Serum Amyloid A1 (SAA1) Revisited: Restricted Leukocyte-Activating Properties of Homogeneous SAA1.

Sara Abouelasrar Salama,Paul Proost,Mirre De Bondt,Vivian Louise Soares Oliveira,Jo Van Damme,Nele Berghmans,Mieke Gouwy,Mieke De Buck,Flavio A Amaral,Sofie Struyf

doi:10.3389/fimmu.2020.00843

Sara Abouelasrar Salama, Paul Proost + Show 8 more

Open Access

https://doi.org/10.3389/fimmu.2020.00843

Copy DOI

Abstract

Infection, sterile injury, and chronic inflammation trigger the acute phase response in order to re-establish homeostasis. This response includes production of positive acute phase proteins in the liver, such as members of the serum amyloid A (SAA) family. In humans the major acute phase SAAs comprise a group of closely related variants of SAA1 and SAA2. SAA1 was proven to be chemotactic for several leukocyte subtypes through activation of the G protein-coupled receptor FPRL1/FPR2. Several other biological activities of SAA1, such as cytokine induction, reported to be mediated via TLRs, have been debated recently. Especially commercial SAA1, recombinantly produced in Escherichia coli, was found to be contaminated with bacterial products confounding biological assays performed with this rSAA1. We purified rSAA1 by RP-HPLC to homogeneity, removing contaminants such as lipopolysaccharides, lipoproteins and formylated peptides, and re-assessed several biological activities attributed to SAA1 (chemotaxis, cytokine induction, MMP-9 release, ROS generation, and macrophage differentiation). The homogeneous rSAA1 (hrSAA1) lacked most cell-activating properties, but its leukocyte-recruiting capacity in vivo and it’s in vitro synergy with other leukocyte attractants remained preserved. Furthermore, hrSAA1 maintained the ability to promote monocyte survival. This indicates that pure hrSAA1 retains its potential to activate FPR2, whereas TLR-mediated effects seem to be related to traces of bacterial TLR ligands in the E. coli-produced human rSAA1.

Highlights

The serum amyloid A (SAA) proteins form a family that is highly conserved in a wide number of species ranging from fish to humans [1]
In order to rule out the possibility that exposure to harsh solvents (50% ACN in 0.1% trifluoroacetic acid (TFA)) would alter the biological activity of homogenous recombinant SAA1 (rSAA1) (hrSAA1), rSAA1 was incubated in 50% ACN in 0.1% TFA for a period of 1 h followed by lyophilization and reconstitution
We purified commercially available rSAA1 expressed in E. coli, which has been used in most studies dealing with its biological activities, via reversed phase-high performance liquid chromatography (RP-HPLC) to produce homogenous rSAA1 free of any residual bacterial contaminants

Summary

Introduction

The serum amyloid A (SAA) proteins form a family that is highly conserved in a wide number of species ranging from fish to humans [1]. The remarkable conservation of SAA throughout evolution points toward a rather important biological role. Humans have four distinct SAA genes giving rise to SAA1, SAA2, SAA3, and SAA4. SAA1 and SAA2 are upregulated during the acute phase response and are referred to as acute-SAA (A-SAA) [2]. Expression of A-SAA primarily occurs in the liver in response to inflammatory cytokines such as interleukin-1β (IL-1β), IL-6 and tumor necrosis factor–α (TNF-α) [2].

Objectives

Methods

Results

Conclusion