Mechanical stiffness augments ligand-dependent progesterone receptor B activation via MEK 1/2 and Rho/ROCK–dependent signaling pathways in uterine fibroid cells

Mechanical stiffness enhanced PRB-dependent gene activation via MEK 1/2 and Rho/ROCK signaling pathways, providing an initial framework for understanding the relationship between mechanical stiffness and PR signaling.

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VOLUME 116, ISSUE 1, P255-265

Authors:

Christina N. Cordeiro Mitchell, M.D., Md Soriful Islam, Ph.D., Sadia Afrin, Ph.D., Joshua Brennan, M.P.H., Kevin J. Psoter, Ph.D., James H. Segars, M.D. 

Abstract:

Objective

To test whether mechanical substrate stiffness would influence progesterone receptor B (PRB) signaling in fibroid cells. Uterine fibroids feature an excessive extracellular matrix, increased stiffness, and altered mechanical signaling. Fibroid growth is stimulated by progestins and opposed by anti-progestins, but a functional interaction between progesterone action and mechanical signaling has not been evaluated.


Design

Laboratory studies.


Setting

Translational science laboratory.


Patient(s)/Animal(s)

Human fibroid cell lines and patient-matched fibroid and myometrial cell lines.


Intervention(s)

Progesterone receptor B–dependent reporter assays and messenger RNA quantitation in cells cultured on stiff polystyrene plates (3GPa) or soft silicone plates (930KPa). Pharmacologic inhibitors of extracellular signal-related protein kinase (ERK) kinase 1/2 (MEK 1/2; PD98059), p38 mitogen-activated protein kinase (SB202190), receptor tyrosine kinases (RTKs; nintedanib), RhoA (A13), and Rho-associated coiled-coil kinase (ROCK; Y27632).


Main Outcome Measure(s)

Progesterone-responsive reporter activation.


Result(s)

Fibroid cells exhibited higher PRB-dependent reporter activity with progesterone (P4) in cells cultured on stiff vs. soft plates. Mechanically induced PRB activation with P4 was decreased 62% by PD98059, 78% by nintedanib, 38% by A13, and 50% by Y27632. Overexpression of the Rho-guanine nucleotide exchange factor (Rho-GEF), AKAP13, significantly increased PRB-dependent reporter activity. Collagen 1 messenger RNA levels were higher in fibroid cells grown on stiff vs. soft plates with P4.


Conclusion(s)

Cells cultured on mechanically stiff substrates had enhanced PRB activation via a mechanism that required MEK 1/2 and AKAP13/RhoA/ROCK signaling pathways. These studies provide a framework to explore the mechanisms by which mechanical stiffness affects progesterone receptor activation.

Fertility and Sterility

Editorial Office, American Society for Reproductive Medicine

Fertility and Sterility® is an international journal for obstetricians, gynecologists, reproductive endocrinologists, urologists, basic scientists and others who treat and investigate problems of infertility and human reproductive disorders.