HDL Glycoprotein Composition and Site-Specific Glycosylation Differentiates Between Clinical Groups and Affects IL-6 Secretion in Lipopolysaccharide-Stimulated Monocytes

Sridevi Krishnan,Atul N Parikh,Michiko Shimoda,Carlito B Lebrilla,Viviane N Ngassam,Emanual Maverakis,Jennifer T Smilowitz,Angela M Zivkovic,George A Kaysen,Barbara Grimes,Glenn M Chertow,Kirsten Johansen,Muchena J Kailemia,Romina Sacchi,Guillaume Luxardi

doi:10.1038/srep43728

Abstract

The goal of this pilot study was to determine whether HDL glycoprotein composition affects HDL’s immunomodulatory function. HDL were purified from healthy controls (n = 13), subjects with metabolic syndrome (MetS) (n = 13), and diabetic hemodialysis (HD) patients (n = 24). Concentrations of HDL-bound serum amyloid A (SAA), lipopolysaccharide binding protein (LBP), apolipoprotein A-I (ApoA-I), apolipoprotein C-III (ApoC-III), α-1-antitrypsin (A1AT), and α-2-HS-glycoprotein (A2HSG); and the site-specific glycovariations of ApoC-III, A1AT, and A2HSG were measured. Secretion of interleukin 6 (IL-6) in lipopolysaccharide-stimulated monocytes was used as a prototypical assay of HDL’s immunomodulatory capacity. HDL from HD patients were enriched in SAA, LBP, ApoC-III, di-sialylated ApoC-III (ApoC-III2) and desialylated A2HSG. HDL that increased IL-6 secretion were enriched in ApoC-III, di-sialylated glycans at multiple A1AT glycosylation sites and desialylated A2HSG, and depleted in mono-sialylated ApoC-III (ApoC-III1). Subgroup analysis on HD patients who experienced an infectious hospitalization event within 60 days (HD+) (n = 12), vs. those with no event (HD−) (n = 12) showed that HDL from HD+ patients were enriched in SAA but had lower levels of sialylation across glycoproteins. Our results demonstrate that HDL glycoprotein composition, including the site-specific glycosylation, differentiate between clinical groups, correlate with HDL’s immunomodulatory capacity, and may be predictive of HDL’s ability to protect from infection.

Highlights

A recent analysis of multiple genome-wide association studies of inflammatory diseases comprising a total of over 200,000 individuals found multiple shared risk loci between HDL and immune pathways[4]
The primary objectives of this study were to determine whether HDL glycoprotein composition: (1) differentiates between clinical groups, (2) modifies the effect of HDL on response phenotype in monocytes stimulated with lipopolysaccharide (LPS), and (3) is associated with risk for infectious hospitalization in a high-risk group
Using our in-house developed technology, a detailed analysis of HDL composition was conducted on each sample including the concentrations of the HDL bound proteins; ApoC-III, α-1-antitrypsin (A1AT), and α- 2-HS-glycoprotein (A2HSG), the quantification of the site-specific glycovariations of these proteins, as well as the concentrations of apolipoprotein A-I (ApoA-I), serum amyloid A (SAA), LPS binding protein (LBP), cholesterol, and measurement of HDL particle size by dynamic light scattering

Summary

Introduction

A recent analysis of multiple genome-wide association studies of inflammatory diseases comprising a total of over 200,000 individuals found multiple shared risk loci between HDL and immune pathways[4]. Correlating HDL composition with function is a daunting task because the highly complex HDL proteome has over 100 proteins, many of which are associated with cholesterol transport and lipid metabolism How these proteins are associated with the acute phase response, immunity, and inflammation is an area of active investigation[17,18,19]. Using our in-house developed technology, a detailed analysis of HDL composition was conducted on each sample including the concentrations of the HDL bound proteins; ApoC-III, α-1-antitrypsin (A1AT), and α- 2-HS-glycoprotein (A2HSG), the quantification of the site-specific glycovariations of these proteins, as well as the concentrations of ApoA-I, serum amyloid A (SAA), LPS binding protein (LBP), cholesterol, and measurement of HDL particle size by dynamic light scattering. Our results demonstrate that HDL composition, including the site-specific glycosylation, differentiates between clinical groups, correlates with HDL’s ability to modulate the LPS-induced monocyte cytokine response, and may be predictive of HDL’s ability to protect against infection

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Discussion

Conclusion