While increased levels of high‐density lipoprotein (HDL)‐cholesterol correlate with protection against cardiovascular disease, recent findings demonstrate that HDL function, rather than HDL‐cholesterol levels, may be a better indicator of cardiovascular risk. One mechanism by which HDL function can be compromised is through modification by reactive aldehydes such as acrolein (Acro), 4‐hydroxynonenal, and malondialdehyde (MDA). In this study, we tested the hypothesis that modification of HDL with reactive aldehydes would impair HDL’s athero‐protective functions in macrophages. Compared to native HDL, Acro‐ and MDA‐modified HDL have impaired abilities to promote migration of primary peritoneal macrophages isolated from C57BL6/J mice. Incubation of macrophages with MDA‐HDL also led to an increased ability to generate reactive oxygen species. Our studies revealed that the changes in HDL function following aldehyde modification are likely not through activation of canonical nuclear factor‐kappa B signaling pathways. Consistent with this finding, treatment of either noncholesterol‐loaded macrophages or foam cells with modified forms of HDL does not lead to significant changes in expression levels of inflammatory markers. Importantly, our data also demonstrate that changes in HDL function are dependent on the type of modification present on the HDL particle. Our findings suggest that modification of HDL with reactive aldehydes can impair some, but not all, of HDL’s athero‐protective functions in macrophages.

Purpose of Review The binding of high-density lipoprotein (HDL) to its primary receptor, scavenger receptor class B type 1 (SR-B1), is critical for lowering plasma cholesterol levels and reducing cardiovascular disease risk. This review provides novel insights into how the structural elements of SR-B1 drive efficient function with an emphasis on bidirectional cholesterol transport. Recent Findings We have generated a new homology model of full-length human SR-B1 based on the recent resolution of the partial structures of other class B scavenger receptors. Interrogating this model against previously published observations allows us to generate structurally informed hypotheses about SR-B1’s ability to mediate HDL-cholesterol (HDL-C) transport. Furthermore, we provide a structural perspective as to why human variants of SR-B1 may result in impaired HDL-C clearance. Summary A comprehensive understanding of SR-B1’s structure–function relationships is critical to the development of therapeutic agents targeting SR-B1 and modulating cardiovascular disease risk.

Scavenger receptor Class B type 1 (SR-B1) is a glycosylated integral membrane protein that serves as the primary receptor for high density lipoprotein (HDL) and plays a central role in the reverse cholesterol transport pathway (RCT). In RCT, HDL removes cholesterol within atherosclerotic plaques and delivers it to the liver for excretion. HDL-cholesterol is delivered into hepatocytes through interaction with SR-B1, highlighting the need to improve our understanding of SR-B1 structure-function relationships to effectively lower plasma cholesterol levels and cardiovascular disease risk. Currently, structural information about SR-B1 remains limited to a structure of short transmembrane-spanning peptide segment, homology models, and transient transfection studies in cultured cells. Towards our long-term goal of resolving a full-length structure of human SR-B1, we have expressed and purified full-length human SR-B1 from an Sf9 insect cell baculoviral infection system. To verify function, human full-length SR-B1 expressed in plated Sf9 cells displayed a significant increase in DiI-HDL binding and DiI-lipid uptake compared to empty vector controls. Using PFO-PAGE, we demonstrated that SR-B1 forms dimers and higher order oligomers in Sf9 cells. After functional verification, we successfully incorporated purified protein into detergent micelles. Thermal shift assays revealed purified SR-B1 remained stable over a 6-week period. Purified SR-B1 also maintained the ability to bind to apolipoprotein A-I (the main protein component of HDL) and holoparticle HDL with high affinity (K D = 42.77 ± 7.15 μg/mL) by microscale thermophoresis (MST). Additionally, SR-B1 was able to bind oxidized low density lipoprotein (oxLDL). Further, while glycosylation remains important for SR-B1 expression, we are the first to demonstrate by MST that SR-B1’s glycosylation status did not impact HDL binding affinity (K D of 42.77 ± 7.15 μg/mL for glycosylated vs. 41.25 ± 4.92 μg/mL for unglycosylated SR-B1). Availability of functional purified human SR-B1 is a critical step towards a high-resolution structure and these studies lay the foundation for understanding the dynamics of SR-B1-mediated cholesterol transport and novel ways to modulate cardiovascular disease risk.

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