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Estradiol promotes the development of a fibrotic phenotype and is increased in the serum of patients with systemic sclerosis.

Aida-Yasuoka K, Peoples C, Yasuoka H, Hershberger P, Thiel K, Cauley JA, Medsger TA, Feghali-Bostwick CA - Arthritis Res. Ther. (2013)

Bottom Line: We confirmed the fibrotic effect of E2 in human skin using an ex vivo organ culture model.Propyl-pyrazole-triol, but not genistein, significantly increased FN expression.The effects of E2 were abrogated by ICI 182,780.

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ABSTRACT

Introduction: Systemic sclerosis (SSc) is more prevalent in women. Our goal is to determine the effects of 17β-estradiol (E2) on the development of fibrosis and to compare circulating levels of estrogens in SSc patients and healthy controls.

Methods: Using primary human dermal fibroblasts, we evaluated the effect of E2 on fibronectin (FN) expression with and without the estrogen receptor (ER) antagonist ICI 182,780, inhibitors of signaling, propyl-pyrazole-triol, an ERα specific ligand, and genistein, an ERβ selective ligand, to identify the signaling pathways mediating E2's effect. We confirmed the fibrotic effect of E2 in human skin using an ex vivo organ culture model. Lastly, we measured levels of E2 and estrone in serum samples from SSc patients with diffuse cutaneous involvement and healthy controls using mass spectrometry.

Results: E2 increased expression of FN in dermal fibroblasts. ICI 182,780, inositol-1,4,5-triphosphate inhibitor, and p38 mitogen-activated protein kinase inhibitor blocked the effects of E2 on FN. Propyl-pyrazole-triol, but not genistein, significantly increased FN expression. Ex vivo, E2 induced fibrosis of human skin. The effects of E2 were abrogated by ICI 182,780. Circulating levels of E2 and estrone were significantly increased in sera of patients with diffuse cutaneous SSc.

Conclusion: Our findings implicate estrogens in the fibrotic process and may explain the preponderance of SSc in women. ICI 182,780 or other ER signaling antagonists may be effective agents for the treatment of fibrosis.

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Estradiol promotes fibronectin production via specific signaling cascades. (A) Fibronectin (FN) protein levels in the cellular lysates of normal skin fibroblasts. Normal skin fibroblasts were stimulated with 17β-estradiol (E2) for 48 hours in the presence or absence of the following chemical inhibitors: MEK inhibitor (MEKi), phosphoinositol 3-kinase inhibitor (PI3Ki), and p38 kinase inhibitor (p38Ki). Cellular lysates were analyzed by western blot using anti-EDA-FN, estrogen receptor (ER)α, ERβ, and GAPDH antibodies. (B) Effect of E2 ligands on the expression and deposition of FN in the extracellular matrix (ECM) of normal skin fibroblasts. PPT, propyl-pyrazole-triol. Primary fibroblasts were cultured with vehicle (dimethylsulfoxide (DMSO), ethanol (EtOH)), E2, PPT, or genistein for 48 hours. ECM was analyzed by western blot using anti-EDA-FN and anti-vitronectin antibodies. Ethanol was used as vehicle for E2 and PPT. DMSO was used as a vehicle for genistein.
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Figure 2: Estradiol promotes fibronectin production via specific signaling cascades. (A) Fibronectin (FN) protein levels in the cellular lysates of normal skin fibroblasts. Normal skin fibroblasts were stimulated with 17β-estradiol (E2) for 48 hours in the presence or absence of the following chemical inhibitors: MEK inhibitor (MEKi), phosphoinositol 3-kinase inhibitor (PI3Ki), and p38 kinase inhibitor (p38Ki). Cellular lysates were analyzed by western blot using anti-EDA-FN, estrogen receptor (ER)α, ERβ, and GAPDH antibodies. (B) Effect of E2 ligands on the expression and deposition of FN in the extracellular matrix (ECM) of normal skin fibroblasts. PPT, propyl-pyrazole-triol. Primary fibroblasts were cultured with vehicle (dimethylsulfoxide (DMSO), ethanol (EtOH)), E2, PPT, or genistein for 48 hours. ECM was analyzed by western blot using anti-EDA-FN and anti-vitronectin antibodies. Ethanol was used as vehicle for E2 and PPT. DMSO was used as a vehicle for genistein.

Mentions: To investigate the mechanism mediating E2 induction of FN, we pretreated skin fibroblasts with vehicle, MEK inhibitor, PI3K inhibitor, or p38 MAPK inhibitor for 1 hour prior to the addition of E2. FN protein levels were assessed by western blot analysis 48 hours post treatment. PI3K inhibitor and p38 MAPK inhibitor attenuated the E2-mediated increase of FN (Figure 2A). MEK inhibitor had a more modest effect on E2 induction of FN. We also examined the effect of the chemical inhibitors on ERα and ERβ. ERα was increased by E2 and this increase was blocked by PI3K inhibitor, p38 MAPK inhibitor, and MEK inhibitor. There was no significant difference in the expression of ERβ under the same conditions (Figure 2A).


Estradiol promotes the development of a fibrotic phenotype and is increased in the serum of patients with systemic sclerosis.

Aida-Yasuoka K, Peoples C, Yasuoka H, Hershberger P, Thiel K, Cauley JA, Medsger TA, Feghali-Bostwick CA - Arthritis Res. Ther. (2013)

Estradiol promotes fibronectin production via specific signaling cascades. (A) Fibronectin (FN) protein levels in the cellular lysates of normal skin fibroblasts. Normal skin fibroblasts were stimulated with 17β-estradiol (E2) for 48 hours in the presence or absence of the following chemical inhibitors: MEK inhibitor (MEKi), phosphoinositol 3-kinase inhibitor (PI3Ki), and p38 kinase inhibitor (p38Ki). Cellular lysates were analyzed by western blot using anti-EDA-FN, estrogen receptor (ER)α, ERβ, and GAPDH antibodies. (B) Effect of E2 ligands on the expression and deposition of FN in the extracellular matrix (ECM) of normal skin fibroblasts. PPT, propyl-pyrazole-triol. Primary fibroblasts were cultured with vehicle (dimethylsulfoxide (DMSO), ethanol (EtOH)), E2, PPT, or genistein for 48 hours. ECM was analyzed by western blot using anti-EDA-FN and anti-vitronectin antibodies. Ethanol was used as vehicle for E2 and PPT. DMSO was used as a vehicle for genistein.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3672719&req=5

Figure 2: Estradiol promotes fibronectin production via specific signaling cascades. (A) Fibronectin (FN) protein levels in the cellular lysates of normal skin fibroblasts. Normal skin fibroblasts were stimulated with 17β-estradiol (E2) for 48 hours in the presence or absence of the following chemical inhibitors: MEK inhibitor (MEKi), phosphoinositol 3-kinase inhibitor (PI3Ki), and p38 kinase inhibitor (p38Ki). Cellular lysates were analyzed by western blot using anti-EDA-FN, estrogen receptor (ER)α, ERβ, and GAPDH antibodies. (B) Effect of E2 ligands on the expression and deposition of FN in the extracellular matrix (ECM) of normal skin fibroblasts. PPT, propyl-pyrazole-triol. Primary fibroblasts were cultured with vehicle (dimethylsulfoxide (DMSO), ethanol (EtOH)), E2, PPT, or genistein for 48 hours. ECM was analyzed by western blot using anti-EDA-FN and anti-vitronectin antibodies. Ethanol was used as vehicle for E2 and PPT. DMSO was used as a vehicle for genistein.
Mentions: To investigate the mechanism mediating E2 induction of FN, we pretreated skin fibroblasts with vehicle, MEK inhibitor, PI3K inhibitor, or p38 MAPK inhibitor for 1 hour prior to the addition of E2. FN protein levels were assessed by western blot analysis 48 hours post treatment. PI3K inhibitor and p38 MAPK inhibitor attenuated the E2-mediated increase of FN (Figure 2A). MEK inhibitor had a more modest effect on E2 induction of FN. We also examined the effect of the chemical inhibitors on ERα and ERβ. ERα was increased by E2 and this increase was blocked by PI3K inhibitor, p38 MAPK inhibitor, and MEK inhibitor. There was no significant difference in the expression of ERβ under the same conditions (Figure 2A).

Bottom Line: We confirmed the fibrotic effect of E2 in human skin using an ex vivo organ culture model.Propyl-pyrazole-triol, but not genistein, significantly increased FN expression.The effects of E2 were abrogated by ICI 182,780.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: Systemic sclerosis (SSc) is more prevalent in women. Our goal is to determine the effects of 17β-estradiol (E2) on the development of fibrosis and to compare circulating levels of estrogens in SSc patients and healthy controls.

Methods: Using primary human dermal fibroblasts, we evaluated the effect of E2 on fibronectin (FN) expression with and without the estrogen receptor (ER) antagonist ICI 182,780, inhibitors of signaling, propyl-pyrazole-triol, an ERα specific ligand, and genistein, an ERβ selective ligand, to identify the signaling pathways mediating E2's effect. We confirmed the fibrotic effect of E2 in human skin using an ex vivo organ culture model. Lastly, we measured levels of E2 and estrone in serum samples from SSc patients with diffuse cutaneous involvement and healthy controls using mass spectrometry.

Results: E2 increased expression of FN in dermal fibroblasts. ICI 182,780, inositol-1,4,5-triphosphate inhibitor, and p38 mitogen-activated protein kinase inhibitor blocked the effects of E2 on FN. Propyl-pyrazole-triol, but not genistein, significantly increased FN expression. Ex vivo, E2 induced fibrosis of human skin. The effects of E2 were abrogated by ICI 182,780. Circulating levels of E2 and estrone were significantly increased in sera of patients with diffuse cutaneous SSc.

Conclusion: Our findings implicate estrogens in the fibrotic process and may explain the preponderance of SSc in women. ICI 182,780 or other ER signaling antagonists may be effective agents for the treatment of fibrosis.

Show MeSH
Related in: MedlinePlus