During the first trimester, placental development occurs in a low oxygen environment that is known to stimulate angiogenesis and proliferation via upregulation of vascular endothelial growth factor (VEGF). Expression of the placental renin-angiotensin system (RAS) is highest in early pregnancy when oxygen tension is low. Components of the RAS, including angiotensin II type 1 receptor (AGTR1), are significantly upregulated in low oxygen. This is also associated with increased VEGF expression in a first trimester extravillous trophoblast cell line (HTR-8/SVneo) which can be inhibited using losartan (AGTR1 antagonist). Changes in miRNAs known to target these RAS genes also occur. We postulated that low oxygen would increase the expression of AGTR1 as well as VEGF in first trimester human chorionic villous explants and that this would be associated with decreased expression of miRNAs that target AGTR1.
Human first trimester chorionic villous explants (6-12 weeks gestational age) were cultured in 1%, 5% or 20% O2 in DMEM-HG (supplemented with 10% FCS, 1 mg/ml antibiotic-antimycotic and 1% L-glutamine) for 48h (n=8). AGTR1 and VEGF mRNA levels as well as expression of miRNAs predicted to target AGTR1 were determined by qPCR.
Culture in low oxygen (1% O2) significantly increased the expression of AGTR1 mRNA compared with culture in 5% O2 (p=0.03). VEGF mRNA expression was similarly enhanced by culture in 1% O2 compared with culture in 20% O2 (p=0.01). miR-483-3p, which when overexpressed in HTR-8/SVneo cells significantly reduces AGTR1 expression (p=0.03), was significantly reduced in villi cultured in 1% O2 when compared with culture in 20% O2 (p=0.028). Thus, this reduction in miR-483-3p may be responsible for the oxygen-induced upregulation of AGTR1 and subsequent stimulation of angiogenic factors in first trimester chorionic villi.
This study highlights the potential role of miRNAs in promoting the low oxygen-induced AGTR1 expression in placental angiogenesis.