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. 2021 Oct 29;10(21):5086.
doi: 10.3390/jcm10215086.

Sildenafil Citrate Downregulates PDE5A mRNA Expression in Women with Recurrent Pregnancy Loss without Altering Angiogenic Factors-A Preliminary Study

Monika Kniotek  1 , Aleksander Roszczyk  1 , Michał Zych  1 , Małgorzata Wrzosek  2   3 , Monika Szafarowska  4 , Radosław Zagożdżon  1   5 , Małgorzata Jerzak  2   3   6
Affiliations

Sildenafil Citrate Downregulates PDE5A mRNA Expression in Women with Recurrent Pregnancy Loss without Altering Angiogenic Factors-A Preliminary Study

Monika Kniotek et al. J Clin Med. .

Abstract

In our previous study, we showed that sildenafil citrate (SC), a selective PDE5A blocker, modulated NK cell activity in patients with recurrent pregnancy loss, which correlated with positive pregnancy outcomes. It was found that NK cells had a pivotal role in decidualization, angiogenesis, spiral artery remodeling, and the regulation of trophoblast invasion. Thus, in the current study, we determined the effects of SC on angiogenic factor expression and production, as well as idNK cell activity in the presence of nitric synthase blocker L-NMMA. Methods: NK cells (CD56+) were isolated from the peripheral blood of 15 patients and 15 fertile women on MACS columns and cultured in transformation media containing IL-15, TGF-β, and AZA-a methylation agent-for 7 days in hypoxia (94% N2, 1% O2, 5% CO2). Cultures were set up in four variants: (1) with SC, (2) without SC, (3) with NO, a synthase blocker, and (4) with SC and NO synthase blocker. NK cell activity was determined after 7 days of culturing as CD107a expression after an additional 4h of stimulation with K562 erythroleukemia cells. The expression of the PDE5A, VEGF-A, PIGF, IL-8, and RENBP genes was determined with quantitative real-time PCR (qRT-PCR) using TaqMan probes and ELISA was used to measure the concentrations of VEGF-A, PLGF, IL-8, Ang-I, Ang-II, IFN-γ proteins in culture supernatants after SC supplementation. Results: SC downregulated PDE5A expression and had no effect on other studied angiogenic factors. VEGF-A expression was increased in RPL patients compared with fertile women. Similarly, VEGF production was enhanced in RPL patients' supernatants and SC increased the concentration of PIGF in culture supernatants. SC did not affect the expression or concentration of other studied factors, nor idNK cell activity, regardless of NO synthase blockade.

Keywords: NK cells; PDE5A; RPL; VEGF-A; angiotensin; sildenafil.

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Conflict of interest statement

The authors declare that there is no conflict of interest regarding the publication of this paper.

Figures

Figure 1
Figure 1
PDE5A, VEGF-A, ANG-I, PIGF, IL-8 expression after SC treatment in the idNK cells of healthy and RPL women, data shown as the mean ± SD compared with idNK cells maintained without SC (green points, CG—control group, n = 10; red points—RPL patients, n = 8; statistical significance * p < 0.05).
Figure 2
Figure 2
The expression of PDE5A and VEGF-A is upregulated in the idNK cells of patients with RPL. SC downregulated PDE5A gene expression in RPL group, (a) a genomic meta-analysis of PDE5A gene expression in idNK cells, (b) a genomic meta-analysis of VEGF-A gene expression in idNK cells (c) a genomic meta-analysis of RNEBP gene expression in idNK cells, (d) a genomic meta-analysis of PIGF gene expression in idNK cells, (e) a genomic meta-analysis of IL-8 gene expression in idNK cells; data are shown as the median and interquartile range, CG—control group n = 10, RPL patients n = 8, circles and triangles represents single samples; SC—sildenafil citrate 400 ng/mL, statistical significance marked as star—* p < 0.05.
Figure 3
Figure 3
The concentrations of angiogenic factors and cytokines in idNK cultures with and without sildenafil citrate: (a) VEGF-A, (b) PIGF, (c) Ang I, (d) Ang II, (e) IL-8; (f) IFN-γ; Control group n = 9, RPL patients n = 8; dots on the figures reflects numbers of determined samples, SC—sildenafil citrate 400 ng/mL, statistical significance marked as star—* p < 0.05.
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