S AGGARWAL 1,2, B XU 2, A MAKRIS 1,2,3, A HENNESSY 1,2,4
1School of Medicine, Western Sydney University, Campbeltown, Australia, 2The Heart Research Institute, Newtown, Australia, 3Renal unit, Liverpool Hospital, Liverpool, Australia, 4Campbeltown Hospital, Campbeltown, Australia
Background: Angiotensin II type 1 receptor antibodies (AT1AA) (an agonistic antibody to Angiotensin II (Ang II) have been implicated in the pathophysiology of preeclampsia.
Aims: A two-cell model of endovascular invasion that mimics human placentation was used to assess the effects of AT1AA and Ang II on cellular networks and angiogenic markers.
Methods: Uterine microvascular endothelial cells were cultured on Matrigel™. After endothelial cellular networks formed, HTR8/SVNeo trophoblasts,, which had been co-cultured with purified human AT1AA, Ang II or combination polyclonal rabbit AT1AA and Ang II were added. Human AT1AA was purified from sera of women with preeclampsia using MicroLink protein coupling columns. Co-cultures were incubated with either 2% O2 or 21% O2. Images were captured by fluorescence microscopy. Trophoblast integration into endothelial cells was calculated using Image J software. Soluble fms like tyrosine kinase-1 (sFlt-1) was measured in conditioned media using an ELISA. Quantitative PCR for sFlt-1, soluble endoglin (sEng) and hypoxia inducible factor 1 alpha (HIF-1α) were conducted on cell pellets. All data analysed using Graphpad Prism.
Results: Addition of human AT1AA to co-cultures decreased cellular integration in 2% O2 (p=0.03) but not in 21% O2 (n=4). Ang II alone did not affect integration, however the combination of Ang II and AT1AA further decreased cellular integration in 2% O2 compared to AT1AA alone. mRNA expression of sFLT-1, sEng and HIF-1α and sFlt-1 concentrations in cultured media were not altered by the addition of AT1AA+/- Ang II.
Conclusion: The presence of AT1AA inhibits trophoblast invasion in a setting of physiological hypoxia (2% O2), there is further inhibition when AT1AA is combined with Ang II. These findings indicate that AT1AA may mediate early preeclampsia pathophysiology.
Shikha Aggarwal is a renal physician currently working as a VMO at Northern beaches hospital and The Mater hospital in Sydney. She has a special interest in Obstetric renal medicine. She is currently in her final year of a PHD through Western Sydney University studying the role of Angiotension II type 1 receptor antibodies in preeclampsia.