The SspA subtilisin-like protease of Streptococcus suis triggers a pro-inflammatory response in macrophages through a non-proteolytic mechanism

Laetitia Bonifait,Daniel Grenier

doi:10.1186/1471-2180-11-47

Abstract

BackgroundStreptococcus suis is a major swine pathogen worldwide that causes meningitis, septicemia, arthritis, and endocarditis. Using animal models, a surface-associated subtilisin-like protease (SspA) has recently been shown to be an important virulence factor for S. suis. In this study, we hypothesized that the S. suis SspA subtilisin-like protease may modulate cytokine secretion by macrophages thus contributing to the pathogenic process of meningitis.ResultsPhorbol 12-myristate 13-acetate-differentiated U937 macrophages were stimulated with recombinant SspA prior to monitor cytokine secretion by ELISA. Our results indicated that the recombinant SspA was able to dose-dependently induce IL-1β, IL-6, TNF-α, CXCL8 and CCL5 secretion in macrophages. The heat-inactivated protease was still able to induce cytokine secretion suggesting a non-proteolytic mechanism of macrophage activation. Using specific kinase inhibitors, evidence were bought that cytokine secretion by macrophages stimulated with the recombinant SspA involves the mitogen-activated protein kinase signal transduction pathway. While stimulation of macrophages with low concentrations of recombinant SspA was associated to secretion of high amounts of CCL5, the use of recombinant SspA at a high concentration resulted in low amounts of CCL5 detected in the conditioned medium. This was found to be associated with a proteolytic degradation of CCL5 by SspA. The ability of SspA to induce cytokine secretion in macrophages was confirmed using a mutant of S. suis deficient in SspA expression.ConclusionIn conclusion, this study identified a new mechanism by which the S. suis SspA may promote central nervous system inflammation associated with meningitis.

Highlights

Streptococcus suis is a major swine pathogen worldwide that causes meningitis, septicemia, arthritis, and endocarditis
Prior to determine the capacity of the recombinant subtilisin-like protease (SspA) of S. suis to induce an inflammatory response in PMAdifferentiated U937 macrophages, its effect on cell viability was evaluated
The levels of CXCL8 (Figure 1D) increased by 17-fold while that of CCL5 (Figure 1E) increased by 15-fold when the recombinant SspA was used at 0.33 μg/ml (Figure 1D-E)

Summary

Introduction

Streptococcus suis is a major swine pathogen worldwide that causes meningitis, septicemia, arthritis, and endocarditis. A surface-associated subtilisin-like protease (SspA) has recently been shown to be an important virulence factor for S. suis. We hypothesized that the S. suis SspA subtilisin-like protease may modulate cytokine secretion by macrophages contributing to the pathogenic process of meningitis. Our laboratory recently reported on the cloning of a 170 kDa subtilisin-like protease (SspA) found on the cell surface of S. suis [15]. This protease was found to possesses a high protein cleavage specificity and can degrade the Aa chain of fibrinogen preventing thrombin-mediated fibrin formation [15]. The exact contribution of the SspA in the pathogenic process of S. suis infections has not been clearly defined

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