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Down-regulation of the inhibitor of growth family member 4 (ING4) in different forms of pulmonary fibrosis.

Tzouvelekis A, Aidinis V, Harokopos V, Karameris A, Zacharis G, Mikroulis D, Konstantinou F, Steiropoulos P, Sotiriou I, Froudarakis M, Pneumatikos I, Tringidou R, Bouros D - Respir. Res. (2009)

Bottom Line: ING4 was also found down-regulated in IPF patients compared to COP and control subjects.Our data suggest a potential role for ING4 in lung fibrogenesis.ING4 down-regulation may facilitate aberrant vascular remodelling and fibroblast proliferation and migration leading to progressive disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pneumonology, University Hospital of Alexandroupolis, Medical school, Democritus University of Thrace, Greece. atzouvelekis@yahoo.gr

ABSTRACT

Background: Recent evidence has underscored the role of hypoxia and angiogenesis in the pathogenesis of idiopathic fibrotic lung disease. Inhibitor of growth family member 4 (ING4) has recently attracted much attention as a tumor suppressor gene, due to its ability to inhibit cancer cell proliferation, migration and angiogenesis. The aim of our study was to investigate the role of ING4 in the pathogenesis of pulmonary fibrosis both in the bleomycin (BLM)-model and in two different types of human pulmonary fibrosis, including idiopathic pulmonary fibrosis (IPF) and cryptogenic organizing pneumonia (COP).

Methods: Experimental model of pulmonary fibrosis was induced by a single tail vein injection of bleomycin in 6- to 8-wk-old C57BL/6mice. Tissue microarrays coupled with qRT-PCR and immunohistochemistry were applied in whole lung samples and paraffin-embedded tissue sections of 30 patients with IPF, 20 with COP and 20 control subjects.

Results: A gradual decline of ING4 expression in both mRNA and protein levels was reported in the BLM-model. ING4 was also found down-regulated in IPF patients compared to COP and control subjects. Immunolocalization analyses revealed increased expression in areas of normal epithelium and in alveolar epithelium surrounding Masson bodies, in COP lung, whereas showed no expression within areas of active fibrosis within IPF and COP lung. In addition, ING4 expression levels were negatively correlated with pulmonary function parameters in IPF patients.

Conclusion: Our data suggest a potential role for ING4 in lung fibrogenesis. ING4 down-regulation may facilitate aberrant vascular remodelling and fibroblast proliferation and migration leading to progressive disease.

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Related in: MedlinePlus

Decreased ING4 expression in bleomycin (BLM)- induced pulmonary fibrosis (PF) following disease progression. (A) Representative immunohistochemistry with an anti-ING4 antibody on lung paraffin sections from BLM-treated mice (7, 15, and 23, days post-administration). ING4 was mainly expressed in alveolar epithelium (days 7 and 15) whereas showed weak staining within areas of dense fibrosis and collagen deposition at late disease stages (day 21). (B) Computerized image analysis of immunostained sections. *p < 0.05, **p < 0.005, ***p < 0.001. (One way ANOVA and unpaired t-test with Bonferroni correction, F = 71,126).
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Figure 2: Decreased ING4 expression in bleomycin (BLM)- induced pulmonary fibrosis (PF) following disease progression. (A) Representative immunohistochemistry with an anti-ING4 antibody on lung paraffin sections from BLM-treated mice (7, 15, and 23, days post-administration). ING4 was mainly expressed in alveolar epithelium (days 7 and 15) whereas showed weak staining within areas of dense fibrosis and collagen deposition at late disease stages (day 21). (B) Computerized image analysis of immunostained sections. *p < 0.05, **p < 0.005, ***p < 0.001. (One way ANOVA and unpaired t-test with Bonferroni correction, F = 71,126).

Mentions: As angiogenesis[15,18,19] and apoptosis[20,21] represent two of the major pathogenetic hallmarks of pulmonary fibrosis and since HIF-1a, the major transcription factor of hypoxia-related genes involved in angiogenesis and apoptosis, has been recently implicated in the pathogenesis of fibrotic lung disease we sought to investigate the expression of its inhibitor, ING4, both in mRNA and protein level using qRT-PCR and immunohistochemistry analysis, respectively, in a well characterized model of pulmonary fibrosis. Surprisingly, following disease progression, ING4 expression was found downregulated, both in mRNA and protein level, as shown in Figures 1 and 2. In particular, qRT-PCR analysis demonstrated that Ing4 gene expression was downregulated upon administration of BLM and the development of pulmonary inflammation and fibrosis (Figure 1). Experimental findings were further extended by immunohistochemistry analysis for ING4 expression on lung paraffin sections from BLM treated mice (7, 15 and 23 days post administration) which confirmed decreased expression during disease progression. ING4 was extensively expressed in normal epithelium in control lung samples, as well as in early stages of disease (day 7) where inflammation is prominent and fibrosis is almost absent (Figure 2).


Down-regulation of the inhibitor of growth family member 4 (ING4) in different forms of pulmonary fibrosis.

Tzouvelekis A, Aidinis V, Harokopos V, Karameris A, Zacharis G, Mikroulis D, Konstantinou F, Steiropoulos P, Sotiriou I, Froudarakis M, Pneumatikos I, Tringidou R, Bouros D - Respir. Res. (2009)

Decreased ING4 expression in bleomycin (BLM)- induced pulmonary fibrosis (PF) following disease progression. (A) Representative immunohistochemistry with an anti-ING4 antibody on lung paraffin sections from BLM-treated mice (7, 15, and 23, days post-administration). ING4 was mainly expressed in alveolar epithelium (days 7 and 15) whereas showed weak staining within areas of dense fibrosis and collagen deposition at late disease stages (day 21). (B) Computerized image analysis of immunostained sections. *p < 0.05, **p < 0.005, ***p < 0.001. (One way ANOVA and unpaired t-test with Bonferroni correction, F = 71,126).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2662808&req=5

Figure 2: Decreased ING4 expression in bleomycin (BLM)- induced pulmonary fibrosis (PF) following disease progression. (A) Representative immunohistochemistry with an anti-ING4 antibody on lung paraffin sections from BLM-treated mice (7, 15, and 23, days post-administration). ING4 was mainly expressed in alveolar epithelium (days 7 and 15) whereas showed weak staining within areas of dense fibrosis and collagen deposition at late disease stages (day 21). (B) Computerized image analysis of immunostained sections. *p < 0.05, **p < 0.005, ***p < 0.001. (One way ANOVA and unpaired t-test with Bonferroni correction, F = 71,126).
Mentions: As angiogenesis[15,18,19] and apoptosis[20,21] represent two of the major pathogenetic hallmarks of pulmonary fibrosis and since HIF-1a, the major transcription factor of hypoxia-related genes involved in angiogenesis and apoptosis, has been recently implicated in the pathogenesis of fibrotic lung disease we sought to investigate the expression of its inhibitor, ING4, both in mRNA and protein level using qRT-PCR and immunohistochemistry analysis, respectively, in a well characterized model of pulmonary fibrosis. Surprisingly, following disease progression, ING4 expression was found downregulated, both in mRNA and protein level, as shown in Figures 1 and 2. In particular, qRT-PCR analysis demonstrated that Ing4 gene expression was downregulated upon administration of BLM and the development of pulmonary inflammation and fibrosis (Figure 1). Experimental findings were further extended by immunohistochemistry analysis for ING4 expression on lung paraffin sections from BLM treated mice (7, 15 and 23 days post administration) which confirmed decreased expression during disease progression. ING4 was extensively expressed in normal epithelium in control lung samples, as well as in early stages of disease (day 7) where inflammation is prominent and fibrosis is almost absent (Figure 2).

Bottom Line: ING4 was also found down-regulated in IPF patients compared to COP and control subjects.Our data suggest a potential role for ING4 in lung fibrogenesis.ING4 down-regulation may facilitate aberrant vascular remodelling and fibroblast proliferation and migration leading to progressive disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pneumonology, University Hospital of Alexandroupolis, Medical school, Democritus University of Thrace, Greece. atzouvelekis@yahoo.gr

ABSTRACT

Background: Recent evidence has underscored the role of hypoxia and angiogenesis in the pathogenesis of idiopathic fibrotic lung disease. Inhibitor of growth family member 4 (ING4) has recently attracted much attention as a tumor suppressor gene, due to its ability to inhibit cancer cell proliferation, migration and angiogenesis. The aim of our study was to investigate the role of ING4 in the pathogenesis of pulmonary fibrosis both in the bleomycin (BLM)-model and in two different types of human pulmonary fibrosis, including idiopathic pulmonary fibrosis (IPF) and cryptogenic organizing pneumonia (COP).

Methods: Experimental model of pulmonary fibrosis was induced by a single tail vein injection of bleomycin in 6- to 8-wk-old C57BL/6mice. Tissue microarrays coupled with qRT-PCR and immunohistochemistry were applied in whole lung samples and paraffin-embedded tissue sections of 30 patients with IPF, 20 with COP and 20 control subjects.

Results: A gradual decline of ING4 expression in both mRNA and protein levels was reported in the BLM-model. ING4 was also found down-regulated in IPF patients compared to COP and control subjects. Immunolocalization analyses revealed increased expression in areas of normal epithelium and in alveolar epithelium surrounding Masson bodies, in COP lung, whereas showed no expression within areas of active fibrosis within IPF and COP lung. In addition, ING4 expression levels were negatively correlated with pulmonary function parameters in IPF patients.

Conclusion: Our data suggest a potential role for ING4 in lung fibrogenesis. ING4 down-regulation may facilitate aberrant vascular remodelling and fibroblast proliferation and migration leading to progressive disease.

Show MeSH
Related in: MedlinePlus