Dysregulated cell mechanical properties of endometrial stromal cells from endometriosis patients

Abstract

Endometriosis, diagnosed with ectopically implanted endometrial stromal cells (ESC) and epithelial cells to a location outside the uterine cavity, seriously threaten the quality of life and reproductive ability of women, yet the mechanisms and the pathophysiology of the disease remain unclear. Specially, the functional changes of ESC during endometriosis progression need in-depth investigation. In this study, we characterized mechanical properties of normal ESC (NESC) from healthy women and eutopic ESC (EuESC) and ectopic ESC (EcESC) from endometriosis patients. We found the collagen lattice contractile ability of EuESC was significantly stronger than that of NESC, and the cell mobility of EuESC and EcESC was significantly greater than that of NESC. Furthermore, the expression of F-actin and vinculin in NESC, EuESC and EcESC cells progressively increased, and the Rho GTPase activity, of which RhoA exhibited the highest activity, in the three cells gradually increased. Collectively, these results suggest that the mechanical characteristics of NESC, EuESC and EcESC cells exhibited progressive abnormalities. Therefore, the biomechanics of endometrial stromal cells may be a potent target for intervention in patients with endometriosis.

Keywords: Endometriosis; RhoA; cell mechanics; contraction; motility.

Figure 1

Comparison of the area of the collagen lattice containing NESC or EuESC after 24 hours of 3-dimensional culture. The diagram above shows the representative collagen contraction of NESC and EuESC. The diagram below demonstrates the statistical analysis of the collagen lattice area. The white spots outlined the gels. The experiment was performed in triplicate and repeated 3 times using NESC and EuESC from different individuals.

Figure 2

The cell motility of NESC, EuESC and EcESC. The pictures left show representative cell movement of 3 types of cells after 24 hours of culture. The dotted line outlined cell boundary at 0 hour and the black line outlined cell boundary at 24 hour of culture. The area between the lines is the increased area covered by the outward movement of the cells and was analyzed on the right panel. The experiment was performed in triplicate and repeated 3 times using NESC, EuESC and EcESC cells from different individuals.

Figure 3

Immunostaining of F-actin and vinculin in NESC, EuESC and EcESC. The upper 3 pictures represent phalloidin-stained green F-actin, and the lower 3 pictures represent vinculin-stained red focal adhesion complexes. In all 3 types of cells, the phalloidin and vinculin staining intensities gradually increased.

Figure 4

RhoA, Rac1 and Cdc42 expression in NESC, EuESC and EcESC. The first row demonstrates the expression levels of the active small GTPase, the second row shows the total GTPase expression level, and the third row demonstrates the internal reference (ACTB). The left column reports the RhoA expression. The EcESC exhibited the highest level of activity, followed by EuESC, and the activity of NESC was the weakest. The diagrams below were densitometric analysis of activated GTPases from above western images.