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.

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.

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.

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.

Assembly of exogenous fibronectin into type II cell extracellular matrix

1997 ◽  
Vol 272 (5) ◽  
pp. L908-L915 ◽  
Author(s):  
J. W. Swisher ◽  
D. E. Rannels
Keyword(s):  
Extracellular Matrix ◽  
Calf Serum ◽  
Cell Types ◽  
Biologically Active ◽  
Matrix Proteins ◽  
Type Ii ◽  
Attachment Protein ◽  
Pulmonary Epithelial Cells ◽  
Type Ii Cell ◽  
Exogenous Origin

Type II pulmonary epithelial cells (T2P) in primary culture assemble a biologically active extracellular matrix (ECM) from endogenously synthesized components, including fibronectin. Fibronectin is a well-recognized attachment protein that mediates cell adhesion, migration, and cytodifferentiation. In some cell types, exogenous fibronectin also is incorporated into ECM. The latter pathway of ECM assembly was thus investigated in T2P. Cells were cultured for 3-days in Dulbecco's modified Eagle's medium (DMEM) with or without 10% fetal calf serum (FCS), a source of exogenous fibronectin. Cell and matrix fractions were harvested on culture days 1, 2, and 3 to determine synthesis of cell and matrix proteins and matrix fibronectin content. During 3 days in DMEM containing 10% FCS, T2P flattened and spread to confluence more rapidly than cells in DMEM; they also produced ECM with higher fibronectin content than did cells in DMEM alone. On culture days 2 and 3, 10% FCS doubled (on average) synthesis of ECM fibronectin; in contrast, ECM fibronectin content increased nearly 10-fold. These observations suggest that cultured type II cells incorporate exogenous fibronectin into newly assembled ECM to a greater extent than the newly synthesized glycoprotein. Components of both endogenous and exogenous origin may therefore contribute to T2P assembly of a biologically active ECM.

Type II epithelial cells are critical target for hyperoxia-mediated impairment of postnatal lung development

2006 ◽  
Vol 291 (5) ◽  
pp. L1101-L1111 ◽  
Author(s):  
Min Yee ◽  
Peter F. Vitiello ◽  
Jason M. Roper ◽  
Rhonda J. Staversky ◽  
Terry W. Wright ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Epithelial Cells ◽  
Lung Development ◽  
Type I ◽  
Type Ii Cells ◽  
Type Ii ◽  
Newborn Mice ◽  
Type I Cells ◽  
Type Ii Cell ◽  
I Cells

Type II epithelial cells are essential for lung development and remodeling, as they are precursors for type I cells and can produce vascular mitogens. Although type II cell proliferation takes place after hyperoxia, it is unclear why alveolar remodeling occurs normally in adults whereas it is permanently disrupted in newborns. Using a line of transgenic mice whose type II cells could be identified by their expression of enhanced green fluorescent protein and endogenous expression of surfactant proteins, we investigated the age-dependent effects of hyperoxia on type II cell proliferation and alveolar repair. In adult mice, type II cell proliferation was low during room air and hyperoxia exposure but increased during recovery in room air and then declined to control levels by day 7. Eight weeks later, type II cell number and alveolar compliance were indistinguishable from those in room air controls. In newborn mice, type II cell proliferation markedly increased between birth and postnatal day 7 before declining by postnatal day 14. Exposure to hyperoxia between postnatal days 1 and 4 inhibited type II cell proliferation, which resumed during recovery and was aberrantly elevated on postnatal day 14. Eight weeks later, recovered mice had 70% fewer type II cells and 30% increased lung compliance compared with control animals. Recovered mice also had higher levels of T1α, a protein expressed by type I cells, with minimal changes detected in genes expressed by vascular cells. These data suggest that perinatal hyperoxia adversely affects alveolar development by disrupting the proper timing of type II cell proliferation and differentiation into type I cells.

scholarly journals Intermediate filaments in the developing type II cell of fetal monkey lung.

1985 ◽  
Vol 33 (11) ◽  
pp. 1161-1168 ◽  
Author(s):  
E Y Chi ◽  
A M Gown ◽  
A M Vogel ◽  
E C Teh
Keyword(s):  
Monoclonal Antibody ◽  
Epithelial Cells ◽  
Intermediate Filaments ◽  
Type I ◽  
Type Ii Cells ◽  
Type Ii ◽  
Lung Maturation ◽  
Filament Bundles ◽  
Type Ii Cell ◽  
Parallel Filament

Anticytokeratin monoclonal antibody was used to study epithelial cell development in fetal monkey lungs taken from animals of different ages. It is well established that the overall maturity of fetal lung depends greatly on the maturation of type II epithelial cells in the alveolus. In this study, we have correlated the cytokeratin phenotype of mammalian epithelial cells with pneumocyte maturation. We show that differentiation and maturation of the type II cell is related to intermediate filament expression. Twenty-four fetal monkeys (Macaca nemestrina) were delivered by cesarean section at a gestational age of 135-140 days (term = 168 days) and divided into two groups. One group of animals was sacrificed during the first 3 hr of life, and the other group was maintained in incubators for 92-120 hr. Anticytokeratin monoclonal antibody recognizes only alveolar type I and type II epithelial cells. In the first 3 hr of life, the cytokeratin was localized only at the alveolar surface and at the cytoplasmic periphery of the type II cells of these premature animals. However, at the age of 92-120 hr, the epithelia in the lungs reacted more intensely than they did during the first 3 hr. Electron microscopy revealed and confirmed that the type II cells were matured and abundant intermediate filaments appeared in the cytoplasm. The filaments appeared to form either aggregates or parallel filament bundles and few were closely associated with the lamellar bodies. In the immature type II cells at 0-3 hr of life, few intermediate filaments could be localized in the cytoplasm, and no parallel filament bundle was observed, though many appeared in the 92-120 hr lungs. This suggests that the intermediate filaments have a functional significance in the development and maturation of the type II cell. The location and stability of keratin filaments in type II cells may confer the structural strength necessary for cells covering a free surface in the alveoli during lung maturation.

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.

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.

P2u purinoceptor stimulation of surfactant secretion coupled to phosphatidylcholine hydrolysis in type II cells

1994 ◽  
Vol 267 (5) ◽  
pp. L625-L633 ◽  
Author(s):  
L. I. Gobran ◽  
Z. X. Xu ◽  
Z. Lu ◽  
S. A. Rooney
Keyword(s):  
Phospholipase D ◽  
Receptor Gene ◽  
Primary Cultures ◽  
Cell Receptor ◽  
Northern Analysis ◽  
Type Ii Cells ◽  
Type Ii ◽  
Type Ii Cell ◽  
Subsequent Activation ◽  
Phosphatidylcholine Secretion

ATP is known to stimulate surfactant phospholipid secretion in type II cells, and there is evidence that this effect is mediated by a P2 purinoceptor. At least five subtypes of the P2 receptor have been reported, but it is not clear which one exists on the type II cell. To determine whether it is the P2u subtype, at which UTP is equipotent with ATP, we have compared the effects of ATP and UTP on phosphatidylcholine secretion and second messenger formation in primary cultures of rat type II cells. ATP and UTP were equally potent in stimulating phosphatidylcholine secretion and phospholipase D activation. The potency order, UTP = ATP > ADP > 2-methylthio-ATP, was the same as that reported for the P2u receptor. UTP stimulated diacylglycerol and phosphatidic acid formation to the same extent as ATP. ATP also increased choline formation. Formation of diacylglycerol was biphasic, and the first peak in response to ATP was previously shown to be associated with inositol trisphosphate formation. Northern analysis showed that the P2u receptor gene was expressed to a greater extent in type II cells than in whole lung. These data suggest that ATP and UTP act via a P2u receptor that is coupled to phosphoinositide-specific phospholipase C with subsequent activation of phospholipase D acting on phosphatidylcholine. ATP has also been reported to act at an additional type II cell receptor coupled to adenylate cyclase. In contrast, UTP did not promote adenosine 3',5'-cyclic monophosphate formation and therefore does not act at that receptor.

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.

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