bluuebunny/crossref_metadata_2025 huggingface dataset:
Platelet-activating factor increases platelet-dependent glycoconjugate secretion from tracheal submucosal gland
Authors: Sasaki T. , Shimura S. , Ikeda K. , Sasaki H. , Takishima T.
Published: December 2017

Using isolated glands from feline trachea, we examined the effect of platelet-activating factor (PAF) on radiolabeled glycoconjugate release and glandular contraction by measuring induced tension in the absence or presence of platelets. PAF alone did not produce any significant glandular contraction nor any significant change in glycoconjugate release from isolated glands. In the presence of purified platelets containing no plasma, PAF (10(-8) to 10(-5) M) produced significant glycoconjugate secretion in a dose-dependent fashion, but it produced no significant glandular contraction. PAF-evoked glycoconjugate secretion was time dependent, reaching a peak response of 277% of control 15-30 min after the exposure of isolated glands to 10(-5) M PAF in the presence of platelets and returning to 135% of controls at 2 h. Platelets alone did not produce any significant stimulation in glycoconjugate release. CV-3988, a known PAF antagonist, inhibited the secretory response to PAF. Methysergide, a known antagonist to receptors for 5-hydroxytryptamine, did not alter PAF-evoked glycoconjugate secretion. Both indomethacin and SQ 29,548, a thromboxane receptor antagonist, abolished the PAF-evoked glycoconjugate secretion from isolated submucosal glands. Epithiomethanothromboxane A2, a stable thromboxane A2 analogue, produced a significant increase in glycoconjugate secretion in a dose-dependent fashion. These findings indicate that PAF increases glycoconjugate release in the presence of platelets and that the increase is dependent on some aspect of platelet function, namely thromboxane generation.
Glucocorticoids regulate surfactant protein synthesis in a pulmonary adenocarcinoma cell line
Authors: O'Reilly M. A. , Gazdar A. F. , Clark J. C. , Pilot-Matias T. J. , Wert S. E. , Hull W. M. , Whitsett J. A.
Published: December 2017

Synthesis of pulmonary surfactant proteins SP-A, SP-B, and SP-C was demonstrated in a cell line derived from a human adenocarcinoma of the lung. The cells contained numerous lamellar inclusion bodies and formed organized groups of cells containing well-developed junctional complexes and apical microvillous membranes. Synthesis of SP-A was detected in the cells by enzyme-linked immunoabsorbent assay and by immunoprecipitation of [35S]methionine-labeled protein. SP-A was identified as an Mr 31,000-36,000 polypeptide containing asparagine-linked carbohydrate. Northern blot analysis detected SP-A mRNA of 2.2 kb. Dexamethasone (1-10 nM) enhanced the relative abundance of SP-A mRNA. Despite stimulation of SP-A mRNA, intracellular SP-A content was unaltered or inhibited by dexamethasone. SP-B and SP-C mRNAs and synthesis of the SP-B and SP-C precursors were markedly induced by dexamethasone. ProSP-B was synthesized and secreted primarily as an Mr 42,000-46,000 polypeptide. Proteolysis of the proSP-B resulted in the generation of endoglycosidase F-sensitive Mr = 19,000-21,000 and 25,000-27,000 peptides, which were detected both intra- and extracellularly. SP-C proprotein of Mr = 22,000 and smaller SP-C fragments were detected intracellularly but were not detected in the media. Mature forms of SP-B (Mr = 8,000) and SP-C (Mr = 4,000) were not detected. Glucocorticoids directly enhance the relative synthesis and mRNA of the surfactant proteins SP-A, SP-B, and SP-C. Discrepancies among SP-A mRNA, its de novo synthesis, and cell content suggest that glucocorticoid may alter both pre- and posttranslational factors modulating SP-A expression.
Oxidant-mediated activation of phospholipase A2 in pulmonary endothelium
Authors: Chakraborti S. , Gurtner G. H. , Michael J. R.
Published: December 2017

Exposure of bovine pulmonary arterial endothelial cells to the oxidant tert-butyl hydroperoxide (t-bu-OOH) caused a dose-dependent increase in the release of [14C]arachidonic acid and synthesis of the cyclooxygenase products, thromboxane, prostaglandin E2, prostaglandin D2, and prostacyclin. There was no detectable production of peptide leukotrienes before or after administration of t-bu-OOH. Pretreatment with the oxygen radical scavenger 4-hydroxy-2,2,6,6-tetramethylpiperidino radical (HTP) or the antioxidants vitamin E and dithiothreitol prevented the increased arachidonic acid (AA) release caused by t-bu-OOH. t-bu-OOH increased the activity of phospholipase A2 by increasing its apparent maximum velocity without affecting its Michaelis constant. The increased AA release caused by t-bu-OOH did not appear to require new RNA or protein synthesis, because pretreatment of the cells with actinomycin D or cycloheximide did not reduce the increased release of AA or activation of phospholipase A2 caused by t-bu-OOH. Dexamethasone pretreatment of the cells prevented the increase in phospholipase A2 activity, and AA release produced by t-bu-OOH. t-bu-OOH increased the activity of phospholipase A2 and release of AA in both the presence and absence of extracellular calcium (Ca2+). Pretreatment with a nominal Ca2+-free buffer, the Ca2+ chelator ethylene glycolbis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid, nifedipine, or verapamil did not reduce t-bu-OOH-stimulated AA release. In contrast, treatment with the intracellular Ca2+ chelator 8-N,N-diethyamino octyl 3,4,5-trimethoxybenzoate (TMB-8) prevented t-bu-OOH-stimulated AA release in both the presence and absence of extracellular Ca2+. Treatment with calmodulin antagonists also prevented the increased release of AA caused by t-bu-OOH.
Gastrin-releasing peptide stimulates glycoconjugate release from feline trachea
Authors: Lundgren J. D. , Baraniuk J. N. , Ostrowski N. L. , Kaliner M. A. , Shelhamer J. H.
Published: December 2017

The effect of gastrin-releasing peptide (GRP) on respiratory glycoconjugate (RGC) secretion was investigated in a feline tracheal organ culture model. RGC secretion was stimulated by GRP in a dose-dependent fashion at concentrations from 10(-8) to 10(-5) M (range 15-38% increase above control) with a peak effect within 0.5-1 h of incubation. GRP-(14-27), the receptor binding portion of GRP, and the related molecule, bombesin, also stimulated RGC secretion by approximately 20% above control. Acetyl-GRP-(20-27) stimulated RGC release by 10%, whereas GRP-(1-16) was inactive. Autoradiographic studies with 125I-GRP revealed that specific binding was restricted to the submucosal glands and the surface epithelium. A specific radioimmunoassay showed the content of GRP in feline trachea after extraction with ethanol-acetic acid to be 156 +/- 91 fmol/g wet wt. Indirect immunohistochemistry indicated that ganglion cells located just outside the cartilage contained GRP-immunoreactive materials. GRP is a novel mucus secretagogue that may participate in regulating airway mucosal gland secretion.
Calcitonin gene-related peptide in human nasal mucosa
Authors: Baraniuk J. N. , Lundgren J. D. , Goff J. , Mullol J. , Castellino S. , Merida M. , Shelhamer J. H. , Kaliner M. A.
Published: December 2017

To explore the potential range of functions for calcitonin gene-related peptide (CGRP) in human mucosa, we quantified human inferior turbinate nasal mucosal CGRP content by radioimmunoassay, localized CGRP-immunoreactivity by immunohistochemistry, detected 125I-CGRP binding sites by autoradiography, and tested the ability of CGRP to induce submucosal gland secretion in short-term explant culture of human nasal mucosa. Nasal mucosa contained 0.45-0.54 pmol CGRP/g wet wt (n = 18). Immunoreactive CGRP was found in nerve fibers that densely innervated the walls of small muscular arteries arterioles. Venules and venous sinusoids were innervated by individual CGRP staining fibers. Occasional CGRP-containing nerve fibers were also noted adjacent to submucosal gland acini, near the epithelial basement membrane, and between epithelial cells. Specific 125I-CGRP binding sites were concentrated on small muscular arteries and arterioles. CGRP (4 microM) did not stimulate glycoconjugate or lactoferrin release from mucosal explants. These results indicate that in the human nasal mucosa, CGRP is present in nerve fibers, which most likely represent nociceptive sensorimotor nerves that innervate vascular structures (muscular arteries, arterioles, veins and venous sinusoids). It is likely that CGRP release from sensory neurons may play a role in the regulation of vasomotor responses, but no evidence for a role of CGRP in glandular secretion was found.
Recent stereological methods for cell biology: a brief survey
Authors: Cruz-Orive L. M. , Weibel E. R.
Published: December 2017

With the advent of many new tools over the last five years, stereology has become simpler, assumption-free, and more efficient but, at the same time, new terms and concepts have proliferated, which risk overwhelming potential users. The present review is intended to meet the urgent need for a structured classification and evaluation of the newest stereological methods. Being fairly comprehensive, the exposition is necessarily succinct: the reader is referred to selected references for the necessary details and examples.
Liposome-mediated augmentation of antioxidant defenses in fetal rat pneumocytes
Authors: Tanswell A. K. , Olson D. M. , Freeman B. A.
Published: December 2017

Cultured pneumocytes, prepared from fetal rat lung, are growth inhibited and have increased lactate dehydrogenase release and prostaglandin synthesis in response to 50 and 95% O2 exposure. The uptake of cationic liposomes by these fetal cells is more rapid and extensive than is the case with cultured adult pneumocytes. Protection of fetal pneumocytes against the cytotoxic effects of 50 or 95% O2 by liposome-entrapped antioxidant enzymes requires a liposome phospholipid concentration of only 1 nmol/cm2, compared with 45 nmol/cm2 for adult cells, which is a cytotoxic phospholipid concentration for the fetal cells. Despite this capacity of low concentrations of liposomes containing superoxide dismutase and catalase to increase endogenous antioxidant enzyme content, and to protect against cell death, such treatment does not attenuate O2-mediated alterations of cell growth or prostaglandin release. Inhibition of pneumocyte DNA synthesis, by elevated O2 concentrations, cannot be attributed to an autocrine effect of enhanced prostaglandin synthesis, because the addition of 50 microM ibuprofen to inhibit prostaglandin synthesis does not prevent O2-mediated effects on DNA synthesis.
Prostaglandin and leukotriene production by alveolar type II cells in vitro
Authors: Cott G. R. , Westcott J. Y. , Voelkel N. F.
Published: December 2017

Alveolar type II cells were isolated from adult rats, cultured for 22 h, and individual eicosanoids in the media from unstimulated and stimulated cells were quantified by immunoassay. Stimulation with the calcium ionophore A23187 significantly increased the media levels of prostaglandins (prostaglandin and 6-keto-prostaglandin F1 alpha greater than thromboxane B2). In contrast to previous reports, increased media levels of leukotrienes were also recovered from cells incubated with A23187, but only for cells in culture for less than or equal to 24 h. The production of leukotriene C4 was confirmed by a combination of high-performance liquid chromatography and spectrophotometric analysis. The profile of eicosanoids produced by cultures of alveolar type II cells was distinctly different than that of similarly cultured alveolar macrophages. Finally, stimulation of alveolar type II cell cultures with either a phorbol ester or phospholipase C increased media prostaglandin levels but failed to increase leukotriene levels. We conclude that primary cultures of alveolar type II cells are capable of the de novo metabolism of arachidonic acid to both cyclooxygenase and lipoxygenase products and that the production of leukotrienes is dependent on both time in culture and agonist. Thus alveolar type II cells are a potential source for the production of these eicosanoids in vivo, and the particular lipid mediators produced may vary depending on the pathophysiologic stimulus.
Histamine stimulation of surfactant secretion from rat type II pneumocytes
Authors: Chen M. , Brown L. A.
Published: December 2017

The present study examined the effect of histamine on the secretion of phosphatidylcholine, the predominant component of pulmonary surfactant from adult rat alveolar type II pneumocytes in primary culture. Histamine stimulated surfactant secretion in a time- and dose-dependent manner. At a concentration of 10 microM, histamine stimulated surfactant release by 5.2-fold over the basal secretory rate. The concentration producing half the maximal response for histamine-induced secretion was 70 nM. Histamine-induced secretion was blocked by both the selective histamine1 receptor antagonist pyrilamine and the selective histamine2 receptor antagonist cimetidine, but not by the beta-adrenergic antagonist alprenolol. Histamine activated adenylate cyclase and intracellular adenosine 3',5'-cyclic monophosphate (cAMP) production. These effects on adenylate cyclase and cAMP induced by histamine were inhibited by the H2 antagonist cimetidine. This would suggest that surfactant secretion was stimulated by histamine through H2 receptors and by cAMP. When histamine and isoproterenol were added concomitantly, the effects on phosphatidylcholine secretion were additive; however, there was no additive effect on adenylate cyclase activation. This coupled with H2 antagonist pyrilamine inhibition of histamine-induced secretion would suggest that histamine stimulated surfactant secretion through another mechanism in addition to a cAMP-dependent mechanism.
Bioelectric properties of fetal alveolar epithelial monolayers
Authors: O'Brodovich H. , Rafii B. , Post M.
Published: December 2017

To investigate the bioelectric properties on one of the cell types that line the distal lung unit, we isolated type II alveolar epithelium from 18- to 20-day gestation fetal rats (term = 22 days) and grew them on collagen-coated nitrocellulose filters. Amiloride impaired ion transport in a dose-dependent fashion (10(-4) to 10(-6) M) with 10(-4) M decreasing potential difference (PD) (mean +/- SE, 2.0 +/- 0.49 to 0.9 +/- 0.26 mV, P less than 0.01, lumen negative) and short-circuit current (Isc) (7.0 +/- 1.0 to 2.4 +/- 0.64 uA/cm2, P less than 0.01) without affecting resistance (R) (241 +/- 33 to 216 +/- 41 omega. cm2). Benzamil (10(-5) M) but not dimethylamiloride (10(-5) M) decreased Isc. Terbutaline (10(-3) M) increased PD from 1.2 +/- 0.13 to 3.3 +/- 0.40 mV (P less than 0.01), and application of amiloride (10(-4) M) after terbutaline reduced PD and Isc to less than initial base-line values. The Na(+)-K(+)-2Cl- cotransport inhibitors bumetanide (10(-4) M) and furosemide (10(-3) M) had no effect on PD and Isc either before or after terbutaline. Neither the Cl- channel blocker diphenylamine-2-carboxylate (10(-3) M) nor the Na(+)-glucose cotransport inhibitor phloridzin (10(-3) M) affected the bioelectric properties. Fetal type II alveolar epithelium in primary culture actively transport ions and, on the basis of inhibitor-agonist experiments, probably do not secrete Cl- but absorb Na+ through Na+ channels.
Lung epinephrine synthesis
Authors: Kennedy B. , Elayan H. , Ziegler M. G.
Published: December 2017

We studied in vitro and in vivo epinephrine (E) synthesis by rat lung. Nine days after removal of the adrenal medullas, circulating E was reduced to 7% of levels found in sham-operated rats but 30% of lung E remained. Treatment of demedullated rats with 6 hydroxydopamine plus reserpine did not further reduce lung E. In the presence of S-[3H]adenosylmethionine lung homogenates readily N-methylated norepinephrine (NE) to form [3H]E. The rate of E synthesis by lung homogenates was progressively more rapid with increasing NE up to a concentration of 3 mM, above which it declined. The rate of E formation was optimal at an incubation pH of 8 and at temperatures of approximately 55 degrees C. We compared the E-forming enzyme(s) of lung homogenates with those of adrenal and cardiac ventricle. The adrenal contains mainly phenylethanolamine N-methyltransferase (PNMT), which is readily inhibited by SKF 29661 and methylates dopamine (DA) very poorly. Cardiac ventricles contain mainly nonspecific N-methyltransferase (NMT), which is poorly inhibited by SKF 29661 and readily methylates both DA and NE. Lung homogenates were inhibited by SKF 29661 about half as well as adrenal but more than ventricle. We used the rate of E formation from NE as an index of PNMT-like activity and deoxyepinephrine synthesis from DA as an index of NMT-like activity. PNMT and NMT activity in rat lung homogenates were not correlated with each other, displayed different responses to change in temperature, and were affected differently by glucocorticoids.(ABSTRACT TRUNCATED AT 250 WORDS)
Hydrogen peroxide-induced potentiation of contractile responses in isolated rat airways
Authors: Szarek J. L. , Schmidt N. L.
Published: December 2017

We hypothesized that metabolites of O2 may play a role in the development of airway hyperreactivity and undertook this study to examine the effects of one of these metabolites, hydrogen peroxide (H2O2), on electrical field stimulation-induced contractile responses of isolated rat intrapulmonary bronchi. Exposure to H2O2 (1 mM) elicited a transient contractile response with a peak response equivalent to 18.1 +/- 2.0% of the reference contraction obtained to electrical stimulation. The H2O2-induced contraction was attenuated by pretreatment of tissues with indomethacin and superoxide dismutase, but abolished by catalase and mianserin. Subsequent to H2O2 exposure, electrical field stimulation-induced contractile responses were potentiated (P less than 0.0001), whereas acetylcholine-induced contractions were not. The potentiating effects of H2O2 were inhibited by catalase and mianserin. Addition of 5-hydroxytryptamine (5-HT) to the bath similarly potentiated contractions to electrical stimulation (P less than 0.0001). Together, these results are consistent with a role for 5-HT in H2O2-induced contraction and the subsequent potentiation of airway smooth muscle contraction elicited by cholinergic nerve activation. Thus endogenous metabolites of O2 may be important in modulating airway smooth muscle tone.
Interactions between Ca2+ and cAMP messenger system in regulation of airway smooth muscle contraction
Authors: Rasmussen H. , Kelley G. , Douglas J. S.
Published: December 2017

Cholinergic stimuli acting via muscarinic receptors stimulate airway smooth muscle to contract and beta-adrenergic stimuli induce it to relax. Cholinergic agonists stimulate intracellular Ca2+ release and Ca2+ influx, and beta-adrenergic agonists stimulate adenosine 3',5'-cyclic monophosphate (cAMP) production. Since it was found that cAMP alters cholinergic agonist-induced changes in cellular Ca2+ metabolism, it was postulated that cAMP brought about a relaxation of this smooth muscle primarily by altering Ca2+ metabolism. More recently, it has become clear that when cholinergic agonists, as well as histamine and serotonin, act, the hydrolysis of polyphosphoinositols is stimulated, diacylglycerol production is increased, and coordinated changes in Ca2+ metabolism and protein kinase function occur. These agonists also act to inhibit adenylate cyclase activity. Conversely, it has been found that in the absence of agonist activation, cAMP increases Ca2+ influx without stimulating contraction and that in the presence of muscarinic agonists, it acts to inhibit polyphosphoinositide hydrolysis. In addition, cAMP has been shown to alter the state of phosphorylation of a number of cellular proteins, an effect that is independent of any effect on cellular Ca2+ metabolism. Based on these newer data, an expanded model of Ca2(+)-cAMP interactions in the regulation of airway smooth muscle tone is presented. A key feature of this model is the operation of dual, reciprocal inhibitory signals by which the Ca2+ and cAMP messenger systems modulate each other's expression.
Activation of protein kinase C inhibits extraction of serotonin by perfused rat lung in situ
Authors: Weng W. , Pitt B. R.
Published: December 2017

Pulmonary endothelial extraction (E) of serotonin (5-HT) is decreased after exposure to phorbol myristate acetate (PMA) in intact (D. Riggs, A. M. Havill, B. R. Pitt, and C. N. Gillis. J. Appl. Physiol. 64: 2508–2516, 1988.) or perfused (C. L. Myers and B. R. Pitt. J. Appl. Physiol. 65: 377–384, 1988.) lungs. Although the mechanism underlying this change is unclear, we hypothesized, based on studies in cultured pulmonary arterial endothelial cells [C. L. Myers, J. S. Lazo, and B. R. Pitt. Am. J. Physiol. 258 (Lung Cell. Mol. Physiol. 1): L253-L258, 1989] that activation of protein kinase C (PKC) by PMA inhibits this uptake process. Accordingly, we studied the ability of staurosporine, a potent inhibitor of PKC, to block the acute effect of PMA on E(5-HT) by rat lungs perfused at 10 ml/min with Krebs-bicarbonate with 3% albumin. Pulmonary E(5-HT) was measured by indicator-dilution techniques using 5-[3H]HT and [14C]dextran. Both PMA and mezerein (nonphorbol PKC activator) caused dose-dependent decreases in E(5-HT) and increases in perfusion pressure (Ppa). Staurosporine, alone, did not significantly affect either E(5-HT) or Ppa; however, staurosporine (100 nM) completely inhibited the effects of PMA (100 nM) on the above parameters. Papaverine, a nonspecific vasodilator, was able to partially inhibit the pressor response to PMA while not affecting the inhibition of E(5-HT) by PMA, suggesting that the effect on E(5-HT) was not secondary to vasoconstriction (and derecruitment). These data support the hypothesis that activation of PKC leads to prominent pulmonary vascular effects including vasoconstriction and inhibition of endothelial cell 5-HT uptake.
Postnatal development of lung alveoli: suppression by 13% O2 and a critical period
Authors: Massaro G. D. , Olivier J. , Dzikowski C. , Massaro D.
Published: December 2017

We studied the effect of 13% O2 on the development of the lung's gas-exchange region. Rats, acclimatized to 13% O2, remained in 13% O2 while pregnant, and were kept with their pups in 13% O2 until the pups were killed or were placed in air at age 15 days; other rats were always in air. Pups kept in 13% O2, unlike air pups, did not decrease mean chord length (Lm) or increase surface-to-volume ratio (S/V) of gas-exchange air spaces between age 2 and 14 days or by age 40 days. Rats placed in air at age 15 days did not change Lm or S/V even in air. Rats kept in 13% O2, and rats placed in air at age 15 days had fewer alveolar attachments to bronchioles than air rats. Gas-exchange air volume (VA) in 13% O2 rats was equal to or greater than in air rats; VA/kg was larger in 13% O2 than air rats. We conclude that maintenance of rats in 13% O2 during gestation and during the period alveoli are formed by septation blocks septation in a seemingly irrevocable manner. We suggest diminished septation decreases radial traction on conducting airways leading to increased VA/kg.
Muscarinic receptor subtypes in canine trachea
Authors: Brichant J. F. , Warner D. O. , Gunst S. J. , Rehder K.
Published: December 2017

Prejunctional and postjunctional muscarinic receptor subtypes were characterized in canine trachealis muscle strips. In vitro contractile responses of muscle strips to acetylcholine or electric field stimulation were determined in the absence and the presence of gallamine, pirenzepine, and 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP). Gallamine had no effect on the contractile response to acetylcholine but enhanced the contractile response to electric field stimulation. Pirenzepine and 4-DAMP reduced the contractile response to acetylcholine and electric field stimulation. The pA2 value for pirenzepine vs. acetylcholine [7.18 +/- 0.59 (SD)] was consistent with the affinity of pirenzepine for M2 or M3-receptors; whereas the pA2 value for 4-DAMP vs. acetylcholine (8.92 +/- 0.42) and the extremely low affinity of gallamine indicated postjunctional muscarinic receptors of the M3 subtype. The enhancement of the contractile response to electric field stimulation by gallamine suggested the presence of M2-prejunctional receptors.
Enhancement of tumor necrosis factor-induced endothelial cell injury by cycloheximide
Authors: Nolop K. B. , Ryan U. S.
Published: December 2017

Tumor necrosis factor (TNF), a potent polypeptide mediator released by activated monocytes and macrophages, has a number of proinflammatory effects on endothelial cells. TNF is cytotoxic to tumor cells in vivo and in vitro, but TNF-induced toxicity to endothelial cells is less well established. We now report that cycloheximide (CHX), an inhibitor of protein synthesis, renders endothelial cells highly susceptible to TNF-induced lysis. TNF alone did not change the overall rate of protein synthesis by endothelial cells, whereas the addition of CHX completely abolished protein synthesis. Endothelial cells incubated in TNF alone in high concentrations (up to 1,000 U/ml) showed minimal rounding up and release of 51Cr. Likewise, CHX alone (5 micrograms/ml) had no significant effect on endothelial cell morphology and release of 51Cr. However, incubation of endothelial cells in both CHX and TNF caused injury in a dose-dependent manner. Morphological evidence of cell retraction, rounding, and detachment began within 2 h, but specific 51Cr release did not begin to rise until after 4 h. These changes were not observed when endothelial cells were incubated with TNF/CHX at 4 degrees C. The combination of TNF/CHX was lethal to all endothelial cells tested (bovine pulmonary artery, human umbilical vein, and human aorta), with human aortic cells showing the most pronounced changes. We conclude that healthy endothelial cells are resistant to TNF-induced lysis, but inhibition of their ability to make protein renders them highly susceptible.
The regulation of lung elastin synthesis
Authors: Foster J. A. , Curtiss S. W.
Published: December 2017

The important role that elastin plays in the development and proper function of lung has long been recognized. Also, the intimate connection between pulmonary emphysema and the destruction of alveolar elastin has been well established. Understanding the mechanisms regulating pulmonary elastin synthesis is crucial to fully understanding these normal and pathological processes. In this article, we review recent literature on elastin structure, the elastin gene and its multiple RNA transcripts, and the different tropoelastin isoforms that are translated from these mRNAs. The similarity of lung and aortic elastin and the cellular origin of lung elastin are also discussed. We next examine the few studies addressing regulation of elastin expression during lung development, maturation, and aging. The search for modulators of pulmonary elastogenesis, which has yielded mostly negative results, is then reviewed. Finally, we present a cell culture model that has been developed to study the molecular basis of lung injury in pulmonary emphysema.
Plasticity in the airway epithelium
Authors: Basbaum C. , Jany B.
Published: December 2017

Normal cell turnover as well as the response to injury require cell proliferation and differentiation. The airway epithelium maintains these processes throughout adult life. Controlled homeostatically, cell proliferation and differentiation usually restore, as an end point, the pseudostratified architecture of the normal mucociliary epithelium. After injury, however, cell proliferation and differentiation sometimes establish, as an end point, regions of metaplastic cells. In this brief review, we have tried to summarize research findings that 1) describe the development of metaplastic lesions in morphological terms, 2) identify cells the proliferation of which forms the basis of these lesions, and 3) identify molecular changes within these cells that control development of the metaplastic phenotype.
Stimulation of prostaglandin synthesis by hyperoxia in perinatal rat lung cells
Authors: Lee D. S. , Bevan M. G. , Olson D. M.
Published: December 2017

Prostaglandins (PGs) have been implicated in the development of pulmonary oxygen toxicity. We tested the hypothesis that hyperoxia modulates PG synthesis in a differentiation-arrested primary lung cell culture model in the rat at three developmental ages: day-20 gestation (term = 22 days), days 1 and 3 after birth. The time courses of the response to hyperoxia were defined in preconfluent lung cells as well as in growth-arrested, confluent cells. From days 4-8 after plating in growth medium containing 10% carbonstripped fetal bovine serum, exposure to 95% O2, in contrast to 1% O2, inhibited cell proliferation but significantly enhanced the production of PGI2 and, to a lesser extent, PGE2 at all three ages. The capacity to metabolize exogenous arachidonic acid (AA) to PGI2 was also increased two-to threefold (P less than 0.01). Cellular release of lactate dehydrogenase, a measure of O2 toxicity, remained unchanged during exposure to 1% O2 but increased fivefold between 48 and 96 h after exposure to hyperoxia (from 2% total to 10.5%, P less than 0.01). In confluent, growtharrested cells, under serum-free conditions, exposure to hyperoxia for 24–48 h resulted in a similar induction of PG synthesis. Our results suggest that hyperoxia stimulates PG synthesis in the perinatal rat lung and that this effect is independent of cell growth or the presence of serum. We speculate that this hyperoxia-induced PG synthesis is a relatively early response to oxidant stress and may serve as an useful early marker for O2 toxicity in perinatal lung cells.
Strategies for analysis of gene expression: pulmonary surfactant proteins
Authors: Stripp B. R. , Whitsett J. A. , Lattier D. L.
Published: December 2017

Gene transcription is regulated by the formation of protein-DNA complexes that influence the rate of specific initiation of transcription by RNA polymerase. Recent experimental advances allowing the identification of cis regulatory sequences that specify the binding of trans acting protein factors have made significant contributions to our understanding of the mechanistic complexities of transcriptional regulation. These methodologies have prompted the use of similar strategies to elucidate transcriptional control mechanisms involved in the tissue specific and developmental regulation of pulmonary surfactant protein gene expression. The purpose of this review is to describe various methodologies by which molecular biologists identify and subsequently assay regions of nucleic acids presumed to be integral in gene regulation at the level of transcription. It is well established that genes encoding surfactant proteins are subject to regulation by hormones, cytokines, and a variety of biologically active reagents. Perhaps future studies utilizing molecular tools outlined in this review will be valuable in identification of DNA sequences and protein factors required for the regulation of lung surfactant genes.
Acrolein stimulates eicosanoid release from bovine airway epithelial cells
Authors: Doupnik C. A. , Leikauf G. D.
Published: December 2017

Injury to the airway mucosa after exposure to environmental irritants is associated with pulmonary inflammation and bronchial hyperresponsiveness. To better understand the relationships between mediator release and airway epithelial cell injury during irritant exposures, we studied the effects of acrolein, a low-molecular-weight aldehyde found in cigarette smoke, on arachidonic acid metabolism in cultured bovine tracheal epithelial cells. Confluent airway epithelial cell monolayers, prelabeled with “3H”arachidonic acid, released significant levels of 3H activity when exposed (20 min) to 100 microM acrolein. [3H]arachidonic acid products were resolved using reverse-phase high-performance liquid chromatography. Under control conditions the released 3H activity coeluted predominantly with the cyclooxygenase product, prostaglandin (PG) E2. After exposure to acrolein, significant “peaks” in 3H activity coeluted with the lipoxygenase products 12-hydroxyeicosatetraenoic acid (HETE) and 15-HETE, as well as with PGE2, PGF2 alpha, and 6-keto-PGF1 alpha. Dose-response relationships for acrolein-induced release of immunoreactive PGF2 alpha and PGE2 from unlabeled epithelial monolayers demonstrated 30 microM acrolein as the threshold dose, with 100 microM acrolein inducing nearly a fivefold increase in both PGF2 alpha and PGE2. Cellular viability after exposure to 100 microM acrolein, determined by released lactate dehydrogenase activity, was not affected until exposure periods were greater than or equal to 2 h. These results implicate the airway epithelial cell as a possible source of eicosanoids after exposure to acrolein.
Uptake of instilled radiolabeled lamellar bodies from alveolar compartment of the rat
Authors: Nicholas T. E. , Barr H. A. , Power J. H. , Jones M. E.
Published: December 2017

Following the instillation of lamellar bodies containing dipalmitoyl phosphatidyl-[3H]choline (DPPC) down the trachea of adult rats, we found that the half-life of alveolar [3H]-DPPC was 85 min and the time constant was 120 min. As much as 85% was recycled. When we labeled the DPPC with both [3H]choline and [14C]acetate, the ratio 3H/14C increased in the alveolar compartment and then increased further in the lamellar body fraction of the recipient lungs, suggesting that some deacylation-reacylation was occurring. Further evidence of degradation was an increase in free [3H]choline in the microsomal fraction. Whereas hyperpnea induced by breathing 5% CO2-13% O2–82% N2 increased the reuptake of DPPC, reuptake did not appear to be enhanced in the rest period immediately after hyperpnea induced by swimming, when alveolar DPPC was still markedly elevated. Propranolol did not affect reuptake, suggesting that beta-adrenoreceptors were not essential. We suggest that reuptake is coupled more to release than to the amount of surfactant in the alveolar compartment.
Tumor necrosis factor infusions in humans prime neutrophils for hypochlorous acid production
Authors: Wewers M. D. , Rinehart J. J. , She Z. W. , Herzyk D. J. , Hummel M. M. , Kinney P. A. , Davis W. B.
Published: December 2017

Five cancer patients undergoing intravenous infusions of human recombinant tumor necrosis factor (TNF)alpha were evaluated for the effects these infusions had on the priming of circulating neutrophils for hypochlorous acid (HOCl) production. These patients were also studied for changes in temperature, circulating white blood cell counts, blood pressure, and spontaneous monocyte interleukin 1 beta (IL-1 beta) and TNF production. As predicted by previous in vitro studies, patient neutrophils increased their HOCl production to unopsonized zymosan from a baseline of 29.2 +/- 5.9 nmol I- oxidized/4 x 10(6) cells to a peak of 64.2 +/- 9.8 nmol I- oxidized/4 x 10(6) cells at 4 h after TNF infusion (P less than 0.01). Similar increases were also seen at 4 h with phorbol myristic acetate and opsonized zymosan as the stimuli. The priming effect could be reproduced in neutrophils from a normal individual by incubating them with the 30-min serum samples from the infused patients. The ability of this serum to prime neutrophils was completely blocked by a monoclonal anti-TNF alpha-antibody but not by an anti-IL-1 beta antibody. In addition to the priming of their neutrophils, patients also experienced fever, marked hypotension, and an initial fall, followed by rebound to an elevation, in circulating white blood cell counts. The TNF infusions did not produce detectable circulating IL-1 beta nor did they induce significant production of TNF or IL-1 beta by circulating blood monocytes. These studies confirm the role of TNF in producing the signs of sepsis such as hypotension, fever, and leukopenia followed by leukocytosis.(ABSTRACT TRUNCATED AT 250 WORDS)
Antibody against leukocyte integrin (CD18) prevents reperfusion-induced lung vascular injury
Authors: Horgan M. J. , Wright S. D. , Malik A. B.
Published: December 2017

We studied the effects of pretreatment of rabbits with a monoclonal antibody (MoAb) IB4 directed against an epitope on the common beta-chain of leukocyte adhesion glycoprotein CD18 on the development of pulmonary edema following pulmonary artery occlusion and reperfusion. A balloon catheter occluded the right pulmonary artery in rabbits for 24 h, and the balloon was then deflated to allow reperfusion for 2 h. The lungs were removed, attached to a weighing balance, and perfused with Ringer-albumin solution (0.5 g/100 ml). IB4 (0.5 mg/kg) was infused 45 min before the start of reperfusion. In another experiment, we infused OKM-1 (0.5 mg/kg), a control MoAb directed against an irrelevant epitope on the alpha-chain of CD11b/CD18. IB4 prevented rabbit neutrophil adherence to pulmonary artery endothelial cells in vitro in response to a variety of stimuli, whereas OKM-1 was ineffective. Pulmonary artery occlusion and reperfusion increased the lung myeloperoxidase (MPO) content, whereas IB4 prevented the increase in MPO content. The increase in lung weight in the IB4-treated group (400 +/- 40 mg from baseline) was less (P less than 0.001) than in the untreated control group (1,400 +/- 90 mg) and the OKM-1-treated control group (1,320 +/- 125 mg). IB4 pretreatment also prevented the increase in pulmonary capillary filtration coefficient (a measure of capillary permeability to water) occurring in control and OKM-1-treated groups. The results indicate that neutrophil sequestration in the pulmonary microcirculation mediated by CD18 glycoprotein is a major determinant of reperfusion-induced lung vascular injury.
Intracellular calcium concentration of acinar cells in feline tracheal submucosal glands
Authors: Ishihara H. , Shimura S. , Sato M. , Masuda T. , Ishide N. , Miura M. , Sasaki T. , Sasaki H. , Takishima T.
Published: December 2017

We measured the intracellular free calcium ion concentration [( Ca2+]i) of acinar cells in isolated feline tracheal submucosal glands in response to secretagogues using the Ca2(+)-sensitive fluorescent dye fura-2. The secretagogues included cholinergic, adrenergic agonists, substance P (SP), and vasoactive intestinal polypeptide (VIP) which induce mucus glycoprotein secretion from feline tracheal submucosal glands. Methacholine (MCh) produced a significant increase in [Ca2+]i of up to 9.8 times that of control in a dose-dependent fashion at concentrations of 10(-8) to 10(-3) M. [Ca2+]i increase by MCh reached a peak within 30 s after stimulation and thereafter showed a sustained rise. In Ca2(+)-free medium, MCh produced an initial transient rise, which was less than 30% of that in a Ca2(+)-containing solution, and which lasted for 60 s with no prolonged sustained rise in [Ca2+]i. Atropine abolished MCh-evoked [Ca2+]i increase. Phenylephrine and SP produced a prolonged increase in [Ca2+]i without an initial transient increase. Phenylephrine (up to 10(-4) M) evoked an increase in [Ca2+]i by up to 240% that of control, which was abolished by prazosin. SP (up to 10(-4) M) also evoked an increase in [Ca2+]i by 155% that of control, which was abolished by atropine. By contrast, both isoproterenol (up to 10(-5) M) and VIP (up to 10(-5) M) failed to alter [Ca2+]i. These findings indicate that the mucus glycoprotein secretion evoked by muscarinic cholinergic, alpha-adrenergic agonist or SP can be mediated by intracellular Ca2+, whereas that by beta-adrenergic agonists or VIP cannot.
Ultrastructural analysis of regional type II cell development within fetal and neonatal lungs
Authors: Lin Y. , Lechner A. J.
Published: December 2017

Type II cells within guinea pig lungs were studied beginning when lamellar bodies (LB) were first identifiable, to determine the pattern of their ultrastructural maturation through parturition and to search for regional differences suggested for other species in their numbers or maturity within the same lung. Fetuses were examined at gestational ages of 55, 60, and 65 days, as were newborn within 2 h of birth at 68 days. The stratified sampling system divided left lungs into nine blocks from each cranial and caudal lobe for electron-microscopic morphometry. Type II cells were compared among blocks within lobes, between lobes within age groups, and among ages for volume densities [VV(ep2, t)], numerical densities [NV(ep2, t)], and surface densities [SV(ep2, t)] per block; total tissue volumes [V(ep2)], numbers [N(ep2)], and cumulative surface areas [S(ep2)] per lung; and the VV(lb, ep2) and V(lb) of their LB. Within lobes, there were no significant differences among blocks for any parameter. Within age groups, significant interlobar differences were limited only to NV(ep2, t) (cranial greater than caudal at day 55, P less than 0.001). In age-by-age comparisons, the V(ep2), NV(ep2), and S(ep2) each increased dramatically from day 55 to 60 (P less than 0.001) but then remained unchanged through parturition. However, both VV(lb, ep2) and V(lb) increased linearly with fetal age (day 55 less than 60 less than 65 less than newborn), with VV(lb, ep2) achieving 18% type II cell volume in the postpartum group.(ABSTRACT TRUNCATED AT 250 WORDS)
Electrophysiological properties of neurons in guinea pig bronchial parasympathetic ganglia
Authors: Myers A. C. , Undem B. J. , Weinreich D.
Published: December 2017

Active and passive membrane membrane properties of parasympathetic neurons were examined in vitro in a newly localized ganglion on the right bronchus of the guinea pig. Neurons could be classified as “tonic” or “phasic” based on their action potential discharge response to suprathreshold depolarizing constant current steps. Tonic neurons (39%) responded with repetitive action potentials sustained throughout the current step, whereas phasic neurons (61%) responded with an initial burst of action potentials at the onset of the step but then accommodated. Tonic and phasic neurons could not be differentiated by other active or passive membrane properties. Electrical stimulation of the vagus nerve elicited one to three temporally distinct fast nicotinic excitatory potentials, and tetanic stimulation of the vagus nerve evoked slow depolarizing (10% of neurons) and hyperpolarizing (25% of neurons) potentials; the latter was mimicked by muscarinic receptor activation. Similar slow and fast postsynaptic potentials were observed in both tonic and phasic neurons. We suggest neurons within the bronchial ganglion possess membrane and synaptic properties capable of integrating presynaptic stimuli.
Heterogeneity of intact granules purified from canine mastocytoma cell lines
Authors: Thomas P. S. , Schreck R. E. , Ruoss S. J. , Lazarus S. C.
Published: December 2017

We investigated the observation that some mast cells exhibit asynchronous release of granule-associated neutral proteases. Intact mast cell granules were isolated, purified, and studied with respect to their histamine, neutral protease, and proteoglycan content. Studies of two canine mastocytoma cell lines demonstrated differences in storage and packaging of granules within one of the cell lines (G) with respect to the neutral proteases and density, resulting in two distinct subpopulations of granules. By use of identical techniques no such differences were found in the other cell line (BR). These observations suggest that granule subpopulations in some mast cells may have a role in the differential release of mediators.
Role of endothelin 1 in regulating rabbit airway contractility
Authors: Grunstein M. M. , Chuang S. T. , Schramm C. M. , Pawlowski N. A.
Published: December 2017

Endothelin 1 (ET-1) is a potent vasoconstrictor peptide recently isolated from vascular endothelial cells. Because its role and mechanisms of action in regulating airway contractility remain to be identified, we examined the contractile effects of ET-1 in isolated rabbit tracheal smooth muscle (TSM) segments. In TSM under passive tension, ET-1 elicited dose-dependent contractions with a mean +/- SE -log 50% of maximal response value of 7.82 +/- 0.13 vs. a value of 5.61 +/- 0.07 -log M for acetylcholine (ACh). In TSM half-maximally contracted with ACh, however, ET-1 exerted dual and opposing contractile effects. Lower doses of ET-1 (less than or equal to 10(-9) M) produced a 74.2 +/- 16.6% decrease in active TSM tension. This relaxant response to ET-1 was associated with an accelerated accumulation of prostaglandin (PG) I2 and PGE2 and was attenuated by cyclooxygenase inhibition with indomethacin (10(-5) M). The combination of indomethacin and removal of the airway epithelium completely inhibited the TSM relaxant response to ET-1. In contrast, higher doses of ET-1 (greater than 10(-9) M) induced airway contractions that were attenuated by the Ca2+ channel blockers nifedipine (10(-5) M) and diltiazem (10(-5) M) and ablated in Ca2(+)-free buffer. Moreover, ET-1-induced TSM contractions were inhibited by the protein kinase C (PK-C) antagonists 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine, and staurosporine.(ABSTRACT TRUNCATED AT 250 WORDS)
Amiloride-inhibitable Na+ conductive pathways in alveolar type II pneumocytes
Authors: Matalon S. , Bridges R. J. , Benos D. J.
Published: December 2017

The purpose of these studies was to document the existence of electrogenic Na+ uptake by membrane vesicles of rabbit alveolar type II (ATII) cells and the extent to which this process was inhibited by amiloride. ATII cells (greater than 85% pure) were obtained by elastase digestion of lung tissue followed by Percoll centrifugation, and an enriched plasma membrane vesicle fraction was obtained by differential centrifugation. 22Na+ uptake into these vesicles was measured in the presence of a negative inside membrane potential, produced by the addition of the K+ ionophore valinomycin (10 microM) after all external K+ was removed. Electrogenic (valinomycin-sensitive) Na+ uptake (ELNa) was defined as the difference in uptake in the presence and absence of valinomycin. ELNa, normalized per milligram protein, was twice as high across ATII cells than alveolar macrophage membrane vesicles, was inhibited by amiloride (50% inhibitory concentration = 10 microM), and was decreased in the presence of an outwardly directed proton gradient (pHin 6.8; pHout 7.8), suggesting that it was not mediated by Na(+)-H+ antiport. Furthermore, ELNa was equally inhibited by increasing concentrations of amiloride and benzamil but was more sensitive to 5-(N-ethyl-N-isopropyl)-2'-4'-amiloride in concentrations of 10–1,000 microM. These findings indicate that a fraction of Na+ transport across ATII membrane vesicles occurs through a conductive pathway, probably a channel, that has different sensitivity to amiloride and its analogues than the previously described epithelial high amiloride-affinity Na+ channel.