bluuebunny/crossref_metadata_2025 huggingface dataset:
A LEARNING-BASED FRAMEWORK FOR GRAPH MATCHING
Authors: VAN WYK M. A. , VAN WYK B. J.
Published: May 2004

This paper presents a unifying review of a learning-based framework for kernel-based attributed graph matching. The framework, which includes as special cases the RKHS Interplator-Based Graph Matching (RIGM) and Interpolator-Based Kronecker Product Graph Matching (IBKPGM) algorithms, incorporates a general approach where no assumption is made about the adjacency structure of the graphs to be matched. Corresponding pairs of attributed adjacency matrices and attribute vectors of an input and reference graph are used as the input–output training set of a constrained multi-input multi-output multi-variable mapping to be learned. It is shown that a Reproducing Kernel Hilbert Space (RKHS) based interpolator can be used to infer this mapping. Partially constraining the inferred mapping by the generation of additional consistency input–output training pairs and the use of polynomial feature augmentation lead to improved performance. The proposed learning-based framework avoids the explicit calculation of compatibility values.
THE ROLE OF RELATIONAL CONSTRAINTS IN REGION MATCHING
Authors: AHMADYFARD ALIREZA , KITTLER JOSEF
Published: May 2004

We propose a graph-based representation for the elliptic region shape descriptors introduced by Tuytelaars et al.13 In this representation we use image profiles to describe the relation between a pair of image regions. This new representation and a graph matching technique proposed in Ref. 1 are the basis of an object recognition method. An experimental comparative study between the original method and the new graph-based method is carried out. The results show that the graph-based method is more robust to scaling than the original method. Moreover, the misclassification rate using the graph-based method is considerably lower than that yielded by the original method.
EXPERIMENTAL STUDY OF CFOA-BASED INDUCTORLESS CHUA'S CIRCUIT
Authors: KILIÇ RECAI
Published: May 2004

The current feedback op amps (CFOAs), with some significant advantages over the conventional op amps, have been used instead of voltage op amps (VOAs) in new implementations of Chua's circuit. In our previous study, after providing a comparative investigation of CFOA-based realizations of Chua's circuit in the literature, we have also presented an alternative inductorless CFOA-based realization of Chua's circuit, and the circuit's chaotic behavior by Pspice simulations. In this paper, we investigate CFOA-based Chua's circuit by constructing an experimental setup, and testing the performance of the proposed implementation at different frequencies. Its excellent high frequency performance was experimentally verified.
DETECTION OF CHAOS IN SOME LOCAL REGIONS OF PHASE PORTRAITS USING SHANNON ENTROPIES
Authors: LING BINGO WING-KUEN , HO CHARLOTTE YUK-FAN , TAM PETER KWONG-SHUN
Published: May 2004

This letter demonstrates the use of Shannon entropies to detect chaos exhibited in some local regions on the phase portraits. When both eigenvalues of the second-order digital filters with two's complement arithmetic are outside the unit circle, the Shannon entropies of the state variables are independent of the initial conditions and the filter parameters, except for some special values of the filter parameters. At these special values, the Shannon entropies of the state variables are relatively small. The state trajectories corresponding to these filter parameters either exhibit random-like chaotic behaviors in some local regions or converge to some fixed points on the phase portraits. Hence, by measuring the Shannon entropies of the state variables, these special state trajectory patterns can be detected. For completeness, we extend the investigation to the case when the eigenvalues of the second-order digital filters with two's complement arithmetic are complex and are inside or on the unit circle. It is found that the Shannon entropies of the symbolic sequences for the type II trajectories may be higher than that for the type III trajectories, even though the symbolic sequences of the type II trajectories are periodic and have limit cycle behaviors, while that of the type III trajectories are aperiodic and have chaotic behaviors.
PITFALLS IN DISTRIBUTED NONBLOCKING CHECKPOINTING
Authors: NI WEIGANG , VRBSKY SUSAN V. , RAY SIBABRATA
Published: May 2004

Coordinated checkpointing has low stable storage requirements and simplifies the recovery process by reserving a set of consistent global checkpoints. Unfortunately, most algorithms that were proposed either incurred a high communication overhead or blocked all processes. Then, a coordinated algorithm was presented which was nonblocking and which forced only a subset of all processes to participate in a checkpointing event. This algorithm was shown to create inconsistencies in some situations and new algorithms to take consistent checkpoints were proposed. However, we found that these algorithms can still result in inconsistencies when typical behavior in a distributed environment is considered, such as multiple forced checkpoints and multiple concurrent checkpoint initiations. In this paper we identify the inconsistencies that can occur and present an efficient nonblocking algorithm that collects consistent global checkpoints and avoids some of the pitfalls in distributed nonblocking checkpointing.
Impact of PPARγ overexpression and activation on pancreatic islet gene expression profile analyzed with oligonucleotide microarrays
Authors: Parton Laura E. , Diraison Frédérique , Neill Suzanne E. , Ghosh Sujoy K. , Rubino Mark A. , Bisi John E. , Briscoe Celia P. , Rutter Guy A.
Published: May 2004

Peroxisome proliferator-activated receptor-γ (PPARγ) serves as a target for the thiazolidinedione class of antidiabetic drugs and is an important regulator of adipose tissue differentiation. By contrast, the principal target genes for PPARγ in the pancreatic islet and the impact of their induction on insulin secretion are largely undefined. Here, we show that mRNAs encoding both isoforms of rodent PPARγ, γ1 and γ2, are expressed in primary rat islets and are upregulated by overexpresssion of the lipogenic transcription factor sterol response element-binding protein 1c. Unexpectedly, however, oligonucleotide microarray analysis demonstrates that graded activation of PPARγ achieved with 1) the thiazolidinedione GW-347845, 2) transduction with adenoviral PPARγ1, or 3) a combination of both treatments progressively enhances the expression of genes involved in fatty acid oxidation and transport. Moreover, maximal activation of PPARγ1 reduces islet triglyceride levels and enhances the oxidation of exogenous palmitate while decreasing glucose oxidation, cellular ATP content, and glucose-, but not depolarization-stimulated, insulin secretion. We conclude that, in the context of the pancreatic islet, the principal response to PPARγ expression and activation is the activation of genes involved in the disposal, rather than the synthesis, of fatty acids. Although fatty acid oxidation may have beneficial effects on β-cell function in the longer term by countering β-cell “lipotoxicity,” the acute response to this metabolic shift is a marked inhibition of insulin secretion.
Leptin treatment markedly increased plasma adiponectin but barely decreased plasma resistin of ob/ob mice
Authors: Delporte Marie-Laure , El Mkadem Samira Ait , Quisquater Muriel , Brichard Sonia M.
Published: May 2004

Adiponectin (ApN) and leptin are two adipocytokines that control fuel homeostasis, body weight, and insulin sensitivity. Their interplay is still poorly studied. These hormones are either undetectable or decreased in obese, diabetic ob/ob mice. We examined the effects of leptin treatment on ApN gene expression, protein production, secretion, and circulating levels of ob/ob mice. We also briefly tackled the influence of this treatment on resistin, another adipocytokine involved in obesity-related insulin resistance. Leptin-treated (T) obese mice (continuous sc infusion for 6 days) were compared with untreated lean (L), untreated obese (O), and untreated pair-fed obese (PF) mice. Blood was collected throughout the study. At day 3 or day 6, fat pads were either directly analyzed (mRNA, ApN content) or cultured for up to 24 h (ApN secretion). The direct effect of leptin was also studied in 3T3-F442A adipocytes. Compared with L mice, ApN content of visceral or subcutaneous fat and ApN secretion by adipose explants were blunted in obese mice. Accordingly, plasma ApN levels of O mice were decreased by 50%. Leptin treatment of ob/ob mice increased ApN mRNAs, ApN content, and secretion from the visceral depot by 50–80%. Leptin also directly stimulated ApN mRNAs and secretion from 3T3-F442A adipocytes. After 6 days of treatment, plasma ApN of ob/ob mice increased 2.5-fold, a rise that did not occur in PF mice. Plasma resistin of T mice was barely decreased. Leptin treatment, but not mere calorie restriction, corrects plasma ApN in obese mice by restoring adipose tissue ApN concentrations and secretion, at least in part, via a direct stimulation of ApN gene expression. Such a treatment only minimally affects circulating resistin. ApN restoration could, in concert with leptin, contribute to the metabolic effects classically observed during leptin administration.
Pregnancy impairs the counterregulatory response to insulin-induced hypoglycemia in the dog
Authors: Connolly Cynthia C. , Aglione Lisa N. , Smith Marta S. , Lacy D. Brooks , Moore Mary Courtney
Published: May 2004

The impact of pregnancy on the counterregulatory response to insulin-induced hypoglycemia was examined in six nonpregnant (NP) and six pregnant (P; 3rd trimester) conscious dogs by tracer and arteriovenous difference techniques. After basal sampling, insulin was infused intraportally at 30 pmol·kg−1·min−1 for 180 min. Insulin rose from 70 ± 15 to 1,586 ± 221 pmol/l and 27 ± 4 to 1,247 ± 61 pmol/l in the 3rd h in NP and P, respectively. Arterial glucose fell from 5.9 ± 0.2 to 2.3 ± 0.2 mmol/l in P. Glucose was infused in NP to equate the rate of fall of glucose and the steady-state concentrations in the groups (5.9 ± 0.2 to 2.3 ± 0.1 mmol/l in NP). Glucagon was 32 ± 6, 69 ± 11, and 48 ± 10 ng/l (basal and 1st and 3rd h) in NP, but the response was attenuated in P (34 ± 5, 46 ± 6, 41 ± 9 ng/l). Cortisol and epinephrine rose similarly in both groups, but norepinephrine rose more in NP (Δ3.01 ± 0.46 and Δ1.31 ± 0.13 nmol/l, P < 0.05). Net hepatic glucose output (NHGO; μmol·kg−1·min−1) increased from 10.6 ± 1.8 to 21.2 ± 3.3 in NP (3rd h) but did not increase in P (15.1 ± 1.5 to 15.3 ± 2.8 μmol·kg−1·min−1, P < 0.05 between groups). The glycogenolytic contribution to NHGO in NP increased from 5.8 ± 0.7 to 10.4 ± 2.5 μmol·kg−1·min−1 by 90 min but steadily declined in P. The increase in glycerol levels and the gluconeogenic contribution to NHGO were 50% less in P than in NP, but ketogenesis did not differ. The glucagon and norepinephrine responses to insulin-induced hypoglycemia are blunted in late pregnancy in the dog, impacting on the magnitude of the metabolic responses to the fall in glucose.
Seeing the trees in the forest: selective electroporation of adipocytes within adipose tissue
Authors: Granneman James G. , Li Pipeng , Lu Yuyan , Tilak Jacqueline
Published: May 2004

Electroporation has been recently adapted for the transfer of macromolecules into cells of tissues in vivo. Although mature adipocytes constitute <20% of cells residing in adipose tissue, we hypothesized that fat cells might be susceptible to selective electrotransfer of plasmid DNA owing to their large size relative to other cells in the tissue. Results demonstrate the feasibility of electroporating DNA into mature fat cells with >99% selectivity over other cells in the tissue. Further experiments used the “adiporation” technique to image the subcellular targeting of fluorescent bioreporter molecules to the nucleus, mitochondria, and lipid droplets of adipocytes within intact adipose tissue. Finally, we utilized fluorescent bioreporters to examine the effects of constitutive activation of the β-adrenergic signaling pathway in adipocytes. These results demonstrate that overexpression of rat β1-adrenergic receptors alters the cellular morphology of white adipocytes in a fashion that mimics the effects of systemic infusion of β3-adrenergic receptor agonists. Hallmarks of the altered morphology include pronounced fragmentation of the single lipid droplet, repositioning of the nucleus, and induction of mitochondrial biogenesis. These results indicate that activation of β-adrenergic signaling within adipocytes is sufficient to induce a phenotype that resembles typical brown adipocytes and suggest that in vivo electroporation will allow molecular dissection of the mechanisms involved.
Correction of motion artifact in transmembrane voltage-sensitive fluorescent dye emission in hearts
Authors: Tai Dean C.-S. , Caldwell Bryan J. , LeGrice Ian J. , Hooks Darren A. , Pullan Andrew J. , Smaill Bruce H.
Published: May 2004

Fast voltage-sensitive dyes are widely used to image cardiac electrical activity. Typically, the emission spectrum of these fluorochromes is wavelength shifted with altered membrane potential, but the optical signals obtained also decay with time and are affected by contraction. Ratiometry reduces, but may not fully remove, these artifacts. An alternate approach has been developed in which the time decay in simultaneously acquired short- and long-wavelength signals is characterized nonparametrically and removed. Motion artifact is then identified as the time-varying signal component common to both decay-corrected signals and subtracted. Performance of this subtraction technique was compared with ratiometry for intramural optical signals acquired with a fiber-optic probe in an isolated, Langendorff-perfused pig heart preparation ( n = 4) stained with di-4-ANEPPS. Perfusate concentration of 2,3-butanedione monoxime was adjusted (7.5–12.5 mM) to alter contractile activity. Short-wavelength (520–600 nm) and long-wavelength (>600 nm) signals were recorded over 8–16 cardiac cycles at 6 sites across the left ventricular free wall in sinus rhythm and during pacing. A total of 451 such data sets were acquired. Appreciable wall motion was observed in 225 cases, with motion artifact classed as moderate (less than modulation due to action potential) in 187 and substantial (more than modulation due to action potential) in 38. In all cases, subtraction performed as well as, or better than, ratiometry in removing motion artifact and decay. Action potential morphology was recovered more faithfully by subtraction than by ratiometry in 58 of 187 and 31 of 38 cases with moderate and substantial motion artifact, respectively. This novel subtraction approach may therefore provide a means of reducing the concentration of uncoupling agents used in cardiac optical mapping studies.
Salmeterol, a β2-receptor agonist, attenuates lipopolysaccharide-induced lung inflammation in mice
Authors: Maris Nico A. , van der Sluijs Koenraad F. , Florquin Sandrine , de Vos Alex F. , Pater Jennie M. , Jansen Henk M. , van der Poll Tom
Published: May 2004

Lipopolysaccharide is ubiquitously present in the environment. To determine the effect of salmeterol, a long-acting β2-receptor agonist, on lipopolysaccharide-induced lung inflammation, mice received lipopolysaccharide (10 μg) intranasally with or without salmeterol intraperitoneally (5 mg/kg) 30 min earlier and 12 h thereafter. Salmeterol dose- and time-dependently inhibited the lipopolysaccharide-induced influx of neutrophils into bronchoalveolar lavage fluid and lung tissue, and these pulmonary neutrophils displayed a reduced expression of CD11b at their surface. To determine the contribution of the salmeterol effect on neutrophil CD11b in the attenuated neutrophil recruitment, we treated mice intranasally exposed to lipopolysaccharide with salmeterol with or without a blocking anti-CD11b antibody. Anti-CD11b profoundly reduced lipopolysaccharide-induced neutrophil influx in bronchoalveolar lavage fluid, an effect that was modestly enhanced by concurrent salmeterol treatment. These data suggest that salmeterol inhibits lipopolysaccharide-induced neutrophil recruitment to the lungs by a mechanism that possibly in part is mediated by an effect on neutrophil CD11b.
Effects of vascular endothelial growth factor on isolated fetal alveolar type II cells
Authors: Raoul William , Chailley-Heu Bernadette , Barlier-Mur Anne-Marie , Delacourt Christophe , Maître Bernard , Bourbon Jacques R.
Published: May 2004

Previous investigations gained from in vivo or lung explant studies suggested that VEGF is an autocrine proliferation and maturation factor for developing alveolar type II cells. The objective of this work was to determine whether VEGF exerted its growth and maturation effects directly on isolated type II cells. These were isolated from 19-day fetal rat lung and cultured in defined medium. The presence of VEGF receptor-2 was assessed in cultured cells at the pre- and posttranslational levels. Recombinant VEGF165, formerly found to be active on lung explants, failed to enhance type II cell proliferation estimated by thymidine and 5-bromo-2′-deoxy-uridine incorporation. It increased choline incorporation in saturated phosphatidylcholine by 27% but did not increase phospholipid surfactant pool size. VEGF (100 ng/ml) left unchanged the transcript level of surfactant proteins (SP)-A, SP-C, and SP-D but increased SP-B transcripts to four times the control steady-state level. VEGF slightly retarded, but did not prevent, the in vitro transdifferentiation of type II into type I cells, as assessed by immunolabeling of the type I cell marker T1α. We conclude that, with the exception of SP-B expression, which appears to be controlled directly, the previously observed effects of this VEGF isoform on type II cells are likely to be exerted indirectly through reciprocal paracrine interactions involving other lung cell types.
Matrix GLA protein modulates branching morphogenesis in fetal rat lung
Authors: Gilbert Kirk A. , Rannels Stephen R.
Published: May 2004

The regulation of matrix γ-carboxyglutamic acid protein (MGP) expression during the process of lung branching morphogenesis and development was investigated. MGP mRNA expression was determined over an embryonic and postnatal time course and shown to be developmentally regulated. Immunohistochemical analysis revealed increased staining for MGP in peripheral mesenchyme surrounding distal epithelial tubules. Fetal lung explants were used as an in vitro growth model to examine expression and regulation of MGP during branching morphogenesis. MGP mRNA expression over the culture interval mimicked the in vivo time course. Explants cultured in the presence of antibodies against MGP showed gross dilation and reduced terminal lung bud counts, accompanied by changes in MGP, sonic hedgehog, and patched mRNA expression. Similarly, antifibronectin antibody treatment resulted in explant dilation and reduced MGP expression, providing evidence for an interaction with MGP and fibronectin. Conversely, intraluminal microinjection of anti-MGP antibodies had no effect either on explant growth or MGP expression, supporting the hypothesis that MGP exerts its effects through the mesenchyme. Taken together, the results suggest that MGP plays a role in lung growth and development, likely via temporally and spatially specific interactions with other branching morphogenesis-related proteins to influence growth processes.
Greater vascularity, lowered HIF-1/DNA binding, and elevated GSH as markers of adaptation to in vivo chronic hypoxia
Authors: Tissot van Patot M. C. , Bendrick-Peart J. , Beckey V. E. , Serkova N. , Zwerdlinger L.
Published: May 2004

Vascularity is increased in placentas from high- compared with low-altitude pregnancies. An angiogenic response to hypoxia may protect an organ from further hypoxic insult by increasing blood flow and oxygen delivery to the tissue. We hypothesized that increased placental vascularity is sufficient to adapt to high altitude. Therefore, indexes of hypoxic stress would not be present in placentas from successful high-altitude pregnancies. Full-thickness placental biopsies were 1) collected and frozen in liquid nitrogen within 5 min of placental delivery and 2) fixed in formalin for stereologic analyses at high (3,100 m, n = 10) and low (1,600 m, n = 10) altitude. Hypoxia-inducible transcription factor (HIF-1) activity was analyzed by ELISA. Western blot analyses were used to evaluate HIF-1α, HIF-1β, HIF-2α, von Hippel-Lindau protein, VEGF, Flt-1, enolase, and GAPDH. Magnetic resonance spectroscopy was used to evaluate endogenous metabolism. The ratio of placental capillary surface density to villous surface density was 70% greater at high compared with low altitude. HIF-1 activity and HIF-1-associated proteins were unchanged in placentas from high- vs. low-altitude pregnancies. Placental expression of HIF-1-mediated proteins VEGF, Flt-1, enolase, and GAPDH were unchanged at high vs. low altitude. Succinate, GSH, phosphomonoesters, and ADP were elevated in placenta from high compared with low altitude. Placentas from uncomplicated high-altitude pregnancies have greater vascularity and no indication of significant hypoxic stress at term compared with placentas from low altitude.
5-HT induces an arachidonic acid-sensitive calcium influx in rat small intrapulmonary artery
Authors: Guibert Christelle , Marthan Roger , Savineau Jean-Pierre
Published: May 2004

5-Hydroxytryptamine (5-HT) is a potent pulmonary vasoconstrictor and contributes to hypoxic pulmonary vasoconstriction and pulmonary arterial hypertension. Small intrapulmonary vessels are very sensitive to hypoxia and play a major role for blood flow regulation in the lung. Thus we have investigated the mechanisms involved in the calcium signal to 5-HT in rat small intrapulmonary artery (IPA). Effects of 5-HT were examined in isolated IPA (external diameter <250 μm) from rat. Digital imaging with fura-PE3 was used to record intracellular calcium concentration ([Ca2+]i) and to follow external diameter of the vessels. 5-HT induced a sustained [Ca2+]i variation that was sensitive to the inhibitor of the 5-HT2A receptors, ketanserin, and insensitive to voltage-dependent L-type calcium channel blockers (nitrendipine and nicardipine) or voltage-independent calcium channel antagonists (LOE-908, SKF-96365, and gadolinium). The calcium response to 5-HT was also not modified by a sarcoplasmic reticulum Ca2+-ATPase inhibitor (cyclopiazonic acid; CPA), which depletes intracellular calcium stores. CPA alone activated a capacitative calcium channel that was sensitive to LOE-908 and insensitive to SKF-96365 and gadolinium. The sustained calcium signal to 5-HT was partly blocked by inhibitors of arachidonic acid production (RHC-80267 and isotetrandrine) and mimicked by application of exogenous arachidonic acid. These results suggest that activation of a noncapacitative, arachidonic acid-sensitive, receptor-operated calcium channel contributes to 5-HT-induced sustained calcium increase in small IPA.
Increased pulmonary blood flow does not alter surfactant protein gene expression in lambs within the first week of life
Authors: Lee Jae W. , Ovadia Boaz , Azakie Anthony , Salas Sonia , Goerke Jon , Fineman Jeffrey R. , Gutierrez Jorge A.
Published: May 2004

Neonates and infants with congenital heart disease with increased pulmonary blood flow suffer morbidity from poor oxygenation and decreased lung compliance. In a previous experiment involving 4-wk-old lambs with pulmonary hypertension secondary to increased pulmonary blood flow following an in utero placement of an aortopulmonary vascular graft, we found a decrease in surfactant protein (SP)-A gene expression as well as a decrease in SP-A and SP-B protein contents. To determine the timing of these changes, the objective of the present study was to characterize the effect of increased pulmonary blood flow and pulmonary hypertension on SP-A, -B, and -C gene expressions and protein contents within the first week of life. Of eight fetal lambs that underwent the in utero placement of the shunt, there was no difference in the expression of SP-A, -B, and -C mRNA levels or SP-A and -B protein contents compared with age-matched controls. The results showed that, in this model of congenital heart disease with pulmonary hypertension and increased pulmonary blood flow, the effect of the shunt on SP gene expression and protein content was not apparent within the first week of life.
Proinflammatory activity of anti-IL-8 autoantibody:IL-8 complexes in alveolar edema fluid from patients with acute lung injury
Authors: Krupa Agnieszka , Kato Hiroyuki , Matthay Michael A. , Kurdowska Anna K.
Published: May 2004

A significant fraction of IL-8 in lung fluids from patients with the acute lung injury (ALI) is associated with anti-IL-8 autoantibodies (anti-IL-8:IL-8 complexes), and lung fluid concentrations of these complexes correlate with development and outcome of ALI. In this study, we examined whether anti-IL-8:IL-8 complexes exhibit proinflammatory activity in vitro. These complexes were purified from pulmonary edema fluid samples obtained from patients with ALI. First, we found that IL-8 bound to the autoantibody retained its ability to trigger chemotaxis of neutrophils, whereas control antibody did not have significant chemotactic activity. Next, we examined the ability of anti-IL-8:IL-8 complexes to induce neutrophil activation, i.e., neutrophil respiratory burst and degranulation. Anti-IL-8:IL-8 complexes triggered superoxide and myeloperoxidase release from human neutrophils, and in contrast, the control antibody had no effect. We also demonstrated that IgG receptor, FcγRIIa, is the receptor involved in cellular activation mediated by these complexes. Blockade of FcγRIIa completely reverses activity of the complexes with the exception of chemotaxis. Both FcγRIIa and IL-8 receptors mediate chemotactic activity of anti-IL-8:IL-8 complexes, with FcγRIIa being, however, a predominant receptor. Furthermore, activity of the complexes is partially dependent on the activation of the mitogen-activated protein kinases, i.e., ERK and p38, important components of the FcγRIIa signaling cascade. Anti-IL-8:IL-8 complexes may therefore be involved in the pathogenesis of lung inflammation in clinical acute lung injury.
Enhancement of the endotoxin recognition pathway by ventilation with a large tidal volume in rabbits
Authors: Moriyama Kiyoshi , Ishizaka Akitoshi , Nakamura Morio , Kubo Hiroshi , Kotani Toru , Yamamoto Shinichi , Ogawa Eileen N. , Kajikawa Osamu , Frevert Charles W. , Kotake Yoshifumi , Morisaki Hiroshi , Koh Hidefumi , Tasaka Sadatomo , Martin Thomas R. , Takeda Junzo
Published: May 2004

Ventilation with a small tidal volume (Vt) is associated with better clinical outcomes than with a large Vt, particularly in critical settings, including acute lung injury. To determine whether Vtinfluences the lipopolysaccaharide (LPS) recognition pathway, we studied CD14 expression in rabbit lungs and the release of TNF-α by cultured alveolar macrophages after 240 min of ventilation with a large (20 ml/kg) vs. a small (5 ml/kg) Vt. We also applied small or large Vtto lungs instilled with 50 μg/kg of LPS. The alveolar macrophages collected after large Vtventilation revealed a 20-fold increase in LPS-induced TNF-α release compared with those collected after small Vtventilation, whereas TNF-α was undetectable without LPS stimulation. In animals ventilated with a large Vt, the expression of CD14 mRNA in whole lung homogenates and the expression of CD14 protein on alveolar macrophages, assessed by immunohistochemistry, were both significantly increased in the absence of LPS stimulation. A large Vtapplied to LPS-instilled lungs increased the pulmonary albumin permeability and TNF-α release into the plasma. These results suggest that mechanical stress caused by a large Vtsensitizes the lungs to endotoxin, a phenomenon that may occur partially via the upregulation of CD14.
Electron microscopic study of actin polymerization in airway smooth muscle
Authors: Herrera Ana M. , Martinez Eliana C. , Seow Chun Y.
Published: May 2004

Actin polymerization as part of the normal smooth muscle response to various stimuli has been reported. The actin dynamics are believed to be necessary for cytoskeletal remodeling in smooth muscle in its adaptation to external stress and strain and for maintenance of optimal contractility. We have shown in our previous studies in airway smooth muscle that myosins polymerized in response to contractile activation as well as to adaptation at longer cell lengths. We postulated that the same response could be elicited from actins under the same conditions. In the present study, actin filament formation was quantified electron microscopically in cell cross sections. Nanometer resolution allowed us to examine regional distribution of filaments in a cell cross section. Airway smooth muscle bundles were fixed in relaxed and activated states at two lengths; muscle preparations were also fixed after a period of oscillatory strain, a condition known to cause depolymerization of myosin filaments. The results indicate that contractile activation and increased cell length nonsynergistically enhanced actin polymerization; the extent of actin polymerization was substantially less than that of myosin polymerization. Oscillatory strain increased thin filament formation. Although thin filament density was found higher in cytoplasmic areas near dense bodies, contractile activation did not preferentially enhance actin polymerization in these areas. It is concluded that actin thin filaments are dynamic structures whose length and number are regulated by the cell in response to changes in extracellular environment and that polymerization and depolymerization of thin filaments occur uniformly across the whole cell cross section.
Proteomic analysis of pulmonary edema fluid and plasma in patients with acute lung injury
Authors: Bowler Russell P. , Duda Beth , Chan Edward D. , Enghild Jan J. , Ware Lorraine B. , Matthay Michael A. , Duncan Mark W.
Published: May 2004

Proteomics is the large-scale analysis of protein profiles. This approach has not yet been reported in the study of acute lung injury (ALI). This study details protein profiles in plasma and pulmonary edema fluid (EF) from 16 ALI patients and plasma and bronchoalveolar lavage fluid (BALF) from 12 normal subjects. More than 300 distinct protein spots were evident in the EF and BALF of both normal subjects and ALI patients. Of these, 158 were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. In the plasma and EF protein profile of ALI patients, there were multiple qualitative changes. For instance, in all normal subjects, but in only one of the ALI patients, seven distinct surfactant protein A isoforms were evident. Nearly all ALI patients also had protein spots that indicate truncation or other posttranslational modifications. Several of these novel changes could serve as new biomarkers of lung injury.
VE-cadherin-p120 interaction is required for maintenance of endothelial barrier function
Authors: Iyer Seema , Ferreri Deana M. , DeCocco Nina C. , Minnear Fred L. , Vincent Peter A.
Published: May 2004

Interaction of p120 with juxtamembrane domain (JMD) of VE-cadherin has been implicated in regulation of endothelial cell-cell adhesion. We used a number of approaches to alter the level of p120 available for binding to VE-cadherin as a means to investigate the role of p120-VE-cadherin interaction in regulation of barrier function in confluent endothelial monolayers. Expression of an epitope-tagged fragment corresponding to JMD of VE-cadherin resulted in a decrease in endothelial barrier function as assessed by changes in albumin clearance and electrical resistance. Binding of JMD-Flag to p120 resulted in a decreased level of p120. In addition to decreasing p120 level, expression of JMD also decreased level of VE-cadherin. Expression of JMD also caused an increase in MLC phosphorylation and rearrangement of actin cytoskeleton, which, coupled with decreased cadherin, can contribute to loss of barrier function. Reducing p120 by siRNA resulted in a decrease in VE-cadherin, whereas increasing the level of p120 increased the level of VE-cadherin, demonstrating that p120 regulates the level of VE-cadherin. Overexpression of p120 was, however, associated with decreased barrier function and rearrangement of the actin cytoskeleton. Interestingly, expression of p120 was able to inhibit thrombin-induced increases in MLC phosphorylation, suggesting that p120 inhibits activation of Rho/Rho kinase pathway in endothelial cells. Excess p120 also prevented JMD-induced increases in MLC phosphorylation, correlating this phosphorylation with Rho/Rho kinase pathway. These findings show p120 plays a major role in regulating endothelial barrier function, as either a decrease or increase of p120 resulted in disruption of permeability across cell monolayers.
Role of autoimmunity in organ allograft rejection: a focus on immunity to type V collagen in the pathogenesis of lung transplant rejection
Authors: Sumpter Tina L. , Wilkes David S.
Published: May 2004

Lung transplantation is the only definitive treatment modality for many forms of end-stage lung disease. However, the lung is rejected more often than any other type of solid organ allograft due to chronic rejection known as bronchiolitis obliterans (BO). Indeed, BO is the primary reason why the 5- and 7-yr survival rates are worse for the lung than for any other transplanted organ. Alloimmunity to donor antigens is established as the primary mechanism that mediates rejection responses. However, newer immunosuppressive regimens designed to abrogate alloimmune activation have not improved survival. Therefore, these data suggest that other antigens, unrelated to donor transplantation antigens, are involved in rejection. Utilizing human and rodent studies of lung transplantation, our laboratory has documented that a native collagen, type V collagen [col(V)], is a target of the rejection response. Col(V) is highly conserved; therefore, these data indicate that transplant rejection involves both alloimmune and autoimmune responses. The role of col(V) in lung transplant rejection is described in this review article. In addition, the potential role of regulatory T cells that are crucial to modulating autoimmunity and alloimmunity is also discussed.
Activation of the STAT pathway in acute lung injury
Authors: Severgnini Mariano , Takahashi Satoe , Rozo Liliana M. , Homer Robert J. , Kuhn Charles , Jhung Jhung W. , Perides George , Steer Michael , Hassoun Paul M. , Fanburg Barry L. , Cochran Brent H. , Simon Amy R.
Published: May 2004

Acute lung injury (ALI) is a devastating clinical problem with a mortality as high as 60%. It is now appreciated that ALI represents a cytokine excess state that involves the microvasculature of multiple organs. The signal transducers and activators of transcription (STAT) family of transcription factors activate critical mediators of cytokine responses, but there is limited knowledge about their role in mediating ALI. In the present study, we demonstrate that the STAT transcription factors are activated rapidly in the lungs after intraperitoneal and intranasal LPS administration in mice. We also demonstrated that LPS activates both the STAT kinases, Src and JAK, in the lung with kinetics that are consistent with STAT activation. LPS treatment resulted in STAT3 activation throughout the resident lung cells, as well as in the recruited inflammatory cells. Whereas direct LPS treatment did not lead to STAT activation in cultured epithelial or endothelial cells, IL-6 activated STAT3 in both of these cell types. Furthermore, IL-6 was induced by LPS in serum and in the lung with kinetics consistent with STAT3 activation, suggesting that IL-6 may be one mechanism of STAT activation by LPS. In addition, STAT activation required reactive oxygen species, as the overexpression of catalase in mice prevented LPS-mediated STAT activation in the lung. STATs may be a common pathway for mediating ALI, regardless of the inciting factor, as STAT activation also occurred in both a gastric acid aspiration and acute pancreatitis model of ALI. Finally, STATs are activated in the lung long before signs of ALI are present, suggesting that the STAT transcription factors may play a role in initiating the inflammatory response seen in the lung.
Alveolar macrophages from systemic sclerosis patients: evidence for IL-4-mediated phenotype changes
Authors: Hamilton Raymond F. , Parsley Ed , Holian Andrij
Published: May 2004

The mechanism of chronic lung inflammation leading to lung fibrosis is unknown and does not have a characteristic inflammatory macrophage phenotype. This study was undertaken to determine whether a change in macrophage phenotype could account for chronic lung inflammation. In this study, human alveolar macrophages (AM) from subjects with systemic sclerosis (SSc) were obtained from bronchoalveolar lavage (BAL) and characterized on the basis of function (response to LPS), phenotype, and relative cell-surface B7 expression. AM from the subjects' disease-involved and noninvolved lung lobes were compared with each other and to AM from normal volunteer BAL. AM from involved SSc lobes produced significantly more interleukin (IL)-1β and PGE2than AM from uninvolved lobes in response to LPS, but there was no spontaneous production of either mediator. The activator AM phenotype designated by RFD1+ surface epitope was significantly elevated in SSc BAL samples compared with normal BAL, although there were no differences comparing involved vs. noninvolved lobes within SSc subjects. The major histocompatibility complex II costimulatory molecule B7.2 was also significantly elevated in SSc AM compared with normal AM, again with no differences between involved and noninvolved lobes. In an attempt to determine environmental influences on AM phenotypes, normal AM were cultured in vitro with IFN-γ, IL-3, IL-4, IL-10, IL-12, or dexamethasone for 6 days. Of the cytokines examined, only IL-4 induced significant increases in both the activator phenotype RFD1+ and B7.2 expression. Taken together, these results indicate that IL-4 could account for proinflammatory AM phenotype changes and B7 surface-marker shifts, as seen in subjects with SSc.
Interfacial phospholipids inhibit ozone-reactive absorption-mediated cytotoxicity in vitro
Authors: Connor Lydia M. , Ballinger Carol A. , Albrecht Thomas B. , Postlethwait Edward M.
Published: May 2004

The intrapulmonary distribution of inhaled ozone (O3) and induction of site-specific cell injury are related to complex interactions among airflow patterns, local gas-phase concentrations, and the rates of O3 flux into, and reaction and diffusion within, the epithelial lining fluid (ELF). Recent studies demonstrated that interfacial phospholipid films appreciably inhibited NO2 absorption. Because surface-active phospholipids are present on alveolar and airway interfaces, we investigated the effects of interfacial films on O3-reactive absorption and acute cell injury. Compressed films of dipalmitoyl-glycero-3-phosphocholine (DPPC) and rat lung lavage lipids significantly reduced O3-reactive absorption by ascorbic acid, reduced glutathione, and uric acid. Conversely, unsaturated phosphatidylcholine films did not inhibit O3 absorption. We evaluated O3-mediated cell injury using a human lung fibroblast cell culture system, an intermittent tilting exposure regimen to produce a thin covering layer, and nuclear fluorochrome permeability. Exposure produced negligible injury in cells covered with MEM. However, addition of AH2 produced appreciable (<50%) cell injury. Film spreading of DPPC monolayers necessitated the use of untilted regimens. Induction of acute cell injury in untilted cultures required both AH2 plus very high O3 concentrations. Addition of DPPC films significantly reduced cell injury. We conclude that acute cell injury likely results from O3 reaction with ELF substrates. Furthermore, interfacial films of surface-active, saturated phospholipids reduce the local dose of O3-derived reaction products. Finally, because O3 local dose and tissue damage likely correlate, we propose that interfacial phospholipids may modulate intrapulmonary distribution of inhaled O3 and the extent of site-specific cell injury.
Hypoxia-induced inhibition of whole cell membrane currents and ion transport of A549 cells
Authors: Karle Christoph , Gehrig Tobias , Wodopia Ralf , Höschele Sabine , Kreye Volker A. W. , Katus Hugo A. , Bärtsch Peter , Mairbäurl Heimo
Published: May 2004

In excitable cells, hypoxia inhibits K channels, causes membrane depolarization, and initiates complex adaptive mechanisms. It is unclear whether K channels of alveolar epithelial cells reveal a similar response to hypoxia. A549 cells were exposed to hypoxia during whole cell patch-clamp measurements. Hypoxia reversibly inhibited a voltage-dependent outward current, consistent with a K current, because tetraethylamonium (TEA; 10 mM) abolished this effect; however, iberiotoxin (0.1 μM) does not. In normoxia, TEA and iberiotoxin inhibited whole cell current (−35%), whereas the K-channel inhibitors glibenclamide (1 μM), barium (1 mM), chromanol B293 (10 μM), and 4-aminopyridine (1 mM) were ineffective. 86Rb uptake was measured to see whether K-channel modulation also affected transport activity. TEA, iberiotoxin, and 4-h hypoxia (1.5% O2) inhibited total 86Rb uptake by 40, 20, and 35%, respectively. Increased extracellular K also inhibited 86Rb uptake in a dose-dependent way. The K-channel opener 1-ethyl-2-benzimidazolinone (1 mM) increased 86Rb uptake by 120% in normoxic and hypoxic cells by activation of Na-K pumps (+60%) and Na-K-2Cl cotransport (+170%). However, hypoxic transport inhibition was also seen in the presence of 1-ethyl-2-benzimidazolinone, TEA, and iberiotoxin. These results indicate that hypoxia, membrane depolarization, and K-channel inhibition decrease whole cell membrane currents and transport activity. It appears, therefore, that a hypoxia-induced change in membrane conductance and membrane potential might be a link between hypoxia and alveolar ion transport inhibition.
Signal transduction events involved in TPA downregulation of SP-A gene expression
Authors: Miakotina Olga L. , Snyder Jeanne M.
Published: May 2004

Surfactant protein A (SP-A), the most abundant pulmonary surfactant protein, plays a role in innate host defense and blocks the inhibitory effects of serum proteins on surfactant surface tension-lowering properties. SP-A mRNA and protein are downregulated by phorbol esters (TPA) via inhibition of gene transcription. We evaluated the TPA signaling pathways involved in SP-A inhibition in a lung cell line, H441 cells. TPA caused sustained phosphorylation of p44/42 mitogen-activated protein kinase (MAPK), p38 MAPK, and c-Jun-NH2-terminal kinase. An inhibitor of conventional and novel isoforms of protein kinase C (PKC) and two inhibitors of p44/42 MAPK kinase partially or completely blocked the inhibitory effects of TPA on SP-A mRNA levels. In contrast, inhibitors of conventional PKC-α and -β, stress-activated protein kinases, protein phosphatases, protein kinase A, and the phosphatidylinositol 3-kinase pathway had no effect on the TPA-mediated inhibition of SP-A mRNA. TPA also stimulated the synthesis of c-Jun mRNA and protein in a time-dependent manner. Inhibitors of the p44/42 MAPK signaling pathway and PKC blocked the TPA-mediated phosphorylation of p44/42 MAPK and the increase in c-Jun mRNA. We conclude that TPA inhibits SP-A gene expression via novel isoforms of PKC, the p44/42 MAPK pathway, and the activator protein-1 complex.
Elevation of KL-6, a lung epithelial cell marker, in plasma and epithelial lining fluid in acute respiratory distress syndrome
Authors: Ishizaka Akitoshi , Matsuda Tomoyuki , Albertine Kurt H. , Koh Hidefumi , Tasaka Sadatomo , Hasegawa Naoki , Kohno Nobuoki , Kotani Toru , Morisaki Hiroshi , Takeda Junzo , Nakamura Morio , Fang Xiaohui , Martin Thomas R. , Matthay Michael A. , Hashimoto Satoru
Published: May 2004

KL-6 is a pulmonary epithelial mucin more prominently expressed on the surface membrane of alveolar type II cells when these cells are proliferating, stimulated, and/or injured. We hypothesized that high levels of KL-6 in epithelial lining fluid and plasma would reflect the severity of lung injury in patients with acute lung injury (ALI). Epithelial lining fluid was obtained at onset ( day 0) and day 1 of acute respiratory distress syndrome (ARDS)/ALI by bronchoscopic microsampling procedure in 35 patients. On day 0, KL-6 and albumin concentrations in epithelial lining fluid were significantly higher than in normal controls ( P < 0.001), and the concentrations of KL-6 in epithelial lining fluid ( P < 0.002) and in plasma ( P < 0.0001) were higher in nonsurvivors than in survivors of ALI/ARDS. These observations were corroborated by the immunohistochemical localization of KL-6 protein expression in the lungs of nonsurvivors with ALI and KL-6 secretion from cultured human alveolar type II cells stimulated by proinflammatory cytokines. Because injury to distal lung epithelial cells, including alveolar type II cells, is important in the pathogenesis of ALI, the elevation of KL-6 concentrations in plasma and epithelial lining fluid could be valuable indicators for poor prognosis in clinical ALI.
Hepatoma-derived growth factor is a pulmonary endothelial cell-expressed angiogenic factor
Authors: Everett Allen D. , Narron Jill V. , Stoops Tamara , Nakamura Hideji , Tucker Amy
Published: May 2004

Hepatoma-derived growth factor (HDGF) was previously identified as a developmentally regulated cardiovascular and renal gene that is mitogenic for vascular smooth muscle and aortic endothelial cells. As reciprocal interactions of smooth muscle and endothelial cells are necessary for vascular formation, we examined whether HDGF plays a role in angiogenesis. According to immunohistochemistry, HDGF was highly expressed in endothelial cells of nonmuscularized, forming blood vessels of the fetal lung. HDGF was also expressed in endothelial cells of small (20 μm) mature arteries and veins. By Western immunoblotting, HDGF was highly expressed by human pulmonary microvascular endothelial cells in vitro. Adenoviral overexpression of HDGF was mitogenic for human pulmonary microvascular endothelial cells in serum-free medium, stimulating a 1.75-fold increase in bromodeoxyuridine (BrdU) uptake and a twofold increase in cell migration. With the chick chorioallantoic membrane (CAM), a biologic assay for angiogenesis, exogenous recombinant HDGF significantly stimulated blood vessel formation and a dose-dependent reorganization of cells within the CAM into a more compact, linear alignment reminiscent of tube formation. According to double immunostaining for endothelial cells with a transforming growth factor-βII receptor antibody and BrdU as a marker of cell proliferation, exogenous HDGF selectively stimulated endothelial cell BrdU uptake. HDGF also activated specific ERK1/2 signaling and did not overlap with VEGF SAPK/JNK, Akt-mediated pathways. We conclude that HDGF is a highly expressed vascular endothelial cell protein in vivo and is a potent endothelial mitogen and regulator of endothelial cell migration by mechanisms distinct from VEGF.