Abnormal lung growth and the development of pulmonary
hypertension in the Fawn-Hooded rat
Timothy D.
le Cras1,
Dug-Ha
Kim2,
Sarah
Gebb3,
Neil E.
Markham1,
John M.
Shannon3,
Rubin M.
Tuder4, and
Steven H.
Abman1
1 Pediatric Heart Lung Center,
Department of Pediatrics, and
4 Department of Pathology,
University of Colorado School of Medicine and The Children's Hospital,
Denver 80262; 3 National
Jewish Hospital, Denver, Colorado 80206; and
2 Department of Pediatrics, Hallym University,
150-030 Seoul, South Korea
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ABSTRACT |
The Fawn-Hooded
rat (FHR) strain develops accelerated and severe pulmonary hypertension
when exposed to slight decreases in alveolar
PO2. We recently observed that adult
FHR lungs showed a striking pattern of disrupted alveolarization and
hypothesized that abnormalities in lung growth in the perinatal period
predisposes the FHR to the subsequent development of pulmonary
hypertension. We found a reduction in lung weight in the fetus and
1-day- and 1-wk-old FHR compared with a normal rat strain
(Sprague-Dawley). Alveolarization was reduced in infant and adult FHR
lungs. In situ hybridization showed similar patterns of expression of
two epithelial markers, surfactant protein C and 10-kDa Clara cell secretory protein, suggesting that the FHR lung is not characterized by
global delays in epithelial maturation. Barium-gelatin angiograms demonstrated reduced background arterial filling and density in adult
FHR lungs. Perinatal treatment of FHR with supplemental oxygen
increased alveolarization and reduced the subsequent development of
right ventricular hypertrophy in adult FHR. We conclude that the FHR
strain is characterized by lung hypoplasia with reduced alveolarization
and increased risk for developing pulmonary hypertension. We speculate
that altered oxygen sensing may cause impaired lung alveolar and
vascular growth in the FHR.
alveolarization; lung development; lung hypoplasia; bronchopulmonary dysplasia
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INTRODUCTION |
NORMAL LUNG DEVELOPMENT is a highly regulated and
coordinated process of cell growth and differentiation that consists of five morphologically distinct stages. In humans, these stages of lung
development include the embryonic (up to 6 wk of gestation), pseudoglandular (7-16 wk), canalicular (17-28 wk), saccular
(29-35 wk), and alveolar periods (beginning at 36 wk and
continuing during postnatal life; see Ref. 10). During the canalicular
period, the lung undergoes dramatic remodeling, including marked
thinning of the interstitium and increased secondary crest formation,
that signals the onset of alveolarization (10). Although similar stages
of lung development have been identified in other species, the exact
timing of these events varies widely (22). As in the human lung,
alveolarization in the rat lung begins in utero, and alveolar number
increases markedly after birth (22). The first 3 wk of postnatal life
are the critical period of alveolar growth in the rat (7, 26). The
exact mechanisms contributing to alveolar development are incompletely
understood but are influenced by genetic, mechanical, hormonal,
autocrine, and paracrine factors (7-9). Adverse stimuli during
this period, such as hypoxia, hyperoxia, or glucocorticoid treatment,
can disrupt alveolarization and markedly reduce alveolar number (7-9,
25, 27, 29).
The pulmonary circulation develops concomitantly with distal lung air
space growth during late gestation and early postnatal life (15). Early
development of the pulmonary vascular bed involves vasculogenesis and
angiogenesis during the embryonic period, followed primarily by
angiogenesis during lung maturation (16). The precise relationship
between alveolar and vascular development during fetal and early
postnatal life and the mechanisms that coordinate lung vascular growth
with alveolarization are uncertain. However, signals from airway
epithelial cells, such as vascular endothelial cell growth factor
(VEGF), basic fibroblast growth factor, and other mediators, play a
major role in vascular growth and development during fetal life (6, 30,
40). These findings suggest that mechanisms exist that link lung
alveolar growth with vascular development and suggest that disruption
of normal alveolarization may also contribute to altered pulmonary
vascular growth.
The clinical importance of the relationship between alveolarization and
pulmonary vascular development is reflected by the association between
neonatal lung hypoplasia and pulmonary hypertension. Lung hypoplasia
and pulmonary hypertension contribute to high morbidity and mortality
in several neonatal lung diseases including bronchopulmonary dysplasia
(BPD), congenital diaphragmatic hernia (CDH), primary lung hypoplasia,
and Down's syndrome (1, 3, 11, 12, 14, 19, 34, 38).
The Fawn-Hooded rat (FHR) strain is characterized by platelet
abnormalities and systemic hypertension and has been used to study
genetic risk factors for the development of "idiopathic" pulmonary hypertension (20, 33). Unlike other rat strains, FHR develop
severe pulmonary hypertension when exposed to slight decreases in
alveolar PO2, such as at Denver's
altitude, even in the absence of hypoxemia (33). In a previous study, treatment of adult FHR with supplemental oxygen to simulate sea-level alveolar PO2 reduced the development
of pulmonary hypertension (33).
In preliminary studies, we have observed that, in addition to
structural hypertensive vascular disease, histology of the adult FHR
lung is also characterized by a striking pattern of alveolar simplification, reflecting decreased alveolarization and lung hypoplasia (21). Based on these findings, we hypothesized that abnormalities in perinatal lung growth and development could predispose the FHR for subsequent development of pulmonary hypertension. Alternatively, abnormalities of pulmonary vascular development may
contribute to decreased alveolarization. To test these hypotheses, we
characterized maturational changes in lung histology in FHR at fetal
and postnatal ages and determined whether perinatal exposure to
supplemental oxygen during a critical period of lung development would
improve alveolarization and reduce development of pulmonary hypertension. Lung arterial structure and density were studied in adult
FHR using barium-gelatin angiograms. We report that lung weight and
alveolar number were reduced in the FHR fetus and neonate and that
adult FHR had persistent reductions in alveolarization. Barium-gelatin
angiograms showed reduced arterial filling and density in FHR lungs. In
addition, we report that perinatal oxygen treatment during the critical
period of distal lung growth reduced the severity of lung hypoplasia
and right ventricular hypertrophy in adult FHR.
 |
METHODS |
Animals. All procedures and protocols
were approved by the Animal Care and Use Committee at the University of
Colorado Health Science Center. FHR were a gift from Dr. T. Stelzner
(Department of Medicine, University of Colorado School of Medicine),
and a breeding colony was established. Sprague-Dawley rats (SDR) and Fischer (FSR) rats were used as normal rat strains for these studies (Harlan Laboratories, Indianapolis, IN). Pregnant SDR and FSR were
purchased and maintained at Denver's altitude at least 1 wk before
giving birth. Animals were fed ad libitum and were exposed to 12:12-h
day-night cycles. Fetal rat lung tissue was obtained at 20 days of
gestation (term = 21 days) after killing of pregnant females by lethal
injection of pentobarbital sodium. Neonatal rats were also killed for
study by lethal injection at 1 day and at 1, 3, and 10 wk of age. Lung
and body weights and hematocrits were measured.
Oxygen therapy. Rats were placed in a
hyperbaric chamber in which the pressure was increased to 18.5-19
kPa to simulate sea-level alveolar
PO2. The chamber air changes 42 times
every hour. Carbon dioxide and ammonia buildup were prevented by having absorbent systems present in the chamber. A 12:12-h light-dark cycle
was provided by lights in the chamber, controlled by timers. Food and
water were supplied ad libitum. Rats were briefly exposed to Denver's
altitude for <10 min two times a week while the cages were changed
and fresh water and food were supplied. FHR were exposed to oxygen
therapy for 1 wk prenatal (pregnant female placed in chamber) and/or 3 or 10 wk postnatal.
Lung histology and radial alveolar
counts. Rat lungs were fixed for histology by tracheal
installation of 10% buffered Formalin under constant pressure (10 cmH2O). The trachea was ligated
after sustained inflation, and the lungs were excised and immersed in Formalin overnight. Formalin-fixed lung tissue was cut into 4- to
5-mm-thick sections, placed in 10% buffered Formalin, and embedded in
paraffin. Paraffin sections (5 µm thick) were serially mounted on
Superfrost Plus slides (Fisher Scientific, Fair Lawn, NJ) and were
stained with hematoxylin and eosin. At least three lung sections were
assessed from each animal for analysis. Alveolarization was assessed by
the radial alveolar count (RAC) method of Cooney and Thurlbeck (13) and
Emery and Mithal (18). Radial counts were performed by identifying
respiratory bronchioles as described by Randell et al. (29). From the
center of the bronchiole, a perpendicular line was taken to the edge of
the acinus (connective tissue septum or pleura), and the number of
alveoli that intersected the line was counted. For each lung section,
at least 10 counts were performed.
Right ventricular hypertrophy
measurements. At death, hearts from adult rats were
removed and dissected to isolate the free wall of the right ventricle
from the left ventricle and septum. The ratio of right ventricle weight
to left ventricle plus septum weight (RV/LV+S) was used as an index of
right ventricular hypertrophy (23).
Vessel morphometry. Morphometry was
performed on small pulmonary arteries (20-80 µm) on hematoxylin
and eosin-stained lung sections using a Zeiss Interactive Digital
Analysis System. Wall thickness and external diameter were directly
measured; percent wall thickness was calculated as (2 × wall
thickness/vessel diameter) × 100 to assess medial hypertrophy
(2).
In situ hybridization for surfactant protein C and
10-kDa Clara cell secretory protein. Paraffin-embedded
lung tissue sections were hybridized with
[33P]cRNA probes for
surfactant protein C (SP-C) and 10-kDa Clara cell secretory protein
(CC10; see Ref. 17). Hybridized sections were dipped in Kodak NTB-2
emulsion, developed after an appropriate exposure for each probe, and
counterstained with hematoxylin.
Barium-gelatin angiograms and arterial density
counts. Adult FHR and SDR were anesthetized with
pentobarbital sodium, and a rapid thoracotomy was performed. Rats were
heparinized by injection of heparin (500 units) into the right
ventricle. A tracheostomy was performed, and the lungs were inflated
with air. Blood was flushed from the lungs with heparinized saline (1 U/ml) through a main pulmonary artery catheter. A barium-gelatin
mixture was infused at 73 mmHg pressure in the main pulmonary artery
catheter for 3-4 min until surface filling of vessels with barium
was seen uniformly over the surface of the lung as described by deMello et al. (16). The main pulmonary artery was tied off under pressure, and
the lungs were inflation fixed with Formalin. Left lungs were excised
after fixation and imaged to X-ray radiography (16). After radiography,
left lungs were embedded in paraffin, and sections were cut and stained
with hematoxylin and eosin. Barium-filled pulmonary arteries were
counted per high-powered field (×100 magnification). At least
five fields were counted per animal. High-powered fields of peripheral
lung next to the pleural surface were counted. Fields containing large
airways or major vessels were avoided.
Statistical analysis. Data are
presented as means ± SE. Statistical analysis was performed with
the Statview software package (Abacus Concepts, Berkeley, CA).
Statistical comparisons were made using ANOVA and Fisher's protected
least significant difference test.
P < 0.05 was considered significant.
All animal groups contained at least four rats.
 |
RESULTS |
Lung and body weights, hematocrits, and right
ventricular hypertrophy. Lung and body weights were
lower in FHR at all ages compared with SDR
(P < 0.05; Table
1). Lung-to-body weight ratios were reduced
by 32% in fetal, 29% in 1-day-old, and 16% in 1-wk-old FHR compared
with SDR (P < 0.05; Fig.
1). At 3 and 10 wk of age, lung-to-body
weight ratios were not different between study groups (P > 0.05). Hematocrits were not
different for 10-wk-old Denver-raised SDR and FHR (53 ± 1 vs. 52 ± 1, respectively), suggesting that FHR were not chronically
hypoxic at Denver's altitude and that pulmonary hypertension in this
strain is not due to polycythemia. To assess right ventricular
hypertrophy in adult FHR and SDR, we measured RV and LV+S weights.
RV/LV+S was 80% higher in 10-wk-old FHR with to SDR, indicating
significant right ventricular hypertrophy in adult FHR (Fig.
2).

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Fig. 1.
Lung-to-body weight ratios versus age for the Fawn-Hooded rat (FHR) and
Sprague-Dawley rat (SDR) strains. As shown, the ratio of lung to body
weight is reduced during the first week in FHR due to a
disproportionate lung weight.
* P < 0.05 in comparisons of
values between FHR and SDR.
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Fig. 2.
Assessments of right ventricular hypertrophy in the FHR and normal rat
strain (SDR). The ratios of right ventricle to left ventricle plus
septum weights (RV/LV+S), an assessment of right ventricular
hypertrophy, are compared at 10 wk of age in animals raised at
Denver's altitude. As shown, RV/LV+S is markedly increased in adult
FHR compared with SDR. * P < 0.05 in comparisons of values between FHR and SDR.
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Lung histology, morphometry, epithelial marker
expression, and barium-gelatin angiograms and arterial density
counts. Histology revealed striking differences in lung
structure in 3-wk (infant)- and 10-wk (adult)-old FHR compared with SDR
(Fig. 3). Lung histology was also examined
in FSR and was found to be similar to SDR at all ages (data not shown).
Compared with SDR, 3-wk-old FHR lungs showed a pattern of alveolar
simplification characterized by larger and fewer air spaces, with a
thickened interstitium (Fig. 3). At 10 wk, the adult FHR lungs had
persistence of this pattern. Distal air spaces appeared simple and
abnormal (Fig. 3). Lung architecture appears less mature in fetal and
neonatal FHR lungs as reflected by marked interstitial thickening and
cellularity and decreased formation of distal air saccules (Fig.
4). To quantitate changes in alveolar
number, we measured RAC in the lungs of FHR and SDR at ages 1, 3, and
10 wk. RAC were reduced by 42% at 1 wk, 48% at 3 wk, and 50% at 10 wk of age compared with SDR (P < 0.05; Fig. 5). These findings further
demonstrate decreased alveolarization in addition to the striking
histological appearance of lung hypoplasia with alveolar
simplification. RAC could not be performed before 1 wk of age in either
SDR or FHR, since respiratory bronchioles are generally not clearly
identifiable at these early ages.

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Fig. 3.
Alveolar simplification in the FHR. In contrast to the larger number
and smaller size of alveoli in the distal lung of SDR, lung histology
demonstrates a pattern of "alveolar simplification" in 3- and
10-wk-old adult FHR. Micrographs are representative and are at the same
magnification (×40).
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Fig. 4.
Abnormal lung histology in the fetal and newborn FHR. In contrast to a
normal rat strain (SDR), there is a persistent interalveolar
parenchymal thickening in fetal and neonatal (1-day-old and 1-wk-old)
lung of the FHR. Micrographs are representative and are at the same
magnification (×40).
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Fig. 5.
Decreased alveolarization in the FHR. Compared with SDR, alveolar
number, as determined by measurements of radial alveolar counts, is
markedly reduced in FHR at all ages.
* P < 0.05 in comparisons of
values between FHR and SDR.
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To assess lung epithelial maturation, we performed in situ
hybridization of lung sections from fetal and 1- and 10-wk-old FHR and
SDR lungs for CC10 mRNA in bronchial epithelial cells and SP-C mRNA in
type II epithelial cells (Fig. 6). Despite
differences in lung architecture between the strains, the patterns of
SP-C and CC10 expression appeared similar in FHR and SDR lungs at each of the study ages.


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Fig. 6.
Expression of the distal epithelial marker surfactant protein C (SP-C;
A) and proximal epithelial marker
10-kDa Clara cell secretory protein (CC10;
B) in FHR and SDR lung. In situ
hybridization analysis of lung sections shows that the pattern of
expression for SP-C and CC10 mRNAs appears similar in FHR compared with
SDR. Micrographs are representative and are at the same magnification
(×40).
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To examine pulmonary arterial structure and to determine arterial
density, we infused heated barium-gelatin mixtures into the main
pulmonary artery of adult FHR and SDR under high pressure (73 mmHg) as
previously described (16). Angiograms of left lungs showed reduced
barium filling in adult FHR compared with SDR (Fig. 7). Barium-filled pulmonary artery counts
were 35% lower in FHR compared with SDR
(P < 0.05), indicating a
reduction in arterial density (Fig. 8).

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Fig. 7.
Barium-gelatin angiograms of the left lungs of 3 adult SDR compared
with 3 adult FHR. Barium filling was reduced in FHR lungs compared with
SDR.
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Fig. 8.
Pulmonary arterial density is reduced in adult FHR. Pulmonary artery
vessel counts were performed on barium-filled Formalin-fixed left lung
sections. Barium-filled pulmonary arteries were counted per
high-powered field (×100 magnification). Counts were performed on
4 adult SDR and 4 adult FHR lungs.
* P < 0.05 in comparisons of
values between FHR and SDR.
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Effects of brief and prolonged supplemental oxygen
exposure on lung histology, RAC, and right ventricular
hypertrophy. To determine if supplemental oxygen
exposure during the critical period of lung development could increase
alveolarization and reduce pulmonary hypertension in adult FHR, we
studied the effects of prenatal (1 wk) and postnatal brief (3 wk) and
prolonged (10 wk) periods of mild hyperbaria. Compared with controls,
oxygen treatment improved alveolarization and increased RAC in the
10-wk-old FHR lungs (Figs. 9 and
10). Compared with Denver-raised FHR
(control), prenatal oxygen treatment alone did not improve
alveolarization in 10-wk-old FHR (P > 0.05; Fig. 9). However, 3 wk of postnatal supplemental oxygen
increased RAC by 63% (P < 0.05).
Combined prenatal and postnatal supplemental oxygen for 3 or 10 wk
increased RAC by 105 and 114%, respectively
(P < 0.05; Fig. 10).

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Fig. 9.
Supplemental oxygen supplied by hyperbaric exposure improves
alveolarization in the FHR. Lung histology of 10-wk-old Denver-raised
FHR compared with FHR exposed to prenatal and 10 wk of postnatal
supplemental oxygen and 10-wk-old SDR raised at Denver's altitude.
Micrographs are representative and are at the same magnification
(×40).
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Fig. 10.
Alveolarization in FHR improves with supplemental oxygen supplied by
hyperbaric exposure. Alveolar number was increased by 3 wk of postnatal
supplemental oxygen and by a combination of prenatal and 3 or 10 wk of
postnatal supplemental oxygen but not by prenatal supplemental oxygen
alone. NS, not significantly different.
* P < 0.05 vs. prenatal oxygen
group; # P < 0.05 vs. 3-wk postnatal oxygen group.
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Prenatal treatment with supplemental oxygen alone did not reduce right
ventricular hypertrophy as assessed by RV/LV+S (Fig. 11). However, 3 wk of postnatal
supplemental oxygen reduced RV/LV+S 8% and combined prenatal and 3 or
10 wk of postnatal oxygen therapy reduced RV/LV+S by 26 and 29%,
respectively, from control values (Fig. 10). Compared with
Denver-raised FHR (control), the wall thickness of small pulmonary
arteries (with external diameters ranging between 20 and 80 µm) was
reduced 30 and 39%, respectively, by combined prenatal and 3 or 10 wk
of postnatal supplemental oxygen (P < 0.05) but not by prenatal oxygen therapy alone
(P > 0.05; Fig.
12).

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Fig. 11.
Reduced RV/LV+S weights in adult FHR with supplemental oxygen supplied
by hyperbaric exposure. RV/LV+S weights were reduced by 3 wk of
postnatal supplemental oxygen or combined prenatal and 3 or 10 wk of
postnatal supplemental oxygen but not by prenatal supplemental oxygen
alone. * P < 0.05 vs. prenatal
oxygen group;
# P < 0.05 vs. 3-wk postnatal oxygen group.
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Fig. 12.
Reduced small pulmonary artery hypertrophy in adult FHR with
supplemental oxygen supplied by hyperbaric exposure. Small pulmonary
artery (20-80 µm) hypertrophy measured by (2 × wall
thickness/vessel diameter) × 100 is reduced by prenatal and
either 3 or 10 wk of postnatal supplemental oxygen but not by prenatal
supplemental oxygen alone. * P < 0.05 in comparison with Denver-raised control group.
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DISCUSSION |
The FHR is a genetic rat strain that is characterized by the
development of severe pulmonary hypertension in adults (20). In
contrast with other rat strains, the development of pulmonary hypertension in FHR is enhanced by modest decreases in alveolar PO2 despite the absence of chronic
hypoxemia (33). Although the FHR strain has previously been used as a
model to study genetic factors that contribute to idiopathic or
"primary" pulmonary hypertension (33, 37, 39), we now report that alveolarization is markedly reduced in the FHR. We found abnormal lung
development in FHR during the perinatal period, and this resulted in
enlarged distal air spaces in adults. In situ hybridization analysis of
two markers of lung epithelial differentiation, SP-C and CC10, showed
similar patterns of expression in both FHR and SDR, suggesting that the
FHR lung is not characterized by global delays in epithelial maturation
but rather a more specific failure of alveolar formation. In addition,
barium-gelatin angiograms showed reduced filling in adult FHR lungs
compared with SDR, and barium-filled arterial counts indicated a
reduction in arterial density.
We also report that treatment of FHR with supplemental oxygen
(postnatal or combined prenatal and 3 or 10 wk postnatal) to simulate
sea-level PO2 improved
alveolarization and reduced the development of pulmonary hypertension
as reflected by measurements of right ventricular hypertrophy and wall
thickness of small pulmonary arteries. Prenatal treatment with
supplemental oxygen alone had no effect on alveolarization or markers
of pulmonary hypertension, but 3 wk of postnatal treatment increased
alveolarization and reduced right ventricular hypertrophy in 10-wk-old
adult FHR. Combined prenatal with 3 wk of postnatal oxygen treatment
increased alveolarization and reduced RV/LV+S further compared with 3 wk of postnatal treatment alone. However, no further improvement was
seen with combined prenatal and 10 wk of postnatal oxygen treatment
compared with combined prenatal and 3 wk of postnatal treatment. In
SDR, exposure to brief perinatal hypoxia (10% fraction of inspired
oxygen; 9 h prenatal and 2 h postnatal) reduces lung alveolarization at
1 wk of age (27). These findings suggest that transient decreases in
alveolar PO2 in the perinatal period
can have long-lasting effects on alveolarization in normal rats. The
FHR at Denver's altitude show similar findings, and reduced
alveolarization was partially prevented by low levels of supplemental
oxygen in the perinatal period. Based on these observations, we
speculate that the FHR exhibits a marked sensitivity to mild decreases
in alveolar PO2 during a critical
period of lung development that contributes to abnormal lung growth and pulmonary hypertension. The mechanisms that account for this altered oxygen sensitivity in the FHR are unknown.
Although initially characterized by platelet abnormalities and systemic
hypertension (39), it was subsequently recognized that the FHR strain
develops progressive pulmonary hypertension (20), especially after
slight reductions in alveolar PO2 (33). Lung endothelin (ET)-1 peptide levels are elevated in FHR
compared with SDR before the development of pulmonary hypertension (37). Whether increased lung ET-1 levels might also be responsible for
the abnormal lung development that we report in the FHR is unknown. At
least two studies have suggested that the platelet storage disease is
not involved in the pathogenesis of pulmonary hypertension in the FHR
(4, 20). The incidence of increased pulmonary artery pressure in FHR
was different from the incidence of the platelet storage defect (20),
and platelets from both SDR and FHR constricted pulmonary arteries from
FHR (4). A preliminary report has localized the high pulmonary artery
pressure in this strain to a genetic locus, PH1, on chromosome 1 (36). This locus did not co-segregate with ET-1, ET-2, or the ETA
receptor genes, suggesting that these genes may not be involved in the pathogenesis of pulmonary hypertension in the FHR. Whether the genetic
basis for the abnormal lung development that we report also
co-segregates to the PH1 locus is unknown.
Although past studies have considered the FHR strain as a model of
idiopathic pulmonary hypertension, we have observed that the adult FHR
lung is characterized by striking reductions in alveolar number and
that this pattern is already present in the early postnatal period,
well before the earliest age (4 wk) that pulmonary hypertension has
been reported in FHR (33). Based on our observations that FHR strain is
characterized by abnormal alveolarization and marked susceptibility for
the development of progressive pulmonary hypertension, we propose that
there is a relationship between abnormal lung alveolar and vascular
growth with the development of pulmonary hypertension. Interestingly, in our study, perinatal treatment with supplemental oxygen during the
critical period of lung growth improved alveolar number in FHR and
prevented the development of pulmonary hypertension in adult FHR.
This is the first report of developmental abnormalities in lung growth
in a genetic rat strain that is known to develop pulmonary hypertension. Among several animal models that have been developed to
study mechanisms of pulmonary hypertension, the FHR has provided a
unique model of genetic factors that may be linked with the development
of pulmonary hypertension (33, 37). Pulmonary hypertension contributes
significantly to high morbidity and mortality in several human
disorders of lung hypoplasia, including CDH, primary lung hypoplasia,
and lung hypoplasia associated with oligohydramnios or renal
dysfunction (19, 38). In addition, children with chronic lung disease
after premature birth (BPD) and with Down's syndrome (with or without
congenital heart disease) have abnormal lung growth that is
characterized by decreased alveolarization and high risk for pulmonary
hypertension (3, 11, 12, 14, 24, 32, 34). The pathogenic mechanisms
that link abnormal alveolarization with a high risk for the development
of pulmonary hypertension in the setting of lung hypoplasia are
unclear, and few animal models are currently available to study this
complex relationship.
Because reduction of lung surface area is associated with reduced
vascular growth, it is also likely that decreased cross-sectional area
of the pulmonary vascular bed is a significant contributing factor to
the persistence or late development of pulmonary hypertension in these
diseases (3, 11). Although causes of abnormal lung development are
uncertain, it is likely that disruption of alveolarization by adverse
intrauterine stimuli, premature birth, injury, and other mechanisms
contribute to this problem. Past studies have shown that exposure to
hypoxia, hyperoxia, or dexamethasone during the first 3 wk of life in a
rat decreases alveolar number (7, 25, 27-29).
Growth and remodeling of the lung occurs by the successive addition of
new generations of respiratory bronchioles, alveolar ducts and
saccules, and alveoli. Alveoli are the sites of gas exchange in the
lung and are formed by septation of large saccules that constitute the
gas-exchange region of the structurally immature lung (10, 22, 26).
Secondary septa initially form as low ridges that protrude into
primitive air spaces to increase their surface area. This growth
involves close spatial and temporal coordination between several cell
types regarding cell proliferation, differentiation, and matrix
production (31, 35). During the stages of alveolarization, the lung is
also undergoing marked vascular growth and development (5, 16).
Multiple paracrine stimuli, such as platelet-derived growth factor
(PDGF), transforming growth factor-
(TGF-
), VEGF, and the
angiopoietins, contribute to alveolar and vascular growth and
development, providing a "cross talk" of signals between
epithelium and mesenchyme (31, 35). In addition, the process of
septation, which is essential for alveolarization, involves alternate
upfolding of one of the two capillary layers on both sides of the
primary septa (10). This method of formation of alveolar walls suggests
that the failure of capillary network formation and maturation or
disruption of the upfolding of the double capillary network could
potentially cause failed alveolarization. Thus failure of growth or
maturation of the pulmonary microcirculation during the critical period
of alveolarization could potentially cause lung hypoplasia by
decreasing septation. Whether paracrine factors such as VEGF, PDGF, or
TGF-
are responsible for abnormal lung development in the FHR is unknown.
We conclude that lung growth is abnormal in the FHR and suggest that
decreased alveolarization and altered pulmonary vascular growth
contribute to the development of pulmonary hypertension in this strain.
We found that postnatal oxygen therapy during the critical period of
distal lung growth was sufficient to prevent the development of lung
hypoplasia and pulmonary hypertension in adult FHR. We speculate that
these findings may have important implications regarding perinatal
origins of adult cardiopulmonary disease. Epidemiological studies
suggest that factors affecting the fetus and the young may have
long-lasting effects as important causes of later diseases such
as ischemic heart disease, stroke, hypertension, chronic bronchitis,
and emphysema (5). For example, adverse stimuli during critical
periods of lung growth may impair lung structure and function and
may cause cardiopulmonary abnormalities later in life. We speculate
that the FHR strain provides a unique model for further investigation
of this hypothesis and mechanisms linking alveolarization, vascular
growth, and the development of pulmonary hypertension.
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ACKNOWLEDGEMENTS |
We thank Dr. Jen-Ruey Tang for performing hematocrit measurements
and Charles R. Ahrens for performing X-ray chromatography.
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FOOTNOTES |
This work was supported by an American Heart Association (National)
Scientist Development Grant to T. D. Le Cras; a Grant-in-Aid to S. H. Abman; and National Heart, Lung, and Blood Institute Specialized Center
of Research Grant HL-57144 to J. M. Shannon and S. H. Abman.
The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement"
in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Address for reprint requests and other correspondence: T. D. Le Cras,
Dept. of Pediatrics, Box C218, Univ. of Colorado Health Sciences
Center, 4200 E. Ninth Ave., Denver, CO 80262 (E-mail: Timothy.Lecras{at}uchsc.edu).
Received 5 April 1999; accepted in final form 27 May 1999.
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