Background: The syncytiotrophoblast is a large multinucleated cell layer that is maintained by continuous fusion of underlying cytotrophoblast cells in a process known as syncytialisation. The syncytiotrophoblast is the major functional cell layer of the placenta, responsible for regulating feto-maternal nutrient, gas and waste exchange. The placental renin angiotensin system (RAS) is essential for placental development however, little is known about its role in syncytialisation. We therefore aimed to determine the effect of syncytialisation on RAS expression. To do this we used two models of syncytialisation; forskolin-induced syncytialisation of BeWo choriocarcinoma cells and spontaneously fusing human primary trophoblast cells.
Methods: BeWo choriocarcinoma cells were treated with 100mM forskolin 24h post plating and left to syncytialise for 48h before cells and supernatant were collected. Primary trophoblast cells were isolated from human placentae and left to spontaneously syncytialise for 72h, with collection of cells and supernatant every 24h. The mRNA expression of RAS genes including: prorenin (REN), (pro)renin receptor (ATP6AP2), angiotensinogen (AGT), angiotensin converting enzyme 1/2 (ACE/ACE2) and angiotensin II receptor type 1 (AGTR1), were assessed by RT-qPCR.
Results: Forskolin-induced syncytialisation of BeWo cells significantly decreased ATP6AP2 and ACE expression (P=0.002 and <0.0001, respectively), and increased AGT and ACE2 expression (both P<0.001). There were low to undetectable levels of REN and AGTR1 mRNA in BeWo cells and these did not change with forskolin treatment. Spontaneous syncytialisation of trophoblast cells significantly decreased ACE2 expression (P<0.0001) and increased AGT and REN expression (both P<0.05). ATP6AP2, AGTR1 and ACE mRNA expression were unchanged.
Conclusion: BeWo and primary trophoblast cells exhibit different RAS mRNA profiles both before and in response to syncytialisation, highlighting a significant difference between these two in vitro models. BeWo cells predominantly utilise RAS-independent pathways induced by the (P)RR, whereas primary trophoblasts rely more on classical RAS pathways.