Surfactant protein A enhances the binding and deacylation ofE. coliLPS by alveolar macrophages

1999 ◽  
Vol 276 (3) ◽  
pp. L540-L547 ◽  
Author(s):  
Cordula Stamme ◽  
Jo Rae Wright
Keyword(s):  
Alveolar Macrophages ◽  
Protein A ◽  
Surfactant Protein ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Cell Surface Molecule ◽  
Molar Ratios ◽  
A Cell ◽  
First Time ◽  
Lps Binding

Surfactant protein (SP) A and SP-D are involved in multiple immunomodulatory functions of innate host defense partly via their interaction with alveolar macrophages (AMs). In addition, both SP-A and SP-D bind to bacterial lipopolysaccharide (LPS). To investigate the functional significance of this interaction, we first tested the ability of SP-A and SP-D to enhance the binding of tritium-labeled Escherichia coli LPS to AMs. In contrast to SP-D, SP-A enhanced the binding of LPS by AMs in a time-, temperature-, and concentration-dependent manner. Coincubation with surfactant-like lipids did not affect the SP-A-mediated enhancement of LPS binding. At SP-A-to-LPS molar ratios of 1:2–1:3, the LPS binding by AMs reached 270% of control values. Second, we investigated the role of SP-A in regulating the degradation of LPS by AMs. In the presence of SP-A, deacylation of LPS by AMs increased by ∼2.3-fold. Pretreatment of AMs with phosphatidylinositol-specific phospholipase C had no effect on the SP-A-enhanced LPS binding but did reduce the amount of serum-enhanced LPS binding by 50%, suggesting that a cell surface molecule distinct from CD14 mediates the effect of SP-A. Together the results for the first time provide direct evidence that SP-A enhances LPS binding and degradation by AMs.

scholarly journals Rat surfactant protein D enhances the production of oxygen radicals by rat alveolar macrophages

1992 ◽  
Vol 286 (1) ◽  
pp. 5-8 ◽  
Author(s):  
J F Van Iwaarden ◽  
H Shimizu ◽  
P H M Van Golde ◽  
D R Voelker ◽  
L M G Van Golde
Keyword(s):  
Alveolar Macrophages ◽  
Oxygen Radicals ◽  
Protein A ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Surfactant Protein D ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Oxygen Radical Production ◽  
Stimulation Of

Rat surfactant protein D (SP-D) was shown to enhance the production of oxygen radicals by rat alveolar macrophages. This enhancement, which was determined by a lucigenin-dependent chemiluminescence assay, was maximal after 18 min at an SP-D concentration of 0.2 micrograms/ml. Surfactant lipids did not influence the stimulation of alveolar macrophages by SP-D, whereas the oxygen-radical production of these cells induced by surfactant protein A was inhibited by the lipids in a concentration-dependent manner.

SP-A1 and SP-A2 variants differentially enhance association ofPseudomonas aeruginosawith rat alveolar macrophages

2005 ◽  
Vol 288 (1) ◽  
pp. L150-L158 ◽  
Author(s):  
Anatoly N. Mikerov ◽  
Todd M. Umstead ◽  
Weixiong Huang ◽  
Wenlei Liu ◽  
David S. Phelps ◽  
...  
Keyword(s):  
Alveolar Macrophages ◽  
Single Gene ◽  
Protein A ◽  
Surfactant Protein ◽  
Phagocytic Index ◽  
Dependent Manner ◽  
Gene Products ◽  
Concentration Dependent Manner ◽  
Cell Association

Chronic airway inflammation caused by Pseudomonas aeruginosa is an important feature of cystic fibrosis (CF). Surfactant protein A (SP-A) enhances phagocytosis of P. aeruginosa. Two genes, SP-A1 and SP-A2, encode human SP-A. We hypothesized that genetically determined differences in the activity of SP-A1 and SP-A2 gene products exist. To test this, we studied association of a nonmucoid P. aeruginosa strain (ATCC 39018) with rat alveolar macrophages in the presence or absence of insect cell-expressed human SP-A variants. We used two trios, each consisting of SP-A1, SP-A2, and their coexpressed SP-A1/SP-A2 variants. We tested the 6A2and 6A4alleles (for SP-A1), the 1A0and 1A alleles (for SP-A2), and their respective coexpressed SP-A1/SP-A2 gene products. After incubation of alveolar macrophages with P. aeruginosa in the presence of the SP-A variants at 37°C for 1 h, the cell association of bacteria was assessed by light microscopy analysis. We found 1) depending on SP-A concentration and variant, SP-A2 variants significantly increased the cell association more than the SP-A1 variants (the phagocytic index for SP-A1 was ∼52–95% of the SP-A2 activity); 2) coexpressed variants at certain concentrations were more active than single gene products; and 3) the phagocytic index for SP-A variants was ∼18–41% of the human SP-A from bronchoalveolar lavage. We conclude that human SP-A variants in vitro enhance association of P. aeruginosa with rat alveolar macrophages differentially and in a concentration-dependent manner, with SP-A2 variants having a higher activity compared with SP-A1 variants.

scholarly journals Human surfactant protein A with two distinct oligomeric structures which exhibit different capacities to interact with alveolar type II cells

1996 ◽  
Vol 317 (3) ◽  
pp. 939-944 ◽  
Author(s):  
Akiko HATTORI ◽  
Yoshio KUROKI ◽  
Hitoshi SOHMA ◽  
Yoshinori OGASAWARA ◽  
Toyoaki AKINO
Keyword(s):  
Protein A ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Dependent Manner ◽  
Type Ii ◽  
Concentration Dependent Manner ◽  
Lower Affinity ◽  
Alveolar Proteinosis

The lung lavage fluids from patients with pulmonary alveolar proteinosis have been generally used as a source for human surfactant protein A (SP-A). We have recently found that a multimerized form of SP-A oligomer (alveolar proteinosis protein-I, APP-I) exists besides the normal-sized octadecamer (APP-II) in SP-As isolated from the patients. When analysed by Bio-Gel A15m column chromatography in 5 mM Tris buffer (pH 7.4), the apparent molecular masses of APP-I and APP-II were 1.65 MDa and 0.93 MDa, respectively. Gel-filtration analysis also revealed that APP-II is clearly separated from APP-I in the presence of 2 mM Ca2+ and 150 mM NaCl. We investigated the abilities of both SP-A oligomers to regulate phospholipid secretion and to bind to alveolar type II cells. Although APP-I inhibited lipid secretion, it was clearly a less effective inhibitor than APP-II. IC50 for inhibition of lipid secretion was apparently 0.23±0.08 µg/ml (0.14±0.05 nM) and 0.055±0.019 µg/ml (0.059±0.020 nM) for APP-I and APP-II, respectively. Both proteins bound to monolayers of type II cells in a concentration-dependent manner; however, APP-I clearly had a lower affinity to bind to type II cells. The apparent dissociation contants were, Kd = 2.31±0.70 µg/ml (1.40±0.43 nM) and 0.89±0.22 µg/ml (0.95±0.24 nM) for APP-I and APP-II, respectively. Excess unlabelled rat SP-A replaced 45% of 125I-APP-I and 77% of 125I-APP-II for type II cell binding. Although 125I-APP-II competed with excess unlabelled APP-I or APP-II, 125I-APP-I failed to compete and instead its binding rather increased in the presence of unlabelled APPs. The biotinylated APP-I bound to APP-I and APP-II coated on to microtitre wells in a concentration-dependent manner, indicating that APP-I interacts with APPs. This study demonstrates that the multimerized form of human SP-A oligomer exhibits the following attributes: (1) the reduced capacity to regulate phospholipid secretion from type II cells, and (2) lower affinity to bind to type II cells, and that the integrity of a flower-bouquet-like octadecameric structure of SP-A oligomer is important for the expression of full activity of this protein, indicating the importance of the oligomeric structure of mammalian lectins with collagenous domains.

Pulmonary surfactant protein A stimulates chemotaxis of alveolar macrophage

1993 ◽  
Vol 264 (4) ◽  
pp. L338-L344 ◽  
Author(s):  
J. R. Wright ◽  
D. C. Youmans
Keyword(s):  
Pulmonary Surfactant ◽  
Protein A ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Macrophage Migration ◽  
Dependent Manner ◽  
Directed Movement ◽  
Concentration Dependent Manner ◽  
Pulmonary Surfactant Protein ◽  
Checker Board

Pulmonary surfactant modulates several functions of alveolar macrophages including phagocytosis, killing, and chemotaxis. We hypothesized that the reported stimulatory effect of surfactant on macrophage migration was mediated by one of the surfactant proteins, SP-A. We found that macrophage migration was stimulated by SP-A in a concentration-dependent manner. A concentration of 105 micrograms SP-A/ml enhanced migration approximately 10-fold. Heat treatment or reduction and alkylation of SP-A reduced its stimulatory effect. A checker-board analysis showed that SP-A stimulated migration primarily by enhancing chemotaxis (directed movement) rather than chemokinesis (random movement). The interaction of SP-A with macrophages may be mediated at least partly by the collagen-like domain of SP-A. We speculate that SP-A may play a multifunctional role in regulating pulmonary immune response by stimulating multiple macrophage functions.

Role of clathrin-mediated endocytosis of surfactant protein A by alveolar macrophages in intracellular signaling

2009 ◽  
Vol 296 (3) ◽  
pp. L430-L441 ◽  
Author(s):  
Christina Moulakakis ◽  
Cordula Stamme
Keyword(s):  
Alveolar Macrophages ◽  
Intracellular Signaling ◽  
Protein A ◽  
Surfactant Protein ◽  
Kinase Assay ◽  
Coated Vesicle ◽  
Dependent Manner ◽  
Tnf Α

We recently provided evidence that anti-inflammatory macrophage activation, i.e., the inhibition of constitutive and signal-induced NF-κB activity by the pulmonary collectin surfactant protein (SP)-A, critically involves a promoted stabilization of IκB-α, the predominant inhibitor of NF-κB, via posttranscriptional mechanisms comprising the activation of atypical (a)PKCζ. SP-A uptake and degradation by alveolar macrophages (AMφ) occur in a receptor-mediated, clathrin-dependent manner. However, a mutual link between endocytosis of and signaling by SP-A remains elusive. The aim of this study was to investigate whether clathrin-mediated endocytosis (CME) of SP-A by AMφ is a prerequisite for its modulation of the IκB-α/NF-κB pathway. The inhibition of clathrin-coated pit (CCP) formation and clathrin-coated vesicle (CCV) formation/budding abrogates SP-A-mediated IκB-α stabilization and SP-A-mediated inhibition of LPS-induced NF-κB activation in freshly isolated rat AMφ, as determined by Western analysis, fluorescence-activated cell sorting, confocal microscopy, and EMSA. Actin depolymerization and inhibition of CCP formation further abolished SP-A-mediated inhibition of LPS-induced TNF-α release, as determined by ELISA. In addition, SP-A-induced atypical PKCζ activation was abolished by pretreatment of AMφ with CCV inhibitors as determined by in vitro immunocomplex kinase assay. Although CME is classically considered as a means to terminate signaling, our results demonstrate that SP-A uptake via CME by AMφ has to precede the initiation of SP-A signaling.

scholarly journals Surfactant protein A enhances alveolar macrophage phagocytosis of a live, mucoid strain ofP. aeruginosa

1999 ◽  
Vol 277 (4) ◽  
pp. L777-L786 ◽  
Author(s):  
William I. Mariencheck ◽  
Jordan Savov ◽  
Qun Dong ◽  
Michael James Tino ◽  
Jo Rae Wright
Keyword(s):  
Outer Membrane ◽  
Fluorescent Microscopy ◽  
Protein A ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Macrophage Phagocytosis ◽  
Live Bacteria ◽  
Membrane Components

In this study, we investigate the interaction between surfactant protein A (SP-A) and a live, mucoid strain of Pseudomonas aeruginosa and identify a mechanism of clearance of this organism by alveolar macrophages.125I-labeled SP-A bound live, but not heat-killed, P. aeruginosaorganisms in a concentration-dependent manner. Unlabeled SP-A bound live bacteria, protein isolated from whole organisms, and specific proteins of the P. aeruginosa outer membrane. The binding of SP-A to P. aeruginosa and outer membrane components was inhibited by either EDTA or mannose. Phagocytosis assays with fluorescent microscopy demonstrated that the percentage of macrophages with internalized FITC-labeled P. aeruginosa was increased 1.8-fold (19 vs. 35%) by pretreating the live bacteria with SP-A. This finding was confirmed by direct visualization of ingested bacteria by electron microscopy. Adhering macrophages to SP-A-coated surfaces attenuated the increased uptake of P. aeruginosa pretreated with SP-A, suggesting that SP-A acts as an opsonin to stimulate macrophage phagocytosis of this strain of P. aeruginosa.

Lamellar body membrane turnover is stimulated by secretagogues

2000 ◽  
Vol 278 (3) ◽  
pp. L443-L452 ◽  
Author(s):  
Sandra R. Bates ◽  
Jian-Qin Tao ◽  
Susanne Schaller ◽  
Aron B. Fisher ◽  
Henry Shuman
Keyword(s):  
Protein A ◽  
Lamellar Body ◽  
Surfactant Protein ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Dependent Manner ◽  
Type Ii ◽  
Lamellar Bodies ◽  
Membrane Turnover ◽  
Concentration Dependent Manner

Lamellar bodies are specialized cellular organelles used for storage of surfactant by alveolar type II cells of the lung. We utilized monoclonal antibody (MAb) 3C9, which recognizes an integral lamellar body-limiting membrane protein of 180 kDa, to follow lamellar body trafficking. 125I-labeled MAb 3C9 bound to the surface of type II cells and was internalized by the cells in a time- and concentration-dependent manner that was inhibitable by excess unlabeled antibody. The internalized antibody remained undegraded over a 4-h time period. The L2 rat lung cell line that does not have lamellar bodies did not bind iodinated 3C9. Exposure of type II cells to the secretagogues ATP, phorbol 12-myristate 13-acetate, and cAMP resulted in a 1.5- to 2-fold enhancement of binding and uptake of MAb 3C9. Calphostin C inhibited phorbol 12-myristate 13-acetate-stimulated phospholipid secretion and also reduced binding and uptake of MAb 3C9 by type II cells. Treatment of type II cells with phenylarsine oxide to obstruct clathrin-mediated endocytosis had no effect on the internalization of MAb 3C9 while markedly blocking the uptake of surfactant protein A and transferrin. An actin-mediated process was important for lamellar body membrane uptake because incubation with cytochalasin D partially inhibited MAb 3C9 incorporation by type II cells. These studies are compatible with enhanced lamellar body membrane turnover associated with surfactant secretion and indicate that this process can be monitored by the trafficking of the antigen reporter MAb 3C9.

scholarly journals Regulation of Protein Phosphorylation and Pathogen Phagocytosis by Surfactant Protein A

1999 ◽  
Vol 67 (9) ◽  
pp. 4693-4699 ◽  
Author(s):  
Trista L. Schagat ◽  
Michael James Tino ◽  
Jo Rae Wright
Keyword(s):  
Protein Phosphorylation ◽  
Alveolar Macrophages ◽  
Actin Polymerization ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein A ◽  
Sodium Dodecyl ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Dependent Manner ◽  
Cell Functions

ABSTRACT Surfactant protein A (SP-A), a pulmonary member of the collectin family of proteins, facilitates the rapid clearance of pathogens by upregulating immune cell functions in the lungs. SP-A binds to bacteria and targets them for rapid phagocytosis by alveolar macrophages, but the mechanism by which this stimulation occurs is not clear. To characterize the intracellular events that may be involved, we examined the roles of protein phosphorylation and cytoskeletal polymerization in SP-A-stimulated phagocytosis. In rat alveolar macrophages, SP-A stimulated rapid tyrosine phosphorylation of specific proteins in a dose- and time-dependent manner. The pattern of proteins that were phosphorylated in response to SP-A, as resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was similar to that observed for immunoglobulin G (IgG)-stimulated macrophages. Both SP-A and IgG stimulated increases in phagocytosis of Streptococcus pneumoniae above levels in the absence of added protein by 394% ± 81% and 200% ± 25%, respectively. Phagocytosis in both cases was dependent on tyrosine kinases, protein kinase C, and actin polymerization but not on microtubule activity. These studies show that SP-A utilizes pathways similar to those used by IgG to increase macrophage phagocytosis of bacteria.

Killing ofKlebsiella pneumoniaeby human alveolar macrophages

2002 ◽  
Vol 282 (5) ◽  
pp. L944-L956 ◽  
Author(s):  
Judy M. Hickman-Davis ◽  
Philip O'Reilly ◽  
Ian C. Davis ◽  
Janos Peti-Peterdi ◽  
Glenda Davis ◽  
...  
Keyword(s):  
Alveolar Macrophages ◽  
Protein A ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Acetoxymethyl Ester ◽  
Free System ◽  
Colony Forming Units ◽  
Human Alveolar Macrophages ◽  
Intracellular Ca ◽  
A Cell

We investigated putative mechanisms by which human surfactant protein A (SP-A) effects killing of Klebsiella pneumoniae by human alveolar macrophages (AMs) isolated from bronchoalveolar lavagates of patients with transplanted lungs. Coincubation of AMs with human SP-A (25 μg/ml) and Klebsiella resulted in a 68% decrease in total colony forming units by 120 min compared with AMs infected with Klebsiella in the absence of SP-A, and this SP-A-mediated effect was abolished by preincubation with NG-monomethyl-l-arginine. Incubation of transplant AMs with SP-A increased intracellular Ca2+concentration ([Ca2+]i) by 70% and nitrite and nitrate (NOx) production by 45% (from 0.24 ± 0.02 to 1.3 ± 0.21 nmol · 106AMs−1· h−1). Preincubation with 1,2-bis(2-aminophenoxy)ethane- N,N,N′,N′-tetraacetic acid-acetoxymethyl ester inhibited the increase in [Ca2+]iand abrogated the SP-A-mediated Klebsiella phagocytosis and killing. In contrast, incubation of AMs from normal volunteers with SP-A decreased both [Ca2+]iand NOxproduction and did not result in killing of Klebsiella. Significant killing of Klebsiella was also seen in a cell-free system by sustained production of peroxynitrite (>1 μM/min) at pH 5 but not at pH 7.4. These findings indicate that SP-A mediates pathogen killing by AMs from transplant lungs by stimulating phagocytosis and production of reactive oxygen-nitrogen intermediates.

scholarly journals Brassinolide Enhances the Level of Brassinosteroids, Protein, Pigments, and Monosaccharides in Wolffia arrhiza Treated with Brassinazole

Plants ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1311
Author(s):  
Magdalena Chmur ◽  
Andrzej Bajguz
Keyword(s):  
Plant Growth ◽  
Growth And Development ◽  
Inhibitory Effect ◽  
Dependent Manner ◽  
Plant Growth And Development ◽  
Concentration Dependent Manner ◽  
Spectrophotometric Methods ◽  
Synthetic Inhibitor ◽  
Wolffia Arrhiza ◽  
First Time

Brassinolide (BL) represents brassinosteroids (BRs)—a group of phytohormones that are essential for plant growth and development. Brassinazole (Brz) is as a synthetic inhibitor of BRs’ biosynthesis. In the present study, the responses of Wolffia arrhiza to the treatment with BL, Brz, and the combination of BL with Brz were analyzed. The analysis of BRs and Brz was performed using LC-MS/MS. The photosynthetic pigments (chlorophylls, carotenes, and xanthophylls) levels were determined using HPLC, but protein and monosaccharides level using spectrophotometric methods. The obtained results indicated that BL and Brz influence W. arrhiza cultures in a concentration-dependent manner. The most stimulatory effects on the growth, level of BRs (BL, 24-epibrassinolide, 28-homobrassinolide, 28-norbrassinolide, catasterone, castasterone, 24-epicastasterone, typhasterol, and 6-deoxytyphasterol), and the content of pigments, protein, and monosaccharides, were observed in plants treated with 0.1 µM BL. Whereas the application of 1 µM and 10 µM Brz caused a significant decrease in duckweed weight and level of targeted compounds. Application of BL caused the mitigation of the Brz inhibitory effect and enhanced the BR level in duckweed treated with Brz. The level of BRs was reported for the first time in duckweed treated with BL and/or Brz.

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