bluuebunny/crossref_metadata_2025 huggingface dataset:
Endothelial cytoskeletal reorganization in response to PAR1 stimulation is mediated by membrane rafts but not caveolae
Authors: Carlile-Klusacek MaryEllen , Rizzo Victor
Published: March 2007

The vasoactive protease thrombin is a known activator of the protease-activated receptor-1 (PAR1) via cleavage of its NH2terminus. PAR1 activation stimulates the RhoA/Rho kinase signaling cascade, leading to myosin light chain (MLC) phosphorylation, actin stress fiber formation, and changes in endothelial monolayer integrity. Previous studies suggest that some elements of this signaling pathway are localized to caveolin-containing cholesterol-rich membrane domains. Here we show that PAR1 and key components of the PAR-associated signaling cascade localize to membrane rafts and caveolae in bovine aortic endothelial cells (BAEC). To investigate the functional significance of this localization, BAEC were pretreated with filipin (5 μg/ml, 5 min) to ablate lipid rafts before thrombin (100 nM) or PAR agonist stimulation. We found that diphosphorylation of MLC and the actin stress fiber formation normally induced by PAR activation were attenuated after lipid raft disruption. To target caveolae specifically, we used a small interferring RNA approach to knockdown caveolin-1 expression. Thrombin-induced MLC phosphorylation and stress fiber formation were not altered in caveolin-1-depleted cells, suggesting that lipid rafts, but not necessarily caveolae, modulate thrombin-activated signaling pathways leading to alteration of the actin cytoskeleton in endothelial cells.
Modulation of the spontaneous beat-to-beat fluctuations in peripheral vascular resistance during activation of muscle metaboreflex
Authors: Ichinose Masashi , Koga Shunsaku , Fujii Naoto , Kondo Narihiko , Nishiyasu Takeshi
Published: March 2007

Continuous measurement of leg blood flow (LBF) using Doppler ultrasound with simultaneous noninvasive mean arterial blood pressure (MAP) measurement permits beat-to-beat estimates of leg vascular resistance (LVR) in humans. We tested the hypothesis that the beat-to-beat fluctuations in LVR and the dynamic relationship between MAP and LVR are modulated by the activation of muscle metaboreflex. Twelve healthy subjects performed a 1-min isometric handgrip exercise at 50% maximal voluntary contraction, which was followed by a period of imposed postexercise muscle ischemia (PEMI). We then employed transfer function analysis to examine the dynamic relationships between MAP and LBF and between MAP and LVR, both at rest (control) and during PEMI. We found the following. 1) The spectral power for LBF and LVR in low-frequency (∼0.03–0.15 Hz) range significantly increased from control during PEMI without a significant change in the high-frequency (∼0.15–0.35 Hz) power. 2) During PEMI, the transfer function gains for MAP-LBF and MAP-LVR relationships in the low-frequency (∼0.05–0.15 Hz) range were significantly increased during PEMI (vs. control) but were unchanged in the high-frequency (∼0.2–0.3 Hz) range. 3) The phases for MAP-LBF and MAP-LVR relationships were not different during control and PEMI. The phase for MAP-LVR relationship revealed that changes in MAP were followed by directionally similar changes in LVR, which is consistent with the characteristics of intrinsic vascular regulatory mechanisms such as the myogenic response of the resistance arteries. We suggest that, in humans, modulation of the dynamic MAP-LVR relationship during activation of the muscle metaboreflex reflects complex interactions between intrinsic vascular regulatory mechanisms and sympathetic vascular regulation.
Cardiac sympathetic nerve activity and ventricular fibrillation during acute myocardial infarction in a conscious sheep model
Authors: Jardine D. L. , Charles C. J. , Frampton C. M. , Richards A. M.
Published: March 2007

The association between cardiac sympathetic nerve activity (CSNA) and ventricular fibrillation (VF) during acute myocardial infarction (MI) has not been assessed in conscious animal models. During the first 60 min post-MI, mean blood pressure (MBP), heart rate (HR), and CSNA were recorded continuously in 20 conscious sheep. Resistant sheep ( group A, n = 10) were compared with susceptible sheep ( group B, n = 10) who developed fatal VF ( n = 7) or sustained ventricular tachycardia (VT, n = 3). The mean time to VF/VT was 28.1 ± 3.3 min. In group B, MBP, HR, and CSNA were averaged at each consecutive minute from baseline at 14 min before the onset of VF/VT and compared with time-matched values in group A. When compared with those of group A, indexes of CSNA burst size increased before the onset of VF/VT: burst area/minute (F13,208 = 2.17, P = 0.01) and burst area/100 beats (F13,208 = 1.86, P = 0.04). By contrast, burst frequency indexes were not significantly different: burst frequency (F13,208 = 1.6, P = 0.09) and burst incidence (F13,208 = 1.48, P = 0.13). In group A, CSNA burst area/min and burst area/100 beats did not change across this time period (F13,117 = 0.97, P = 0.5, F13,117 = 0.96, P = 0.7) but increased with time in group B (F13,91 = 2.3, P = 0.01; and F13,91 = 2.25, P = 0.01). Between-group comparisons demonstrated no differences in time of onset of ventricular ectopic beats: 18.5 (range 12–24) in group A versus 15.0 min (range 7–22) in group B (Mann-Whitney U-test, P = 0.09). Pre-MI baroreflex slopes were similar: R-R slopes were 11.8 ± 2 and 15.6 ± 1.1 ms/mmHg ( t18 = −1.6, P = 0.14). CSNA slopes were −1.8 ± 0.3 and −2.3 ± 0.2%/mmHg ( t18 = −1.4, P = 0.2). An early increase in CSNA burst size indexes (before 60 min post-MI), mediated by an excitatory sympathetic reflex, is important in the genesis of VF/VT.
A novel signaling pathway for β-adrenergic receptor-mediated activation of phosphoinositide 3-kinase in H9c2 cardiomyocytes
Authors: Yano Naohiro , Ianus Vlad , Zhao Ting C. , Tseng Andy , Padbury James F. , Tseng Yi-Tang
Published: March 2007

Stimulation of cardiac β-adrenergic receptors (β-AR) activates both the Gs- and Gi-coupled signaling cascades, including the phosphoinositide 3 kinase (PI3K) pathway, that have important physiological implications. Multiple isoforms of PI3K exist in the heart. The goals of this study were to examine the intracellular signaling pathways linking β-AR to PI3K and to identify the PI3K isoform mediating this transactivation in a cardiac context. Acute β-AR stimulation with isoproterenol resulted in increased tyrosine kinase-associated PI3K activity and phosphorylation of Akt and p70S6K in H9c2 cardiomyocytes. Cotreatment with ICI-118,551, but not CGP-20712, abolished the increase in PI3K activity, suggesting a β2-AR-mediated event. PI3K activation was also abrogated by cotreatment with pertussis toxin, 4-amino-5-(4-chlorophenyl)-7-( t-butyl)pyrazolol[3,4-d]pyrimidine (PP2, a selective Src-family tyrosine kinases inhibitor), or AG-1296 [selective platelet-derived growth factor receptor (PDGFR) inhibitor] but not with an inhibitor for protein kinase A, protein kinase C, Ras, adenylyl cyclase, epidermal growth factor receptor, or insulin-like growth factor-1 receptor. β-AR stimulation induced an increase in tyrosine phosphorylation of PDGFR, which was abolished by inhibition of Src either by PP2 or small interfering RNA. Moreover, H9c2 cardiomyocytes stably transfected with a vector expressing a Gβγ sequestrant peptide derived from the COOH-terminus of β-AR kinase-1 failed to activate PI3K after β-AR stimulation, suggesting Gβγ is required for the transactivation. Furthermore, acute β-AR stimulation in vivo resulted in increases in PDGFR-associated PI3K and PI3Kα isoform activities but not the activities of other isoforms (PI3Kβ, -δ, -γ) in adult mouse heart. Taken together, these data provide in vitro and in vivo evidence for a novel mechanism of β-AR-mediated transactivation of cardiac PI3Kα via sequential involvement of Gαi/Gβγ, Src, and PDGFR.
TGF-β- and CTGF-mediated fibroblast recruitment influences early outward vein graft remodeling
Authors: Jiang Zhihua , Yu Peng , Tao Ming , Fernandez Chessy , Ifantides Cristos , Moloye Olajompo , Schultz Gregory S. , Ozaki C. Keith , Berceli Scott A.
Published: March 2007

Luminal shearing forces have been shown to impact both geometric remodeling and the development of intimal hyperplasia. Less well studied is the influence of intramural wall stresses on vessel growth and adaptation. Using a vein graft-fistula configuration to isolate the impact of circumferential wall stress, we identify the reorganization of adventitial myofibroblasts as the dominant histological event that limits early outward remodeling of vein grafts in response to elevated wall stress. We hypothesize that increased production of transforming growth factor-β (TGF-β) and connective tissue growth factor (CTGF) induces recruitment of myofibroblasts, promotes adventitial reorganization, and limits early outward remodeling in response to increased intramural wall stress. Vein grafts with a distal arteriovenous fistula in the neck of rabbits were constructed, resulting in a fourfold differential in circumferential wall stress. Using this model, we demonstrate 1) elevated wall stress augments the production of TGF-β and CTGF, 2) increased TGF-β expression and CTGF expression are correlated with the enhanced differentiation from fibroblasts to myofibroblasts, as evidenced by the significant increase in the α-actin-positive cells in adventitia, and 3) the levels of TGF-β, CTGF, and α-actin are inversely correlated with the magnitude of outward remodeling of the graft wall. Increased wall stress after vein graft implantation appears to induce a TGF-β- and CTGF-mediated recruitment of adventitial fibroblasts and a conversion to a myofibroblast phenotype. Although important in the maintenance of wall stability in the face of an increased mechanical load, this adventitial adaptation limits early outward remodeling of the vein conduit and may prove deleterious in maintaining long-term vein graft patency.
The functional consequence of RhoA knockdown by RNA interference in rat cerebral arteries
Authors: Corteling Randolph L. , Brett Suzanne E. , Yin Hao , Zheng Xi-Long , Walsh Michael P. , Welsh Donald G.
Published: March 2007

Uridine triphosphate (UTP) constricts cerebral arteries by activating transduction pathways that increase cytosolic [Ca2+] and myofilament Ca2+ sensitivity. The signaling proteins that comprise these pathways remain uncertain with recent studies implicating a role for several G proteins. To start clarifying which G proteins enable UTP-induced vasoconstriction, a small interfering RNA (siRNA) approach was developed to knock down specified targets in rat cerebral arteries. siRNA directed against Gq and RhoA was introduced into isolated cerebral arteries using reverse permeabilization. Following a defined period of organ culture, arteries were assayed for contractile function, mRNA levels, and protein expression. Targeted siRNA reduced RhoA or Gq mRNA expression by 60–70%, which correlated with a reduction in RhoA but not Gq protein expression. UTP-induced constriction was abolished in RhoA-depleted arteries, but this was not due to a reduction in myosin light chain phosphorylation. UTP-induced actin polymerization was attenuated in RhoA-depleted arteries, which would explain the loss of agonist-induced constriction. In summary, this study illustrates that siRNA approaches can be effectively used on intact arteries to induce targeted knockdown given that the protein turnover rate is sufficiently high. It also demonstrates that the principal role of RhoA in agonist-induced constriction is to facilitate the formation of F-actin, the physical structure to which phosphorylated myosin binds to elicit arterial constriction.
The effects of PM10particles and oxidative stress on macrophages and lung epithelial cells: modulating effects of calcium-signaling antagonists
Authors: Brown David M. , Hutchison Laura , Donaldson Kenneth , Stone Vicki
Published: March 2007

We have previously examined the ability of air pollution particles (PM10) to promote release of the proinflammatory cytokine tumor necrosis factor-α (TNF-α) from human peripheral blood mononuclear cells and demonstrated a role for calcium as a signaling molecule in this process. We have now studied the ability of oxidative stress induced by a synthetic oxidant tert-butyl hydroperoxide (tBHP) to induce TNF-α production via calcium signaling in the mouse macrophage cell line (J774). The oxidant tBHP significantly increased intracellular calcium and the release of TNF-α in J774 cells, an effect that was reduced to control levels by inhibition of calcium signaling with verapamil, BAPTA-AM, and W-7. This study also investigated interactions between PM10-treated macrophages and epithelial cells by using conditioned medium (CM) from PM10-treated mononuclear cells to stimulate the release of the neutrophil chemoattractant chemokine IL-8 from A549 lung epithelial cells. TNF-α protein release was demonstrated in human mononuclear cells after PM10treatment, an effect that was inhibited by calcium antagonists. Treatment of A549 cells with monocyte/PM10CM produced increased IL-8 release that was reduced with CM from monocyte/PM10/calcium antagonist treatments. The expression of ICAM-1 was increased after incubation with CM from monocyte/PM10treatment, and this increase was prevented by treatment with CM from monocyte/PM10/calcium antagonist. These data demonstrate a link between oxidative stress, calcium, and inflammatory mediator production in macrophages and lung epithelial cells.
Hydrogen peroxide-induced Ca2+mobilization in pulmonary arterial smooth muscle cells
Authors: Lin Mo-Jun , Yang Xiao-Ru , Cao Yuan-Ning , Sham James S. K.
Published: March 2007

Reactive oxygen species (ROS) generated from NADPH oxidases and mitochondria have been implicated as key messengers for pulmonary vasoconstriction and vascular remodeling induced by agonists and hypoxia. Since Ca2+mobilization is essential for vasoconstriction and cell proliferation, we sought to characterize the Ca2+response and to delineate the Ca2+pathways activated by hydrogen peroxide (H2O2) in rat intralobar pulmonary arterial smooth muscle cells (PASMCs). Exogenous application of 10 μM to 1 mM H2O2elicited concentration-dependent increase in intracellular Ca2+concentration in PASMCs, with an initial rise followed by a plateau or slow secondary increase. The initial phase was related to intracellular release. It was attenuated by the inositol trisphosphate (IP3) receptor antagonist 2-aminoethyl diphenylborate, ryanodine, or thapsigargin, but was unaffected by the removal of Ca2+in external solution. The secondary phase was dependent on extracellular Ca2+influx. It was unaffected by the voltage-gated Ca2+channel blocker nifedipine or the nonselective cation channel blockers SKF-96365 and La3+, but inhibited concentration dependently by millimolar Ni2+, and potentiated by the Na+/Ca2+exchange inhibitor KB-R 7943. H2O2did not alter the rate of Mn2+quenching of fura 2, suggesting store- and receptor-operated Ca2+channels were not involved. By contrast, H2O2elicited a sustained inward current carried by Na+at −70 mV, and the current was inhibited by Ni2+. These results suggest that H2O2mobilizes intracellular Ca2+through multiple pathways, including the IP3- and ryanodine receptor-gated Ca2+stores, and Ni2+-sensitive cation channels. Activation of these Ca2+pathways may play important roles in ROS signaling in PASMCs.
A role for CFTR in the elevation of glutathione levels in the lung by oral glutathione administration
Authors: Kariya Chirag , Leitner Heather , Min Elysia , van Heeckeren Christiaan , van Heeckeren Anna , Day Brian J.
Published: March 2007

The cystic fibrosis transmembrane conductance regulator (CFTR) protein is the only known apical glutathione (GSH) transporter in the lung. The purpose of these studies was to determine whether oral GSH or glutathione disulfide (GSSG) treatment could increase lung epithelial lining fluid (ELF) GSH levels and whether CFTR plays a role in this process. The pharmacokinetic profile of an oral bolus dose of GSH (300 mg/kg) was determined in mice. Plasma, ELF, bronchoalveolar lavage (BAL) cells, and lung tissue were analyzed for GSH content. There was a rapid elevation in the GSH levels that peaked at 30 min in the plasma and 60 min in the lung, ELF, and BAL cells after oral GSH dosing. Oral GSH treatment produced a selective increase in the reduced and active form of GSH in all lung compartments examined. Oral GSSG treatment (300 mg/kg) resulted in a smaller increase of GSH levels. To evaluate the role of CFTR in this process, Cftr knockout (KO) mice and gut-corrected Cftr KO-transgenic (Tg) mice were given an oral bolus dose of GSH (300 mg/kg) and compared with wild-type mice for changes in GSH levels in plasma, lung, ELF, and BAL cells. There was a twofold increase in plasma, a twofold increase in lung, a fivefold increase in ELF, and a threefold increase in BAL cell GSH levels at 60 min in wild-type mice; however, GSH levels only increased by 40% in the plasma, 60% in the lung, 50% in the ELF, and twofold in the BAL cells within the gut-corrected Cftr KO-Tg mice. No change in GSH levels was observed in the uncorrected Cftr KO mice. These studies suggest that CFTR plays an important role in GSH uptake from the diet and transport processes in the lung.
Nox4 mediates TGF-β1-induced retinoblastoma protein phosphorylation, proliferation, and hypertrophy in human airway smooth muscle cells
Authors: Sturrock Anne , Huecksteadt Thomas P. , Norman Kimberly , Sanders Karl , Murphy Thomas M. , Chitano Pasquale , Wilson Kimberly , Hoidal John R. , Kennedy Thomas P.
Published: March 2007

Transforming growth factor-β1 (TGF-β1) plays a pivotal role in increasing airway smooth muscle mass in severe asthma by inducing proliferation and hypertrophy of human airway smooth muscle. The mechanism(s) for these effects of TGF-β1 have not been fully elucidated. In this study, we demonstrate that TGF-β1 is a potent inducer of expression of the nonphagocyte NAD(P)H oxidase catalytic homolog Nox4, diphenylene iodonium-inhibitable reactive oxygen species production, proliferation, and hypertrophy in cultured human airway smooth muscle cells. By confocal microscopy, TGF-β1-induced Nox4 was localized with the endoplasmic reticulum and the nucleus, implying a role for Nox4 in regulation of both the cell cycle and protein synthesis. Consistent with this hypothesis, TGF-β1 increased retinoblastoma protein phosphorylation at both Ser807/811 and Ser780. Silencing Nox4 prevented TGF-β1-mediated retinoblastoma protein phosphorylation, proliferation, and cell hypertrophy. TGF-β1 also increased phosphorylation of eukaryotic translation initiation factor 4E binding protein-1 at Thr37/46, and this was likewise blocked by silencing Nox4. This is the first report to suggest a functional role for Nox4 in cell cycle transition and to demonstrate that Nox4 influences the pathobiochemistry of asthma by generating reactive oxygen species that promote TGF-β1-induced proliferation and hypertrophy of human airway smooth muscle.
Low fat adiponectin expression is associated with oxidative stress in nondiabetic humans with chronic kidney disease—impact on plasma adiponectin concentration
Authors: Barazzoni Rocco , Bernardi Annamaria , Biasia Franco , Semolic Annamaria , Bosutti Alessandra , Mucci MariaPia , Dore Franca , Zanetti Michela , Guarnieri Gianfranco
Published: March 2007

In spite of association between high plasma adiponectin and high metabolic and cardiovascular (CV) risk, highest adiponectin increments retain CV and metabolic protective effects in advanced chronic kidney disease (CKD). Passive accumulation can favor CKD-associated hyperadiponectinemia but potential additional regulation by adipose tissue remains undefined. Oxidative stress (OS) is associated with metabolic and CV disease and with CKD [increasing from conservative treatment (CT) to maintenance hemodialysis (MHD)], and OS can reduce adiponectin expression in experimental models. OS (in the form of plasma thiobarbituric acid-reactive substances: TBARS), subcutaneous adipose adiponectin mRNA, and plasma adiponectin were studied in CKD patients ( stages 4 and 5) on CT ( n = 7) or MHD ( n = 11). Compared with CT and controls (C: n = 6) MHD had highest TBARS and lowest adiponectin mRNA ( P < 0.05) with lower adipose adiponectin protein ( P < 0.05 vs. CT). MHD also had lower plasma adiponectin than CT, although both had higher adiponectin than C ( P < 0.05). In renal transplant recipients (RT: CKD stage 3; n = 5) normal TBARS were, in turn, associated with normal adiponectin mRNA ( P < 0.05 vs. MHD). In all CKD ( n = 23), adiponectin mRNA was associated positively with adiponectin plasma concentration ( P < 0.01). In all subjects ( n = 29), adiponectin mRNA was related ( P < 0.05) negatively with TBARS after adjusting for plasma C-reactive protein (CRP) or CRP and creatinine. Thus altered OS, adiponectin expression, and plasma concentration represent a novel cluster of metabolic and CV risk factors in MHD that are normalized in RT. The data suggest novel roles of 1) MHD-associated OS in modulating adiponectin expression and 2) adipose tissue in contributing to circulating adiponectin in advanced CKD.
Prostaglandin E2inhibits vasotocin-induced osmotic water permeability in the frog urinary bladder by EP1-receptor-mediated activation of NO/cGMP pathway
Authors: Bachteeva Vera , Fock Ekaterina , Lavrova Elena , Nikolaeva Svetlana , Gambaryan Stepan , Parnova Rimma
Published: March 2007

PGE2is a well-known inhibitor of the antidiuretic hormone-induced increase of osmotic water permeability (OWP) in different osmoregulatory epithelia; however, the mechanisms underlying this effect of PGE2are not completely understood. Here, we report that, in the frog Rana temporaria urinary bladder, EP1-receptor-mediated inhibition of arginine-vasotocin (AVT)-induced OWP by PGE2is attributed to increased generation of nitric oxide (NO) in epithelial cells. It was shown that the inhibitory effect of 17-phenyl-trinor-PGE2(17-ph-PGE2), an EP1agonist, on AVT-induced OWP was significantly reduced in the presence of 7-nitroindazole (7-NI), a neuronal NO synthase (nNOS) inhibitor. NO synthase (NOS) activity in both lysed and intact epithelial cells measured as a rate of conversion of l-[3H]arginine to l-[3H]citrulline was Ca2+dependent and inhibited by 7-NI. PGE2and 17-ph-PGE2, but not M&B-28767 (EP3agonist) or butaprost (EP2agonist), stimulated NOS activity in epithelial cells. The above effect of PGE2was abolished in the presence of SC-19220, an EP1antagonist. 7-NI reduced the stimulatory effect of 17-ph-PGE2on NOS activity. 17-ph-PGE2increased intracellular Ca2+concentration and cGMP in epithelial cells. Western blot analysis revealed an nNOS expression in epithelial cells. These results show that the inhibitory effect of PGE2on AVT-induced OWP in the frog urinary bladder is based at least partly on EP1-receptor-mediated activation of the NO/cGMP pathway, suggesting a novel cross talk between AVT, PGE2, and nNOS that may be important in the regulation of water transport.
Alveolar diffusion-perfusion interactions during high-altitude residence in guinea pigs
Authors: Yilmaz Cuneyt , Merrill Dane D. , Hsia Connie C. W.
Published: March 2007

We previously reported in weanling guinea pigs raised at high altitude (HA; 3,800 m) an elevated lung diffusing capacity estimated by morphometry from alveolar-capillary surface area, harmonic mean blood-gas barrier thickness, and pulmonary capillary blood volume (Vc) compared with litter-matched control animals raised at an intermediate altitude (IA; 1,200 m) (Hsia CCW, Polo Carbayo JJ, Yan X, Bellotto DJ. Respir Physiol Neurobiol 147: 105–115, 2005). To determine if HA-induced alveolar ultrastructural changes are associated with improved alveolar function, we measured lung diffusing capacity for carbon monoxide (DlCO), membrane diffusing capacity for carbon monoxide (DmCO), Vc, pulmonary blood flow, and lung volume by a rebreathing technique in litter-matched male weanling Hartley guinea pigs raised at HA or IA for 4 or 12 mo. Separate control animals were also raised and studied at sea level (SL). Resting measurements were obtained in the conscious nonsedated state. In HA animals compared with corresponding IA or SL controls, lung volume and hematocrit were significantly higher while pulmonary blood flow was lower. At a given pulmonary blood flow, DlCO and DmCO were higher in HA-raised animals than in control animals without a significant change in Vc. We conclude that 1) HA residence enhanced physiological diffusing capacity corresponding to that previously estimated on the basis of structural adaptation, 2) adaptation in diffusing capacity and its components should be interpreted with respect to pulmonary blood flow, and 3) this noninvasive rebreathing technique could be used to follow adaptive responses in small animals.
Individual limb work does not explain the greater metabolic cost of walking in elderly adults
Authors: Ortega Justus D. , Farley Claire T.
Published: March 2007

Elderly adults consume more metabolic energy during walking than young adults. Our study tested the hypothesis that elderly adults consume more metabolic energy during walking than young adults because they perform more individual limb work on the center of mass. Thus we compared how much individual limb work young and elderly adults performed on the center of mass during walking. We measured metabolic rate and ground reaction force while 10 elderly and 10 young subjects walked at 5 speeds between 0.7 and 1.8 m/s. Compared with young subjects, elderly subjects consumed an average of 20% more metabolic energy ( P = 0.010), whereas they performed an average of 10% less individual limb work during walking over the range of speeds ( P = 0.028). During the single-support phase, elderly and young subjects both conserved ∼80% of the center of mass mechanical energy by inverted pendulum energy exchange and performed a similar amount of individual limb work ( P = 0.473). However, during double support, elderly subjects performed an average of 17% less individual limb work than young subjects ( P = 0.007) because their forward speed fluctuated less ( P = 0.006). We conclude that the greater metabolic cost of walking in elderly adults cannot be explained by a difference in individual limb work. Future studies should examine whether a greater metabolic cost of stabilization, reduced muscle efficiency, greater antagonist cocontraction, and/or a greater cost of generating muscle force cause the elevated metabolic cost of walking in elderly adults.
Respiratory syncytial virus infection in anesthetized weanling rather than adult rats prolongs the apneic responses to right atrial injection of capsaicin
Authors: Peng Wenhong , Zhuang Jianguo , Harrod Kevin S. , Xu Fadi
Published: March 2007

Apnea is a common complication in infants infected by respiratory syncytial virus (RSV). A recent study has shown that intranasal inoculation of RSV in conscious weanling rats strengthens the apneic responses to right atrial injection of capsaicin (CAP), leading to 66% mortality. The objectives of the present study were to determine 1) whether RSV infection changes baseline minute ventilation (V̇e) and arterial blood gases in anesthetized rats; 2) what the effects of RSV infection are on the respiratory responses to CAP; and 3) whether the RSV-strengthened apneic responses are age dependent. Our experiments were conducted in anesthetized and spontaneously breathing rats divided into four groups of weanling and adult rats that received either intranasal inoculation of RSV or virus-free medium. Two days after RSV infection (0.7 ml/kg), animal blood gases, baseline V̇e, and V̇e responses to right atrial injection of three doses of CAP (4, 16, and 64 μg/kg) were measured and compared among the four groups. Our results showed that RSV infection increased respiratory frequency (∼25%, P < 0.05) in weanling but not adult rats, with little effect on arterial blood gases. RSV infection amplified the apneic responses to CAP in weanling but not adult rats, characterized by increases in the initial (40%) and the longest apneic duration (650%), the number of apneic episodes (139%), and the total duration of apneas (60%). These amplifications led to 50% mortality ( P < 0.05). We conclude that RSV infection increases respiratory frequency and strengthens the apneic responses to CAP only in anesthetized weanling but not adult rats.
Epidermal Growth Factor-, Transforming Growth Factor-β-, Retinoic Acid- and 1,25-Dihydroxyvitamin D3-Regulated Expression of the Novel Protein PTPIP51 in Keratinocytes
Authors: Stenzinger Albrecht , Schreiner Dietmar , Pfeiffer Thorsten , Tag Claudia , Hofer Hans Werner , Wimmer Monika
Published: March 2007

The novel protein PTPIP51 (protein tyrosine phosphatase-interacting protein 51), which has been found to interact with protein tyrosine phosphatases of the PTP1B/TcPTP subfamily, is expressed in all suprabasal layers of human epidermis. Hence, a human keratinocyte cell line (HaCaT) grown on culture slides was used as a simplified model system to study the influence of hormonal agents on the regulation of PTPIP51 expression. Results were obtained by immunocytochemistry and subsequent statistical analysis. Additionally, immunoblotting was performed to detect the possible occurrence of distinct molecular weight forms as described previously. Subcellular localization of PTPIP51 protein was analyzed by specific staining of cellular organelles. HaCaT cells were subjected to treatment with factors that are crucial for the regulation of proliferation and differentiation of keratinocytes in human epidermis: epidermal growth factor (EGF), transforming growth factor-β(TGF-β), retinoic acid (RA) and 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]. Epidermal growth factor receptor (EGFR) expressed in HaCaT cells was inhibited by PD153035. Only about 35% of untreated HaCaT cells were immunoreactive for the PTPIP51 protein. Whereas cells treated with increasing concentrations of 1,25 (OH)2D3 showed a stepwise numerical increase of PTPIP51-positive cells, treatment with RA did not influence the number of PTPIP51-positive cells except when supraphysiological concentrations were applied. Concentration-dependent increase of cells stained positive for PTPIP51 was also observed when HaCaT cells were subjected to EGF treatment. Additional treatment of these cells with PD153035 led to a slight decrease in the fraction of PTPIP51-positive cells, which was not statistically significant. Immunoblotting results suggest a specific pattern of different molecular weight forms of PTPIP51 being expressed in HaCaT cells. Subcellular analysis revealed an association of the protein with mitochondria in nonconfluent cells, whereas confluent cells lack such correlation. The intracellular distribution of PTPIP51 resembled the localization of its interacting partner TcPTP. Furthermore, PTPIP51 was found to be present in both the nucleus and cytoplasm of HaCaT cells. In summary, the results indicate a possible association of PTPIP51 expression with differentiation as well as with apoptosis of keratinocytes.
In the Ventral Tegmental Area, Progestins’ Membrane-Mediated Actions for Lordosis of Hamsters and Rats Involve Protein Kinase A
Authors: Petralia Sandra M. , Walf Alicia A. , Frye Cheryl A.
Published: March 2007

Progestin-facilitated lordosis of hamsters and rats is enhanced by activation of dopamine type 1 (D1) or GABAA/benzodiazepine receptor complexes (GBRs) in the ventral tegmental area (VTA) and these effects involve G-proteins and second messengers, such as adenosine 3′,5′-monophosphate (cAMP). We examined whether D1- and/or GBR-mediated increases in progestin-facilitated lordosis of female hamsters and rats involve the cAMP-dependent protein kinase, protein kinase A (PKA), in the VTA. In experiment 1, ovariectomized hamsters, primed with estradiol (E2; 10 µg at h 0) + progesterone (P; 100 µg at h 45), were first pre-tested for lordosis and motor behavior (h 48) and then infused with the PKA inhibitor, Rp-cAMP (100 ng/side), or vehicle. Thirty minutes later, hamsters were retested and then received infusions of the D1 agonist, SKF38393 (100 ng/side), the GBR agonist, muscimol (100 ng/side), or vehicle to the VTA. Hamsters were post-tested for lordosis and motor behavior 30 min later. In Experiment 2, ovariectomized rats, primed with E2 (10 µg at h 0), were first pre-tested for lordosis and then infused with Rp-cAMP (100 ng/side) or vehicle to the VTA at h 44. Immediately after testing, rats received infusions of SKF38393 (100 ng/side), muscimol (100 ng/side), or vehicle and were retested for lordosis. Rats were then infused with the neurosteroid, 5α-pregnan-3α-ol-20-one (3α,5α-THP; 100 or 200 ng/side), or β-cyclodextrin vehicle and were post-tested for lordosis and motor behavior 10 and 60 min later. The enhancing effects of progestins or progestins plus D1 or GBR activation on lordosis of E2-primed hamsters and rats were blocked by the PKA inhibitor, Rp-cAMP. Thus, in the VTA, progestins’ membrane actions involving D1 or GBRs are mediated, in part, by PKA.
A de novo Complex Chromosome Rearrangement Involving Chromosomes 2, 3, 5, 9 and 11 Detected Prenatally and Studied Postnatally by Conventional Cytogenetics and Molecular Cytogenetic Analyses
Authors: Mechoso Búrix , Vaglio Alicia , Quadrelli Andrea , Mark Hon Fong L. , Huang Xin-Li , Milunsky Aubrey , Quadrelli Roberto
Published: March 2007

Objective: To describe a de novo complex chromosome rearrangement (CCR) detected prenatally and studied after birth. Methods: Conventional cytogenetics and fluorescent in situ hybridization (FISH) were performed on amniotic fluid and peripheral blood. High-resolution comparative genomic hybridization (HR-CGH) analysis was made postnatally. Results: Prenatal/postnatal cytogenetic, FISH and HR-CGH analyses revealed an apparently balanced de novo CCR ascertained as 46,XY,t(2;3;9)(q21;p24;q22),der(5)inv(5)(?p11q13)t(5;11)(?p13;q25),ins(5;3)(?p13;?p23p24). At 9 months, the child has neither congenital anomalies nor evidence of delayed psychomotor development. Conclusions: Our report describes a rare CCR detected prenatally and shows the usefulness of FISH and CGH in complementing conventional cytogenetics.
Prenatal Diagnosis of Hb Bart’s Hydrops Fetalis Caused by a Genetic Compound Heterozygosity for Two Different αo-Thalassemia Determinants
Authors: Siriratmanawong Nirut , Pinmuang-ngam Charnchai , Fucharoen Goonnapa , Fucharoen Supan
Published: March 2007

Objective: To describe prenatal diagnosis of hemoglobin (Hb) Bart’s hydrops fetalis caused by a previously undescribed condition in Thailand of the interaction of αo-thalassemia with the Southeast Asian (–SEA) and the THAI (–THAI) deletions in a Thai family. Methods: Molecular and hematological features associated with αo-thalassemia carriers found in a pregnant Thai woman and her husband were investigated. Prenatal diagnosis was performed at her third pregnancy on amniotic fluid using a multiplex PCR for simultaneous detection of the (–SEA) and (–THAI) αo-thalassemia determinants. Results: Initial DNA analysis for the common form of αo-thalassemia (–SEA) identified that the pregnant woman was a carrier whereas her husband was negative for this mutation. They had a healthy non-thalassemic daughter from the first pregnancy but the second pregnancy ended unexpectedly with Hb Bart’s hydrops fetalis. Analysis of the archived DNA specimen of the husband demonstrated that he was in fact a carrier of the (–THAI) αo-thalassemia determinant. A successful application of a multiplex PCR for simultaneous detection of these two forms of αo-thalassemia in prenatal diagnosis at the third pregnancy was demonstrated and identified that the fetus was a carrier of the (–SEA) αo-thalassemia. Conclusion: Although relatively rare, the (–THAI) αo-thalassemia should be included in a routine screening to prevent a Hb Bart’s hydrops fetalis syndrome. A simple DNA assay based on a multiplex PCR approach would therefore provide an effective means for αo-thalassemia screening in the region.
The Fetal Dandy Walker Complex: Associated Anomalies, Perinatal Outcome and Postnatal Imaging
Authors: Harper Terry , Fordham Lynn Ansley , Wolfe Honor M.
Published: March 2007

Objective: This study compares the diagnostic accuracy of prenatal ultrasound with postnatal imaging and evaluates for associated fetal anomalies and their impact on immediate neonatal outcome in fetal Dandy Walker complex (DWC). Methods: Cases of fetal DWC diagnosed in a single ultrasound unit from January 2000 through July 2004 were reviewed for associated fetal anomalies, fetal karyotype, immediate neonatal outcome and postnatal head imaging. Results: A total of 55 fetuses with DWC were identified. Of liveborn cases, postnatal imaging confirmed prenatal intracranial findings in 50% of Dandy Walker variant (DWV) and 100% of Dandy Walker malformation (DWM), with additional central nervous system findings noted in 21% of all cases. Additional fetal anomalies were seen in 26/40 (65%) cases of DWV and 15/15 (100%) cases of DWM. Immediate neonatal survival was predicted by karyotype and associated fetal anomalies. Conclusion: The sonographic diagnosis of fetal DWM is accurate. Significant discrepancies exist in prenatal and postnatal diagnosis of DWV. Comprehensive fetal ultrasound and karyotype should be offered for all fetuses with DWC. Postnatal imaging should be performed on all fetal DWC.
A Case of Intra-Amniotic Maternal Hemorrhage in Term Pregnancy
Authors: Sijanovic Sinisa , Selthofer Robert , Abicic-Žuljevic Kristina , Milojkovic Miodrag , Topolovec Zlatko , Sijanovic Ivanka , Kulaš Tomislav
Published: March 2007

Introduction: Intra-amniotic maternal hemorrhage is a rare condition, hard to differentiate from some other conditions in pregnancy. We report an unusual case of intra-amniotic maternal hemorrhage in term pregnancy ending in urgent cesarean section, identified on ultrasound examination. Case Report: A 24-year-old female (gravida 3, para 2, abortion 0) was admitted to hospital at 40 weeks’ gestation for collapse and general weakness. Her blood pressure was 90/60 mm Hg, pulse 112 bpm, temperature 36°C. The fetal heart race tracings were stable and reactive. The cervix was opened 3 cm. Further examination revealed no vaginal bleeding and normal amnioscopic findings. An ultrasound examination confirmed singleton, head-presenting gestation without any visible congenital anomalies, amniotic fluid index was at the 50th percentile, anterior placenta without evidence of previa, abruption or retroplacental hematoma. An inhomogeneous echogenic mass, measuring 12 × 8 cm, was noted within the amniotic cavity, there was no evidence of pathological flow through the mass on color Doppler. After admittance to the hospital, the patient complained of regular pains, weakness and collapsed with signs of hemorrhagic shock. Repeated ultrasound evaluation showed no change in acoustic texture and size, but the amniotic fluid now had a hyperechoic appearance which revealed increasing intra-amniotic hemorrhage. Because of clinical signs of maternal hemorrhagic shock confirmed by laboratory findings of decreasing red blood parameters, an urgent cesarean section was performed. A female infant weighing 3,070 g, Apgar score 5/7, was delivered. After removal of the placenta there was no sign of abruption, which was confirmed at histopathology. Hemoglobin A was detected in the amniotic fluid by Abt’s test. The patient had an uncomplicated postoperative course. The infant developed normally. Conclusion: When there are signs of fetal distress or maternal hemorrhagic shock, an urgent cesarean section should be performed.
Oxidative Stress Alters the Integrity of Cell-Free mRNA Fragments Associated with Placenta-Derived Syncytiotrophoblast Microparticles
Authors: Rusterholz Corinne , Holzgreve Wolfgang , Hahn Sinuhe
Published: March 2007

Objectives: Cell-free fetal RNA from placental origin is present in the peripheral blood of pregnant women. Therefore, its qualitative analysis might provide insights into the physiological condition of the placenta. Here, we examine whether oxidative stress affects the integrity of placentally derived mRNA in vitro. Methods: Placental explants were cultured under normal or oxidative conditions, and the levels of placental and syncytiotrophoblast microparticle associated glyceraldehyde-3-phosphate dehydrogenase 3′ versus 5′ mRNA fragments were analyzed by real-time reverse-transcriptase polymerase chain reaction. Result: The relative ratio of 3′ to 5′ mRNA fragments associated with placental microparticles was significantly altered upon culture under oxidative stress. Conclusions: Our data suggest that oxidative stress reduces the levels of full-length, particle-associated glyceraldehyde-3-phosphate dehydrogenase mRNA transcripts released by the placenta. Therefore, analysis of the microparticle-coupled mRNA integrity in pregnant women might prove useful to diagnose disorders such as preeclampsia, where placental oxidative stress is involved.
False-Positive Quadruple Screen Test for Trisomy 18 in a Patient with a Fetus with Bloom’s Syndrome
Authors: Baxi Laxmi , Brown Stephen , Thaker Harshwardhan M.
Published: March 2007

In the literature, conflicting reports on the significance of false-positive maternal serum multiple marker testing for trisomy 18 are encountered; however, the biology of this finding is discussed infrequently. We present such a case in association with Bloom’s syndrome in the fetus. The fetus had intrauterine growth restriction, seen early in the second trimester, oligohydramnios, and was delivered at 34 weeks of gestation for impending fetal compromise. We propose that the adverse outcome of the pregnancy with false-positive serum analyte testing for trisomy 18 might result from a small-sized placenta and perhaps pathology at receptor level.
Pharmacokinetics of Imipenem in Sheep with Special Reference to Its Hepato-Renal Effects
Authors: EL-Sooud K. Abo , Khammas W. , Kadri M. Nadir
Published: March 2007

Background: Imipenem is a carbapenem antibiotic with a broad spectrum of activity. The aim of this study was to determine the pharmacokinetics of imipenem after single intravenous and intramuscular injections and the effect of repeated intramuscular injections on hepato-renal functions in sheep. Methods: Imipenem concentrations in plasma and urine were determined by a microbiological agar plate assay. Results: Following intramuscular injection, imipenem was rapidly absorbed and the peak plasma concentration was 9.99 µg ml–1 and the systemic bioavailability was 65.97%. Urine concentrations of imipenem were much higher than in plasma. Light and electron microscopy evidenced little changes in the kidney andliver. Conclusion: Imipenem is likely to be efficacious in most urinary tract infections and has no adverse effects on hepato-renal structures and functions.
One-Day Thyroid Surgery: Retrospective Analysis of Safety and Patient Satisfaction on a Consecutive Series of 1,571 Cases over a Three-Year Period
Authors: Materazzi G. , Dionigi G. , Berti P. , Rago R. , Frustaci G. , Docimo G. , Puccini M. , Miccoli P.
Published: March 2007

Short-stay thyroid surgery (<24 h hospital stay) is becoming increasingly popular but some potentially lethal complications are considered strong arguments against shortening hospitalization after thyroidectomy. The authors reviewed the data of 1,571 patients undergoing one-day thyroid surgery over a 3-year period to determine safety and patient satisfaction. There were 1,244 females and 327 males. Mean age was 43 years. Patient satisfaction was evaluated by a questionnaire given on discharge, while post-discharge surgical recovery was analyzed by the PSR scale. Total thyroidectomy was performed in 1,119 patients (71%), hemithyroidectomy in 450 (29%), isthmusectomy in 2. Morbidity occurred in 152 patients (9.6%). Surgical complications were transient hypocalcemia in 112 cases and permanent hypoparathyroidism in 3; monolateral transient nerve palsy occurred in 10 cases, bilateral in 3; definitive monolateral recurrent palsy in 4 cases. Bleeding requiring re-intervention occurred in 10 cases, wound complications in 5 cases, and intraoperative tracheal lesion in 1 patient. Among complicated patients, 129 (84.8%) were treated after discharge as outpatients. Conversion to inpatient treatment occurred in 28 patients (1.7%) (25 for surgical reasons). Four patients (0.2%) required hospital readmission. Patients were very satisfied in 84.2%, satisfied in 9.5%, poorly satisfied in 4.3%, completely unsatisfied in 2%. Postoperative recovery mean score by PSR scale resulted in 85.14% (0–100%). Our results confirm that the one-day surgery model is safe, effective, and highly agreeable in patients undergoing surgery for thyroid disease.
Safety of Rosuvastatin: Update on 16,876 Rosuvastatin-Treated Patients in a Multinational Clinical Trial Program
Authors: Shepherd James , Vidt Donald G. , Miller Elinor , Harris Susan , Blasetto James
Published: March 2007

Background: The safety and tolerability of rosuvastatin were assessed using data from 16,876patients who received rosuvastatin 5–40 mg in a multinational phase II/III/IIIb/IV program, representing 25,670 patient-years of continuous exposure to rosuvastatin. Methods: An integrated database, consisting of 33 trials whose databases were locked up to and including September 16, 2005, was used to examine adverse events and laboratory data. Results: In placebo-controlled trials, adverse events irrespective of causality assessment occurred in 52.1% of patients receiving rosuvastatin 5–40 mg (n = 931) and 51.8% of patients receiving placebo (n = 483). In all controlled clinical trials with comparator statins, rosuvastatin 5–40 mg was associated with an adverse event profile similar to profiles for atorvastatin 10–80 mg, simvastatin 10–80 mg, and pravastatin 10–40 mg. Clinically significant elevations in alanine aminotransferase (>3 times the upper limit of normal [ULN] on at least 2 consecutive occasions) were uncommon (≤0.2%) in the rosuvastatin and comparator statin groups. Elevated creatine kinase >10 times ULN occurred in ≤0.3% of patients receiving rosuvastatin or other statins. Myopathy (creatine kinase >10 times ULN with muscle symptoms) possibly related to treatment occurred in 0.03% of patients taking rosuvastatin at doses ≤40 mg. The frequency of dipstick-positive proteinuria at rosuvastatin doses ≤20 mg was comparable to that seen with other statins, and the development of proteinuria was not predictive of acute or progressive renal disease. Both short- and long-term rosuvastatin treatment were associated with small increases in estimated glomerular filtration rate, with improvements appearing to be somewhat greater in those patients beginning treatment with greater renal impairment. In the phase II–IV program, no deaths were attributed to rosuvastatin; at doses of rosuvastatin ≤40 mg, 1 case of rhabdomyolysis occurred in a patient who received rosuvastatin 20 mg and concomitant gemfibrozil treatment. Conclusion: In summary, rosuvastatin was well tolerated by a broad range of patients with dyslipidemia, and its safety profile was similar to those of comparator statins investigated in the clinical program. (Nota bene: The clinical development program for rosuvastatin initially evaluated rosuvastatin doses up to 80 mg. Following completion of the phase III/IIIb program, a decision was made not to pursue marketing approval for the 80-mg dose because the additional lipid-modifying benefits of this dose did not justify the potential risks for use in the general population of patients with dyslipidemia.)