Turnover of extracellular matrix by type II pulmonary epithelial cells

1995 ◽  
Vol 268 (2) ◽  
pp. L336-L346 ◽  
Author(s):  
S. E. Dunsmore ◽  
D. E. Rannels
Keyword(s):  
Extracellular Matrix ◽  
Epithelial Cells ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Alveolar Epithelium ◽  
Kinetic Studies ◽  
Type Ii Cells ◽  
Type Ii ◽  
Pulmonary Epithelial Cells ◽  
Matrix Stability

Rat type II pulmonary epithelial cells synthesize and assemble a multicomponent extracellular matrix (ECM) which can modulate cellular differentiation in primary culture. This study defines turnover of the type II cell matrix. Turnover kinetics were analyzed in two types of pulse-chase protocols based on loss of radioactive ECM components. To estimate turnover of previously synthesized ECM, type II cells were plated on extracted matrix that was radiolabeled 2, 3, or 6 days; alternatively, ECM was radiolabeled in pulse-chase experiments to measure turnover by the same cells that synthesized the matrix. Rapid initial rates of ECM turnover were evident in both cases. While overall matrix stability appeared to change with culture time, sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed a similar spectrum of proteins in the ECM over the course of kinetic studies. The results reveal rapid turnover of ECM by type II cells and suggest that matrix stability may be regulated. These observations provide a basis for future investigations of the physiological significance of turnover of individual ECM components by the alveolar epithelium.

Adrenal hormone regulation of fibronectin synthesis by type II pulmonary epithelial cells

1997 ◽  
Vol 273 (1) ◽  
pp. L86-L92
Author(s):  
S. E. Dunsmore ◽  
Y. C. Lee ◽  
D. E. Rannels
Keyword(s):  
Extracellular Matrix ◽  
Epithelial Cells ◽  
Cell Phenotype ◽  
Type Ii Cells ◽  
Mrna Levels ◽  
Hormone Regulation ◽  
Type Ii ◽  
Adrenal Hormone ◽  
Pulmonary Epithelial Cells ◽  
Type Ii Cell

Previous work suggested an association between changes in the alveolar extracellular matrix and epithelial cell growth in lungs of adrenalectomized rats in vivo. Other studies demonstrated that adrenal hormones modulate extracellular matrix synthesis by type II pulmonary epithelial cells in vitro. Adrenal hormone regulation of type II cell fibronectin synthesis was thus examined. Fibronectin synthesis was quantitated by immunoprecipitation of the metabolically labeled molecule from cells, extracellular matrix, and culture medium. On day 1 of primary culture, synthesis of matrix fibronectin by type II cells isolated from the lungs of adrenalectomized animals was more than twice that by cells isolated from control rats. Adrenalectomy elevated steady-state fibronectin mRNA levels in primary isolates of type II cells cultured for 1 or 3 days. These results suggest that altered fibronectin synthesis and deposition into the extracellular matrix accompany changes in type II cell phenotype that occur after adrenalectomy.

Extracellular matrix synthesis and turnover by type II pulmonary epithelial cells

1992 ◽  
Vol 262 (5) ◽  
pp. L582-L589 ◽  
Author(s):  
D. E. Rannels ◽  
S. E. Dunsmore ◽  
R. N. Grove
Keyword(s):  
Extracellular Matrix ◽  
Steady State ◽  
Epithelial Cells ◽  
Primary Cultures ◽  
Calf Serum ◽  
Type I ◽  
Type Ii Cells ◽  
Type Ii ◽  
Pulmonary Epithelial Cells ◽  
Type Ii Cell

Both type I and type II pulmonary epithelial cells contact the extracellular matrix (ECM). Type II cell-ECM interactions are bidirectional; they involve matrix-mediated modulation of type II cell differentiation, as well as cellular synthesis and deposition of ECM components. The present experiments examine the kinetics of accumulation of newly synthesized proteins in cell and matrix fractions from primary cultures of type II pneumocytes. Cycloheximide-sensitive incorporation of [3H]leucine into total protein of both the cell and ECM fractions was linear for 24–30 h, when steady-state labeling was reached and maintained to at least day 8. Over this interval, the cells enlarged but did not divide. Newly synthesized proteins recovered in the matrix fraction averaged 1–2% of those in the cells. Relative rates of radiolabeling of matrix proteins peaked at culture day 2 and increased in the absence of serum. In short-pulse studies, initial rates of protein synthesis were equal on culture days 1 and 3; this suggested that the steady-state labeling kinetics above reflected protein turnover. This was supported by rapid loss of radioactivity from the ECM after fresh type II cells were seeded on a prelabeled, cell-free matrix surface. Fresh or conditioned Dulbecco's modified Eagle's medium containing 10% fetal calf serum had little effect on matrix stability. These results demonstrate regulated deposition and turnover of a complex ECM by type II cells and provide a basis for further investigations of factors that control these processes.

Primary translation products of pulmonary surfactant protein D

1991 ◽  
Vol 260 (4) ◽  
pp. L247-L253 ◽  
Author(s):  
E. Crouch ◽  
K. Rust ◽  
A. Persson ◽  
W. Mariencheck ◽  
M. Moxley ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Surfactant Protein ◽  
Proteolytic Processing ◽  
Surfactant Protein D ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Secretory Product ◽  
Type Ii

Surfactant protein D (SP-D) is a collagenous, surfactant-associated, carbohydrate-binding protein that is synthesized by alveolar type II epithelial cells. To further characterize SP-D, we isolated RNA from adult rat lungs and rat type II cells and translated mRNAs in vitro. [35S]methionine-labeled translation products were precipitated with antibodies to rat SP-D, resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and visualized by fluorography. Immune precipitates of translation reactions for rat lung or rat type II cells demonstrated a single collagenous polypeptide (39.3 kDa) that was smaller than surfactant-associated SP-D (43 kDa, reduced) but larger than the mature secreted form of rat SP-A. This component was not identified in translation reactions of rat liver, gut, brain, mammary gland, or rat L2 cell RNA. There was a fivefold enrichment of SP-D mRNA in freshly isolated type II cells relative to lung; however, the levels of translatable SP-D mRNA decreased rapidly during the first 24 h of cell culture. The SP-D translation product migrated faster than the major cellular form of SP-D but approximately 1 kDa slower than cellular SP-D synthesized in the presence of 2,2'-dipyridyl plus tunicamycin. Translation in the presence of canine pancreatic microsomes gave a single glycosylated, endoglycosidase F-sensitive form (40.6 kDa) and demonstrated cleavage of a small signal peptide. These results indicate that SP-D is a secretory product of differentiated type II epithelial cells and that SP-D is secreted in a mature form that does not undergo further proteolytic processing in vivo.

Composition of extracellular matrix of type II pulmonary epithelial cells in primary culture

1995 ◽  
Vol 269 (6) ◽  
pp. L754-L765 ◽  
Author(s):  
S. E. Dunsmore ◽  
C. Martinez-Williams ◽  
R. A. Goodman ◽  
D. E. Rannels
Keyword(s):  
Extracellular Matrix ◽  
Epithelial Cells ◽  
Primary Culture ◽  
Biological Effects ◽  
Primary Cultures ◽  
Matrix Composition ◽  
Type Ii Cells ◽  
Type Ii ◽  
Pulmonary Epithelial Cells ◽  
Culture Surface

Type II pulmonary epithelial cells in primary culture synthesize and deposit an extracellular matrix which has reciprocal biological effects on cellular differentiation. The present work establishes conditions for metabolic labeling of matrix constituents and for separation of cells from the associated matrix; it also defines matrix composition, which does not appear to change qualitatively between days 2 and 6 of primary culture. Type II cells synthesize and deposit a spectrum of radiolabeled components on the culture surface. These include fibronectin, laminin, type IV collagen, and plasminogen activator inhibitor-1, along with additional unidentified proteins. Few radiolabeled proteins in medium conditioned by type II cells bind nonspecifically to the culture surface in the absence of cells. Fibroblasts and macrophages, which may contaminate the primary cultures, do not appear to contribute substantially to the type II cell matrix. These results demonstrate that type II cells synthesize and deposit a complex multicomponent extracellular matrix. The work provides a basis for further investigations of bidirectional interactions between type II cells and the extracellular matrix.

Roles of SP-A, SP-B, and SP-C in modulation of lipid uptake by pulmonary epithelial cells in vitro

1996 ◽  
Vol 270 (1) ◽  
pp. L69-L79 ◽  
Author(s):  
A. D. Horowitz ◽  
B. Moussavian ◽  
J. A. Whitsett
Keyword(s):  
Epithelial Cells ◽  
Lipid Vesicles ◽  
Type Ii Cells ◽  
Type Ii ◽  
Lipid Uptake ◽  
3T3 Cells ◽  
Alveolar Cell ◽  
Pulmonary Epithelial Cells ◽  
Nih 3T3

The effects of the surfactant proteins (SP)-A, SP-B, and SP-C on binding and endocytosis of fluorescently labeled lipid vesicles were studied in rat type II epithelial cells and in MLE-12 cells, a pulmonary adenocarcinoma cell line with alveolar cell characteristics. Incorporation of SP-C in lipid vesicles markedly stimulated binding to the cell membrane at 4 degrees C and endocytosis of lipids at 37 degrees C. SP-C enhanced lipid uptake in MLE-12 cells, type II cells, and NIH 3T3 cells. SP-B stimulated lipid uptake in MLE-12 cells, but to a lesser degree. SP-B decreased the amount of lipid uptake stimulated by SP-C, SP-A did not increase endocytosis of lipids by MLE-12 cells or type II cells, but aggregates of lipid were observed associated with the cell surface in the presence of SP-A. Maintenance of active surfactant in the lung may be achieved through the selective uptake and degradation of surfactant subfractions depleted in SP-A and SP-B.

Expression of glycogen phosphorylase isozymes in developing rat lung

1997 ◽  
Vol 273 (2) ◽  
pp. L389-L394
Author(s):  
S. R. Rannels ◽  
L. Liu ◽  
T. E. Weaver
Keyword(s):  
Enzyme Activity ◽  
Epithelial Cells ◽  
Glycogen Phosphorylase ◽  
Alveolar Epithelium ◽  
Fetal Lung ◽  
Ribonuclease Protection Assay ◽  
Type Ii Cells ◽  
Type Ii ◽  
Adult Type ◽  
The Brain

Glycogen accumulates to significant levels in epithelial cells of the developing respiratory tract. Mobilization of glycogen stores is regulated differentially along the respiratory epithelium such that glycogenolysis in the alveolar epithelium (the site of surfactant synthesis) precedes that in the bronchial and bronchiolar epithelium. The initial step in glycogen degradation is catalyzed by glycogen phosphorylase, which exists as three genetically distinct isozymes referred to as muscle, liver, and brain isoforms. The goal of this study was to characterize the temporal and spatial expression of each of the glycogen phosphorylase isozymes in developing lung to determine which isoform(s) was associated with glycogen mobilization in the fetal type II epithelial cell. RNA levels encoding glycogen phosphorylase were assessed by ribonuclease protection assay using isoform-specific antisense probes. RNAs encoding the brain and liver isozymes were detected in isolated day 20 fetal type II epithelial cells and at lower levels in adult type II cells. The muscle isoform RNA was barely detectable in fetal type II cells and was undetectable in adult type II cells. Expression of brain and liver isoform RNAs was higher in whole fetal lung than in fetal type II cells. Consistent with this result, in situ hybridization studies demonstrated widespread expression of the brain and liver isoforms in developing lung tissues; in contrast, expression of the muscle isoform was restricted to the pulmonary vein. Glycogen phosphorylase enzyme activity corresponding to the brain isoform was clearly detected in isolated fetal type II cells; however, the majority of enzyme activity migrated as two bands with distinct electrophoretic mobilities that may have been the result of isoform heterodimerization. Collectively, these results suggest that the brain and liver isoforms of glycogen phosphorylase may be involved in mobilization of type II cell glycogen during late fetal lung development.

Synthesis of fibronectin and laminin by type II pulmonary epithelial cells

1996 ◽  
Vol 270 (2) ◽  
pp. L215-L223 ◽  
Author(s):  
S. E. Dunsmore ◽  
Y. C. Lee ◽  
C. Martinez-Williams ◽  
D. E. Rannels
Keyword(s):  
Extracellular Matrix ◽  
Epithelial Cells ◽  
Primary Culture ◽  
Messenger Rna ◽  
Type Ii ◽  
Matrix Deposition ◽  
Pulmonary Epithelial Cells ◽  
The Matrix ◽  
Average Proportion ◽  
Type Ii Cell

Previous investigations demonstrated that type II pulmonary epithelial cells regulate extracellular matrix deposition as a function of time in primary culture. In those studies, the matrix fraction was analyzed as a whole. The present work focused on two components of the type II cell matrix, fibronectin and laminin. These glycoproteins have differing effects on differentiation of type II cells in primary culture. Fibronectin synthesis was quantitated between day 1 and day 6 in the cells, matrix, and medium; laminin synthesis was quantitated only in the cells. Although total fibronectin synthesis was regulated as a function of time in culture, reaching its greatest value on day 2, the average proportion of newly synthesized fibronectin in the cells (35%), medium (50%), and matrix (15%) remained constant over a 6-day interval. Between day 2 and day 6, the relative abundance of fibronectin messenger RNA increased 6.5-fold. Rates of cellular laminin synthesis did not vary with time in culture. These results support a role for differential regulation of fibronectin and laminin synthesis to determine the composition of the type II cell extracellular matrix.

Cell-specific expression of human HGF by alveolar type II cells induces remodeling of septal wall tissue in the lung: a morphometric study

2012 ◽  
Vol 113 (5) ◽  
pp. 799-807 ◽  
Author(s):  
Alexandra Leuenberger ◽  
Amiq Gazdhar ◽  
Gudrun Herrmann ◽  
Matthias Ochs ◽  
Thomas Geiser ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Gene Transfer ◽  
Surface Area ◽  
Alveolar Epithelium ◽  
Elastic Fibers ◽  
Type Ii Cells ◽  
Type Ii ◽  
Specific Expression ◽  
Alveolar Epithelial ◽  
Healthy Lung

Hepatocyte growth factor (HGF) is involved in development and regeneration of the lungs. Human HGF, which was expressed specifically by alveolar epithelial type II cells after gene transfer, attenuated the bleomycin-induced pulmonary fibrosis in an animal model. As there are also regions that appear morphologically unaffected in fibrosis, the effects of this gene transfer to normal lungs is of interest. In vitro studies showed that HGF inhibits the formation of the basal lamina by cultured alveolar epithelial cells. Thus we hypothesized that, in the healthy lung, cell-specific expression of HGF induces a remodeling within septal walls. Electroporation of a plasmid of human HGF gene controlled by the surfactant protein C promoter was applied for targeted gene transfer. Using design-based stereology at light and electron microscopic level, structural alterations were analyzed and compared with a control group. HGF gene transfer increased the volume of distal air spaces, as well as the surface area of the alveolar epithelium. The volume of septal walls, as well as the number of alveoli, was unchanged. Volumes per lung of collagen and elastic fibers were unaltered, but a marked reduction of the volume of residual extracellular matrix (all components other than collagen and elastic fibers) and interstitial cells was found. A correlation between the volumes of residual extracellular matrix and distal air spaces, as well as total surface area of alveolar epithelium, could be established. Cell-specific expression of HGF leads to a remodeling of the connective tissue within the septal walls in the healthy lung, which is associated with more pronounced stretching of distal air spaces at a given hydrostatic pressure during instillation fixation.

scholarly journals Identification of vitamin K-dependent carboxylase activity in lung type II cells but not in lung macrophages

1988 ◽  
Vol 250 (2) ◽  
pp. 557-563 ◽  
Author(s):  
R Wallin ◽  
S R Rannels
Keyword(s):  
Epithelial Cells ◽  
Vitamin K ◽  
Alveolar Epithelium ◽  
Type Ii Cells ◽  
Significant Activity ◽  
Type Ii ◽  
Clotting Factors ◽  
Carboxylase Activity ◽  
Type Ii Cell ◽  
Molecular Masses

Fluorography of 14C-labelled glutamic acid residues in vitamin K-dependent protein precursors in lung microsomes (microsomal fractions) shows that the lung has several substrates that are not found in the liver. These precursor proteins unique to the lung have apparent molecular masses of 65, 53, 50, 36, 31 and 13 kDa. Type II epithelial cells appear to synthesize most of the vitamin K-dependent proteins in the lung. The 36 and the 31 kDa precursors also found in Type-II-cell microsomes have a similar molecular mass to those of surfactant-associated proteins, and we have previously shown [Rannels, Gallaher, Wallin & Rannels (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 5952-5956] that the 36 kDa protein is one of the precursors for these proteins. Immunoblotting of membrane fragments of Type-II-cell microsomes with plasma prothrombin antibodies identified two prothrombin-like antigens of apparent molecular masses 68 and 65 kDa. This raises the question as to whether Type II cells are also a potential site for synthesis of prothrombin and possibly other vitamin K-dependent clotting factors. Pulmonary macrophages appear to be devoid of vitamin K-dependent carboxylase activity. However, Type II epithelial cells have significant activity, and this activity was unaltered when these cells were maintained in primary culture for 3 days, suggesting that carboxylase activity is expressed in lung alveolar epithelium independently of culture-induced changes in cellular differentiation. Carboxylase activity in Type II cells was enhanced 2-fold when cells were cultured for 24 h in the presence of 50 microM-warfarin. Type II cells, therefore, resemble hepatocytes with regard to their response to coumarin anticoagulant drugs.

Sign in / Sign up

Export Citation Format

Share Document