Decidual natural killer cells regulate vessel stability: implications for impaired spiral artery remodelling

Abstract

Decidual NK (dNK) cells are present during uterine spiral artery remodelling, an event that is crucial for successful placentation and the provision of an adequate blood supply to the developing fetus. Spiral artery remodelling is impaired in the pregnancy complication pre-eclampsia. Although dNK cells are known to play active roles at the maternal-fetal interface, little is known about their effect on endothelial integrity, an important component of vessel stability. We present a study in which we have modelled dNK-endothelium interactions, using first-trimester dNK cells isolated from both normal pregnancies and those with impaired spiral artery remodelling. dNK cells were isolated from first-trimester pregnancies, screened by uterine artery Doppler ultrasound to determine resistance indices (RI) that relate to the extent of spiral artery remodelling. dNK culture supernatant from normal-RI pregnancies (but not high-RI pregnancies) destabilised endothelial tube-like structures in Matrigel, and normal-RI dNK cells induced endothelial intercellular adhesion molecule-1 and tumour necrosis factor-α expression to a greater extent than high-RI dNK cells. We have established a functional role for dNK cells in the disruption of endothelial structures and have suggested how impairment of this process may be contributing to the reduced vessel remodelling in pregnancies with a high uterine artery resistance index. These findings have implications for our understanding of the pathology of pre-eclampsia and other pregnancy disorders where remodelling is impaired.

Keywords: Decidua; Endothelium; Natural killer; Pre-eclampsia; Remodelling; Spiral artery.

Fig. 1

The effect of dNK cell secreted factors on EC ICAM-1 expression. Cell surface ICAM-1 expression (a marker of EC activation) by SGHEC-7 cells after 24-h incubation with pools of normal-RI or high-RI dNK cell-conditioned medium (CM). Results are mean + SEM of experiments carried out with six different pools of each patient group. *p < 0.05; Wilcoxon matched pairs signed rank test.

Fig. 2

The effect of normal and high-RI dNK secreted factors on EC tube-like structures. SGHEC-7 cells were allowed to form tube-like structures over 6 h, followed by the addition of pooled normal-RI or high-RI dNK-conditioned medium (CM). (A) Average length of EC tube-like structures is determined at 0, 2, 5, 10 and 18 h using Image Pro Plus software. The solid line represents normal-RI dNK CM, the dashed line represents high-RI dNK CM. Results are mean ± SEM of three separate experiments carried out in duplicate. ***p < 0.0001; ANOVA with Bonferroni post-test analysis, normal-RI CM versus high-RI CM at each time point. (B) Images of EC tube-like structures cultured in normal-RI or high-RI dNK CM at 0, 5 and 10 h.

Fig. 3

dNK-induced network destabilisation is not caused by apoptosis. SGHEC-7 cells were allowed to form tube-like structures over 6 h, followed by the addition of pooled normal-RI dNK-conditioned medium (CM) ± 50 μM zVAD-fmk. Average length of EC tube-like structures as determined at 0, 2, 5, 10 and 18 h using Image Pro Plus software. Solid line represents normal-RI dNK CM, dashed line represents normal-RI dNK CM with zVAD-fmk. Results are mean ± SEM of three separate experiments carried out in duplicate. Not significant; analysed using an ANOVA with Bonferroni post-test analysis comparing CM with and without zVAD-fmk at each time point.

Fig. 4

The effect of dNK cell secreted factors on EC TNFα expression. (A) SGHEC-7 cell TNFα expression was determined by ELISA after 24-h incubation with pools of normal-RI or high-RI dNK cell-conditioned medium (CM). Results are mean + SEM of five individual experiments carried out in duplicate. *p < 0.05; paired t-test. (B) The effect of TNFα on EC tube-like structures. SGHEC-7 cells were allowed to form tube-like structures over 6 h, followed by the addition of dNK culture medium (not conditioned medium), containing the indicated concentrations of TNFα. Results are mean + SEM of three separate experiments carried out in duplicate. *p < 0.05;**p < 0.001; ***p < 0.0001; ANOVA with Bonferroni post-test analysis compared with medium alone at each time point.