Honors Program
Date of this Version
5-2024
Document Type
Thesis
Citation
Rohr R. Structure and Heparin Binding Capabilities of Rodent Stabilin-2. 2024. Undergraduate Honors Thesis. University of Nebraska-Lincoln.
Abstract
Background: The clearance of systemic circulatory hyaluronan (HA) is mediated by the scavenger receptor, Stabilin-2. Stabilin-2 is expressed in native tissues as two isoforms: a large monomeric isoform which is encoded by the full-length processed gene and a smaller isoform that is presumably a proteolytically processed product of the large isoform. This smaller isoform was initially cloned and characterized in the rodent model (rat) and named the Hyaluronic Acid Receptor for Endocytosis (HARE) around 25 years ago. Thus, the large isoform is named Stabilin-2, and the smaller isoform is named HARE, though they both perform the same function in human tissues. Stabilin-2/HARE are predominantly expressed in the sinusoidal endothelial cells of the liver (LSECs), lymph node, spleen, and bone marrow. Their expression in the LSECs, which interfaces the blood on one side and the Space of Disse on the other, is what makes these receptors optimal for their clearance function. Human Stabilin-2/HARE are better characterized as clearance receptors for numerous blood-borne ligands such as hyaluronan, heparin, chondroitin sulfate, phosphatidylserine, oxidized low density lipoprotein, advanced glycation end-products, Factor VIII and others. Hypothesis: There has been some controversy in the field as to whether heparin is a ligand for rodent Stabilin-2/HARE, and we set to prove our hypothesis that this receptor does bind heparin. Objective: The objective of this study is to resolve this issue by testing if recombinant Stabilin-2/HARE binds and internalizes unfractionated heparin (UFH). Both Stabilin-2 and HARE were independently expressed in stable Flp-In 293 cell lines and tested for heparin binding and endocytosis using HA as a control. Results: The results indicate that heparin binding and endocytosis to rat Stabilin-2/HARE is comparable with human Stabilin-2/HARE, though the affinity of UFH may be slightly lower for the rat receptors. During these experiments, it was also discovered that the rat receptors have a faster turnover rate and same affinity for HA than the human receptors, and the production of HARE from the Stabilin-2 protein in human recombinant cells is very low compared to what occurs in native rat tissues. Conclusion: Our conclusions are that the rat receptors are similar to the human protein receptors and that the rodent model is a relevant physiological model for human metabolism and disease regarding Stabilin-2/HARE activity.
Comments
Copyright Reed Rohr 2024