MZe786, a hydrogen sulfide-releasing aspirin prevents preeclampsia in heme oxygenase-1 haplodeficient pregnancy under high soluble flt-1 environment

Abstract

Preeclampsia affects one in twelve of the 130 million pregnancies a year. The lack of an effective therapeutic to prevent or treat it is responsible for an annual global cost burden of 100 billion US dollars. Preeclampsia also affects these women later in life as it is a recognised risk factor for cardiovascular disease, stroke and vascular dementia. Our laboratory demonstrated that preeclampsia is associated with high soluble fms-like tyrosine kinase 1 (sFlt-1) and low heme oxygenase-1 (HO1/Hmox1) expression. Here we sought to determine the therapeutic value of a novel H₂S-releasing aspirin (MZe786) in HO-1 haploid deficient (Hmox1^+/-) pregnant mice in a high sFlt-1 environment. Pregnant Hmox1^+/- mice were injected with adenovirus encoding sFlt-1 or control virus at gestation day E11.5. Subsequently, Hmox1^+/- dams were treated daily with a number of treatment regimens until E17.5, when maternal and fetal outcomes were assessed. Here we show that HO-1 compromised mice in a high sFlt-1 environment during pregnancy exhibit severe preeclampsia signs and a reduction in antioxidant genes. MZe786 ameliorates preeclampsia by reducing hypertension and renal damage possibly by stimulating antioxidant genes. MZe786 also improved fetal outcome in comparison with aspirin alone and appears to be a better therapeutic agent at preventing preeclampsia than aspirin alone.

Keywords: Heme oxygenase-1; Hydrogen sulfide; Hypertension; Preeclampsia; sFlt-1.

Graphical abstract

Fig. 1

High sFlt-1 causes a severe preeclampsia phenotype in Hmox1 haploid deficient mice. (A) Mean arterial blood pressure (MAP) measured at E17.5 gestation in Hmox1^+/+ (Black circles) and Hmox1^+/− (black triangle) mice after administration of Ad-CMV (control) or Ad-sFlt-1 (1x10⁹ pfu) at E11.5. (B) Serial sections of representative glomeruli stained with hematoxylin and eosin (HE). Scale bars, 100 μm. (C) Single-blind analysis of glomeruli damage from Hmox1^+/+ (Black column) and Hmox1^+/− (White column) mice treated with Ad-CMV or Ad-sFlt-1. (n = 6 Hmox1^+/+; n = 6 Hmox1^+/−). (D) Representative images of uterine horn (E17.5) showing overexpression of sFlt-1 leads to fetal death (FD). (E) Fetal loss expressed as a percentage of the total pup numbers. (F) Fetal weight distribution of pups and 10th percentile pup weight population mark, representing the fetal growth restriction in the weight curve from Hmox1^+/+ and Hmox1^+/− mice treated with Ad-CMV or Ad-sFlt-1. (n = 13 Hmox1^+/+; n = 14 Hmox1^+/−). Results are expressed as representative or as mean (±SEM). Analysed by Two-way ANOVA.

Fig. 2

MZe786 rescues maternal outcome in Hmox1^+/−pregnant mice in a high sFlt-1 environment. (A) H₂S metabolite, trimethyl sulfonium (TMS) levels measured in urine on day E17.5 in Hmox1^+/− timed pregnant mice injected Ad-sFlt-1 and treated with Carrier (control), MZe786, MZe486 or aspirin. (B) Mean arterial blood pressure (MAP) recorded at E17.5 gestation in Hmox1^+/− timed pregnant mice injected with Ad-CMV or Ad-sFlt-1 and treated with MZe786, MZe486 or aspirin. (C) Representative serial sections of glomeruli stained with hematoxylin and eosin (H&E). Scale bars, 100 μm. (D) Single-blind analysis of glomeruli damage. (E) Urinary kidney injury marker −1 (KIM-1) level. (F) Plasma sEng level measured on day E17.5 gestation in Hmox1^+/− timed pregnant mice injected with Ad-CMV or Ad-sFlt-1 and treated with carrier, MZe786, MZe486 or aspirin (n = 11 Ad-CMV; n = 13 Carrier; n = 13 MZe786; n = 14 MZe486; n = 10 Aspirin group). Results are expressed as representative or as mean (±SEM). Analysed by One-way ANOVA.

Fig. 3

MZe786 improves fetal outcome in a high sFlt-1 environment in Hmox1^+/−mice. (A) Representative images of uterine horn (E17.5) showing rescue of fetal death (FD) in sFlt-1 induced Hmox1^+/− mice by H₂S-releasing molecules. (B) Fetal loss expressed as a percentage over total pup numbers in Hmox1^+/− mice injected with Ad-CMV or Ad-sFlt-1 and treated with MZe786, MZe486 or aspirin. (C) Hmox1^+/− fetal weight distribution and 10th percentile pup weight population mark, representing the fetal growth restriction in the weight curve (D) Hmox1^+/− Fetal:placental ratio in Hmox1^+/− mice injected with Ad-CMV or Ad-sFlt-1 and treated with MZe786, MZe486 or aspirin. Results are expressed as representative or as mean (±SEM). Analysed by One-way ANOVA.

Fig. 4

MZe786 stimulates antioxidant defence in Hmox1^+/−mice overexpressing sFlt-1. (A) Kidney relative mRNA expression of genes; (B) Txn1, (C) Glrx and (D) PGC1α measured by quantitative PCR n Hmox1^+/− mice injected with Ad-CMV or Ad-sFlt-1 and treated with carrier, MZe786 or aspirin. (E) Mitochondrial content measured as the ratio of mtDNA/nDNA in Hmox1^+/− mice injected with Ad-CMV or Ad-sFlt-1 and treated with carrier, MZe786 or aspirin. Results are expressed as representative or as mean (±SEM). Analysed by One-way ANOVA.