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.

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.

Cyclic mechanical stretch inhibits cell proliferation and induces apoptosis in fetal rat lung fibroblasts

2002 ◽  
Vol 282 (3) ◽  
pp. L448-L456 ◽  
Author(s):  
Juan Sanchez-Esteban ◽  
Yulian Wang ◽  
Lawrence A. Cicchiello ◽  
Lewis P. Rubin
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Cell Cycle Progression ◽  
Fetal Lung ◽  
Mechanical Stretch ◽  
Lung Fibroblasts ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Dna Flow Cytometry ◽  
Fetal Rat ◽  
Induced Apoptosis

Development of the pulmonary air sacs is crucial for extrauterine survival. Late fetal lung development is characterized by a thinning of the mesenchyme, which brings pneumocytes and endothelial cells into apposition. We hypothesized that mechanical stretch, simulating fetal breathing movements, plays an important role in this remodeling process. Using a Flexercell Strain Unit, we analyzed the effects of intermittent stretch on cell proliferation and apoptosis activation in fibroblasts isolated from fetal rat lungs during late development. On day 19, intermittent stretch increased cells in G0/G1 by 22% ( P = 0.001) and decreased in S phase by 50% ( P = 0.003) compared with unstretched controls. Cell proliferation analyzed by 5-bromo-2′-deoxyuridine incorporation showed a similar magnitude of cell cycle arrest ( P = 0.04). At this same gestational age, stretch induced apoptosis by two- to threefold over controls, assayed by DNA flow cytometry, terminal deoxynucleotidyl transferase-mediated dUTP-FITC nick-end labeling, and caspase-3 activation. These results indicate that mechanical stretch of fibroblasts isolated during the canalicular stage inhibits cell cycle progression and activates apoptosis. These findings are cotemporal with the mesenchymal thinning that normally occurs in situ.

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.

Identification of Betamethasone-Regulated Target Genes and Cell Pathways in Fetal Rat Lung Mesenchymal Fibroblasts

Endocrinology ◽  
2019 ◽  
Vol 160 (8) ◽  
pp. 1868-1884 ◽  
Author(s):  
Bennet K L Seow ◽  
Annie R A McDougall ◽  
Kelly L Short ◽  
Megan J Wallace ◽  
Stuart B Hooper ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Preterm Birth ◽  
Transcriptome Sequencing ◽  
Target Genes ◽  
Fetal Lung ◽  
Lung Fibroblasts ◽  
Matrix Remodeling ◽  
Rat Lung ◽  
Fetal Rat ◽  
Fetal Rat Lung

Abstract Preterm birth is characterized by severe lung immaturity that is frequently treated antenatally or postnatally with the synthetic steroid betamethasone. The underlying cellular targets and pathways stimulated by betamethasone in the fetal lung are poorly defined. In this study, betamethasone was compared with corticosterone in steroid-treated primary cultures of fetal rat lung fibroblasts stimulated for 6 hours and analyzed by whole-cell transcriptome sequencing and glucocorticoid (GC) receptor (GR) chromatin immunoprecipitation sequencing (ChIP-Seq) analysis. Strikingly, betamethasone stimulated a much stronger transcriptional response compared with corticosterone for both induced and repressed genes. A total of 483 genes were significantly stimulated by betamethasone or corticosterone, with 476 stimulated by both steroids, indicating a strong overlap in regulation. Changes in mRNA levels were confirmed by quantitative PCR for eight induced and repressed target genes. Pathway analysis identified cell proliferation and cytoskeletal/cell matrix remodeling pathways as key processes regulated by both steroids. One target, transglutaminase 2 (Tgm2), was localized to fetal lung mesenchymal cells. Tgm2 mRNA and protein levels were strongly increased in fibroblasts by both steroids. Whole-genome GR ChIP-Seq analysis with betamethasone identified GC response element–binding sites close to the previously characterized GR target genes Per1, Dusp1, Fkbp5, and Sgk1 and near the genes identified by transcriptome sequencing encoding Crispld2, Tgm2, Hif3α, and Kdr, defining direct genomic induction of expression in fetal lung fibroblasts via the GR. These results demonstrate that betamethasone stimulates specific genes and cellular pathways controlling cell proliferation and extracellular matrix remodeling in lung mesenchymal fibroblasts, providing a basis for betamethasone’s treatment efficacy in preterm birth.

scholarly journals Mechanical strain-induced posttranscriptional regulation of fibronectin production in fetal lung cells

1999 ◽  
Vol 277 (1) ◽  
pp. L142-L149 ◽  
Author(s):  
Eric Mourgeon ◽  
Jing Xu ◽  
A. Keith Tanswell ◽  
Mingyao Liu ◽  
Martin Post
Keyword(s):  
Cell Proliferation ◽  
Mechanical Strain ◽  
Fetal Lung ◽  
Northern Analysis ◽  
Normal Lung ◽  
Lung Cells ◽  
Mrna Levels ◽  
Rat Lung ◽  
Fetal Rat ◽  
Fetal Rat Lung

We have shown that intermittent mechanical strain, simulating fetal breathing movements, stimulated fetal rat lung cell proliferation. Because normal lung growth requires proper coordination between cell proliferation and extracellular matrix remodeling, we investigated the effect of strain on fibronectin metabolism. Organotypic cultures of fetal rat lung cells, subjected to intermittent strain, showed increased fibronectin content in the culture media. Fibronectin-degrading activity in media from strained cells was similar to that of static cultures. Northern analysis revealed that strain inhibited fibronectin mRNA accumulation seen during static culture. Synthesis of fibronectin, determined by metabolic labeling, was increased by strain despite lower mRNA levels or presence of actinomycin D. This increase was not mediated via a rapamycin-sensitive mechanism. Strain stimulated prelabeled fibronectin secretion even in the presence of cycloheximide. These results suggest that strain differentially regulates fibronectin production of fetal lung cells at the transcriptional and posttranscriptional levels. Mechanical strain increases soluble fibronectin content by stimulating its synthesis and secretion without increasing fibronectin message levels.

Increased lung expansion alters the proportions of type I and type II alveolar epithelial cells in fetal sheep

2000 ◽  
Vol 278 (6) ◽  
pp. L1180-L1185 ◽  
Author(s):  
S. Flecknoe ◽  
R. Harding ◽  
G. Maritz ◽  
S. B. Hooper
Keyword(s):  
Epithelial Cells ◽  
Fetal Lung ◽  
Alveolar Epithelial Cells ◽  
Type I ◽  
Intermediate Cell ◽  
Type Ii Cells ◽  
Type Ii ◽  
Cell Type ◽  
Alveolar Epithelial ◽  
Lung Expansion

Type I and type II alveolar epithelial cells (AECs) are derived from the same progenitor cell, but little is known about the factors that regulate their differentiation into separate phenotypes. An alteration in lung expansion alters the proportion type II AECs in the fetal lung, indicating that this may be a regulatory factor. Our aim was to quantify the changes in the proportion of type I and type II AECs caused by increased fetal lung expansion and to provide evidence for transdifferentiation of type II into type I cells. Lung tissue samples were collected from ovine fetuses exposed to increased lung expansion induced by 2, 4, or 10 days of tracheal obstruction (TO). The identities and proportions of AEC types were determined with electron microscopy. The proportion of type II cells was reduced from 28.5 ± 2.2% in control fetuses to 9.4 ± 2.3% at 2 days of TO and then to 1.9 ± 0.8% at 10 days. The proportion of type I AECs was not altered at 2 days of TO (63.1 ± 2.3%) compared with that of control cells (64.8 ± 0.5%) but was markedly elevated (to 89.4 ± 0.9%) at 10 days of TO. The proportion of an intermediate AEC type, which displayed characteristics of both type I and type II cells, increased from 5.7 ± 1.3% in control fetuses to 23.8 ± 5.1% by 2 days of TO and was similar to control values at 10 days of TO (7.7 ± 0.9%). Our data show that increases in fetal lung expansion cause time-dependent changes in the proportion of AEC types, including a transient increase in an intermediate cell type. These data provide the first evidence to support the hypothesis that increases in fetal lung expansion induce differentiation of type II into type I AECs via an intermediate cell type.

Disparate cytokine regulation of ICAM-1 in rat alveolar epithelial cells and pulmonary endothelial cells in vitro

1995 ◽  
Vol 269 (1) ◽  
pp. L127-L135 ◽  
Author(s):  
W. W. Barton ◽  
S. Wilcoxen ◽  
P. J. Christensen ◽  
R. Paine
Keyword(s):  
Endothelial Cells ◽  
Epithelial Cells ◽  
Inflammatory Cytokines ◽  
Alveolar Epithelial Cells ◽  
Normal Lung ◽  
Type I ◽  
Type Ii Cells ◽  
Type Ii ◽  
Alveolar Epithelial

Intercellular adhesion molecule-1 (ICAM-1) is expressed at high levels on type I alveolar epithelial cells in the normal lung and is induced in vitro as type II cells spread in primary culture. In contrast, in most nonhematopoetic cells ICAM-1 expression is induced in response to inflammatory cytokines. We have formed the hypothesis that the signals that control ICAM-1 expression in alveolar epithelial cells are fundamentally different from those controlling expression in most other cells. To test this hypothesis, we have investigated the influence of inflammatory cytokines on ICAM-1 expression in isolated type II cells that have spread in culture and compared this response to that of rat pulmonary artery endothelial cells (RPAEC). ICAM-1 protein, determined both by a cell-based enzyme-linked immunosorbent assay and by Western blot analysis, and mRNA were minimally expressed in unstimulated RPAEC but were significantly induced in a time- and dose-dependent manner by treatment with tumor necrosis factor-alpha, interleukin-1 beta, or interferon-gamma. In contrast, these cytokines did not influence the constitutive high level ICAM-1 protein expression in alveolar epithelial cells and only minimally affected steady-state mRNA levels. ICAM-1 mRNA half-life, measured in the presence of actinomycin D, was relatively long at 7 h in alveolar epithelial cells and 4 h in RPAEC. The striking lack of response of ICAM-1 expression by alveolar epithelial cells to inflammatory cytokines is in contrast to virtually all other epithelial cells studied to date and supports the hypothesis that ICAM-1 expression by these cells is a function of cellular differentiation.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Human lung myofibroblast-derived inducers of alveolar epithelial apoptosis identified as angiotensin peptides

1999 ◽  
Vol 277 (6) ◽  
pp. L1158-L1164 ◽  
Author(s):  
Rongi Wang ◽  
Carlos Ramos ◽  
Iravati Joshi ◽  
Alex Zagariya ◽  
Annie Pardo ◽  
...  
Keyword(s):  
Conditioned Medium ◽  
Human Lung ◽  
Smooth Muscle Actin ◽  
Alveolar Epithelial Cells ◽  
Lung Fibroblasts ◽  
Normal Fibroblast ◽  
Ang Ii ◽  
Alveolar Epithelial ◽  
Apoptotic Activity

Earlier work from this laboratory found that fibroblasts isolated from fibrotic human lung [human interstitial pulmonary fibrosis (HIPF)] secrete a soluble inducer(s) of apoptosis in alveolar epithelial cells (AECs) in vitro [B. D. Uhal, I. Joshi, A. True, S. Mundle, A. Raza, A. Pardo, and M. Selman. Am. J. Physiol. 269 ( Lung Cell. Mol. Physiol. 13): L819–L828, 1995]. The cultured human fibroblast strains most active in producing the apoptotic activity contained high numbers of stellate cells expressing α-smooth muscle actin, a myofibroblast marker. The apoptotic activity eluted from gel-filtration columns only in fractions corresponding to proteins. Western blotting of the protein fraction identified immunoreactive angiotensinogen (ANGEN), and two-step RT-PCR revealed expression of ANGEN by HIPF fibroblasts but not by normal human lung fibroblasts. Specific ELISA detected angiotensin II (ANG II) at concentrations sixfold higher in HIPF-conditioned medium than in normal fibroblast-conditioned medium. Pretreatment of the concentrated medium with purified renin plus purified angiotensin-converting enzyme (ACE) further increased the ELISA-detectable ANG II eightfold. Apoptosis of AECs in response to HIPF-conditioned medium was completely abrogated by the ANG II receptor antagonist saralasin (50 μg/ml) or anti-ANG II antibodies. These results identify the protein inducers of AEC apoptosis produced by HIPF fibroblasts as ANGEN and its derivative ANG II. They also suggest a mechanism for AEC death adjacent to HIPF myofibroblasts [B. D. Uhal,, I. Joshi, C. Ramos, A. Pardo, and M. Selman. Am. J. Physiol. 275 ( Lung Cell. Mol. Physiol. 19): L1192–L1199, 1998].

scholarly journals P-Rex1 Cooperates With TGFβR2 to Drive Lung Fibroblast Migration in Pulmonary Fibrosis

2021 ◽  
Vol 12 ◽  
Author(s):  
Qing Liang ◽  
Yanhua Chang ◽  
Jing Liu ◽  
Yan Yu ◽  
Wancheng Qiu ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Lung Fibroblast ◽  
Alveolar Epithelial Cells ◽  
Lung Fibroblasts ◽  
Mouse Lung ◽  
Nucleotide Exchange ◽  
Alveolar Epithelial ◽  
Fibroblast Migration ◽  
Overexpression System ◽  
Tgf Β1

Pulmonary fibrosis is a kind of interstitial lung disease with progressive pulmonary scar formation, leading to irreversible loss of lung functions. The TGF-β1/Smad signaling pathway plays a key role in fibrogenic processes. It is associated with the increased synthesis of extracellular matrix, enhanced proliferation of fibroblasts, and transformation of alveolar epithelial cells into interstitial cells. We investigated P-Rex1, a PIP3-Gβγ–dependent guanine nucleotide exchange factor (GEF) for Rac, for its potential role in TGF-β1–induced pulmonary fibrosis. A high expression level of P-Rex1 was identified in the lung tissue of patients with pulmonary fibrosis than that from healthy donors. Using the P-Rex1 knockdown and overexpression system, we established a novel player of P-Rex1 in mouse lung fibroblast migration. P-Rex1 contributed to fibrogenic processes in lung fibroblasts by targeting the TGF-β type Ⅱ receptor (TGFβR2). The RNA-seq analysis for expression profiling confirmed the modulation of P-Rex1 in cell migration and the involvement of P-Rex1 in TGF-β1 signaling. These results identified P-Rex1 as a signaling molecule involved in TGF-β1–induced pulmonary fibrosis, suggesting that P-Rex1 may be a potential target for pulmonary fibrosis treatment.

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