Biomedical subjects

H Lum

Publications and source records attributed to H Lum.

At least 37 records · Page 2Linked to original sources

PubMed1994

Thrombin receptor activation peptide induces pulmonary vasoconstriction.

We investigated the involvement of the 14-residue thrombin receptor activating peptide SFLLRNPNDKYEPF (TRAP-14) in mediating the pulmonary vasoconstriction in response to alpha-thrombin. Isolated guinea pig lungs were uniformly perfused with Ringer-albumin solution at a constant flow of 28 ml/min. Addition of TRAP-14 or human alpha-thrombin to the perfusate caused dose-dependent increases of pulmonary arterial pressure within 1 min. TRAP-14 at 1 microM increased pulmonary arterial pressure to a similar extent as 10 nM alpha-thrombin (i.e., increase of 7.7 +/- 0.8 and 7.4 +/- 0.9 cmH2(0) from baseline, respectively). The increases in pulmonary venous resistance induced by TRAP-14 and alpha-thrombin were two- to fivefold greater than the increases in pulmonary arterial resistance, indicating that both agonists mediated pulmonary hypertension secondary to pulmonary venoconstriction. Stimulation of cultured guinea pig pulmonary artery smooth muscle cells with 100 microM TRAP-14 or 10 nM alpha-thrombin increased cytosolic Ca2+ concentration about five- to sevenfold over baseline. The increase in cytosolic Ca2+ concentration in smooth muscle cells was not observed with a subsequent challenge with either agonist, indicating desensitization. In the perfused lungs, an initial stimulation with alpha-thrombin or TRAP-14 desensitized the lungs to either agonist. The alpha-thrombin-desensitized lungs remained refractile to alpha-thrombin after 1 h of perfusion with fresh Ringer solution, whereas the TRAP-14-desensitized lungs recovered 79% of the vasoconstrictor response by 10 min and 93% of the response by 30 min.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1994

Regulation of vascular endothelial barrier function.

The increase in endothelial permeability in response to inflammatory mediators such as alpha-thrombin and histamine is accompanied by cell rounding and interendothelial gap formation, implicating that the predominant transport pathway is a diffusive one [i.e., via cellular junctions (paracellular transport)]. However, the possible contribution by vesicle-mediated transport (i.e., via albumin binding protein gp60) to the overall permeability increase needs investigation. Regulation of paracellular transport in endothelial cells is associated with modulation of actin-based systems which anchor the cell to its neighbor or extracellular matrix, thus maintaining endothelial integrity. At the cell-cell junctions, actin is linked indirectly to the plasma membrane by linking proteins (e.g., vinculin, catenins, alpha-actinin) to cadherins, which function in homophilic intercellular adhesion. Cadherins may also play a role in regulating the formation of tight junctions, which also may be associated with actin. At endothelial focal contacts, the transmembrane receptors (integrins) for matrix proteins are linked to actin via linking proteins (i.e., vinculin, talin, alpha-actinin). In response to inflammatory mediators, second messengers signal two regulatory pathways which modulate the actin-based systems, which may lead to impairment of the endothelial barrier integrity. One pathway is based on protein kinase C (PKC) isozyme-specific phosphorylation of linking proteins at the cell-cell and cell-matrix junctions. The increased phosphorylation is associated with actin reorganization, cell rounding, and increased paracellular transport. The other is the activation of myosin light-chain kinase, (MLCK), which causes an actin-myosin-based contraction that may lead to a centripetal retraction of endothelial cells. Current research is in the identification of protein substrates of PKC isozymes, the specific role of their phosphorylation in barrier function, and determining the precise role of MLCK in modulation of endothelial barrier function.

Animals↗

PubMed1993

Modeling vehicle accidents and highway geometric design relationships.

The statistical properties of four regression models--two conventional linear regression models and two Poisson regression models--are investigated in terms of their ability to model vehicle accidents and highway geometric design relationships. Potential limitations of these models pertaining to their underlying distributional assumptions, estimation procedures, functional form of accident rate, and sensitivity to short road sections, are identified. Important issues, such as the treatment of vehicle exposure and traffic conditions, and data uncertainties due to sampling and nonsampling errors, are also discussed. Roadway and truck accident data from the Highway Safety Information System (HSIS), a highway safety data base administered by the Federal Highway Administration (FHWA), have been employed to illustrate the use and the limitations of these models. It is demonstrated that the conventional linear regression models lack the distributional property to describe adequately random, discrete, nonnegative, and typically sporadic vehicle accident events on the road. As a result, these models are not appropriate to make probabilistic statements about vehicle accidents, and the test statistics derived from these models are questionable. The Poisson regression models, on the other hand, possess most of the desirable statistical properties in developing the relationships. However, if the vehicle accident data are found to be significantly overdispersed relative to its mean, then using the Poisson regression models may overstate or understate the likelihood of vehicle accidents on the road. More general probability distributions may have to be considered.

Accidents, Traffic↗

PubMed1993

Thrombin receptor peptide inhibits thrombin-induced increase in endothelial permeability by receptor desensitization.

Thrombin, a potent activator of cellular responses, proteolytically cleaves, and thereby activates its receptor. In the present study, we compared the effects of the thrombin receptor 14-amino acid peptide (TRP-14; SFLLRNPNDKYEPF), which comprises the NH2 terminus after cleavage of the thrombin receptor, and of the native alpha-thrombin on endothelial monolayer permeability. Addition of TRP-14 (1-200 microM) to bovine pulmonary artery endothelial cells increased [Ca2+]i in a dose-dependent manner. The peak increase in [Ca2+]i in response to 100 microM TRP-14 or 0.1 microM alpha-thrombin was similar (i.e., 931 +/- 74 nM and 1032 +/- 80 nM, respectively), which was followed by a slow decrease with t1/2 values of 0.73 and 0.61 min, respectively. Extracellular Ca2+ chelation with 5 mM EGTA abolished the sustained increases in [Ca2+]i induced by either TRP-14 or alpha-thrombin. alpha-thrombin (0.1 microM) increased transendothelial [125I]albumin permeability, whereas TRP-14 (1-100 microM) had no effect. Coincubation of 100 microM TRP-14 with 1 microM DIP-alpha-thrombin also did not increase permeability over control values. Stimulation of BPAEC with 0.1 microM alpha-thrombin induced translocation of protein kinase C (PKC) from the cytosol to the plasma membrane indicative of PKC activation, whereas TRP-14 had no effect at any concentration. TRP-14 at 100 microM desensitized BPAEC to thrombin-induced increases in [Ca2+]i and transendothelial permeability. The Ca2+ desensitization was reversed after approximately 60 min, and this recovery paralleled the recovery of the permeability response. These findings indicate that the TRP-14-induced Ca2+ mobilization in the absence of PKC activation is insufficient to increase endothelial permeability. In contrast, the increase in endothelial permeability after alpha-thrombin occurred in conjunction with Ca2+ mobilization as well as PKC activation. TRP-14 pretreatment prevented the alpha-thrombin-induced increase in endothelial permeability secondary to desensitization of the Ca2+ signal. The results suggest that combined cytosolic Ca2+ mobilization mediated by TRP-14 and PKC activation mediated by a TRP-14-independent pathway are dual signals responsible for the thrombin-induced increase in vascular endothelial permeability.

Amino Acid Sequence↗

PubMed1993

Human orbital fibroblasts in culture bind and respond to endothelin.

Human fibroblasts in primary cell culture were studied for their ability to bind to endothelin (ET), a 21-amino acid peptide with profound vasoconstricting properties. When 125I-labeled ET-1 was incubated with confluent orbital fibroblasts in the presence of increasing concentrations of unlabeled ligand, a single class of binding site was defined with a dissociation constant of 1.42 x 10(-8) M and a maximal binding capacity of 9.1 x 10(-10) mol/micrograms protein. ET-3 was a substantially less potent competitor for 125I-ET-1 binding sites than was unlabeled ET-1. Dermal fibroblasts demonstrated approximately 75% less ET-1 saturation binding activity, on a cellular protein basis, than did those from the orbit. Orbital fibroblasts responded to ET-1 (10(-9) M) with a rapid and transient increase in the free concentration of intracellular Ca2+ ([Ca2+]i) as assessed by monitoring acetoxymethyl ester of fura 2 fluorescence intensity. Rechallenge with the peptide elicited a substantially attenuated response than that seen after the initial treatment. There was no consistent effect of ET-1 on [Ca2+]i in dermal cultures. ET-3 failed to influence [Ca2+]i in either type of fibroblast. It would appear that orbital fibroblasts bind and respond to ET in a manner distinct from that observed in dermal fibroblasts, raising the possibility that the peptide may have site-specific actions in orbital connective tissue.

Calcium↗

PubMed1993

Albumin and Ricinus communis agglutinin decrease endothelial permeability via interactions with matrix.

We studied the effects of albumin and the lectin Ricinus communis agglutinin (RCA) on hydraulic conductivity (Lp) of bovine pulmonary microvascular endothelial cell monolayers (BPMVEC) because of the evidence that albumin and RCA can interfere with transendothelial albumin permeability (Siflinger-Birnboim, A., J. Schnitzer, H. Lum, F. Blumenstock, C. Shen, P. Del Vecchio, and A. Malik. J. Cell. Physiol. 149: 575-584, 1991). BPMVEC were seeded on microporous polycarbonate filters, and the liquid flux was measured by collecting effluent into a tubing of known inner diameter at transendothelial hydrostatic pressures (P) ranging from 5 to 20 cmH2O. Lp was calculated as the slope of the relationship of liquid flux per unit surface area (Jv) vs. P. Addition of RCA (50 micrograms/ml) or albumin (5 mg/ml) to the endothelial cell medium containing albumin-free Hanks' balanced saline solution (HBSS) decreased total Lp (expressed x 10(-6) cm.s-1 x cmH2O-1) from 17.2 +/- 3.6 during HBSS to 4.7 +/- 0.9 during albumin and 5.7 +/- 1.6 during RCA (P < 0.01 for both). The RCA effect, but not that of albumin, was prevented by the addition of D-galactose (0.1 M) (the cognate hapten monosaccharide of RCA). We determined the contribution of the extracellular matrix (ECM) in decreasing the Lp by obtaining ECM after treatment of the monolayers with 0.025 M NH4OH to detach endothelial cells from the ECM. Basal ECM Lp (expressed x 10(-6) cm.s-1 x cmH2O-1) was 57.0 +/- 15.3, and it decreased to 19.7 +/- 4.3 and 17.5 +/- 2.9 during RCA and albumin, respectively (P < 0.01 for both). In contrast, RCA and albumin did not alter the filter Lp values. Another lectin, Ulex europaeus agglutinin, and the protein immunoglobulin G had no effect on Lp values.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Fibrin contact increases endothelial permeability to albumin.

We studied the effects of contact of bovine pulmonary artery endothelial cell monolayers with fibrin on the endothelial barrier function. Fibrin formed by clotting purified fibrinogen (0.5 to 3.0 mg/ml) with alpha-thrombin (1 U/ml) was added to endothelial monolayers and permeability measurements were made after fibrin removal. Fibrin incubation for 3 hours resulted in 2- to 5-fold increases in transendothelial 125I-albumin permeability. Permeability returned to baseline value within 3 hours after fibrin removal. Direct contact with fibrin was necessary for the response, since fibrin separated from the endothelium did not increase permeability. Contact with agarose (2 mg/ml) or fibrinogen (0.5 to 3.0 mg/ml) also did not increase endothelial permeability. Transmission electron microscopic examination indicated normal appearance of interendothelial junctions at a time when albumin permeability was increased and no overt evidence of endothelial injury. Incubation of fibrin with endothelial monolayers at 4 degrees C prevented the increase in albumin permeability. We examined the possibility that increased albumin transcytosis was responsible for fibrin's effect using 14C-sucrose (Mr = 342D), a lipid insoluble tracer. Fibrin increased sucrose flux by 1.5-fold compared to 2- to 5-fold increases in albumin flux. The results indicate that fibrin contact with the endothelial cell increases endothelial permeability. The effect of fibrin may involve activation of temperature-sensitive bulk phase transcytosis of albumin.

Albumins↗

PubMed1992

Time course of thrombin-induced increase in endothelial permeability: relationship to Ca2+i and inositol polyphosphates.

The temporal relationship between the alpha-thrombin-induced increase in transendothelial permeability and the alpha-thrombin-mediated changes in several key transmembrane signaling events was examined in confluent monolayers of bovine pulmonary artery endothelial cells (BPAEC). The time courses of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] generation, changes in cytosolic [Ca2+] ([Ca2+]i), and reorganization of cytoskeletal F-actin were determined to assess the relationship between these events and the onset of the alpha-thrombin-induced increase in endothelial permeability. alpha-Thrombin (10(-7) M) increased the transendothelial 125I-albumin clearance rate half-maximally by approximately 1 min and maximally by approximately 2 min (160% over control level). The increase in permeability occurred concomitantly with reorganization of F-actin cytoskeleton (i.e., loss of peripheral band and increased stress fiber density) and increased actin polymerization. Stimulation of fura-2-loaded BPAEC with 10(-7) M alpha-thrombin produced a typical biphasic rise in [Ca2+]i. The initial rapid increase in [Ca2+]i peaked by approximately 16 s after thrombin challenge and the [Ca2+]i response showed a slow decrease to half-maximal within 50 s. The alpha-thrombin-induced increase in permeability as well as the increase in [Ca2+]i were consistently preceded by increased Ins(1,4,5)P3 generation detectable within 10 s after thrombin challenge. These results indicate that alpha-thrombin triggers a cascade of events (i.e., Ins(1,4,5)P3 generation and the ensuing rise in [Ca2+]i), which may comprise the second messengers that mediate F-actin reorganization and the increase in endothelial permeability.

Actins↗

PubMed1992

Thrombin receptor 14-amino acid peptide binds to endothelial cells and stimulates calcium transients.

We examined the binding characteristics of the recently described thrombin receptor amino-terminal peptide, SFLLRNPNDKYEPF (T. K. H. Vu, D. T. Hung, V. I. Wheaton, and S. R. Coughlin. Cell 64: 1057-1068, 1991), termed TRP-14, and its effect in activating intracellular calcium transients in pulmonary vascular endothelial cells. Binding of 125I-labeled TRP-14 was found to be saturable with a affinity constant of 2 microM and maximum binding of 41 pmol/mg of cell protein. The 125I-labeled TRP-14 also interacted with bovine pulmonary microvessel endothelial cells, human umbilical vein endothelial cells, and porcine pulmonary artery smooth muscle cells. Binding of 125I-labeled diisopropylphosphoryl (DIP)-alpha-thrombin, which is catalytically inactive but binds to thrombin receptors, was not inhibited by TRP-14 or vice versa, indicating that TRP-14 did not compete for the alpha-thrombin binding site(s) on the endothelial cell surface. TRP-14 (> 1 microM) increased the concentration of intracellular calcium ([Ca2+]i) in endothelial cells with kinetics similar to the increase in [Ca2+]i triggered by alpha-thrombin. In contrast, DIP-alpha-thrombin did not increase [Ca2+]i and also did not prevent the rise in [Ca2+]i induced by the subsequent challenge with either TRP-14 or alpha-thrombin. Because the generation of TRP-14 by the proteolytically active forms of thrombin stimulated a rise in endothelial [Ca2+]i, TRP-14 may be the agonist responsible for the activation of the alpha-thrombin receptor in pulmonary vascular endothelial cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1992

Non-predatory ingestive behaviors of the praying mantids Tenodera aridifolia sinensis (Sauss.) and Sphodromantis lineola (Burr.).

Praying mantids are thought to be so strictly predacious that, historically, carnivorousness has been used as a defining characteristics of the taxon Mantodea, and no data exist on other ingestive behaviors. We observed food- and water-deprived male and female Tenodera aridifolia sinensis (Sauss.) and Sphodromantis lineola (Burr.) in various situations and found that their ingestive behaviors are more variable than generally recognized. Both species regularly drink water in response to deprivation, the amount imbibed being correlated with the total (but not daily) percent of initial body weight lost. When presented with stimuli consisting of plastic beads of different reflective quality (shiny clear, opaque frosted, or matte black), lighted by a single direct light source, the shiny stimuli consistently elicited the most drinking-like behavior by the mantids. The preference was consistent, whether stimuli were presented together or singly. This suggests that visual cues can be used to identify water and are alone capable of maintaining drinking-like behavior. When S. lineola were presented with various stimuli, including diced apple and diced banana, in a five-way or a three-way choice test, mantids that chose a stimulus within the allotted time consistently ate the diced banana. When presented one of two stimuli differing only in odor (30 x 20 x 15 mm cloth bags filled with either plastic beads or banana), S. lineola did not attempt to eat the former, but 50% attempted to eat the bag of banana. Eating bouts were always preceded by antennae drumming in the direction of, or over the stimulus.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Reoxygenation of endothelial cells increases permeability by oxidant-dependent mechanisms.

We investigated the effects of hypoxia/reoxygenation exposure on the barrier function of endothelial cell monolayers. Bovine pulmonary microvessel endothelial cells were grown to confluence on microporous filters (0.8-microns pore diameter) and exposed to hypoxia (0.1% O2 or PO2 approximately 1 mm Hg) for 2, 4, 12, or 24 hours, followed by reoxygenation with room air for a period ranging from 16 seconds to 2 hours. The transendothelial clearance rate of 125I-albumin was measured to determine the permeability of endothelial monolayers. Permeability increased twofold or fivefold over control values after 1 hour of reoxygenation in monolayers that had been exposed to either 12 or 24 hours of hypoxia. The response occurred within 5 minutes of reoxygenation, increased maximally by 40 minutes, and remained elevated with continuous reoxygenation for up to 2 hours. The increase in permeability was associated with F-actin reorganization, a change to spindlelike cells, and injured mitochondria. Immunoblot analysis indicated that neither hypoxia alone nor reoxygenation changed CuZn superoxide dismutase (SOD), MnSOD, and catalase levels. However, release of superoxide anions (O2-) into the extracellular medium increased by twofold within 40-60 minutes of reoxygenation. Treatment of endothelial cells with CuZnSOD (100 units/ml) for the 24-hour hypoxia period prevented O2- generation and approximately 50% of the increase in permeability. Higher CuZnSOD concentrations (greater than or equal to 200 units/ml) were not protective. Treatment with catalase (100-1,000 units/ml) inhibited the reoxygenation-induced increase in permeability at the highest catalase concentration (1,000 units/ml), suggesting a critical role of hydrogen peroxide in mediating the response.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1991

Lectin binding to gp60 decreases specific albumin binding and transport in pulmonary artery endothelial monolayers.

The effect of albumin binding to cultured bovine pulmonary artery endothelial cell (BPAEC) monolayers on the transendothelial flux of 125I-labelled bovine serum albumin (BSA) was examined to determine its possible role on albumin transcytosis. The transport of 125I-BSA tracer across BPAEC grown on gelatin- and fibronectin-coated filters (0.8 microns pore diam.) was affected by the presence of unlabelled BSA in the medium in that transendothelial 125I-BSA permeability decreased, reaching a 40% reduction at BSA concentrations equal to or greater than 5 mg/ml. BSA binding to BPAEC monolayers was saturated at concentration of 10 mg/ml with an apparent binding affinity of 6 x 10(-7) M. In contrast, gelatin added to the medium altered neither 125I-BSA binding nor transport. Several lectins were tested for their ability to inhibit 125I-BSA binding and transport. One lectin, Ricinus communis (RCA), reduced 125I-BSA binding by 70% and transport by 40%. Other lectins, Ulex europaeus, Triticum vulgare, and Glycine max decreased neither 125I-BSA binding nor transport. The reduction of 125I-BSA transport by RCA was not observed in the presence of saturating levels of BSA, indicating that RCA influenced only the albumin-dependent component of transport. RCA, but not other lectins, precipitated a 60 kDa plasmalemmal glycoprotein from cell lysates of surface radioiodinated BPAEC monolayers. This 60 kDa glycoprotein appears to be the equivalent of gp60 identified previously as an albumin binding glycoprotein in rat microvascular endothelium. In summary, approximately 40% of albumin transport across BPAEC monolayers is dependent on albumin binding. This component of albumin transport is inhibited by 80% by the binding of RCA to gp60. These results suggest that binding of albumin to gp60 on pulmonary artery endothelial cell membrane is a critical determinant of transendothelial albumin flux involving mechanisms such as plasmalemmal vesicular transcytosis.

Animals↗

PubMed1991

Serum albumin decreases transendothelial permeability to macromolecules.

We examined the effects of serum albumin and other serum proteins on the fluxes of tracer 125I-albumin (MW 69 kDa) and 125I-haptoglobin (MW 100 kDa) across the pulmonary artery endothelial monolayer in vitro to test the role of serum proteins in modulating the endothelial barrier function. Replacement of control complete culture medium (20% fetal calf serum in DMEM) with DMEM alone increased the transendothelial 125I-albumin clearance rate (a measure of 125I-albumin permeability) by 83% of the control value. Repletion with 50% calf serum or with 2.0 g% albumin (i.e., the albumin concentration in 50% serum) decreased 125I-albumin permeability to the control value. This effect of serum or albumin was concentration-dependent since neither 12.5% serum nor 0.5 g% albumin (i.e., albumin concentration in 12.5% serum) altered 125I-albumin permeability from control values. The ammonium sulfate-precipitated serum protein fraction rich in albumin decreased 125I-albumin permeability from the control DMEM value, whereas serum fractions containing predominantly gamma-globulin or depleted of protein did not significantly alter 125I-albumin permeability. Other serum proteins that have been proposed to reduce endothelial permeability, alpha 1-acid glycoprotein (0.035-0.14 g/100 ml) and fibronectin (5 mg/100 ml), did not decrease 125I-albumin permeability from DMEM values. The endothelial permeability of 125I-haptoglobin of 4.63 +/- 0.53 x 10(-6) cm/sec in the presence of DMEM was 30% of the 125I-albumin permeability value. The addition of 2.0 g% albumin or 50% serum decreased 125I-haptoglobin permeability to 57 and 31%, respectively, of the DMEM value. These results indicate the critical role of serum albumin in regulating the restrictiveness of the endothelial barrier to macromolecules.

Animals↗

PubMed1991

Fibrinogen degradation product fragment D increases endothelial monolayer permeability.

We assessed the effects of the two primary high-molecular-weight fibrinogen degradation products (FDP), fragments D and E, on the pulmonary vascular endothelial barrier function. Fragments D and E were purified to homogeneity by QAE Sephadex chromatography followed by gel filtration. Incubation of bovine pulmonary artery endothelial monolayers with 0.5-2.0 microM fragment D for 2 h caused a doubling of transendothelial 125I-albumin clearance rate (a measure of 125I-albumin permeability). Fragment E only produced a 0.6-fold increase in 125I-albumin clearance rate at concentration of 4.0 microM. Both FDP remained active in incubating media with serum. The permeability-increasing effect of fragment D was reversible and was not due to cell detachment or lysis. The fragment-D effect was time dependent and was associated with redistribution of endothelial F-actin microfilaments. The effect was independent of the carboxy-terminal sequence on gamma-chain of fragment D. Fragments D and E binding to pulmonary artery endothelial cells was specific and reversible, but fragment D binding was three-fold greater than fragment E, which may account for the greater permeability increase mediated by fragment D. The results indicate that FDP, especially fragment D, increase endothelial permeability to albumin. The response involves specific binding of fragment D to endothelial cells and redistribution of intracellular actin.

Actins↗

PubMed1990

Morphological evidence for alveolar recruitment during inflation at high transpulmonary pressure.

The effect of continuous inflation of lungs at 30 cmH2O transpulmonary pressure (Ptp) on air-space size was assessed by chord length-frequency distribution analysis. Lungs from gerbils were excised, allowed to collapse freely, and inflated to 30 cmH2O Ptp in a humidified chamber kept at 37 degrees C. When the lungs appeared fully inflated with no observable pleural surface atelectasis, the left lung was occluded while the right was maintained at 30 cmH2O for 10 min longer and then occluded. During this time, the right lung increased its volume from 70 to 100%. Then both lungs were quick frozen, freeze dried, and embedded in glycol methacrylate, and 1- to 2-microns-thick histological sections cut. Lungs from a control group of gerbils were similarly inflated to 30 cmH2O, both left and right were occluded, the left was quick frozen immediately, and the right was frozen 10 min later. Chord lengths of air spaces from cranial and caudal lobes of lungs were acquired using a Dapple Systems image analyzer, and a two-population frequency distribution was generated for analysis with an IBM PC. The results indicate that the volume increase during continuous inflation at 30 cmH2O Ptp was associated with a shift in the chord length distribution toward the smaller chord lengths. A two-population statistical analysis indicated that the inflation resulted in an increase in the relative proportion of smaller chord lengths, with no increase in the mean of this smaller population. We conclude that continuous inflation at 30 cmH2O Ptp results in alveolar recruitment.

Animals↗

PubMed1990

Pulmonary edema induced by phagocytosing neutrophils. Protective effect of monoclonal antibody against phagocyte CD18 integrin.

We studied the changes in pulmonary hemodynamics and lung wet weight induced with opsonized zymosan (OZ) in isolated guinea pig lungs perfused with Ringer-albumin solution containing neutrophils (PMNs). Addition of OZ to the PMN-perfused lungs caused pulmonary vasoconstriction and weight gain; neither OZ nor PMNs added individually to the perfusate altered pulmonary vasomotor tone or wet weight. The steady gain in lung weight by 1,588 +/- 464 mg over the 45-minute study period was associated with pulmonary capillary hypertension and an increase in the capillary filtration coefficient, indicative of increased lung vascular permeability. These responses may not be due to generation of oxygen radicals, because the alterations in pulmonary hemodynamics and lung weight were not reduced by addition of superoxide dismutase, catalase, or superoxide dismutase plus catalase. We examined the basis of the PMN-mediated effects by layering PMNs on bovine pulmonary artery endothelial monolayers. Challenge with OZ resulted in increased endothelial permeability to 125I-albumin. The monoclonal antibody IB4 (directed against CD18, the common beta-subunit of structurally related adhesion receptors on phagocytes, LFA-1, Mac-1, and P150,95) prevented the OZ-mediated increase in PMN adherence to endothelial cells and the increase in endothelial permeability to 125I-albumin. IB4 also inhibited the lung weight gain mediated by the OZ-stimulated PMNs in intact lungs. The protective effect of IB4 could be ascribed neither to inhibition of uptake of OZ by PMNs nor to inhibition of release of oxygen radicals, myeloperoxidase, and elastase.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1989

Nature of thrombin-induced sustained increase in cytosolic calcium concentration in cultured endothelial cells.

It has recently been appreciated that thrombin induces the retraction of endothelial cells resulting in an alteration of the integrity of the monolayers. We studied thrombin-induced changes in cytosolic calcium concentration (Ca2+i) using microfluorometry of fura-2-loaded single cells, cell topography (scanning electron microscopy), and cytoskeleton (rhodamine phalloidin) in endothelial cells. Thrombin caused an initial and sustained phase of an increase in Ca2+i. Pretreatment with pertussis toxin abolished both phases of Ca2+i response. Sustained phase of thrombin effect required extracellular calcium. Pretreatment of endothelial cells with indomethacin protracted the sustained phase, whereas a lipoxygenase inhibitor, nordihydroguaiaretic acid curtailed it. Thrombin caused a marked retraction of confluent endothelial cells coincident with the sustained phase of Ca2+i response. This was paralleled by the formation of gaps in F-actin distribution at the periphery of the cells. Pretreatment of endothelial cells with nordihydroguaiaretic acid blunted the thrombin-induced cell retraction. Microinjection of various putative messengers into the endothelial cells showed that initial Ca2+ mobilization is not sufficient to account for sustained elevation of Ca2+i. The sustained response required microinjection of phospholipase A2 or co-injection of phospholipase A2 with phosphatidylinositol 4,5-bisphosphate-specific phospholipase C, phosphatidylinositol 1,4,5-trisphosphate, or CaCl2, further implying that thrombin receptor(s) can be coupled to both phospholipases C and A2. Sustained elevation of Ca2+i was a necessary prerequisite for the thrombin-induced changes in endothelial cell topography.

Animals↗

PubMed1989

Phallacidin prevents thrombin-induced increases in endothelial permeability to albumin.

Calf pulmonary artery endothelial monolayers cultured on polycarbonate filters were utilized to study 125I-labeled albumin permeability and actin filament distribution in response to thrombin challenge. Thirty-minute exposure to alpha-thrombin (10(-7) M) significantly increased albumin clearance rates. These changes were associated with marked alterations in actin filament distribution, resulting in loss of peripheral actin bands and an increase in the number of cytoplasmic stress fibers. Because the actin peripheral filaments are thought to play an important role in junctional stability, we postulated that stabilization of actin filaments should protect against thrombin-induced barrier disruptions. Pretreatment of cells with 0.3 microM 7-nitrobenz-2-oxa-1,3-diazole (NBD)-phallacidin, a specific actin-stabilizing agent, prevented the changes in actin filament distribution and markedly attenuated the increase in albumin permeability. Because of the potential toxicity of phallatoxins, we evaluated the effects of pretreatment on cell viability and growth parameters. There were no differences in viability, seeding efficiency, or doubling times in cells treated with 0.3 microM NBD-phallacidin in comparison to controls. Our data support the hypothesis that actin filaments, particularly peripheral bands, contribute significantly to the maintenance of barrier function in cultured endothelial cells.

Actins↗