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.

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.

Adrenal hormone regulation of extracellular matrix synthesis by type II cells

1995 ◽  
Vol 268 (6) ◽  
pp. L885-L893 ◽  
Author(s):  
S. E. Dunsmore ◽  
S. R. Rannels ◽  
R. N. Grove ◽  
D. E. Rannels
Keyword(s):  
Extracellular Matrix ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Hormone Regulation ◽  
Type Ii ◽  
Adrenal Steroid ◽  
Adrenal Hormone ◽  
Ecm Proteins ◽  
Cell Growth And Differentiation

Previous studies suggested an association between changes in extracellular matrix (ECM) and pulmonary epithelial cell growth and differentiation in lungs of adrenalectomized rats. The role of adrenal hormones to modulate synthesis of cell and ECM proteins by type II epithelial cells was thus investigated. In cells isolated from lungs of adrenalectomized animals, synthesis of ECM proteins was stimulated to a greater extent than was synthesis of cell proteins. This resulted in increased relative synthesis of matrix components. Conversely, dexamethasone (200 nM) treatment in vitro reduced the effects of adrenalectomy and, in control cells, decreased relative ECM synthesis. The magnitude of these effects depended on time in culture and serum conditions. For example, culture of normal cells in serum-free medium mimicked the effects of adrenalectomy, whereas there were no effects of dexamethasone in the presence of serum. These results indicate a role for adrenal steroid hormones to modulate ECM metabolism by type II pulmonary epithelial cells and suggest that consequent effects on ECM biological activity may modify epithelial cell differentiation or growth.

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.

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.

Connexin expression by alveolar epithelial cells is regulated by extracellular matrix

2001 ◽  
Vol 280 (2) ◽  
pp. L191-L202 ◽  
Author(s):  
Yihe Guo ◽  
Cara Martinez-Williams ◽  
Clare E. Yellowley ◽  
Henry J. Donahue ◽  
D. Eugene Rannels
Keyword(s):  
Extracellular Matrix ◽  
Epithelial Cells ◽  
Gap Junction ◽  
Intercellular Communication ◽  
Translational Regulation ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type Ii ◽  
Gap Junction Intercellular Communication ◽  
Type Ii Cell

Extracellular matrix (ECM) proteins promote attachment, spreading, and differentiation of cultured alveolar type II epithelial cells. The present studies address the hypothesis that the ECM also regulates expression and function of gap junction proteins, connexins, in this cell population. Expression of cellular fibronectin and connexin (Cx) 43 increase in parallel during early type II cell culture as Cx26 expression declines. Gap junction intercellular communication is established over the same interval. Cells plated on a preformed, type II cell-derived, fibronectin-rich ECM demonstrate accelerated formation of gap junction plaques and elevated gap junction intercellular communication. These effects are blocked by antibodies against fibronectin, which cause redistribution of Cx43 protein from the plasma membrane to the cytoplasm. Conversely, cells cultured on a laminin-rich ECM, Matrigel, express low levels of Cx43 but high levels of Cx26, reflecting both transcriptional and translational regulation. Cx26 and Cx43 thus demonstrate reciprocal regulation by ECM constituents.

Enhanced alveolar type II cell growth on a pulmonary extracellular matrix over fibroblasts

1997 ◽  
Vol 272 (3) ◽  
pp. L413-L417 ◽  
Author(s):  
I. Y. Adamson ◽  
L. Young ◽  
J. Bakowska
Keyword(s):  
Extracellular Matrix ◽  
Cell Growth ◽  
Growth Effect ◽  
Lung Fibroblasts ◽  
Cell Contact ◽  
Alveolar Type ◽  
Thymidine Uptake ◽  
Type Ii Cells ◽  
Type Ii ◽  
Type Ii Cell

The growth of alveolar type II cells was studied when these cells were maintained for 2 days on a pulmonary endothelium-derived extracellular matrix (ECM) on a filter with or without lung fibroblasts in the lower chambers of culture wells. Type II cell proliferation was enhanced by the ECM compared with other substrates but was significantly higher with fibroblasts beneath. This was determined by thymidine uptake and cell numbers. The diffusing factor from fibroblasts appeared to be keratinocyte growth factor (KGF), because this cytokine increased type II cell growth in culture and the neutralizing antibody to KGF blocked the observed fibroblast-induced growth increase. None of the antibodies to various cytokines had any effect on the ECM-induced proliferation. Although the type II cells were shown to produce degradative activity for the ECM, there was little secreted enzyme activity in supernatants and there was no demonstrated autocrine-regulated growth effect. The results suggest that type II cell growth may be stimulated by both 1) a matrix-bound factor that acts through a cell contact-mediated process, and 2) a fibroblast-secreted factor that appears to be KGF.

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.

Alveolar type II cell growth on a pulmonary endothelial extracellular matrix

1996 ◽  
Vol 270 (6) ◽  
pp. L1017-L1022 ◽  
Author(s):  
I. Y. Adamson ◽  
L. Young
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Cell Growth ◽  
Alveolar Type ◽  
Thymidine Uptake ◽  
Type I ◽  
Type Ii Cells ◽  
Type Ii ◽  
Lamellar Bodies ◽  
Type Ii Cell

Most of the alveolar epithelium overlies a fused basement membrane produced by epithelial and endothelial cells. To determine how this type of matrix influences type II cell growth and function, we studied the effects of culturing isolated rat alveolar type II cells on an extracellular matrix (ECM) freshly produced by pulmonary vascular endothelial cells grown 5 days in culture. Type II cells from the same rats were cultured on plastic or Matrigel for comparison. A large increase in mitotic activity was seen in type II cells grown on the endothelial ECM at 2 days only; thereafter cells spread rapidly to confluence and lost their lamellar bodies. Cells grown on Matrigel remained cuboidal with lamellar bodies but grew more slowly, as judged by [3H]thymidine uptake and cell numbers. Incorporation of labeled choline into disaturated phosphatidylcholine (DSPC) was used as a marker of surfactant synthesis. After the rapid, brief burst of proliferation, type II cells on endothelial ECM showed a sudden decline in DSPC-DNA by day 4 compared with cells grown on matrigel. Binding of the lectin Bauhinia purpurea (BPA) indicated that after a phase of division, cells on endothelial ECM developed as type I epithelium by 4 days of culture, when > 70% of cells stained positively for BPA binding, whereas few cuboidal cells on Matrigel were stained. The results indicate that type II cells respond briefly to growth factors in pulmonary endothelial ECM; then this type of matrix promotes cell spreading with loss of type II function as cells subsequently resemble type I epithelium.

Effects of RSV infection on pulmonary surfactant protein SP-A in cultured human type II cells: contrasting consequences on SP-A mRNA and protein

2005 ◽  
Vol 289 (6) ◽  
pp. L1113-L1122 ◽  
Author(s):  
Joseph L. Alcorn ◽  
James M. Stark ◽  
Constance L. Chiappetta ◽  
Gaye Jenkins ◽  
Giuseppe N. Colasurdo
Keyword(s):  
Gene Expression ◽  
Protein Gene ◽  
Surfactant Protein ◽  
Type Ii Cells ◽  
Protein Homeostasis ◽  
Mrna Levels ◽  
Type Ii ◽  
Human Type ◽  
Rsv Infection ◽  
Type Ii Cell

Respiratory syncytial virus (RSV) is the most important cause of serious lower respiratory illness in infants and children. Surfactant proteins A (SP-A) and D (SP-D) play critical roles in lung defense against RSV infections. Alterations in surfactant protein homeostasis in the lung may result from changes in production, metabolism, or uptake of the protein within the lung. We hypothesized that RSV infection of the type II cell, the primary source of surfactant protein, may alter surfactant protein gene expression. Human type II cells grown in primary culture possess lamellar bodies (a type II cell-specific organelle) and the ability to express surfactant protein mRNA. These cells were infected with RSV (by morphology and antibody binding). Surfactant protein mRNA levels determined by quantitative RT-PCR indicated a marked increase in SP-A mRNA levels (3-fold) 24 h after RSV exposure, whereas SP-D mRNA levels were unaffected. In contrast to mRNA levels, total SP-A protein levels (determined by Western blot analysis) were decreased 40% after RSV infection. The percentage of secreted SP-A was 43% of the total SP-A in the RSV-infected cells, whereas the percentage of secreted SP-A was 61% of the total SP-A in the uninfected cells. These changes in SP-A transcript levels and protein secretion in cultured human cells were recapitulated in RSV-infected mouse lung. Our findings suggest that type II cells are potentially important targets of RSV lower respiratory infection and that alterations in surfactant protein gene expression and SP-A protein homeostasis in the lung may arise via direct effects of RSV.

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