Hoxa-5 in mouse developing lung: cell-specific expression and retinoic acid regulation

2000 ◽  
Vol 279 (5) ◽  
pp. L863-L871 ◽  
Author(s):  
Chiwan Kim ◽  
Heber C. Nielsen
Keyword(s):  
Retinoic Acid ◽  
Molecular Mechanisms ◽  
Homeobox Gene ◽  
Fetal Lung ◽  
Lung Epithelial Cells ◽  
Lung Fibroblasts ◽  
Mouse Lung ◽  
Dependent Manner ◽  
Specific Expression ◽  
Protein Expressions

Hoxa-5 is a homeobox gene that is highly expressed in the developing mouse lung. However, little is known about the molecular mechanisms controlling expression. We characterized the ontogeny of Hoxa-5 gene and protein expressions during lung development and then studied the cell-specific effects of retinoic acid (RA) on Hoxa-5 mRNA in fetal lung fibroblasts and MLE-12 mouse lung epithelial cells. Strong but constant Hoxa-5 gene and protein expressions were detected from mouse lung on embryonic day 13.5 to postnatal day 2. At baseline, the gene was strongly expressed in the fibroblasts of day 17.5 fetal mouse lungs. A very weak but reproducible expression was present in the MLE-12 cells. RA stimulated gene expression in both cell types in a time- and dose-dependent manner. Peak expression occurred much later in the MLE-12 cells compared with that in fibroblasts. Cycloheximide and actinomycin D treatment studies suggested that the differences in RA effect on each cell type may involve the presence of a repressor that can be overcome by RA.

Hypoxia results in an HIF-1-dependent induction of brain-specific aldolase C in lung epithelial cells

2006 ◽  
Vol 291 (5) ◽  
pp. L950-L956 ◽  
Author(s):  
Jyh-Chang Jean ◽  
Celeste B. Rich ◽  
Martin Joyce-Brady
Keyword(s):  
Hypoxia Inducible Factor ◽  
Fetal Lung ◽  
Lung Epithelial Cells ◽  
Mouse Lung ◽  
Developmentally Regulated ◽  
Specific Expression ◽  
Aldolase C ◽  
Expression Studies ◽  
Lung Epithelial ◽  
Responsive Element

Aldolase C (EC 4.1.2.13 ) is a brain-specific aldolase isoform and a putative target of the transcription factor hypoxia-inducible factor (HIF)-1. We identified aldolase C as a candidate hypoxia-regulated gene in mouse lung epithelial (MLE) cells using differential display. We show that the message accumulates in a robust fashion when MLE cells are exposed to 1% oxygen and is inversely related to oxygen content. Induction in hypoxia is dependent on protein synthesis. We localized a hypoxia-responsive element (HRE) in the aldolase C promoter using a series of deletion and heterologous expression studies. The HRE overlaps with a region of the proximal aldolase C promoter that is also related to its brain-specific expression. The HRE contains an Arnt (HIF-1β) and an HIF-1α site. We show that induction in hypoxia is dependent on the HIF-1 site and that HIF-1α protein is present, by gel-shift assay, within nuclear complexes of MLE cells in hypoxia. Aldolase C mRNA expression is developmentally regulated in the fetal lung, rapidly downregulated in the newborn lung at birth, and inducible in the adult lung when exposed to hypoxia. This pattern of regulation is not seen in the brain. This preservation of this HRE in the promoters of four other species suggests that aldolase C may function as a stress-response gene.

Retinoic acid stimulates immature lung fibroblast growth via a PDGF-mediated autocrine mechanism

2000 ◽  
Vol 279 (1) ◽  
pp. L81-L90 ◽  
Author(s):  
Abraham Liebeskind ◽  
Suseela Srinivasan ◽  
David Kaetzel ◽  
Margaret Bruce
Keyword(s):  
Retinoic Acid ◽  
Growth Factor ◽  
Thymidine Incorporation ◽  
Neutralizing Antibody ◽  
Receptor Expression ◽  
Lung Fibroblasts ◽  
Transcriptional Level ◽  
Dependent Manner ◽  
Adult Rat ◽  
Autocrine Mechanism

all trans-retinoic acid (RA) enhances alveolarization in neonates and reinitiates alveolarization in emphysematous adult rat lungs, suggesting that RA may stimulate cell proliferation by upregulating growth factor ligand and/or receptor expression either indirectly or directly by acting on RA-responsive genes encoding growth factors. We report that RA and 1,25-dihydroxyvitamin D3(Vit D), alone and in combination, significantly increase [3H]thymidine incorporation in cultured fetal and postnatal rat lung fibroblasts ( P < 0.05). The greatest increase (11-fold) was seen in 4-day cells treated with the two agents in combination ( P < 0.0001). [3H]thymidine incorporation was age dependent. The greatest response to RA occurred in 4-day fibroblasts ( P < 0.01), whereas the response to Vit D was greatest in embryonic day 20 fibroblasts ( P < 0.001). Neutralizing antibody to platelet-derived growth factor (PDGF)-AB decreased [3H]thymidine incorporation in response to RA alone or in combination with Vit D, indicating a role for PDGF. Expression of mRNAs for PDGF-A and PDGF receptor (PDGFR)-α and -β was upregulated at the transcriptional level in an age- and treatment-dependent manner. Thus exogenous RA may influence alveolarization by stimulating fibroblast proliferation through a PDGF-mediated autocrine mechanism, which is enhanced when RA and Vit D are administered in combination.

scholarly journals Retinoic acid-receptor activation of SP-B gene transcription in respiratory epithelial cells

1998 ◽  
Vol 275 (2) ◽  
pp. L239-L246 ◽  
Author(s):  
Cong Yan ◽  
Manely Ghaffari ◽  
Jeffrey A. Whitsett ◽  
Xin Zeng ◽  
Zvjezdana Sever ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Epithelial Cells ◽  
Gene Transcription ◽  
Receptor Activation ◽  
Pulmonary Adenocarcinoma ◽  
Lung Epithelial Cells ◽  
Mouse Lung ◽  
Target Cells ◽  
Adenocarcinoma Cells

Retinoids are known to play important roles in organ development of the lung. Retinoids exert their activity by modulating the expression of numerous genes, generally influencing gene transcription, in target cells. In the present work, the mechanism by which retinoic acid (RA) regulates surfactant protein (SP) B expression was assessed in vitro. RA (9- cis-RA) enhanced SP-B mRNA in pulmonary adenocarcinoma cells (H441 cells) and increased transcriptional activity of the SP-B promoter in both H441 and mouse lung epithelial cells (MLE-15). Cotransfection of H441 cells with retinoid nuclear receptor (RAR)-α, -β, and -γ and retinoid X receptor (RXR)-γ further increased the response of the SP-B promoter to RA. Treatment of H441 cells with RA increased immunostaining for the SP-B proprotein and increased the number of cells in which the SP-B proprotein was detected. An RA responsive element mediating RA stimulation of the human SP-B promoter was identified. RAR-α and -γ and RXR-α but not RAR-β or RXR-β and -γ were detected by immunohistochemical analysis of H441 cells. RA, by activating RAR activity, stimulated the transcription and synthesis of SP-B in pulmonary adenocarcinoma cells.

Regulation of c-ret expression by retinoic acid in rat metanephros: implication in nephron mass control

1998 ◽  
Vol 275 (6) ◽  
pp. F938-F945 ◽  
Author(s):  
Evelyne Moreau ◽  
José Vilar ◽  
Martine Lelièvre-Pégorier ◽  
Claudie Merlet-Bénichou ◽  
Thierry Gilbert
Keyword(s):  
Retinoic Acid ◽  
Molecular Mechanisms ◽  
Kidney Development ◽  
Mrna Level ◽  
Cell Surface Receptor ◽  
Dependent Manner ◽  
All Trans Retinoic Acid ◽  
Surface Receptor ◽  
Dose Dependent

Vitamin A and its derivatives have been shown to promote kidney development in vitro in a dose-dependent fashion. To address the molecular mechanisms by which all- trans-retinoic acid (RA) may regulate the nephron mass, rat kidneys were removed on embryonic day 14( E14) and grown in organ culture under standard or RA-stimulated conditions. By using RT-PCR, we studied the expression of the glial cell line-derived neurotrophic factor (GDNF), its cell surface receptor-α (GDNFR-α), and the receptor tyrosine kinase c-ret, known to play a major role in renal organogenesis. Expression of GDNF and GDNFR-α transcripts was high at the time of explantation and remained unaffected in culture with or without RA. In contrast, c-ret mRNA level, which was low in E14 metanephros and dropped rapidly in vitro, was increased by RA in a dose-dependent manner. The same is true at the protein level. Exogenous GDNF barely promotes additional nephron formation in vitro. Thus the present data establish c-ret as a key target of retinoids during kidney organogenesis.

Bradykinin augments fibroblast-mediated contraction of released collagen gels

2001 ◽  
Vol 281 (1) ◽  
pp. L164-L171 ◽  
Author(s):  
Tadashi Mio ◽  
Xiangde Liu ◽  
Myron L. Toews ◽  
Yuichi Adachi ◽  
Debra J. Romberger ◽  
...  
Keyword(s):  
Receptor Antagonist ◽  
Type I Collagen ◽  
Inflammatory Diseases ◽  
Fetal Lung ◽  
Lung Fibroblasts ◽  
Type I ◽  
Dependent Manner ◽  
Collagen Gels ◽  
Protein Kinase C Inhibitors

Bradykinin is a multifunctional mediator of inflammation believed to have a role in asthma, a disorder associated with remodeling of extracellular connective tissue. Using contraction of collagen gels as an in vitro model of wound contraction, we assessed the effects of bradykinin tissue on remodeling. Human fetal lung fibroblasts were embedded in type I collagen gels and cultured for 5 days. After release, the floating gels were cultured in the presence of bradykinin. Bradykinin significantly stimulated contraction in a concentration- and time-dependent manner. Coincubation with phosphoramidon augmented the effect of 10−9 and 10−8 M bradykinin. A B2 receptor antagonist attenuated the effect of bradykinin, whereas a B1 receptor antagonist had no effect, suggesting that the effect is mediated by the B2 receptor. An inhibitor of intracellular Ca2+mobilization abolished the response; addition of EGTA to the culture medium attenuated the contraction of control gels but did not modulate the response to bradykinin. In contrast, the phospholipase C inhibitor U-73122 and the protein kinase C inhibitors staurosporine and GF-109203X attenuated the responses. These data suggest that by augmenting the contractility of fibroblasts, bradykinin may have an important role in remodeling of extracellular matrix that may result in tissue dysfunction in chronic inflammatory diseases, such as asthma.

scholarly journals Disrupted TSH Receptor Expression in Female Mouse Lung Fibroblasts Alters Subcellular IGF-1 Receptor Distribution

Endocrinology ◽  
2015 ◽  
Vol 156 (12) ◽  
pp. 4731-4740 ◽  
Author(s):  
Stephen J. Atkins ◽  
Stephen I. Lentz ◽  
Roshini Fernando ◽  
Terry J. Smith
Keyword(s):  
Molecular Mechanisms ◽  
Flow Cytometric Analysis ◽  
Receptor Expression ◽  
Lung Fibroblasts ◽  
Mouse Lung ◽  
Suitable Model ◽  
Mrna Levels ◽  
Cytoplasmic Staining ◽  
Orbital Fibroblasts ◽  
Nuclear Signal

A relationship between the actions of TSH and IGF-1 was first recognized several decades ago. The close physical and functional associations between their respective receptors (TSHR and IGF-1R) has been described more recently in thyroid epithelium and human orbital fibroblasts as has the noncanonical behavior of IGF-1R. Here we report studies conducted in lung fibroblasts from female wild-type C57/B6 (TSHR+/+) mice and their littermates in which TSHR has been knocked out (TSHR−/−). Flow cytometric analysis revealed that cell surface IGF-1R levels are substantially lower in TSHR−/− fibroblasts compared with TSHR+/+ fibroblasts. Confocal immunofluorescence microscopy revealed similar divergence with regard to both cytoplasmic and nuclear IGF-1R. Western blot analysis demonstrated both intact IGF-1R and receptor fragments in both cellular compartments. In contrast, IGF-1R mRNA levels were similar in fibroblasts from mice without and with intact TSHR expression. IGF-1 treatment of TSHR+/+ fibroblasts resulted in reduced nuclear and cytoplasmic staining for IGF-1Rα, whereas it enhanced the nuclear signal in TSHR−/− cells. In contrast, IGF-1 enhanced cytoplasmic IGF-1Rβ in TSHR−/− fibroblasts while increasing the nuclear signal in TSHR+/+ cells. These findings indicate the intimate relationship between TSHR and IGF-1R found earlier in human orbital fibroblasts also exists in mouse lung fibroblasts. Furthermore, the presence of TSHR in these fibroblasts influenced not only the levels of IGF-1R protein but also its subcellular distribution and response to IGF-1. They suggest that the mouse might serve as a suitable model for delineating the molecular mechanisms overarching these two receptors.

scholarly journals All-transretinoic acid modulates radiation-induced proliferation of lung fibroblasts via IL-6/IL-6R system

2006 ◽  
Vol 290 (3) ◽  
pp. L597-L606 ◽  
Author(s):  
Chiharu Tabata ◽  
Hajime Kubo ◽  
Rie Tabata ◽  
Manabu Wada ◽  
Keiichiro Sakuma ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Retinoic Acid ◽  
Intraperitoneal Administration ◽  
Lung Fibroblasts ◽  
High Dose ◽  
Dependent Manner ◽  
Antitumor Effects ◽  
Receptor System ◽  
Lung Cancers ◽  
All Trans Retinoic Acid

Although high-dose thoracic radiotherapy is an effective strategy for some malignancies including lung cancers and malignant lymphomas, it often causes complications of radiation fibrosis. To study the mechanism initiating tissue fibrosis, we investigated irradiation-induced cytokine production from human lung fibroblastic cells and found that IL-6 production was stimulated by irradiation. IL-6 is an autocrine growth factor for human myeloma cells, and retinoic acid is reported to inhibit their growth. Thus we evaluated the effect of all- trans retinoic acid (ATRA) on cell proliferation of lung fibroblasts along with the cytokine/receptor system. Irradiation-dependent stimulation of IL-6 production was correlated with increased NF-κB activity, and ATRA reduced this effect. Irradiation also increased the levels of mRNA for IL-6R and gp130, which were blocked by coexisting ATRA. Furthermore, IL-6 stimulated cell proliferation in dose-dependent manner but was overcome by pharmacological concentration of ATRA. These effects of ATRA were inhibited by rottlerin, which suggests ATRA abolished irradiation-induced stimulation through a PKCδ-dependent pathway. Finally, we demonstrated that IL-6 transcripts in the lung were upregulated at 2 mo after irradiation, and the effect was inhibited by the intraperitoneal administration of ATRA. ATRA is expected to have an advantage for radiotherapy in its antitumor effects, as reported previously, and to prevent radiotherapy-induced pulmonary injury.

Paraquat increases connective tissue growth factor expression and impairs lung fibroblast proliferation and viscoelasticity

2014 ◽  
Vol 33 (12) ◽  
pp. 1232-1240 ◽  
Author(s):  
N Zhang ◽  
Y-P Xie ◽  
L Pang ◽  
X-X Zang ◽  
J Wang ◽  
...  
Keyword(s):  
Growth Factor ◽  
Connective Tissue ◽  
Connective Tissue Growth Factor ◽  
Molecular Mechanisms ◽  
Tissue Growth ◽  
Lung Fibroblasts ◽  
Proliferation Index ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dependent Manner ◽  
Paraquat Poisoning

This in vitro study was designed to investigate the molecular mechanisms of paraquat-induced damage using cultured human fetal lung fibroblasts (MRC-5 cells), in order to promote the development of improved therapies for paraquat poisoning. Paraquat’s effects on proliferation were examined by flow cytometry, on viscoelasticity by the micropipette aspiration technique, and on connective tissue growth factor (CTGF) expression by real-time polymerase chain reaction and enzyme-linked immunosorbent assay. Paraquat was found to significantly reduce the proliferation index of MRC-5 cells in a concentration-dependent manner ( p < 0.05) and to significantly impair the viscoelastic properties in a time-independent manner ( p < 0.05). Exposure to paraquat led to a significant and time-dependent increase in CTGF expression ( p < 0.05) and induced changes in the morphology and biomechanical characteristics of the MRC-5 cells. These findings not only provide novel insights into the mechanisms of paraquat-induced lung fibrosis but may represent useful targets of improved molecular-based therapies for paraquat poisoning.

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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