The role of adhering platelets on villous trophoblast

Due to their small size and absence of a cell nucleus, maternal platelets adhering to villous surface of human placenta are frequently disregarded in routine microscopic surveys. However, adhering maternal platelets may represent an underappreciated source of modulating factors, which could affect placental trophoblast cells and their interaction with maternal circulating blood cells. We have recently shown maternal platelets on villous surface of human first trimester placenta, indicating that adherence of maternal platelets is a common process even in early stages of gestation. Moreover, our previous analysis of placental tissue specimens and their released factors revealed considerable levels of well-described platelet-derived signaling proteins. Based on these preliminary observations, this project will test the hypothesis whether the relative proportion of maternal platelets adhering to placental villous surface increases over gestation and whether this correlates with the degree of fibrinoid deposition, representing accumulation of amorphous, proteinaceous material. In this context, the current project will provide data on the incidence of adhering maternal platelets in placental tissue from pregnancies complicated by preeclampsia, which is characterized by high blood pressure and a large amount of protein in the urine. For this purpose platelets and fibrinoid deposits on placental surface will be analyzed in placenta tissues from healthy and preeclampsia patients by detection of specific platelet and fibrinoid markers. Since factors released from adhering platelets have manifold effects on various cell types, another research question of this project will test whether or not platelet- derived factors can affect proliferation, differentiation and hormone secretion of placental trophoblast cells, and moreover, whether platelet-derived factors can activate passing maternal immune cells. These study questions will be addressed by cultivation of placental trophoblast cells and tissue specimens as well as maternal blood cells with platelet-derived factors. Enhanced release of platelet- derived factors and activation of maternal circulating blood cells may have adverse effects on placental trophoblast cells and may overall contribute to a proinflammatory microenvironment in the human placenta. Proinflammatory conditions program early placenta functions and growth very early in pregnancy long before any changes become clinically apparent. The role of adhering maternal platelets on villous trophoblast cells has not been addressed before and represents scientific innovation. Thus, data from this project may open new mechanistic and therapeutic avenues for pregnancy complications related to enhanced placental incidence of adhering maternal platelets and could be the basis for further translational studies.

Data from this project suggest that maternal platelets can enter the intervillous space of human placenta very early in pregnancy, since they can be detected on the surface of the placental villi and in intercellular gaps of trophoblast cell columns from gestational week 5 onwards. Furthermore, results show that activation of maternal platelets at the maternal- placental interface impairs the expression and secretion of the classical pregnancy related hormone human chorionic gonadotropin (hCG) in human first trimester placenta. The impaired hCG synthesis is not the consequence of hampered morphological differentiation, as shown by analysis of differentiation-associated genes and electron microscopy of first trimester placental tissue and a trophoblast cell line. Inhibitor experiments suggest an interaction of two cellular signaling pathways, including a potential sequestration of transcription co-activators, which consequently results in a limitation of hCG production. While the presence of adhering maternal platelets on the surface of placental villi can impair hCG production, the significance of platelets in intercellular gaps of trophoblast cell columns remained undetermined. Immunohistochemistry suggests that platelets flow through vascular-like channels in distal parts of trophoblast columns, and spread into adjacent intercellular trophoblast gaps by leakage of maternal plasma flow. In these distal parts of columns, trophoblasts are loosely cohesive connected by desmosomes and have numerous surface projections extending into wide intercellular gaps as determined by transmission electron microscopy. Within these gaps, maternal platelets were identified by their characteristic open canalicular system (OCS), glycogen particles and organelles, such as dense bodies and a-granules. The observed amoeboid shape of the platelets as well as the appearance of fine-grained material in the OCS and cell surface invaginations suggest activation of platelets and extrusion of granules content into channels of the OCS and to the platelet exterior. Cell culture experiments suggest that the major histocompatibility complex, class I, G (HLA-G), which is primarily expressed by trophoblasts in cell columns, does not affect aggregation and adhesion of platelets. However, evidence for platelet activation in these areas tempts to speculate on a contributing role of platelet-derived factors in the differentiation processes of trophoblasts into the invasive phenotype.