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Lee, Kim, Park, Choi, Chung, and Moon: The effect of estrogen on extracellular signal-regulated kinases activation in uterine artery smooth muscle cells during pregnancy
Original Article
Korean Journal of Obstetrics & Gynecology

Korean Journal of Obstetrics & Gynecology 2011; 54(9): 499-507.

Published online: 26 September 2011

DOI: https://doi.org/10.5468/KJOG.2011.54.9.499

The effect of estrogen on extracellular signal-regulated kinases activation in uterine artery smooth muscle cells during pregnancy

Yong Woo Lee, MD, Ho Yeon Kim, MD, Sang Min Park, MD, Soo Ran Choi, MD, Ji Youn Chung, MD, Chong Soo Moon, MD

Department of Obstetrics and Gynecology, Hallym University College of Medicine, Seoul, Korea.

Corresponding author: Chong Soo Moon, MD, PhD. Department of Obstetrics and Gynecology, Kangdong Sacred Heart Hospital, Hallym University College of Medicine, 445 Gil-dong, Gangdong-gu, Seoul 134-701, Korea. Tel: +82-2-2224-2259, Fax: +82-2-2224-2265, obgyn25@hallym.or.kr

Received 30 May 2011       Revised 27 June 2011       Accepted 27 June 2011

Copyright © 2011. Korean Society of Obstetrics and Gynecology

(open-access):

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Objective

To explore the endothelium-independent mechanisms of estrogen induced uterine vasodilatation, this study was performed to determine whether uterine artery smooth muscle (UASM) cells are direct targets of estrogen, and estradiol (E2) stimulates extracellular signal-regulated kinases (ERK2/1) in endothelium denuded UASMs.

Methods

The uterine arteries were obtained from late gestation pregnant sheep and the endothelium was denuded with collagenase digestion. The uterine artery smooth muscle segments were digested, collected and cultured. Endothelial integrity and smooth muscle status were assessed by Double immunofluorescence staining and flow cytometry. The UASM cells were treated with increasing concentrations of E2 (10-14 to 10-6 M), and pretreated with ICI 182,780 followed by different concentrations (10-10 and 10-7 M) of E2. Western blot analysis of ERK2/1 phosphorylation with a phospho-mitogen activated protein kinases (MAPKs) antibody were carried out to total cell extracts.

Results

The loss of endothelial function and adequacy of smooth muscle integrity were confirmed. When challenged with increasing concentrations of E2, a bi-phase ERK2/1 phosphorylation was observed. Treatment with low doses (10-14 to 10-10 M) of E2, ERK2/1 phosphorylation was dose-dependently increased, whereas high doses (10-9 to 10-8 M) did not phosphorylate ERK2/1. However, treatment with pharmacological doses (10-7 to 10-6 M) drastically phosphorylated ERK2/1. In the presence of ICI 182,780, E2 induced ERK2/1 phosphorylation were abolished in both.

Conclusion

It suggests that UASM is the target tissue of estrogen during uterine vasodilatation, and estrogen stimulation of ERK2/1 activation is mediated by an estrogen receptor-dependent mechanism. It also is presumed that endothelium independent mechanism exists in estrogen induced vasodilatation.

Keywords: Estrogen, Uterine artery smooth muscle, Vasodilatation, ERK2/1

Figures and Tables

Fig. 1
Morphology of uterine artery smooth muscle cells in culture and double immunofluorescence staining of α-actin and caveolin-1 (×400). SM-α-actin, smooth muscle α-actin.
kjog-54-499-g001
Fig. 2
Flow cytometric analysis of uterine artery smooth muscle cells. SSC-H, stained smooth muscle cells; FSC-H, fluorescence activated cells; FL1-H, flow cytometric analysis; eNOS mAb, endothelial nitric oxide synthase monoclonal antibody; SM-α-actin, smooth muscle α-actin; rIgG, anti rabbit immunoglobulin G fluorescein isothiocyanate conjugate; Cav-1 pAb, caveolin-1 polyclonal antibody.
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Fig. 3
Western blot analysis of estrogen receptor α and β in uterine artery smooth muscle and uterine artery smooth muscle cell. UASM, uterine artery smooth muscle; UASMC, uterine artery smooth muscle cell; ER, estrogen receptor.
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Fig. 4
Effects of estrogen on extracellular signal-regulated kinases (ERK2/1) activation in uterine artery smooth muscle cell. WB: p-ERK pAb, western blot phospho-extracellular signal-regulated kinases polyclonal antibodies; pERK, phospho-ERK; M, molecular weight.
kjog-54-499-g004
Fig. 5
Role of estrogen receptor in estrogen induced extracellular signal-regulated kinases (ERK2/1) activation in uterine artery smooth muscle cell. WB: p-ERK pAb, western blot phospho-extracellular signal-regulated kinases polyclonal antibodies; pERK, phospho-ERK; ICI, ICI 182,780; E2, estrdiol; WB:Pan-ERK mAb, western blot phospho-extracellular signal-regulated kinases monolonal antibodies.
kjog-54-499-g005

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