Relationships Between H2S and OT/OTR Systems in Preeclampsia

Abstract

Pre-eclampsia (PE) is a hypertensive pregnancy complication. Oxidative stress is hypothesized to contribute to the pathophysiology of PE. Both the hydrogen sulfide (H₂S) and oxytocin (OT) systems might play a role in the pathophysiology of PE, like their antioxidant and hypotensive effects. Thus, the role of the interaction of the OT and H₂S systems in the context of PE was further elucidated in the present clinical case-control study "NU-HOPE" (Nürnberg-Ulm: The role of H₂S and Oxytocin Receptor in Pre-Eclampsia; ethical approval by the Landesärztekammer Bayern, file number 19033, 29 August 2019), comparing uncomplicated pregnancies, early onset PE (ePE, onset < 34 weeks gestational age) and late onset PE (lPE, onset > 34 weeks gestational age). Routine clinical data, serum H₂S and homocysteine levels, and tissue protein expression, as well as nitrotyrosine formation, were determined. The main findings were (i) unchanged plasma sulfide levels, (ii) significantly elevated homocysteine levels in ePE, but not lPE, (iii) significantly elevated expression of H₂S enzymes and OT receptor in the placenta in lPE, and (iv) significantly elevated nitrotyrosine formation in the lPE myometrium. Taken together, these findings suggest a role for the interaction of the endogenous H₂S- and OT/OTR systems in the pathophysiology of pre-eclampsia, possibly linked to impaired antioxidant protection.

Keywords: cystathionine-β-synthase; cystathionine-γ-lyase; heme-oxygenase 1; homocysteine; oxidative stress.

Figure 1

Recruitment and sampling procedures. (A) Trial profile. Over a period of 6 months, all pregnant women who had an appointment for a scheduled cesarean section and all pregnant women with suspected pre-eclampsia were approached and all patients fulfilling the inclusion criteria were recruited for the study after written informed consent. Some patients dropped out of the study because they did not show up for follow-up appointments; one control patient had to be reassigned to the case group because of developing pre-eclampsia. Three patients from the case group were dropped from the study because the initial diagnosis of pre-eclampsia was not confirmed at a later timepoint. The case group was further divided into early and late onset pre-eclampsia. (B) Time course of the study.

Figure 2

Homocysteine and sulfide levels. Homocysteine p = 0.008 (Kruskal–Wallis test); ** p < 0.01. Plasma sulfide p = 0.59 (one-way ANOVA).

Figure 3

Representative pictures with the IHC target stained in red (left) and quantification (% positive tissue) (right) of protein expression in the placenta. CSE, CBS and nitrotyrosine were statistically analyzed with Kruskal–Wallis test. OTR was statistically analyzed with one-way ANOVA. The N per group is subject to variation, since some samples could not be used for the final analysis due to technical reasons (i.e., artificial IHC signal due to lifting of tissue or compromised tissue quality). This was not consistent throughout the groups and/or for a specific target protein, which is why the N for the individual analyses is given within the figure.

Figure 4

Representative pictures with the IHC target stained in red (left) and quantification (% positive tissue) (right) of protein expression in the myometrium of uterine samples. CSE and OTR were statistically analyzed with one-way ANOVA. CBS and nitrotyrosine were statistically analyzed with Kruskal–Wallis test. The N per group is subject to variation, since some samples could not be used for the final analysis due to technical reasons (i.e., artificial IHC signal due to lifting of tissue or compromised tissue quality). This was not consistent throughout the groups and/or for a specific target protein, which is why the N for the individual analyses is given within the figure.

Figure 5

Quantification of protein expression via Western blots. Top: HO-1, CSE and OTR expression in PBMC. PBMC from a healthy volunteer was used as naïve control. Bottom: HO-1 and CSE expression in the placenta. Protein extracts from HeLa cells were used as control. Bottom right: exemplary Western blots. HO-1, CSE and OTR in PBMC were statistically analyzed with the Kruskal–Wallis test. CSE and HO-1 in the placenta were statistically analyzed with one-way ANOVA. ePE = early onset pre-eclampsia, lPE = late onset pre-eclampsia.

Figure 6

Correlation analysis (Pearson) of placental CBS and OTR expression with linear regression (top left), placental CSE and OTR expression with linear regression (top right), uterine CBS and OTR expression with linear regression (middle left), uterine CSE and OTR expression ((middle right), no correlation), PBMC and placental OTR expression ((bottom left), no correlation) and PBMC and uterine OTR expression with linear regression (bottom right).