VDUP1: a potential mediator of expansion-induced lung growth and epithelial cell differentiation in the ovine fetus

2006 ◽  
Vol 290 (2) ◽  
pp. L250-L258 ◽  
Author(s):  
C. E. Filby ◽  
S. B. Hooper ◽  
F. Sozo ◽  
V. A. Zahra ◽  
S. J. Flecknoe ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cell Differentiation ◽  
Epithelial Cell ◽  
Lung Development ◽  
Fetal Lung ◽  
Normal Lung ◽  
Mrna Levels ◽  
Lung Growth ◽  
Fetal Sheep ◽  
Lung Expansion

The degree of fetal lung expansion is a critical determinant of fetal lung growth and alveolar epithelial cell (AEC) differentiation, although the mechanisms involved are unknown. As VDUP1 (vitamin D3-upregulated protein 1) can modulate cell proliferation, can induce cell differentiation, and is highly expressed in the lung, we have investigated the effects of fetal lung expansion on VDUP1 expression and its relationship to expansion-induced fetal lung growth and AEC differentiation in fetal sheep. Alterations in fetal lung expansion caused profound changes in VDUP1 mRNA levels in lung tissue. Increased fetal lung expansion significantly reduced VDUP1 mRNA levels from 100 ± 8% in control fetuses to 37 ± 4, 46 ± 4, and 45 ± 9% of control values at 2, 4, and 10 days of increased fetal lung expansion, respectively. Reduced fetal lung expansion increased VDUP1 mRNA levels from 100 ± 16% in control fetuses to 162 ± 16% of control values after 7 days. VDUP1 was localized to airway epithelium in small bronchioles, AECs, and some mesenchymal cells. Its expression was inversely correlated with cell proliferation during normal lung development ( R2 = 0.972, P < 0.002) as well as in response to alterations in fetal lung expansion ( R2 = 0.956, P < 0.001) and was positively correlated with SP-B expression during normal lung development ( R2 = 0.803, P < 0.0001) and following altered lung expansion ( R2 = 0.817, P < 0.001). We suggest that VDUP1 may be an important mediator of expansion-induced lung cell proliferation and AEC differentiation in the developing lung.

The oncogeneTrop2regulates fetal lung cell proliferation

2011 ◽  
Vol 301 (4) ◽  
pp. L478-L489 ◽  
Author(s):  
Annie R. A. McDougall ◽  
Stuart B. Hooper ◽  
Valerie A. Zahra ◽  
Foula Sozo ◽  
Camden Y. Lo ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Fetal Lung ◽  
Alveolar Epithelial Cells ◽  
Lung Fibroblasts ◽  
Normal Lung ◽  
Lung Growth ◽  
Ki 67 ◽  
Alveolar Epithelial ◽  
Fetal Rat ◽  
Lung Expansion

The factors regulating growth of the developing lung are poorly understood, although the degree of fetal lung expansion is critical. The oncogene Trop2 (trophoblast antigen 2) is upregulated during accelerated fetal lung growth, and we hypothesized that it may regulate normal fetal lung growth. We investigated Trop2 expression in the fetal and neonatal sheep lung during accelerated and delayed lung growth induced by alterations in fetal lung expansion, as well as in response to glucocorticoids. Trop2 expression was measured using real-time PCR and localized spatially using in situ hybridization and immunofluorescence. During normal lung development, Trop2 expression was higher at 90 days gestational age (GA; 4.0 ± 0.8) than at 128 days GA (1.0 ± 0.1), decreased to 0.5 ± 0.1 at 142 days GA (full term ∼147 days GA), and was positively correlated to lung cell proliferation rates ( r = 0.953, P < 0.005). Trop2 expression was regulated by fetal lung expansion, but not by glucocorticoids. It was increased nearly threefold by 36 h of increased fetal lung expansion ( P < 0.05) and was reduced to ∼55% of control levels by reduced fetal lung expansion ( P < 0.05). Trop2 expression was associated with lung cell proliferation during normal and altered lung growth, and the TROP2 protein colocalized with Ki-67-positive cells in the fetal lung. TROP2 was predominantly localized to fibroblasts and type II alveolar epithelial cells. Trop2 small interfering RNA decreased Trop2 expression by ∼75% in cultured fetal rat lung fibroblasts and decreased their proliferation by ∼50%. Cell viability was not affected. This study demonstrates that TROP2 regulates lung cell proliferation during development.

Regulation of alveolar epithelial cell phenotypes in fetal sheep: roles of cortisol and lung expansion

2004 ◽  
Vol 287 (6) ◽  
pp. L1207-L1214 ◽  
Author(s):  
Sharon J. Flecknoe ◽  
Rochelle E. Boland ◽  
Megan J. Wallace ◽  
Richard Harding ◽  
Stuart B. Hooper
Keyword(s):  
Epithelial Cell ◽  
Alveolar Epithelial Cell ◽  
Fetal Lung ◽  
Saline Infusion ◽  
Type I ◽  
Mrna Levels ◽  
Type Ii ◽  
Fetal Sheep ◽  
Alveolar Epithelial ◽  
Lung Expansion

Our aim was to determine whether cortisol's effect on alveolar epithelial cell (AEC) phenotypes in the fetus is mediated via a sustained alteration in lung expansion. Chronically catheterized fetal sheep were exposed to 1) saline infusion, 2) cortisol infusion (122–131 days' gestation, 1.5–4.0 mg/day), 3) saline infusion plus reduced lung expansion, or 4) cortisol infusion plus reduced lung expansion. The proportions of type I and II AECs were determined by electron microscopy, and surfactant protein (SP)-A, -B, and -C mRNA levels were determined by Northern blot analysis. Cortisol infusions significantly increased type II AEC proportions (to 38.2 ± 2.2%), compared with saline-infused fetuses (23.8 ± 2.4%), and reduced type I AEC proportions (to 59.0 ± 2.2%), compared with saline-infused fetuses (70.4 ± 2.4%). Reduced lung expansion also increased type II AEC proportions (to 52.9 ± 3.5%) and decreased type I AEC proportions (to 34.2 ± 3.7%), compared with control, saline-infused fetuses. The infusion of cortisol into fetuses exposed to reduced lung expansion tended to further increase type II (to 60.3 ± 2.1%, P = 0.066) and reduce type I AEC (to 26.6 ± 2.3%, P = 0.07) proportions. SP-A, -B, and -C mRNA levels changed in parallel with the changes in type II AEC proportions. These results indicate that cortisol alters the proportion of type I and type II AECs via a mechanism unrelated to the degree of fetal lung expansion. However, reductions in fetal lung expansion appear to have a greater impact on the proportion of AECs than cortisol.

Alterations in lung expansion affect surfactant protein A, B, and C mRNA levels in fetal sheep

1999 ◽  
Vol 276 (2) ◽  
pp. L239-L245 ◽  
Author(s):  
A. Lines ◽  
L. Nardo ◽  
I. D. Phillips ◽  
F. Possmayer ◽  
S. B. Hooper
Keyword(s):  
Lung Tissue ◽  
Blot Analysis ◽  
Fetal Lung ◽  
Surfactant Protein ◽  
Mrna Levels ◽  
Lung Growth ◽  
Fetal Sheep ◽  
Protein Levels ◽  
Tracheal Obstruction ◽  
Lung Expansion

Obstruction of the fetal trachea is a potent stimulus for fetal lung growth, and it has been suggested that this procedure may be used therapeutically to reverse lung growth deficits in human fetuses with lung hypoplasia. However, little is known about the effects of increased lung expansion on other aspects of lung development. Our aim was to determine the effect of increased and decreased lung expansion on the mRNA levels encoding surfactant protein (SP) A, SP-B, and SP-C in ovine fetal lungs. Lung tissue samples were collected from fetuses exposed to 2, 4, or 10 days of increased lung expansion caused by tracheal obstruction. The mRNA levels for SP-A, SP-B, and SP-C were determined by Northern blot analysis with specific ovine cDNA probes; SP-A protein levels were determined by Western blot analysis. Compared with age-matched (128-day gestational age) control fetuses, SP-A, SP-B, and SP-C mRNA levels in fetal lung tissue were significantly reduced at 2 days of tracheal obstruction and remained reduced at 4 and 10 days. However, SP-A protein levels were not reduced at 2 days of tracheal obstruction, tended to be reduced at 4 days, and were almost undetectable at 10 days. In contrast to tracheal obstruction, 7 days of lung liquid drainage significantly increased SP-C, but not SP-A, mRNA levels in fetal lung tissue compared with age-matched control fetuses. Our results demonstrate that increases in fetal lung expansion, induced by obstruction of the fetal trachea, cause large simultaneous reductions in SP-A, SP-B, and SP-C mRNA levels in the fetal lung as well as a decrease in SP-A protein levels. These data suggest that expression of the genes encoding SPs in the fetal lung are specifically responsive to the degree of lung expansion.

scholarly journals Increased expansion of the lung stimulates calmodulin 2 expression in fetal sheep

2002 ◽  
Vol 282 (3) ◽  
pp. L440-L447 ◽  
Author(s):  
A. M. Gillett ◽  
M. J. Wallace ◽  
M. T. Gillespie ◽  
S. B. Hooper
Keyword(s):  
Northern Blot Analysis ◽  
Fetal Lung ◽  
Mrna Levels ◽  
Lung Growth ◽  
Coding Region ◽  
Fetal Sheep ◽  
Protein Levels ◽  
Lung Expansion ◽  
Rapid Amplification

Obstruction of the fetal trachea causes the lungs to expand with accumulated liquid. Although this is a potent stimulus for lung growth, the mechanisms involved are unknown. Our aim was to identify genes that are differentially expressed as a result of increased fetal lung expansion. Using differential display RT-PCR, we isolated a cDNA fragment partially encoding calmodulin 2 (CALM2) and identified the remainder of the coding region by 5′-rapid amplification of cDNA ends. Differential expression of CALM2 was confirmed by Northern blot analysis; CALM2 mRNA levels were increased to 161 ± 5% of control at 2 days of increased lung expansion, induced by tracheal obstruction (TO), and had returned to control levels at days 4 and 10. Using in situ hybridization analysis, we found that the proportion of CALM2-labeled cells increased from 10.3 ± 1.0% to 21.4 ± 6.8% by 2 days of TO. This increase in CALM2 expression was reflected by a tendency for calmodulin protein levels to increase from 122.7 ± 17.3 to 156.5 ± 17.7 at 2 days of TO. Thus increases in fetal lung expansion result in time-dependent changes in CALM2 mRNA levels, which closely parallels the changes in lung DNA synthesis rates. As calmodulin is essential for cell proliferation, increased CALM2 mRNA levels may reflect an important role for calmodulin in expansion-induced fetal lung growth.

Increased lung expansion alters lung growth but not alveolar epithelial cell differentiation in newborn lambs

2007 ◽  
Vol 292 (2) ◽  
pp. L454-L461 ◽  
Author(s):  
S. J. Flecknoe ◽  
K. J. Crossley ◽  
G. M. Zuccala ◽  
J. E. Searle ◽  
B. J. Allison ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Epithelial Cell ◽  
Alveolar Epithelial Cell ◽  
Surfactant Protein ◽  
Mrna Levels ◽  
Lung Growth ◽  
Proliferating Cells ◽  
Alveolar Epithelial ◽  
Epithelial Cell Differentiation ◽  
Lung Expansion

Although increased lung expansion markedly alters lung growth and epithelial cell differentiation during fetal life, the effect of increasing lung expansion after birth is unknown. We hypothesized that increased basal lung expansion, caused by ventilating newborn lambs with a positive end-expiratory pressure (PEEP), would stimulate lung growth and alter alveolar epithelial cell (AEC) proportions and decrease surfactant protein mRNA levels. Two groups of lambs were sedated and ventilated with either 0 cmH2O PEEP (controls, n = 5) or 10 cmH2O PEEP ( n = 5) for 48 h beginning at 15 ± 1 days after normal term birth. A further group of nonventilated 2-wk-old lambs was used for comparison. We determined wet and dry lung weights, DNA and protein content, a labeling index for proliferating cells, surfactant protein mRNA expression, and proportions of AECs using electron microscopy. Although ventilating lambs for 48 h with 10 cmH2O PEEP did not affect total lung DNA or protein, it significantly increased the proportion of proliferating cells in the lung when compared with nonventilated 2-wk-old controls and lambs ventilated with 0 cmH2O PEEP (control: 2.6 ± 0.5%; 0 PEEP: 1.9 ± 0.3%; 10 PEEP: 3.5 ± 0.3%). In contrast, no differences were observed in AEC proportions or surfactant protein mRNA levels between either of the ventilated groups. This study demonstrates that increases in end-expiratory lung volumes, induced by the application of PEEP, lead to increased lung growth in mechanically ventilated 2-wk-old lambs but do not alter the proportions of AECs.

Changes in lung expansion alter pulmonary DNA synthesis and IGF-II gene expression in fetal sheep

1993 ◽  
Vol 265 (4) ◽  
pp. L403-L409 ◽  
Author(s):  
S. B. Hooper ◽  
V. K. Han ◽  
R. Harding
Keyword(s):  
Gene Expression ◽  
Dna Synthesis ◽  
Fetal Lung ◽  
Liquid Volume ◽  
Mrna Levels ◽  
Pulmonary Tissue ◽  
Fetal Sheep ◽  
Tracheal Obstruction ◽  
Lung Expansion ◽  
Lung Liquid

Our aim was to determine the effect of short-term (7 days) alterations in fetal lung liquid volume on pulmonary DNA synthesis rates and insulin-like growth factor-II (IGF-II) mRNA levels. Fifteen chronically catheterized fetal sheep were divided into three groups. In one, the trachea was obstructed, in another lung liquid was drained by gravity, and the third group served as controls. After 7 days, [3H]thymidine was injected into each fetus and 8 h later fetal tissues were collected. Fetal lung-to-body weight ratios and total lung DNA contents were greatly increased in fetuses with tracheal obstruction compared with control fetuses, whereas the drainage of lung liquid did not affect these measurements. DNA synthesis rates in pulmonary tissue were significantly reduced from a mean control value of 153.3 +/- 25.1 disintegrations per minute (dpm)/microgram DNA to 57.2 +/- 8.6 dpm/microgram DNA by lung liquid drainage (P < 0.05) and were significantly increased to 236.0 +/- 24.0 dpm/microgram DNA by tracheal obstruction (P < 0.05). Following tracheal obstruction, lung IGF-II mRNA levels were increased to 177.0 +/- 18.2% (P < 0.05) of the mean value for control fetuses, whereas they were reduced to 56.1 +/- 7.1% of control in lung liquid-drained fetuses. We conclude that altering fetal lung expansion has a potent and rapid effect on pulmonary DNA synthesis and that this effect may, in part, be mediated by an alteration in IGF-II gene expression.

Sustained changes in lung expansion alter tropoelastin mRNA levels and elastin content in fetal sheep lungs

2003 ◽  
Vol 284 (4) ◽  
pp. L643-L649 ◽  
Author(s):  
Belinda J. Joyce ◽  
Megan J. Wallace ◽  
Richard A. Pierce ◽  
Richard Harding ◽  
Stuart B. Hooper
Keyword(s):  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Fetal Lung ◽  
Elastic Fibers ◽  
Mrna Levels ◽  
Fetal Sheep ◽  
Time Points ◽  
Tracheal Obstruction ◽  
Lung Expansion

Our objective was to determine the effects of sustained alterations in fetal lung expansion on pulmonary elastin synthesis. In fetal sheep, lung expansion was either decreased between 111 and 131 days' gestation (term ∼147 days) by tracheal drainage or increased for 2, 4, 7, or 10 days by tracheal obstruction, ending at 128 days' gestation. Lung tropoelastin mRNA levels were assessed by Northern blot analysis, total elastin content was measured biochemically, and staining of lung sections was used to assess the localization and form of elastic fibers. Tracheal obstruction significantly elevated pulmonary tropoelastin mRNA levels 2.5-fold at 2 days, but values were not different from controls at 4, 7, and 10 days; elastin content tended to be increased at all time points. A sustained decrease in lung expansion by tracheal drainage reduced pulmonary tropoelastin mRNA levels 2.5-fold; elastin content was also decreased compared with controls, and tissue localization was altered. Our results indicate that the degree of lung expansion in the fetus influences elastin synthesis, content, and tissue deposition.

scholarly journals Influence of growth hormone on the lung growth response to tracheal obstruction in fetal sheep

2000 ◽  
Vol 278 (3) ◽  
pp. L453-L459 ◽  
Author(s):  
L. Nardo ◽  
I. R. Young ◽  
S. B. Hooper
Keyword(s):  
Growth Hormone ◽  
Dna Content ◽  
Fetal Lung ◽  
Control Group ◽  
Lung Growth ◽  
Lung Weight ◽  
Fetal Sheep ◽  
Tracheal Obstruction ◽  
Lung Expansion ◽  
Lung Liquid

Obstructing the fetal trachea is a potent stimulus for fetal lung growth, but little is known about the factors that regulate this process. Our aim was to determine the role of growth hormone (GH) in regulating the increase in lung growth induced by obstruction of the trachea in fetal sheep. Twenty chronically catheterized fetal sheep, nine of which were hypophysectomized, were divided into four experimental groups: 1) control group ( n = 4), 2) a group in which the fetal trachea was obstructed for 3 days (3-day obstructed; n = 6), 3) a 3-day obstructed group in which the pituitary was removed [hypophysectomized (HX)] and the fetus was given maintenance infusions of ACTH, thyroxine, and human GH (hGH; HX hGH 3-day obstructed; n = 5), and 4) a HX 3-day obstructed group in which the fetus was given maintenance infusions of ACTH and thyroxine ( n = 5). Tracheal obstruction significantly increased fetal lung liquid volumes from 37.2 ± 3.2 ml/kg in control fetuses to 75.6 ± 9.0 ml/kg in 3-day obstructed fetuses, and the presence or absence of GH did not affect this increase. Similarly, the presence or absence of GH did not affect the increase in lung weight or protein content induced by 3 days of tracheal obstruction. However, in the absence of GH, 3 days of tracheal obstruction failed to increase total lung DNA content above unobstructed control values (107.9 ± 5.3 and 94.1 ± 7.0 mg/kg for control and HX 3-day obstructed groups, respectively). In contrast, 3 days of tracheal obstruction increased total lung DNA content to a similar extent in fetuses with an intact pituitary and HX fetuses that received GH replacement (126.0 ± 4.4 and 126.7 ± 4.0 mg/kg for 3-day obstructed and HX hGH 3-day obstructed groups, respectively). These data indicate that the absence of GH either abolishes or delays the acceleration in cell division caused by an increase in fetal lung expansion.

scholarly journals Glucocorticoid signalling drives reduced versican levels in the fetal mouse lung

2020 ◽  
Vol 64 (3) ◽  
pp. 155-164
Author(s):  
Kelly L Short ◽  
A Daniel Bird ◽  
Bennet K L Seow ◽  
Judy Ng ◽  
Annie R A McDougall ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Fetal Lung ◽  
Lung Fibroblasts ◽  
Normal Lung ◽  
Mouse Lung ◽  
Mrna Levels ◽  
Lung Maturation ◽  
Fetal Mouse ◽  
Distal Lung

Glucocorticoid (GC) signaling via the glucocorticoid receptor (GR) is essential for lung maturation in mammals. Previous studies using global or conditional mouse model knockouts of the GR gene have established that GR-mediated signaling in the interstitial mesenchyme of the fetal lung is critical for normal lung development. Screens for downstream GC-targets in conditional mesenchymal GR deficient mouse lung (GRmesKO) identified Versican (Vcan), an important extracellular matrix component and cell proliferation regulator, as a potential GR-regulated target. We show that, of the five major VCAN isoforms, the VCAN-V1 isoform containing the GAGβ domain is the predominant VCAN isoform in the fetal mouse lung distal mesenchyme at both E16.5 and E18.5, whereas the GAGα-specific VCAN-V2 isoform was only localized to the smooth muscle surrounding proximal airways. Both Vcan-V1 mRNA and protein levels were strongly overexpressed in the GRmesKO lung at E18.5. Finally, we investigated the GC regulation of the ECM protease ADAMTS 12 and showed that Adamts 12 mRNA levels were markedly reduced at E18.5 in GRmesKO fetal mouse lung and were strongly induced by both cortisol and betamethasone in cultures of primary rat fetal lung fibroblasts. ADAMTS12 protein immunoreactivity was also strongly increased in the distal lung at E18.5, after dexamethasone treatment in utero. In summary, glucocorticoid signaling via GR represses GAGβ domain-containing VCAN isoforms in distal lung mesenchyme in vivo by repressing Vcan gene expression and, in part, by inducing the ECM protease ADAMTS12, thereby contributing to the control of ECM remodelling and lung cell proliferation prior to birth.

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