bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of oxidant stress on inflammation and survival of iNOS knockout mice after marrow transplantation
Authors: Yang Shuxia , Porter Valerie A. , Cornfield David N. , Milla Carlos , Panoskaltsis-Mortari Angela , Blazar Bruce R. , Haddad Imad Y.
Published: December 2017

In a model of idiopathic pneumonia syndrome after bone marrow transplantation (BMT), injection of allogeneic T cells induces nitric oxide (·NO), and the addition of cyclophosphamide (Cy) generates superoxide (O[Formula: see text]·) and a tissue-damaging nitrating oxidant. We hypothesized that ·NO and O[Formula: see text]· balance are major determinants of post-BMT survival and inflammation. Inducible nitric oxide synthase (iNOS) deletional mutant mice (−/−) given donor bone marrow and spleen T cells (BMS) exhibited improved survival compared with matched BMS controls. Bronchoalveolar lavage fluids obtained on day 7post-BMT from iNOS(−/−) BMS mice contained less tumor necrosis factor-α and interferon-γ, indicating that ·NO stimulated the production of proinflammatory cytokines. However, despite suppressed inflammation and decreased nitrotyrosine staining, iNOS(−/−) mice given both donor T cells and Cy (BMS + Cy) died earlier than iNOS-sufficient BMS + Cy mice. Alveolar macrophages from iNOS(−/−) BMS + Cy mice did not produce ·NO but persisted to generate strong oxidants as assessed by the oxidation of the intracellular fluorescent probe 2′,7′-dichlorofluorescin. We concluded that ·NO amplifies T cell-dependent inflammation and addition of Cy exacerbates ·NO-dependent mortality. However, the lack of ·NO during Cy-induced oxidant stress decreases survival of T cell-recipient mice, most likely by generation of ·NO-independent toxic oxidants.
Hypoxia decreases lung neprilysin expression and increases pulmonary vascular leak
Authors: Carpenter Todd C. , Stenmark Kurt R.
Published: December 2017

Although prior studies suggest that hypoxia may increase pulmonary vascular permeability, the mechanisms responsible for that effect remain uncertain. Neprilysin (neutral endopeptidase) is a cell surface metallopeptidase that degrades several vasoactive peptides including substance P and bradykinin. We hypothesized that hypoxia could reduce lung neprilysin expression, leading to increased vascular leak. Weanling rats were exposed to normobaric hypoxia (inspired O2 fraction = 0.1). Lung neprilysin activity was significantly decreased after 24 and 48 h of hypoxia ( P < 0.006). The decrease in enzyme activity was associated with decreased lung neprilysin protein content and decreased lung neprilysin mRNA expression. Immunohistochemistry showed a predominantly perivascular distribution of neprilysin, with clear reductions in neprilysin immunoreactivity after exposure to hypoxia. Exposure to hypoxia for 24 h also caused marked increases in vascular leak ( P = 0.008), which were reversed by the administration of recombinant neprilysin. The hypoxia-induced increase in leak was also reversed by substance P and bradykinin receptor antagonists. We conclude that in young rats hypoxia decreases lung neprilysin expression, which contributes to increased pulmonary vascular leak via substance P and bradykinin receptors.
Inhaled carbon monoxide and hyperoxic lung injury in rats
Authors: Clayton Carolyn E. , Carraway Martha Sue , Suliman Hagir B. , Thalmann Edward D. , Thalmann Katherine N. , Schmechel Donald E. , Piantadosi Claude A.
Published: December 2017

Because carbon monoxide (CO) has been proposed to have anti-inflammatory properties, we sought protective effects of CO in pulmonary O2toxicity, which leads rapidly to lung inflammation and respiratory failure. Based on published studies, we hypothesized that CO protects the lung against O2by selectively increasing expression of antioxidant enzymes, thereby decreasing oxidative injury and inflammation. Rats exposed to O2with or without CO [50–500 parts/million (ppm)] for 60 h were compared for lung wet-to-dry weight ratio (W/D), pleural fluid volume, myeloperoxidase (MPO) activity, histology, expression of heme oxygenase-1 (HO-1), and manganese superoxide dismutase (Mn SOD) proteins. The brains were evaluated for histological evidence of damage from CO. In O2-exposed animals, lung W/D increased from 4.8 in normal rats to 6.3; however, only CO at 200 and 500 ppm decreased W/D significantly (to 5.9) during O2exposure. Large volumes of pleural fluid accumulated in all rats, with no significant CO treatment effect. Lung MPO values increased after O2and were not attenuated by CO treatment. CO did not enhance lung expression of oxidant-responsive proteins Mn SOD and HO-1. Animals receiving O2and CO at 200 or 500 ppm showed significant apoptotic cell death in the cortex and hippocampus by immunochemical staining. Thus significant protection by CO against O2-induced lung injury could not be confirmed in rats, even at CO concentrations associated with apoptosis in the brain.
Synergistic effects of tumor necrosis factor-α and thrombin in increasing endothelial permeability
Authors: Tiruppathi Chinnaswamy , Naqvi Tabassum , Sandoval Raudel , Mehta Dolly , Malik Asrar B.
Published: December 2017

Because activation of the coagulation cascade and the generation of thrombin coexist with sepsis and the release of tumor necrosis factor (TNF)-α, we determined the effects of TNF-α on the mechanism of thrombin-induced increase in endothelial permeability. We assessed Ca2+signaling in human umbilical vein endothelial cells. In human umbilical vein endothelial cells exposed to TNF-α for 2 h, thrombin produced a rise in the intracellular Ca2+concentration ([Ca2+]i) lasting up to 10 min. In contrast, thrombin alone produced a rise in [Ca2+]ilasting for 3 min, whereas TNF-α alone had no effect on [Ca2+]i.Thrombin-induced inositol 1,4,5-trisphosphate generation was not different between control and TNF-α-exposed cells. In the absence of extracellular Ca2+, thrombin produced similar increases in [Ca2+]iin both control and TNF-α-exposed cells. In TNF-α-exposed cells, the thrombin-induced Ca2+influx after intracellular Ca2+store depletion was significantly greater and prolonged compared with control cells. Increased Ca2+entry was associated with an approximately fourfold increase in Src activity and was sensitive to the Src kinase inhibitor PP1. After TNF-α exposure, thrombin caused increased tyrosine phosphorylation of junctional proteins and actin stress fiber formation as well as augmented endothelial permeability. These results suggest that TNF-α stimulation of endothelial cells results in amplification of the thrombin-induced Ca2+influx by an Src-dependent mechanism, thereby promoting loss of endothelial barrier function.
An immediate endothelial cell signaling response to lung ischemia
Authors: Song Chun , Al-Mehdi Abu B. , Fisher Aron B.
Published: December 2017

Abrupt cessation of lung perfusion induces a rapid endothelial response that is not associated with anoxia but reflects loss of normal shear stress. This response includes membrane depolarization, H2O2generation, and increased intracellular Ca2+. We evaluated these parameters immediately upon nonhypoxic ischemia using fluorescence videomicroscopy to image in situ endothelial cells in isolated, ventilated rat lungs. Lungs labeled with 4-{2-[6-(dioctylamino)-2-naphthalenyl]ethenyl}1-(3-sulfopropyl)-pyridinium (di-8-ANEPPS; a membrane potential probe), Amplex Red (an extracellular H2O2probe), or fluo 3-AM (a Ca2+indicator) were subjected to control perfusion followed by global ischemia. Endothelial di-8-ANEPPS fluorescence increased significantly within the first second of ischemia and stabilized at 15 s, indicating membrane depolarization by ∼17 mV; depolarization was blocked by preperfusion with the K+channel agonist lemakalim. Increased H2O2, inhibitable by catalase, was detected in the vascular space at 1–2 s after the onset of ischemia. Increased intracellular Ca2+was detected 10–15 s after the onset of ischemia; the initial increase was inhibited by preperfusion with thapsigargin. Thus the temporal sequence of the initial response of endothelial cells in situ to loss of shear stress (i.e., ischemia) is as follows: membrane depolarization, H2O2release, and increased intracellular Ca2+.
Keratinocyte growth factor reduces alveolar epithelial susceptibility to in vitro mechanical deformation
Authors: Oswari Jane , Matthay Michael A. , Margulies Susan S.
Published: December 2017

Keratinocyte growth factor (KGF) is a potent mitogen that prevents lung epithelial injury in vivo. We hypothesized that KGF treatment reduces ventilator-induced lung injury by increasing the alveolar epithelial tolerance to mechanical strain. We evaluated the effects of in vivo KGF treatment to rats on the response of alveolar type II (ATII) cells to in vitro controlled, uniform deformation. KGF (5 mg/kg) or saline (no-treatment control) was instilled intratracheally in rats, and ATII cells were isolated 48 h later. After 24 h in culture, both cell groups were exposed to 1 h of continuous cyclic strain (25% change in surface area); undeformed wells were included as controls. Cytotoxicity was evaluated quantitatively with fluorescent immunocytochemistry. There was >1% cell death in undeformed KGF-treated and control groups. KGF pretreatment significantly reduced deformation-related cell mortality to only 2.2 ± 1.3% (SD) from 49 ± 5.5% in control wells ( P < 0.001). Effects of extracellular matrix, actin cytoskeleton, and phenotype of KGF-treated and control cells were examined. The large reduction in deformation-induced cell death demonstrates that KGF protects ATII cells by increasing their strain tolerance and supports KGF treatment as a potential preventative measure for ventilator-induced lung injury.
Inhibition of voltage-gated K+currents by endothelin-1 in human pulmonary arterial myocytes
Authors: Shimoda Larissa A. , Sylvester J. T. , Booth Gregory M. , Shimoda Tenille H. , Meeker Sonya , Undem Bradley J. , Sham James S. K.
Published: December 2017

Recent studies demonstrate that endothelin-1 (ET-1) constricts human pulmonary arteries (PA). In this study, we examined possible mechanisms by which ET-1 might constrict human PA. In smooth muscle cells freshly isolated from these arteries, whole cell patch-clamp techniques were used to examine voltage-gated K+(KV) currents. KVcurrents were isolated by addition of 100 nM charybdotoxin and were identified by current characteristics and inhibition by 4-aminopyridine (10 mM). ET-1 (10−8M) caused significant inhibition of KVcurrent. Staurosporine (1 nM), a protein kinase C (PKC) inhibitor, abolished the effect of ET-1. Rings of human intrapulmonary arteries (0.8–2 mm OD) were suspended in tissue baths for isometric tension recording. ET-1-induced contraction was maximal at 10−8M, equal to that induced by KVchannel inhibition with 4-aminopyridine, and attenuated by PKC inhibitors. These data suggest that ET-1 constricts human PA, possibly because of myocyte depolarization via PKC-dependent inhibition of KV. Our results are consistent with data we reported previously in the rat, suggesting similar mechanisms may be operative in both species.
Upregulation of transcription factors in lung in the early phase of postpneumonectomy lung growth
Authors: Landesberg Leonard J. , Ramalingam Ramachandran , Lee Kenneth , Rosengart Todd K. , Crystal Ronald G.
Published: December 2017

In the adult rodent, pneumonectomy results in compensatory lung growth characterized by cell proliferation. The molecular mechanisms governing this response remain unknown. We hypothesized that, in the early period postpneumonectomy, upregulated expression of transcription factors drives the growth process. We utilized a cDNA expression array to screen for upregulated transcription factors after left pneumonectomy in adult C57BL/6 mice, using unoperated mice as controls. Quantification of mRNA expression in the remaining lung at 2 h demonstrated a twofold or greater upregulation of six transcription factors: early growth response gene-1 (Egr-1), Nurr77, tristetraprolin, the primary inhibitor of nuclear factor-κB (IκB-α), gut-enriched Krüppel-like factor (GKLF), and LRG-21. Northern analysis was used to quantify the upregulation of expression of these genes relative to sham thoracotomy and unoperated controls. The largest increase was in Egr-1 (4.7-fold > naive). Time-course analysis over the first 24 h confirmed the transient nature of the early upregulation. In the context that postpneumonectomy lung growth is associated with cell proliferation and that genes such as Egr-1, Nurr77, LRG-21, and tristetraprolin have known roles in stress response, vascular biology, embryology, and cellular development, these data support the concept that transcription factors function early in the cascade of events leading to the compensatory response.
Translational activation and repression by distinct elements within the 5′-UTR of ENaC α-subunit mRNA
Authors: Otulakowski Gail , Freywald Tanya , Wen Yanxia , O'Brodovich Hugh
Published: December 2017

The rat amiloride-sensitive epithelial Na+channel (rENaC), the rate-limiting step in epithelial Na+transport, consists of three subunits, α, β, and γ. We hypothesized that α-rENaC translation is regulated via its 5′-untranslated region (UTR). Transient transfections of α-rENaC promoter-reporter constructs in representative epithelial cell lines demonstrated up to fivefold differences in activity among constructs containing different amounts of the α-rENaC 5′-UTR sequence. Differences in reporter protein activity did not parallel differences in reporter mRNA, demonstrating that 5′-UTR regulation must be at the level of translation. Specifically, translation was enhanced by a region extending from +53 to +211 bp downstream from the transcription start site and repressed by the region between +367 and +499 bp. Examination of the 5′-UTR sequence revealed an out-of-frame initiation codon within the repressive region, 43 bp upstream from the start of the α-rENaC open reading frame. Mutational analysis of this upstream start codon indicated that it plays, at most, a minor role in impeding translation both in vitro and in vivo, suggesting that additional mechanisms of translational regulation are operative.
Direct and indirect bacterial killing functions of neutrophil defensins in lung explants
Authors: Porro Giuliana A. , Lee Jin-Hwa , de Azavedo Joyce , Crandall Ian , Whitehead Thomas , Tullis Elizabeth , Ganz Tomas , Liu Mingyao , Slutsky Arthur S. , Zhang Haibo
Published: December 2017

Studies of the antimicrobial activity of neutrophil defensins have mostly been carried out in microbiological media, and their effects on the host defense in physiological conditions are unclear. We examined 1) the antibacterial activity of defensins in physiological media with and without lung tissue present, 2) the effect of defensins on hydrogen peroxide (H2O2) production by lung tissue that had been exposed to bacteria, and 3) the effect of diphenyleneiodonium (DPI), an inhibitor of reactive oxygen species formation, on the antibacterial activity of defensins in the presence of lung tissue. Defensins were incubated with Escherichia colior Pseudomonas aeruginosa in the absence or presence of primary cultured mouse lung explants. Defensins reduced bacterial counts by ∼65-fold and ∼25-fold, respectively, at 48 h; bacterial counts were further decreased by ∼600-fold and ∼12,000-fold, respectively, in the presence of lung tissue. Defensins induced H2O2production by lung tissue, and the rate of killing of E. coli by defensins was reduced by ∼2,500-fold in the presence of 10 μM DPI. We conclude that defensins exert a significant antimicrobial effect under physiological conditions and that this effect is enhanced in the presence of lung tissue by a mechanism that involves the production of reactive oxygen species.
Lung fibroblasts inhibit activation-induced death of T cells through PGE2-dependent mechanisms
Authors: Yarovinsky Timur O. , Hunninghake Gary W.
Published: December 2017

Activation-induced cell death (AICD) is a regulatory mechanism eliminating excess activated T cells, mainly through a Fas/Fas ligand-dependent mechanism. The goal of this study was to determine whether mouse primary lung fibroblasts are capable of modulating AICD. Using T cell hybridoma DO11.10, we found that fibroblasts in coculture rescue T cells from AICD. Fibroblast-conditioned medium (FCM) also inhibited apoptosis of T cells activated with immobilized anti-CD3 antibody. The effects of lung fibroblasts are mediated, in part, by secreted prostaglandin E2(PGE2) because treatment of fibroblasts with indomethacin decreased antiapoptotic activity of FCM. Addition of exogenous PGE2to FCM from fibroblast cultures treated with indomethacin restored the inhibitory activity of FCM. Expression of Fas receptor and Fas ligand by anti-CD3-activated DO11.10 cells was not affected by PGE2. However, the same concentrations of PGE2significantly downregulated activation of caspase-8 and caspase-3. These results demonstrate that lung fibroblasts inhibit the AICD of T cells by secreting PGE2, which downregulates caspase activation and apoptosis.
Nuclear factor-κB augments β2-adrenergic receptor expression in human airway epithelial cells
Authors: Aksoy Mark O. , Bin Wei , Yang Yi , Yun-You Duan , Kelsen Steven G.
Published: December 2017

Interleukin (IL)-1β increases β2-adrenergic receptor (β2-AR) mRNA and density by protein kinase C (PKC)-dependent mechanisms in human airway epithelial cells. The present study examined the role of several nuclear transcription factors in the PKC-activated upregulation of β2-AR expression. BEAS-2B cells were exposed to the PKC activator phorbol 12-myristate 13-acetate (PMA; 0.1 μM for 2–18 h). PMA had no effect on activator protein (AP)-2 or cAMP response element binding protein DNA binding activity but markedly increased nuclear factor (NF)-κB and AP-1 binding as assessed by electrophoretic gel mobility shift assay. PMA also increased the activity of a β2-AR promoter-luciferase reporter construct in transiently transfected cells. These effects were inhibited by the PKC inhibitors Ro-31-8220 and calphostin C. Furthermore, with increasing Ro-31-8220, β2-AR promoter-reporter activity correlated closely with both NF-κB and AP-1 activities ( r > 0.89 for both). Finally, the selective NF-κB inhibitor MG-132 dose dependently reduced NF-κB binding and β2-AR promoter activity but increased AP-1 binding. We conclude that PKC-induced upregulation of β2-AR expression in human airway epithelial cells appears to be mediated, at least in part, by increases in NF-κB activity.
Preventing mediastinal shift after pneumonectomy impairs regenerative alveolar tissue growth
Authors: Hsia C. C. W. , Wu E. Y. , Wagner E. , Weibel E. R.
Published: December 2017

To examine the effects of mechanical lung strain on regenerative growth of alveolar septal tissue after pneumonectomy (PNX), we replaced the right lungs of adult dogs with a custom-shaped inflatable silicone prosthesis. The prosthesis was either inflated (Inf) to maintain the mediastinum at the midline or deflated to allow mediastinal shift. The animals were euthanized ∼15 mo later, and the lungs were fixed at a constant distending pressure. With the Inf prostheses, lung expansion, alveolar septal tissue volumes, surface areas, and diffusing capacity of the tissue-plasma barrier were significantly lower than with the deflated prostheses; the expected post-PNX tissue responses were impaired by 30–60%. Capillary blood volume was significantly higher with Inf prostheses, consistent with microvascular congestion. Measurements in the Inf group remained consistently and significantly higher than those expected for a normal left lung, indicating persistence of partial compensation. In one dog, delayed deflation of the prosthesis 9–10 mo after PNX led to vigorous lung expansion and septal tissue growth, particularly of type II epithelial cells. We conclude that mechanical lung strain is a major signal for regenerative lung growth; however, other signals are also implicated, accounting for a significant fraction of the compensatory response to PNX.
Rapid alveolar liquid removal by a novel convective mechanism
Authors: Wang P. M. , Ashino Y. , Ichimura H. , Bhattacharya J.
Published: December 2017

Although alveoli clear liquid by active transport, the presence of surface-active material on the alveolar surface suggests that convective mechanisms for rapid liquid removal may exist. To determine such mechanisms, we held the isolated blood-perfused rat lung at a constant alveolar pressure (Pa). Under videomicroscopy, we micropunctured a single alveolus to infuse saline or Ringer solution in ∼10 adjacent alveoli. Infused alveoli were lost from view. However, as the infused liquid cleared, the alveoli reappeared and their diameters could be quantified. Hence the time-dependent determination of alveolar diameter provided a means for quantifying the time to complete liquid removal (C t ) in single alveoli. All determinations were obtained at an Pa of 5 cmH2O. C t , which related inversely to alveolar diameter, averaged 4.5 s in alveoli with the fastest liquid removal. Injections of dye-stained liquid revealed that the liquid flowed from the injected alveoli to adjacent air-filled alveoli. Lung hyperinflations instituted by cycling Pa between 5 and 15 cmH2O decreased C t by 50%. Chelation of intracellular Ca2+ prolonged C t and abolished the inflation-induced enhancement of liquid removal. We conclude that when liquid is injected in a few alveoli, it rapidly flows to adjacent air-filled alveoli. The removal mechanisms are dependent on alveolar size, inflation, and intracellular Ca2+. We speculate that removal of liquid from the alveolar surface is determined by the curvature and surface-active properties of the air-liquid interface.
Platelet lipid(s) bound to albumin increases endothelial electrical resistance: mimicked by LPA
Authors: Minnear Fred L. , Patil Sandeep , Bell Donald , Gainor Jonathan P. , Morton Christine A.
Published: December 2017

The objectives were to determine whether the permeability-decreasing activity of platelet-conditioned medium (PCM) is associated with a lipid bound to albumin and whether lysophosphatidic acid (LPA) is present in the PCM. A decrease in permeability was assessed by an increase in electrical resistance across endothelial cell monolayers derived from bovine pulmonary arteries and microvessels. The Sephacryl S-200 fraction of PCM that contained albumin, the albumin immunoprecipitate from the PCM, and the methanol extract from the albumin immunoprecipitate all increased endothelial electrical resistance. Increased electrical resistance induced by PCM was not abolished by boiling and was mimicked by 1-oleoyl-LPA and 1-palmitoyl-LPA. Analysis of a methanol-chloroform extract of one sample of PCM by electrospray mass spectrometry revealed many fatty acids, ceramide, diacylglycerol, phosphatidic acid, and palmitoyl-LPA, but analysis of a second sample of PCM and the methanol extract of its albumin immunoprecipitate revealed no LPA, only lipids. These findings indicate that a bioactive lipid(s), possibly LPA, released from platelets and subsequently bound to albumin forms an active complex that decreases endothelial permeability.
Functional effects of protease-activated receptor-2 stimulation on human airway smooth muscle
Authors: Chambers Linda S. , Black Judith L. , Poronnik Philip , Johnson Peter R. A.
Published: December 2017

The protease-activated receptor (PAR)-2 is present on the smooth muscle and epithelium of human airways and can be activated by mast cell tryptase, trypsin, or the PAR-2 activating peptide (AP). Trypsin and the PAR-2 AP induced contractions in human isolated airways, and these contractions were potentiated in the presence of the cyclooxygenase (COX) inhibitor indomethacin. Trypsin also increased the contractions to histamine in airways from sensitized (allergic) patients but not from nonsensitized (nonallergic) patients. Tryptase purified from human lung, skin and lung recombinant β-tryptases, trypsin, and the PAR-2 AP all increased DNA synthesis in human airway smooth muscle (HASM) cells. Activation of PAR-2 by tryptase, trypsin, and the PAR-2 AP did not induce PGE2release from HASM cells. Trypsin and the PAR-2 AP increased the levels of intracellular calcium in HASM cells, with desensitization evident after treatment with either agonist. In conclusion, activation of PAR-2 can induce contractions of human airways, potentiate contractions to histamine, and induce proliferation and therefore may contribute to airway diseases such as asthma.
Induction of MMP-9 in normal human bronchial epithelial cells by TNF-α via NF-κB-mediated pathway
Authors: Hozumi Atsuko , Nishimura Yoshihiro , Nishiuma Teruaki , Kotani Yoshikazu , Yokoyama Mitsuhiro
Published: December 2017

In this study, we determined whether the proinflammatory cytokines tumor necrosis factor (TNF)-α and interleukin-1β contribute to the regulation of matrix metalloproteinase (MMP)-9 in human bronchial epithelial cells and whether the induction of MMP-9 is regulated by the transcription factor nuclear factor (NF)-κB. We demonstrated that TNF-α induced MMP-9 at both the protein and mRNA levels in human bronchial epithelial cells and that interleukin-1β did not. In contrast, induction of the tissue inhibitor of metalloproteinase-1 by TNF-α was less than that of interleukin-1β. Increased expression of MMP-9 and NF-κB activation induced by TNF-α were inhibited by pyrrolidine dithiocarbamate and N-acetyl-l-cysteine but were not inhibited by curcumin. These results suggest that TNF-α induces the expression of MMP-9 in human bronchial epithelial cells and that this induction is mediated via the NF-κB-mediated pathway.
Surfactant protein D enhances bacterial antigen presentation by bone marrow-derived dendritic cells
Authors: Brinker Karen G. , Martin Emily , Borron Paul , Mostaghel Elahe , Doyle Carolyn , Harding Clifford V. , Wright Jo Rae
Published: December 2017

Surfactant protein (SP) D functions as a soluble pattern recognition molecule to mediate the clearance of pathogens by phagocytes in the innate immune response. We hypothesize that SP-D may also interact with dendritic cells, the most potent antigen presenting cell, to enhance uptake and presentation of bacterial antigens. Using mouse bone marrow-derived dendritic cells, we show that SP-D binds to immature dendritic cells in a dose-, carbohydrate-, and calcium-dependent manner, whereas SP-D binding to mature dendritic cells is reduced. SP-D also binds to Escherichia coli HB101 and enhances its association with dendritic cells. Additionally, SP-D enhances the antigen presentation of an ovalbumin fusion protein expressed in E. coli HB101 to ovalbumin-specific major histocompatibility complex class II T cell hybridomas. The enhancement of antigen presentation by SP-D is dose dependent and is not shared by other collectin-like proteins tested. These studies demonstrate that SP-D augments antigen presentation by dendritic cells and suggest that innate immune molecules such as SP-D may help initiate an adaptive immune response for the purpose of resolving an infection.
Sialyl Lewisxhybridized complement receptor type 1 moderates acid aspiration injury
Authors: Kyriakides Constantinos , Wang Yong , Austen William G. , Favuzza Joanne , Kobzik Lester , Moore Francis D. , Hechtman Herbert B.
Published: December 2017

The potentially enhanced anti-inflammatory effects of the sialyl Lewisx(sLex)-decorated version of soluble complement receptor type 1 (sCR1) in moderating acid aspiration injury are examined. HCl was instilled in tracheostomy tubes placed in mice, and extravasation of125I-labeled albumin in bronchoalveolar lavage (BAL) fluid was used to calculate the vascular permeability index (PI). Neutrophil counts in BAL fluid and immunohistochemistry were performed. PI was moderated by 82% after treatment with sCR1sLexcompared with 54% in sCR1-untreated mice ( P < 0.05). Respective reductions in PI in mice treated 0.5 and 1 h after acid aspiration with sCR1sLexof 70 and 57% were greater than the decreases in PI of 45 and 38% observed in respective sCR1-treated groups ( P < 0.05). BAL fluid neutrophil counts in sCR1sLex-treated mice were significantly less than those in sCR1-treated animals, which were similar to those in untreated mice. Immunohistochemistry stained for sCR1 only on the pulmonary vascular endothelium of sCR1sLex- but not sCR1-treated mice. In conclusion, sCR1sLexmoderates permeability by antagonizing complement activation and neutrophil adhesion. Delayed complement and neutrophil antagonism significantly reduces injury.
Differential regulation of diverse physiological responses to VEGF in pulmonary endothelial cells
Authors: Becker Patrice M. , Verin Alexander D. , Booth Mary Ann , Liu Feng , Birukova Anna , Garcia Joe G. N.
Published: December 2017

The mechanisms responsible for the divergent physiological responses of endothelial cells to vascular endothelial growth factor (VEGF) are incompletely understood. We hypothesized that VEGF elicits increased endothelial permeability and cell migration via differential activation of intracellular signal transduction pathways. To test this hypothesis, we established a model of VEGF-induced endothelial barrier dysfunction and chemotaxis with bovine pulmonary endothelial cells. We compared the effects of VEGF on transendothelial electrical resistance (TER), actin cytoskeletal remodeling, and chemotaxis of lung endothelial cells and then evaluated the role of the mitogen-activated protein kinases (MAPKs) p38 and extracellular signal-regulated kinase (ERK)1/2 in VEGF-mediated endothelial responses. The dose response of pulmonary arterial and lung microvascular endothelial cells to VEGF differed when barrier regulation and chemotaxis were evaluated. Inhibition of tyrosine kinase, phosphoinositol 3-kinase, or p38 MAPK significantly attenuated VEGF-mediated TER, F-actin remodeling, and chemotaxis. VEGF-mediated decreased TER was also significantly attenuated by inhibition of ERK1/2 MAPK but not by inhibition of fetal liver kinase-1 (flk-1) or Src kinase. In contrast, VEGF-mediated endothelial migration was not attenuated by ERK1/2 inhibition but was abolished by inhibition of either flk-1 or Src kinase. These data suggest potential mechanisms by which VEGF may differentially mediate physiological responses in vivo.
Lung lining liquid modifies PM2.5 in favor of particle aggregation: a protective mechanism
Authors: Kendall Michaela , Tetley Teresa D. , Wigzell Edward , Hutton Bernie , Nieuwenhuijsen Mark , Luckham Paul
Published: December 2017

The health effects of particle inhalation including urban air pollution and tobacco smoke comprise a significant public health concern worldwide, although the mechanisms by which inhaled particles cause premature deaths remain undetermined. In this study, we assessed the physicochemical interactions of fine airborne particles (PM2.5) and lung lining liquid using scanning electron microscopy, atomic force microscopy, and X-ray photon spectroscopy. We provide experimental evidence to show that lung lining liquid modifies the chemistry and attractive forces at the surface of PM2.5, which leads to enhanced particle aggregation. We propose that this is an important protective mechanism that aids particle clearance in the lung.
Nitrogen dioxide exposure: effects on airway and blood cells
Authors: Frampton Mark W. , Boscia Joseph , Roberts Norbert J. , Azadniv Mitra , Torres Alfonso , Cox Christopher , Morrow Paul E. , Nichols Joan , Chalupa David , Frasier Lauren M. , Gibb F. Raymond , Speers Donna M. , Tsai Ying , Utell Mark J.
Published: December 2017

This study examined the effects of nitrogen dioxide (NO2) exposure on airway inflammation, blood cells, and antiviral respiratory defense. Twenty-one healthy volunteers were exposed on separate occasions to air and 0.6 and 1.5 ppm NO2for 3 h with intermittent moderate exercise. Phlebotomy and bronchoscopy were performed 3.5 h after each exposure, and recovered cells were challenged with respiratory viruses in vitro. Blood studies revealed a 4.1% NO2dose-related decrease in hematocrit ( P = 0.003). Circulating total lymphocytes ( P = 0.024) and T lymphocytes ( P = 0.049) decreased with NO2exposure. Exposure to NO2increased the blood lymphocyte CD4+-to-CD8+ratio from 1.74 ± 0.11 to 1.85 ± 0.12 in males but decreased it from 1.88 ± 0.19 to 1.78 ± 0.19 in females ( P < 0.001 for gender difference). Polymorphonuclear leukocytes in bronchial lavage increased with NO2exposure ( P = 0.003). Bronchial epithelial cells obtained after exposure to 1.5 ppm NO2released 40% more lactate dehydrogenase after challenge with respiratory syncytial virus than with air exposure ( P = 0.024). In healthy subjects, exposures to NO2at levels found indoors cause mild airway inflammation, effects on blood cells, and increased susceptibility of airway epithelial cells to injury from respiratory viruses.
Chronic hypertension impairs flow-induced vasodilation and augments the myogenic response in fetal lung
Authors: Storme Laurent , Parker Thomas A. , Kinsella John P. , Rairigh Robyn L. , Abman Steven H.
Published: December 2017

We hypothesized that altered vasoreactivity in perinatal pulmonary hypertension (PH) is characterized by abnormal responses to hemodynamic stress, including the loss of flow-induced vasodilation and an augmented myogenic response. Therefore, we studied the acute hemodynamic effects of brief compression of the ductus arteriosus (DA) in control fetal lambs and in lambs during exposure to chronic PH. In both groups, acute DA compression decreased pulmonary vascular resistance (PVR) by 20% at baseline ( day 0). After 2 days of hypertension, acute DA compression paradoxically increased PVR by 50% in PH lambs, whereas PVR decreased by 25% in controls. During the 8-day study period, PVR increased during acute DA compression in PH lambs, whereas acute DA compression continued to cause vasodilation in controls. Brief treatment with the nitric oxide (NO) synthase inhibitor nitro-l-arginine (l-NA) increased basal PVR in control but not PH lambs, suggesting decreased NO production in PH lambs. Chronic hypertension increased the myogenic response afterl-NA in PH lambs, whereas the myogenic response remained unchanged in controls. The myogenic response was inhibited by nifedipine in PH lambs, suggesting that the myogenic response is dependent upon the influx of extracellular calcium. We conclude that chronic PH impairs flow-induced vasodilation and increases the myogenic response in fetal lung. We speculate that decreased NO signaling and an augmented myogenic response contributes to abnormal vasoreactivity in PH.
Lysophosphatidic acid upregulates the epidermal growth factor receptor in human airway smooth muscle cells
Authors: Ediger Tracy L. , Danforth Bryan L. , Toews Myron L.
Published: December 2017

Human airway smooth muscle cells treated with lysophosphatidic acid (LPA) and epidermal growth factor (EGF) exhibit synergistic stimulation of mitogenesis (Ediger TL and Toews ML. J Pharmacol Exp Ther 294: 1076–1082, 2000). The effects of LPA treatment of human airway smooth muscle cells on EGF receptor (EGFR) regulation have now been investigated. LPA treatment for 12–24 h resulted in a twofold increase in 125I-EGF binding and EGFR protein levels as assessed by Western blot analysis. Competition binding assays indicated single-site binding with an affinity of 3 nM, and the affinity was not changed by LPA treatment. EGFR upregulation was blocked by cycloheximide and actinomycin D, suggesting that LPA influences transcriptional regulation of EGFR expression. Inhibitor studies revealed a prominent role for activation of mitogen-activated protein kinase and p70 ribosomal S6 kinase. Both synergism and EGFR upregulation increased with increased cell density, whereas EGFR expression in control cells decreased. The similar requirements for exposure time, LPA concentrations, and cell confluence suggest that EGFR upregulation may be one contributing factor to the synergistic stimulation of mitogenesis seen with LPA plus EGF.
Leptin mediates the parathyroid hormone-related protein paracrine stimulation of fetal lung maturation
Authors: Torday J. S. , Sun H. , Wang L. , Torres E.
Published: December 2017

Developing rat lung lipofibroblasts express leptin beginning on embryonic day (E) 17, increasing 7- to 10-fold by E20. Leptin and its receptor are expressed mutually exclusively by fetal lung fibroblasts and type II cells, suggesting a paracrine signaling “loop.” This hypothesized mechanism is supported by the following experimental data: 1) leptin stimulates the de novo synthesis of surfactant phospholipid by both fetal rat type II cells (400% · 100 ng−1· ml−1· 24 h−1) and adult human airway epithelial cells (85% · 100 ng−1· 24 h−1); 2) leptin is secreted by lipofibroblasts in amounts that stimulate type II cell surfactant phospholipid synthesis in vitro; 3) epithelial cell secretions such as parathyroid hormone-related protein (PTHrP), PGE2, and dexamethasone stimulate leptin expression by fetal rat lung fibroblasts; 4) PTHrP or leptin stimulate the de novo synthesis of surfactant phospholipid (2- to 2.5-fold/24 h) and the expression of surfactant protein B (SP-B; >25-fold/24 h) by fetal rat lung explants, an effect that is blocked by a leptin antibody; and 5) a PTHrP receptor antagonist inhibits the expression of leptin mRNA by explants but does not inhibit leptin stimulation of surfactant phospholipid or SP-B expression, indicating that PTHrP paracrine stimulation of type II cell maturation requires leptin expression by lipofibroblasts. This is the first demonstration of a paracrine loop that functionally cooperates to induce alveolar acinar lung development.
Renal alpha-adrenoceptors and sodium excretion in the dog
Authors: Fildes R. D. , Eisner G. M. , Calcagno P. L. , Jose P. A.
Published: December 2017

The purpose of this study was to characterize the age-dependent role of alpha-adrenergic activity on renal vascular resistance (RVR) and renal blood flow (RBF) and the role of alpha 2-receptors in sodium transport. Intrarenal infusion of phentolamine in hydropenic anesthetized pups and adult dogs increased RBF and glomerular filtration rate (GFR) only in pups. The effect was most marked in the youngest pups (15.67 +/- 0.67 days). At 1.0 micrograms X kg-1 X min-1, phentolamine increased RBF from 1.38 +/- 0.04 to 1.64 +/- 0.04 ml X min-1 X g kidney wt-1 and GFR from 0.19 +/- 0.01 to 0.23 +/- 0.01 ml X min-1 X g kidney wt-1. Absolute (UNaV) and fractional (FENa) sodium excretions increased in all animals, but mean percent increases were greatest in adult dogs. Intrarenal yohimbine infusion in adult dogs (10-100 micrograms X kg-1 X min-1) produced a dose-related increase in UNaV and FENa without affecting RBF and GFR. UNaV increased from 0.22 +/- 0.05 to 0.54 +/- 0.12 mueq X min-1 X g kidney wt-1 and FENa increased from 0.32 +/- 0.05 to 0.63 +/- 0.06% at the dose of 100 micrograms X kg-1 X min-1. These studies confirm a modest role for alpha-adrenoreceptors in the high RVR characteristic of newborn pups and provide evidence for a role of alpha 2-adrenoceptors in the renal transport of sodium; the extent of the contribution of renal alpha-adrenergic system could not be tested in this experiment.
Aldosterone-stimulated transmethylations are linked to sodium transport
Authors: Wiesmann W. P. , Johnson J. P. , Miura G. A. , Chaing P. K.
Published: December 2017

The effect of aldosterone (Aldo) on phospholipid (PL) biosynthesis in cultured toad bladder epithelial cells was studied in cells incubated with [1,2-14C]choline and [methyl-3H]methionine over a 5-h period. Aldo (10(-7) M) did not alter the uptake of either precursor but significantly stimulated the incorporation of both labels into phosphatidylcholine (PC), the only PL labeled. 3H labeling of PC increased 29% and 14C incorporation into PC increased 34% in cells exposed to Aldo. A similar 30% increase in protein carboxymethylation occurred in cells treated with Aldo. 3-Deazaadenosine (DZA), a methylation inhibitor, abolished the Aldo-stimulated increase in PC labeling from [3H]methionine. PC labeling from [1,2-14C]choline was not affected by DZA. Basal and Aldo-stimulated protein carboxy-methylation were inhibited by DZA. DZA (300 microM) caused a mild decrease in basal short-circuit current (ISC) but completely inhibited the ISC response to 10(-7) M Aldo. Inhibition was complete when DZA was added up to 2 h following exposure to Aldo, and was reversible. Cells previously exposed to Aldo showed a significant increase in ISC within 2 h following removal of DZA. We conclude that Aldo stimulates PL methylation, protein carboxymethylation, and turnover of PC from choline. Inhibition of methylation reactions coincides with the inhibition of ISC response to Aldo.
Glomerular ultrafiltration dynamics: historical perspective
Authors: Arendshorst W. J. , Gottschalk C. W.
Published: December 2017

Our knowledge of the structure and function of the renal glomerulus is reviewed in a historical context. The glomerular corpuscles were first described by Malpighi in 1666. Subsequent injection studies led to conflicting claims concerning a glomerular-tubular connection. This connection was accepted only after the convincing demonstration of the anatomical relationship essentially as we now know it by Bowman in 1842. Ludwig was the first to propose that the mechanism of separation of fluid in the glomeruli was by ultrafiltration. Estimates of the ultrafiltration forces in mammals led to conflicting speculation as to whether or not filtration-pressure equilibrium was reached in glomerular capillaries. Results of direct determinations in some Munich-Wistar rats indicate filtration pressure equilibrium, an ultrafiltration coefficient (Kf) of 0.08 nl X s-1 X mmHg-1, and a strong influence of plasma flow on filtration rate (GFR). In contrast, evidence has been presented that filtration dynamics in other Munich-Wistar rats and several other strains of rats are characterized by filtration disequilibrium, a Kf of 0.04 nl X s-1 X mmHg-1, and a weak dependence of GFR on plasma flow. In conscious and anesthetized rats, kidney GFR is usually relatively stable in the presence of renal vasodilation. Filtration disequilibrium is reported in the dog and equilibrium in the squirrel monkey. Although predictions for humans suggest filtration disequilibrium, final conclusions await an in-depth analysis of direct measurements.
Mechanism for inhibition of renin release by acute plasma volume expansion in the dog
Authors: Roy M. W. , Ott C. E. , Welch W. J. , Downs J. H. , Kotchen T. A.
Published: December 2017

Plasma volume expansion alters renal tubular sodium chloride transport and renal nerve activity. The purpose of this study was to determine the mechanism(s) for inhibition of renin secretion by acute volume expansion with albumin in the anesthetized dog. In dogs with a single intact kidney, albumin infusion decreased renin release by 86% and significantly increased renal blood flow, glomerular filtration rate, and sodium excretion. Albumin volume expansion inhibited renin secretion to a lesser extent in dogs with denervated filtering kidneys and in dogs with innervated nonfiltering kidneys. In dogs with denervated nonfiltering kidneys, albumin infusion did not change renin secretion. Comparable volume expansion was produced in all groups. Thus, inhibition of renin release by acute plasma volume expansion is dependent on both a renal tubular mechanism and the integrity of the renal nerves. Partial inhibition of renin release was observed with interruption of either one of the mechanisms, whereas interruption of both mechanisms totally abolished the effect of acute plasma volume expansion on renin secretion.
Determination of the sodium transport pool in epithelia from tracer fluxes: a simplified approach
Authors: Turnheim K. , Plass H.
Published: December 2017

The epithelial content of tracer Na can be calculated either from the exponential increase of unidirectional transepithelial Na tracer fluxes observed after addition of labeled Na to the solution bathing the luminal side of isolated epithelia or from the linear portion of the rate of accumulation of label in the serosal solution. The second method is not only simpler, because fewer data points are required and no logarithmic transformations are necessary, but also more accurate, as shown in experiments with stripped epithelia of rabbit descending colon. From the difference in tissue Na tracer content in the absence and presence of amiloride, which blocks luminal Na uptake, a Na concentration in the transport pool of 8-10 mM is obtained, assuming that all cells participate in active transport. From a comparison of the intracellular Na concentration derived from current-voltage relations of the apical Na entry step with the Na concentration in the epithelial Na transport pool, an estimate may be obtained of what fraction of the cell mass is involved in active Na transport.
Role of prostaglandins in renin suppression during acute potassium loading
Authors: Kirchner K. A.
Published: December 2017

Acute K loading suppresses plasma renin activity (PRA) and may alter prostaglandin biosynthesis. To determine whether the effect of K on PRA was modulated through changes in the prostaglandin system, dietary NaCl-restricted rats were infused with prostaglandin I2. PRA was then determined before and 90 min after a KNO3 infusion. Control rats received only the prostaglandin infusion. PRA increased in rats receiving prostaglandins alone. K loading prevented the increase in PRA. To further exclude prostaglandins in PRA suppression during K loading, the effects of K on PRA were determined during prostaglandin inhibition with meclofenamate or indomethacin. K loading also suppressed PRA during prostaglandin inhibition. In neither study could the changes in PRA be attributed to changes in mean arterial pressure, glomerular filtration rate, plasma volume expansion, or urinary NaCl excretion rate. We conclude that the effects of K on PRA are independent of the prostaglandin system.
Reabsorption and secretion of alpha-ketoglutarate along the rat nephron: a micropuncture study
Authors: Ferrier B. , Martin M. , Baverel G.
Published: December 2017

The transport of alpha-ketoglutarate (alpha-KG) across the luminal membrane of the rat nephron was studied by micropuncture and microassay techniques. In normal and acidotic rats, approximately 75% of the filtered alpha-KG was reabsorbed in the proximal tubule and 20% in the pars recta and/or loop of Henle at endogenous plasma concentration of alpha-KG. A progressive elevation to steady-state levels of plasma alpha-KG resulted in a progressive reduction of the fractional reabsorption of alpha-KG in the proximal tubule as well as in a progressive increase in the fractional reabsorption of alpha-KG in the pars recta and/or loop of Henle. At plasma alpha-KG concentration 20-40 times above normal, reabsorption of alpha-KG was found to be limited by a maximal tubular capacity. In alkalotic rats, net secretion of alpha-KG in the early proximal convoluted tubule, net reabsorption in the remainder of the proximal convoluted tubule, and net secretion in the pars recta and/or loop of Henle were observed. These micropuncture data indicate that, depending on the acid-base conditions, net reabsorption or net secretion of alpha-KG may occur in at least two distinct sites along the rat nephron.