Introduction
Pre-eclampsia (PE) is a systemic vascular Fdisorder affecting 4-5% of pregnant women globally1. PE is a leading cause of maternal and foetal morbidity and mortality, and confers a life- long risk to maternal and infant’s cardiovascular health2-5. Classically, PE manifests as new-onset hypertension and proteinuria after 20 weeks of gestation6. Nevertheless, symptoms, severity, time of onset, and outcome can vary significantly and early diagnosis remains challenging 6, 7.
The wide range of clinical phenotypes makes PE more fitting to be a syndrome instead of a single disease. Especially the distinction early- and late-onset PE (EO-PE and LO-PE) is frequently applied and both are likely to have distinct aetiologies8, 9. Nonetheless, the exact definition of EO-PE and LO-PE is controversial. The current stratification only focusses on time of onset of clinical symptoms, gestational age at time of birth, or a combination of these, without referring to underlying causes6, 9, 10.
Quantifying altered placental morphology in different clinical presentations of PE is key to distinguish the clinical syndrome into distinct pathological phenotypes. Currently, 2D histological evaluation is the gold standard for placental investigation11-13. Yet, none of the histological findings are specific for PE, i.e., are also observed in other placental syndromes like intra-uterine growth restriction (IUGR)14-16.
More advanced imaging methods such as confocal microscopy have been used for imaging terminal villous vasculature17-20. However, the villous tree is built up of stem villi (of which the diameter can be up to 3000 µm), intermediate villi, and terminal villi21.
Multiphoton microscopy (MPM) is a tomographic (3D imaging) technique with subcellular resolution. It is based on multiphoton excited fluorescence and features deep tissue penetration (up to 1 μm)22. Therefore, thick tissue slabs (containing different components of the villous tree) can be visualized, closely to their native form. Moreover, MPM is efficient in exciting autofluorescence signal, suitable for label free imaging of imaging cytoplasm and connective tissue even in vivo23.
MPM has been applied to investigate placental membrane architecture24, 25. But, to our knowledge, phenotype quantification of the villous tree and vasculature of healthy and PE placentas with MPM has not been described yet. The aim of this study was to develop a MPM placental imaging protocol and to demonstrate quantifiable imaging targets. Quantification of these targets could aid in a more unequivocal characterization of different PE subtypes and improve disease stratification.