Immunomodulatory Siglecs are controlled by their glycoprotein and glycolipid ligands. Siglec-glycolipid interactions are often studied outside the context of a lipid bilayer, missing the complex behaviors of glycolipids in a membrane. Through optimizing a liposomal formulation to dissect Siglec–glycolipid interactions, it is shown that Siglec-6 can recognize glycolipids independent of its canonical binding pocket, suggesting that Siglec-6 possesses a secondary binding pocket tailored for recognizing glycolipids in a bilayer. A panel of synthetic neoglycolipids is used to probe the specificity of this glycolipid binding pocket on Siglec-6, leading to the development of a neoglycolipid with higher avidity for Siglec-6 compared to natural glycolipids. This neoglycolipid facilitates the delivery of liposomes to Siglec-6 on human mast cells, memory B-cells and placental syncytiotrophoblasts. A physiological relevance for glycolipid recognition by Siglec-6 is revealed for the binding and internalization of extracellular vesicles. These results demonstrate a unique and physiologically relevant ability of Siglec-6 to recognize glycolipids in a membrane.

The syncytiotrophoblast is a single giant multinucleate epithelial cell that forms the maternal surface of the human placenta and is bathed in maternal blood. It acts as a physical barrier between the maternal and fetal compartments and facilitates nutrient and waste exchange. As such, syncytiotrophoblast dysfunction is a key feature of pregnancy pathologies, such as preeclampsia. An understudied feature of dysfunctional syncytiotrophoblast is the loss of apical microvilli. But a paucity of data exists about the mechanisms regulating syncytiotrophoblast microvilli maintenance. Atypical protein kinase-c (aPKC) isoforms are evolutionarily conserved apical polarity regulators, which are known to play a role in the regulation of intestinal microvilli. Thus, we hypothesized that aPKC isoforms regulate syncytiotrophoblast microvilli and apical surface structure. Using human placental explant culture and primary human trophoblasts, we found that aPKCs regulate the structure, permeability, and endocytic function of the syncytiotrophoblast apical surface in a spatially restricted manner. A heightened inflammatory environment is often involved in the pathogenesis of placental pathologies, and the pro-inflammatory cytokine TNF-α can decrease aPKC-ι expression in intestinal cells. Here we establish that TNF-α exposure leads to reduced expression of the aPKC-ι isoform in syncytiotrophoblast and profoundly alters ST apical structure and permeability via regionalized pyroptosis, a highly pro-inflammatory form of cell death. Therefore, this is the first work to identify a regulator of apical-surface structure and the induction of the pyroptotic cascade at the maternal surface of the human placenta.

The syncytiotrophoblast is a human epithelial cell that is bathed in maternal blood on the maternal-facing surface of the human placenta. It therefore acts as a barrier and exchange interface between the mother and fetus. Syncytiotrophoblast dysfunction is a feature of pregnancy pathologies, like preeclampsia. Dysfunctional syncytiotrophoblasts display a loss of microvilli, which is a marker of aberrant apical–basal polarization, but little data exist about the regulation of syncytiotrophoblast polarity. Atypical PKC isoforms are conserved polarity regulators. Thus, we hypothesized that aPKC isoforms regulate syncytiotrophoblast polarity. Using human placental explant culture and primary trophoblasts, we found that loss of aPKC activity or expression induces syncytiotrophoblast gasdermin-E-dependent pyroptosis, a form of programmed necrosis. We also establish that TNF-α induces an isoform-specific decrease in aPKC expression and gasdermin-E-dependent pyroptosis. Therefore, aPKCs are homeostatic regulators of the syncytiotrophoblast function and a pathogenically relevant pro-inflammatory cytokine leads to the induction of programmed necrosis at the maternal–fetal interface. Hence, our results have important implications for the pathobiology of placental disorders like preeclampsia.