Discipline: Biological Sciences
Subcategory: Biochemistry (not Cell and Molecular Biology and Genetics)
Session: 2
Leah Rowe - University of Arkansas at Pine Bluff
Co-Author(s): Dr. Linda S. May-Zhang, Department of Pharmacology, Vanderbilt University ; Dr. Sean S. Davies, Department of Pharmacology, Vanderbilt University
Background: While cardiovascular disease (CVD) is inversely associated with high density lipoprotein cholesterol (HDL-C), pharmacological interventions aimed to increase HDL-C have failed to reduce disease risk. Recent evidence suggest that CVD risk is more closely related to HDL function than HDL-C. In CVD, increased oxidative stress generates reactive lipid species that alter HDL function. Our laboratory has recently reported that isolevuglandins (IsoLGs), highly reactive lipid dicarbonyls generated in parallel to isoprostanes, cause deleterious consequences to HDL structure and function, including rendering macrophages more inflammatory [May-Zhang LS, et al. (2018) J Biol Chem, 293: 9176-9187]. Whether this gain-of-function is due to modification of proteins versus lipid in HDL is unknown. Hypothesis & Aim: Since IsoLG modified phosphatidylethanolamines (PEs) are reported to activate macrophages [Guo L, et al. (2015) Antioxid Redox Signal, 22: 1633-1645], we hypothesize that IsoLG modification of PE in HDL creates an inflammatory particle. The aim of this study is to determine the differential effects of modified apoA-I versus PE on particle size, composition, and function using synthetically engineered HDL. Methods: Experimental groups of synthetic HDL particles with IsoLG modified ApoA-I and/or PE compared with unmodified particles as controls were engineered from recombinant ApoA-I, phosphatidylcholine, and free cholesterol, and PE. Size and composition were characterized by native gel electrophoresis and fast protein liquid chromatography. Function was assessed by its ability to protect against lipopolysaccharide (LPS)-induced cytokine expression in primary murine macrophages. Results: IsoLG-modification of synthetic HDL creates a wider size distribution with an average of smaller particles. Inclusion of IsoLG-PE does not alter particle size, composition, or electrophoretic mobility. While IsoLG inhibited HDL’s protection against LPS-stimulated TNFα and IL-1β mRNA expression, particles containing IsoLG-PE causes a dramatic increase cytokine response compared to LPS alone. Conclusions & Significance: While IsoLG modification of both PE and ApoA-I contribute to dysfunctional HDL, our results suggest that IsoLG-modified PE causes a gain-of-inflammatory function. Future directions include testing the synthetic spherical HDL particles as well as testing for other functions including cholesterol efflux. Understanding the cause of HDL dysfunction will lead to the development of new diagnostic and therapeutic approaches to atherosclerosis.
Funder Acknowledgement(s): The study was funded by NIH grants P01 HL-116263 (SSD) and HL138745 (LSM). This research was performed under the mentorship of Drs. May-Zhang and Davies.
Faculty Advisor: Dr.Linda May-Zhang, linda.zhang.2@vanderbilt.edu
Role: During this research experiment, I started by learning how to make the synthetic HDL particles from previously obtained materials. I further learned how to make each component and I isolated the primary protein (ApoA-1) needed to construct the syntheic HDL particles for HDL modification. Once I made the HDL particles, I measured protein concentration and characterized the particles by native gel imagery.