Molecular regulation of endothelial glycocalyx-mediated transcytosis

Endothelial cells are the primary regulatory interface for the exchange of substances between blood and tissues. The luminal (blood-facing) surface of endothelial cells contains a complex, carbohydrate rich structure called the glycocalyx which is the first surface of interactions between circulating components and endothelial cells. However, the involvement of glycocalyx components in the binding, uptake, and transport of substances across endothelial barriers is inadequately understood, particularly in vivo. Our research over the last 5 years has elucidated fundamental regulatory aspects of glycocalyx-mediated transendothelial transport of chemokines and viral proteins. We have observed that there are tissue-specific differences in transport regulation, and that transport can occur selectively in different vascular zones, suggesting that the molecular machinery of the glycocalyx that regulates transport also varies by tissue and vascular zone. A particular strength of our approach is our unique ability to conduct highly sensitive and rigorous radiochemical-based assays which enable us to calculate blood-to-tissue transport rates of proteins in vivo and in situ, as well as other pharmacological parameters such as vascular binding, biodistribution, and clearance. We have also developed cell-based assays that are amenable for studies of glycocalyx-dependent transendothelial transport, with intact HS-dependent chemokine transport processes resembling those in vivo. In the next 5 years, our main goal is to characterize the molecular components that regulate the basic biological process of glycocalyx-mediated transendothelial transcytosis, including those that confer tissue specificity of transport functions. As heparan sulfate (HS) is a major component of the glycocalyx which regulates the transendothelial transport of chemokines, we will focus on the molecular components of HS- mediated chemokine transport as an initial example. We hypothesize that chemokines undergo endothelial transcytosis by binding to HS on membrane-attached HS proteoglycans (HSPGs i.e. syndecans or glypicans), which are subsequently endocytosed and routed through vesicular pathways to the abluminal side, where they are released. Our research plan entails the implementation of spatial transcriptomics, proteomics, glycomics, and cell biology to relate our in vivo functional results to the expression and composition of endothelial HSPGs in different tissues and vascular zones. We will pair these results with functional assays in vitro using pharmacologic and molecular approaches to identify the HSPGs and attached HS moieties that are regulating transendothelial transport. We will additionally characterize transcytotic routing mechanisms of HSPGs and their bound ligands in vitro using fluorescent imaging, pharmacologic, and molecular approaches. Achievement of these goals would advance the general knowledge of endothelial cell biology and endocytic/transcytotic transport mechanisms. Our findings would also have broad applications in studying biological processes that involve HS/ligand interactions in health and disease, and for drug development. Project Number: 1R35GM161201-01 | Fiscal Year: 2026 | NIH Institute/Center: National Institute of General Medical Sciences (NIGMS) | Principal Investigator: Michelle Erickson | Institution: SEATTLE INST FOR BIOMEDICAL/CLINICAL RES, SEATTLE, WA | Award Amount: $374,550 | Activity Code: R35 | Study Section: Special Emphasis Panel[ZRG1 CDB-N (55)] View on NIH RePORTER: https://reporter.nih.gov/project-details/11258364