Inhibition of lung calcium-independent phospholipase A2 by surfactant protein A

1994 ◽  
Vol 267 (3) ◽  
pp. L335-L341 ◽  
Author(s):  
A. B. Fisher ◽  
C. Dodia ◽  
A. Chander
Keyword(s):  
Phospholipase A2 ◽  
Chemical Reduction ◽  
Lung Surfactant ◽  
Protein A ◽  
Lamellar Body ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Lamellar Bodies ◽  
Pla2 Activity ◽  
Body Protein

The effect of lung surfactant protein A (SP-A) on lung phospholipase A2 (PLA2) activity was investigated. SP-A was purified from bovine surfactant obtained by lung lavage. PLA2 was assayed using radiolabeled 1,2-dipalmitoyl phosphatidylcholine (DPPC) in surfactant-like unilamellar liposomes with Ca(2+)-free acidic (pH 4) or 10 mM Ca2+, alkaline (pH 8.5) buffer. SP-A significantly inhibited Ca(2+)-independent acidic PLA2 of rat lung homogenate or isolated lamellar bodies but had no effect on the Ca(2+)-dependent alkaline enzyme. Lamellar body PLA2 was inhibited by 50% with 0.25 micrograms SP-A/microgram lamellar body protein. Similar inhibition by SP-A was observed when 1-palmitoyl,2-oleoyl PC (POPC) was the substrate. Binding assay showed binding of 125I-labeled SP-A to DPPC but not to POPC, indicating that removal of substrate was not the mechanism for inhibition of the enzyme by SP-A. Chemical reduction or alkylation of SP-A abolished its inhibitory effect on PLA2 activity. Inactivation of endogenous SP-A in isolated lamellar bodies or surfactant increased Ca(2+)-independent PLA2 activity in these fractions. The presence of SP-A in liposomes stimulated the uptake of DPPC by isolated granular pneumocytes in primary culture but significantly inhibited its degradation. These results indicate that the Ca(2+)-independent acidic PLA2 has a role in the metabolism of internalized surfactant phospholipid and that SP-A can modulate the activity of this enzyme.

scholarly journals Immunocytochemical localization of lysozyme and surfactant protein A in rat type II cells and extracellular surfactant forms.

1992 ◽  
Vol 40 (10) ◽  
pp. 1491-1500 ◽  
Author(s):  
E M Haller ◽  
S A Shelley ◽  
M R Montgomery ◽  
J U Balis
Keyword(s):  
Lung Surfactant ◽  
Protein A ◽  
Lamellar Body ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Peripheral Compartment ◽  
Type Ii Cells ◽  
Type Ii ◽  
Lamellar Bodies ◽  
Body Protein

Using immunogold labeling of fixed, cryosubstituted tissue sections, we compared the distribution of lysozyme, an oxidant-sensitive lamellar body protein, with that of surfactant protein A (SP-A) in rat Type II cells, extracellular surfactant forms, and alveolar macrophages. Morphometric analysis of gold particle distribution revealed that lysozyme and SP-A were present throughout the secretory and endosomal pathways of Type II cells, with prominent localization of lysozyme in the peripheral compartment of lamellar bodies. All extracellular surfactant forms were labeled for both proteins with preferential labeling of tubular myelin and unilamellar vesicles. Labeling of tubular myelin for SP-A was striking when compared with that of lamellar bodies and other extracellular surfactant forms. Lamellar body-like forms and multilamellar structures were uniformly labeled for lysozyme, suggesting that this protein is rapidly redistributed within these forms after secretion of lysozyme-laden lamellar bodies. By contrast, increased labeling for SP-A was observed over peripheral membranes of lamellar body-like forms and multilamellar structures, apparently reflecting progressive SP-A enrichment of these membranes during tubular myelin formation. The results indicate that lysozyme is an integral component of the lamellar body peripheral compartment and secreted surfactant membranes, and support the concept that lysozyme may participate in the structural organization of lung surfactant.

Pulmonary surfactant protein A-mediated uptake of phosphatidylcholine by alveolar type II cells

1993 ◽  
Vol 265 (2) ◽  
pp. L193-L199 ◽  
Author(s):  
A. Tsuzuki ◽  
Y. Kuroki ◽  
T. Akino
Keyword(s):  
Pulmonary Surfactant ◽  
Protein A ◽  
Lamellar Body ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Type Ii ◽  
Lamellar Bodies ◽  
Pulmonary Surfactant Protein

Pulmonary surfactant protein A (SP-A)-mediated uptake of phosphatidylcholine (PC) by alveolar type II cells was investigated. SP-A enhanced the uptake of liposomes containing dipalmitoylphosphatidylcholine (DPPC), 1-palmitoyl-2-linoleoyl phosphatidylcholine (PLPC), or 1,2-dihexadecyl-sn-glycero-3-phosphocholine (DPPC-ether), a diether analogue of DPPC, but about twice as much DPPC was taken up by type II cells as PLPC or DPPC-ether. When subcellular distribution was analyzed, 51.3 +/- 2.9% (mean +/- SD, n = 3) of cell-associated radiolabeled DPPC was recovered in the lamellar body-rich fraction in the presence of SP-A, whereas only 19.3 +/- 1.9% (mean +/- SD, n = 3) was found to this fraction in the absence of SP-A. When type II cells were incubated either with DPPC at 0 degree C or with DPPC-ether at 37 degrees C, or no cells were included, low proportions of the cell-associated lipids were present in the fractions corresponding to lamellar bodies even in the presence of SP-A. Anti-SP-A antibody significantly reduced the radioactivity incorporated into the lamellar body fraction. Phosphatidylcholine that had been incorporated into lamellar bodies remained largely intact when SP-A was present. Subcellular fractionations of type II cells with radiolabeled SP-A and DPPC revealed that the sedimentation characteristics of cell-associated SP-A are different from those of DPPC, although a small broad peak of radiolabeled SP-A was found in the lamellar body fraction.(ABSTRACT TRUNCATED AT 250 WORDS)

Surfactant protein A metabolism in preterm ventilated lambs

1992 ◽  
Vol 262 (6) ◽  
pp. L765-L772 ◽  
Author(s):  
M. Ikegami ◽  
J. F. Lewis ◽  
B. Tabor ◽  
E. D. Rider ◽  
A. H. Jobe
Keyword(s):  
Gestational Age ◽  
Protein A ◽  
Lamellar Body ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Lamellar Bodies ◽  
Specific Activities ◽  
Kinetics Of

Surfactant protein A (SP-A) metabolism was studied in vivo in 33 preterm ventilated lambs at 138 +/- 1 days gestational age by measuring recoveries of exogenously administered surfactant containing both radiolabeled SP-A and labeled saturated phosphatidylcholine (Sat PC) given via the trachea at birth. Endogenously secreted SP-A was also labeled with [35S]methionine and followed over 24 h. The exogenously labeled SP-A left the alveolar pool more rapidly than did Sat PC over the first 5 h of life (P less than 0.05), and both exogenously labeled SP-A and Sat PC were detected within lamellar bodies by 2 h, indicating uptake from the airspaces. The quantity of SP-A in alveolar washes increased about twofold from birth to 5 h of age, whereas alveolar Sat PC pools were constant over 24 h. The SP-A endogenously labeled with [35S]methionine was recovered at highest specific activities in the alveolar washes at 10 and 45 min after birth with no labeled SP-A detectable in lamellar body fractions until 2 h. The curve for endogenous SP-A labeling of lamellar bodies was similar to that for exogenous labeling, indicating that SP-A was initially secreted by a pathway independent of lamellar bodies with subsequent SP-A labeling of lamellar bodies. The kinetics of SP-A metabolism were very different than for Sat PC in preterm lambs.

ATP and adenosine 3',5'-cyclic monophosphate stimulate the synthesis of surfactant protein A in rat lung

1993 ◽  
Vol 264 (5) ◽  
pp. L431-L437 ◽  
Author(s):  
A. Wali ◽  
M. F. Beers ◽  
C. Dodia ◽  
S. I. Feinstein ◽  
A. B. Fisher
Keyword(s):  
Total Protein ◽  
Protein A ◽  
Lamellar Body ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Enzyme Linked Immunosorbent Assay ◽  
Translational Efficiency ◽  
Rat Lung ◽  
Body Protein ◽  
Cyclic Monophosphate

Synthesis and secretion of surfactant protein A (SP-A) were studied in the isolated perfused rat lung using Trans35S-label (approximately 85% methionine, 15% cysteine) in the perfusate with or without 1 mM ATP or 0.1 mM 8-bromoadenosine 3',5',-cyclic monophosphate (8-BrcAMP) for up to 6 h of perfusion. By enzyme-linked immunosorbent assay, the SP-A content was 36 +/- 0.3% of total protein in extracellular surfactant and 10.8 +/- 1.9% of total protein in lamellar bodies of control lungs; these relativr proportions were maintained in the presence of ATP or 8-BrcAMP. Incorporation of [35S]methionine (cysteine) into the surfactant and lamellar body protein fraction could be detected at 4 h of perfusion. At 6 h, specific activity of total protein [disintegrations per minute (dpm)/micrograms)] was significantly increased in both the surfactant (54%) and lamellar body fractions (30%) under the influence of either secretagogue compared with control conditions. In the presence of ATP, there was a significant increase in the SP-A immunoprecipitable counts of 61 and 72% in extra- and intracellular compartments, respectively. However, no significant change was observed in the relative abundance of SP-A mRNA between control and secretagogue-treated lungs. This dissociation of SP-A mRNA abundance and label incorporation into protein indicates that alteration in translational efficiency or posttranslational factors may be involved in the secretagogue-induced stimulation of SP-A synthesis.

scholarly journals Pathway to lamellar bodies for surfactant protein A

2010 ◽  
Vol 299 (1) ◽  
pp. L51-L58 ◽  
Author(s):  
Aron B. Fisher ◽  
Chandra Dodia ◽  
Peter Ruckert ◽  
Jian-Qin Tao ◽  
Sandra R. Bates
Keyword(s):  
Specific Activity ◽  
Protein A ◽  
Lamellar Body ◽  
Lavage Fluid ◽  
Surfactant Protein ◽  
Lung Lavage ◽  
Surfactant Protein A ◽  
Coated Vesicle ◽  
Lamellar Bodies ◽  
Vesicle Budding

Alveolar surfactant protein A (SP-A) is endocytosed by type II epithelial cells through clathrin-dependent uptake and targeted to lamellar bodies for resecretion. However, the mechanism for secretion of newly synthesized SP-A, whether regulated exocytosis of lamellar bodies or constitutive secretion, is unresolved. If it is the latter, lamellar body SP-A would represent endocytosed protein. Amantadine, an inhibitor of clathrin-coated vesicle budding, was used to evaluate the role of endocytosis in accumulation of SP-A in lamellar bodies. In isolated rat lungs, amantadine (10 mM) inhibited uptake of endotracheally instilled 35S-labeled biosynthesized surfactant proteins by >80%. To study trafficking of newly synthesized SP-A, lungs were perfused for up to 6 h with [35S]methionine, and surfactant was isolated from lung lavage fluid and lamellar bodies were isolated from lung homogenate. With control lungs, the mean specific activity of [35S]SP-A (disintegrations per minute per microgram of SP-A) increased linearly with time of perfusion: it was significantly higher in isolated lamellar bodies than in surfactant and was increased in both compartments by 50–60% in the presence of 0.1 mM 8-bromo-cAMP. These results suggest a precursor-product relationship between lamellar body and extracellular [35S]SP-A. Specific activities in both compartments were unaffected by addition of amantadine (10 mM) to the lung perfusate, indicating that uptake from the alveolar space was not responsible for the increase in lamellar body [35S]SP-A. Thus the pathway for secretion of newly synthesized SP-A is by transfer from the site of synthesis to the storage/secretory organelle prior to lamellar body exocytosis.

Trafficking of surfactant protein A in fetal rabbit lung in organ culture

1993 ◽  
Vol 264 (1) ◽  
pp. L27-L35 ◽  
Author(s):  
J. L. Alcorn ◽  
C. R. Mendelson
Keyword(s):  
Organ Culture ◽  
Protein A ◽  
Lamellar Body ◽  
Fetal Lung ◽  
Surfactant Protein ◽  
Surfactant Protein A ◽  
Major Protein ◽  
Lamellar Bodies ◽  
Rabbit Lung ◽  
Tissue Homogenates

Surfactant protein A (SP-A), a major protein component of pulmonary surfactant, is a developmentally and hormonally regulated sialoglycoprotein expressed in type II pneumonocytes. Surfactant proteins and glycerophospholipids are transported to multilamellar structures termed lamellar bodies, which serve to store surfactant lipoprotein until secretion by exocytosis into the alveolar lumen. The cellular mechanism(s) for targeting of SP-A and other surfactant components to lamellar bodies is unknown. In the present study, we have investigated the transport of SP-A to lamellar bodies in fetal rabbit lung tissue in organ culture using pulse-chase analysis of [35S]-methionine-labeled SP-A protein. SP-A accumulated in lamellar bodies within 1–3 h of synthesis; lamellar body SP-A was found to be endoglycosidase H resistant and represented 30–40% of the radiolabeled SP-A recovered from the tissue for periods of up to 12 h postlabeling. Based on our estimates of lamellar body recovery from tissue homogenates, lamellar body-associated SP-A may account for 60–80% of the SP-A present in the fetal lung explants. Treatment of fetal rabbit lung explants with inhibitors of oligosaccharide addition (tunicamycin) and processing (castanospermine), which act within the endoplasmic reticulum, significantly reduced the rate of transport of newly synthesized SP-A to lamellar bodies. An inhibitor of oligosaccharide processing that acts on a processing step that takes place within the Golgi apparatus (swainsonine) reduced the rate of transport of radiolabeled SP-A to lamellar bodies by approximately 50%.(ABSTRACT TRUNCATED AT 250 WORDS)

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