observe β-adrenergic stimulation-induced apoptosis in isolated adult rat cardiomyocytes while simultaneously recording intracellular Ca2+concentration and cell length. Trains of Ca2+transients and corresponding rhythmic contractions and relaxations (beating) were observed in apoptotic cells. Frequencies of Ca2+transients and beating gradually increased with time and were accompanied by cellular shrinkage. As the cells shrank, amplitudes of Ca2+transients declined and diastolic intracellular Ca2+concentration increased until the transients were lost. Beating and progression of apoptosis were significantly inhibited by antagonists against the L-type Ca2+channel (nifedipine), ryanodine receptor (ryanodine), inositol 1,4,5-trisphosphate receptor (heparin), sarco(endo)plasmic Ca2+-ATPase (thapsigargin), and Na+/Ca2+exchanger (KB-R7943). Electron-microscopic examination of beating cardiomyocytes revealed progressive breakdown of Z disks. Immunohistochemical analysis and Western blot confirmed that disappearance of Z disk constituent proteins (α-actinin, desmin, and tropomyosin) preceded degradation of other cytoskeletal proteins. It thus appears that, in adult cardiomyocyte apoptosis, Ca2+transients mediate apoptotic beating and efficient sarcomere destruction initiated by Z disk breakdown.
Angiotensin II hypertension is attenuated in interleukin-6 knockout mice
Authors: Lee Dexter L. , Sturgis Lashon C. , Labazi Hicham , Osborne James B. , Fleming Cassandra , Pollock Jennifer S. , Manhiani Marlina , Imig John D. , Brands Michael W.
Published: November 2005

Plasma levels of IL-6 correlate with high blood pressure under many circumstances, and ANG II has been shown to stimulate IL-6 production from various cell types. This study tested the role of IL-6 in mediating the hypertension caused by high-dose ANG II and a high-salt diet. Male C57BL6 and IL-6 knockout (IL-6 KO) mice were implanted with biotelemetry devices and placed in metabolic cages to measure mean arterial pressure (MAP), heart rate (HR), sodium balance, and urinary albumin excretion. Baseline MAP during the control period averaged 114 ± 1 and 109 ± 1 mmHg for wild-type (WT) and IL-6 KO mice, respectively, and did not change significantly when the mice were placed on a high-salt diet (HS; 4% NaCl). ANG II (90 ng/min sc) caused a rapid increase in MAP in both groups, to 141 ± 9 and 141 ± 4 in WT and KO mice, respectively, on day 2. MAP plateaued at this level in KO mice (134 ± 2 mmHg on day 14 of ANG II) but began to increase further in WT mice by day 4, reaching an average of 160 ± 4 mmHg from days 10 to 14 of ANG II. Urinary albumin excretion on day 4 of ANG II was not different between groups (9.18 ± 4.34 and 8.53 ± 2.85 μg/2 days for WT and KO mice). By day 14, albumin excretion was nearly fourfold greater in WT mice, but MAP dropped rapidly back to control levels in both groups when the ANG II was stopped after 14 days. Thus the ∼30 mmHg greater ANG II hypertension in the WT mice suggests that IL-6 contributes significantly to ANG II-salt hypertension. In addition, the early separation in MAP, the albumin excretion data, and the rapid, post-ANG II recovery of MAP suggest an IL-6-dependent mechanism that is independent of renal injury.
Effect of acute sympathetic nervous system activation on flow-mediated dilation of brachial artery
Authors: Dyson Kenneth S. , Shoemaker J. Kevin , Hughson Richard L.
Published: November 2005

We tested the hypothesis that flow-mediated dilation (FMD) of the brachial artery would be impaired by acute increases in sympathetic nervous system activity (SNA) in models where similar peak shear stress stimulus was achieved by varying the duration of forearm muscle ischemia. Eleven healthy young men were studied under four different conditions, each with its own control: lower body suction (LBS), cold pressor test (CPT), mental arithmetic task (MAT), and activation of muscle chemoreflex (MCR). The duration of ischemia before observation of FMD by ultrasound imaging was 5 min each for control, LBS, and CPT; 3 min for MAT; and 2-min for MCR. Peak shear rate was not different between control and any of the SNA conditions, although total shear in the first minute was reduced in MAT. MCR was the only condition in which brachial artery vasoconstriction was observed before forearm occlusion [4.38 (SD 0.53) vs. control 4.60 (SD 0.53) mm, P < 0.05]; however, diameter increased to the same absolute value as that of the control, so the percent FMD was greater for MCR [9.85 (SD 2.33) vs. control 5.29 (SD 1.50)%]. Blunting of the FMD response occurred only in the CPT model [1.51 (SD 1.20)%]. During SNA, the increase in plasma cortisol from baseline was significant only for MCR; the increase in plasma norepinephrine was significant for MCR, LBS, and CPT; and the increase in epinephrine was significant only for MCR. These results showed that the four models employed to achieve increases in SNA had different effects on baseline brachial artery diameter and that blunted FMD is not a general response to increased SNA.
Time-dependent modulation of arterial baroreflex control of muscle sympathetic nerve activity during isometric exercise in humans
Authors: Ichinose Masashi , Saito Mitsuru , Kondo Narihiko , Nishiyasu Takeshi
Published: November 2005

We investigated the time-dependent modulation of arterial baroreflex (ABR) control of muscle sympathetic nerve activity (MSNA) that occurs during isometric handgrip exercise (IHG). Thirteen healthy subjects performed a 3-min IHG at 30% maximal voluntary contraction, which was followed by a period of imposed postexercise muscle ischemia (PEMI). The ABR control of MSNA (burst incidence and strength and total activity) was evaluated by analyzing the relationship between spontaneous variations in diastolic arterial pressure (DAP) and MSNA during supine rest, at each minute of IHG, and during PEMI. We found that 1) the linear relations between DAP and MSNA variables were shifted progressively rightward until the third minute of IHG (IHG3); 2) 2 min into IHG (IHG2), the DAP-MSNA relations were shifted upward and were shifted further upward at IHG3; 3) the sensitivity of the ABR control of total MSNA was increased at IHG2 and increased further at IHG3; and 4) during PEMI, the ABR operating pressure was slightly higher than at IHG2, and the sensitivity of the control of total MSNA was the same as at IHG2. During PEMI, the DAP-burst strength and DAP-total MSNA relations were shifted downward from the IHG3 level to the IHG2 level, whereas the DAP-burst incidence relation remained at the IHG3 level. These results indicate that during IHG, ABR control of MSNA is modulated in a time-dependent manner. We suggest that this modulation of ABR function is one of the mechanisms underlying the progressive increase in blood pressure and MSNA during the course of isometric exercise.
Stimulation of A2A-adenosine receptors after myocardial infarction suppresses inflammatory activation and attenuates contractile dysfunction in the remote left ventricle
Authors: Toufektsian Marie-Claire , Yang Zequan , Prasad Konkal M. , Overbergh Lutgart , Ramos Susan I. , Mathieu Chantal , Linden Joel , French Brent A.
Published: November 2005

Following myocardial infarction (MI), contractile dysfunction develops not only in the infarct zone but also in noninfarcted regions of the left ventricle remote from the infarct zone. Inflammatory activation secondary to MI stimulates inducible nitric oxide synthase (iNOS) induction with excess production of nitric oxide. We hypothesized that the anti-inflammatory effects of selective A2A-adenosine receptor (A2AAR) stimulation would suppress inflammation and preserve cardiac function in the remote zone early after MI. A total of 53 mice underwent 60 min of coronary occlusion followed by 24 h of reperfusion. The A2AAR agonist (ATL146e, 2.4 μg/kg) was administered intraperitoneally 1, 3, and 6 h postreperfusion. Because of the 1-h delay in treatment after MI, ATL146e had no effect on infarct size, as demonstrated by contrast-enhanced cardiac MRI ( n = 18) performed 24 h post-MI. ATL146e did however preserve global cardiac function at that time by limiting contractile dysfunction in remote regions [left ventricle wall thickening: 51 ± 4% in treated ( n = 9) vs. 29 ± 3% in nontreated groups ( n = 9), P < 0.01]. RT-PCR, immunohistochemistry, and Western blot analysis indicated that iNOS mRNA and protein expression were significantly reduced by ATL146e treatment in both infarcted and noninfarcted zones. Similarly, elevations in plasma nitrate-nitrite after MI were substantially blunted by ATL146e ( P < 0.01). Finally, treatment with ATL146e reduced NF-κB activation in the myocardium by over 50%, not only in the infarct zone but also in noninfarcted regions ( P < 0.05). In conclusion, A2AAR stimulation after MI suppresses inflammatory activation and preserves cardiac function, suggesting the potential utility of A2AAR agonists against acute heart failure in the immediate post-MI period.
Effect of angiotensin II receptor blockade on autonomic nervous system function in patients with essential hypertension
Authors: Krum Henry , Lambert Elisabeth , Windebank Emma , Campbell Duncan J. , Esler Murray
Published: November 2005

It has long been proposed that the renin-angiotensin system exerts a stimulatory influence on the sympathetic nervous system, including augmentation of central sympathetic outflow and presynaptic facilitation of norepinephrine release from sympathetic nerves. We tested this proposition in 19 patients with essential hypertension, evaluating whether the angiotensin receptor blockers (ARBs) eprosartan and losartan had identifiable antiadrenergic properties. This was done in a prospective, randomized, three-way placebo-controlled study of crossover design. Patients were randomized to 600 mg of eprosartan daily, 50 mg of losartan daily, or placebo. The treatment period was 4 wk, with 2-wk washout periods. Multiunit firing rates in efferent sympathetic nerves distributed to skeletal muscle vasculature (muscle sympathetic nerve activity, MSNA) were measured with microneurography, testing whether ARBs inhibit central sympathetic outflow. In parallel, isotope dilution methodology was used to measure whole body norepinephrine spillover to plasma. Mean blood pressure on placebo was 151/98 mmHg, with both ARBs causing reductions of ∼11 mmHg systolic and 6 mmHg diastolic pressure, placebo corrected. Both MSNA [35 ± 12 bursts/min (mean ± SD) on placebo] and whole body norepinephrine spillover [366 ± 247 ng/min] were unchanged by ARB administration, indicating that the ARBs did not materially inhibit central sympathetic outflow or act presynaptically to reduce norepinephrine release at existing rates of nerve firing. These findings contrast with the easily demonstrable reduction in sympathetic nervous activity produced by antihypertensive drugs of the imidazoline-binding class, which are known to act within the brain to inhibit sympathetic nervous outflow. We conclude that sympathetic nervous inhibition is not a major component of the blood pressure-lowering action of ARBs in essential hypertension.
Lowering of interstitial fluid pressure in rat submandibular gland: a novel mechanism in saliva secretion
Authors: Berggreen Ellen , Wiig Helge
Published: November 2005

The submandibular gland transports fluid at a high rate through the interstitial space during salivation, but the exact level of all forces governing transcapillary fluid transport has not been established. In this study, our aim was to measure the relation between interstitial fluid volume (Vi) and interstitial fluid pressure (Pif) in salivary glands during active secretion and after systemically induced passive changes in gland hydration. We tested whether interstitial fluid could be isolated by tissue centrifugation to enable measurement of interstitial fluid colloid osmotic pressure. During control conditions, Vi averaged 0.23 ml/g wet wt (SD 0.014), with a corresponding mean Pif measured with micropipettes of 3.0 mmHg (SD 1.3). After induction of secretion by pilocarpine, Pif dropped by 3.8 mmHg (SD 1.5) whereas Vi was unchanged. During dehydration and overhydration of up to 20% increase of Vi above control, a linear relation was found between volume and pressure, resulting in a compliance (ΔVi/ΔPif) of 0.012 ml·g wet wt−1·mmHg−1. Interstitial fluid was isolated, and interstitial fluid colloid osmotic pressure averaged 10.4 mmHg (SD 1.2), which is 64% of the corresponding level in plasma. We conclude that Pif drops during secretion and, thereby, increases the net transcapillary pressure gradient, a condition that favors fluid filtration and increases the amount of fluid available for secretion. The reduction in Pif is most likely induced by contraction of myoepithelial cells and suggests an active and new role for these cells in salivary secretion. The relatively low interstitial compliance of the organ will enhance the effect of the myoepithelial cells on Pif during reduced Vi.
Relationship among diastolic intraventricular pressure gradients, relaxation, and preload: impact of age and fitness
Authors: Popović Zoran B. , Prasad Anand , Garcia Mario J. , Arbab-Zadeh Armin , Borowski Allen , Dijk Erika , Greenberg Neil L. , Levine Benjamin D. , Thomas James D.
Published: November 2005

Diastolic intraventricular pressure gradients (IVPGs) are a measure of the ability of the ventricle to facilitate its filling using diastolic suction. We assessed 15 healthy young but sedentary subjects, aged <50 yr (young subjects; age, 35 ± 9 yr); 13 healthy but sedentary seniors, aged >65 yr with known reductions in ventricular compliance (elderly sedentary subjects; age, 70 ± 4 yr); and 12 master athletes, aged >65 yr, previously shown to have preserved ventricular compliance (elderly fit subjects; age, 68 ± 3 yr). Pulmonary capillary wedge pressure (PCWP) and echocardiography measurements were performed at baseline, during load manipulation by lower body negative pressure at −15 and −30 mmHg, and after saline infusion of 10 and 20 ml/kg (elderly) or 15 and 30 ml/kg (young). IVPGs were obtained from color M-mode Doppler echocardiograms. Baseline IVPGs were lower (1.2 ± 0.4 vs. 2.4 ± 0.7 mmHg, P < 0.0001), and the time constant of pressure decay (τ0) was longer (60 ± 10 vs. 46 ± 6 ms, P < 0.0001) in elderly sedentary than in young subjects, with no difference in PCWP. Although PCWP changes during load manipulations were similar ( P = 0.70), IVPG changes were less prominent in elderly sedentary than in young subjects ( P = 0.02). Changes in stroke volume and IVPGs during loading manipulations correlated ( r = 0.96, P = 0.0002). PCWP and τ0 were strong multivariate correlates of IVPGs ( P < 0.001, for both). IVPG response to loading interventions in elderly sedentary and elderly fit subjects was similar ( P = 0.33), despite known large differences in ventricular compliance. The ability to regulate IVPGs during changes in preload is impaired with aging. Preserving ventricular compliance during aging by lifelong exercise training does not prevent this impairment.
Exercise training induces respiratory substrate-specific decrease in Ca2+-induced permeability transition pore opening in heart mitochondria
Authors: Marcil Mariannick , Bourduas Karine , Ascah Alexis , Burelle Yan
Published: November 2005

The purpose of this study was to determine whether regular exercise (treadmill running, 10 wk) alters the susceptibility of rat isolated heart mitochondria to Ca2+-induced permeability transition pore (PTP) opening and whether this could be associated with changes in the modulation of PTP opening by selected physiological effectors. Basal leak-driven and ADP-stimulated respiration in the presence of substrates for complex I, II, and IV were not affected by training. Fluorimetric studies revealed that in the control and exercise-trained groups, the amount of Ca2+required to trigger PTP opening was greater in the presence of complex II vs. I substrates (230 ± 12 vs. 134 ± 7 nmol Ca2+/mg protein, P < 0.01; pooled average of control and trained groups). In addition, with a substrate feeding the complex II, training increased by 45% ( P < 0.01) the amount of Ca2+required to trigger PTP opening both in the presence and absence of the PTP inhibitor cyclosporin A. However, membrane potential, reactive oxygen species production, NAD(P)H ratio, and Ca2+uptake kinetics were not different in mitochondria from both groups. Together, these results suggest the existence of a substrate-specific regulation of the PTP in heart mitochondria and suggest that regular exercise results in a reduced sensitivity to Ca2+-induced PTP opening in presence of complex II substrates.
Connexin40 and connexin43 in mouse aortic endothelium: evidence for coordinated regulation
Authors: Isakson Brant E. , Damon David N. , Day Kathleen H. , Liao Yongbo , Duling Brian R.
Published: November 2005

In the vessel wall, endothelial cells are metabolically and electrically coupled to each other and to the adjacent smooth muscle cells by gap junctions composed of connexins. Gap junctions may be formed from combinations of several different connexin proteins, and deletion of one connexin can lead to modification of the expression of another. To reveal a possible interaction between connexin40 (Cx40) and connexin43 (Cx43) in endothelium, we studied their distribution in vessels from C57Bl/6 and Cx40 knockout mice (Cx40−/−) using immunoblots and immunocytochemistry on aortic cross sections and en face whole mounts. En face preparations from C57Bl/6 mice revealed two distinct pools of Cx43, one pericellular and the other intracellular. Cx40 was largely restricted to the periphery of the cells, and in Cx40−/− mice it was, as expected, undetectable. In the Cx40−/− mice, total Cx43 protein was also modestly reduced (immunoblots), but there was a major redistribution of the protein within the cell. The pericellular component of Cx43 was rendered virtually undetectable, and the intracellular compartments were normal or even slightly elevated. Smooth muscle Cx43 was also reduced in the Cx40−/− animals. These findings indicate that the cellular distribution of Cx43 is dependent on the presence of Cx40, and in view of the profound effects on the pericellular pool of the Cx43, the findings suggest that interactions between Cx40 and Cx43 regulate communication between endothelial cells and perhaps between smooth muscle and endothelial cells as well.
Arterial compliance of rowers: implications for combined aerobic and strength training on arterial elasticity
Authors: Cook Jill N. , DeVan Allison E. , Schleifer Jessica L. , Anton Maria M. , Cortez-Cooper Miriam Y. , Tanaka Hirofumi
Published: November 2005

Regular endurance exercise increases central arterial compliance, whereas resistance training decreases it. It is not known how the vasculature adapts to a combination of endurance and resistance training. Rowing is unique, because its training encompasses endurance- and strength-training components. We used a cross-sectional study design to determine arterial compliance of 15 healthy, habitual rowers [50 ± 9 (SD) yr, 11 men and 4 women] and 15 sedentary controls (52 ± 8 yr, 10 men and 5 women). Rowers had been training 5.4 ± 1.2 days/wk for 5.7 ± 4.0 yr. The two groups were matched for age, body composition, blood pressure, and metabolic risk factors. Central arterial compliance (simultaneous ultrasound and applanation tonometry on the common carotid artery) was higher ( P < 0.001) and carotid β-stiffness index was lower ( P < 0.001) in rowers than in sedentary controls. There were no group differences for measures of peripheral (femoral) arterial stiffness. The higher central arterial compliance in rowers was associated with a greater cardiovagal baroreflex sensitivity, as estimated during a Valsalva maneuver ( r = 0.54, P < 0.005). In conclusion, regular rowing exercise in middle-aged and older adults is associated with a favorable effect on the elastic properties of the central arteries. Our results suggest that simultaneously performed endurance training may negate the stiffening effects of strength training.
Oxygen tension modulates the expression of pulmonary vascular BKCachannel α- and β-subunits
Authors: Resnik Ernesto , Herron Jean , Fu Rao , Ivy D. Dunbar , Cornfield David N.
Published: November 2005

At birth, the lung environment changes from low to relatively high O2tension. Pulmonary blood flow increases and pulmonary artery pressure decreases. Recent data suggest that pulmonary vascular calcium-sensitive K+channel (BKCa) activation mediates perinatal pulmonary vasodilation. Although BKCachannel expression is developmentally regulated, the molecular mechanisms responsible for BKCaexpression remain unknown. We tested the hypothesis that the low-O2tension environment of the normal fetus modulates BKCachannel expression. We analyzed BKCaexpression under conditions of hypoxia and normoxia both in vitro and in vivo. BKCaα-subunit mRNA expression increased twofold in ovine pulmonary artery smooth muscle cell (PASMC) primary cultures maintained in hypoxia. In vivo, BKCaexpression was similarly affected by hypoxia. When adult Sprague-Dawley rats were placed in hypobaric hypoxic chambers for 3 wk, hypoxic animals showed an increase of threefold in the expression of BKCaα- and more than twofold in the expression of BKCaβ1-subunit mRNA. Immunochemical staining was consistent with the genetic data. To assess transcriptional activation of the β-subunit of the BKCa, both BKCaβ1- and β2-subunit luciferase (KCaβ:luc+) reporter genes were constructed. Hypoxia increased PASMC KCaβ1:luc+reporter expression by threefold and KCaβ2:luc+expression by 35%. Fetal PASMC treated with the hypoxia-inducible factor-1 mimetic deferoxamine showed a 63 and 41% increase in BKCachannel α- and β1-subunit expression, respectively. Together, these results suggest that oxygen tension modulates BKCachannel subunit mRNA expression, and the regulation is, at least in part, at the transcriptional level.
Protective effect of endogenous β-adrenergic tone on lung fluid balance in acute bacterial pneumonia in mice
Authors: Su Xiao , Robriquet Laurent , Folkesson Hans G. , Matthay Michael A.
Published: November 2005

Some investigators have reported that endogenous β-adrenoceptor tone can provide protection against acute lung injury. Therefore, we tested the effects of β-adrenoceptor inhibition in mice with acute Escherichia coli pneumonia. Mice were pretreated with propranolol or saline and then intratracheally instilled with live E. coli (107 colony-forming units). Hemodynamics, arterial blood gases, plasma catecholamines, extravascular lung water, lung permeability to protein, bacterial counts, and alveolar fluid clearance were measured. Acute E. coli pneumonia was established after 4 h with histological evidence of acute pulmonary inflammation, arterial hypoxemia, a threefold increase in lung vascular permeability, and a 30% increase in extravascular lung water as an increase in plasma catecholamine levels. β-Adrenoceptor inhibition resulted in a marked increase in extravascular lung water that was explained by both an increase in lung vascular permeability and a reduction in net alveolar fluid clearance. The increase in extravascular lung water with propranolol pretreatment was not explained by an increase in systemic or vascular pressures. The increase in lung vascular permeability was explained in part by anti-inflammatory effects of β-adrenoceptor stimulation because plasma macrophage inflammatory protein-2 levels were higher in the propranolol pretreatment group compared with controls. The decrease in alveolar fluid clearance with propranolol was explained by a decrease in catecholamine-stimulated fluid clearance. Together, these results indicate that endogenous β-adrenoceptor tone has a protective effect in limiting accumulation of extravascular lung water in acute severe E. coli pneumonia in mice by two mechanisms: 1) reducing lung vascular injury and 2) upregulating the resolution of alveolar edema.
Zinc modulates airway epithelium susceptibility to death receptor-mediated apoptosis
Authors: Bao Shenying , Knoell Daren L.
Published: November 2005

The accelerated loss of lung epithelium through activation of extrinsic apoptosis is believed to play a causative role in lung pathogenesis. Previous investigations have shown that zinc is required to sustain lung epithelial cell viability under stress conditions and that depletion of intracellular zinc predisposes cells to apoptosis. In this investigation, we determined whether intracellular zinc deficiency enhanced the susceptibility of primary, differentiated cultures of human lung epithelium to death receptor-mediated apoptosis, leading to barrier dysfunction. Cultures obtained from multiple donors were exposed to stimuli that provoke death receptor-mediated apoptosis and depleted of intracellular zinc with a zinc-specific chelating agent. Transepithelial resistance, paracellular transport, caspase-8 and caspase-3 activity, and apoptosis were measured. Activation of extrinsic apoptosis or zinc chelation alone resulted in a nominal increase in caspase function and apoptosis without major evidence of barrier disruption. Activation of extrinsic apoptosis in addition to zinc depletion resulted in an abrupt decrease in transepithelial resistance, a substantial increase in apoptosis, and an increased paracellular leak. Cultures were rescued by supplementation with zinc sulfate. Further analysis revealed that exogenous zinc facilitates cell survival through activation of the phosphatidylinositol 3-kinase/Akt signaling pathway. We conclude that intracellular zinc is a vital factor in lung epithelium that protects cells from death receptor-mediated apoptosis and barrier dysfunction.
Peroxynitrite mediates TNF-α-induced endothelial barrier dysfunction and nitration of actin
Authors: Neumann Paul , Gertzberg Nancy , Vaughan Erin , Weisbrot Joshua , Woodburn Renee , Lambert William , Johnson Arnold
Published: November 2005

We tested the hypothesis that tumor necrosis factor (TNF)-α induces a peroxynitrite (ONOO−)-dependent increase in permeability of pulmonary microvessel endothelial monolayers (PMEM) that is associated with generation of nitrated β-actin (NO2-β-actin). The permeability of PMEM was assessed by the clearance rate of Evans blue-labeled albumin. β-Actin was extracted from PMEM lysate with a DNase-Sepharose column. The extracted β-actin was quantified in terms of its nitrotyrosine/β-actin ratio with antinitrotyrosine and anti-β-actin antibodies, sequentially, by dot-blot assays. The cellular compartmentalization of NO2-β-actin was displayed by showing confocal localization of nitrotyrosine-immunofluorescence with β-actin-immunofluorescence but not with F-actin fluorescence. Incubation of PMEM with TNF (100 ng/ml) for 0.5 and 4.0 h resulted in increases in permeability to albumin. There was an increase in the nitrotyrosine/β-actin ratio at 0.5 h with minimal association of the NO2-β-actin with F-actin polymers. The TNF-induced increase in the nitrotyrosine/β-actin ratio and permeability were prevented by the anti-ONOO− agent Urate. The data indicate that TNF induces an ONOO−-dependent barrier dysfunction, which is associated with the generation of NO2-β-actin.
Inhibition of phosphodiesterase 1 augments the pulmonary vasodilator response to inhaled nitric oxide in awake lambs with acute pulmonary hypertension
Authors: Evgenov Oleg V. , Busch Cornelius J. , Evgenov Natalia V. , Liu Rong , Petersen Bodil , Falkowski George E. , Pethő Beata , Vas Ádám , Bloch Kenneth D. , Zapol Warren M. , Ichinose Fumito
Published: November 2005

Phosphodiesterase 1 (PDE1) modulates vascular tone and the development of tolerance to nitric oxide (NO)-releasing drugs in the systemic circulation. Any role of PDE1 in the pulmonary circulation remains largely uncertain. We measured the expression of genes encoding PDE1 isozymes in the pulmonary vasculature and examined whether or not selective inhibition of PDE1 by vinpocetine attenuates pulmonary hypertension and augments the pulmonary vasodilator response to inhaled NO in lambs. Using RT-PCR, we detected PDE1A, PDE1B, and PDE1C mRNAs in pulmonary arteries and veins isolated from healthy lambs. In 13 lambs, the thromboxane A2analog U-46619 was infused intravenously to increase mean pulmonary arterial pressure to 35 mmHg. Four animals received an intravenous infusion of vinpocetine at incremental doses of 0.3, 1, and 3 mg·kg−1·h−1. In nine lambs, inhaled NO was administered in a random order at 2, 5, 10, and 20 ppm before and after an intravenous infusion of 1 mg·kg−1·h−1vinpocetine. Administration of vinpocetine did not alter pulmonary and systemic hemodynamics or transpulmonary cGMP or cAMP release. Inhaled NO selectively reduced mean pulmonary arterial pressure, pulmonary capillary pressure, and pulmonary vascular resistance index, while increasing transpulmonary cGMP release. The addition of vinpocetine enhanced pulmonary vasodilation and transpulmonary cGMP release induced by NO breathing without causing systemic vasodilation but did not prolong the duration of pulmonary vasodilation after NO inhalation was discontinued. Our findings demonstrate that selective inhibition of PDE1 augments the therapeutic efficacy of inhaled NO in an ovine model of acute chemically induced pulmonary hypertension.
Dopamine regulation of amiloride-sensitive sodium channels in lung cells
Authors: Helms My N. , Chen Xi-Juan , Ramosevac Semra , Eaton Douglas C. , Jain Lucky
Published: November 2005

Dopamine increases lung fluid clearance. This is partly due to activation of basolateral Na-K-ATPase. However, activation of Na-K-ATPase by itself is unlikely to produce large changes in transepithelial transport. Therefore, we examined apical and basolateral dopamine's effect on apical, highly selective sodium channels [epithelial sodium channels (ENaC)] in monolayers of an alveolar type 2 cell line (L2). Dopamine increased channel open probability ( Po) without changing the unitary current. The D1receptor blocker SCH-23390 blocked the dopamine effect, but the D2receptor blocker sulpiride did not. The dopamine-mediated increase in ENaC activity was not a secondary effect of dopamine stimulation of Na-K-ATPase, since ouabain applied to the basolateral surface to block the activity of Na-K-ATPase did not alter dopamine-mediated ENaC activity. Protein kinase A (PKA) was not responsible for dopamine's effect since a PKA inhibitor, H89, did not reduce dopamine's effect. However, cpt-2-O-Me-cAMP, which selectively binds and activates EPAC (exchange protein activated by cAMP) but not PKA, increased ENaC Po. An Src inhibitor, PP2, and the phosphatidylinositol-3-kinase inhibitor, LY-294002, blocked dopamine's effect on ENaC. In addition, an MEK blocker, U0126, an inhibitor of phospholipase A2, and a protein phosphatase inhibitor also blocked the effect of dopamine on ENaC Po. Finally, since the cAMP-EPAC-Rap1 pathway also activates DARPP32 (32-kDa dopamine response protein phosphatase), we confirmed that dopamine phosphorylates DARPP32, and okadaic acid, which blocks phosphatases (DARPP32), also blocks dopamine's effect. In summary, dopamine increases ENaC activity by a cAMP-mediated alternative signaling pathway involving EPAC and Rap1, signaling molecules usually associated with growth-factor-activated receptors.
Defending body mass during food restriction inAcomys russatus: a desert rodent that does not store food
Authors: Gutman Roee , Choshniak Itzhak , Kronfeld-Schor Noga
Published: November 2005

Golden spiny mice, which inhabit rocky deserts and do not store food, must therefore employ physiological means to cope with periods of food shortage. Here we studied the physiological means used by golden spiny mice for conserving energy during food restriction and refeeding and the mechanism by which food consumption may influence thermoregulatory mechanisms and metabolic rate. As comparison, we studied the response to food restriction of another rocky desert rodent, Wagner’s gerbil, which accumulates large seed caches. Ten out of 12 food-restricted spiny mice (resistant) were able to defend their body mass after an initial decrease, as opposed to Wagner’s gerbils ( n = 6). Two of the spiny mice (nonresistant) kept losing weight, and their food restriction was halted. In four resistant and two nonresistant spiny mice, we measured heart rate, body temperature, and oxygen consumption during food restriction. The resistant spiny mice significantly ( P < 0.05) reduced energy expenditure and entered daily torpor. The nonresistant spiny mice did not reduce their energy expenditure. The gerbils’ response to food restriction was similar to that of the nonresistant spiny mice. Resistant spiny mice leptin levels dropped significantly ( n = 6, P < 0.05) after 24 h of food restriction, and continued to decrease throughout food restriction, as did body fat. During refeeding, although the golden spiny mice gained fat, leptin levels were not correlated with body mass ( r2= 0.014). It is possible that this low correlation allows them to continue eating and accumulate fat when food is plentiful.
Blunted hypertensive effect of combined fructose and high-salt diet in gene-targeted mice lacking functional serum- and glucocorticoid-inducible kinase SGK1
Authors: Huang Dan Yang , Boini Krishna M. , Friedrich Björn , Metzger Marco , Just Lothar , Osswald Hartmut , Wulff Peer , Kuhl Dietmar , Vallon Volker , Lang Florian
Published: November 2005

Serum- and glucocorticoid-inducible kinase (SGK1) is transcriptionally upregulated by mineralocorticoids and activated by insulin. The kinase stimulates the renal epithelial Na+channel and may thus participate in blood pressure regulation. Hyperinsulinemia is triggered by dietary fructose, which sensitizes blood pressure for salt intake. The role of SGK1 in hypertensive effects of combined fructose and high-salt intake was thus explored in SGK1 knockout mice ( sgk1−/−) and their wild-type littermates ( sgk1+/+). Renal SGK1 transcript levels of sgk1+/+mice were significantly elevated after fructose diet. Under control diet, fluid intake, urinary flow rate, urinary Na+, K+, and Cl−excretion, and blood pressure were similar in sgk1−/−and sgk1+/+mice. Addition of 10% fructose to drinking water increased fluid intake and urinary flow rate in both genotypes, and did not significantly alter urinary Na+, K+, and Cl−output in either genotype. Additional high NaCl diet (4% NaCl) did not significantly alter fluid intake and urine volume but markedly increased urinary output of Na+and Cl−, approaching values significantly ( P < 0.05) larger in sgk1−/−than in sgk1+/+mice (Na+: 2,572 ± 462 vs. 1,428 ± 236; Cl−: 2,364 ± 388 vs. 1,379 ± 225 μmol/24 h). Blood pressure was similar in sgk1+/+and sgk1−/−mice at control diet or fructose alone but increased only in sgk1+/+mice (115 ± 1 vs. 103 ± 0.7 mmHg, P < 0.05) after combined fructose and high-salt intake. Acute intravenous insulin infusion (during glucose clamp) caused antinatriuresis in sgk1+/+mice, an effect significantly blunted in sgk1−/−mice. The observations reveal a pivotal role of SGK1 in insulin-mediated sodium retention and the salt-sensitizing hypertensive effect of high fructose intake.
Ischemic acute renal failure induces the expression of a wide range of nephrogenic proteins
Authors: Villanueva Sandra , Céspedes Carlos , Vio Carlos P.
Published: November 2005

Ischemia-induced acute renal failure (ARF) is a disorder with high morbidity and mortality. ARF is characterized by a regeneration phase, yet its molecular basis is still under study. Changes in gene expression have been reported in ARF, and some of these genes are specific for nephrogenic processes. We tested the hypothesis that the regeneration process developed after ischemia-induced ARF can be characterized by the reexpression of important regulatory proteins of kidney development. The distribution pattern and levels of nephrogenic proteins in rat kidneys after ischemia were studied by immunohistochemistry and immunoblot analysis. Ischemic damage was assessed by conventional morphology, serum creatinine, and the apoptotic markers terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) and caspase 3. The hypoxia levels induced by ischemia were assessed by specific markers: hypoxia induced factor (HIF)-1α and 2-pimonidazole. In kidneys with ARF, an important initial damage was observed through periodic acid Schiff staining, by the induction of damage markers α-smooth muscle actin (α-SMA) and macrophages (ED-1) and by apoptosis induction. In agreement with diminishing renal damage at the initial reparation phase, the expression of the mesenchymal proteins vimentin, neural cell adhesion molecules (Ncam), and the epithelial markers, Pax-2, Noggin, and basic fibroblast growth factor was observed; after, in a second phase, the tubular markers bone morphogen protein 7, Engrailed, and Lim-1, as well as the transcription factors Smad and p-Smad, were observed. Additionally, the endothelial markers VEGF and Tie-2 were induced at the initial and middle stages of regeneration phase, respectively. The expression of these proteins was restricted in time and space, as well as spatially and temporally. Because all of these proteins are important in maintaining a functional kidney, these results suggest that during the regeneration process after induced hypoxia, these nephrogenic proteins can be reexpressed in a similar fashion to that observed during development, thus restoring mature kidney function.
Opiate slowing of feline respiratory rhythm and effects on putative medullary phase-regulating neurons
Authors: Lalley Peter M.
Published: November 2005

Opiates have effects on respiratory neurons that depress tidal volume and air exchange, reduce chest wall compliance, and slow rhythm. The most dose-sensitive opioid effect is slowing of the respiratory rhythm through mechanisms that have not been thoroughly investigated. An in vivo dose-response analysis was performed on medullary respiratory neurons of adult cats to investigate two untested hypotheses related to mechanisms of opioid-mediated rhythm slowing: 1) Opiates suppress intrinsic conductances that limit discharge duration in medullary inspiratory and expiratory neurons, and 2) opiates delay the onset and lengthen the duration of discharges postsynaptically in phase-regulating postinspiratory and late-inspiratory neurons. In anesthetized and unanesthetized decerebrate cats, a threshold dose (3 μg/kg) of the μ-opioid receptor agonist fentanyl slowed respiratory rhythm by prolonging discharges of inspiratory and expiratory bulbospinal neurons. Additional doses (2–4 μg/kg) of fentanyl also lengthened the interburst silent periods in each type of neuron and delayed the rate of membrane depolarization to firing threshold without altering synaptic drive potential amplitude, input resistance, peak action potential frequency, action potential shape, or afterhyperpolarization. Fentanyl also prolonged discharges of postinspiratory and late-inspiratory neurons in doses that slowed the rhythm of inspiratory and expiratory neurons without altering peak membrane depolarization and hyperpolarization, input resistance, or action potential properties. The temporal changes evoked in the tested neurons can explain the slowing of network respiratory rhythm, but the lack of significant, direct opioid-mediated membrane effects suggests that actions emanating from other types of upstream bulbar respiratory neurons account for rhythm slowing.