Prostaglandin E2 integrates
the effects of fluid distension and glucocorticoid on lung
maturation
J. S.
Torday,
H.
Sun, and
J.
Qin
Division of Neonatology, Department of Pediatrics, University of
Maryland Medical School, Baltimore, Maryland 21201
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ABSTRACT |
Both
glucocorticoids and alveolar fluid distension affect the rate of fetal
lung maturation, possibly representing a common cellular pathway. In an
explant culture, there is a spontaneous increase in triglyceride
incorporation into saturated phosphatidylcholine over time. This
mechanism is stimulated by prostaglandin (PG) E2, blocked by both bumetanide and
indomethacin, and overridden by exogenous
PGE2. Type II cells synthesized
and produced PGE2 between
days 16 and
21 postconception, increasing fourfold
between days 19 and
21. Fetal rat lung fibroblasts
released triglyceride in response to
PGE2, increasing 10- to 14-fold
between days 19 and
21 postconception; phloretin (1 × 10
5 M) completely
blocked this effect of PGE2 on
triglyceride release. Dexamethasone stimulated both type II cell
PGE2 synthesis (threefold) and
fibroblast triglyceride release in response to
PGE2 (60%) by
day 20 cells. Stretching type II cells
also increased PGE2 synthesis
(~100% at 1, 2, and 3 h vs. static cultures). Recombination of
[3H]triglyceride-labeled
fibroblasts with type II cells in an organotypic culture resulted in
progressive incorporation of label into saturated phosphatidylcholine
by type II cells. This process was also blocked by the addition of
indomethacin and overridden by exogenous
PGE2. These data suggest that the
combined effects of alveolar fluid dilatation and glucocorticoids may
coordinate the timely transfer of triglyceride from fibroblasts to type
II cells for augmented surfactant production through their effects on
PGE2 production and action as term
approaches.
stretch; surfactant; triglyceride
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INTRODUCTION |
HORMONAL EFFECTS on fetal lung development have
provided important tools for understanding the cellular and molecular
mechanisms that orchestrate and time this complex
epithelial-mesenchymal cascade (22). However, the paradoxical
observation that the lung can develop structurally and functionally in
explant culture in the absence of added hormones (6, 8, 16) has left
open the question as to what is the nature of the central regulatory mechanism(s) mediating alveolar differentiation. One potential clue is
the observation that fluid distension and the resultant stretching of
lung tissue are necessary for normal lung development (1, 5, 17),
leaving open the question as to whether the stretch-mediated and
hormone-regulated pathways share common regulatory elements. The
existence of a common mechanism was suggested by the study of Skinner
et al. (20), in which cortisol was shown to potentiate the effect of
prostaglandin (PG) E2, a
stretch-activated factor (25), on fetal rat lung fibroblast adenosine
3',5'-cyclic monophosphate production. In this context,
Hume et al. (9-11) and Ballard et al. (2) have identified
PGE2 as an endogenous mediator of
the automaturational mechanism of lung explant development. We now
provide experimental evidence that both dexamethasone (Dex) and stretch
stimulate PGE2 synthesis by type
II cells and surfactant substrate (triglyceride) release by
fibroblasts, integrating key epithelial-mesenchymal interactions
involved in surfactant phospholipid metabolism in a timely manner.
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METHODS AND MATERIALS |
Reagents.
PGE2, Dex, bumetanide,
indomethacin, phloretin, triolein, and phospholipid standards were all
purchased from Sigma Chemical (St. Louis, MO).
[9,10-3H(N)]triolein
(30 Ci/mmol) and
[5,6,8,9,10,11,12,14,15-3H(N)]arachidonic
acid (100 Ci/mmol) were purchased from New England Nuclear (Boston,
MA). Trypsin (1:250), Dulbecco's modified Eagle's medium (DMEM), and
antibiotics for tissue culture were obtained from GIBCO (Grand Island,
NY). Collagen type I was purchased from Collaborative Biomedical
Products (Bedford, MA). A PGE2
enzyme-linked immunosorbent assay was bought from Cayman Chemical (Ann
Arbor, MI).
Explant culture of fetal rat lung.
Time-mated (vaginal smear positive = day
0) Sprague-Dawley rats were obtained from Charles River Breeders (Kingston, NY). The pups were weighed to confirm that
they were within the appropriate weight-for-age limits established in
this laboratory. The dams were killed by decapitation, and the pups
were removed from the uterus by laparotomy and kept on ice. The lungs
were removed en bloc in a laminar flow hood with sterile technique,
dissected free of adjoining structures, and put into ice-cold sterile
Hanks' balanced salt solution without calcium or magnesium (HBSS).
Lungs (2/well) were planted in 24-well plates (1-cm diameter) in a
minimal volume of culture medium for 60 min at 37°C in 5%
CO2-95% air to promote adherence
to the plastic surface. At the end of the incubation period, the medium
used to transfer the explants to the wells was aspirated, and culture medium was added to cover the explants (1 ml). The explants were cultured in Waymouth MB-752/1 medium containing penicillin (100 µg/ml), streptomycin (100 µg/ml), and amphotericin B (5 µg/ml) (7).
Technique for harvesting lung fibroblasts by
differential adherence. Five to ten time-mated dams
were used per preparation depending on the number of experimental
variables to be tested. The fetal lungs were removed into HBSS as
described in Explant culture of fetal rat
lung. The HBSS was decanted, and five
volumes of 0.05% trypsin were added to the lung preparation. The lungs were dissociated in a 37°C water bath with a Teflon stirring bar to
disrupt the tissue mechanically. Once the tissue was dispersed into a
unicellular suspension (~20 min), the cells were pelleted at 500 g for 10 min at room temperature in a
50-ml polystyrene centrifuge tube. The supernatant was decanted, and
the pellet was resuspended in DMEM containing 20% fetal bovine serum
(FBS) to yield a mixed cell suspension of ~3 × 108 cells (determined by a Coulter
particle counter, Hialeah, FL). The cell suspension was then added to
culture flasks (25 or 80 cm2)
for 30-60 min to allow for differential adherence of lung
fibroblasts (21). These cells are >95% pure fibroblasts on the basis
of vimentin staining.
Isolation of fetal type II cells by Nycodenz
gradient. The minced lungs from four to six litters
were pooled, washed with 50 ml of a mixture of 140 mM NaCl, 5 mM KCl,
2.5 mM
Na2HPO4,
10 mM
N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic
acid (HEPES), 6 mM glucose, and 0.2 mM ethylene
glycol-bis(
-aminoethyl
ether)-N,N,N',N'-tetraacetic acid, pH 7.4 (solution I), and
filtered through 100-µm nylon mesh. The minced lungs were transferred
to a trypsinizing flask containing 40 ml of a mixture of 140 mM NaCl, 5 mM KCl, 2.5 mM
Na2HPO2,
10 mM HEPES, 2.0 mM CaCl2, and 1.3 mM MgSO4, pH 7.4 (solution II) with elastase (30 orcein-elastase units/ml) and deoxyribonuclease (250 µg/ml) and
stirred for 20 min at 37°C. The digestion was terminated by adding
5 ml of carbon-stripped (hormone-depleted) FBS. The cell suspension was
filtered sequentially through 2- and 4-ply gauze and then 37- and
15-µm nylon mesh and was washed with additional
solution II to a final volume of 40 ml. The cells were centrifuged at 130 g for 10 min at 20°C and
resuspended in DMEM with 2% FBS for the addition to Nycodenz gradients
(26). Identity of fetal type II cells was documented by staining for the presence of glycogen and by the reaction with specific type II cell
markers, Maclura pomifera lectin, and antibody to
cytokeratins 8 and 18 and was found to be ~90% pure by these
criteria.
Organotypic culture. Organotypic
cultures were prepared as previously described (23).
Cell stretch apparatus. This technique
is an adaptation of the method of Wirtz and Dobbs (27). Freshly
isolated rat fetal lung type II cells (day
20 postconception) were seeded onto Silastic membranes
(Dow Corning, Midland, MI) coated with type I collagen. The Silastic
membranes were mounted in a 25-mm filter holder (Gelman Sciences, Ann
Arbor, MI), and DMEM (0.7 ml) was added to the upper chamber to
maintain the cell layer. To stretch the cell monolayer, a 0.5-ml bolus
of air was introduced into the lower chamber through a plastic tube
that was then clamped to maintain positive pressure. The preparations
were then placed in a CO2
incubator at 37°C in an atmosphere of 5%
CO2-95% air for 3 h. At the end
of the incubation, the conditioned media were collected and frozen at
70°C until analyzed for
PGE2. The cell layers were
harvested from the Silastic membranes with 0.1% trypsin; an aliquot of
the cell suspension was taken to determine cell number.
Uptake and release of
[3H]triolein by fibroblasts.
Culture medium was aspirated from confluent fibroblast cultures and
replaced with DMEM containing 20% triglyceride (vol/vol) mixed with
[3H]triolein (5 µCi/ml) that was prepared by first drying the
[3H]triolein under a
stream of nitrogen, resuspending it in 50 µl of ethanol, and then
adding the DMEM plus serum and vortexing the mixture thoroughly. The
cells were incubated at 37°C in 5% CO2-95% air for 4-12 h and
subsequently analyzed for their triglyceride content. For the
triglyceride release assay, the fibroblasts were incubated with the
[3H]triolein mixture
for 24 h, washed three times with 3 ml of DMEM to remove excess
radiolabeled solution, and then incubated with PGE2 (200 ng/ml) in DMEM at
37°C in 5% CO2-95% air for 2 h. At the end of the incubation, the media were aspirated and analyzed for triglyceride content (18).
Fibroblast triglyceride incorporation into type II
cell phospholipid. The method for determining the rate
of fibroblast
[3H]triglyceride
incorporation into type II cell phospholipids has been previously
described (23). Briefly, day 20 fetal
rat lung fibroblasts were incubated with
[3H]triolein (5 µCi/ml) and 200 µg/ml of serum triglyceride for 24 h and were then
recombined with day 20 fetal rat type
II cells in organotypic culture for 24 h. At the end of the incubation, type II cells were reisolated and analyzed for their phospholipid content.
Rate of [3H]arachidonic acid
incorporation into
[3H]PGE2.
This assay was based on the method of Polgar and Taylor
(19). Briefly, type II cells in monolayer culture were
incubated with
[3H]arachidonic acid
(5 µCi/ml) in DMEM in a CO2
incubator for 24 h. At the end of the incubation, the culture medium
was extracted with ethyl acetate, the extract was evaporated under a
stream of nitrogen at 55°C, and the extract was chromatographically
separated by thin-layer chromatography in ethyl
acetate-isooctane-acetic acid-water (11:5:2:10). The area on the
chromatogram corresponding to authentic
PGE2 was scraped from the plate,
and its radioactive content was determined by liquid scintillation
spectrometry. PGE2 represented at
least 80% of the radioactivity on the chromatogram.
PGE2 assay.
PGE2 in culture medium was assayed
with a commercial enzyme-linked immunosorbent assay.
DNA assay. DNA was assayed by the
method of Burton (4).
Statistical analysis. Data were
analyzed either by Student's t-test
or one- or two-way analysis of variance; differences between groups
were determined by the method of least significant difference, as
indicated.
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RESULTS |
Day 20 postconception fetal rat lung
explants were maintained in culture for up to 3 days in the presence of
either bumetanide (1 × 10
5 M), an inhibitor of
chloride ion secretion, indomethacin (5 × 10
8 M), and/or
PGE2 (200 ng/ml) as indicated
(Fig. 1). At the time of assay, the
cultures were incubated with
[3H]triolein to
monitor for saturated phosphatidylcholine (SPC). As can be seen in Fig.
1, during the course of the 3-day culture period, there was a threefold
increase in triolein incorporation into SPC by the explants. Treatment
of the explants with PGE2 further
enhanced the rate of
[3H]triolein
incorporation into SPC (30%). In contrast to the effects of time in
culture and PGE2 treatment, both
bumetanide and indomethacin blocked the spontaneous increase in
triglyceride incorporation into SPC; the inhibitory effects of both
bumetanide and indomethacin were completely reversed by exogenous
PGE2.

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Fig. 1.
[3H]triolein
incorporation into saturated phosphatidylcholine (SPC) by fetal rat
lung explants. Day 17 postconception
fetal rat lung explants were cultured for 3 days (d3) in serum-free
medium plus bumetanide (Bu; 5 × 10 5 M), indomethacin (IND;
5 × 10 8 M),
prostaglandin E2
(PGE2; 200 ng/ml),
Bu/PGE2, or Bu/IND and
subsequently incubated with
[3H]triolein for 15 h.
dpm, Disintegrations/min. Values are means ± SD of 5 experiments;
n = 20 lungs. Significantly different
from control culture (day 0):
* P < 0.05;
** P < 0.01 (by 2-way analysis
of variance).
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PGE2 synthesis by type II cells.
When primary cultures of fetal rat type II cells were incubated with
[3H]arachidonic acid,
the predominant metabolite (>80%) was
PGE2, which was largely found in
the extracellular medium (>95%). As can be seen in Fig.
2A,
PGE2 synthesis was observed at all
gestational ages (16-21 days postconception), although there were
obvious developmental changes: there was basal synthesis of
[3H]PGE2
between days 16 and
18 postconception; beginning on
day 20 postconception, there was a
2.5-fold increase in de novo PGE2 synthesis (P < 0.05) that increased
to 4-fold by day 21 postconception (P < 0.01). A similar developmental
pattern was observed when conditioned medium from confluent type II
cell monolayers was assayed for
PGE2 content (Fig.
2B): basal production of
PGE2 (100-140 ng · 106
cells
1 · 24 h
1) was observed between
days 16 and
19 postconception, followed by a
saltatory increase on days 20 and
21 postconception (324 ± 102 and
609 ± 170 ng · 106
cells
1 · 24 h
1,
respectively).

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Fig. 2.
Ontogeny of PGE2 synthesis by
fetal rat lung type II cells. A: type
II cells were incubated with
[3H]arachidonic acid
(5 µCi/ml for 2 h), and media were analyzed for
[3H]PGE2.
Values are means ± SD of 5 experiments;
n = 20 lungs. Significantly different
from previous day: * P < 0.05;
** P < 0.01 (by 1-way analysis
of variance). B: type II cells were
incubated in serum-free medium for 24 h, and conditioned medium was
harvested and assayed for PGE2
content by enzyme-linked immunosorbent assay. Values are means ± SD
of 3-5 experiments; n = 15-20 lungs. Significantly different from previous day:
* P < 0.05;
** P < 0.01 (by 1-way analysis
of variance).
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Effect of PGE2 on triglyceride release by
fetal rat lung fibroblasts.
Using a concentration of PGE2
comparable to the amount of PGE2
produced by day 20 postconception type
II cells/24 h (i.e., 200 ng/ml), we observed a saltatory increase in
triglyceride release by fetal rat lung fibroblasts (Fig.
3) beginning on day
19 postconception, which then increased progressively
on days 20 (6-fold) and
21 postconception (10- to 14-fold).
Basal triglyceride release remained unchanged over this entire period
in the absence of added PGE2 (<5% of intracellular triglyceride). Release of triglyceride by day 21 postconception fibroblasts in
response to PGE2 increased linearly over a 15-h time course (Fig. 4);
coincubation with phloretin (1 × 10
5 M), a
PGE2-receptor antagonist,
completely blocked the effect of
PGE2 on triglyceride release by
these cells.

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Fig. 3.
Ontogeny of PGE2-stimulated
triglyceride release by fetal rat lung fibroblasts. Fibroblasts were
incubated with
[3H]triolein (5 µCi/ml + 200 µg of nonradiolabeled triglyceride) for 24 h and
subsequently challenged with (+) or without ( )
PGE2 (200 ng/ml) for 15 h.
[3H]triglyceride
released into medium was quantitated. Each data point represents mean ± SD of 3-5 experiments; n = 15-20 lungs. Significantly different from previous day:
* P < 0.05;
** P < 0.01 (by 1-way analysis
of variance).
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Fig. 4.
Phloretin inhibition of
PGE2-stimulated triglyceride
release by fibroblasts. Day 21 fibroblasts were preincubated with
[3H]triolein (5 µCi/ml + 200 µg of nonradiolabeled triglyceride) for 24 h. Cells
were subsequently challenged with
PGE2 (200 ng/ml) with and without
phloretin (1 × 10 5 M)
for up to 15 h. Culture medium was analyzed for
[3H]triglyceride. Each
data point represents mean ± SD of 2 experiments;
n = 10 lungs. Significantly different
from previous data point: * P < 0.05; ** P < 0.01;
*** P < 0.001 (by 1-way
analysis of variance).
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Effects of Dex on PGE2 synthesis and
action.
Day 20 postconception fetal rat type
II cells in a monolayer culture were exposed to Dex (1 × 10
7 M/15 h) in serum-free
medium for 15 h and subsequently incubated with
[3H]arachidonic acid
for 2 h (Fig. 5). Analysis of the culture
medium revealed a threefold increase in
[3H]PGE2
synthesized as a result of Dex treatment. Assay of the same media for
PGE2 content revealed a 70%
increase in production in response to Dex (290 ± 80 vs. 415 ± 38 ng · 106
cells
1 · 24 h
1;
P < 0.02 for control vs. Dex
cultures by Student's t-test). Treatment of day 20 postconception
fetal rat lung fibroblasts with Dex (1 × 10
7 M/15 h) resulted in a
60% increase in triglyceride release in response to
PGE2 (Fig.
6). Note that there was only a 5-10%
triglyceride release in the control and Dex-treated cultures.

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Fig. 5.
Effect of dexamethasone (Dex) on
PGE2 synthesis by fetal rat lung
type II cells. Type II cells were incubated with Dex (1 × 10 7 M) in serum-free MEM
for 15 h and subsequently incubated with
[3H]arachidonic acid
(5 µCi/ml for 2 h). Cells and medium were analyzed for
[3H]PGE2.
Values are means ± SD of 3 experiments;
n = 15 lungs. * Significantly
different from control cells, P < 0.01 (by Student's t-test).
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Fig. 6.
Effect of Dex on PGE2-mediated
triglyceride release by fetal rat lung fibroblasts. Fibroblasts were
initially incubated with
[3H]triolein (5 µCi/ml + 200 µg of nonradiolabeled triolein) for 24 h, followed by
a 15-h exposure to Dex (1 × 10 7 M). Cells were
subsequently challenged with PGE2
(200 ng/ml) for 15 h, and culture medium was analyzed for
[3H]triglyceride
released. Values are means ± SD of 3 experiments;
n = 15 lungs. * Significantly
different from control cells, P < 0.01. ** Significantly different from Dex-treated cells,
P < 0.05 (both by Student's
t-test).
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Effect of stretch on type II cell PGE2
synthesis.
Both static and stretched day 20 type
II cells exhibited progressive increases in
[3H]arachidonic acid
incorporation into
[3H]PGE2
over a 3-h period (Fig. 7), although the
stretched cells synthesized approximately twice as much
PGE2 as nonstretched cells per
unit time. Because a similar increase has been observed when type II
cells are cultured on plastic (data not shown), we assume this is a
reflection of baseline PG synthesis unrelated to culture on Silastic.
The stretched cells were evaluated for damage and found to be >98%
viable by trypan blue dye exclusion; assay for lactic dehydrogenase
content of conditioned medium revealed no differences between static
and stretched cultures.

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Fig. 7.
Effect of stretch on type II cell
PGE2 synthesis. Type II cells were
cultured on collagen-coated Silastic membranes and stretched. Cells
were incubated with
[3H]arachidonic acid
(5 µCi/ml) for the times indicated, and cells and media were analyzed
for
[3H]PGE2
synthesized. Values are means ± SD of 5 experiments;
n = 15 lungs. Significantly different
from control cells: * P < 0.05; ** P < 0.01 (by
Student's t-test).
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Effect of indomethacin on fibroblast triglyceride
incorporation into type II cell SPC. To directly
evaluate the effect of endogenous type II cell
PGE2 production on fibroblast
triglyceride incorporation into type II cell SPC, day
20 fetal rat lung fibroblasts were prelabeled with
[3H]triglyceride for
24 h and recombined with day 20 fetal
rat type II cells in an organotypic culture. As can be seen in Fig.
8, there was a linear increase in the
amount of
[3H]triglyceride
incorporated into
[3H]SPC by type II
cells over the 8-h time course. In contrast to these observations, the
addition of indomethacin (5 × 10
8 M) to the incubation
medium inhibited the incorporation of fibroblast triglyceride into type
II cell SPC to a rate comparable to that of
[3H]triglyceride
incorporation by type II cells alone; simultaneous addition of
PGE2 (200 ng/ml) neutralized the
indomethacin effect. In separate studies, we have established that
indomethacin has no effect on
[3H]triglyceride
incorporation into SPC by type II cells.

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Fig. 8.
Effect of IND on fibroblast triglyceride incorporation into type II
cell SPC. Day 20 postconception fetal
rat lung fibroblasts (fib) were prelabeled with
[3H]triglyceride (5 µCi/ml + 200 µg of serum triglyceride/24 h) and recombined with
day 20 postconception fetal rat type
II cells (TII) in organotypic culture. Cultures were treated with IND
(5 × 10 8 M) and
PGE2 (200 ng/ml) as indicated.
Type II cells were harvested at the times indicated and analyzed for
[3H]SPC. Each data
point represents mean ± SD of 3 experiments;
n = 15 lungs. * Significantly
different from TII cells, P < 0.0001 (by Student's t-test).
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 |
DISCUSSION |
In the present series of experiments, we have observed that fluid
distension of the developing lung in explant culture (3, 14, 15) is
associated with increased triglyceride incorporation into surfactant
phospholipids, a process that was blocked by the addition of either
bumetanide or indomethacin, implicating fluid distension and PG
synthesis in this complex cellular mechanism. The addition of exogenous
PGE2 to these cultures in the
presence or absence of bumetanide stimulated triglyceride incorporation into SPC, indicating that PGE2
stimulates this mechanism through a stretch-related process. We
subsequently found that type II cells synthesize and produce
PGE2 beginning on
days 19-20 postconception and
that PGE2 [at a
concentration similar to that produced by static day
20 postconception type II cells (200 ng/ml)]
stimulates triglyceride release from cultured fetal rat lung
fibroblasts. This sequence provides a cellular mechanism for type II
cell PGE2-mediated mobilization of
triglycerides from lung fibroblasts. Phloretin inhibition of
PGE2-stimulated triglyceride
release by fibroblasts indicates that the PG effect is receptor
mediated (4a). Furthermore, both the synthesis of
PGE2 by type II cells and its
stimulation of triglyceride release by
PGE2 are enhanced by exposure to
Dex. Thus the timing of PGE2
elaboration and augmentation by steroids and stretch is coincident with
the spontaneous surge of surfactant production in utero (7, 23).
Additionally, when type II cells are stretched in culture, the de novo
synthesis of PGE2 is stimulated. These data suggest that fluid distension stimulates
PGE2 elaboration by type II cells,
triggering the incorporation of triglyceride into surfactant
phospholipids.
Lung tissue has the capacity to differentiate spontaneously in vitro,
forming all of the cellular elements of the mature lung (2-5,
9-11), particularly the ontogeny of the alveolar type II cells (2,
4) that flatten and develop surfactant synthetic capacity. This process
of cellular development occurs in conjunction with fluid dilatation of
the terminal air sacs that has been found to play a central role in
cellular growth and differentiation. Fluid distension of developing
alveoli has been shown to be necessary for normal lung growth and
differentiation both in vivo (6-8) and in vitro (9, 10, 12). The
distension of the alveoli and the attendant stretching of the cells
lining them has been implicated in cell growth (12, 13) and
differentiation (24), particularly as they relate to PG synthesis (2,
8-11).
In the present series of studies, the explant experiments indicate that
automaturation, as reflected by the spontaneous increase in
incorporation of triglyceride into surfactant phospholipid with time in
culture, can be inhibited by either bumetanide, which blocks chloride
ion secretion and alveolar fluid formation (14, 15), or indomethacin,
an inhibitor of PG synthesis. Furthermore, the inhibitory effects of
both bumetanide and indomethacin can be overcome by the addition of
exogenous PGE2 to the explant
cultures, suggesting that the effect of alveolar dilatation is
mediated, at least in part, by PGs. Similar effects of
PGE2 on the structural maturation
of human fetal lung were shown by both Ballard et al. (2) and Hume et
al. (9-11) with regard to formation of alveolar lumina and
cellular maturation. We have now addressed one of the underlying
mechanisms that mediate the stretch-dependent cell-cell interactions
leading to augmented surfactant synthesis.
In previous studies, Nunez and Torday (18) found that the
accumulation of triglycerides by fetal lung fibroblasts is a
glucocorticoid-regulated mechanism that may be important for surfactant
phospholipid synthesis because these triglycerides can act as a
substrate for type II cell surfactant phospholipid synthesis in cell
culture as found by Torday et al. (23). The transit of triglyceride
from fibroblasts to type II cells in an organotypic culture is
glucocorticoid stimulated (18), inferring that either the synthesis of
PGE2, its action on fibroblast
triglyceride release, or both are stimulated by glucocorticoids. In the
present series of experiments, we have shown that Dex stimulates both
PGE2 synthesis by type II cells and fibroblast triglyceride release in response to
PGE2, which would explain the
observed effect of Dex on triglyceride mobilization in the organotypic
cultures. These effects of Dex on
PGE2 synthesis and action in the
fetal lung are somewhat surprising at first because steroids are
generically anti-inflammatory (and prostanoids are proinflammatory).
However, this is true in mature, homeostatically regulated systems,
whereas the present studies are of a developmental mechanism. In
support of this seemingly paradoxical effect of steroid on the
developing rat lung, Tsai et al. (24) have observed Dex stimulation of
PG synthesis by fetal rat lung in vivo, and Skinner et al. (20) have
observed that cortisol enhances the fetal rat lung fibroblast adenosine
3',5'-cyclic monophosphate response to
PGE2.
In the present series of experiments, we have demonstrated that
1) triglyceride metabolism by fetal
lung explants depends on alveolar fluid distension and PG synthesis,
2) type II cells develop the ability
to synthesize PGE2,
3) fibroblasts develop the capacity
to respond to PGE2,
4) fibroblasts develop the capacity to respond to PGE2 with regard to
triglyceride secretion, 5) Dex stimulates both 2 and
3, and
6) stretch stimulates de novo
PGE2 synthesis by type II cells.
Therefore, we conclude that fluid distension plays a central role in
the organized development of the primordial lung, a process that can be
modulated by steroid exposure.
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ACKNOWLEDGEMENTS |
This study was presented at the meeting of the Society for
Pediatric Research, Washington, DC, May 1996.
 |
FOOTNOTES |
Address reprint requests to J. S. Torday.
Received 20 August 1996; accepted in final form 2 October 1997.
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